U.S. patent application number 12/127930 was filed with the patent office on 2008-11-27 for metaxalone products, method of manufacture, and method of use.
This patent application is currently assigned to MUTUAL PHARMACEUTICAL COMPANY, INC.. Invention is credited to Matthew W. Davis, Jie Du, Richard H. Roberts.
Application Number | 20080292584 12/127930 |
Document ID | / |
Family ID | 40072602 |
Filed Date | 2008-11-27 |
United States Patent
Application |
20080292584 |
Kind Code |
A1 |
Roberts; Richard H. ; et
al. |
November 27, 2008 |
METAXALONE PRODUCTS, METHOD OF MANUFACTURE, AND METHOD OF USE
Abstract
Disclosed herein are methods of using metaxalone. In one
embodiment, the method comprises determining that a patient in need
metaxalone therapy is taking a substance that is a n inhibitor or
an inducer of a cytochrome p450 isozyme, wherein the cytochrome
P450 is CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or CYP3A4,
and adjusting administration to the patient of metaxalone or the
substance to avoid an adverse event associated with metaxalone. In
another embodiment, the method comprises informing a user that
metaxalone ais metabolized by a cytochrome p450 isozyme, wherein
the cytochrome P450 is CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6,
CYP2E1, or CYP3A4. Also included are articles of manufacture
comprising a container containing a dosage form of metaxalone,
wherein the container is associated with published material
informing that metaxalone affects activity of a cytochrome p450
isozyme. Also disclosed are a method of treatment and a method of
manufacturing a metaxalone product.
Inventors: |
Roberts; Richard H.;
(Lakewood, NJ) ; Du; Jie; (Lansdale, PA) ;
Davis; Matthew W.; (Erwinna, PA) |
Correspondence
Address: |
CANTOR COLBURN, LLP
20 Church Street, 22nd Floor
Hartford
CT
06103
US
|
Assignee: |
MUTUAL PHARMACEUTICAL COMPANY,
INC.
Philadelphia
PA
|
Family ID: |
40072602 |
Appl. No.: |
12/127930 |
Filed: |
May 28, 2008 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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11349534 |
Feb 6, 2006 |
|
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12127930 |
|
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60726861 |
Oct 14, 2005 |
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Current U.S.
Class: |
424/85.4 ;
424/730; 424/736; 424/751; 514/178; 514/211.05; 514/230.8;
514/252.18; 514/253.01; 514/253.06; 514/253.09; 514/254.07;
514/256; 514/29; 514/307; 514/365; 514/376 |
Current CPC
Class: |
A61K 31/47 20130101;
A61K 31/7042 20130101; A61K 36/752 20130101; A61K 31/421 20130101;
A61K 36/38 20130101; A61P 21/02 20180101; A61K 36/38 20130101; A61K
31/554 20130101; A61K 36/752 20130101; A61K 36/81 20130101; A61K
31/56 20130101; A61P 21/00 20180101; A61K 31/5375 20130101; A61K
36/81 20130101; A61K 31/506 20130101; A61K 2300/00 20130101; A61K
2300/00 20130101; A61K 31/426 20130101; A61K 2300/00 20130101; A61K
31/496 20130101 |
Class at
Publication: |
424/85.4 ;
514/376; 514/253.09; 514/253.01; 514/307; 514/365; 514/230.8;
514/29; 514/211.05; 514/178; 424/736; 514/252.18; 514/254.07;
514/253.06; 514/256; 424/730; 424/751 |
International
Class: |
A61K 38/21 20060101
A61K038/21; A61K 31/421 20060101 A61K031/421; A61K 31/496 20060101
A61K031/496; A61K 31/47 20060101 A61K031/47; A61K 31/426 20060101
A61K031/426; A61K 31/5375 20060101 A61K031/5375; A61K 36/38
20060101 A61K036/38; A61P 21/02 20060101 A61P021/02; A61K 36/81
20060101 A61K036/81; A61K 31/7042 20060101 A61K031/7042; A61K
31/554 20060101 A61K031/554; A61K 31/56 20060101 A61K031/56; A61K
36/752 20060101 A61K036/752; A61K 31/506 20060101 A61K031/506 |
Claims
1.-13. (canceled)
14. A method of preventing sedation in a patient in need of
metaxalone comprising determining a dosing regimen for metaxalone
to be administered to a patient in need thereof, determining that a
substance that is a known inhibitor of a cytochrome P450 isozyme
(CYP) selected from CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6,
CYP2E1, and CYP3A4 is currently administered or will be
administered to the patient, and altering dosing of metaxalone
administered to the patient from the determined dosing regimen
while the known inhibitor of the CYP is coadministered to the
patient to prevent sedation.
15. The method of claim 14 further comprising monitoring plasma
concentration of metaxalone in the patient after administration of
metaxalone to the patient.
16. The method of claim 14, wherein altering dosing of metaxalone
comprises decreasing a dose of metaxalone administered to the
patient; or decreasing a number of doses per day administered to
the patient.
17. The method of claim 14, wherein the determined dosing regimen
is 800 mg, three or four times daily.
18. The method of claim 14 wherein the CYP is CYP3A4 and the known
inhibitor of CYP3A4 is an HIV antiviral, delavirdine, indinavir,
nelfinavir, ritonavir; amiodarone, aprepitant, cinchloramphenicol,
cimetidine, clarithromycin, diethyl-dithiocarbamate, diltiazem,
erythromycin, fluconazole, fluvoxamine, gestodene, grapefruit
juice, Seville orange juice, imatinib, itraconazole, ketoconazole,
mifepristone, nefazodone, norfloxacin, norfluoxetine, mibefradil,
star fruit, verapamil, or voriconazole; the CYP is CYP1A2 and the
known inhibitor of CYP1A2 is amiodarone, cimetidine, a
fluoroquinolone, fluvoxamine, furafylline, interferon, methoxsalen,
or mibefradil; or the CYP is CYP2E1 and the known inhibitor of
CYP2E1 is diethyl-dithiocarbamate or disulfuram.
19. A method of optimizing the dose of metaxalone to administer to
a patient in need of a skeletal muscle relaxant, comprising
determining that a substance that is a known inducer or a known
inhibitor of a cytochrome P450 isozyme (CYP) selected from CYP1A2,
CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, and CYP3A4 is administered
to a patient in need of a skeletal muscle relaxant; and optimizing
a dose of metaxalone administered to the patient by administering a
dose of metaxalone greater than a standard dose if a known inducer
of the CYP is administered to the patient or administering a dose
less than the standard dose of metaxalone if a known inhibitor of
the CYP is administered to the patient.
20. The method of 19, wherein the standard dose of metaxalone is
800 mg.
21. The method of claim 19, further comprising optimizing a
frequency per day at which the optimized dose is taken by the
patient.
22. The method of claim 19, further comprising monitoring plasma
concentration of metaxalone in the patient to determine if the
patient is at risk of a subtherapeutic outcome or of a metaxalone
toxicity; and optimizing the dose of metaxalone administered to the
patient to alleviate the risk.
23.-37. (canceled)
38. A method of avoiding an adverse event when administering
metaxalone, comprising determining that a patient in need of
metaxalone therapy is taking a substance that is a known inhibitor
or a known inducer of cytochrome P450 isozyme (CYP) selected from
CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, and CYP3A4, and
adjusting administration to the patient of metaxalone or the
substance to avoid an adverse event associated with a change in the
metabolism of metaxalone.
39. The method of claim 38, wherein the method further comprises
informing the patient or the patient's medical care worker that
administration of metaxalone with a substance that is a known
inhibitor of the CYP can result in increased plasma concentration
of metaxalone; informing the patient or the patient's medical care
worker that administration of metaxalone with a substance that is a
known inducer of the CYP can result in decreased plasma
concentration of metaxalone; or monitoring the patient.
40.-64. (canceled)
65. The method of claim 19, wherein the CYP is CYP1A2, CYP2E1, or
CYP3A4.
66. The method of claim 19, wherein the CYP is CYP3A4, and the
substance is an inhibitor of CYP3A4 selected from an HIV antiviral;
amiodarone, aprepitant, cinchloramphenicol, cimetidine,
clarithromycin, diethyl-dithiocarbamate, diltiazem, erythromycin,
fluconazole, fluvoxamine, gestodene, grapefruit juice, Seville
orange juice, imatinib, itraconazole, ketoconazole, mifepristone,
nefazodone, norfloxacin, norfluoxetine, mibefradil, star fruit,
verapamil, and voriconazole; or an inducer of CYP3A4 selected from
an HIV Antiviral; a barbiturate, carbamazepine, efavirenz, a
glucocorticoid, modafinil, nevirapine, phenobarbital, phenyloin,
rifampin, St. John's wort, troglitazone, oxcarbazepine,
pioglitazone, and rifabutin.
67. The method of claim 19, wherein the CYP is CYP1A2, and the
substance is an inhibitor of CYP1A2 selected from amiodarone,
cimetidine, a fluoroquinolone, fluvoxamine, furafylline,
interferon, methoxsalen, and mibefradil; or an inducer of CYP1A2
selected from insulin, methyl cholanthrene, modafinil, nafcillin,
beta-naphthoflavone, omeprazole, and tobacco.
68. The method of claim 19, wherein the CYP is CYP2E1, and the
substance is an inhibitor of CYP2E1 selected from
diethyl-dithiocarbamate and disulfuram; or an inducer of CYP2E1
selected from ethanol and isoniazid.
69. The method of claim 39, wherein monitoring the patient
comprises: monitoring the patient's plasma concentration of
metaxalone; monitoring the patient for a symptom of an active agent
interaction between the substance and metaxalone; monitoring the
patient for an adverse reaction resulting from coadministration of
the substance and metaxalone; monitoring the patient for an adverse
reaction associated with metaxalone; monitoring the patient for a
metaxalone-associated toxicity; monitoring the patient for a
sub-therapeutic outcome for metaxalone; monitoring the patient for
reduced efficacy of metaxalone; monitoring the patient for an
adverse reaction or sub-therapeutic outcome associated with reduced
plasma concentration of metaxalone; or monitoring the patient for
an adverse reaction associated with an elevated plasma
concentration of metaxalone.
Description
CROSS REFERENCE TO RELATED APPLICATION
[0001] This application is a continuation in part of U.S.
application Ser. No. 11/349,534 filed Feb. 6, 2006, which claims
the benefit of U.S. Provisional Application Ser. No. 60/726,861
filed Oct. 14, 2005, each of which is hereby incorporated by
reference in its entirety.
BACKGROUND
[0002] This application relates to metaxalone products for
therapeutic purposes, and in particular to improved methods of use
of metaxalone.
[0003] Metaxalone, 5-[(3,5-dimethylphenoxy)methyl]-2-oxazolidinone,
is used as a skeletal muscle relaxant. The mechanism of action of
metaxalone in humans has not been established but may be due to
general central nervous system depression.
[0004] Metaxalone was approved by the U.S. Food and Drug
Administration (FDA) in 1962 as an adjunct to rest, physical
therapy, and other measures for the relief of discomforts
associated with acute, painful musculoskeletal conditions, such as
muscles in spasm. Metaxalone is marketed in the United States under
the brand name SKELAXIN.RTM.. The dosage forms currently approved
for marketing are tablets containing 400 milligrams (mg) or 800 mg
of metaxalone. The currently recommended dose for adults and
children over 12 years of age is 800 mg, three to four times a
day.
[0005] Food can affect gastric emptying, and may also alter the
release of an active agent from a dosage form, the solubilization
of the active agent, and the transport of the active agent across
the intestinal wall. For lipophilic, water-insoluble active agents,
fatty meals can increase gastric residence time thereby increasing
the time available for solubilization and also may enhance the
solubilization of the active agent by the lipids contained in the
meal. According to U.S. Pat. No. 6,407,128, evaluation of the
effect of food on the pharmacokinetics of metaxalone showed that
food increased the rate and extent of absorption of a 400 mg oral
dosage form in humans.
[0006] Studies directed to possible interactions of metaxalone with
other active agents have been limited. There have been no detailed
studies of the specific enzymes involved in metabolism of
metaxalone or of the inhibitory or inducing effects of metaxalone
on any Phase I or Phase II metabolic enzymes. In particular, there
appear to be no published studies of the inhibitory and inducing
effects of metaxalone on particular human cytochrome p450 isozymes
or the possible metabolism of metaxalone by particular human
cytochrome p450 isozymes.
[0007] Several major enzymes and pathways are involved in drug
metabolism. Pathways of drug biotransformation are usually divided
into two major groups of reactions: Phase I and Phase II
metabolism.
[0008] Some typical examples of Phase I metabolism include
oxidation, hydrolysis and reduction. Examples of Phase I enzymes
involved in oxidation reactions are the cytochrome p450
monooxygenase system, the flavin-containing monooxygenase system,
alcohol dehydrogenase and aldehyde dehydrogenase, monoamine
oxidase, and peroxidases for co-oxidation. Examples of Phase I
enzymes involved in reduction are NADPH-cytochrome p450 reductase
and reduced (ferrous) cytochrome p450. Examples of Phase I
hydrolysis enzymes are epoxide hydrolase, esterases and
amidases.
[0009] Phase II metabolism involves conjugation reactions. Typical
conjugation reactions are glucuronidation, sulfation, amino acid
conjugation, acetylation, methylation, and mercapturic acid
conjugation. Examples of Phase II metabolic enzymes are glutathione
S-transferases (GSTs), mercapturic acid biosynthetic enzymes
(transpeptidases, peptidases, and N-acetylases),
UDP-glucoron(os)yltransferases, N-acetyltransferases, amino acid
N-acyl transferases, and sulfotransferases.
[0010] One of the most important groups of Phase I enzymes are the
cytochrome p450 monooxygenase system enzymes. The cytochrome p450
enzymes are a highly diverse superfamily of enzymes. NADPH is
required as a coenzyme and oxygen is used as a substrate. Each
enzyme is termed an isoform or isozyme since each derives from a
different gene.
[0011] Many members of the cytochrome p450 family are known to
metabolize active agents in humans. Active agent interactions
associated with metabolism by cytochrome p450 isoforms generally
result from enzyme inhibition or enzyme induction. Enzyme
inhibition often involves competition between two active agents for
the substrate binding site of the enzyme, although other mechanisms
for inhibition exist. Enzyme induction occurs when an active agent
activates an enzyme or stimulates the synthesis of more enzyme
protein, enhancing the enzyme's metabolizing capacity.
[0012] Cytochrome p450 isozymes identified as important in active
agent metabolism are CYP1A2, CYP2A6, CYP2B6, CYP2C9, CYP2C19,
CYP2D6, CYP2E1, and CYP3A4. Examples of cytochrome p450 enzymes
known to be involved in active agent interactions are the CYP3A
subfamily, which is involved in many clinically significant active
agent interactions, including those involving non-sedating
antihistamines and cisapride, and CYP2D6, which is responsible for
the metabolism of many psychotherapeutic agents, such as
thioridazine. CYP3A4 and CYP1A2 enzymes are involved in active
agent interactions involving theophylline. CYP2C9, CYP1A2, and
CYP2C19 are involved in active agent interactions involving
warfarin. Phenyloin and fosphenyloin are metabolized by CYP1A2,
CYP2C9, CYP2C19, and CYP3A4; mexiletine is metabolized by CYP2D6
and CYP1A2; and propafenone is metabolized by CYP2D6, CYP3A4, and
CYP1A2.
[0013] Additionally, several cytochrome p450 isozymes are known to
be genetically polymorphic, leading to altered substrate
metabolizing ability in some individuals. Allelic variants of
CYP2D6 are the best characterized, with many resulting in an enzyme
with reduced, or no, catalytic activity. Gene duplication also
occurs. As a result, four phenotypic subpopulations of metabolizers
of CYP2D6 substrates exist: poor (PM), intermediate (IM), extensive
(EM), and ultrarapid (UM). The genetic polymorphisms vary depending
on the population in question. For example, Caucasian populations
contain a large percentage of individuals who are poor
metabolizers, due to a deficiency in CYP2D6--perhaps 5-10% of the
population, while only 1-2% of Asians are PMs. CYP2C9, which
catalyzes the metabolism of a number of commonly used active
agents, including that of warfarin and phenyloin, is also
polymorphic. The two most common CYP2C9 allelic variants have
reduced activity (5-12%) compared to the wild-type enzyme. Genetic
polymorphism also occurs in CYP2C19, for which at least 8 allelic
variants have been identified that result in catalytically inactive
protein. About 3% of Caucasians are poor metabolizers of active
agents metabolized by CYP2C19, while 13-23% of Asians are poor
metabolizers of active agents metabolized by CYP2C19.
[0014] Previous in vivo data from Bruce et al. (Bruce R B et al.
(1966) Metabolism of metaxalone. J Med Chem. 9, 286 288) showed
that 27% of metaxalone was converted to a carboxylic acid
derivative that appeared to be subsequently glucuronidated to an
additional metabolite. Bruce et al did not identify the metabolic
enzymes involved in this process. However, the FDA has suggested
that identification of the particular cytochrome P450 isozymes
involved is warranted when cytochrome P450 enzymes account for at
least 25% of the total clearance of an active agent, as observed
for metaxalone, since in vitro identification of the cytochrome
P450 isozymes that metabolize a particular active agent helps
predict the potential for in vivo active agent interactions, the
impact of polymorphic enzyme activity on active agent disposition,
and the formation of toxic or active metabolites.
[0015] Active agent interactions present a health risk to patients
and a medical challenge for all medical care workers. Various
studies of adverse reactions from exposure to active agents have
found that 6.5-23% of the adverse reactions result from active
agent interactions. Unfortunately, each year a number of deaths
occur as the direct result of patients taking a new prescription
pharmaceutical product in combination with their existing
medication regimen. By understanding the unique functions and
characteristics of Phase I and Phase II metabolic enzymes, such as
the cytochrome p450 enzyme superfamily, medical care workers such
as physicians and pharmacists may better anticipate, avoid, or
safely manage active agent interactions and may predict or explain
an individual's response to a particular therapeutic regimen.
[0016] There accordingly remains a need in the art for improved
methods for the administration and use of metaxalone, in particular
methods that take into account the effects of metaxalone on
activity of Phase I and Phase II metabolic enzymes, including the
cytochrome P450 isozymes.
SUMMARY
[0017] Disclosed herein are methods of using metaxalone. Metaxalone
can be used in the treatment of various diseases or conditions,
including, for example, musculoskeletal conditions and head
pain.
[0018] In an embodiment, the method comprises administering
metaxalone and a substance that is a known inhibitor or a known
inducer of CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or
CYP3A4 to a patient; and monitoring the patient during
administration of metaxalone and the substance.
[0019] In an embodiment, the method comprises determining that a
patient in need metaxalone therapy is taking a substance that is a
known inhibitor of a cytochrome p450 isozyme, wherein the
cytochrome P450 is CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1,
or CYP3A4, and adjusting administration to the patient of
metaxalone or the substance to avoid an adverse event associated
with metaxalone.
[0020] In an embodiment, the method comprises administering
metaxalone and an inhibitor of a cytochrome p450 isozyme, wherein
the cytochrome P450 is CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6,
CYP2E1, or CYP3A4 to a patient; and altering dosing of the
inhibitor or metaxalone for the patient if metaxalone plasma
concentration of the patient increases during coadministration with
the inhibitor.
[0021] In an embodiment, the method comprises administering
metaxalone and an inducer of CYP3A4 to a patient; and altering
dosing of the inducer of CYP3A4 or metaxalone for the patient if
metaxalone plasma concentration of the patient decreases during
coadministration with the inducer of CYP3A4.
[0022] In an embodiment, the method comprises administering
metaxalone and an inhibitor of CYP1A2, CYP2C8, CYP2C9, CYP2C19,
CYP2D6, CYP2E1, or CYP3A4 to a patient; determining that the
patient experiences a metaxalone-associated toxicity during
coadministration with the inhibitor; and altering dosing of the
inhibitor or metaxalone such that the metaxalone-associated
toxicity is reduced.
[0023] In an embodiment, the method comprises administering
metaxalone and an inducer of CYP1A2, CYP2C8, CYP2C9, CYP2C19,
CYP2D6, CYP2E1, or CYP3A4 to a patient; determining that the
patient experiences a subtherapeutic outcome for metaxalone during
coadministration with the inducer; and altering dosing of the
inducer or metaxalone such that efficacy is achieved.
[0024] In an embodiment, the method comprises determining that a
patient in need metaxalone therapy is taking a substance that is a
known inducer of CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1,
or CYP3A4, and adjusting administration to the patient of
metaxalone or the substance to avoid a subtherapeutic outcome with
metaxalone.
[0025] In an embodiment, the method comprises administering
metaxalone to a patient in need of metaxalone therapy; determining
that a known inhibitor of CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6,
CYP2E1, or CYP3A4 is administered to the patient; and monitoring
the patient during administration of metaxalone and the known
inhibitor for an adverse event associated with metaxalone.
[0026] In an embodiment, the method comprises administering
metaxalone to a patient in need of metaxalone therapy; determining
that a known inducer of CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6,
CYP2E1, or CYP3A4 is administered to the patient; and monitoring
the patient during administration of metaxalone and the known
inducer for reduced efficacy of metaxalone.
[0027] In an embodiment, the method comprises determining for a
patient to whom metaxalone is going to be administered or is being
administered whether a substance that is currently being or will be
administered to the patient is an inhibitor of a cytochrome P450
isozyme (CYP) selected from CYP1A2, CYP2C8, CYP2C9, CYP2C19,
CYP2D6, CYP2E1, and CYP3A4; and determining risk for the patient of
a metaxalone-associated toxicity resulting from inhibition of
metaxalone metabolism by the CYP during coadministration of
metaxalone and the substance.
[0028] In an embodiment, the method comprises determining for a
patient to whom metaxalone is going to be administered or is being
administered whether a substance that is currently being or will be
administered to the patient is an inducer of a cytochrome P450
isozyme (CYP) selected from CYP1A2, CYP2C8, CYP2C9, CYP2C19,
CYP2D6, CYP2E1, and CYP3A4; and determining risk for the patient of
a subtherapeutic outcome for metaxalone resulting from induction of
metaxalone metabolism by the CYP during coadministration of
metaxalone and the substance.
[0029] In an embodiment, the method comprises determining a dosing
regimen for metaxalone to be administered to a patient in need
thereof, determining that a substance that is a known inhibitor of
a cytochrome P450 isozyme (CYP) selected from CYP1A2, CYP2C8,
CYP2C9, CYP2C19, CYP2D6, CYP2E1, and CYP3A4 is currently
administered or will be administered to the patient, and altering
dosing of metaxalone administered to the patient from the
determined dosing regimen while the known inhibitor of the CYP is
coadministered to the patient to prevent sedation.
[0030] In an embodiment, the method comprises determining that a
substance that is a known inducer or a known inhibitor of a
cytochrome P450 isozyme (CYP) selected from CYP1A2, CYP2C8, CYP2C9,
CYP2C19, CYP2D6, CYP2E1, and CYP3A4 is administered to a patient in
need of a skeletal muscle relaxant; and optimizing a dose of
metaxalone administered to the patient by administering a dose of
metaxalone greater than a standard dose if a known inducer of the
CYP is administered to the patient or administering a dose less
than a standard dose of metaxalone if a known inhibitor of the CYP
is administered to the patient.
[0031] In another embodiment, the method comprises informing a user
that metaxalone is metabolized by a cytochrome p450 isozyme,
wherein the cytochrome P450 is CYP1A2, CYP2C8, CYP2C9, CYP2C19,
CYP2D6, CYP2E1, or CYP3A4.
[0032] In another embodiment, the method comprises informing a user
that metaxalone inhibits a cytochrome p450 isozyme.
[0033] In yet another embodiment, the method comprises informing a
user that metaxalone induces a cytochrome p450 isozyme.
[0034] In another embodiment, the method comprises informing a user
that metaxalone is metabolized by a cytochrome P450 isozyme (CYP),
wherein the CYP is CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1,
or CYP3A4; and that there is a potential active agent interaction
for metaxalone with an active agent that is a substrate, inhibitor,
or inducer of the CYP or that taking metaxalone with the active
agent can affect the plasma concentration, bioavailability, safety,
efficacy, or a combination comprising at least one of the foregoing
of metaxalone or the active agent.
[0035] In another embodiment, the method comprises informing a user
that metaxalone is an inhibitor or an inducer of a cytochrome p450
isozyme and administration of metaxalone with a substance that is a
substrate of the cytochrome p450 isozyme can affect the plasma
concentration, bioavailability, safety, efficacy, or a combination
comprising at least one of the foregoing of the substance.
[0036] In yet another embodiment, the method comprises obtaining
metaxalone from a container providing information that metaxalone
affects activity of a cytochrome p450 isozyme.
[0037] In yet another embodiment, the method comprises obtaining
metaxalone from a container providing information that metaxalone
is metabolized by a cytochrome p450 isozyme.
[0038] In yet another embodiment, the method comprises obtaining
metaxalone from a container providing information that metaxalone
is an inhibitor or an inducer of a cytochrome p450 isozyme.
[0039] In yet another embodiment, the method comprises
administering to a patient metaxalone and an active agent; and
informing the patient that metaxalone affects activity of a
cytochrome p450 isozyme.
[0040] In yet another embodiment, the method comprises informing a
user that metaxalone affects activity of a cytochrome p450 isozyme;
that administration of metaxalone with a substance can affect the
plasma concentration, bioavailability, safety, efficacy, or a
combination comprising at least one of the foregoing of metaxalone
or the substance; and that any effect on the plasma concentration,
bioavailability, safety, efficacy, or a combination comprising at
least one of the foregoing of metaxalone or the substance can vary
with administration of metaxalone with or without food.
[0041] In another embodiment, the method comprises obtaining
metaxalone from a container providing information that metaxalone
affects activity of a cytochrome p450 isozyme; that administration
of metaxalone with a substance can affect the plasma concentration,
bioavailability, safety, efficacy, or a combination comprising at
least one of the foregoing of metaxalone or the substance; and that
any effect on the plasma concentration, bioavailability, safety,
efficacy, or a combination comprising at least one of the foregoing
of metaxalone or the substance can vary with administration of
metaxalone with or without food.
[0042] Also disclosed herein are methods of manufacturing a
metaxalone product.
[0043] In one embodiment, the method comprises packaging a
metaxalone dosage form with information that metaxalone affects
activity of a cytochrome p450 isozyme.
[0044] In another embodiment, the method comprises packaging a
metaxalone dosage form with information that metaxalone is
metabolized by a cytochrome p450 isozyme.
[0045] In another embodiment, the method comprises packaging a
metaxalone dosage form with information that metaxalone is an
inhibitor or an inducer of a cytochrome p450 isozyme.
[0046] In yet another embodiment, the method comprises packaging a
metaxalone dosage form with information that metaxalone affects
activity of a cytochrome p450 isozyme; that administration of
metaxalone with a substance can affect the plasma concentration,
safety, or efficacy of the metaxalone or the substance; and that
any effect on the plasma concentration, bioavailability, safety,
efficacy, or a combination comprising at least one of the foregoing
of metaxalone or the substance can vary with administration of
metaxalone with or without food.
[0047] Also disclosed herein are articles of manufacture comprising
a container containing a dosage form of metaxalone.
[0048] In one embodiment, the container is associated with
published material informing that metaxalone affects activity of a
cytochrome p450 isozyme.
[0049] In another embodiment, the container is associated with
published material informing that metaxalone is metabolized by a
cytochrome p450 isozyme.
[0050] In another embodiment, the container is associated with
published material informing that metaxalone is an inhibitor or an
inducer of a cytochrome p450 isozyme.
[0051] In yet another embodiment, the container is associated with
published material informing that metaxalone affects activity of a
cytochrome p450 isozyme; that administration to a patient of
metaxalone with a substance can affect plasma concentration,
bioavailability, safety, efficacy, or a combination comprising at
least one of the foregoing of metaxalone or the substance; and that
any effect on the plasma concentration, bioavailability, safety,
efficacy, or a combination comprising at least one of the foregoing
of metaxalone or the substance can vary with administration of
metaxalone with or without food.
[0052] In yet another embodiment, the article comprises a container
comprising a dosage form of metaxalone, and published material. In
one embodiment, the published material informs that there is a
potential active agent interaction with warfarin; or that
administration with warfarin can affect the plasma concentration,
bioavailability, safety, efficacy, or a combination comprising at
least one of the foregoing of metaxalone or warfarin. In another
embodiment, the published material informs that metaxalone is a
substrate of CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or
CYP3A4, or that metaxalone is an inhibitor of CYP1A2, CYP2B6,
CYP2C9, CYP2C19, CYP2D6, CYP2E1, or CYP3A4, or that metaxalone is
an inducer of CYP1A2 or CYP3A4.
[0053] Also disclosed herein is an article of manufacture
comprising packaging material and a product contained within the
packaging material, wherein the product comprises, as at least one
active ingredient, metaxalone, and wherein the packaging material
comprises a label approved by a regulatory agency for the product
which states that metaxalone affects activity of a cytochrome p450
isozyme.
[0054] Also disclosed herein is a method of using an active agent
that is a known substrate, inhibitor, or inducer of a cytochrome
P450 isozyme (CYP) selected from CYP1A2, CYP2C8, CYP2C9, CYP2C19,
CYP2D6, CYP2E1, and CYP3A4 or a substrate of a cytochrome p450
isozyme.
[0055] In one embodiment, the method comprises informing a user
that metaxalone affects activity of a cytochrome p450 isozyme and
that administration of the active agent with metaxalone can affect
the plasma concentration, bioavailability, safety, efficacy, or a
combination comprising at least one of the foregoing of the active
agent or metaxalone.
[0056] In another embodiment, the method comprises obtaining an
active agent that is a known substrate, inhibitor, or inducer of a
cytochrome P450 isozyme (CYP) selected from CYP1A2, CYP2C8, CYP2C9,
CYP2C19, CYP2D6, CYP2E1, and CYP3A4 or a substrate of a cytochrome
p450 isozyme from a container providing information that metaxalone
affects activity of a cytochrome p450 isozyme and that
administration of the active agent with metaxalone can affect
plasma concentration, bioavailability, safety, efficacy, or a
combination comprising at least one of the foregoing of the active
agent or metaxalone.
[0057] Also disclosed herein is a method of manufacturing a
pharmaceutical product comprising an active agent that is a known
substrate, inhibitor, or inducer of a cytochrome P450 isozyme (CYP)
selected from CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, and
CYP3A4 or a substrate of a cytochrome p450 isozyme.
[0058] In one embodiment, the method comprises packaging a dosage
form of the active agent that is a known substrate, inhibitor, or
inducer of a cytochrome P450 isozyme (CYP) selected from CYP1A2,
CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, and CYP3A4 or a substrate
of a cytochrome p450 isozyme with information that metaxalone
affects activity of a cytochrome p450 isozyme and that
administration of the active agent with metaxalone can affect
plasma concentration, bioavailability, safety, efficacy, or a
combination comprising at least one of the foregoing of the active
agent or metaxalone.
[0059] Also disclosed herein is an article of manufacture
comprising a container containing a dosage form of an active agent
that is a known substrate, inhibitor, or inducer of a cytochrome
P450 isozyme (CYP) selected from CYP1A2, CYP2C8, CYP2C9, CYP2C19,
CYP2D6, CYP2E1, and CYP3A4 or a substrate of a cytochrome p450
isozyme. The container is associated with published material
informing that metaxalone affects activity of a cytochrome p450
isozyme and that administration to a patient of the active agent
and metaxalone can affect plasma concentration, bioavailability,
safety, efficacy, or a combination comprising at least one of the
foregoing of the active agent or metaxalone.
[0060] These and other embodiments, advantages and features of the
present invention become clear when detailed description and
examples are provided in subsequent sections.
DETAILED DESCRIPTION
[0061] Disclosed herein are methods of using metaxalone and
metaxalone products. The inventors have determined that metaxalone
is metabolized by several cytochrome P450 isozymes and identified
risks associated with administration of metaxalone with a substance
which is an inhibitor or an inducer of this isozyme. With the
knowledge of the particular information, a medical care worker can
better avoid or safely manage an active agent interaction in a
patient between metaxalone and the substance, and its resultant
effects on efficacy or safety of metaxalone. Specifically,
knowledge that metaxalone is metabolized by the particular
cytochrome P450 isozymes permits the administration of metaxalone
and a substance which is an inhibitor or an inducer of this isozyme
to a patient to be optimized for the patient by a medical care
worker to provide safe use of metaxalone, while oftentimes reducing
or minimizing side effects or adverse events resulting from
interactions with the substance. Knowledge of the particular
information permits a medical care worker to use metaxalone to
treat a patient that is taking another substance which is an
inhibitor or an inducer of the particular cytochrome P450 isozymes
such that a side effect, an adverse reaction, or an active agent
interaction between metaxalone and the substance can be avoided in
the patient. Knowledge of the particular information allows proper
dosing, dispensing, and administration of metaxalone or the
substance to the patient by the patient's medical care worker to
avoid, or reduce risk of occurrence of a sub-therapeutic effect, a
side effect, an adverse reaction, or an active agent interaction
between metaxalone and the substance and alerts the patient and the
patient's medical care worker to the need to monitor the patient
for symptoms of a sub-therapeutic effect, a side effect, an adverse
reaction, or an active agent interaction between metaxalone and the
substance.
[0062] Metaxalone therapy can be considered optimal when effective
plasma levels are reached when required. In addition, peak plasma
values (C.sub.max) should be as low as possible so as to reduce the
incidence and severity of possible side effects.
[0063] The terms "a" and "an" do not denote a limitation of
quantity, but rather denote the presence of at least one of the
referenced item. The term "or" means "and/or". The terms
"comprising", "having", "including", and "containing" are to be
construed as open-ended terms (i.e., meaning "including, but not
limited to"). Recitation of ranges of values are merely intended to
serve as a shorthand method of referring individually to each
separate value falling within the range, unless otherwise indicated
herein, and each separate value is incorporated into the
specification as if it were individually recited herein. The
endpoints of all ranges are included within the range and
independently combinable. All methods described herein can be
performed in a suitable order unless otherwise indicated herein or
otherwise clearly contradicted by context. The use of any and all
examples, or exemplary language (e.g., "such as"), is intended
merely to better illustrate the invention and does not pose a
limitation on the scope of the invention unless otherwise claimed.
No language in the specification should be construed as indicating
any non-claimed element as essential to the practice of the
invention as used herein. Unless defined otherwise, technical and
scientific terms used herein have the same meaning as is commonly
understood by one of skill in the art to which this invention
belongs.
[0064] An "active agent" means a compound (including metaxalone),
element, or mixture that when administered to a patient, alone or
in combination with another compound, element, or mixture, confers,
directly or indirectly, a physiological effect on the patient. The
indirect physiological effect may occur via a metabolite or other
indirect mechanism. When the active agent is a compound, then
salts, solvates (including hydrates) of the free compound or salt,
crystalline forms, non-crystalline forms, and any polymorphs of the
compound are included. Compounds may contain one or more asymmetric
elements such as stereogenic centers, stereogenic axes and the
like, e.g., asymmetric carbon atoms, so that the compounds can
exist in different stereoisomeric forms. These compounds can be,
for example, racemates or optically active forms. For compounds
with two or more asymmetric elements, these compounds can
additionally be mixtures of diastereomers. For compounds having
asymmetric centers, all optical isomers in pure form and mixtures
thereof are encompassed. In addition, compounds with carbon-carbon
double bonds may occur in Z- and E-forms, with all isomeric forms
of the compounds. In these situations, the single enantiomers,
i.e., optically active forms can be obtained by asymmetric
synthesis, synthesis from optically pure precursors, or by
resolution of the racemates. Resolution of the racemates can also
be accomplished, for example, by conventional methods such as
crystallization in the presence of a resolving agent, or
chromatography, using, for example a chiral HPLC column. All forms
are contemplated herein regardless of the methods used to obtain
them.
[0065] All forms (for example solvates, optical isomers,
enantiomeric forms, polymorphs, free compound and salts of an
active agent) of metaxalone or other active agent may be employed
either alone or in combination.
[0066] "Active agent interaction" refers to a change in the
metabolism of an active agent in a patient that can occur with
co-administration of a second active agent. A "potential active
agent interaction" refers to an active agent interaction between
two active agents that is theoretically possible based on knowledge
that one of the active agents is metabolized by a given cytochrome
p450 isozyme and that the second of the active agents is a
substrate, inhibitor, or inducer of that cytochrome p450
isozyme.
[0067] "Administering metaxalone with a substance", "administering
metaxalone and a substance", or "co-administering metaxalone and a
substance" means metaxalone and the substance are administered
simultaneously in a single dosage form, administered concomitantly
in separate dosage forms, or administered in separate dosage forms
separated by some amount of time that is within the time in which
both metaxalone and the substance are within the blood stream of a
patient. The metaxalone and the substance need not be prescribed
for a patient by the same medical care worker. The substance need
not require a prescription. Administration of metaxalone or the
substance can occur via any appropriate route, for example, oral
tablets, oral capsules, oral liquids, inhalation, injection,
suppositories or topical contact.
[0068] "Adverse event" means any untoward medical occurrence in a
patient administered an active agent and which does not necessarily
have to have a causal relationship with this treatment. An adverse
event (AE) can therefore be any unfavorable and unintended sign
(including an abnormal laboratory finding, for example), symptom,
or disease temporally associated with the use of the active agent,
whether or not considered related to the active agent.
[0069] "Adverse reaction" means a response to an active agent which
is noxious and unintended and which occurs at doses normally used
in humans for prophylaxis, diagnosis, or therapy of disease or for
modification of physiological function. The unintended response can
be an unexpected diminished or enhanced pharmacologic activity or
toxicity of the active agent, e.g., a carisoprodol-associated
toxicity. An adverse reaction also includes any undesirable or
unexpected event requiring discontinuation of the active agent,
modification of the dose, prolonged hospitalization, or the
administration of supportive treatment. "Affects" includes an
increase or decrease in degree, level, or intensity; a change in
time of onset or duration; a change in type, kind, or effect, or a
combination comprising at least one of the foregoing.
[0070] As used herein, "allelic variant" means one of the
alternative forms at a genetic locus on a single chromosome. For
loci in most of the human genome, a human has two chromosomes,
which may carry the same or two different allelic variants.
[0071] "Adjusting administration of an active agent", "altering
administration of an active agent", "adjusting dosing of an active
agent", or "altering dosing of an active agent" are all equivalent
and mean making no change in the dose or dosing regimen of the
active agent; tapering off, reducing, or increasing the dose or the
dosing interval of the active agent; ceasing to administer the
active agent to the patient; or substituting a different active
agent for the active agent.
[0072] "Dosing regimen" means the dose of an active agent taken at
a first time by a patient and the interval (time or symptomatic) at
which any subsequent doses of the active agent are taken by the
patient. The additional doses of the active agent can be different
from the dose taken at the first time. For metaxalone, the standard
dosing regimen for adults and children over 12 years of age is 800
mg, three or four times daily.
[0073] A "dose" means the measured quantity of an active agent to
be taken at one time by a patient.
[0074] "Bioavailability" means the extent or rate at which an
active agent is absorbed into a living system or is made available
at the site of physiological activity. For active agents that are
intended to be absorbed into the bloodstream, bioavailability data
for a given formulation may provide an estimate of the relative
fraction of the administered dose that is absorbed into the
systemic circulation. "Bioavailability" can be characterized by one
or more pharmacokinetic parameters.
[0075] A "dosage form" means a unit of administration of an active
agent. Examples of dosage forms include tablets, capsules,
injections, suspensions, liquids, emulsions, creams, ointments,
suppositories, inhalable forms, transdermal forms, and the
like.
[0076] The term "effective amount" or "therapeutically effective
amount" means an amount effective, when administered to a patient,
to provide any therapeutic benefit. A therapeutic benefit may be an
amelioration of symptoms, e.g., an amount effective to decrease the
symptoms of an acute musculoskeletal condition, such as muscle
spasms. The amount that is "effective" will vary from subject to
subject, depending on the age and general condition of the
individual, the particular active agent, and the like. Thus, it is
not always possible to specify an exact "effective amount."
However, an appropriate "effective" amount in any individual case
may be determined by one of ordinary skill in the art using routine
experimentation. In certain circumstances a patient may not present
symptoms of a condition for which the patient is being treated. A
therapeutically effective amount of an active agent may also be an
amount sufficient to provide a significant positive effect on any
indicium of a disease, disorder, or condition, e.g. an amount
sufficient to significantly reduce the frequency and severity of
muscle spasms. A significant effect on an indicium of a disease,
disorder, or condition is statistically significant in a standard
parametric test of statistical significance, for example Student's
T-test, where p.gtoreq.0.05. An "effective amount or
"therapeutically effective amount" of metaxalone may also be an
amount of about 3600 mg per day or less, about 3200 mg per day or
less, about 50 mg to about 3600 mg per day, or of any dosage amount
approved by a governmental authority such as the US FDA, for use in
treatment. In some embodiments amounts of 3200 mg metaxalone per
day, 800 mg metaxalone per unit dosage form, or 400 mg metaxalone
or less per unit dosage form is an "effective amount" or
"therapeutically effective amount" of metaxalone.
[0077] "Efficacy" means the ability of an active agent administered
to a patient to produce a therapeutic effect in the patient.
[0078] As used herein "food" means a solid food with sufficient
bulk and fat content that it is not rapidly dissolved and absorbed
in the stomach. More specifically, the food is a meal, such as
breakfast, lunch, or dinner. A dosage of metaxalone administered to
a patient "with food" or in a "fed" state is administered to the
patient between about 30 minutes prior to about 2 hours after
eating a meal; more specifically, the dosage is administered within
15 minutes of eating a meal. The terms "without food" or "fasted"
are defined to mean the condition of not having consumed solid food
for about one hour prior to until about 2 hours after such
consumption.
[0079] "Head pain" includes any painful conditions of the head, but
particularly includes headaches, such as migraines, cluster
headaches, tension headaches, or tension related migraines. Head
pain further includes painful facial conditions such as TMJ
(temporomandibular joint) disorders.
[0080] "Informing" means referring to or providing, published
material, for example, providing an active agent with published
material to a user; or presenting information orally, for example,
by presentation at a seminar, conference, or other educational
presentation, by conversation between a pharmaceutical sales
representative and a medical care worker, or by conversation
between a medical care worker and a patient; or demonstrating the
intended information to a user for the purpose of
comprehension.
[0081] "Labeling" means all labels or other means of written,
printed, graphic, electronic, verbal, or demonstrative
communication that is upon a pharmaceutical product or a dosage
form or that accompanies such pharmaceutical product or dosage
form.
[0082] As used herein, an enzyme "metabolizing" a substance means
the enzyme can chemically transform the substance.
[0083] A "medical care worker" means a worker in the health care
field who may need or utilize information regarding an active
agent, including a dosage form thereof, including information on
safety, efficacy, dosing, administration, or pharmacokinetics.
Examples of medical workers include physicians, pharmacists,
physician's assistants, nurses, aides, caretakers (which can
include family members or guardians), emergency medical workers,
and veterinarians.
[0084] As used herein, "metaxalone therapy" refers to medical
treatment of a symptom, disorder, or condition by administration of
metaxalone.
[0085] The term "musculoskeletal condition" includes any condition
affecting the muscles, tendons, ligaments, bones, joints, and
associated tissues that move the body and maintain its form. Such
conditions include conditions that originate in the muscles,
tendons, ligaments, or bones and associated tissues or conditions
that originate elsewhere in the body, for example in the central or
peripheral nervous system, that are manifested in the muscles,
tendons, ligaments, bones, joints or associated tissues.
[0086] A substance having a "narrow therapeutic index" (NTI) means
a substance falling within any definition of narrow therapeutic
index as promulgated by the U.S. Food and Drug Administration or
any successor agency thereof, for example, a substance having a
less than 2-fold difference in median lethal dose (LD50) and median
effective dose (ED50) values, or having a less than 2-fold
difference in the minimum toxic concentration and minimum effective
concentration in the blood; and for which safe and effective use of
the substance requires careful titration and patient
monitoring.
[0087] "Oral dosage form" includes a dosage form for oral
administration.
[0088] A "patient" means a human or non-human animal in need of
medical treatment. Medical treatment can include treatment of an
existing condition, such as a disease or disorder, prophylactic or
preventative treatment, or diagnostic treatment. In some
embodiments the patient is a human patient.
[0089] A "pharmaceutical supplier" means a person (other than a
medical care worker), business, charitable organization,
governmental organization, or other entity involved in the transfer
of active agent, including a dosage form thereof, between entities,
for profit or not. Examples of pharmaceutical suppliers include
pharmaceutical distributors, pharmaceutical wholesalers,
pharmaceutical benefits managers, pharmacy chains, pharmacies
(online or physical), hospitals, HMOs, supermarkets, the Veterans
Administration, or foreign businesses or individuals importing
active agent into the United States.
[0090] "Pharmacokinetic parameters" describe the in vivo
characteristics of an active agent (or surrogate marker for the
active agent) over time, such as plasma concentration (C),
C.sub.max, C.sub.n, C.sub.24, T.sub.max, and AUC. "C.sub.max" is
the measured concentration of the active agent in the plasma at the
point of maximum concentration. "C.sub.n" is the measured
concentration of an active agent in the plasma at about n hours
after administration. "C.sub.24" is the measured concentration of
an active agent in the plasma at about 24 hours after
administration. The term "T.sub.max" refers to the time at which
the measured concentration of an active agent in the plasma is the
highest after administration of the active agent. "AUC" is the area
under the curve of a graph of the measured concentration of an
active agent (typically plasma concentration) vs. time, measured
from one time point to another time point. For example AUC.sub.0-t
is the area under the curve of plasma concentration versus time
from time 0 to time t. The AUC.sub.0-.infin. or AUC.sub.0-INF is
the calculated area under the curve of plasma concentration versus
time from time 0 to time infinity.
[0091] "Phenotype" means an observable trait of an organism
resulting from the interplay of environment and genetics. Examples
include apparent rate of metabolism of substrates by a cytochrome
p450 isozyme of an organism, such as the "poor metabolizer" (PM) or
"ultrarapid metabolizer" (UM) phenotypes identified in humans for
metabolism of substrates metabolized by CYP2D6.
[0092] "Polymorphism" means the differences in DNA sequences that
occur naturally in a population. Single nucleotide substitutions,
insertions, and deletions of nucleotides and repetitive sequences
(microsatellites) are all examples of a polymorphism.
[0093] A "product" or "pharmaceutical product" means a dosage form
of an active agent plus published material, and optionally
packaging.
[0094] "Providing" means giving, administering, selling,
distributing, transferring (for profit or not), manufacturing,
compounding, or dispensing.
[0095] "Published material" means a medium providing information,
including printed, audio, visual, or electronic medium, for example
a flyer, an advertisement, a product insert, printed labeling, an
internet web site, an internet web page, an internet pop-up window,
a radio or television broadcast, a compact disk, a DVD, an audio
recording, or other recording or electronic medium.
[0096] "Product insert" means the professional labeling
(prescribing information) for a pharmaceutical product, a patient
package insert for the pharmaceutical product, or a medication
guide for the pharmaceutical product.
[0097] "Professional labeling" or "prescribing information" means
the official description of a pharmaceutical product approved by a
regulatory agency (e.g., FDA or EMEA) regulating marketing of the
pharmaceutical product, which includes a summary of the essential
scientific information needed for the safe and effective use of the
drug, such as, for example indication and usage; dosage and
administration; who should take it; adverse events (side effects);
instructions for use in special populations (pregnant women,
children, geriatric, etc.); safety information for the patient, and
the like.
[0098] "Patient package insert" means information for patients on
how to safely use a pharmaceutical product that is part of the
FDA-approved labeling. It is an extension of the professional
labeling for a pharmaceutical product that may be distributed to a
patient when the product is dispensed which provides
consumer-oriented information about the product in lay language,
for example it may describe benefits, risks, how to recognize
risks, dosage, or administration.
[0099] "Medication Guide" means an FDA-approved patient labeling
for a pharmaceutical product conforming to the specifications set
forth in 21 CFR 208 and other applicable regulations which contains
information for patients on how to safely use a pharmaceutical
product. A medication guide is scientifically accurate and is based
on, and does not conflict with, the approved professional labeling
for the pharmaceutical product under 21 CFR 201.57, but the
language need not be identical to the sections of approved labeling
to which it corresponds. A medication guide is typically available
for a pharmaceutical product with special risk management
information.
[0100] "Risk" means the probability or chance of adverse reaction,
injury, or other undesirable outcome arising from a medical
treatment. An "acceptable risk" of an adverse reaction means that
individuals or groups in society are willing to take or be
subjected to the risk that the adverse reaction might occur since
the adverse reaction is one whose probability of occurrence is
small, or whose consequences are so slight, or the benefits
(perceived or real) of the active agent are so great. An
"unacceptable risk" of an adverse reaction means that individuals
or groups in society are unwilling to take or be subjected to the
risk that the adverse reaction might occur upon weighing the
probability of occurrence of the adverse reaction, the consequences
of the adverse reaction, and the benefits (perceived or real) of
the active agent. "At risk" means in a state or condition marked by
a high level of risk or susceptibility.
[0101] Herein, an "acceptable risk" means a measure of the risk of
harm, injury, or disease arising from a medical treatment that will
be tolerated by a person or group. Whether a risk is "acceptable"
will depend upon the advantages that the person or group perceives
to be obtainable in return for taking the risk, whether they accept
whatever scientific and other advice is offered about the magnitude
of the risk, and numerous other factors, both political and
social.
[0102] Risk assessment consists of identifying and characterizing
the nature, frequency, and severity of the risks associated with
the use of a product.
[0103] "Safety" means the incidence or severity of adverse events
associated with administration of an active agent, including
adverse effects associated with patient-related factors (e.g., age,
gender, ethnicity, race, target illness, abnormalities of renal or
hepatic function, co-morbid illnesses, genetic characteristics such
as metabolic status, or environment) and active agent-related
factors (e.g., dose, plasma level, duration of exposure, or
concomitant medication).
[0104] "Salts" as used herein describes "pharmaceutically
acceptable salts" of metaxalone and other active agents discussed
herein and also includes solvates and hydrates of such active
agents. The active agent may be modified by making non-toxic acid
or base addition salts thereof. Examples of pharmaceutically
acceptable salts include mineral or organic acid addition salts of
basic residues such as amines; alkali or organic addition salts of
acidic residues; and the like, and combinations comprising one or
more of the foregoing salts. The pharmaceutically acceptable salts
include non-toxic salts and the quaternary ammonium salts of the
metaxalone. For example, non-toxic acid salts include those derived
from inorganic acids such as hydrochloric, hydrobromic, sulfuric,
sulfamic, phosphoric, nitric and the like; other acceptable
inorganic salts include metal salts such as sodium salt, potassium
salt, cesium salt, and the like; and alkaline earth metal salts,
such as calcium salt, magnesium salt, and the like, and
combinations comprising one or more of the foregoing salts.
Pharmaceutically acceptable organic salts includes salts prepared
from organic acids such as acetic, propionic, succinic, glycolic,
stearic, lactic, malic, tartaric, citric, ascorbic, pamoic, maleic,
hydroxymaleic, phenylacetic, glutamic, benzoic, salicylic, mesylic,
esylic, besylic, sulfanilic, 2-acetoxybenzoic, fumaric,
toluenesulfonic, methanesulfonic, ethane disulfonic, oxalic,
isethionic, HOOC--(CH2)n-COOH where n is 0-4, and the like; organic
amine salts such as triethylamine salt, pyridine salt, picoline
salt, ethanolamine salt, triethanolamine salt, dicyclohexylamine
salt, N,N'-dibenzylethylenediamine salt, and the like; and amino
acid salts such as arginate, asparginate, glutamate, and the like;
and combinations comprising one or more of the foregoing salts.
[0105] A "sensitive plasma concentration profile active agent"
means an active agent for which a moderate change in plasma
concentration can have a deleterious effect on the prescribed
therapeutic intent.
[0106] "Side effect" means a secondary effect resulting from taking
an active agent. The secondary effect can be a negative
(unfavorable) effect or a positive (favorable) effect.
[0107] Solid dosage forms of metaxalone comprise up to about 3600
mg metaxalone, specifically about 50 to about 3200 mg metaxalone,
more specifically about 100 to about 800 mg metaxalone. In one
embodiment, the solid dosage form is an oral dosage form, for
example, a tablet. In some embodiments, the tablets comprise about
400 mg, about 450 mg, or about 800 mg metaxalone. Amounts in dosage
forms are given for metaxalone free base, however equivalent
amounts of other forms of metaxalone can be used.
[0108] A "substance" taken or administered with metaxalone means a
substance that affects the safety, bioavailability, plasma
concentration, efficacy, or a combination comprising at least one
of the foregoing of metaxalone or the substance. A "substance" can
be an active agent, an herbal supplement, a nutritional supplement,
a vitamin, a xenobiotic, or an environmental contaminant.
[0109] A substance is a "substrate" of enzyme activity when it can
be chemically transformed by action of the enzyme on the substance.
"Enzyme activity" refers broadly to the specific activity of the
enzyme (i.e., the rate at which the enzyme transforms a substrate
per mg or mole of enzyme) as well as the metabolic effect of such
transformations. Thus, a substance is an "inhibitor" of enzyme
activity when the specific activity or the metabolic effect of the
specific activity of the enzyme can be decreased by the presence of
the substance, without reference to the precise mechanism of such
decrease. For example a substance can be an inhibitor of enzyme
activity by competitive, non-competitive, allosteric or other type
of enzyme inhibition, by decreasing expression of the enzyme, or
other direct or indirect mechanisms. Similarly, a substance is an
"inducer" of enzyme activity when the specific activity or the
metabolic effect of the specific activity of the enzyme can be
increased by the presence of the substance, without reference to
the precise mechanism of such increase. For example a substance can
be an inducer of enzyme activity by increasing reaction rate, by
increasing expression of the enzyme, by allosteric activation or
other direct or indirect mechanisms. It is possible for a substance
to be a substrate, inhibitor, or inducer of an enzyme activity. For
example, the substance can be an inhibitor of enzyme activity by
one mechanism and an inducer of enzyme activity by another
mechanism. The function (substrate, inhibitor, or inducer) of the
substance with respect to activity of an enzyme can depend on
environmental conditions.
[0110] "Subtherapeutic outcome" means a response to an active agent
that is less than that anticipated from a dosing regimen of the
active agent used for treatment of disease or for modification of
physiological function.
[0111] The terms "treating" and "treatment" mean implementation of
therapy with the intention of reducing in severity or frequency
symptoms, elimination of symptoms or underlying cause, prevention
of the occurrence of symptoms or their underlying cause, and
improvement or remediation of damage.
[0112] A "user" means a patient, a medical care worker, or a
pharmaceutical supplier.
[0113] The cytochrome p450 enzymes are a highly diverse superfamily
of enzymes. Each cytochrome p450 enzyme is termed an "isoform" or
"isozyme" since each derives from a different gene. Cytochrome p450
enzymes are categorized into families and subfamilies by amino acid
sequence similarities. These enzymes are designated by the letters
"CYP" followed by an Arabic numeral representing the family, a
letter representing the sub-family and another Arabic numeral
representing a specific gene (e.g., CYP2D6). Particular isozymes
discussed herein are named as per the recommendations of the P450
Gene Superfamily Nomenclature Committee (see e.g., "P450
superfamily: Update on new sequences, gene mapping, accession
numbers, and nomenclature" Pharmacogenetics 6, 1-42 1996, part A
pp. 1-21.). Herein, the designation for a cytochrome p450 isozyme
may encompass the homolog from any species identified as having
such an isozyme. For example, CYP1A2 genes are known in at least
rat, human, rabbit, hamster, dog, guinea pig, mouse and chicken and
the designation "CYP1A2" includes the CYP1A2 protein from each
species known to have a CYP1A2 gene. In some embodiments, the
designation for a cytochrome p450 isozyme is the human isozyme.
[0114] In one embodiment, CYP1A2 is human CYP1A2 (Entrez Gene ID:
1544; reference protein sequence Genbank NP.sub.--000752), and
includes any allelic variants. Specifically, CYP1A2 includes any
allelic variants included in the list of human CYP1A2 allelic
variants maintained by the Human Cytochrome P450 (CYP) Allele
Nomenclature Committee; more specifically it includes any of the *1
through *16 alleles. Additional reference amino acid sequences for
human CYP1A2 include Genbank AAK25728, AAY26399, AAA35738,
AAA52163, AAA52163, AAF13599, AAH67424, AAH67425, AAH67426,
AAH67427, AAH67428, AAH67429, AAA52154, AAA52146, CAA77335, P05177,
Q6NWU3, Q6NWU5, Q9BXX7, and Q9UK49.
[0115] In one embodiment, CYP2A6 is human CYP2A6 (Entrez Gene ID:
1548; reference protein sequence Genbank NP.sub.--000753), and
includes any CYP2A6 allelic variants. Specifically, CYP2A6 includes
any allelic variants included in the list of human CYP2A6 allelic
variants maintained by the Human Cytochrome P450 (CYP) Allele
Nomenclature Committee; more specifically it includes any of the *1
through *22 alleles. Additional reference amino acid sequences for
human CYP2A6 include Genbank AAG45229, AAB40518, AAF13600,
AAH96253, AAH96254, AAH96255, AAH96256, AAA52067, CAA32097,
CAA32117, P11509, Q13120, and Q4VAU0.
[0116] In one embodiment, CYP2B6 is human CYP2B6 (Entrez Gene ID:
1555; reference protein sequence Genbank NP.sub.--000758), and
includes any CYP2B6 allelic variants. Specifically, CYP2B6 includes
any allelic variants included in the list of human CYP2B6 allelic
variants maintained by the Human Cytochrome P450 (CYP) Allele
Nomenclature Committee; more specifically it includes any of the *1
through *25 alleles. Additional reference amino acid sequences for
human CYP2B6 include Genbank AAF32444, AAD25924, ABB84469,
AAF13602, AAH67430, AAH67431, AAA52144, P20813, Q6NWU1, Q6NWU2, and
Q9UNX8.
[0117] In one embodiment, CYP2C8 is human CYP2C8 (Entrez Gene ID:
1558; reference protein sequence Genbank NP.sub.--110518), and
includes any CYP2C8 allelic variants. Specifically, CYP2B8 includes
any allelic variants included in the list of human CYP2C8 allelic
variants maintained by the Human Cytochrome P450 (CYP) Allele
Nomenclature Committee; more specifically it includes any of the *1
through *10 alleles. Additional reference amino acid sequences for
human CYP2C8 include Genbank CAH71307, AAR89907, CAA38578,
AAH20596, AAA35739, AAA35740, AAA52160, AAA52161, CAA35915,
CAA68550, P10632, Q5VX93, Q8WWB1, and Q9UCZ9.
[0118] In one embodiment, CYP2C9 is human CYP2C9 (Entrez Gene ID:
1559; reference protein sequence Genbank NP.sub.--000762), and
includes any CYP2C9 allelic variants. Specifically, CYP2CP includes
any allelic variants included in the list of human CYP2C9 allelic
variants maintained by the Human Cytochrome P450 (CYP) Allele
Nomenclature Committee; more specifically it includes any of the *1
through *24 alleles. Additional reference amino acid sequences for
human CYP2C9 include Genbank CAH71303, AAP88931, AAT94065,
AAW83816, AAD13466, AAD13467, AAH20754, AAH70317, BAA00123,
AAA52159, AAB23864, P11712, Q5EDC5, Q5VX92, Q6IRV8, Q8WW80, Q9UEH3,
and Q9UQ59.
[0119] In one embodiment, CYP2C19 is human CYP2C19 (Entrez Gene ID:
1557; reference protein sequence Genbank NP.sub.--000760), and
includes any CYP2C19 allelic variants. Specifically, CYP2C19
includes any allelic variants included in the list of human CYP2C19
allelic variants maintained by the Human Cytochrome P450 (CYP)
Allele Nomenclature Committee; more specifically it includes any of
the *1 through *21 alleles. Additional reference amino acid
sequences for human CYP2C19 include Genbank BAD02827, CAH73444,
CAH74068, AAV41877, AAL31347, AAL31348, AAA36660, AAB59426,
CAA46778, P33261, Q16743, Q767A3, Q8WZB1, and Q8WZB2.
[0120] In one embodiment, CYP2D6 is human CYP2D6 (Entrez Gene ID:
1565; reference protein sequence Genbank NP.sub.--000097), and
includes any CYP2D6 allelic variants. Specifically, it CYP2D6
includes any allelic variants included in the list of human CYP2D6
allelic variants maintained by the Human Cytochrome P450 (CYP)
Allele Nomenclature Committee; more specifically it includes any of
the *1 through *58 alleles. Additional reference amino acid
sequences for human CYP2D6 include Genbank AAS55001, ABB01370,
ABB01371, ABB01372, ABB01373, AAA35737, AAA53500, BAD92729,
AAU87043, AAH66877, AAH67432, AAH75023, AAH75024, AAI06758,
AAI06759, CAG30316, AAA52153, AAA36403, CAA30807, and P10635.
[0121] In one embodiment, CYP2E1 is human CYP2E1 (Entrez Gene ID:
1571; reference protein sequence Genbank NP.sub.--000764), and
includes any CYP2E1 allelic variants. Specifically, CYP2E1 includes
any allelic variants included in the list of human CYP2E1 allelic
variants maintained by the Human Cytochrome P450 (CYP) Allele
Nomenclature Committee; more specifically it includes any of the *1
through *7 alleles. Additional reference amino acid sequences for
human CYP2E1 include Genbank CAH70047, BAA00902, BAA08796,
AAA52155, AAD13753, AAF13601, CA147002, AAH67433, AAH67435,
AAZ77710, AAA35743, AAD14267, PO.sub.5181, Q16868, Q5VZD5, Q6LER5,
Q6NWT7, and Q6NWT9.
[0122] In one embodiment, CYP3A4 is human CYP3A4 (Entrez Gene ID:
1576; reference protein sequence Genbank NP.sub.--059488), and
includes any CYP3A4 allelic variants. Specifically, CYP3A4 includes
any allelic variants included in the list of human CYP3A4 allelic
variants maintained by the Human Cytochrome P450 (CYP) Allele
Nomenclature Committee; more specifically it includes any of the *1
through *20 alleles. Additional reference amino acid sequences for
human CYP3A4 include Genbank AAF21034, AAG32290, AAG53948,
EAL23866, AAF13598, CAD91343, CAD91645, CAD91345, AAH69418,
AAI01632, BAA00001, AAA35747, AAA35742, AAA35744, AAA35745,
CAA30944, P05184, P08684, Q6GRK0, Q7Z448, Q86SK2, Q86SK3, and
Q9BZM0.
[0123] The ability of metaxalone to act as a substrate, inhibitor,
or inducer of various cytochrome p450 isozymes was determined in
studies described below. A summary of the findings of the studies
is provided in Table 1.
TABLE-US-00001 TABLE 1 Summary of metaxalone effects on cytochrome
p450 isozymes. CYP isozyme Substrate Inhibitor Inducer/Inhibitor
1A2 + + + 2A6 0 0 0 2B6 0 + 0 2C8 + 0 ND 2C9 + 0 - 2C19 + + 0 2D6 +
+ - 2E1 + + 0 3A4 + + +
[0124] For each possible function of metaxalone (i.e., substrate,
inhibitor, or inducer), there is a column in the table. A "+" in a
particular column and row indicates that the study found that
metaxalone functioned in that capacity with respect to the
cytochrome p450 isozyme represented in that row, while a "0"
indicates that the results did not support that metaxalone
functioned in that capacity with respect to the cytochrome p450
isozyme represented in that row. In the column labeled
Inducer/Inhibitor, a "+" denotes that the metaxalone functioned as
an inducer of the CYP isozyme, while a "-" denotes that metaxalone
functioned as an inhibitor of the CYP isozyme. For example,
metaxalone was found to be a substrate, inhibitor, and inducer of
CYP1A2 activity, and was found to be an inhibitor of CYP2C9
activity. The symbol "ND" indicates that no experiment was
performed.
[0125] As summarized in Table 1, metaxalone was found to be a
substrate for CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, and
CYP3A4, and therefore can also act as a competitor of other
substrates for these isozymes. Additionally, metaxalone was
determined to be an inhibitor of the cytochrome p450 isozymes
CYP1A2, CYP2B6, CYP2C9, CYP2C19, CYP2D6, CYP2E1, and CYP3A4 and an
inducer of CYP1A2 and CYP3A4.
[0126] Enzymes involved in Phase I and Phase II active agent
metabolism, such as the cytochrome p450 isozymes, respond to the
constantly changing types and amounts of substrate active agents
they encounter. For example, changes in active agent metabolism due
to competition for the same cytochrome p450 isoform can change the
clinical effectiveness or safety of an active agent by altering the
plasma concentration of the active agent or its metabolite(s).
Similarly, inhibition or induction of the cytochrome p450 isoform
that metabolizes a particular active agent can change the clinical
effectiveness or safety of that active agent. Therefore, for any
cytochrome p450 for which metaxalone acts as a substrate,
inhibitor, or inducer, the administration of metaxalone with a
substance that is a substrate, inhibitor, or inducer of that
cytochrome p450 can affect the metabolism of the metaxalone or the
substance. For the case in which the substance is a narrow
therapeutic index active agent, such as warfarin or phenyloin, too
little of the active agent in the blood stream can lead to
insufficient therapeutic activity, while a too large dose of the
active agent can lead to excessive therapeutic activity or
toxicity, both of which can be detrimental.
[0127] The invention provides methods of using metaxalone. These
methods include using metaxalone in the treatment of various
diseases or conditions, including, for example, musculoskeletal
conditions, specifically acute and painful musculoskeletal
conditions, muscle sprains, muscle spasms, spasticity, low back
pain and stiffness, acute lumbosacral pain, cervical stiffness or
torticohis; as well as head pain, including migraines, cluster
headaches, tension headaches, or tension related migraines. Using
metaxalone in the treatment or prevention of a disease or condition
in a patient can include administering metaxalone to a patient,
dispensing metaxalone to a patient, or dispensing metaxalone to a
medical care worker for administering to a patient.
[0128] In one embodiment, the method comprises informing a user
that metaxalone affects activity of a cytochrome p450 isozyme. The
cytochrome p450 isozyme may be any cytochrome p450 isozyme. For
example the cytochrome p450 isozyme may be CYP1A2 CYP2B6, CYP2C9,
CYP2C19, CYP2D6, CYP2E1, or CYP3A4. In some embodiments the
cytochrome p450 isozyme is CYP1A2, CYP3A4, or CYP2C19. In certain
embodiments the cytochrome p450 isozyme is a human enzyme. In some
embodiments, the method further comprises providing the user with
metaxalone. If the user is a patient, the method can further
comprise dosing the patient to improve safety or efficacy of
metaxalone such that the dosing minimizes adverse events or side
effects of metaxalone, specifically when metaxalone is
co-administered to the patient with another substance, such as
another active agent.
[0129] Informing the user that metaxalone affects the activity of a
cytochrome p450 isozyme includes providing a user with information
about any effect of metaxalone on the activity of any cytochrome
p450 isozyme. Informing the user that metaxalone affects the
activity of a cytochrome p450 isozyme includes informing a user of
any of the following: that metaxalone is metabolized by a
cytochrome p450 isozyme; that metaxalone is an inducer of activity
of a cytochrome p450 isozyme; that a cytochrome p450 isozyme
metabolizing metaxalone is CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6,
CYP2E1, or CYP3A4; that metaxalone is metabolized by CYP1A2,
CYP2E1, or CYP3A4; that metaxalone is a competitive inhibitor of
CYP1A2, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or CYP3A4; that metaxalone
is a substrate of CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1,
and CYP3A4; that there is a potential active agent interaction
between metaxalone and an active agent that is a substrate,
inhibitor, or inducer of CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6,
CYP2E1, and CYP3A4; that metaxalone is an inhibitor of a cytochrome
p450 isozyme; that caution is recommended when metaxalone and a
substrate of CYP2B6, CYP2C9, CYP2C19, or CYP2D6 are administered to
a patient known to have a poor metabolizer phenotype for or that
has reduced activity of CYP2B6, CYP2C9, CYP2C19, or CYP2D6; that
caution is recommended when administering metaxalone with the
substance when the substance is an active agent having a narrow
therapeutic index; that the allelic variants of CYP2B6, CYP2C9,
CYP2C19, or CYP2D6 present in the patient can further affect the
potential active agent interaction between metaxalone and an active
agent; that there is a potential active agent interaction of
metaxalone with an active agent that is a substrate of the
cytochrome p450 isozyme; that there is a potential active agent
interaction of metaxalone with warfarin; that metaxalone affects
the activity of CYP1A2, CYP2B6, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or
CYP3A4; that there is a potential active agent interaction of
metaxalone with a substance that is a substrate of CYP1A2, CYP2B6,
CYP2C9, CYP2C19, CYP2D6, CYP2E1, or CYP3A4; that metaxalone is an
inhibitor of CYP1A2, CYP2B6, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or
CYP3A4; and that metaxalone is an inducer of CYP1A2 or CYP3A4
activity; that there is a potential active agent interaction of
metaxalone with a substance that is a substrate of CYP1A2 or
CYP3A4.
[0130] The method can further comprise informing the user that
administration of metaxalone with a substance can affect the plasma
concentration, bioavailability, safety, efficacy, or a combination
comprising at least one of the foregoing of metaxalone or the
substance. In some embodiments, the method further comprises
providing the user with the substance.
[0131] The effect of coadministration of metaxalone and the
substance can be determined by comparison of the plasma
concentration, bioavailability, safety, efficacy, or a combination
comprising at least one of the foregoing of the substance with and
without coadministration of metaxalone or by comparison of the
plasma concentration, bioavailability, safety, efficacy, or a
combination comprising at least one of the foregoing of metaxalone
with and without coadministration of the substance.
[0132] Informing the user that administration of metaxalone with a
substance can affect the plasma concentration, bioavailability,
safety, efficacy, or a combination comprising at least one of the
foregoing of metaxalone or the substance includes providing a user
with information about any effect of metaxalone on plasma
concentration, bioavailability, safety, efficacy, or a combination
comprising at least one of the foregoing of metaxalone or the
substance. This includes informing a user of any of the following:
that taking metaxalone with an active agent can affect the
bioavailability, safety, or efficacy of the active agent or
metaxalone; that administration of metaxalone and a substance that
is a substrate, inhibitor, or inducer of CYP1A2, CYP2C8, CYP2C9,
CYP2C19, CYP2D6, CYP2E1, or CYP3A4 can affect plasma concentration,
bioavailability, safety, efficacy, or a combination comprising at
least one of the foregoing of metaxalone or the substance; that
administration of metaxalone with a substance that is a CYP1A2,
CYP2C9, CYP2C19, CYP2D6, CYP2E1, and CYP3A4 substrate can increase
the plasma concentration of the substance; that taking metaxalone
with an active agent that is a substrate, inhibitor, or inducer of
CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or CYP3A4 can
affect the plasma concentration, bioavailability, safety, efficacy,
or a combination comprising at least one of the foregoing of
metaxalone or the active agent; that administration of metaxalone
with an active agent that is a cytochrome p450 isozyme substrate
having a narrow therapeutic index can affect plasma concentration,
bioavailability, safety, efficacy, or a combination comprising at
least one of the foregoing of the active agent; that administration
of metaxalone with an active agent that is a CYP1A2, CYP2C8,
CYP2C9, CYP2C19, CYP2D6, CYP2E1, or CYP3A4 substrate having a
narrow therapeutic index can affect plasma concentration,
bioavailability, safety, efficacy, or a combination comprising at
least one of the foregoing of the active agent; that metaxalone can
affect the plasma concentration, bioavailability, safety, efficacy,
or a combination comprising at least one of the foregoing of an
active agent that is a substrate of the cytochrome p450 isozyme;
that administration of metaxalone with an active agent that is a
substrate of the cytochrome p450 isozyme and that has a narrow
therapeutic index can increase plasma concentration of the active
agent; that a substance that induces CYP1A2, CYP2C8, CYP2C9,
CYP2C19, CYP2D6, CYP2E1, or CYP3A4 activity can decrease metaxalone
plasma concentration; that co-administration of metaxalone and a
substance that inhibits CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6,
CYP2E1, or CYP3A4 may result in an increase in metaxalone plasma
concentration, whereas co-administration of metaxalone and a
substance that induces CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6,
CYP2E1, or CYP3A4 may decrease metaxalone plasma concentration;
that a substance that inhibits CYP1A2, CYP2C8, CYP2C9, CYP2C19,
CYP2D6, CYP2E1, or CYP3A4 activity can increase metaxalone plasma
concentration; that a substance that is a substrate of CYP1A2,
CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or CYP3A4 can increase
plasma concentration of metaxalone or the substance; that
administration of metaxalone with warfarin can affect the plasma
concentration, bioavailability, safety, efficacy, or a combination
comprising at least one of the foregoing of metaxalone or warfarin;
that administration of metaxalone with an active agent that is a
known substrate, inhibitor, or inducer of YP1A2, CYP2C8, CYP2C9,
CYP2C19, CYP2D6, CYP2E1, or CYP3A4 or that is a substrate of
CYP2B6, CYP2C9, CYP2D6, CYP2E1, or CYP3A4 can affect plasma
concentration, bioavailability, safety, efficacy, or a combination
comprising at least one of the foregoing of the active agent or
metaxalone; that the plasma concentration of a substance that is a
substrate of CYP1A2, CYP2B6, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or
CYP3A4 can decrease when the substance is administered with
metaxalone; that administration of metaxalone with a substance that
is a substrate of CYP1A2, CYP2B6, CYP2C9, CYP2C19, CYP2D6, CYP2E1,
or CYP3A4 can affect plasma concentration, bioavailability, safety,
efficacy, or a combination comprising at least one of the foregoing
of the substance; that administration of metaxalone with an active
agent that is a CYP1A2, CYP2B6, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or
CYP3A4 substrate having a narrow therapeutic index can affect
plasma concentration, bioavailability, safety, efficacy, or a
combination comprising at least one of the foregoing of the active
agent; that the plasma concentration of a substance that is a
substrate of CYP1A2 or CYP3A4 can decrease when the substance is
administered with metaxalone; that administration of metaxalone and
a substance that is a substrate of CYP1A2 or CYP3A4 activity can
affect plasma concentration, bioavailability, safety, efficacy, or
a combination comprising at least one of the foregoing of the
substance.
[0133] In another embodiment, the method comprises informing a user
that metaxalone is metabolized by a cytochrome p450 isozyme. The
cytochrome p450 isozyme metabolizing metaxalone is CYP1A2, CYP2C8,
CYP2C9, CYP2C19, CYP2D6, CYP2E1, or CYP3A4; specifically the
cytochrome p450 isozyme metabolizing metaxalone is CYP1A2, CYP2E1,
or CYP3A4. In some embodiments, the method further comprises
informing the user that administration of metaxalone and a
substance that is a substrate, inhibitor, or inducer of CYP1A2,
CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or CYP3A4 can affect
plasma concentration, bioavailability, safety, efficacy, or a
combination comprising at least one of the foregoing of metaxalone
or the substance. Methods provided herein include informing a user
that the substance or metaxalone is a substrate, inhibitor, or
inducer of CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or
CYP3A4. The substance can inhibit CYP1A2, CYP2C8, CYP2C9, CYP2C19,
CYP2D6, CYP2E1, or CYP3A4 activity and the effect can be an
increase in metaxalone plasma concentration, or the substance can
induce CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or CYP3A4
activity and the effect can be a decrease in metaxalone plasma
concentration. In yet another embodiment, the user is informed that
the substance is a substrate of CYP1A2, CYP2C8, CYP2C9, CYP2C19,
CYP2D6, CYP2E1, or CYP3A4 and plasma concentration of the substance
or metaxalone can increase. In yet another embodiment, the method
comprises informing the user that taking metaxalone and a substance
that is a substrate of CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6,
CYP2E1, or CYP3A4 can increase plasma concentration of metaxalone
or the substance.
[0134] The method also comprises informing a user that metaxalone
is an inhibitor or an inducer of a cytochrome p450 isozyme.
Cytochrome p450 isozymes inhibited by metaxalone include CYP1A2,
CYP2B6, CYP2C9, CYP2C19, CYP2D6, CYP2E1, and CYP3A4. Cytochrome
p450 isozymes that are induced by metaxalone include CYP1A2 and
CYP3A4. In some embodiments the method further comprises informing
a user that administration of metaxalone and a substance that is a
substrate of the cytochrome p450 isozyme can affect plasma
concentration, bioavailability, safety, efficacy, or a combination
comprising at least one of the foregoing of the substance. In some
embodiments, the method can further comprise informing that
metaxalone is an inhibitor of the cytochrome p450 isozyme or that
the effect on the substance can be an increase in plasma
concentration. In other embodiments, the method can further
comprise informing that metaxalone is an inducer of CYP1A2 or
CYP3A4 or that the effect on the substance can be a decrease in
plasma concentration.
[0135] In some embodiments, the method can further comprise
providing the user with metaxalone. Other embodiments include
administering metaxalone or another substance. Administration may
be to a patient by the patient, a medical worker, or other user.
Metaxalone can be administered in a therapeutically effective
amount. In some embodiments, the method can further comprise
providing the user with metaxalone or informing the user that
caution is recommended when administering metaxalone with the
substance when the substance is an active agent having a narrow
therapeutic index.
[0136] In an embodiment, the method includes informing a user that
there is a potential active agent interaction between metaxalone
and a substance that is a substrate, an inhibitor, or an inducer of
CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or CYP3A4;
administering metaxalone to a patient, wherein the patient is
receiving a substance that is a substrate, an inhibitor, or an
inducer of CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, and
CYP3A4; and monitoring the patient. Monitoring the patient can be
monitoring the patient's blood plasma level of metaxalone or the
substance; monitoring the patient for a symptom of an active agent
interaction between the substance and metaxalone; monitoring the
patient for an adverse reaction or side effect resulting from
coadministration of the substance and metaxalone; monitoring the
patient for an adverse reaction or side effect associated with
metaxalone; monitoring the patient for a subtherapeutic outcome
associated with a reduced plasma concentration of metaxalone, or
monitoring the patient for an adverse reaction or side effect
associated with an elevated or reduced plasma concentration of
metaxalone. The method can further include adjusting administration
of metaxalone or the substance in response to the monitoring. The
substance can be an active agent.
[0137] In some embodiments, the method of using metaxalone can
further comprise administering metaxalone or a substance.
Administration may be to a patient by the patient, a medical care
worker, or other user. Metaxalone can be administered in a
therapeutically effective amount. The substance can be an active
agent. The active agent can have a sensitive plasma concentration
profile or a narrow therapeutic index. The method can also comprise
monitoring a patient, for example, monitoring the patient for an
adverse reaction, a side effect, a subtherapeutic outcome, or a
symptom of an active agent interaction or monitoring a patient's
plasma concentration of metaxalone or the substance. The method can
also comprise adjusting administration or dosing of the substance
or metaxalone for the patient based on the results of monitoring,
for example a determined plasma concentration of the active agent
or metaxalone.
[0138] In all of the embodiments herein, a medical care worker can
determine the plasma concentration of a substance such as an active
agent, including metaxalone, by performing or ordering the
performance of any suitable method. For example, the medical care
worker could order a test using blood drawn from the patient for
determining the plasma concentration of metaxalone or the
substance.
[0139] The information provided to a user can comprise any
combination of information disclosed herein concerning the effects
of metaxalone on the activity of a cytochrome p450 isozyme or on
the plasma concentration, bioavailability, safety, efficacy, or a
combination comprising at least one of the foregoing of metaxalone
or a substance. The information may also comprise any combination
of information disclosed herein concerning the effects of a
substance on the activity of a cytochrome p450 isozyme or on the
plasma concentration, bioavailability, safety, efficacy, or a
combination comprising at least one of the foregoing of metaxalone
or a substance when the substance is used with metaxalone.
[0140] Medical information provided in any of the methods described
herein concerning the effects of administering metaxalone with an
additional substance may alternatively be provided in layman's
terms, so as to be better understood by patients or non-medical
professionals. Those of skill in the medical art are familiar with
the various layman's terms that can be used to describe the effects
of active agent interactions.
[0141] In yet another embodiment, the method of using metaxalone
comprises obtaining metaxalone from a container providing
information that metaxalone affects activity of a cytochrome p450.
Information can also be provided that administering metaxalone with
a substance can affect plasma concentration, bioavailability,
safety, efficacy, or a combination comprising at least one of the
foregoing of the substance or metaxalone. The method also comprises
providing metaxalone in the container providing such information.
The method may also comprise providing a substance, such as an
active agent, in a container providing information that metaxalone
affects activity of a cytochrome p450 or that administering
metaxalone with the substance may affect plasma concentration,
bioavailability, safety, efficacy, or a combination comprising at
least one of the foregoing of the substance or metaxalone. The
provided information may be any information disclosed herein
concerning the effects of metaxalone or a substance on the activity
of a cytochrome p450 isozyme or any information disclosed herein
concerning the effects of metaxalone when administered with a
substance on the plasma concentration, bioavailability, safety,
efficacy, or a combination comprising at least one of the foregoing
of the substance or metaxalone. The method can further comprise
ingesting the metaxalone or the substance.
[0142] The method may also comprise informing the user or providing
information that, when an active agent and metaxalone are
administered to a patient, that it is recommended that a medical
care worker determine the patient's plasma concentration of the
active agent; and alter dosing of the active agent for the patient
based on the determined active agent plasma concentration.
Additionally, the method can comprise determining the metabolizer
phenotype of the patient or the allelic variant of the patient for
a cytochrome p450 isozyme; specifically the cytochrome p450 isozyme
is CYP2B6, CYP2C9, CYP2C19, or CYP2D6.
[0143] Also disclosed herein are methods of administering
metaxalone.
[0144] In an embodiment, the method comprises determining for a
patient to whom metaxalone is going to be administered or is being
administered whether a substance that is currently being or will be
administered to the patient is an inhibitor of a cytochrome P450
isozyme (CYP), wherein the CYP is CYP1A2, CYP2C8, CYP2C9, CYP2C19,
CYP2D6, CYP2E1, and CYP3A4; and determining risk for the patient of
a metaxalone-associated toxicity resulting from inhibition of
metaxalone metabolism by the CYP during coadministration of
metaxalone and the substance.
[0145] In an embodiment, the method comprises determining for a
patient to whom metaxalone is going to be administered or is being
administered whether a substance that is currently being or will be
administered to the patient is an inducer of a cytochrome P450
isozyme (CYP), wherein the CYP is CYP1A2, CYP2C8, CYP2C9, CYP2C19,
CYP2D6, CYP2E1, and CYP3A4; and determining risk for the patient of
a subtherapeutic outcome for metaxalone resulting from induction of
metaxalone metabolism by the CYP during coadministration of
metaxalone and the substance.
[0146] Depending on the determined risk of a metaxalone-related
toxicity or a subtherapeutic outcome, the methods can further
comprise administering metaxalone or the substance to the patient.
For example, if there is no risk of metaxalone-associated toxicity
or of a subtherapeutic outcome for the metaxalone treatment, or if
the risk is determined to be acceptable, metaxalone and the
substance can be administered to the patient. Alternatively, if
there is a risk of metaxalone-associated toxicity or of a
subtherapeutic outcome for the metaxalone treatment, or if the risk
is determined to be unacceptable, metaxalone can be administered to
the patient, but not the substance.
[0147] The method can further comprise determining that the patient
has a poor metabolizer phenotype for CYP2C9, CYP2C19, or CYP2D6; or
that the patient belongs to an Asiatic or Oceanic ethnic group.
[0148] Determining risk of an adverse reaction, such as a toxicity
or a subtherapeutic outcome, for metaxalone and a substance with
which it may be administered is based on an appropriate set of risk
parameters. As will be evident to those of skill in the art, the
risk parameters to be considered will be based upon factors which
influence the risk that a known or suspected adverse reaction will
occur if the patient receives metaxalone with or without the
substance, and will vary depending upon the substance in question
for coadministration with metaxalone. Factors that may define the
relevant risk parameters include effect of the substance on
activity of the relevant cytochrome P450 isozyme(s), e.g. CY3A4 or
CYP2C19; the likelihood that certain preexisting conditions may
exist in the patient; information collected from the patient
including information relating to the patient's conduct; the
patient's past or ongoing medical treatment, such as other
procedures or medication which the patient may have received or is
still receiving; results of certain diagnostic tests which have
been performed; and the like. Information collected from the
patient for determining risk may be obtained prior to the initial
dispensation of metaxalone or the substance to the patient or may
be obtained from the patient on a periodic basis. For example,
after treatment with metaxalone and the substance is begun,
information on the onset of certain symptoms which may be
indicative of the need for changes in the patient's treatment
regimen may be obtained from the patient on a periodic basis.
[0149] Diagnostic tests may be probative of the concentration of
one or more active agents, including a prescribed active agent, to
assure that appropriate dosing is maintained in the patient. Such
diagnostic testing may be conducted on any bodily fluid or waste
product of the patient, including the blood, serum, plasma, saliva,
semen or urine, as well as the feces. Diagnostic testing may also
be performed on a biopsy of any tissue of the patient or may
include genetic testing, which may be indicative of a genetic
predisposition to a particular adverse side effect. Other forms of
diagnostic testing, such as diagnostic imaging, or tests which may
be probative of the proper functioning of any tissue, organ, or
system are also contemplated. Preferably, appropriate information
or diagnostic test results are obtained and considered in
determining risk.
[0150] Where the relevant risk parameters indicate that the risk of
the adverse reaction occurring outweighs the potential benefit of
metaxalone or the substance, the risk can be deemed
unacceptable.
[0151] Determining risk can comprise accessing a computer-hosted
database. The database may be in the form of a look-up table, or
similar structure, that provides output information based on the
input of information.
[0152] Alternatively, determining risk can comprise obtaining
information from standard treatment guidelines, textbooks,
compendial literature, journals, drug manufacturer guidelines,
internet websites providing information on active agent
interactions (e.g., "Drug Interaction Checker" at the MEDScape
website or the drug interaction website maintained by Dr. D.
Flockhart, Indiana University School of Medicine); or FDA labeling
for particular active agents.
[0153] In an embodiment, the method comprises informing a patient
receiving an active agent or the patient's medical care worker that
metaxalone is a substrate of CYP1A2, CYP2C8, CYP2C9, CYP2C19,
CYP2D6, CYP2E1, or CYP3A4; and adjusting administration of
metaxalone or the active agent to the patient as a result of the
informing to avoid an adverse event associated with metaxalone, to
avoid a subtherapeutic outcome for metaxalone in the patient, or to
produce a treatment response in the patient. The active agent is a
known substrate, inhibitor, or inducer of CYP1A2, CYP2C8, CYP2C9,
CYP2C19, CYP2D6, CYP2E1, or CYP3A4.
[0154] In an embodiment, the method comprises administering
metaxalone to a patient; and monitoring the patient during
administration of metaxalone if the patient is taking a substance
that is a known substrate, inhibitor, or inducer of activity of
CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or CYP3A4.
[0155] In an embodiment, the method comprises determining that a
substance that is a known substrate, inhibitor, or inducer of
activity of CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or
CYP3A4 is administered to the patient; and adjusting administration
of metaxalone or the substance to the patient to avoid a
metaxalone-related adverse reaction or subtherapeutic outcome.
[0156] In an embodiment, the method comprises determining that
metaxalone is a substrate of a cytochrome P450, wherein the CYP is
CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or CYP3A4;
administering metaxalone to a patient; and monitoring the patient
during administration of metaxalone if a substance that is a known
substrate, inhibitor, or inducer of activity of the CYP is
coadministered to the patient. The method can further comprise
determining that a substance that is a known substrate, inhibitor,
or inducer of activity of the CYP is administered to the
patient.
[0157] Such methods can include informing a user that metaxalone is
a substrate of the CYP. The method can include informing the user
that administration of metaxalone with a substance can affect the
plasma concentration, bioavailability, safety, efficacy, or a
combination comprising at least one of the foregoing of metaxalone
or the substance. The method can include informing the user of any
information disclosed herein about metaxalone metabolism and any
information disclosed herein about the effect of metaxalone or the
substance on the plasma concentration, bioavailability, safety,
efficacy, or a combination comprising at least one of the foregoing
of metaxalone or the substance when metaxalone is used with the
substance.
[0158] Determining that a substance that is a known substrate,
inhibitor, or inducer of CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6,
CYP2E1, or CYP3A4 is administered to a patient in need of
metaxalone therapy can be performed by consulting with the patient
regarding substances, e.g., medications, taken in by the patient, a
medical care worker administering medications to the patient, a
prescription database including medications prescribed to the
patient, or by any other method known in the art.
[0159] Determining that metaxalone is a substrate of a cytochrome
p450 isozyme or determining that co-administration of metaxalone
and a substance that inhibits that cytochrome P450 may result in an
increased metaxalone plasma concentration, or that
co-administration of metaxalone and a substance that induces that
cytochrome P450 may result in a decreased metaxalone plasma
concentration can be performed by consulting the package insert for
the metaxalone product administered to the patient or consulting a
database including prescribing information and potential risks for
metaxalone, or by any other method known in the art.
[0160] Monitoring the patient can comprise monitoring the patient's
plasma concentration of metaxalone or the substance; monitoring the
patient for symptoms of an active agent interaction between the
substance and metaxalone; monitoring the patient for an adverse
reaction (e.g., a toxicity or a subtherapeutic outcome) resulting
from administration of the substance and metaxalone; monitoring the
patient for an adverse reaction (e.g., a toxicity or a
subtherapeutic outcome) associated with metaxalone; or monitoring
the patient for decreased efficacy of metaxalone.
[0161] Monitoring the patient can be monitoring any appropriate
patient-specific, disease-specific, or substance-specific parameter
appropriate to avoid or safely manage an active agent interaction.
Monitoring the patient can be, for example, monitoring the patient
for an adverse reaction, a subtherapeutic outcome, a side effect,
or a symptom of an active agent interaction for example by physical
examination or visual identification; monitoring the blood level of
metaxalone or the substance in the patient; monitoring clinical
laboratory tests appropriate for metaxalone, the substance, or a
medical diagnosis for the patient; monitoring therapeutic effect of
metaxalone or the substance on the patient's condition; monitoring
occurrence in the patient of a known side effect or adverse
reaction of metaxalone or the substance; monitoring the patient for
occurrence of an unexpected response during treatment; monitoring
changes in control, signs, or symptoms of a condition of the
patient, or determining a complete list of medical diagnoses for
the patient. Monitoring the patient can be performed by the patient
or by a medical care worker.
[0162] For metaxalone, the most frequently reported adverse
reactions include CNS reactions: drowsiness, dizziness, headache,
and nervousness or "irritability"; and digestive reactions: nausea,
vomiting, and gastrointestinal upset. Other possible adverse
reactions include immune system reactions such as hypersensitivity
reaction and rash with or without pruritus; hematologic reactions
such as leukopenia and hemolytic anemia; and hepatobiliary
reactions such as jaundice. Though rare, anaphylactoid reactions
have been reported with metaxalone. Examples of
metaxalone-associated toxicities include drowsiness, dizziness,
impairment of a mental activity, impairment of a physical activity,
central nervous system depression, respiratory depression, or
coma.
[0163] Determining that a patient experiences an adverse reaction,
such as a metaxalone-associated toxicity or a subtherapeutic
outcome, can be performed by obtaining information from the patient
regarding onset of certain symptoms which may be indicative of the
adverse reaction, results of diagnostic tests indicative of the
adverse reaction, and the like.
[0164] Determining the level of metabolism of metaxalone in a
subject may be performed for example by determining plasma
concentrations of metaxalone or appropriate metaxalone metabolites,
and any other methods known in the art.
[0165] Adjusting administration of metaxalone or the substance to
the patient to avoid an adverse reaction or a subtherapeutic
outcome, or adjusting dosing regimens can be performed by one of
ordinary skill in the art, taking into consideration the physiology
of the patient, including such factors as the age, sex, and health
of the patient, as well as active agents the patient may be taking
at the time. Optionally, the patient can be monitored at the
initial, or a subsequent, stage of treatment to ensure therapeutic
plasma levels of metaxalone or the substance are achieved or
maintained.
[0166] Various laboratory methods are known, including ones that
are commercially available, for detecting the presence of allelic
variants of cytochrome p450 isozymes in an individual or
determining the metabolizer phenotype of an individual for a
particular cytochrome p450 isozyme. Any suitable method known in
the art may be used. Methods include analyzing a blood sample from
the individual to determine the allelic variant of a particular
cytochrome p450 isozyme gene present in the individual (for example
by genotyping or haplotyping DNA or RNA from the gene using mass
spectrometry, gel electrophoresis, or TAQMAN assays; or analyzing
the protein sequence expressed by the gene). The metabolizer
phenotype of the individual can be inferred based on the known
properties of the allelic variants determined to be present in the
individual. Alternatively, the blood sample can be used to measure
enzyme activity of the cytochrome p450 isozyme using a suitable
assay amd isozyme-selective substrate. Among suitable
isozyme-selective substrates are those used in the studies herein,
or those suggested in FDA guidelines directed to collecting
cytochrome p450 isozyme data for regulatory submissions relating to
an active agent.
[0167] Food may alter the release of an active agent from a dosage
form, the solubilization of the active agent, and the transport of
the active agent across the intestinal wall. According to U.S. Pat.
No. 6,407,128, pharmacokinetic studies of metaxalone indicate that
food increases the rate and extent of absorption of a 400 mg oral
dosage form in humans. In that study, food increased peak plasma
concentrations (C.sub.max), and extent of absorption (AUC.sub.0-t,
AUC.sub.0-inf) relative to a fasted treatment with observed
increases of 177.5%, 123.5%, and 115.4%. Based on that study,
administration of metaxalone with food increases the
bioavailability of metaxalone and therefore a particular oral dose
given with food may physiologically correspond to a higher plasma
concentration of metaxalone than the same oral dose given in a
fasted state. Consequently, any effect on plasma concentration,
bioavailability, safety, efficacy, or a combination comprising at
least one of the foregoing of administration of metaxalone with an
additional substance which is a substrate, inhibitor, or inducer of
a cytochrome p450 isozyme for which metaxalone is a substrate,
inhibitor, or inducer can be further affected by whether or not the
metaxalone was administered with food.
[0168] Methods of using metaxalone comprise informing a user that
metaxalone affects the activity of a cytochrome p450; that
administration of metaxalone with a substance can affect the plasma
concentration, bioavailability, safety, efficacy, or a combination
comprising at least one of the foregoing of metaxalone or the
substance; and that any effect on the plasma concentration,
bioavailability, safety, efficacy, or a combination comprising at
least one of the foregoing of metaxalone or the substance can vary
with administration of metaxalone with or without food.
[0169] In another embodiment, the method of using metaxalone
comprises obtaining metaxalone from a container providing
information that metaxalone affects activity of a cytochrome p450;
that administration of metaxalone with a substance can affect the
plasma concentration, bioavailability, safety, or efficacy of
metaxalone or the substance; and that any effect on the plasma
concentration, bioavailability, safety, or efficacy of metaxalone
or the substance can vary with administration of metaxalone with or
without food. The method also includes providing metaxalone in the
container providing information.
[0170] In one embodiment, the metaxalone is always administered
with food. In another embodiment, the metaxalone is always
administered without food. In yet another embodiment, the
metaxalone is sometimes administered with food and sometimes
administered without food.
[0171] Also disclosed herein are methods of manufacturing a
metaxalone pharmaceutical product.
[0172] In one embodiment, the method comprises packaging a
metaxalone dosage form with information that metaxalone affects
activity of a cytochrome p450 isozyme. The information may also
advise that administration of metaxalone with a substance can
affect the plasma concentration, bioavailability, safety, efficacy,
or a combination comprising at least one of the foregoing of
metaxalone or the substance. The information may also include any
information disclosed herein about the effect of metaxalone or a
substance on the activity of a cytochrome p450 isozyme and any
information disclosed herein about the effect of metaxalone or a
substance on the plasma concentration, bioavailability, safety,
efficacy, or a combination comprising at least one of the foregoing
of metaxalone or the substance.
[0173] In an embodiment, the method comprises packaging a
metaxalone dosage form with information that metaxalone is
metabolized by a cytochrome p450 isozyme. The cytochrome p450
isozyme metabolizing metaxalone is CYP1A2, CYP2C8, CYP2C9, CYP2C19,
CYP2D6, CYP2E1, or CYP3A4. The information may also advise that
administration of metaxalone and a substance that is a substrate,
inhibitor, or inducer of CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6,
CYP2E1, or CYP3A4 can affect plasma concentration, bioavailability,
safety, efficacy, or a combination comprising at least one of the
foregoing of metaxalone or the substance.
[0174] In an embodiment, the method comprises packaging a
metaxalone dosage form with information that administration of
metaxalone with an active agent that is a CYP1A2, CYP2C8, CYP2C9,
CYP2C19, CYP2D6, CYP2E1, or CYP3A4 substrate having a narrow
therapeutic index can affect plasma concentration, bioavailability,
safety, efficacy, or a combination comprising at least one of the
foregoing of the active agent.
[0175] In another embodiment, the method comprises packaging a
metaxalone dosage form with information that metaxalone is an
inhibitor or an inducer of a cytochrome p450 isozyme. The
information may further advise that administration of metaxalone
with an active agent that is a substrate of the cytochrome p450
isozyme can affect the plasma concentration, bioavailability,
safety, efficacy, or a combination comprising at least one of the
foregoing of the active agent. The cytochrome p450 isozyme is
CYP1A2, CYP2B6, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or CYP3A4. In some
embodiments, the active agent is a substrate of the cytochrome p450
isozyme inhibited by metaxalone and the plasma concentration of the
active agent can increase; in other embodiments, the active agent
is a substrate of the cytochrome p450 isozyme induced by metaxalone
and the plasma concentration of the active agent can decrease.
[0176] In yet another embodiment, the method comprises packaging a
metaxalone dosage form with information that metaxalone affects
activity of a cytochrome p450 isozyme and that administration of
metaxalone with a substance can affect the plasma concentration,
bioavailability, safety, efficacy, or a combination comprising at
least one of the foregoing of the metaxalone or the substance; and
that any effect on the plasma concentration, bioavailability,
safety, efficacy, or a combination comprising at least one of the
foregoing of metaxalone or the substance can vary with
administration of metaxalone with or without food. In one
embodiment, the metaxalone is always administered with food. In
another embodiment, the metaxalone is always administered without
food. In yet another embodiment, the metaxalone is sometimes
administered with food and sometimes administered without food.
[0177] Another aspect of the invention is a method of using an
active agent that is a known substrate, inhibitor, or inducer of
CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or CYP3A4 or that
is a substrate of a cytochrome p450 isozyme.
[0178] In one embodiment, the method comprises informing a user
that metaxalone affects activity of a cytochrome p450 isozyme and
that administration of the active agent and metaxalone can affect
the plasma concentration, bioavailability, safety, efficacy, or a
combination comprising at least one of the foregoing of the active
agent or the metaxalone. The cytochrome p450 isozyme is CYP1A2,
CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or CYP3A4. In some
embodiments, the method further comprises providing the user with
the active agent or metaxalone.
[0179] In another embodiment, the method comprises obtaining an
active agent that is a known substrate, inhibitor, or inducer of
CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or CYP3A49 or that
is a substrate of a cytochrome p450 isozyme from a container
providing information that metaxalone affects activity of a
cytochrome p450 isozyme and that the administration of the active
agent with metaxalone can affect plasma concentration,
bioavailability, safety, efficacy, or a combination comprising at
least one of the foregoing of the active agent or the metaxalone.
The method may also comprise providing the active agent in the
container providing information.
[0180] Also disclosed herein is a method of manufacturing a
pharmaceutical product of an active agent that is a known
substrate, inhibitor, or inducer of CYP1A2, CYP2C8, CYP2C9,
CYP2C19, CYP2D6, CYP2E1, or CYP3A4 or that is a substrate of a
cytochrome p450 isozyme.
[0181] In one embodiment, the method comprises packaging a dosage
form of an active agent that is a known substrate, inhibitor, or
inducer of CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or
CYP3A4 or that is a substrate of a cytochrome p450 isozyme with
information that metaxalone affects activity of a cytochrome p450
isozyme.
[0182] In each of the methods for using an active agent that is a
known substrate, inhibitor, or inducer of CYP1A2, CYP2C8, CYP2C9,
CYP2C19, CYP2D6, CYP2E1, or CYP3A4 or that is a substrate of a
cytochrome p450 isozyme or the methods of manufacturing a
pharmaceutical product of such an active agent, the information
provided to the user or with the dosage form may include any
information disclosed herein about the effect of metaxalone or the
active agent on the activity of a cytochrome p450 isozyme and any
information disclosed herein about the effect of metaxalone or the
active agent on the plasma concentration, bioavailability, safety,
efficacy, or a combination comprising at least one of the foregoing
of metaxalone or the active agent.
[0183] The invention provides articles of manufacture.
[0184] In some embodiments, the article of manufacture comprises a
container containing a dosage form of metaxalone.
[0185] Metaxalone can be formulated as a dosage form for
administration where the formulation generally contains metaxalone
and a pharmaceutically acceptable excipient. As used herein,
"pharmaceutically acceptable excipient" means any other component
added to the pharmaceutical formulation other than the active
agent. Excipients may be added to facilitate manufacture, enhance
stability, control release, enhance product characteristics,
enhance bioavailability, enhance patient acceptability, etc.
Pharmaceutical excipients include carriers, fillers, binders,
disintegrants, lubricants, glidants, compression aids, colors,
sweeteners, preservatives, suspending agents, dispersing agents,
film formers, flavors, printing inks, buffer agents, pH adjusters,
preservatives etc.
[0186] In one embodiment, the container is associated with
published material informing that metaxalone affects activity of a
cytochrome p450 isozyme. The published material can further inform
that administration of metaxalone with a substance that is a
substrate, inhibitor, or inducer of the cytochrome p450 isozyme can
affect plasma concentration, bioavailability, safety, efficacy, or
a combination comprising at least one of the foregoing of
metaxalone or the substance. The published material may be in the
form of printed labeling, or in some other form. The cytochrome
p450 can be CYP1A2, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6,
CYP2E1, or CYP3A4. The published material comprising the article of
manufacture may also include any information disclosed herein about
the effect of metaxalone or a substance on the activity of a
cytochrome p450 isozyme and any information disclosed herein about
the effect of metaxalone or a substance on the plasma
concentration, bioavailability, safety, efficacy, or a combination
comprising at least one of the foregoing of metaxalone or the
substance.
[0187] In another embodiment, the container is associated with
published material informing that metaxalone is metabolized by a
cytochrome p450 isozyme. The cytochrome p450 isozyme metabolizing
metaxalone is CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or
CYP3A4. In some embodiments, the published material further informs
that administration of metaxalone with a substance that is a
substrate, inhibitor, or inducer of CYP1A2, CYP2C8, CYP2C9,
CYP2C19, CYP2D6, CYP2E1, or CYP3A4 can affect plasma concentration,
bioavailability, safety, efficacy, or a combination comprising at
least one of the foregoing of metaxalone or the substance. In other
embodiments, the published material further informs that a
substance that induces CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6,
CYP2E1, or CYP3A49 activity can decrease metaxalone plasma
concentration, that a substance that inhibits CYP1A2, CYP2C8,
CYP2C9, CYP2C19, CYP2D6, CYP2E1, or CYP3A4 activity can increase
metaxalone plasma concentration, or that a substance that is a
substrate of CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or
CYP3A4 can increase plasma concentration of metaxalone or the
substance.
[0188] In yet another embodiment, the container is associated with
published material informing that metaxalone is an inhibitor or an
inducer of a cytochrome p450 isozyme. The published material may
further inform that administration of metaxalone can affect the
plasma concentration, bioavailability, safety, efficacy, or a
combination comprising at least one of the foregoing of substances
that are substrates of the cytochrome p450 isozyme. The cytochrome
p450 isozyme is CYP1A2, CYP2B6, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or
CYP3A4.
[0189] In another embodiment, the container is associated with
published material that includes information that caution is
recommended when administering metaxalone with the substrate,
wherein the substrate has a narrow therapeutic index.
[0190] In yet another embodiment, the container is associated with
published material informing that metaxalone affects activity of a
cytochrome p450 isozyme; that administration to a patient of
metaxalone with a substance can affect plasma concentration,
bioavailability, safety, efficacy, or a combination comprising at
least one of the foregoing of metaxalone or the substance; and that
any effect on the plasma concentration, bioavailability, safety,
efficacy, or a combination comprising at least one of the foregoing
of metaxalone or the substance can vary with administration of
metaxalone with or without food.
[0191] In yet another embodiment, the article comprises a container
comprising a dosage form of metaxalone, and published material. In
one embodiment, the published material provides information that
there is a potential active agent interaction with warfarin; or
that administration with warfarin can affect the bioavailability,
safety, or efficacy of metaxalone or warfarin. In another
embodiment, the published material informs that metaxalone affects
activity of CYP1A2, CYP2B6, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or
CYP3A4. The published material may further inform that there is a
potential active agent interaction with a substance that is a
substrate of CYP1A2, CYP2B6, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or
CYP3A4 or that administration of metaxalone with a substance that
is a substrate of CYP1A2, CYP2B6, CYP2C9, CYP2C19, CYP2D6, CYP2E1,
or CYP3A4 can affect plasma concentration, bioavailability, safety,
efficacy, or a combination comprising at least one of the foregoing
of the substance. In another embodiment, the published material
informs that metaxalone is a substrate of CYP1A2, CYP2C8, CYP2C9,
CYP2C19, CYP2D6, CYP2E1, or CYP3A4. The published material may also
inform that there is a potential active agent interaction with a
substance that is a substrate, inhibitor, or inducer of CYP1A2,
CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or CYP3A4 or that
administration of metaxalone with a substance that is a substrate,
inhibitor, or inducer of CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6,
CYP2E1, or CYP3A4 can affect plasma concentration, bioavailability,
safety, efficacy, or a combination comprising at least one of the
foregoing of metaxalone or the substance. In yet another
embodiment, the published material informs that metaxalone is an
inhibitor of activity of CYP1A2, CYP2B6, CYP2C9, CYP2C19, CYP2D6,
CYP2E1, or CYP3A4. In yet another embodiment, the published
material informs that metaxalone is an inducer of activity of
CYP1A2 or CYP3A4. In each of these latter embodiments, the
published material may further inform that there is a potential
active agent interaction with a substance that is a substrate of
CYP1A2, CYP2B6, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or CYP3A4 or that
administration of metaxalone with a substance that is a substrate
of CYP1A2, CYP2B6, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or CYP3A4 can
affect plasma concentration, bioavailability, safety, efficacy, or
a combination comprising at least one of the foregoing of the
substance. In some embodiments, the published material can be
printed labeling.
[0192] Also disclosed herein is an article of manufacture
comprising packaging material and a dosage form contained within
the packaging material, wherein the dosage form comprises, as at
least one active ingredient, metaxalone, and wherein the packaging
material comprises a label approved by a regulatory agency for the
product. The label may inform that metaxalone affects activity of a
cytochrome p450 isozyme; that a cytochrome p450 isozyme
metabolizing metaxalone is CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6,
CYP2E1, or CYP3A4; that metaxalone is an inhibitor of activity of
CYP1A2, CYP2B6, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or CYP3A4; or that
metaxalone is an inducer of activity of CYP1A2 or CYP3A4. Examples
of regulatory agencies are the US FDA or the European Agency for
the Evaluation of Medicinal Products (EMEA).
[0193] The invention further includes an article of manufacture
comprising a container holding a dosage form of metaxalone
associated with published material informing that there is a
potential active agent interaction with warfarin, or that
administration with warfarin can affect the bioavailability,
safety, or efficacy of the metaxalone or the warfarin. The
published material may further comprise instructions regarding
measuring the Prothrombin Time/International Normalized Ratio
daily, every other day, weekly, every other week, monthly, or
according to another schedule or time criteria, or instructions to
monitor the blood levels of warfarin as AUC.sub.0-t, AUC.sub.0-INF,
C.sub.MAX, or a combination comprising one or more of the foregoing
pharmacokinetic parameters.
[0194] The invention includes articles of manufacture in which the
substance administered with metaxalone is phenyloin. In one
embodiment, the article of manufacture comprises a container
holding a dosage form of metaxalone associated with published
material informing that there is a potential active agent
interaction with phenyloin, or that administration of metaxalone
with phenyloin can affect the bioavailability, safety, efficacy or
a combination comprising at least one of the foregoing of the
metaxalone or the phenyloin. The published material may further
comprise instructions to monitor the blood levels of phenyloin as
AUC.sub.0-t, AUC.sub.0-INF, C.sub.MAX, or a combination comprising
one or more of the foregoing pharmacokinetic parameters.
[0195] Also disclosed herein is an article of manufacture
comprising a container containing a dosage form of an active agent
that is a known substrate, inhibitor, or inducer of CYP1A2, CYP2C8,
CYP2C9, CYP2C19, CYP2D6, CYP2E1, or CYP3A or that is a substrate of
a cytochrome p450 isozyme. The container is associated with
published material informing that metaxalone affects activity of a
cytochrome p450 isozyme and administration to a patient of the
active agent and metaxalone can affect plasma concentration,
bioavailability, safety, efficacy, or a combination comprising at
least one of the foregoing of the active agent or metaxalone. In
one embodiment of any of these methods or articles involving an
active agent that is a known substrate, inhibitor or inducer of
CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or CYP3A or that
is a substrate of a cytochrome p450 isozyme, the active agent is an
inducer of CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or
CYP3A and plasma concentration of metaxalone can decrease. In
another embodiment the active agent is an inhibitor of CYP1A2,
CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or CYP3A and plasma
concentration of metaxalone can increase. In yet another
embodiment, the active agent is a substrate of CYP1A2, CYP2C8,
CYP2C9, CYP2C19, CYP2D6, CYP2E1, or CYP3A and plasma concentration
of the active agent or metaxalone can increase. In yet another
embodiment, the active agent is a substrate of CYP1A2, CYP2B6,
CYP2C9, CYP2C19, CYP2D6, CYP2E1, or CYP3A4 and plasma concentration
of the active agent can increase. In yet another embodiment, the
active agent is a substrate of CYP1A2 or CYP3A4 and plasma
concentration of the active agent can decrease. In any of these
embodiments, the active agent can have a narrow therapeutic index.
The published material comprising the article of manufacture may
also include any information disclosed herein about the effect of
metaxalone or the active agent on the activity of a cytochrome p450
isozyme and any information disclosed herein about the effect of
metaxalone or the active agent on the plasma concentration,
bioavailability, safety, efficacy, or a combination comprising at
least one of the foregoing of metaxalone or the active agent.
[0196] In embodiments of the articles of manufacture, the dosage
form will typically be contained in a suitable container capable of
holding and dispensing the dosage form and which will not
significantly interact with the active agent(s) in the dosage form.
Further, the container will be in physical relation with the
published material. The published material may be associated with
the container by any means that maintains physical proximity of the
two. By way of example, the container and the published material
can both be contained in a packaging material such as a box or
plastic shrink wrap. Alternatively, the published material can be
bonded to the container, such as with glue that does not obscure
the published material, or with other bonding or holding means. Yet
another alternative is that the published material is placed within
the container with the dosage form.
[0197] In other embodiments of the article, someone hands the
published material to the patient, for example a pharmacist can
hand a product insert to a patient in conjunction with dispensing
the dosage form. The published material may be a product insert,
flyer, brochure, or a packaging material for the dosage form such
as a bag, or the like.
[0198] In any of the embodiments disclosed herein the published
material or information associated with or provided by a container
can be contained in any fixed and tangible medium. For example, the
information can be part of a leaflet, brochure, or other printed
material provided with a container or separate from a container.
The information can also take the form of a flyer, advertisement,
or the label for marketing the active agent approved by a
regulatory agency. The information can also be recorded on a
compact disk, DVD or any other recording or electronic medium.
[0199] The container can be in the form of bubble or blister pack
cards, optionally arranged in a desired order for a particular
dosing regimen. Suitable blister packs that can be arranged in a
variety of configurations to accommodate a particular dosing
regimen are well known in the art or easily ascertained by one of
ordinary skill in the art.
[0200] Metaxalone dosage forms existing as liquids, solutions,
emulsions, or suspensions can be packaged in a container for
convenient dosing of pediatric or geriatric patients. For example,
prefilled droppers (such as eye droppers or the like), prefilled
syringes, and similar containers housing the liquid, solution,
emulsion, or suspension form are contemplated.
[0201] The substance used with metaxalone in the methods and
articles of manufactures described herein may have certain effects,
direct or indirect, on the activity of a cytochrome p450 enzyme.
The substance or metaxalone can be a substrate, inhibitor, or
inducer of a Phase I or Phase II metabolic enzyme; specifically,
the substance or metaxalone is a substrate, inhibitor, or inducer
of a cytochrome p450 isozyme. More specifically, the substance can
be a substrate of CYP1A2, CYP2B6, CYP2C9, CYP2C19, CYP2D6, CYP2E1,
or CYP3A4, or an inhibitor or inducer of CYP1A2, CYP2C8, CYP2C9,
CYP2C19, CYP2D6, CYP2E1, or CYP3A. For example in certain
embodiments the substance is: a substrate, inhibitor, or inducer of
a cytochrome p450 isozyme; an active agent; a substrate, inhibitor,
or inducer of CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or
CYP3A activity; an active agent with a narrow therapeutic index; an
inducer of CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or
CYP3A activity and plasma concentration of metaxalone can decrease;
an inhibitor of CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or
CYP3A and plasma concentration of metaxalone can increase; a
substrate of CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or
CYP3A and plasma concentration of the substance or metaxalone can
increase; a substrate of CYP1A2, CYP2B6, CYP2C9, CYP2C19, CYP2D6,
CYP2E1, or CYP3A4 and plasma concentration of the substance can
increase; or the substance is a substrate of CYP1A2 or CYP3A4 and
plasma concentration of the substance can decrease when the
substance is administered with metaxalone.
[0202] In any of the above methods or articles, the substance can
be an active agent.
[0203] Examples of substrates of CYP1A2 include amitriptyline,
caffeine, clomipramine, clozapine, cyclobenzaprine, estradiol,
fluvoxamine, haloperidol, imipramine, mexiletine, naproxen,
olanzapine, ondansetron, phenacetin, acetaminophen, propranolol,
riluzole, ropivacaine, tacrine, theophylline, tizanidine,
verapamil, (R)-warfarin, zileuton, and zolmitriptan. Examples of
active agents that are inhibitors of CYP1A2 include amiodarone,
cimetidine, a fluoroquinolone (e.g., ciprofloxacin, gatifloxacin,
levofloxacin, lomefloxacin, moxifloxacin, norfloxacin, or
ofloxacin), fluvoxamine, furafylline, interferon, methoxsalen, and
mibefradil. Examples of inducers of CYP1A2 include chemicals
released from digestion of broccoli, brussel sprouts, and
char-grilled meat; chemicals inhaled when smoking tobacco; insulin,
methyl cholanthrene, modafinil, nafcillin, beta-naphthoflavone, and
omeprazole.
[0204] Examples of substrates of CYP2C19 include the proton pump
inhibitors: lansoprazole, omeprazole, pantoprazole, and E-3810; the
anti-epileptics: diazepam, phenyloin, fosphenyloin, S-mephenyloin,
and phenobarbitone (Phenobarbital); as well as amitriptyline,
carisoprodol, citalopram, clomipramine, cyclophosphamide,
hexobarbital, imipramine, indomethacin, R-mephobarbital,
moclobemide, nelfinavir, nilutamide, primidone, progesterone,
proguanil, propranolol, teniposide, and R-warfarin. Examples of
inhibitors of CYP2C19 include chloramphenicol, cimetidine,
felbamate, fluoxetine, fluvoxamine, indomethacin, ketoconazole,
lansoprazole, modafinil, omeprazole, oxcarbazepine, probenicid,
ticlopidine, and topiramate. Examples of inducers of CYP2C19
include carbamazepine, norethindrone, prednisone, and rifampin
(rifampicin).
[0205] Examples of substrates of CYP2B6 include bupropion,
cyclophosphamide, efavirenz, ifosfamide, and methadone.
[0206] Examples of inhibitors of CYP2C8 include quercetin, a
glitazone (e.g., rosiglitazone or pioglitazone), gemfibrozil,
montelukast, and trimethoprim. Examples of inducers of CYP2C8
include rifampin.
[0207] Examples of substrates of CYP2C9 include diclofenac,
ibuprofen, meloxicam, S-naproxen, piroxicam, suprofen, tolbutamide,
glipizide, losartan, irbesartan, glyburide (glibenclamide),
glipizide, glimepiride, amitriptyline, celecoxib, fluoxetine,
fluvastatin, nateglinide, phenyloin, rosiglitazone, tamoxifen,
torsemide, and S-warfarin. Examples of inhibitors of CYP2C9 include
amiodarone, fenofibrate, fluconazole, fluvastatin, fluvoxamine,
isoniazid, lovastatin, phenylbutazone, probenicid, sertraline,
sulfamethoxazole, sulfaphenazole, teniposide, voriconazole, and
zafirlukast. Examples of inducers of CYP2C9 include rifampin and
secobarbital.
[0208] Examples of substrates of CYP2D6 include carvedilol,
S-metoprolol, propafenone, timolol; amitriptyline, clomipramine,
desipramine, imipramine, paroxetine; haloperidol, perphenazine,
risperidone, thioridazine; alprenolol, amphetamine, aripiprazole,
atomoxetine, bufuralol, chlorpheniramine, chlorpromazine, codeine,
debrisoquine, dexfenfluramine, dextromethorphan, duloxetine,
encamide, flecamide, fluoxetine, fluvoxamine, lidocaine,
metoclopramide, methoxyamphetamine, mexiletine, minaprine,
nebivolol, nortriptyline, ondansetron, perhexyline, phenacetin,
phenformin, propranolol, sparteine, tamoxifen, tramadol, and
venlafaxine. Examples of inhibitors of CYP2D6 include amiodarone,
bupropion, celecoxib, chlorpromazine, chlorpheniramine, cimetidine,
citalopram, clomipramine, cocaine, doxepin, doxorubicin,
duloxetine, escitalopram, fluoxetine, halofantrine,
red-haloperidol, levomepromazine, metoclopramide, methadone,
mibefradil, midodrine, moclobemide, paroxetine, quinidine,
ranitidine, ritonavir, sertraline, terbinafine, ticlopidine,
histamine H1 receptor antagonists, diphenhydramine,
chlorpheniramine, clemastine, perphenazine, hydroxyzine, and
tripelennamine. Examples of inducers of CYP2D6 include rifampicin
and dexamethasone.
[0209] Examples of substrates of CYP2E1 include enflurane,
halothane, isoflurane, methoxyflurane, sevoflurane; acetaminophen,
aniline, benzene, chlorzoxazone, ethanol, N,N-dimethyl formamide,
and theophylline. Examples of inhibitors of CYP2E1 include
diethyl-dithiocarbamate and disulfuram. Examples of inducers of
CYP2E1 include ethanol and isoniazid.
[0210] Examples of substrates of CYP3A4 include clarithromycin,
erythromycin, telithromycin: quinidine; alprazolam, diazepam,
midazolam, triazolam; cyclosporine, tacrolimus (FK506); indinavir,
nelfinavir, ritonavir, saquinavir; cisapride; astemizole,
chlorpheniramine, terfenadine; amlodipine, diltiazem, felodipine,
lercanidipine, nifedipine, nisoldipine, nitrendipine, verapamil;
atorvastatin, cerivastatin, lovastatin, simvastatin; estradiol,
hydrocortisone, progesterone, testosterone; alfentanyl,
aripiprazole, buspirone, cafergot, caffeine, cilostazol, cocaine,
codeine, dapsone, dextromethorphan, docetaxel, domperidone,
eplerenone, fentanyl, finasteride, gleevec, haloperidol,
irinotecan, Levo-Alpha Acetyl Methadol (LAAM), lidocaine,
methadone, nateglinide, odanestron, pimozide, propranolol, quinine,
salmeterol, sildenafil, sirolimus, tamoxifen, taxol, terfenadine,
trazodone, vincristine, zaleplon, and zolpidem. Examples of
inhibitors of CYP3A4 include HIV Antivirals (e.g., delavirdine,
indinavir, nelfinavir, and ritonavir); amiodarone, aprepitant,
cinchloramphenicol, cimetidine, clarithromycin,
diethyl-dithiocarbamate, diltiazem, erythromycin, fluconazole,
fluvoxamine, gestodene, grapefruit juice, Seville orange juice,
imatinib, itraconazole, ketoconazole, mifepristone, nefazodone,
norfloxacin, norfluoxetine, mibefradil, star fruit, verapamil, and
voriconazole. Examples of inducers of CYP3A4 include HIV Antivirals
(e.g., efavirenz, and nevirapine); barbiturates (e.g.,
allobarbital, amobarbital, aprobarbital, alphenal, barbital,
brallobarbital, mephobarbital, secobarbital, and phenobarbital),
carbamazepine, efavirenz, glucocorticoids (e.g., prednisone,
prednisilone, methylprednisilone, dexamethasone, and
hydrocortisone), modafinil, nevirapine, phenyloin, rifampin, St.
John's wort, troglitazone, oxcarbazepine, pioglitazone, and
rifabutin.
[0211] In any of the embodiments described herein, the substance
can be a sensitive plasma concentration profile active agent.
Examples of a sensitive plasma concentration profile active agent
include cyclophosphamide, efavirenz, fosphenyloin, glimepiride,
mexiletine, phenyloin, progesterone, tamoxifen, theophylline,
warfarin, and any active agent having a narrow therapeutic
index.
[0212] In any of the embodiments described herein, the substance
can be an active agent having a narrow therapeutic index. Examples
of narrow therapeutic index active agents include warfarin,
phenyloin, fosphenyloin, thioridazine, theophylline, cyclosporine,
and pimozide.
[0213] In some embodiments, the active agent comprises warfarin.
Warfarin, 3-.alpha.-acetonylbenzyl)-4-hydroxycoumarin, is an
anticoagulant, which is eliminated by metabolism by cytochrome p450
isoforms including CYP2C9, CYP2C19, CYP2C8, CYP2C18, CYP1A2, and
CYP3A4. Warfarin has a narrow therapeutic index such that too
little can lead to excessive clotting, while excessive warfarin can
lead to excessive bleeding. The dosing of warfarin is
individualized according to the patient's sensitivity to the active
agent as indicated, for example, by the Prothrombin
Time/International Normalized Ratio (PT/INR). The PT/INR gives an
indication of how fast blood is clotting. The recommended initial
dose is 2-5 mg/day, with 2-10 mg/day as the maintenance dose.
Warfarin tablets for oral administration include tablets comprising
1, 2, 2.5, 3, 4, 5, 6, 7.5, and 10 mg of warfarin. The INR may be
adjusted to 2.0-4.5, or 2.0-3.0 or 2.5-3.5 depending on whether the
warfarin is being administered to treat venous thromboembolism,
non-valvular atrial fibrillation, post-myocardial infarction, heart
valve prophylaxis, or recurrent systemic embolism.
[0214] In the PT test, a reagent which induces coagulation is added
to a sample of the patient's plasma. The reagent typically
primarily comprises thromboplastin and calcium chloride. Many
commercially available PT reagents contain crude thromboplastin
extracted from natural sources, e.g., rabbit brain, rabbit
brain/lung mixtures, human placenta, or bovine brain, although
recombinant thromboplastin may also be employed. Prothrombin time
assays are performed by mixing the plasma sample and reagent at a
constant temperature such as 37.degree. C., and monitoring the
progress of the reaction until a perceptible clot (or "gel clot")
is detected. The development of a gel clot is the end point of the
reaction. This end point may be detected in various ways such as by
viscosity change, by electrode reaction, and, most commonly, by
photometric means. The test result is generally compared to a
result using a normal (control) plasma and converted to an INR.
[0215] The International Normalized Ratio, or INR, was developed to
standardize PT values, so that test results from different
thromboplastins and coagulation analyzers become equivalent. Under
the INR system, a thromboplastin is assigned an International
Sensitivity Index (ISI) value. The ISI indicates the relative
sensitivity of the thromboplastin compared to an international
reference thromboplastin. If a thromboplastin has the same
sensitivity as the reference thromboplastin, then its ISI is 1.0. A
higher ISI value indicates that a thromboplastin is less sensitive
than the reference thromboplastin. The ISI is used in the following
formula to calculate an INR value from a PT value: INR=(patient
PT/mean normal PT).sup.ISI. The ISI is usually determined by the
thromboplastin manufacturer. Different ISI values are assigned for
different models or classes of coagulation analyzers.
[0216] In another embodiment, the active agent comprises phenyloin.
Phenyloin, 5,5-diphenylhydantoin, is an antiepileptic active agent
useful in the treatment of epilepsy which is eliminated by
metabolism by cytochrome p450 isoforms including CYP1A2, CYP2C9,
CYP2C19, and CYP3A4. Phenyloin has a narrow therapeutic index such
that too little can lead to insufficient results and excessive
phenyloin can lead to phenyloin toxicity. The typical clinically
effective serum level is about 10 to about 20 .mu.g/mL. The
recommended initial dose is one 100 mg capsule 3 to 4 times per
day, with 300 mg/day dose in three divided doses or one single dose
per day. The dosing of phenyloin can be individualized according to
the patient's sensitivity to the active agent by measuring plasma
concentration of phenyloin.
[0217] Methods of treating a musculoskeletal condition or head pain
with metaxalone are provided herein. Such methods include informing
a user that metaxalone affects the activity of a cytochrome p450
isozyme. The method may further include informing the user that
administration of metaxalone with a substance can affect the plasma
concentration, bioavailability, safety, efficacy, or a combination
comprising at least one of the foregoing of metaxalone or the
substance. The method may also include informing the user of any
information disclosed herein about the effect of metaxalone or the
substance on the activity of a cytochrome p450 isozyme and any
information disclosed herein about the effect of metaxalone or the
substance on the plasma concentration, bioavailability, safety,
efficacy, or a combination comprising at least one of the foregoing
of metaxalone or the substance. Methods of treatment may also
include providing a user with metaxalone or administering
metaxalone to a patient.
[0218] Methods of treatment include methods in which the user is a
patient and additionally comprising administering metaxalone and an
active agent to the patient. The patient may be, for example, a
human patient, a patient in need of treatment of a musculoskeletal
condition or head pain, a patient receiving prophylactic metaxalone
treatment, or a patient undergoing metaxalone therapy. The amount
of metaxalone administered may be a therapeutically effective
amount.
[0219] Methods of treatment may additionally include monitoring the
patient's plasma concentration of the active agent as
AUC.sub.0-INF, AUC.sub.0-t, C.sub.MAX, or a combination of any of
the foregoing pharmacokinetic parameters. When metaxalone is
administered together with another active agent, methods of
treatment can include determining the plasma concentration of the
active agent and altering dosing of the active agent for the
patient based on the determined active agent plasma
concentration.
[0220] In another embodiment, a method of treatment comprises
administering to a patient in need of both a skeletal muscle
relaxant and an anticoagulant, for example, metaxalone and
warfarin, and monitoring the Prothrombin Time/International
Normalized Ratio. Monitoring the Prothrombin Time/International
Normalized Ratio may be performed for example daily, every other
day, weekly, every other week, monthly, or according to another
schedule or time criteria. The method may further comprise
providing to the patient or medical care worker instructions
regarding measuring the Prothrombin Time/International Normalized
Ratio.
[0221] When the substance administered with metaxalone is an NTI
active agent, methods using a blood test to monitor plasma levels
of the NTI active agent comprise administering to a patient
metaxalone and the NTI active agent, and monitoring the blood
levels of the NTI active agent as AUC.sub.0-t, AUC.sub.0-INF,
C.sub.MAX, or a combination comprising one or more of the foregoing
pharmacokinetic parameters.
[0222] In one embodiment, a method of using a blood test to monitor
warfarin levels comprises administering to a patient in need of
both a skeletal muscle relaxant and an anticoagulant both
metaxalone and warfarin, and monitoring the blood levels of
warfarin as AUC.sub.0-t, AUC.sub.0-INF, C.sub.MAX, or a combination
comprising one or more of the foregoing pharmacokinetic
parameters.
[0223] In another embodiment, the substance is phenyloin, and a
method using a blood test to monitor plasma levels of phenyloin
comprise administering to a patient metaxalone and phenyloin, and
monitoring the blood levels of phenyloin as AUC.sub.0-t,
AUC.sub.0-INF, C.sub.MAX, or a combination comprising one or more
of the foregoing pharmacokinetic parameters.
[0224] In all of the embodiments herein, a medical care worker can
determine the plasma concentration of an active agent by performing
or ordering the performance of any suitable method. For example,
the medical care worker could order a test using blood drawn from
the patient for determining the plasma concentration of the active
agent.
[0225] The invention is further illustrated by the following
examples.
EXAMPLE 1
Determination of Human Cytochrome p450 Isozymes Using Metaxalone as
a Substrate
[0226] The study of this example was performed to determine the
metabolism of metaxalone by human cytochrome p450 isoforms CYP1A2,
CYP2A6, CYP2C9, CYP2C19, CYP2D6, CYP2E1, and CYP3A4. Microsomes
containing singly-expressed human CYP isoforms were incubated in
the presence of metaxalone. The metabolism of metaxalone was
evaluated by measuring the disappearance of metaxalone by
high-performance liquid chromatography (HPLC).
[0227] Commercially available microsomes from baculovirus-infected
insect cells containing singly-expressed recombinant wild-type (*1
allele) human CYP enzymes and cDNA-expressed human cytochrome p450
oxidoreductase [BD SUPERSOMES Enzymes; BD Biosciences Discovery
Labware (Woburn, Mass.)] were used. For CYP2A6, CYP2C9, CYP2C19,
and CYP2E1, the SUPERSOMES also expressed human cytochrome b5 in
addition to human cytochrome p450 oxidoreductase and the human CYP
isozyme.
[0228] Metaxalone stock solutions were prepared in methanol at 100
times (100.times.) the final concentration. The stock solutions
were added to incubation mixtures to obtain final concentrations of
0.5, 2.5, and 25 .mu.M (corresponding to 111, 552, and 5530 ng
metaxalone/mL, respectively), each containing 1% methanol. All
incubations were conducted at 37.+-.1.degree. C. in a shaking water
bath with a sample size of N=3 replicates for each experimental
group. Incubation mixtures of microsomes (corresponding to 10 pmol
p450) and metaxalone were prepared in 0.1 M Tris buffer. After a
5-minute pre-incubation, an NADPH regenerating system (NRS) was
added to the incubation mixtures to initiate reactions, with a
final incubation volume of 0.5 mL. Incubations were continued for
30 minutes, and then terminated, except for those for CYP2C19,
which were incubated for 36 minutes prior to termination. Samples
were then analyzed for metaxalone.
[0229] Positive controls with a suitable isoform-selective
substrate were performed for each CYP isoform to verify metabolic
activity. Concentration of metabolites formed from CYP
isoform-selective substrates in the positive control samples was
analyzed using liquid chromatography/mass spectrometry (LC/MS) or
HPLC, as appropriate. A table of the substrate, substrate
concentration, solvent, metabolite formed, and metabolite assay
method for each CYP isozyme studied is below.
TABLE-US-00002 CYP Isoform-selective Substrate Metabolite isoform
substrate concentration Solvent Metabolite formed Assay CYP1A2
Phenacetin 50 .mu.M ACN Acetaminophen LC/MS CYP2A6 Coumarin 8 .mu.M
ACN 7-hydroxy coumarin HPLC-UV CYP2C9 Tolbutamide 150 .mu.M ACN
4'-methylhydroxytolbutamide LC/MS CYP2C19 S-Mephenytoin 50 .mu.M
ACN 4'-hydroxy mephenytoin LC/MS CYP2D6 Dextromethorphan 5 .mu.M
Water dextrorphan LC/MS CYP2E1 Chlorzoxazone 50 .mu.M ACN 6-hydroxy
chlorzoxazone LC/MS CYP3A4 Testosterone 100 .mu.M ACN
6.beta.-hydroxy testosterone HPLC-UV
[0230] Matrix controls were performed to determine the background
signal from the matrix components (microsomes (10 pmol p450), 01N
Tris buffer, NRS, and 1% methanol). Additionally metabolic negative
controls were performed to distinguish potential nonenzymatic
metabolism of metaxalone from p450-mediated metabolism. Incubation
mixtures were prepared in 0.1 M Tris buffer with SUPERSOMES (10
pmol P450) and metaxalone (at each concentration). After a 5-minute
pre-incubation, 2% sodium bicarbonate solution was added to the
incubation mixtures. Incubation was for 30 minutes at a final
volume of 0.5 mL. Matrix and metabolic negative controls were
terminated by adding an equal volume of methanol. The matrix
control and metabolic negative control samples were analyzed for
metaxalone by HPLC. Analysis of samples was subsequent to storage
at -70.degree. C.
[0231] Results are presented for each studied human cytochrome p450
isozyme in Tables 2-8.
TABLE-US-00003 TABLE 2 Metabolism of Metaxalone by Expressed
Recombinant Human CYP1A2 Metaxalone Metaxalone Present Percent of
Metabolic Concentration Raw Adjusted (.mu.M) Negative Control
(.mu.M) (.mu.M) Individual Mean .+-. SD Individual Mean .+-. SD MNC
0.20195 0.404 0.391 .+-. 0.0113 103 100 .+-. 2.88 (0.5) 0.19430
0.389 99.3 0.19097 0.382 97.6 0.5 0.15087 0.302 0.352 .+-. 0.0761
77.1 89.9 .+-. 19.4 0.21975 0.440 112 0.15734 0.315 80.4 MNC
0.65183 1.30 1.33 .+-. 0.0221 98.3 100 .+-. 1.67 (2.5) 0.66350 1.33
100 0.67394 1.35 102 2.5 0.52700 1.05 1.07 .+-. 0.0167 79.5 80.4
.+-. 1.26 0.52908 1.06 79.8 0.54235 1.08 81.8 MNC 10.11453 20.2
19.8 .+-. 0.360 102 100 .+-. 1.82 (25) 9.76568 19.5 98.5 9.86156
19.7 99.5 25 8.20521 16.4 16.6 .+-. 0.337 82.8 83.7 .+-. 1.70
8.19232 16.4 82.6 8.49030 17.0 85.6 MXC 0.00000.sup.a N/A N/A .+-.
N/A N/A N/A .+-. N/A (0) 0.00000.sup.a N/A N/A 0.00000.sup.a N/A
N/A Abbreviations: SD, standard deviation; MNC, metabolic negative
control; MXC, matrix control; N/A, not applicable .sup.aThe Raw
value (.mu.M) was below the lowest concentration on the standard
curve (0.05 .mu.M). Note: For all calculations above, the resulting
values are shown with at least three significant figures for
display purposes only.
TABLE-US-00004 TABLE 3 Metabolism of Metaxalone by Expressed
Recombinant Human CYP2A6 Metaxalone Metaxalone Present Percent of
Metabolic Concentration Raw Adjusted (.mu.M) Negative Control
(.mu.M) (.mu.M) Individual Mean .+-. SD Individual Mean .+-. SD MNC
0.15455 0.309 0.311 .+-. 0.00446 99.4 100 .+-. 1.43 (0.5) 0.15795
0.316 102 0.15375 0.308 98.9 0.5 0.15457 0.309 0.299 .+-. 0.0124
99.5 96.1 .+-. 3.99 0.15112 0.302 97.2 0.14253 0.285 91.7 MNC
0.74261 1.49 1.52 .+-. 0.0353 97.9 100 .+-. 2.33 (2.5) 0.75568 1.51
99.6 0.77755 1.56 102 2.5 0.79130 1.58 1.61 .+-. 0.0373 104 106
.+-. 2.46 0.79791 1.60 105 0.82642 1.65 109 MNC 7.74594 15.5 15.3
.+-. 0.147 101 100 .+-. 0.959 (25) 7.64948 15.3 99.8 7.60163 15.2
99.2 25 7.76399 15.5 15.6 .+-. 0.0975 101 102 .+-. 0.636 7.85044
15.7 102 7.84628 15.7 102 MXC 0.00000.sup.a N/A N/A .+-. N/A N/A
N/A .+-. N/A (0) 0.00000.sup.a N/A N/A 0.00000.sup.a N/A N/A
Abbreviations: SD, standard deviation; MNC, metabolic negative
control; MXC, matrix control; N/A, not applicable .sup.aThe Raw
value (.mu.M) was below the lowest concentration on the standard
curve (0.05 .mu.M) Note: For all calculations above, the resulting
values are shown with at least three significant figures for
display purposes only.
TABLE-US-00005 TABLE 4 Metabolism of Metaxalone by Expressed
Recombinant Human CYP2C9 Metaxalone Metaxalone Present Percent of
Metabolic Concentration Raw Adjusted (.mu.M) Negative Control
(.mu.M) (.mu.M) Individual Mean .+-. SD Individual Mean .+-. SD MNC
0.17052 0.341 0.348 .+-. 0.00997 97.9 100 .+-. 2.86 (0.5) 0.17229
0.345 98.9 0.17990 0.360 103 0.5 0.18004 0.360 0.355 .+-. 0.00608
103 102 .+-. 1.75 0.17784 0.356 102 0.17403 0.348 99.9 MNC 0.93197
1.86 1.93 .+-. 0.0605 96.8 100 .+-. 3.14 (2.5) 0.96526 1.93 100
0.99235 1.98 103 2.5 0.96842 1.94 1.92 .+-. 0.0246 101 99.7 .+-.
1.28 0.96593 1.93 100 0.94597 1.89 98.2 MNC 10.31249 20.6 21.3 .+-.
0.620 97.1 100 .+-. 2.92 (25) 10.63201 21.3 100 10.93245 21.9 103
25 10.66111 21.3 21.5 .+-. 0.144 100 101 .+-. 0.675 10.80454 21.6
102 10.72836 21.5 101 MXC 0.00000.sup.a N/A N/A .+-. N/A N/A N/A
.+-. N/A (0) 0.00000.sup.a N/A N/A 0.00000.sup.a N/A N/A
Abbreviations: SD, standard deviation; MNC, metabolic negative
control; MXC, matrix control; N/A, not applicable .sup.aThe Raw
value (.mu.M) was below the lowest concentration on the standard
curve (0.05 .mu.M) Note: For all calculations above, the resulting
values are shown with at least three significant figures for
display purposes only.
TABLE-US-00006 TABLE 5 Metabolism of Metaxalone by Expressed
Recombinant Human CYP2C19 Metaxalone Metaxalone Present Percent of
Metabolic Concentration Raw Adjusted (.mu.M) Negative Control
(.mu.M) (.mu.M) Individual Mean .+-. SD Individual Mean .+-. SD MNC
0.18718 0.374 0.370 .+-. 0.00898 101 100 .+-. 2.43 (0.5) 0.18763
0.375 102 0.17964 0.359 97.2 0.5 0.16773 0.335 0.345 .+-. 0.0104
90.8 93.4 .+-. 2.82 0.17180 0.344 93.0 0.17808 0.356 96.4 MNC
0.72720 1.45 1.39 .+-. 0.0560 105 100 .+-. 4.03 (2.5) 0.67562 1.35
97.2 0.68261 1.37 98.2 2.5 0.67218 1.34 1.34 .+-. 0.00561 96.7 96.5
.+-. 0.404 0.67254 1.35 96.7 0.66751 1.34 96.0 MNC 9.84488 19.7
20.1 .+-. 1.03 97.8 100 .+-. 5.13 (25) 9.69255 19.4 96.3 10.65287
21.3 106 25 9.34508 18.7 18.6 .+-. 0.120 92.9 92.6 .+-. 0.597
9.35948 18.7 93.0 9.24903 18.5 91.9 MXC 0.00000.sup.a N/A N/A .+-.
N/A N/A N/A .+-. N/A (0) 0.00000.sup.a N/A N/A 0.06454 N/A N/A
Abbreviations: SD, standard deviation; MNC, metabolic negative
control; MXC, matrix control; N/A, not applicable .sup.aThe Raw
value (.mu.M) was below the lowest concentration on the standard
curve (0.05 .mu.M) Note: For all calculations above, the resulting
values are shown with at least three significant figures for
display purposes only.
TABLE-US-00007 TABLE 6 Metabolism of Metaxalone by Expressed
Recombinant Human CYP2D6 Metaxalone Metaxalone Present Percent of
Metabolic Concentration Raw Adjusted (.mu.M) Negative Control
(.mu.M) (.mu.M) Individual Mean .+-. SD Individual Mean .+-. SD MNC
0.14509 0.290 0.292 .+-. 0.00220 99.4 100 .+-. 0.755 (0.5) 0.14716
0.294 101 0.14547 0.291 99.7 0.5 0.18683 0.374 0.319 .+-. 0.0477
128 109 .+-. 16.3 0.14857 0.297 102 0.14305 0.286 98.0 MNC 0.79025
1.58 1.56 .+-. 0.0184 101 100 .+-. 1.18 (2.5) 0.78433 1.57 100
0.77221 1.54 98.7 2.5 0.75826 1.52 1.53 .+-. 0.0111 96.9 97.7 .+-.
0.707 0.76852 1.54 98.2 0.76697 1.53 98.0 MNC 9.63762 19.3 19.2
.+-. 0.0994 100 100 .+-. 0.517 (25) 9.54788 19.1 99.4 9.62976 19.3
100 25 9.52577 19.1 19.2 .+-. 0.436 99.2 99.9 .+-. 2.27 9.84529
19.7 103 9.42917 18.9 98.2 MXC 0.00000.sup.a N/A N/A .+-. N/A N/A
N/A .+-. N/A (0) 0.00000.sup.a N/A N/A 0.00000.sup.a N/A N/A
Abbreviations: SD, standard deviation; MNC, metabolic negative
control; MXC, matrix control; N/A, not applicable .sup.aThe Raw
value (.mu.M) was below the lowest concentration on the standard
curve (0.05 .mu.M) Note: For all calculations above, the resulting
values are shown with at least three significant figures for
display purposes only.
TABLE-US-00008 TABLE 7 Metabolism of Metaxalone by Expressed
Recombinant Human CYP2E1 Metaxalone Metaxalone Present Percent of
Metabolic Concentration Raw Adjusted (.mu.M) Negative Control
(.mu.M) (.mu.M) Individual Mean .+-. SD Individual Mean .+-. SD MNC
0.18358 0.367 0.355 .+-. 0.0104 103 100 .+-. 2.92 (0.5) 0.17510
0.350 98.6 0.17416 0.348 98.1 0.5 0.17871 0.357 0.352 .+-. 0.00648
101 99.0 .+-. 1.83 0.17235 0.345 97.0 0.17662 0.353 99.4 MNC
0.89075 1.78 1.69 .+-. 0.117 105 100 .+-. 6.89 (2.5) 0.77998 1.56
92.2 0.86695 1.73 102 2.5 0.88299 1.77 1.76 .+-. 0.00318 104 104
.+-. 0.188 0.87990 1.76 104 0.88209 1.76 104 MNC 9.11125 18.2 17.8
.+-. 0.410 103 100 .+-. 2.30 (25) 8.70811 17.4 98.0 8.84728 17.7
99.5 25 8.73183 17.5 19.2 .+-. 2.71 98.2 108 .+-. 15.3 11.15149
22.3 125 8.87878 17.8 99.9 MXC 0.00000.sup.a N/A N/A .+-. N/A N/A
N/A .+-. N/A (0) 0.00000.sup.a N/A N/A 0.00000.sup.a N/A N/A
Abbreviations: SD, standard deviation; MNC, metabolic negative
control; MXC, matrix control; N/A, not applicable .sup.aThe Raw
value (.mu.M) was below the lowest concentration on the standard
curve (0.05 .mu.M) Note: For all calculations above, the resulting
values are shown with at least three significant figures for
display purposes only.
TABLE-US-00009 TABLE 8 Metabolism of Metaxalone by Expressed
Recombinant Human CYP3A4 Metaxalone Metaxalone Present Percent of
Metabolic Concentration Raw Adjusted (.mu.M) Negative Control
(.mu.M) (.mu.M) Individual Mean .+-. SD Individual Mean .+-. SD MNC
0.16014 0.320 0.318 .+-. 0.00502 101 100 .+-. 1.58 (0.5) 0.15592
0.312 98.2 0.16039 0.321 101 0.5 0.15978 0.320 0.320 .+-. 0.00333
101 101 .+-. 1.05 0.16159 0.323 102 0.15826 0.317 99.6 MNC 0.85285
1.71 1.72 .+-. 0.0127 99.3 100 .+-. 0.741 (2.5) 0.86553 1.73 101
0.85828 1.72 99.9 2.5 0.85730 1.71 1.68 .+-. 0.0289 99.8 98.0 .+-.
1.68 0.82923 1.66 96.5 0.83738 1.67 97.5 MNC 8.65154 17.3 17.4 .+-.
0.0906 99.4 100 .+-. 0.521 (25) 8.71767 17.4 100 8.73830 17.5 100
25 8.53809 17.1 17.1 .+-. 0.192 98.1 98.1 .+-. 1.10 8.44686 16.9
97.1 8.63905 17.3 99.3 MXC 0.00000.sup.a N/A N/A .+-. N/A N/A N/A
.+-. N/A (0) 0.00000.sup.a N/A N/A 0.00000.sup.a N/A N/A
Abbreviations: SD, standard deviation; MNC, metabolic negative
control; MXC, matrix control; N/A, not applicable .sup.aThe Raw
value (.mu.M) was below the lowest concentration on the standard
curve (0.05 .mu.M) Note: For all calculations above, the resulting
values are shown with at least three significant figures for
display purposes only.
[0232] Tables 2 and 5 show the results for human CYP1A2 and
CYP2C19, respectively. The results for these two cytochrome p450
isozymes show that metaxalone is a substrate for the enzymatic
activity of both CYP1A2 and CYP2C19.
[0233] Disappearance of metaxalone was detected following
incubation with CYP1A2 in the presence of the NADPH-regenerating
system. Disappearance of metaxalone ranged from 10.1% to 19.6%
(Table 2). The difference from the starting amount is statistically
significant at 2.5 and 25 .mu.M using an unpaired two-tailed t-test
(p.ltoreq.0.05). These results indicate that CYP1A2 is involved in
the metabolism of metaxalone.
[0234] In the experiments with CYP2C19, metaxalone disappearance
was evident following incubation with metaxalone at all three
concentrations (Table 5). The mean disappearance of metaxalone was
6.6% for the reaction using 0.5 .mu.M metaxalone; the reduction in
the mean amount of metaxalone from the value for the corresponding
metabolic negative control was statistically significant
(p.ltoreq.0.05) using an unpaired two-tailed t-test. The amount of
the disappearance of metaxalone observed at 2.5 or 25 .mu.M was not
statistically significant (p>0.05) compared to the mean values
for the corresponding metabolic negative controls using a
two-tailed t-test. These results indicate that CYP2C19 is also
involved in the metabolism of metaxalone, though to a lesser extent
than CYP1A2.
[0235] Experiments with the other tested cytochrome p450 isozymes
(Tables 3-4 and 6-8) failed to show any statistically significant
disappearance of metaxalone following incubation at the standard
conditions, indicating that, within the limits of detection for
these experiments, metaxalone was not used as a substrate by the
other tested cytochrome p450 isozymes: CYP2A6, CYP2C9, CYP2D6,
CYP2E1, and CYP3A4.
EXAMPLE 2
Metaxalone Inhibition of Cytochrome p450 Isozymes in Human
Microsomes
[0236] The study of this example was performed to determine the
potential of metaxalone to inhibit the activities of cytochrome
p450 (CYP) isoforms CYP1A2, CYP2A6, CYP2B6, CYP2C8, CYP2C9,
CYP2C19, CYP2D6, CYP2E1, and CYP3A4 in human liver microsomes.
Human liver microsomes were incubated in the presence of metaxalone
and a substrate selective for each CYP isoform. A table of the
substrate, substrate concentration, solvent, metabolite formed and
metabolite assay method for each CYP isozyme studied is below.
TABLE-US-00010 CYP Isoform-selective Substrate Metabolite isoform
substrate concentration Solvent Metabolite formed Assay CYP1A2
Phenacetin 50 .mu.M ACN acetaminophen LC/MS CYP2A6 Coumarin 8 .mu.M
ACN 7-hydroxy coumarin HPLC-UV CYP2B6 S-Mephenytoin 1 mM ACN
nirvanol LC/MS CYP2C8 Paclitaxel 5 .mu.M ACN 6-hydroxy paclitaxel
LC/MS CYP2C9 Tolbutamide 150 .mu.M ACN 4'-methylhydroxytolbutamide
LC/MS CYP2C19 S-Mephenytoin 50 .mu.M ACN 4'-hydroxy mephenytoin
LC/MS CYP2D6 Dextromethorphan 5 .mu.M Water dextrorphan LC/MS
CYP2E1 Chlorzoxazone 50 .mu.M ACN 6-hydroxy chlorzoxazone LC/MS
CYP3A4 Testosterone 100 .mu.M ACN 6.beta.-hydroxy testosterone
HPLC-UV
[0237] Metaxalone stock solutions were prepared in methanol at 100
times (100.times.) the final concentration and added to incubation
mixtures to obtain final concentrations of 0.3, 1, 3, 30, and 100
.mu.M (corresponding to 66.3, 221, 663, 6630 and 22,100 ng
metaxalone/mL, respectively), each containing 1% methanol.
[0238] Microsomes were prepared by differential centrifugation of
liver homogenates pooled from at least ten human donors.
[0239] All metaxalone incubations were conducted at 37.+-.1.degree.
C. in a shaking water bath using a sample size of N=3 replicates
for experimental groups. Incubation mixtures were prepared in 0.1 M
Tris buffer and contained microsomes (0.25 mg protein/mL for
CYP2C9, CYP2D6, CYP2E1, and CYP3A4; 0.5 mg protein/mL for CYP1A2,
CYP2A6, CYP2B6, CYP2C8, and CYP2C19), metaxalone (at each
concentration), and a CYP isoform-selective substrate. After a 5
minute preincubation, NADPH regenerating system (NRS) was added to
initiate the reaction. CYP2A6 and CYP3A4 incubations were for 10
minutes. All other incubations were for 30 minutes.
[0240] Incubations for CYP2C8 were terminated by adding 1.0 mL of
ACN, while all other incubations were terminated by adding 1.0 mL
of methanol. Samples were transferred to cryovials and analyzed
after storage at -70.degree. C. Triplicate replicates were
performed for each concentration of metaxalone for each cytochrome
p450 isozyme.
[0241] To verify that the test system was responsive to inhibitors,
a positive control using 1 .mu.M ketoconazole, a selective
inhibitor of CYP3A4, was added to CYP3A4 microsome incubations with
100 .mu.M testosterone. Four replicates were performed. The test
system was considered responsive to inhibitors since the mean
specific activity of CYP3A4 in the positive control samples treated
with ketoconazole was <14% of the mean specific activity in the
corresponding vehicle control samples.
[0242] Vehicle control experiments were performed to establish a
baseline value for enzyme activity. Incubation mixtures were
prepared in 0.1 M Tris buffer with microsomes (0.25 mg protein/mL
for CYP2C9, CYP2D6, CYP2E1, and CYP3A4; 0.5 mg protein/mL for
CYP1A2, CYP2A6, CYP2B6, CYP2C8, and CYP2C19), 1% methanol, and a
CYP isoform-selective substrate. Four replicates were
performed.
[0243] Metaxalone interference control samples were also included
to eliminate the possibility of interference by metaxalone or its
metabolites in detection of the metabolite formed from an
isoform-selective substrate. Two replicates were performed.
Incubation mixtures containing microsomes (0.25 mg protein/mL for
CYP2C9, CYP2D6, CYP2E1, and CYP3A4; 0.5 mg protein/mL for CYP1A2,
CYP2A6, CYP2B6, CYP2C8, and CYP2C19), 100 .mu.M metaxalone, and 1%
substrate solvent were prepared in 0.1 M Tris buffer. No
interference was detected in any of the metabolite assay methods
used.
[0244] Results for each CYP isoform, in the presence and absence of
metaxalone, are reported in Tables 9-17.
TABLE-US-00011 TABLE 9 CYP1A2 Activity in Pooled Human Microsomes
Acetaminophen formation Specific Activity Metaxalone Raw Adjusted
(.mu.M) (pmol/min/mg protein) Percent (.mu.M) (.mu.M) Individual
Mean .+-. SD Individual Mean .+-. SD of VC 0 0.23653 0.237 0.216
.+-. 0.0138 31.5 28.8 .+-. 1.84 100 (VC) 0.21124 0.211 28.2 0.21156
0.212 28.2 0.20568 0.206 27.4 0.3 0.21120 0.211 0.210 .+-. 0.00536
28.2 28.0 .+-. 0.715 97.2 0.21487 0.215 28.6 0.20431 0.204 27.2 1
0.19966 0.200 0.200 .+-. 0.00246 26.6 26.6 .+-. 0.327 92.3 0.19709
0.197 26.3 0.20200 0.202 26.9 3 0.19900 0.199 0.195 .+-. 0.00589
26.5 26.0 .+-. 0.785 90.3 0.18839 0.188 25.1 0.19813 0.198 26.4 30
0.18924 0.189 0.194 .+-. 0.00544 25.2 25.9 .+-. 0.725 89.8 0.19323
0.193 25.8 0.20000 0.200 26.7 100 0.17757 0.178 0.177 .+-. 0.000206
23.7 23.6 .+-. 0.0275 82.0 0.17733 0.177 23.6 0.17716 0.177 23.6
Abbreviations: SD, standard deviation; VC, vehicle control (1%
Methanol). Note: For all calculations above, the resulting values
are shown with at least three significant figures for display
purposes only.
TABLE-US-00012 TABLE 10 CYP2A6 Activity in Pooled Human Microsomes
7-Hydroxyycoumarin formation Specific Activity Metaxalone Raw
Adjusted (.mu.M) (pmol/min/mg protein) Percent (.mu.M) (.mu.M)
Individual Mean .+-. SD Individual Mean .+-. SD of VC 0 1.03214
1.03 1.06 .+-. 0.0356 413 426 .+-. 14.2 100 (VC) 1.04464 1.04 418
1.06891 1.07 428 1.11282 1.11 445 0.3 1.07439 1.07 1.03 .+-. 0.0399
430 413 .+-. 16.0 96.9 0.99553 0.996 398 1.02457 1.02 410 1 0.99854
0.999 1.02 .+-. 0.0184 399 407 .+-. 7.36 95.7 1.02269 1.02 409
1.03468 1.03 414 3 1.05100 1.05 1.09 .+-. 0.0402 420 436 .+-. 16.1
102 1.13132 1.13 453 1.08822 1.09 435 30 1.08205 1.08 1.14 .+-.
0.0493 433 455 .+-. 19.7 107 1.15129 1.15 461 1.17736 1.18 471 100
0.98864 0.989 1.01 .+-. 0.0416 395 404 .+-. 16.6 94.8 0.98209 0.982
393 1.05713 1.06 423 Abbreviations: SD, standard deviation; VC,
vehicle control (1% Methanol) Note: For all calculations above, the
resulting values are shown with at least three significant figures
for display purposes only.
TABLE-US-00013 TABLE 11 CYP2B6 Activity in Pooled Human Microsomes
Nirvanol formation Specific Activity Metaxalone Raw Adjusted
(.mu.M) (pmol/min/mg protein) Percent (.mu.M) (.mu.M) Individual
Mean .+-. SD Individual Mean .+-. SD of VC 0 0.23500 0.235 0.225
.+-. 0.0120 31.3 29.9 .+-. 1.60 100 (VC) 0.23266 0.233 31.0 0.22199
0.222 29.6 0.20877 0.209 27.8 0.3 0.20942 0.209 0.203 .+-. 0.00904
27.9 27.0 .+-. 1.21 90.2 0.19234 0.192 25.6 0.20601 0.206 27.5 1
0.20438 0.204 0.223 .+-. 0.0201 27.3 29.8 .+-. 2.68 99.5 0.22144
0.221 29.5 0.24442 0.244 32.6 3 0.19695 0.197 0.203 .+-. 0.00751
26.3 27.1 .+-. 1.00 90.6 0.20166 0.202 26.9 0.21166 0.212 28.2 30
0.21681 0.217 0.217 .+-. 0.00162 28.9 28.9 .+-. 0.216 96.6 0.21548
0.215 28.7 0.21871 0.219 29.2 100 0.18648 0.186 0.188 .+-. 0.00436
24.9 25.1 .+-. 0.581 83.7 0.18463 0.185 24.6 0.19293 0.193 25.7
Abbreviations: SD, standard deviation; VC, vehicle control (1%
Methanol) Note: For all calculations above, the resulting values
are shown with at least three significant figures for display
purposes only.
TABLE-US-00014 TABLE 12 CYP2C8 Activity in Pooled Human Microsomes
6-Hydroxypaclitaxel formation Specific Activity Metaxalone Raw
Adjusted (.mu.M) (pmol/min/mg protein) Percent (.mu.M) (.mu.M)
Individual Mean .+-. SD Individual Mean .+-. SD of VC 0 0.13462
0.135 0.136 .+-. 0.00522 17.9 18.2 .+-. 0.696 100 (VC) 0.14017
0.140 18.7 0.14074 0.141 18.8 0.12965 0.130 17.3 0.3 0.14476 0.145
0.126 .+-. 0.0163 19.3 16.8 .+-. 2.18 92.7 0.11377 0.114 15.2
0.12042 0.120 16.1 1 0.13927 0.139 0.140 .+-. 0.00305 18.6 18.7
.+-. 0.406 103 0.13749 0.137 18.3 0.14343 0.143 19.1 3 0.15034
0.150 0.149 .+-. 0.00174 20.0 19.9 .+-. 0.232 109 0.14945 0.149
19.9 0.14698 0.147 19.6 30 0.14949 0.149 0.138 .+-. 0.0114 19.9
18.4 .+-. 1.52 101 0.13724 0.137 18.3 0.12667 0.127 16.9 100
0.13170 0.132 0.133 .+-. 0.0207 17.6 17.8 .+-. 2.76 97.8 0.15467
0.155 20.6 0.11340 0.113 15.1 Abbreviations: SD, standard
deviation; VC, vehicle control (1% Methanol) Note: For all
calculations above, the resulting values are shown with at least
three significant figures for display purposes only.
TABLE-US-00015 TABLE 13 CYP2C9 Activity in Pooled Human Microsomes
4'-Methylhydroxytolbutamide formation Specific Activity Metaxalone
Raw Adjusted (.mu.M) (pmol/min/mg protein) Percent (.mu.M) (.mu.M)
Individual Mean .+-. SD Individual Mean .+-. SD of VC 0 0.17476
0.175 0.166 .+-. 0.0208 46.6 44.3 .+-. 5.54 100 (VC) 0.14904 0.149
39.7 0.14954 0.150 39.9 0.19164 0.192 51.1 0.3 0.13620 0.136 0.135
.+-. 0.00106 36.3 36.1 .+-. 0.283 81.4 0.13415 0.134 35.8 0.13565
0.136 36.2 1 0.15107 0.151 0.136 .+-. 0.0187 40.3 36.2 .+-. 4.98
81.6 0.14080 0.141 37.5 0.11485 0.115 30.6 3 0.13051 0.131 0.135
.+-. 0.0103 34.8 36.0 .+-. 2.75 81.2 0.12759 0.128 34.0 0.14670
0.147 39.1 30 0.14975 0.150 0.151 .+-. 0.00841 39.9 40.3 .+-. 2.24
91.0 0.14376 0.144 38.3 0.16037 0.160 42.8 100 0.16269 0.163 0.145
.+-. 0.0150 43.4 38.8 .+-. 4.00 87.4 0.13711 0.137 36.6 0.13627
0.136 36.3 Abbreviations: SD, standard deviation; VC, vehicle
control (1% Methanol) Note: For all calculations above, the
resulting values are shown with at least three significant figures
for display purposes only.
TABLE-US-00016 TABLE 14 CYP2C19 Activity in Pooled Human Microsomes
4'-Hydroxymephenytoin formation Specific Activity Metaxalone Raw
Adjusted (.mu.M) (pmol/min/mg protein) Percent (.mu.M) (.mu.M)
Individual Mean .+-. SD Individual Mean .+-. SD of VC 0 0.16904
0.169 0.168 .+-. 0.00550 22.5 22.4 .+-. 0.733 100 (VC) 0.17373
0.174 23.2 0.16915 0.169 22.6 0.16055 0.161 21.4 0.3 0.13971 0.140
0.142 .+-. 0.00299 18.6 19.0 .+-. 0.399 84.6 0.14558 0.146 19.4
0.14164 0.142 18.9 1 0.11367 0.114 0.113 .+-. 0.00140 15.2 15.0
.+-. 0.186 67.0 0.11336 0.113 15.1 0.11111 0.111 14.8 3 0.11597
0.116 0.114 .+-. 0.00238 15.5 15.2 .+-. 0.317 67.7 0.11127 0.111
14.8 0.11423 0.114 15.2 30 0.08336 0.0834 0.107 .+-. 0.0211 11.1
14.3 .+-. 2.82 63.8 0.12339 0.123 16.5 0.11502 0.115 15.3 100
0.10857 0.109 0.109 .+-. 0.00205 14.5 14.5 .+-. 0.274 64.9 0.11132
0.111 14.8 0.10730 0.107 14.3 Abbreviations: SD, standard
deviation; VC, vehicle control (1% Methanol) Note: For all
calculations above, the resulting values are shown with at least
three significant figures for display purposes only.
TABLE-US-00017 TABLE 15 CYP2D6 Activity in Pooled Human Microsomes
Dextrorphan formation Specific Activity Adjusted (.mu.M)
(pmol/min/mg protein) Metaxalone (.mu.M) Raw (.mu.M) Individual
Mean .+-. SD Individual Mean .+-. SD Percent of VC 0 0.18550 0.186
0.183 .+-. 0.00342 49.5 48.9 .+-. 0.911 100 (VC) 0.18569 0.186 49.5
0.18424 0.184 49.1 0.17843 0.178 47.6 0.3 0.14820 0.148 0.149 .+-.
0.00258 39.5 39.8 .+-. 0.688 81.3 0.14716 0.147 39.2 0.15206 0.152
40.5 1 0.15910 0.159 0.154 .+-. 0.00482 42.4 41.2 .+-. 1.28 84.2
0.14949 0.149 39.9 0.15485 0.155 41.3 3 0.16116 0.161 0.164 .+-.
0.00353 43.0 43.7 .+-. 0.940 89.3 0.16267 0.163 43.4 0.16788 0.168
44.8 30 0.15533 0.155 0.156 .+-. 0.00335 41.4 41.6 .+-. 0.893 85.1
0.15983 0.160 42.6 0.15328 0.153 40.9 100 0.15992 0.160 0.158 .+-.
0.00255 42.6 42.0 .+-. 0.680 85.9 0.15489 0.155 41.3 0.15813 0.158
42.2 Abbreviations: SD, standard deviation; VC, vehicle control (1%
Methanol) Note: For all calculations above, the resulting values
are shown with at least three significant figures for display
purposes only.
TABLE-US-00018 TABLE 16 CYP2E1 Activity in Pooled Human Microsomes
6-Hydroxychlorzoxazone formation Specific Activity Adjusted (.mu.M)
(pmol/min/mg protein) Metaxalone (.mu.M) Raw (.mu.M) Individual
Mean .+-. SD Individual Mean .+-. SD Percent of VC 0 0.85346 0.853
0.852 .+-. 0.0127 228 227 .+-. 3.39 100 (VC) 0.86925 0.869 232
0.84615 0.846 226 0.83969 0.840 224 0.3 0.73634 0.736 0.710 .+-.
0.0228 196 189 .+-. 6.08 83.3 0.69947 0.699 187 0.69469 0.695 185 1
0.72701 0.727 0.716 .+-. 0.0194 194 191 .+-. 5.18 84.0 0.72685
0.727 194 0.69326 0.693 185 3 0.76089 0.761 0.755 .+-. 0.0110 203
201 .+-. 2.94 88.6 0.74221 0.742 198 0.76169 0.762 203 30 0.71716
0.717 0.733 .+-. 0.0145 191 196 .+-. 3.88 86.1 0.74538 0.745 199
0.73733 0.737 197 100 0.74969 0.750 0.743 .+-. 0.0175 200 198 .+-.
4.66 87.2 0.75620 0.756 202 0.72321 0.723 193 Abbreviations: SD,
standard deviation; VC, vehicle control (1% Methanol) Note: For all
calculations above, the resulting values are shown with at least
three significant figures for display purposes only.
TABLE-US-00019 TABLE 17 CYP3A4 Activity in Pooled Human Microsomes
6.beta.-Hydroxytestosterone formation Specific Activity Adjusted
(.mu.M) (pmol/min/mg protein) Metaxalone (.mu.M) Raw (.mu.M)
Individual Mean .+-. SD Individual Mean .+-. SD Percent of VC 0
0.12662* N/A 0.742 .+-. 0.00679 N/A 594 .+-. 5.43 100 (VC) 0.74589
0.746 597 0.74640 0.746 597 0.73440 0.734 588 0.3 0.64318 0.643
0.647 .+-. 0.0130 515 517 .+-. 10.4 87.1 0.66083 0.661 529 0.63550
0.636 508 1 0.65762 0.658 0.654 .+-. 0.00353 526 523 .+-. 2.83 88.1
0.65446 0.654 524 0.65057 0.651 520 3 0.67154 0.672 0.668 .+-.
0.00420 537 534 .+-. 3.36 90.0 0.66336 0.663 531 0.66907 0.669 535
30 0.62513 0.625 0.633 .+-. 0.0370 500 506 .+-. 29.6 85.2 0.67282
0.673 538 0.59996 0.600 480 100 0.63960 0.640 0.596 .+-. 0.0454 512
477 .+-. 36.3 80.3 0.59940 0.599 480 0.54904 0.549 439
Abbreviations: SD, standard deviation; VC, vehicle control (1%
Methanol) *Sample has been removed from all calculations due to the
incorrect volume being added to the sample to stop the reaction.
Note: For all calculations above, the resulting values are shown
with at least three significant figures for display purposes
only.
[0245] Under these experimental conditions, metaxalone inhibited
activities of CYP1A2, CYP2B6, CYP2C19, CYP2D6, CYP2E1, and CYP3A4
in human liver microsomes at one or more of the tested metaxalone
concentrations at a statistically significant level (p.ltoreq.0.05
using an unpaired two-tailed t-test). The inhibition ranged from
12.8% (CYP2E1) to 35.1% (CYP2C19) at a metaxalone concentration of
100 .mu.M.
[0246] Under these experimental conditions, no tested concentration
of metaxalone inhibited activity of CYP2A6, CYP2C8, or CYP2C9 in
human liver microsomes at a statistically significant level
(p>0.05 using an unpaired two-tailed t-test).
EXAMPLE 3
Metaxalone Induction/Inhibition of Cytochrome p450 Isozymes
[0247] The study of this example was performed to determine if
there is induction or inhibition by metaxalone of cytochrome p450
isozymes CYP1A2, CYP2A6, CYP2B6, CYP2C9, CYP2C19, CYP2D6, CYP2E1,
and CYP3A4. These induction/inhibition studies used cryopreserved
human hepatocytes and compared enzymatic activity levels for each
of these cytochrome p450 isozymes, using an appropriate enzyme
substrate, in the human hepatocytes following in vitro exposure for
48.+-.3 hrs in the presence or absence of metaxalone.
[0248] Hepatocytes from three human donors were obtained from a
cryopreserved hepatocyte bank (In Vitro Technologies, Inc., USA).
After thawing, viable hepatocytes were transferred to
collagen-coated 48-well plates for attachment in plating medium
(DMEM stock (Dulbecco's modified Eagle's medium, supplemented with
bovine serum albumin, fructose,
N-(2-hydroxyethyl)piperazine-N'-(2-ethanesulfonate) (HEPES), and
sodium bicarbonate), supplemented with antibiotics, bovine serum,
hydrocortisone, insulin and minimum essential medium (MEM)
nonessential amino acids). After attachment to the collagen matrix,
plating medium was replaced with sandwich medium (plating medium
supplemented with VITROGEN) and incubated until use. All
incubations were conducted at 37.+-.1.degree. C., 95% air/5%
CO.sub.2 and saturating humidity.
[0249] After establishment of the hepatocyte culture, sandwich
medium was removed and the hepatocytes were incubated with
incubation solution (DMEM stock supplemented with antibiotics,
hydrocortisone, insulin, and MEM non-essential amino acids)
containing 0.4, 4.0, or 40 .mu.M metaxalone for 24.+-.1.5 hrs.
Incubation solution was aspirated and replaced with incubation
solution containing the same concentration of metaxalone and
incubated for an additional 24.+-.1.5 hrs. After the metaxalone
treatment period, the incubation solution was replaced with 150
.mu.L Krebs-Henseleit (KHB) buffer supplemented with antibiotics,
calcium chloride, heptanoic acid, HEPES, and sodium bicarbonate
(supplemented KHB) and incubated for 10 minutes. The supplemented
KHB was replaced with 150 .mu.L supplemented KHB containing the
appropriate isoform-selective substrate and incubated for 4 hrs
prior to termination by adding 150 .mu.L ice-cold methanol, except
for the CYP2C8 incubations which were terminated by adding 150
.mu.L acetonitrile. Samples were transferred to cryovials and
analyzed after storage at -70.degree. C. Three induction replicates
were performed at each metaxalone concentration for each cytochrome
p450 isozyme.
[0250] A table of the substrate, substrate concentration,
metabolite formed, and metabolite assay method for each CYP isozyme
studied is provided below. All substrates were dissolved in
acetonitrile.
TABLE-US-00020 Isoform-selective Substrate Metabolite CYP isoform
substrate concentration Metabolite formed Assay CYP1A2 Phenacetin
100 .mu.M acetaminophen LC/MS CYP2A6 Coumarin 100 .mu.M
7-hydroxycoumarin, 7- HPLC-UV hydroxycoumaringlucuronide, 7-
hydroxycoumarin sulfate CYP2B6 S-Mephenytoin 1 mM nirvanol LC/MS
CYP2C9 Tolbutamide 50 .mu.M 4'-methylhydroxytolbutamide LC/MS
CYP2C19 S-Mephenytoin 100 .mu.M 4'-hydroxy mephenytoin LC/MS CYP2D6
Dextromethorphan 16 .mu.M dextrorphan LC/MS CYP2E1 Chlorzoxazone
300 .mu.M 6-hydroxychlorzoxazone LC/MS CYP3A4 Testosterone 125
.mu.M 6.beta.-hydroxy testosterone HPLC-UV
[0251] Metaxalone 100.times. stock solutions were prepared in
methanol as described above and diluted with incubation medium to
produce incubation solutions with 0.4, 4.0, and 40 .mu.M
metaxalone.
[0252] Replicate trials and controls were performed. Positive
controls (n=4) were performed to verify that the test system was
sensitive to known inducers by testing induction of CYP1A2 and
CYP3A4 using 50 .mu.M omeprazole and 25 .mu.M rifampicin,
respectively, as inducers with the appropriate isoform-selective
substrate. Both positive control test systems showed .gtoreq.200%
induction. Additionally, reference control samples were included to
evaluate inducibility of CYP2B6, CYP2C9, and CYP2C19 in the test
system. The reference controls included 1 mM Phenobarbital (CYP2B6)
or 25 .mu.M rifampicin as the reference inducer. The reference
controls showed a statistically significant amount of induction for
each hepatocyte donor for CYP2B6 and CYP2C9, although the amount of
induction varied between the three hepatocyte donors for each
isozyme. For CYP2C19, rifampin induced CYP2C19 activity in donor 1
and donor 3, but did not induce CYP2C19 activity in donor 2 at a
statistically significant level (p<0.05 using an unpaired
two-tailed t-test).
[0253] Results for each cytochrome p450 isozyme are shown in Tables
18-25. Significant induction was observed at these experimental
conditions in all three donors for CYP1A2 and in one donor for
CYP3A4 at the highest tested concentration. Additionally,
significant inhibition in enzyme activity was observed in all three
donors for CYP2C9 and in two donors for CYP2D6. Under these
experimental conditions, no significant effects on activity of
CYP2A6, CYP2B6, CYP2C19, or CYP2E1 were observed after exposure to
any of the tested concentrations of metaxalone. Significance of a
change in specific activity from that measured for the vehicle
control (0 .mu.M metaxalone) was determined using a two-tailed
t-test. Mean specific activity values with associated
p-values.ltoreq.0.05 were deemed to be statistically
significant.
TABLE-US-00021 TABLE 18 CYP1A2 Activity in Cryopreserved Human
Hepatocytes Acetaminophen formation Specific Activity Adjusted
(.mu.M) (pmol/min/million cells) Metaxalone (.mu.M) Raw (.mu.M)
Individual Mean .+-. SD Individual Mean .+-. SD Percent of VC Donor
1 0 0.05388 0.0539 0.0487 .+-. 0.00543 0.481 0.435 .+-. 0.0485 100
(VC) 0.05227 0.0523 0.467 0.04658 0.0466 0.416 0.04203 0.0420 0.375
0.4 0.05121 0.0512 0.0537 .+-. 0.00309 0.457 0.479 .+-. 0.0276 110
0.05264 0.0526 0.470 0.05714 0.0571 0.510 4 0.07410 0.0741 0.0638
.+-. 0.0193 0.662 0.570 .+-. 0.172 131 0.07581 0.0758 0.677 0.04160
0.0416 0.371 40 0.15156 0.152 0.161 .+-. 0.0133 1.35 1.44 .+-.
0.119 332 0.15617 0.156 1.39 0.17659 0.177 1.58 Donor 2 0 0.03023
0.0302 0.0300 .+-. 0.00305 0.270 0.267 .+-. 0.0272 100 (VC) 0.03210
0.0321 0.287 0.03193 0.0319 0.285 0.02556 0.0256 0.228 0.4 0.03165
0.0317 0.0323 .+-. 0.000850 0.283 0.289 .+-. 0.00759 108 0.03208
0.0321 0.286 0.03329 0.0333 0.297 4 0.03346 0.0335 0.0340 .+-.
0.00198 0.299 0.304 .+-. 0.0177 113 0.03619 0.0362 0.323 0.03234
0.0323 0.289 40 0.06015 0.0602 0.0589 .+-. 0.00795 0.537 0.526 .+-.
0.0710 197 0.06616 0.0662 0.591 0.05040 0.0504 0.450 Donor 3 0
0.04357 0.0436 0.0410 .+-. 0.00447 0.389 0.366 .+-. 0.0399 100 (VC)
0.04576 0.0458 0.409 0.03607 0.0361 0.322 0.03849 0.0385 0.344 0.4
0.04030 0.0403 0.0438 .+-. 0.00361 0.360 0.391 .+-. 0.0322 107
0.04347 0.0435 0.388 0.04750 0.0475 0.424 4 0.04411 0.0441 0.0443
.+-. 0.000214 0.394 0.396 .+-. 0.00191 108 0.04453 0.0445 0.398
0.04425 0.0443 0.395 40 0.12276 0.123 0.122 .+-. 0.00365 1.10 1.09
.+-. 0.0326 297 0.11776 0.118 1.05 0.12487 0.125 1.11
Abbreviations: SD, standard deviation; VC, vehicle control (1%
Methanol) Note: For all calculations above, the resulting values
are shown with at least three significant figures for display
purposes only.
TABLE-US-00022 TABLE 19 CYP2A6 Activity in Cryopreserved Human
Hepatocytes Total Metabolite formation Specific Activity Adjusted
(.mu.M) (pmol/min/million cells) Metaxalone (.mu.M) Raw (.mu.M)
Individual Mean .+-. SD Individual Mean .+-. SD Percent of VC Total
Metabolite Formation: Donor 1 0 0.0171.sup.d <0.300 <0.300
.+-. 0.000 <2.68 <2.68 .+-. 0.000 100 (VC) 0.000.sup.d
<0.300 <2.68 0.000.sup.d <0.300 <2.68 0.000.sup.d
<0.300 <2.68 0.4 0.000.sup.d <0.300 <0.300 .+-. 0.000
<2.68 <2.68 .+-. 0.000 100 0.000.sup.d <0.300 <2.68
0.000.sup.d <0.300 <2.68 4 0.000.sup.d <0.300 <0.300
.+-. 0.000 <2.68 <2.68 .+-. 0.000 100 0.000.sup.d <0.300
<2.68 0.000.sup.d <0.300 <2.68 40 0.000.sup.d <0.300
<0.300 .+-. 0.000 <2.68 <2.68 .+-. 0.000 100 0.000.sup.d
<0.300 <2.68 0.000.sup.d <0.300 <2.68 Total Metabolite
Formation: Donor 2 0 0.0381.sup.d <0.300 <0.300 .+-. 0.000
<2.68 <2.68 .+-. 0.000 100 (VC) 0.0413.sup.d <0.300
<2.68 0.0365.sup.d <0.300 <2.68 0.0320.sup.d <0.300
<2.68 0.4 0.0225.sup.d <0.300 <0.300 .+-. 0.000 <2.68
<2.68 .+-. 0.000 100 0.0381.sup.d <0.300 <2.68
0.0381.sup.d <0.300 <2.68 4 0.0344.sup.d <0.300 <0.300
.+-. 0.000 <2.68 <2.68 .+-. 0.000 100 0.0353.sup.d <0.300
<2.68 0.0297.sup.d <0.300 <2.68 40 0.0293.sup.d <0.300
<0.300 .+-. 0.000 <2.68 <2.68 .+-. 0.000 100 0.0266.sup.d
<0.300 <2.68 0.0333.sup.d <0.300 <2.68 Total Metabolite
Formation: Donor 3 0 0.000.sup.d <0.300 <0.300 .+-. 0.000
<2.68 <2.68 .+-. 0.000 100 (VC) 0.0196.sup.d <0.300
<2.68 0.0237.sup.d <0.300 <2.68 0.000.sup.d <0.300
<2.68 0.4 0.0216.sup.d <0.300 <0.300 .+-. 0.000 <2.68
<2.68 .+-. 0.000 100 0.0182.sup.d <0.300 <2.68
0.0182.sup.d <0.300 <2.68 4 0.000.sup.d <0.300 <0.300
.+-. 0.000 <2.68 <2.68 .+-. 0.000 100 0.0197.sup.d <0.300
<2.68 0.0162.sup.d <0.300 <2.68 40 0.000.sup.d <0.300
<0.300 .+-. 0.000 <2.68 <2.68 .+-. 0.000 100 0.0188.sup.d
<0.300 <2.68 0.000.sup.d <0.300 <2.68 Abbreviations:
SD, standard deviation; VC, vehicle control (1% Methanol) .sup.dThe
observed analyzed value (.mu.M) for all metabolites were below the
lowest concentration on the corresponding standard curve. Note: For
all calculations above, the resulting values are shown with at
least three significant figures for display purposes only.
TABLE-US-00023 TABLE 20 CYP2B6 Activity in Cryopreserved Human
Hepatocytes Nirvanol formation Specific Activity Adjusted (.mu.M)
(pmol/min/million cells) Metaxalone (.mu.M) Raw (.mu.M) Individual
Mean .+-. SD Individual Mean .+-. SD Percent of VC Donor 1 0
0.03230 0.0323 0.0319 .+-. 0.00156 0.288 0.285 .+-. 0.0139 100 (VC)
0.03384 0.0338 0.302 0.03014 0.0301 0.269 0.03141 0.0314 0.280 0.4
0.03380 0.0338 0.0340 .+-. 0.000883 0.302 0.304 .+-. 0.00789 107
0.03329 0.0333 0.297 0.03501 0.0350 0.313 4 0.02742 0.0274 0.0305
.+-. 0.00272 0.245 0.273 .+-. 0.0243 95.7 0.03241 0.0324 0.289
0.03178 0.0318 0.284 40 0.03233 0.0323 0.0310 .+-. 0.00204 0.289
0.277 .+-. 0.0182 97.1 0.03203 0.0320 0.286 0.02866 0.0287 0.256
Donor 2 0 0.02927 0.0293 0.0289 .+-. 0.00230 0.261 0.258 .+-.
0.0205 100 (VC) 0.02920 0.0292 0.261 0.03137 0.0314 0.280 0.02582
0.0258 0.231 0.4 0.02544 0.0254 0.0306 .+-. 0.00559 0.227 0.273
.+-. 0.0499 106 0.02986 0.0299 0.267 0.03654 0.0365 0.326 4 0.02852
0.0285 0.0281 .+-. 0.000884 0.255 0.250 .+-. 0.00790 97.0 0.02703
0.0270 0.241 0.02860 0.0286 0.255 40 0.00341.sup.a <0.0250
<0.0250 .+-. 0.000 <0.223 <0.223 .+-. 0.000 <86.5
0.00320.sup.a <0.0250 <0.223 0.00330.sup.a <0.0250
<0.223 Donor 3 0 0.02349.sup.a <0.0250 <0.0252 .+-.
0.000435 <0.223 <0.225 .+-. 0.00388 100 (VC) 0.02587 0.0259
0.231 0.02376.sup.a <0.0250 <0.223 0.02236.sup.a <0.0250
<0.223 0.4 0.02177.sup.a <0.0250 <0.0250 .+-. 0.000
<0.223 <0.223 .+-. 0.000 99.1 0.02343.sup.a <0.0250
<0.223 0.02326.sup.a <0.0250 <0.223 4 0.02392.sup.a
<0.0250 <0.0250 .+-. 0.000 <0.223 <0.223 .+-. 0.000
99.1 0.02490.sup.a <0.0250 <0.223 0.02229.sup.a <0.0250
<0.223 40 0.02005.sup.a <0.0250 <0.0250 .+-. 0.000
<0.223 <0.223 .+-. 0.000 99.1 0.01976.sup.a <0.0250
<0.223 0.02169.sup.a <0.0250 <0.223 Abbreviations: SD,
standard deviation; VC, vehicle control (1% Methanol) .sup.aThe
observed analyzed value (.mu.M) was below the lowest concentration
on the standard curve (0.025 .mu.M). Note: For all calculations
above, the resulting values are shown with at least three
significant figures for display purposes only.
TABLE-US-00024 TABLE 21 CYP2C9 Activity in Cryopreserved Human
Hepatocytes 4'-Methylhydroxytolbutamide formation Specific Activity
Adjusted (.mu.M) (pmol/min/million cells) Percent Metaxalone
(.mu.M) Raw (.mu.M) Individual Mean .+-. SD Individual Mean .+-. SD
of VC Donor 1 0 0.01215 0.0122 0.0137 .+-. 0.00161 0.108 0.122 .+-.
0.0144 100 (VC) 0.01502 0.0150 0.134 0.01513 0.0151 0.135 0.01245
0.0125 0.111 0.4 0.01557 0.0156 0.0147 .+-. 0.000753 0.139 0.132
.+-. 0.00672 108 0.01410 0.0141 0.126 0.01455 0.0146 0.130 4
0.01331 0.0133 0.0137 .+-. 0.00136 0.119 0.122 .+-. 0.0121 100
0.01523 0.0152 0.136 0.01261 0.0126 0.113 40 0.00931.sup.a
<0.0100 <0.0100 .+-. 0.0000346 <0.0893 <0.0895 .+-.
0.000309 <73.2 0.00952.sup.a <0.0100 <0.0893 0.01006
0.0101 0.0898 Donor 2 0 0.05192 0.0519 0.0491 .+-. 0.00479 0.464
0.438 .+-. 0.0428 100 (VC) 0.04864 0.0486 0.434 0.05325 0.0533
0.475 0.04250 0.0425 0.379 0.4 0.04819 0.0482 0.0474 .+-. 0.00223
0.430 0.423 .+-. 0.0200 96.6 0.04489 0.0449 0.401 0.04915 0.0492
0.439 4 0.04634 0.0463 0.0456 .+-. 0.000864 0.414 0.407 .+-.
0.00772 92.9 0.04581 0.0458 0.409 0.04465 0.0447 0.399 40 0.02917
0.0292 0.0296 .+-. 0.000651 0.260 0.265 .+-. 0.00581 60.4 0.02936
0.0294 0.262 0.03038 0.0304 0.271 Donor 3 0 0.02021 0.0202 0.0181
.+-. 0.00206 0.180 0.162 .+-. 0.0184 100 (VC) 0.01700 0.0170 0.152
0.01952 0.0195 0.174 0.01586 0.0159 0.142 0.4 0.02067 0.0207 0.0201
.+-. 0.00125 0.185 0.179 .+-. 0.0111 111 0.02096 0.0210 0.187
0.01867 0.0187 0.167 4 0.01807 0.0181 0.0187 .+-. 0.00235 0.161
0.167 .+-. 0.0210 103 0.02129 0.0213 0.190 0.01671 0.0167 0.149 40
0.01364 0.0136 0.0142 .+-. 0.000560 0.122 0.127 .+-. 0.00500 78.4
0.01432 0.0143 0.128 0.01475 0.0148 0.132 Abbreviations: SD,
standard deviation; VC, vehicle control (1% Methanol) .sup.aThe
observed analyzed value (.mu.M) was below the lowest concentration
on the standard curve (0.01 .mu.M). Note: For all calculations
above, the resulting values are shown with at least three
significant figures for display purposes only.
TABLE-US-00025 TABLE 22 CYP2C19 Activity in Cryopreserved Human
Hepatocytes 4'-Hydroxymephenytoin formation Specific Activity
Adjusted (.mu.M) (pmol/min/million cells) Metaxalone (.mu.M) Raw
(.mu.M) Individual Mean .+-. SD Individual Mean .+-. SD Percent of
VC Donor 1 0 0.00025.sup.a <0.0500 <0.0500 .+-. 0.000
<0.446 <0.446 .+-. 0.000 100 (VC) 0.00058.sup.a <0.0500
<0.446 0.00114.sup.a <0.0500 <0.446 0.00058.sup.a
<0.0500 <0.446 0.4 0.00708.sup.a <0.0500 <0.0500 .+-.
0.000 <0.446 <0.446 .+-. 0.000 100 0.01319.sup.a <0.0500
<0.446 0.01861.sup.a <0.0500 <0.446 4 0.01649.sup.a
<0.0500 <0.0500 .+-. 0.000 <0.446 <0.446 .+-. 0.000 100
0.00029.sup.a <0.0500 <0.446 0.00064.sup.a <0.0500
<0.446 40 0.00057.sup.a <0.0500 <0.0500 .+-. 0.000
<0.446 <0.446 .+-. 0.000 100 0.00031.sup.a <0.0500
<0.446 0.00037.sup.a <0.0500 <0.446 Donor 2 0 N/A* N/A
<0.0500 .+-. 0.000 N/A <0.446 .+-. 0.000 100 (VC)
0.01146.sup.a <0.0500 <0.446 0.01456.sup.a <0.0500
<0.446 N/A* N/A N/A 0.4 0.00765.sup.a <0.0500 <0.0500 .+-.
0.000 <0.446 <0.446 .+-. 0.000 100 0.00779.sup.a <0.0500
<0.446 0.00808.sup.a <0.0500 <0.446 4 0.00775.sup.a
<0.0500 <0.0500 .+-. 0.000 <0.446 <0.446 .+-. 0.000 100
0.00744.sup.a <0.0500 <0.446 0.00773.sup.a <0.0500
<0.446 40 0.00697.sup.a <0.0500 <0.0500 .+-. 0.000
<0.446 <0.446 .+-. 0.000 100 0.00840.sup.a <0.0500
<0.446 0.00790.sup.a <0.0500 <0.446 Donor 3 0
0.00026.sup.a <0.0500 <0.0500 .+-. 0.000 <0.446 <0.446
.+-. 0.000 100 (VC) 0.00000.sup.a <0.0500 <0.446
0.00000.sup.a <0.0500 <0.446 0.00000.sup.a <0.0500
<0.446 0.4 0.00000.sup.a <0.0500 <0.0500 .+-. 0.000
<0.446 <0.446 .+-. 0.000 100 0.00000.sup.a <0.0500
<0.446 0.00023.sup.a <0.0500 <0.446 4 0.00000.sup.a
<0.0500 <0.0500 .+-. 0.000 <0.446 <0.446 .+-. 0.000 100
0.00000.sup.a <0.0500 <0.446 0.00000.sup.a <0.0500
<0.446 40 0.00191.sup.a <0.0500 <0.0500 .+-. 0.000
<0.446 <0.446 .+-. 0.000 100 0.00000.sup.a <0.0500
<0.446 0.00000.sup.a <0.0500 <0.446 Abbreviations: SD,
standard deviation; VC, vehicle control (1% Methanol) .sup.aThe
observed analyzed value (.mu.M) was below the lowest concentration
on the standard curve (0.05 .mu.M). *Sample lost after preparation.
Note: For all calculations above, the resulting values are shown
with at least three significant figures for display purposes
only.
TABLE-US-00026 TABLE 23 CYP2D6 Activity in Cryopreserved Human
Hepatocytes Dextrorphan formation Specific Activity Adjusted
(.mu.M) (pmol/min/million cells) Metaxalone (.mu.M) Raw (.mu.M)
Individual Mean .+-. SD Individual Mean .+-. SD Percent of VC Donor
1 0 0.00772.sup.a <0.0100 <0.0100 .+-. 0.000 <0.0893
<0.0893 .+-. 0.000 100 (VC) 0.00796.sup.a <0.0100 <0.0893
0.00736.sup.a <0.0100 <0.0893 0.00724.sup.a <0.0100
<0.0893 0.4 0.00809.sup.a <0.0100 <0.0100 .+-. 0.000
<0.0893 <0.0893 .+-. 0.000 100 0.00749.sup.a <0.0100
<0.0893 0.00853.sup.a <0.0100 <0.0893 4 0.00832.sup.a
<0.0100 <0.0100 .+-. 0.000 <0.0893 <0.0893 .+-. 0.000
100 0.00721.sup.a <0.0100 <0.0893 0.00744.sup.a <0.0100
<0.0893 40 0.00398.sup.a <0.0100 <0.0100 .+-. 0.000
<0.0893 <0.0893 .+-. 0.000 100 0.00205.sup.a <0.0100
<0.0893 0.00520.sup.a <0.0100 <0.0893 Donor 2 0 0.01286
0.0129 0.0139 .+-. 0.00152 0.115 0.124 .+-. 0.0136 100 (VC) 0.01432
0.0143 0.128 0.01581 0.0158 0.141 0.01247 0.0125 0.111 0.4 0.01302
0.0130 0.0133 .+-. 0.000485 0.116 0.119 .+-. 0.00433 95.9 0.01302
0.0130 0.116 0.01386 0.0139 0.124 4 0.01361 0.0136 0.0143 .+-.
0.000589 0.122 0.128 .+-. 0.00526 103 0.01468 0.0147 0.131 0.01457
0.0146 0.130 40 0.00998.sup.a <0.0100 <0.0102 .+-. 0.000260
<0.0893 <0.0906 .+-. 0.00232 <73.2 0.00956.sup.a
<0.0100 <0.0893 0.01045 0.0105 0.0933 Donor 3 0 0.07011
0.0701 0.0665 .+-. 0.00607 0.626 0.594 .+-. 0.0542 100 (VC) 0.05856
0.0586 0.523 0.07219 0.0722 0.645 0.06505 0.0651 0.581 0.4 0.06218
0.0622 0.0657 .+-. 0.00305 0.555 0.586 .+-. 0.0272 98.8 0.06688
0.0669 0.597 0.06789 0.0679 0.606 4 0.06071 0.0607 0.0597 .+-.
0.00164 0.542 0.533 .+-. 0.0146 89.8 0.06060 0.0606 0.541 0.05782
0.0578 0.516 40 0.05087 0.0509 0.0489 .+-. 0.00347 0.454 0.436 .+-.
0.0310 73.5 0.05088 0.0509 0.454 0.04486 0.0449 0.401
Abbreviations: SD, standard deviation; VC, vehicle control (1%
Methanol) .sup.aThe observed analyzed value (.mu.M) was below the
lowest concentration on the standard curve (0.01 .mu.M). Note: For
all calculations above, the resulting values are shown with at
least three significant figures for display purposes only.
TABLE-US-00027 TABLE 24 CYP2E1 Activity in Cryopreserved Human
Hepatocytes 6-Hydroxychlorzoxazone formation Specific Activity
Adjusted (.mu.M) (pmol/min/million cells) Metaxalone (.mu.M) Raw
(.mu.M) Individual Mean .+-. SD Individual Mean .+-. SD Percent of
VC Donor 1 0 0.28067 0.281 0.283 .+-. 0.00460 2.51 2.53 .+-. 0.0411
100 (VC) 0.28793 0.288 2.57 0.28627 0.286 2.56 0.27817 0.278 2.48
0.4 0.28854 0.289 0.277 .+-. 0.0279 2.58 2.47 .+-. 0.249 97.8
0.29749 0.297 2.66 0.24529 0.245 2.19 4 0.28784 0.288 0.295 .+-.
0.0236 2.57 2.64 .+-. 0.210 104 0.27623 0.276 2.47 0.32160 0.322
2.87 40 0.28453 0.285 0.294 .+-. 0.00876 2.54 2.63 .+-. 0.0782 104
0.29753 0.298 2.66 0.30121 0.301 2.69 Donor 2 0 0.07385 0.0739
0.0748 .+-. 0.00211 0.659 0.668 .+-. 0.0188 100 (VC) 0.07610 0.0761
0.679 0.07690 0.0769 0.687 0.07229 0.0723 0.645 0.4 0.07071 0.0707
0.0776 .+-. 0.00753 0.631 0.693 .+-. 0.0673 104 0.07649 0.0765
0.683 0.08565 0.0857 0.765 4 0.06315 0.0632 0.0670 .+-. 0.00355
0.564 0.598 .+-. 0.0317 89.6 0.06775 0.0678 0.605 0.07013 0.0701
0.626 40 0.06247 0.0625 0.0745 .+-. 0.0141 0.558 0.665 .+-. 0.126
99.6 0.07091 0.0709 0.633 0.09003 0.0900 0.804 Donor 3 0 0.05899
0.0590 0.0570 .+-. 0.00420 0.527 0.509 .+-. 0.0375 100 (VC) 0.06077
0.0608 0.543 0.05718 0.0572 0.511 0.05110 0.0511 0.456 0.4 0.05031
0.0503 0.0517 .+-. 0.00140 0.449 0.462 .+-. 0.0125 90.7 0.05310
0.0531 0.474 0.05169 0.0517 0.462 4 0.05245 0.0525 0.0500 .+-.
0.00389 0.468 0.446 .+-. 0.0348 87.7 0.05202 0.0520 0.464 0.04550
0.0455 0.406 40 0.05260 0.0526 0.0535 .+-. 0.00164 0.470 0.478 .+-.
0.0146 93.9 0.05541 0.0554 0.495 0.05254 0.0525 0.469
Abbreviations: SD, standard deviation; VC, vehicle control (1%
Methanol) Note: For all calculations above, the resulting values
are shown with at least three significant figures for display
purposes only.
TABLE-US-00028 TABLE 25 CYP3A4 Activity in Cryopreserved Human
Hepatocytes 6.beta.-Hydroxytestosterone formation Specific Activity
Adjusted (.mu.M) (pmol/min/million cells) Metaxalone (.mu.M) Raw
(.mu.M) Individual Mean .+-. SD Individual Mean .+-. SD Percent of
VC Donor 1 0 0.05693.sup.a <0.100 <0.100 .+-. 0.000 <0.893
<0.893 .+-. 0.000 100 (VC) 0.05726.sup.a <0.100 <0.893
0.05367.sup.a <0.100 <0.893 0.04590.sup.a <0.100 <0.893
0.4 0.05415.sup.a <0.100 <0.100 .+-. 0.000 <0.893
<0.893 .+-. 0.000 100 0.06053.sup.a <0.100 <0.893
0.05911.sup.a <0.100 <0.893 4 0.05783.sup.a <0.100
<0.100 .+-. 0.000 <0.893 <0.893 .+-. 0.000 100
0.05948.sup.a <0.100 <0.893 0.05705.sup.a <0.100 <0.893
40 0.06888.sup.a <0.100 <0.100 .+-. 0.000 <0.893 <0.893
.+-. 0.000 100 0.06424.sup.a <0.100 <0.893 0.06511.sup.a
<0.100 <0.893 Donor 2 0 0.12401 0.124 <0.117 .+-. 0.0141
1.11 <1.05 .+-. 0.126 100 (VC) 0.13222 0.132 1.18 0.07973.sup.a
<0.100 <0.893 0.11219 0.112 1.00 0.4 0.12083 0.121 0.134 .+-.
0.0122 1.08 1.20 .+-. 0.109 >115 0.14424 0.144 1.29 0.13828
0.138 1.23 4 0.10953 0.110 0.116 .+-. 0.00524 0.978 1.03 .+-.
0.0468 >98.7 0.11883 0.119 1.06 0.11837 0.118 1.06 40 0.14198
0.142 0.141 .+-. 0.00273 1.27 1.26 .+-. 0.0244 >121 0.14356
0.144 1.28 0.13824 0.138 1.23 Donor 3 0 0.06064.sup.a <0.100
<0.100 .+-. 0.000 <0.893 <0.893 .+-. 0.000 100 (VC)
0.05981.sup.a <0.100 <0.893 0.06402.sup.a <0.100 <0.893
0.08660.sup.a <0.100 <0.893 0.4 0.05106.sup.a <0.100
<0.100 .+-. 0.000 <0.893 <0.893 .+-. 0.000 100
0.08255.sup.a <0.100 <0.893 0.05998.sup.a <0.100 <0.893
4 0.06298.sup.a <0.100 <0.100 .+-. 0.000 <0.893 <0.893
.+-. 0.000 100 0.05381.sup.a <0.100 <0.893 0.07264.sup.a
<0.100 <0.893 40 0.05587.sup.a <0.100 <0.101 .+-.
0.00238 <0.893 <0.905 .+-. 0.0213 101 0.10413 0.104 0.930
0.08088.sup.a <0.100 <0.893 Abbreviations: SD, standard
deviation; VC, vehicle control (1% Methanol); .sup.aThe observed
analyzed value (.mu.M) was below the lowest concentration on the
standard curve (0.1 .mu.M). Note: For all calculations above, the
resulting values are shown with at least three significant figures
for display purposes only.
[0254] Table 18 presents the results for CYP1A2. Under these
experimental conditions, exposure to metaxalone at 40 .mu.M induced
CYP1A2 activity in human hepatocytes prepared from Donors 1, 2, and
3. For each of the three donors, the increases in CYP1A2 activity
by metaxalone at 0.4 and 4 .mu.M were not statistically significant
(p>0.05; unpaired two-tailed t test).
[0255] Table 25 presents the results for CYP3A4. Metaxalone at the
concentration of 40 M induced CYP3A4 activity by about 21% in one
of three donors tested, Donor 2. Therefore under these experimental
conditions, exposure to metaxalone at 40 .mu.M induced CYP3A4
activity in human hepatocytes prepared from Donor 2. The increase
in CYP3A4 activity following treatment with metaxalone at 0.4 .mu.M
for Donor 2 was not statistically significant (p>0.05; unpaired
two-tailed t test). CYP3A4 activity in the vehicle controls for
Donor 1 and Donor 3 were below the lower limit of quantitation.
Exposure of hepatocytes from Donors 1 and 3 to metaxalone at the
concentrations tested did not induce CYP3A4 activity since the
activity following treatment with metaxalone was still below the
lower limit of quantitation at each tested concentration.
[0256] Table 21 presents the results for CYP2C9. Under these
experimental conditions, exposure to metaxalone at 40 .mu.M
significantly reduced CYP2C9 activity in human hepatocytes prepared
from Donors 1, 2, and 3. The observed changes in CYP2C9 activity
following exposure to metaxalone at 0.4 and 4 .mu.M were not
statistically significant (p>0.05; two-tailed t test). Thus,
under these experimental conditions, exposure to metaxalone at 40
.mu.M inhibited CYP2C9 activity.
[0257] Table 23 presents the results for CYP2D6. CYP2D6 activity
was below the lower limit of quantitation in the vehicle controls
and for the metaxalone-exposed samples for Donor 1. However, under
these experimental conditions, exposure to metaxalone at 40 .mu.M
significantly reduced CYP2D6 activity in human hepatocytes prepared
from Donors 2 and 3. The observed changes in CYP2D6 activity
following exposure to metaxalone at 0.4 and 4 .mu.M were not
statistically significant (p>0.05; two-tailed t test). Thus,
under these experimental conditions, exposure to metaxalone at 40
.mu.M inhibited CYP2D6 activity.
EXAMPLE 4
Metabolic Phenotyping of Metaxalone
[0258] A follow-up metabolic phenotyping study was performed in
light of the study of Example 1 to clarify the metabolic profile of
metaxalone and the relative contribution of specific CYP isozymes
to its overall metabolic clearance. Two complementary methodologies
to determine metabolism of metaxalone were performed to corroborate
results: determination of metabolism in pooled human liver
microsomes in the presence of isozyme-selective enzyme inhibitors
and a repeated determination of metabolism by singly expressed
human recombinant CYP isozymes. The microsomal system approximates
the in vivo distribution of hepatic enzymes. When the test compound
is metabolized by multiple CYP isozymes, selective chemical
inhibitors are used in human hepatic microsomes to identify the
relative contribution of each isozyme. When combined with knowledge
of the relative abundance of each P450 isoform in the pool of human
liver microsomes being used, the relative contribution of different
P450 enzymes in the metabolism of the compound of interest in human
liver microsomes can be better evaluated
[0259] Metaxalone (MW=221.26, Lot #P-1055-116A1, purity=99.8%)
stock solutions at 1.875 mM were prepared in methanol and stored at
-20.degree. C. Stocks were diluted daily in the appropriate buffers
such that the final organic solvent concentration was <1%.
[0260] Potassium phosphate monobasic, potassium phosphate dibasic,
NADPH tetrasodium salts, and other reagents were purchased from
Sigma Chemical Co. or equivalent vendors. Methanol (HPLC grade),
water (HPLC grade), ethyl acetate, and other solvents were
purchased from Fisher, Burdick & Jackson, J. T. Baker,
Mallinckrodt, or equivalent vendors. All inhibitors were of the
highest purity available. Furafylline, pilocarpine, thio-TEPA,
quercetin, sulfaphenazole, ticlopidine, quinidine, clomethiazole,
and ketoconazole were obtained from Sigma Chemical Co.
[0261] Human liver microsomes pooled from 15 individuals (male and
female) (Pool HMMC-PL020; CellzDirect, Inc., In Vitro Products and
Services Division) were used in some experiments of this study.
This pool of human liver microsomes was characterized by
CellzDirect, Inc with respect to donor medical history, major
cytochrome P450 enzyme activities and kinetic parameters, as well
as for polymorphic forms for CYP2C9, CYP2C19, and CYP2D6 present in
the individual donors of the pool.
[0262] In these follow-up studies, metaxalone concentrations were
chosen to be 0.75 and 7.5 .mu.M, approximately 1.times. and
0.1.times. of C.sub.max based on the estimated clinical plasma
concentrations of metaxalone provided on the August 2003 version of
the package insert for SKELAXIN.RTM..
[0263] An isocratic LC-MS/MS was developed to allow for
chromatographic resolution and quantitation of metaxalone contained
within an incubation matrix for this concentration range. The
following LC-MS/MS method was used for metaxalone quantitation:
TABLE-US-00029 TABLE 26 LC/MS method for metaxalone quantitation.
Metaxalone Substrate:Metabolite: Metaxalone Standard Metabolite
0.078 to 10.0 .mu.M Range: Mobile Phase A 2% Methanol, 98% Water,
and 0.1% Acetic Acid Mobile Phase B 80% Methanol, 20% Water, and
0.1% Acetic Acid Gradient 0.400 mL/min, initial: 50% Mobile Phase
B, 0.01 min: 50% Mobile Phase B, 2.00 min: 80% Mobile Phase B, 2.10
min: 50% Mobile Phase B, and 4.00 min: Stop Detection Method:
LC-MS/MS HPLC Column: WATERS AQ12 4 .times. 23 mm Flow rate
(approx.): 0.400 mL/min Source: Electrospray (positive ion) Run
Time (approx.): 4.0 minutes MRM 222.14 .fwdarw. 160.83
(Sulfamethoxazole): Quantitation: Least Squares Regression
1/X.sup.2 Weighting
[0264] Micromass MASSLYNX.RTM. software (version 3.4, Manchester,
UK) was used to collect and process chromatographic data. Data were
graphed and analyzed using the software program Microsoft
EXCEL.RTM. 2003 (Redmond, Wash.). Percent turnover of metaxalone
was calculated using the following equation:
% Turnover=100-{[TA(final)]/[mean of TA(0 min)]}.times.100,
where TA(sample group)=Test Article (i.e., metaxalone) and the
particular sample group is noted within the parentheses. Percent
inhibition of turnover of metaxalone was calculated using the
following equation:
% Inhibition of Turnover=100{1-([TA(0
min)]-[TA(final-inhibitor)])/([TA(0 min)]-[TA(final-no
inhibitor)])}.
[0265] Pilot human liver microsomal incubations were performed in
order to establish appropriate protein concentration and time
points for linear reaction conditions for metaxalone turnover.
Metaxalone at 0.75 or 7.5 .mu.M was incubated with 0.1, 0.25, 0.5,
and 1.0 mg/mL microsomal protein for 60 minutes at 37.degree. C.
Also, incubations of metaxalone at 0.75 or 7.5 .mu.M with 0.5 mg/mL
microsomal protein were run for 0, 15, 30, 45, and 60 minutes at
37.degree. C. The reactions, in 0.1 M phosphate buffer, pH 7.4,
were initiated by addition of 1 mM NADPH. Negative controls (no
NADPH and heat-treated microsomes) were included to account for any
non-enzymatic or non-NADPH dependent reactions. The reactions were
terminated at the appropriate time points by addition of 1 volume
(relative to total reaction volume) of methanol. The samples were
centrifuged at approximately 3000 rpm and the clear supernatant was
transferred to a clean tube and analyzed by the LC-MS/MS method
described above.
[0266] Linear disappearance of metaxalone from the pooled human
liver microsomal incubations was observed for both the time- and
microsomal protein-dependence incubation series. A maximum turnover
of 38.0% was determined at 60 minutes with 1 mg/mL protein. This
turnover was NADPH-dependent, indicating that disappearance of
metaxalone under these conditions was due to CYP450 enzymatic
activity. Based on these results, all subsequent assays used a
microsomal protein concentration of 1 mg/mL with an incubation time
of 60 minutes to maximize the dynamic range of metaxalone
turnover.
[0267] Selective chemical inhibitors were then used to evaluate the
effects of individual CYP450s in human liver microsomes on the
metabolism of metaxalone. The CYP isozyme-selective chemical
inhibitors and concentrations used are shown in Table 27 below.
TABLE-US-00030 TABLE 27 Selective Chemical Inhibitors and
Concentrations Used in Protocols for in vitro Assessment of
Chemical Inhibition in Human Liver Microsomes. P450 Isozyme
Selective chemical inhibitor Concentration CYP1A2 Furafylline 50
.mu.M CYP2A6 Pilocarpine 100 .mu.M CYP2B6 Thio-TEPA 75 .mu.M CYP2C8
Quercetin 10 .mu.M CYP2C9 Sulfaphenazole 20 .mu.M CYP2C19
Ticlopidine 1 .mu.M CYP2D6 Quinidine 10 .mu.M CYP2E1 Clomethiazole
100 .mu.M CYP3A4 Ketoconazole 1 .mu.M
[0268] The inhibitors or vehicle control were incubated with the
pooled human liver microsomes (1 mg/mL) and NADPH (1 mM) in 0.1 M
phosphate buffer, pH 7.4. The reaction was initiated by addition of
0.75 or 7.5 .mu.M metaxalone and incubated for 60 minutes.
Incubations were conducted in triplicate and terminated by addition
of organic solvent. Samples were extracted and analyzed by LC-MS/MS
as described above. The rates of metaxalone depletion were compared
to controls without inhibitors.
[0269] The data from these studies are provided in Tables 28 and
29, below.
TABLE-US-00031 TABLE 28 Chemical Inhibitor Data in Human Liver
Microsomes with 0.75 .mu.M Metaxalone % inhibition Mean of turnover
Conc. Conc. % % Turnover % inhibition Std Inhibitor CYP Sample ID
(.mu.M) (.mu.M) Turnover Mean Std. Error of turnover Mean Error
0.75uM-T0-P1.0-1 0.730 0.75uM-T0-P1.0-2 0.815 0 min.
0.75uM-T0-P1.0-3 0.770 0.772 0.75uM-T60-P1.0-1 0.536 30.5
0.75uM-T60-P1.0-2 0.561 27.3 60 min. 0.75uM-T60-P1.0-3 0.529 0.542
31.5 29.8 1.3 0.75uM-T60-P1.0-F-1 0.924 -19.7 166
0.75uM-T60-P1.0-F-2 0.767 0.60 98.0 Furafylline CYP1A2
0.75uM-T60-P1.0-F-3 0.701 0.797 9.2 -3.29 8.57 69.1 111 29
0.75uM-T60-P1.0-P-1 0.664 13.9 53.2 0.75uM-T60-P1.0-P-2 0.658 14.7
50.6 Pilocarpine CYP2A6 0.75uM-T60-P1.0-P-3 0.622 0.648 19.4 16.0
1.7 34.9 46.2 5.7 0.75uM-T60-P1.0-TT-1 0.678 12.1 59.3
0.75uM-T60-P1.0-TT-2 0.684 11.3 62.0 Thio-TEPA CYP2B6
0.75uM-T60-P1.0-TT-3 0.665 0.676 13.8 12.4 0.7 53.7 58.3 2.4
0.75uM-T60-P1.0-Qr-1 0.661 14.3 51.9 0.75uM-T60-P1.0-Qr-2 0.769
0.39 98.7 Quercetin CYP2C8 0.75uM-T60-P1.0-Qr-3 0.728 0.719 5.6
6.78 4.06 81.1 77.2 13.6 0.75uM-T60-P1.0-S-1 0.730 5.4 81.9
0.75uM-T60-P1.0-S-2 0.728 5.6 81.1 Sulfaphenazole CYP2C9
0.75uM-T60-P1.0-S-3 0.729 0.729 5.6 5.53 0.08 81.3 81.4 0.3
0.75uM-T60-P1.0-Ti-1 0.698 9.6 67.9 0.75uM-T60-P1.0-Ti-2 0.714 7.5
74.7 Ticlopidine CYP2C19 0.75uM-T60-P1.0-Ti-3 0.715 0.709 7.4 8.16
0.70 75.2 72.6 2.3 0.75uM-T60-P1.0-Qi-1 0.694 10.0 66.4
0.75uM-T60-P1.0-Qi-2 0.920 -19.2 165 Quinidine CYP2D6
0.75uM-T60-P1.0-Qi-3 0.692 0.769 10.3 0.376 9.796 65.4 98.7 32.9
0.75uM-T60-P1.0-Clo-1 0.780 -1.1 104 0.75uM-T60-P1.0-Clo-2 0.896
-16.1 154 Clomethiazole CYP2E1 0.75uM-T60-P1.0-Clo-3 0.729 0.802
5.6 -3.87 6.39 81.4 113 21 0.75uM-T60-P1.0-K-1 0.844 -9.4 132
0.75uM-T60-P1.0-K-2 0.807 -4.6 116 Ketoconazole CYP3A4
0.75uM-T60-P1.0-K-3 0.842 0.831 -9.1 -7.71 1.54 130 126 5
TABLE-US-00032 TABLE 29 Chemical Inhibitor Data in Human Liver
Microsomes with 7.5 .mu.M Metaxalone % Turnover Conc. Mean % Std. %
inhibition Std Inhibitor CYP Sample ID (.mu.M) Conc. (.mu.M)
Turnover Mean Error of turnover Mean Error 7.5uM-T0-P1.0-1 8.34
7.5uM-T0-P1.0-2 8.55 0 min. 7.5uM-T0-P1.0-3 8.65 8.51
7.5uM-T60-P1.0-1 6.40 24.8 7.5uM-T60-P1.0-2 6.48 23.9 60 min.
7.5uM-T60-P1.0-3 6.38 6.42 25.1 24.6 0.3 7.5uM-T60-P1.0-F-1 8.07
5.2 78.8 7.5uM-T60-P1.0-F-2 8.21 3.6 85.4 Furafylline CYP1A2
7.5uM-T60-P1.0-F-3 8.05 8.11 5.5 4.77 0.59 77.7 80.6 2.4
7.5uM-T60-P1.0-P-1 7.76 8.8 64.2 7.5uM-T60-P1.0-P-2 7.28 14.5 41.2
Pilocarpine CYP2A6 7.5uM-T60-P1.0-P-3 7.41 7.48 13.0 12.1 1.7 47.1
50.8 6.9 7.5uM-T60-P1.0-TT-1 7.86 7.7 68.7 7.5uM-T60-P1.0-TT-2 7.26
14.8 40.0 Thio-TEPA CYP2B6 7.5uM-T60-P1.0-TT-3 6.86 7.33 19.4 14.0
3.4 21.1 43.3 13.8 7.5uM-T60-P1.0-Qr-1 8.67 -1.8 107
7.5uM-T60-P1.0-Qr-2 8.48 0.45 98.2 Quercetin CYP2C8
7.5uM-T60-P1.0-Qr-3 7.43 8.19 12.7 3.77 4.50 48.5 84.7 18.3
7.5uM-T60-P1.0-S-1 7.49 12.1 51.0 7.5uM-T60-P1.0-S-2 7.65 10.2 58.5
Sulfaphenazole CYP2C9 7.5uM-T60-P1.0-S-3 7.44 7.52 12.6 11.6 0.7
48.8 52.8 2.9 7.5uM-T60-P1.0-Ti-1 8.02 5.8 76.5 7.5uM-T60-P1.0-Ti-2
7.58 11.0 55.5 Ticlopidine CYP2C19 7.5uM-T60-P1.0-Ti-3 9.50 8.37
-11.6 1.71 6.83 147 93.1 27.7 7.5uM-T60-P1.0-Qi-1 7.62 10.5 57.5
7.5uM-T60-P1.0-Qi-2 7.80 8.4 65.9 Quinidine CYP2D6
7.5uM-T60-P1.0-Qi-3 7.57 7.66 11.1 10.0 0.8 55.1 59.5 3.3
7.5uM-T60-P1.0-Clo-1 8.29 2.7 89.2 7.5uM-T60-P1.0-Clo-2 8.54 -0.25
101 Clomethiazole CYP2E1 7.5uM-T60-P1.0-Clo-3 8.63 8.48 -1.3 0.356
1.197 105 98.6 4.9 7.5uM-T60-P1.0-K-1 8.49 0.28 98.8
7.5uM-T60-P1.0-K-2 9.78 -14.9 161 Ketoconazole CYP3A4
7.5uM-T60-P1.0-K-3 9.12 9.13 -7.1 -7.26 4.39 129 130 18
[0270] Mean metaxalone disappearance after 60 minutes of incubation
with 1 mg/mL microsomal protein was observed to be 24.6.+-.0.3% at
7.5 .mu.M (Table 29) vs. 29.8.+-.1.3% at 0.75 .mu.M (Table 28) at
the conditions examined.
[0271] Inhibition of metaxalone disappearance in human liver
microsomes at either 0.75 or 7.5 .mu.M metaxalone was observed upon
addition of the CYP1A2 selective inhibitor furafylline (111.+-.29%
or 80.6.+-.2.4%), the CYP3A4 inhibitor ketoconazole (126.+-.5% or
130.+-.18%), the CYP2E1 inhibitor clomethiazole (113.+-.21% or
98.6.+-.4.9%), the CYP2C19 inhibitor ticlopidine (72.6.+-.2.3% or
93.11.+-.27.7%), the CYP2D6 inhibitor quinidine (98.7.+-.32.9% or
59.5.+-.4.9%), the CYP2C9 inhibitor sulfaphenazole (81.4.+-.0.3% or
52.8.+-.2.9%), and the CYP2C8 inhibitor quercetin (77.2.+-.13.6% or
84.7.+-.18.3%) (See Tables 28-29). Smaller inhibition of turnover
was also observed upon addition of the CYP2A6 inhibitor pilocarpine
(46.2.+-.5.7% or 50.8.+-.6.9%) or the CYP2B6 inhibitor thio-TEPA
(58.3.+-.2.4% or 43.3.+-.13.8%), however the observed inhibition of
metaxalone metabolism by these two inhibitors in the pooled human
liver microsomal system was not supported by the singly expressed
recombinant CYP isozyme data.
[0272] Overall, this study in pooled human liver microsomes
indicates that metaxalone can be metabolized to some extent by each
of CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, and CYP3A4,
with CYP1A2, CYP2E1, and CYP3A4 appearing to be the major CYP450
enzymes involved.
[0273] An additional set of experiments using singly-expressed
recombinant human CYP enzymes were conducted to corroborate which
specific CYP450s are capable of metabolizing colchicines.
[0274] Commercially available microsomes from baculovirus-infected
insect cells containing singly-expressed recombinant human CYP
enzymes and cDNA-expressed human cytochrome p450 oxidoreductase [BD
SUPERSOMES.RTM. Enzymes; BD Biosciences Discovery Labware (Woburn,
Mass.)] were used. For CYP2A6, CYP2C9, CYP2C19, and CYP2E1, the
SUPERSOMES.RTM. also expressed human cytochrome b5 in addition to
human cytochrome p450 oxidoreductase and the human CYP isozyme. For
CYP2C9 and CYP2D6, SUPERSOMES singly-expressing different allelic
variants of the cytochrome p450 isozyme were commercially
available. For each of CYP2C9 and CYP2D6, only the *1 allele was
tested in these experiments.
[0275] Incubation mixtures containing CYP1A2, CYP2A6, CYP2B6,
CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, and CYP3A4
singly-expressed enzymes at 5 and 20 pmol cytochrome P450 were
incubated in 0.1 M phosphate buffer, pH 7.4 with 1 mM NADPH and
0.75 or 7.5 .mu.M metaxalone for 60 minutes at 37.degree. C. The
incubations, performed in triplicate, were terminated by addition
of 1 volume (relative to total reaction volume) of methanol. The
samples were extracted and analyzed by LC-MS/MS, as described
above. The amount of metaxalone depletion by the CYP were compared
to the amount of depletion in a control for native activity
(microsomes expressing no recombinant CYP450 enzyme).
[0276] An additional control for each isoform included verification
that the isoform was active by incubating mixtures with the
universal CYP substrate, phenanthrene, and monitoring turnover of
phenanthrene fluorometrically at 254 nm (excitation) and 378 nm
(emission). These controls confirmed that each recombinant enzyme
microsomal preparation was active (data not shown).
[0277] The results measuring metaxalone disappearance at either
0.75 or 7.5 .mu.M metaxalone in incubations containing either 5 or
20 pmol of a recombinant cytochrome P450 isozyme is summarized
below in Tables 30-32.
TABLE-US-00033 TABLE 30 Metabolism of Metaxalone (0.75 .mu.M) by
Expressed Recombinant Human Cytochromes P450 at 5 or 20 pmol 5 pmol
cyp 20 pmol cyp [metaxalone] % Turnover [metaxalone] % Turnover
.mu.M Mean Std dev p % Mean Std Error .mu.M Mean Std dev p % Mean
Std Error control 0.786 0.786 control 0.802 0.802 control 0.785
0.791 0.10 0.785 0.791 0.10 1A2 0.653 17.5 0.540 31.7 1A2 0.742 6.2
0.511 35.5 1A2 0.761 0.719 0.06 0.099 3.9 9.2 4.2 0.496 0.516 0.02
<0.0001 37.3 34.8 1.6 2A6 0.777 1.8 0.837 -5.7 2A6 0.824 -4.1
0.762 3.7 2A6 0.769 0.790 0.03 0.96 2.8 0.14 2.16 0.753 0.784 0.05
0.81 4.8 0.91 3.33 2B6 0.878 -11.0 0.748 5.5 2B6 0.813 -2.8 0.845
-6.7 2B6 0.843 0.845 0.03 0.052 -6.6 -6.8 2.4 0.823 0.805 0.05 0.66
-4.0 -1.8 3.7 2C8 0.812 -2.6 0.790 0.19 2C8 0.877 -10.9 0.743 6.1
2C8 0.905 0.865 0.05 0.059 -14.3 -9.3 3.5 0.771 0.768 0.02 0.19 2.5
2.9 1.7 2C9 0.743 6.1 0.722 8.7 2C9 0.749 5.4 0.760 4.0 2C9 0.845
0.779 0.06 0.74 -6.8 1.6 4.2 0.834 0.772 0.06 0.60 -5.4 2.4 4.2
control 0.738 0.738 control 0.697 0.697 control 0.701 0.712 0.023
0.701 0.712 0.023 2C19 0.696 2.2 0.659 7.4 2C19 0.658 7.6 0.618
13.3 2C19 0.634 0.662 0.031 0.091 11.0 6.9 2.5 0.591 0.623 0.034
0.020 16.9 12.5 2.8 2D6 0.649 8.8 0.546 23.3 2D6 0.629 11.6 0.593
16.7 2D6 0.647 0.642 0.011 0.0084 9.1 9.9 0.9 0.581 0.573 0.024
0.0020 18.5 19.5 2.0 2E1 0.693 2.7 0.608 14.6 2E1 0.604 15.1 0.622
12.6 2E1 0.596 0.631 0.054 0.074 16.3 11.3 4.4 0.560 0.597 0.033
0.0073 21.3 16.2 2.6 3A4 0.567 20.3 0.486 31.7 3A4 0.582 18.3 0.434
39.0 3A4 0.605 0.585 0.019 0.0017 15.0 17.9 1.6 0.488 0.470 0.031
0.0004 31.4 34.0 2.5
TABLE-US-00034 TABLE 31 Metabolism of Metaxalone (7.5 .mu.M) by
Expressed Recombinant Human Cytochromes P450 at 5 or 20 pmol 5 pmol
cyp 20 pmol cyp [metaxalone] % Turnover [metaxalone] % Turnover
.mu.M Mean Std dev p % Mean Std Error .mu.M Mean Std dev p % Mean
Std Error control 8.52 8.52 control 8.41 8.41 control 8.40 8.44
0.07 8.40 8.45 0.07 -- 1A2 7.24 14.3 5.63 33.3 1A2 NA NA 5.72 32.3
1A2 7.97 7.60 -- -- 5.7 10.0 4.3 6.29 5.88 0.36 0.0003 25.5 30.4
2.4 2A6 7.69 9.0 8.58 -1.6 2A6 8.08 4.3 7.93 6.1 2A6 8.64 8.14 0.48
0.16 -2.3 3.7 3.3 9.27 8.59 0.67 0.72 -9.8 -1.8 4.6 2B6 8.56 -1.3
8.95 -6.0 2B6 8.19 3.0 8.29 1.8 2B6 7.84 8.20 0.36 0.31 7.2 3.0 2.5
8.14 8.46 0.43 0.95 3.6 -0.20 2.93 2C8 7.58 10.3 8.09 4.3 2C8 8.43
0.21 7.59 10.2 2C8 8.58 8.20 0.54 0.48 -1.6 3.0 3.7 7.31 7.66 0.40
0.028 13.4 9.3 2.7 2C9 7.24 14.3 8.54 -1.1 2C9 8.20 2.9 8.31 1.6
2C9 7.90 7.78 0.49 0.08 6.4 7.9 3.4 7.54 8.13 0.52 0.36 10.7 3.7
3.6 control 7.36 7.36 control 7.85 7.85 control 7.93 7.71 0.31 --
7.93 7.71 2C19 7.69 0.25 6.16 20.2 2C19 7.17 7.1 6.32 18.0 2C19
6.82 7.23 0.44 0.19 11.6 6.3 3.3 6.34 6.27 0.10 0.0015 17.8 18.7
0.8 2D6 6.89 10.7 5.87 23.9 2D6 6.75 12.5 5.90 23.5 2D6 6.60 6.74
0.15 0.0080 14.5 12.6 1.1 5.61 5.79 0.16 0.0007 27.3 24.9 1.2 2E1
6.41 16.9 6.33 18.0 2E1 6.42 16.7 6.25 19.0 2E1 6.04 6.29 0.22
0.0028 21.7 18.4 1.6 6.60 6.39 0.18 0.0031 14.5 17.2 1.4 3A4 6.36
17.5 5.07 34.3 3A4 6.43 16.7 5.18 32.8 3A4 6.50 6.43 0.07 0.0022
15.7 16.6 0.5 5.40 5.22 0.17 0.0003 30.0 32.4 1.2 NA--NOT INCLUDED
IN CALCULATIONS; NO PEAK DETECTED.
[0278] In these experiments, several CYP450s at both 5 and 20
pmoles were shown to be capable of metabolizing metaxalone at 0.75
.mu.M including CYP3A4, CYP2E1, CYP2D6, CYP2C19, and CYP1A2 which
showed percent turnover of 17.9, 11.3, 9.9, 6.9 and 9.2% at 5 pmol
CYP450, and 34.0, 16.2, 19.5, 12.5, and 34.8% at 20 pmol,
respectively. At 7.5 .mu.M metaxalone CYP3A4, CYP2E1, CYP2D6,
CYP2C19, and CYP1A2 were most effective with percent turnovers of
16.6, 18.4, 12.6, 6.3, and 10.0% at 5 pmol CYP450, and 32.4, 17.2,
24.9, 18.7, and 30.4% at 20 pmol CYP450, respectively. At the
higher metaxalone concentration, CYP2C8 and CYP2C9 also moderately
metabolized metaxalone (ranging from 3.0% to 9.3% turnover). These
data indicate that multiple CYP450s can contribute to the
metabolism of metaxalone, with CYP1A2, CYP3A4, CYP2E1, CYP2C19, and
CYP2D6 producing the largest percentages for metaxalone metabolism
under these conditions.
[0279] By combining both approaches to analysis of metaxalone
metabolism, selective chemical inhibitors against metabolism by
pooled human liver microsomes and individual recombinant CYP450
expressed in microsomes, metaxalone appears to be a substrate for
CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, and CYP3A4, with
metabolism in the human liver likely to be primarily by hepatic
cytochrome P450 isozymes CYP1A2, CYP2E1, and CYP3A4, with possible
additional contributions to metabolism by hepatic cytochrome P450
isozymes CYP2C19, CYP2C8, CYP2D6, and CYP2C9.
[0280] Preferred embodiments of this invention are described
herein, including the best mode known to the inventors for carrying
out the invention. Variations of those preferred embodiments may
become apparent to those of ordinary skill in the art upon reading
the foregoing description. The inventors expect skilled artisans to
employ such variations as appropriate, and the inventors intend for
the invention to be practiced otherwise than as specifically
described herein. Accordingly, this invention includes all
modifications and equivalents of the subject matter recited in the
claims appended hereto as permitted by applicable law. Moreover,
any combination of the above-described elements in all possible
variations thereof is encompassed by the invention unless otherwise
indicated herein or otherwise clearly contradicted by context.
* * * * *