U.S. patent number RE47,404 [Application Number 15/644,637] was granted by the patent office on 2019-05-28 for viral hepatitis treatment.
This patent grant is currently assigned to ROMARK LABORATORIES, L.C.. The grantee listed for this patent is Romark Laboratories, L.C.. Invention is credited to Jean-Francois Rossignol.
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United States Patent |
RE47,404 |
Rossignol |
May 28, 2019 |
Viral hepatitis treatment
Abstract
The present disclosure relates to methods for treating viral
hepatitis, compounds useful in the treatment of viral hepatitis,
and pharmaceutical compositions comprising such compounds. In one
embodiment, pharmaceutical compositions comprising nitazoxanide,
tizoxanide, or derivatives and/or mixtures thereof are provided, as
well as methods of treating hepatitis C using such
compositions.
Inventors: |
Rossignol; Jean-Francois (St.
Petersburg, FL) |
Applicant: |
Name |
City |
State |
Country |
Type |
Romark Laboratories, L.C. |
Tampa |
FL |
US |
|
|
Assignee: |
ROMARK LABORATORIES, L.C.
(Tampa, FL)
|
Family
ID: |
38257004 |
Appl.
No.: |
15/644,637 |
Filed: |
July 7, 2017 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
Issue Date |
|
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11651672 |
Jan 12, 2014 |
8633230 |
|
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60757036 |
Jan 9, 2006 |
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Reissue of: |
14137280 |
Dec 20, 2013 |
9107913 |
Aug 18, 2015 |
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Current U.S.
Class: |
1/1 |
Current CPC
Class: |
A61K
31/7068 (20130101); A61K 31/426 (20130101); A61K
38/212 (20130101); A61K 31/7068 (20130101); A61P
31/12 (20180101); A61K 38/21 (20130101); A61K
45/06 (20130101); A61P 31/14 (20180101); A61K
38/212 (20130101); A61P 1/16 (20180101); A61P
3/10 (20180101); A61K 45/06 (20130101); A61K
31/426 (20130101); A61K 38/21 (20130101); A61K
38/21 (20130101); A61K 38/21 (20130101); A61K
2300/00 (20130101); A61K 2300/00 (20130101); A61K
31/426 (20130101); A61K 31/426 (20130101); A61K
2300/00 (20130101); A61K 2300/00 (20130101); A61K
38/212 (20130101); A61K 38/212 (20130101); A61K
2300/00 (20130101); A61K 2300/00 (20130101) |
Current International
Class: |
A61K
31/425 (20060101); A61K 31/426 (20060101); A61K
31/70 (20060101); A61K 38/00 (20060101); A61K
31/7068 (20060101); A61K 45/06 (20060101); A61K
38/21 (20060101) |
Field of
Search: |
;514/370 |
References Cited
[Referenced By]
U.S. Patent Documents
Foreign Patent Documents
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0755386-81 |
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EP |
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0755386 |
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EP |
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1213029 |
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EP |
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1222921 |
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Jul 2002 |
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EP |
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1005342-81 |
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Dec 2002 |
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EP |
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1005342 |
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Dec 2002 |
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EP |
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WO-95028393 |
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Oct 1995 |
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WO |
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WO-04041295 |
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May 2004 |
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WO |
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WO-05049065 |
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Jun 2005 |
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WO |
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WO-06031566 |
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Mar 2006 |
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WO |
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WO-06110814 |
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Oct 2006 |
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WO |
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WO-07081974 |
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Jul 2007 |
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WO |
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Other References
"Romark to Develop Alina.RTM. (nitazoxanide) as New Treatment for
Chronic Hepatits C." Romark Laboratories. Jan. 10, 2006. Web. Aug.
5, 2009. www.natap.orq/2006/HCV/011006 02.htm. cited by applicant
.
"What is Viral Hepatits?" Centers for Disease Control and
Prevention. Apr. 1, 2008. Web. Dec. 14, 2009.
www.cdc.aov/hepatitis/PublicInfo.htm. cited by applicant .
Stockis et al. "Nitazoxanide Pharmacokinetics and Tolerability in
Man After Single Ascending Oral Doses." Int. J. Clin. Pharmacol.
Ther. 40.5(2002):213-220. cited by applicant .
Stockis et al. "Nitazoxanide Pharmacokinetics and Tolerability in
Man During 7 Days of 0.5 g and 1 q b.i.d. Dosing." Int. J. Clin.
Pharmacol. Ther. 40.5(2002):221-227. cited by applicant .
Fabris et al. "Type 1 Diabetes Mellitus in Patients with Chronic
Hepatitis C Before and After Interferon Therapy." Ailment
Pharmacol. Ther. 18(2003):549-558. cited by applicant .
Fung et al. "Viral Hepatitis in 2003." Curr. Opin. Gastroenterol.
20.3(2004):241-247. cited by applicant .
Ortiz et al. "Randomized Clinical Study of Nitazoxanide Compared to
Metronidazole in the Treatment of Symptomatic Giardiasis in
Children from Northern Peru." Ailment Pharmacol. Ther.
15(2001):1409-1415. cited by applicant .
Rossignol et al. "Effect of Nitazoxanide for Treatment of Severe
Rotavirus Diarrhea: Randomised Double-Blind Placebo-Controlled
Trial." Lancet. 368.9530(2006):124-129. cited by applicant .
Rossignol et al. "Effect of Nitazoxanide in Persistent Diarrhea and
Enteritis Associated With Blastocystis hominis." Clin.
Gastroenterol. Hepatol. 3(2005):987-991. cited by applicant .
Rossignol et al. "Nitazoxanide in the Treatment of Viral
Gastroenteritis: A Randomized Double-Blind Placebo-Controlled
Clinical Trial." Ailment Pharmacol. Ther. 24(2006):1423-1430. cited
by applicant .
Rossignol et al. "Nitazoxanide in Treating Chronic Hepatitis C: in
Vitro Activity and a Clinical Case Report." Gastroenterol.
130(2006):A-841. (Abstract #T1821). cited by applicant .
Rossignol et al. "Treatment of Diarrhea Caused by Cryptosporidium
parvum: A Prospective Randomized, Double-Blind, Placebo-Controlled
Study of Nitazoxanide." J. Infect. Dis. 184(2001):103-106. cited by
applicant .
Rossignol et al. "Treatment of Diarrhea Caused by Giardia
intestinalis and Entamoeba histolytica or E. dispar: A Randomized,
Double-Blind, Placebo-Controlled Study of Nitazoxanide." J. Infect.
Dis. 184(2004):381-384. cited by applicant.
|
Primary Examiner: Kugel; Timothy J.
Attorney, Agent or Firm: Foley & Lardner LLP
Government Interests
GOVERNMENT RIGHTS
The U.S. Government has certain rights in this invention pursuant
to Contract No. NO1-AI-30046 awarded by the NIAID.
Parent Case Text
CROSS-REFERENCE TO RELATED APPLICATIONS
This application .Iadd.is a Reissue Application of U.S. Pat. No.
9,107,913, which issued on Aug. 18, 2015 from U.S. application Ser.
No. 14/137,280, which is a Continuation of U.S. application Ser.
No. 11/651,672, which issued as U.S. Pat. No. 8,633,230, which
.Iaddend.claims priority under 35 U.S.C. .[..sctn.119.].
.Iadd..sctn. 119 .Iaddend.to Provisional U.S. Patent Application
Ser. No. 60/757,036, filed Jan. 9, 2006, the disclosure of which is
incorporated by reference herein.
Claims
What is claimed is:
1. A method of treating hepatitis C in a patient suffering from
hepatitis C, the method comprising administering to the patient a
compound selected from nitazoxanide and tizoxanide or a mixture
thereof, in an amount effective to reduce serum hepatitis C virus
RNA in the serum of the patient to undetectable levels after
administering the compound to the patient for a first period of
time between about 3 days and about 24 weeks, wherein the amount is
from 100 mg to 2000 mg per day.
2. The method of claim 1, wherein the compound is administered in
the form of a composition further comprising a pharmaceutically
acceptable carrier.
3. The method of claim 2, wherein the composition comprises a
mixture of nitazoxanide and tizoxanide.
4. The method of claim 2, wherein the composition further comprises
one or more additional biologically active agents selected from the
group consisting of an interferon, an anti-diabetic agent,
ribavirin and 2-methyl cytidine.
5. The method of claim 1 or 3, further comprising, after the first
period of time, administering the compound or composition and an
interferon to the patient for a second period of time of between
about 1 week and about 48 weeks.
6. The method of claim 1, wherein the compound is administered to
the patient for a period of between about 3 days and about 2
years.
7. The method of claim 1 or 3, wherein the method further comprises
administering one or more additional active agents selected from
the group consisting of an interferon, an anti-diabetic agent,
ribavirin and 2-methyl cytidine.
8. The method of claim 7, wherein the one or more additional active
agents comprises an interferon.
9. The method of claim 8, wherein the interferon is formulated
separately from the compound.
10. The method of claim 8, wherein the interferon is interferon
.alpha.-2a, interferon .alpha.-2b, or a polyethylene glycol
conjugate of interferon .alpha.-2a or interferon .alpha.-2b.
11. The method of claim 8, wherein the interferon is administered
to the patient for a period of about 1 week to about 48 weeks.
12. The method of claim 11, wherein the interferon is administered
to the patient for a period of about 1 week to about 4-12
weeks.
13. The method of claim 8, wherein the interferon is administered
to the patient between 1 and 3 times each week.
14. The method of claim 8, wherein administration of the interferon
is initiated after the first period of time.
15. The method of claim 8, wherein the first period of time is
between about 1 week and about 4 weeks.
16. The method of claim 1, wherein the compound is administered to
the patient one to three times each day during the period of
treatment.
17. The method of claim 7, wherein the one or more additional
active agents comprises an anti-diabetes agent.
18. The method of claim 17, wherein the anti-diabetes agent is
formulated separately from the compound.
19. The method of claim 4, wherein the composition comprises an
anti-diabetes agent.
20. The method of claim 4, wherein the composition comprises an
interferon.
21. The method of claim 4, wherein the composition comprises an
interferon and an anti-diabetes agent.
22. A method of treating a patient suffering from hepatitis C, the
method comprising (a) pretreating the patient for a predetermined
period of time with an amount of nitazoxanide, tizoxanide or a
mixture thereof, said amount being in the range of 100 mg to 2000
mg per day, and (b) after the predetermined period of time,
administering to the patient an amount of an interferon, wherein
the method is effective to reduce serum hepatitis C virus RNA in
the serum of the patient to undetectable levels.
23. The method of claim 22, wherein the predetermined period of
time is between about 3 days and about 3 months.
24. The method of claim 23, wherein the predetermined period of
time is between about 1 week and about 4 weeks.
25. The method of claim 22, wherein interferon is selected from
interferon .alpha.-2a, interferon .alpha.-2b, and a polyethylene
glycol conjugate of interferon .alpha.-2a or interferon
.alpha.-2b.
26. The method of claim 25, wherein step (b) further comprises
administering a compound selected from nitazoxanide, tizoxanide or
a mixture thereof.
.Iadd.27. A method of treating liver fibrosis comprising
administering to a subject in need thereof an effective amount of a
compound selected from nitazoxanide, tizoxanide and a mixture
thereof..Iaddend.
.Iadd.28. The method of claim 27, wherein the compound is
administered in the form of a composition further comprising a
pharmaceutically acceptable carrier..Iaddend.
.Iadd.29. The method of claim 28, wherein the composition comprises
a mixture of nitazoxanide and tizoxanide..Iaddend.
.Iadd.30. The method of claim 27, wherein the fibrosis is caused by
hepatitis..Iaddend.
.Iadd.31. The method of claim 30, wherein the hepatitis is
hepatitis C..Iaddend.
.Iadd.32. The method of claim 27, wherein the effective amount is
from about 100 mg to about 2000 mg per day..Iaddend.
.Iadd.33. The method of claim 27, wherein the effective amount is
from 250 mg to 1000 mg per day..Iaddend.
.Iadd.34. The method of claim 27, wherein the effective amount
achieves and maintains in the subject a blood level of tizoxanide
between 0.1 .mu.g/ml and 10 .mu.g/ml..Iaddend.
.Iadd.35. The method of claim 27, wherein the compound is
administered orally in an oral dosage form..Iaddend.
.Iadd.36. The method of claim 35, wherein the oral dosage form is a
tablet, a capsule, a caplet or a particulate..Iaddend.
.Iadd.37. The method of claim 35, wherein the oral dosage form
provides a sustained release of the compound over an extended
period of time..Iaddend.
.Iadd.38. The method of claim 35, wherein the subject is a
human..Iaddend.
.Iadd.39. The method of claim 27, wherein the fibrosis comprises
fibrous portal expansion..Iaddend.
.Iadd.40. The method of claim 27, wherein the fibrosis is bridging
fibrosis..Iaddend.
.Iadd.41. The method of claim 27, wherein said administering slows
down progression of the fibrosis..Iaddend.
Description
TECHNICAL FIELD
The present disclosure relates to methods for treating viral
hepatitis, compounds useful in the treatment of viral hepatitis,
and pharmaceutical compositions comprising such compounds.
BACKGROUND
Hepatitis refers to a variety of conditions that involve
inflammation of the liver. Viral hepatitis, of which there are
several types (e.g., hepatitis A, B, C, D, and E), is an
inflammation of the liver due to a viral infection. Each type of
viral hepatitis may exhibit different symptoms and may be
characterized by different approaches to treatment and prevention.
For example, vaccines have been developed for hepatitis A and B,
but not for hepatitis C or E.
The main goal of treatment of chronic hepatitis C is to eliminate
detectable viral RNA from the blood. Patients lacking detectable
hepatitis C virus RNA in the blood 24 weeks after completing
therapy typically have a favorable prognosis and may be considered
to be cured of the virus. Such a condition is known as a sustained
virologic response. For patients not achieving a sustained
virologic response, there may be other more subtle benefits of
treatment, such as slowing the progression of liver scarring
(fibrosis).
Treatment of hepatitis C virus (HCV) commonly involves
administration of injectable interferon (or injectable pegylated
interferon), ribavirin, or a combination thereof. Interferon alpha
is a naturally occurring glycoprotein that is secreted by cells in
response to viral infections. It exerts its effects by binding to a
membrane receptor. Receptor binding initiates a series of
intracellular signaling events that ultimately leads to enhanced
expression of certain genes. This leads to the enhancement and
induction of certain cellular activities including augmentation of
target cell killing by lymphocytes and inhibition of virus
replication in infected cells. Ribavirin is a synthetic nucleoside
that has activity against a broad spectrum of viruses.
Interferon alpha, with or without ribavirin, is associated with may
side effects. Flu-like symptoms, depression, rashes, other unusual
reactions and abnormal blood counts are common examples of such
side effects. Ribavirin is associated with a significant risk of
abnormal fetal development. Accordingly, women who are potentially
pregnant should not begin therapy until a report of a negative
pregnancy test has been obtained. Female patients are advised to
avoid becoming pregnant during treatment. Patients using interferon
alpha and ribavirin are advised to have blood tests approximately
once a month, and somewhat more frequently at the beginning of
treatment. Certain groups of patients cannot take ribavirin, for
example those with anemia, heart disease or kidney disease. In such
cases, pegylated interferon alpha is typically prescribed alone.
Some patients with hepatitis C (e.g., patients also having advanced
liver disease) are advised not to take interferon alpha or
pegylated interferon alpha because of the risk of serious side
effects. For such patients, no previously available method of
treatment is recognized as being effective and safe for treating
hepatitis C.
There is therefore a need in the art to develop an effective method
of treatment of hepatitis C. An ideal method of treatment would
achieve a sustained virologic response in a wide range of patients.
Such a treatment would employ readily available active agents and
would have minimal side effects. When co-administration of
interferon alpha is employed, an ideal method of treatment would
require reduced amounts of interferon alpha (i.e., reduced
frequency of administration, reduced amount per administration, or
both) as compared with traditional methods of treatment.
SUMMARY OF THE DISCLOSURE
The present invention is directed at addressing one or more of the
abovementioned drawbacks of known methods for treating viral
hepatitis C.
In one embodiment, then, the disclosure describes a method of
treating a patient suffering from hepatitis C. The method comprises
administering to the patient a therapeutically effective amount of
a compound selected from nitazoxanide, tizoxanide, derivatives of
nitazoxanide, and derivatives of tizoxanide.
In another embodiment, the disclosure describes a method for
treating a patient suffering from viral hepatitis. The method
comprises administering to the patient a therapeutically effective
amount of a first compound having the structure of formula I:
R1-NHCO--R2. In formula I, R1 and R2 are independently selected
from moieties that provide improved stability of the NHCO group in
biological fluid and tissue. In one aspect of the embodiment, the
first compound is neither nitazoxanide nor tizoxanide.
In yet another embodiment, the disclosure describes an improvement
in a method for treating a patient suffering from hepatitis C
comprising administering to the patient a therapeutically effective
amount of nitazoxanide, tizoxanide, or mixtures thereof.
In yet another embodiment, the disclosure describes a method of
treating a patient suffering from hepatitis C. The method comprises
pretreating the patient by administering to the patient for a
predetermined period of time a first composition comprising a
therapeutically effective amount of a compound selected from
nitazoxanide, tizoxanide, derivatives of nitazoxanide, and
derivatives of tizoxanide, or mixtures thereof. The method further
comprises administering to the patient, after the predetermined
period of time, a therapeutically effective amount of a second
composition comprising an active agent.
In yet another embodiment, the disclosure describes a composition
comprising: (a) one or more compounds selected from nitazoxanide,
tizoxanide, derivatives of nitazoxanide, and derivatives of
tizoxanide; (b) an interferon; and (c) an anti-diabetes agent.
In yet another embodiment, the disclosure describes compositions
effective in the methods of treatment disclosed herein.
BRIEF DESCRIPTION OF THE DRAWINGS
FIGS. 1a and 1b are graphs illustrating the synergistic activity of
nitazoxanide with interferon alpha-2b or 2'-C-methyl cytidine
against HCV replication in an HCV replicon containing cell
line.
FIGS. 2a and 2b are graphs illustrating synergistic activity when
an HCV replicon-containing cell line is treated first with
nitazoxanide and then with nitazoxanide plus interferon
alpha-2b.
FIG. 3 is a patient disposition chart showing the selection of
participants for the experiment described in Example 5.
FIG. 4, described in Example 5, is a graph showing mean
quantitative serum HCV RNA levels over time for different treatment
groups.
FIG. 5, described in Example 5, is a graph showing quantitative
serum HCV RNA levels over time for different patients.
FIG. 6 is a patient disposition chart showing the selection of
participants for the experiment described in Example 6.
FIG. 7, described in Example 6, is a graph showing platelet count
versus time for patients administered pegylated interferon alpha-2b
plus either Alinia.RTM. or a placebo.
FIG. 8, described in Example 6, is a graph showing neutrophil count
versus time for patients administered pegylated interferon alpha-2b
plus either Alinia.RTM. or a placebo.
DETAILED DESCRIPTION OF THE INVENTION
Definitions and Nomenclature
Before describing the present invention in detail, it is to be
understood that unless otherwise indicated, this invention is not
limited to particular dosages, formulations or methods of use, as
such may vary. It is also to be understood that the terminology
used herein is for the purpose of describing particular embodiments
only, and is not intended to be limiting.
It must be noted that, as used in this specification and the
appended claims, the singular forms "a," "an" and "the" include
plural referents unless the context clearly dictates otherwise.
Thus, for example, "a dosage form" refers not only to a single
dosage form but also to a combination of two or more different
dosage forms, "an active agent" refers to a combination of active
agents as well as to a single active agent, and the like.
As used in the specification and the appended claims, the terms
"for example," "for instance," "such as," "including" and the like
are meant to introduce examples that further clarify more general
subject matter. Unless otherwise specified, these examples are
provided only as an aid for understanding the invention, and are
not meant to be limiting in any fashion.
Unless defined otherwise, all technical and scientific terms used
herein have the meaning commonly understood by one of ordinary
skill in the art to which the invention pertains. Although any
methods and materials similar or equivalent to those described
herein may be useful in the practice or testing of the present
invention, preferred methods and materials are described below.
Specific terminology of particular importance to the description of
the present invention is defined below.
When referring to a compound of the invention, and unless otherwise
specified, the term "compound" is intended to encompass not only
the specified molecular entity but also its pharmaceutically
acceptable, pharmacologically active analogs, including, but not
limited to, salts, polymorphs, esters, amides, prodrugs, adducts,
conjugates, active metabolites, and the like, where such
modifications to the molecular entity are appropriate.
The terms "treating" and "treatment" as used herein refer to
reduction in severity and/or frequency of symptoms, elimination of
symptoms and/or underlying cause, prevention of the occurrence of
symptoms and/or their underlying cause (e.g., prophylactic
therapy), improvement or remediation of damage, or reduction in
intensity of infection.
By the terms "effective amount" and "therapeutically effective
amount" of a compound of the invention is meant a nontoxic but
sufficient amount of the drug or agent to provide the desired
effect.
By "pharmaceutically acceptable" is meant a material that is not
biologically or otherwise undesirable, i.e., the material may be
incorporated into a pharmaceutical composition administered to a
patient without causing any undesirable biological effects or
interacting in a deleterious manner with any of the other
components of the composition in which it is contained. When the
term "pharmaceutically acceptable" is used to refer to a
pharmaceutical carrier or excipient, it is implied that the carrier
or excipient has met the required standards of toxicological and
manufacturing testing or that it is included on the Inactive
Ingredient Guide prepared by the U.S. Food and Drug
administration.
By "patient," or "subject" is meant any animal for which treatment
is desirable. Patients may be mammals, and typically, as used
herein, a patient is a human individual.
The present disclosure includes compounds of formula I:
R1-NHCO--R2, as well as their use in the treatment of hepatitis,
particularly hepatitis C, and pharmaceutical compositions
comprising them.
In one embodiment of formula I, R1 and R2 are independently
selected from moieties that stabilize (i.e., provide improved
stability of) the NHCO group. By "stabilize" is meant that the NHCO
group is less prone to reaction in biological fluid and tissue as
compared with an unsubstituted NHCO group (e.g., NH.sub.2COH,
R1-NHCOH, NH.sub.2CO--R2, and the like), that is, as compared with
the analogous compound having hydrogen as either R1 or R2. Such
reactions include cleavage of the NHCO group (e.g., breakage of the
nitrogen-carbon bond), addition to the NHCO group, substitution
reactions, hydrogenation reactions, hydration reactions, oxidation
reactions, reduction reactions, and the like.
In one embodiment, the compounds of formula I exclude nitazoxanide
and tizoxanide. In another embodiment, the compounds of formula I
include nitazoxanide and tizaxanide.
In another embodiment, R1 and R2 are each a substituted or
unsubstituted cyclic group. Such groups may be heterocyclic groups
or a carbocyclic group such as an aryl or cycloalkyl group. In one
example, R1 is a heterocyclic ring and R2 is an aryl, optionally
substituted by one to three substituents. Another example group of
compounds of formula I includes compounds wherein R1 and R2 are
both benzene, each optionally substituted by one to three
substituents.
In yet another embodiment, R1 is selected from thiazole and
thiadiazole substituted by one to three substituents, and R2 is
benzene substituted by one to three substituents.
Examples of substituents for R1 and R2 include OH, alkoxy, halo,
alkyl, fluoroalkyl, ester, thioalkyl, and functional groups.
Specific examples include fluoro, bromo, OAc, CH.sub.3, CF.sub.3,
NO.sub.2, CH.sub.2CO.sub.2Et, SCH.sub.3, OCH.sub.3 and the
like.
Examples of the heterocyclic groups for R1 and R2 include aromatic
heterocyclic groups or saturated or unsaturated non-aromatic
heterocyclic groups (alicyclic heterocyclic group). Such groups
contain, besides carbon atoms, at least one heteroatom (preferably
1 to 4 heteroatom(s), more preferably, 1 to 2 heteroatom(s)), and
may contain from 1 to 3 different kind of heteroatoms, (preferably
1 to 2 kinds of heteroatom(s)). As used herein, the term
"heteroatom" is meant to include oxygen atoms, sulfur atoms, and
nitrogen atoms.
Examples of the "aromatic heterocyclic group" include an aromatic
monocyclic heterocyclic group such as a 5 or 6-membered aromatic
monoyclic heterocyclic group (e.g., furyl, thienyl, pyrrolyl,
oxazolyl, isooxazolyl, thiazolyl, thiadiazolyl, isothiazolyl,
imidazolyl, pyrazolyl, 1,2,3-oxadiazolyl, 1,2,4-oxadiazolyl,
1,3,4-oxadiazolyl, furazanyl, 1,2,3-thiadiazolyl,
1,2,4-thiadiazolyl, 1,3,4-thiadiazolyl, 1,2,3-triazolyl,
1,2,4-triazolyl, tetrazolyl, pyridyl, pyridazinyl, pyrimidinyl,
pyrazinyl, triazinyl, etc.); an aromatic fused heterocyclic group
such as a 8 to 12-membered aromatic fused heterocyclic group (e.g.,
benzofuranyl, isobenzofuranyl, benzothienyl, indolyl, isoindolyl,
1H-indazolyl, benzindazolyl, benzoxazolyl, 1,2-benzoisooxazolyl,
benzothiazolyl, benzopyranyl, 1,2-benzoisothiazolyl,
1H-benzotriazolyl, quinolyl, isoquinolyl, cinnolinyl, quinazolinyl,
quinoxalinyl, phthalazinyl, naphthylidinyl, purinyl, pteridinyl,
carbazolyl, alpha-carbolinyl, beta-carbolinyl, gamma-carbolinyl,
acridinyl, phenoxazinyl, phenothiazinyl, phenazinyl, phenoxathinyl,
thianthrenyl, phenanthridinyl, phenanthrolinyl, indolizinyl,
pyrrolo[1,2-b]pyridazinyl, pyrazolo[1,5-a]pyridyl,
imidazo[1,2-a]pyridyl, imidazo[1,5-a]pyridyl,
imidazo[1,2-b]pyridazinyl, imidazo[1,2-a]pyrimidinyl,
1,2,4-triazolo[4,3-a]pyridyl, 1,2,4-triazolo[4,3-b]pyridaizinyl);
preferably, a heterocyclic group consisting of the above-mentioned
5- or 6-membered aromatic monocyclic heterocyclic group fused with
a benzene ring or heterocyclic group consisting of the
above-mentioned 5- or 6-membered aromatic monocyclic heterocyclic
group fused with the same or different above-mentioned 5- or
6-membered aromatic monocyclic heterocyclic group.
Examples of the "non-aromatic heterocyclic group" include a 3 to
8-membered (preferably 5 or 6-membered) saturated or unsaturated
(preferably saturated) non-aromatic heterocyclic group (aliphatic
heterocyclic group) such as oxiranyl, azetidinyl, oxetanyl,
thiethanyl, pyrrolidinyl, tetrahydrofuryl, thiolanyl, piperidinyl,
tetrahydropyranyl, morpholinyl, thiomorpholinyl, piperazinyl.
In one embodiment of compounds having the structure of formula I,
R1 is heterocyclic. In another embodiment, R1 is heterocyclic
comprising 2 or 3 heteroatoms. In yet another embodiment, R1 is
substituted heterocyclic and comprises 2 or 3 heteroatoms. In yet
another embodiment, R1 is heterocyclic, substituted with 1, 2, or 3
groups selected from hydroxide, halogen (i.e., iodo, chloro, bromo,
or fluoro), alkoxy (e.g., OCH.sub.3), fluoroalkyl (e.g., CF.sub.3),
ester (e.g., CH.sub.2CO.sub.2Et), thioalkyl (e.g., SCH.sub.3), OAc,
and alkyl (e.g., CH.sub.3). For example, R1 is thiazole or
substituted thiazole.
In one embodiment of compounds having the structure of formula I,
R2 is aryl. In another embodiment, R2 is substituted aryl. In yet
another embodiment, R2 is aryl that comprises 2, 3, or 4
substituents. In another embodiment, R2 is aryl and comprises
substituents in the ortho and meta positions (relative to the point
of attachment of the aryl group to the carbonyl group of formula
I). In still another embodiment, R2 is aryl comprising 2 or more
substituents selected from hydroxide, halogen (i.e., iodo, chloro,
bromo, or fluoro), alkoxy (e.g., OCH.sub.3), fluoroalkyl (e.g.,
CF.sub.3), ester (e.g., CH.sub.2CO.sub.2Et), thioalkyl (e.g.,
SCH.sub.3), OAc, and alkyl (e.g., CH.sub.3).
In one embodiment of the compounds having the structure of formula
I, the reactivity of the NHCO group in the compound is reduced
toward cleavage reactions compared with the reactivity of the
analogous compound having hydrogen as either R1 or R2.
In another embodiment, R1 and R2 are independently selected from
substituted cyclic groups, unsubstituted cyclic groups, substituted
heterocyclic groups, and unsubstituted heterocyclic groups, wherein
either R1, R2, or both R1 and R2 are optionally aromatic. In yet
another embodiment, R1 and R2 are selected from aryl, substituted
aryl, heteroaryl, substituted heteroaryl, alicyclic, substituted
alicyclic, heterocyclic, and substituted heterocyclic. In yet
another embodiment, R1 and R2 are each substituted with from one to
three substituents independently selected from OH, NO.sub.2, alkoxy
(such as methoxy), halo (such as F and Br), alkyl (such as methyl),
fluoroalkyl (such as fluoromethyl), ester (such as OAc, and
CH.sub.2CO.sub.2Et), and thioalkyl (such as thiomethyl). In a still
further embodiment, at least one of R1 and R2 is heterocyclic. In a
still further embodiment, at least one of R1 and R2 comprises
between 1 and 3 heteroatoms. In yet another embodiment, at least
one of R1 and R2 comprises a heterocyclic group selected from
aromatic monocyclic heterocycles, aromatic fused heterocycles, and
non-aromatic heterocycles. In a still further embodiment, at least
one of R1 and R2 comprises a heterocyclic group selected from
furyl, thienyl, pyrrolyl, oxazolyl, isooxazolyl, thiazolyl,
thiadiazolyl, isothiazolyl, imidazolyl, pyrazolyl,
1,2,3-oxadiazolyl, 1,2,4-oxadiazolyl, 1,3,4-oxadiazolyl, furazanyl,
1,2,3-thiadiazolyl, 1,2,4-thiadiazolyl, 1,3,4-thiadiazolyl,
1,2,3-triazolyl, 1,2,4-triazolyl, tetrazolyl, pyridyl, pyridazinyl,
pyrimidinyl, pyrazinyl, triazinyl, benzofuranyl, isobenzofuranyl,
benzothienyl, indolyl, isoindolyl, 1H-indazolyl, benzindazolyl,
benzoxazolyl, 1,2-benzoisooxazolyl, benzothiazolyl, benzopyranyl,
1,2-benzoisothiazolyl, 1H-benzotriazolyl, quinolyl, isoquinolyl,
cinnolinyl, quinazolinyl, quinoxalinyl, phthalazinyl,
naphthylidinyl, purinyl, pteridinyl, carbazolyl, alpha-carbolinyl,
beta-carbolinyl, gamma-carbolinyl, acridinyl, phenoxazinyl,
phenothiazinyl, phenazinyl, phenoxathinyl, thianthrenyl,
phenanthridinyl, phenanthrolinyl, indolizinyl,
pyrrolo[1,2-b]pyridazinyl, pyrazolo[1,5-a]pyridyl,
imidazo[1,2-a]pyridyl, imidazo[1,5-a]pyridyl,
imidazo[1,2-b]pyridazinyl, imidazo[1,2-a]pyrimidinyl,
1,2,4-triazolo[4,3-a]pyridyl, 1,2,4-triazolo[4,3-b]pyridaizinyl),
oxiranyl, azetidinyl, oxetanyl, thiethanyl, pyrrolidinyl,
tetrahydrofuryl, thiolanyl, piperidinyl, tetrahydropyranyl,
morpholinyl, thiomorpholinyl, and piperazinyl, any of which may be
optionally substituted with 1 to 3 substituents. In another
embodiment, R1 is a heterocyclic group optionally substituted with
1 to 3 substituents and R2 is aryl optionally substituted with 1 to
3 substituents. In yet another embodiment, R1 is thiazole or
thiadiazole optionally substituted with 1 to 3 substituents. In a
still further embodiment, R2 is phenyl optionally substituted with
1 to 3 substituents. In a still further embodiment, R1 and R2 are
both aryl, each optionally substituted with 1 to 3
substituents.
Examples of compounds that have the structure of formula I include
nitazoxanide, tizoxanide, RM-4803, RM-4819, RM-4832, and RM-4850,
wherein nitazoxanide, tizoxanide, RM-4819, RM-4832, and RM-4850 are
particularly preferred. The structures of these compounds are shown
in the following list:
##STR00001##
As described below, the compositions of the current disclosure
comprise, as an active agent, compounds having the structure of
formula I in a pharmaceutically acceptable form. If desired, the
compositions may further comprise one or more additional active
agents (also described in detail below). Where it is appropriate,
any of the active agents may be administered in the form of the
compound per se, and/or in the form of a salt, polymorph, ester,
amide, prodrug, derivative, or the like, provided the salt,
polymorph, ester, amide, pro-drug or derivative is suitable
pharmacologically. Where it is appropriate, salts, esters, amides,
prodrugs and other derivatives of the active agents may be prepared
using standard procedures known to those skilled in the art of
synthetic organic chemistry and described, for example, by J.
March, Advanced Organic Chemistry: Reactions, Mechanisms and
Structure, 4th Ed. (New York: Wiley-Interscience, 1992). For any
active agents that may exist in enantiomeric forms, the active
agent may be incorporated into the present compositions either as
the racemate or in enantiomerically enriched form.
Compounds having the structure of formula I may be prepared
according to literature methods. For example, the preparation of
compounds 1a and 1b are described in U.S. Pat. No. 3,950,351, and
WO 95/28393, respectively. Synthetic methods for the preparation of
analogues and derivatives of 1a and 1b, as well as other compounds
having structures that fall within the scope of formula I, employ
known procedures that will be apparent to the skilled artisan.
Pharmaceutical compositions according to this disclosure comprise a
compound having the structure of formula I, as described herein.
Such pharmaceutical compositions may also comprise: (1) one or more
additional compounds having the structure of formula (I); (2) one
or more pharmaceutically acceptable carriers as disclosed herein;
and (3) one or more additional components as described herein. The
compositions may contain from 0.05% to 95% by weight of the active
agent(s), with the pharmaceutically acceptable carrier(s) and any
additional components forming the 5% to 99.95% by weight that
remains.
One or more additional active agents may be included in the
pharmaceutical compositions and methods of treatment described
herein. In one embodiment, the additional active agent is effective
in treating hepatitis. For example, the compositions may include
one or more additional therapeutic agents useful in treating
hepatitis C such as ribavirin and immune-stimulating agents such as
interferons, including interferon .alpha.-2b, a derivative of
interferon .alpha.-2b such as a polyethylene glycol-conjugated form
of interferon .alpha.-2b, interferon .alpha.-2a, or interferon
alfacon-1. Specific examples also include Omega IFN (BioMedicines
Inc., Emeryville, Calif.); BILN-2061 (Boehringer Ingelheim Pharma
KG, Ingelheim, Germany); Summetrel (Endo Pharmaceuticals Holdings
Inc., Chadds Ford, Pa.); Roferon A, Pegasys, Pegasys and Ribavirin,
and CellCept (F. Hoffmann-La Roche LTD, Basel, Switzerland);
Wellferon (GlaxoSmithKline plc, Uxbridge, UK) Albuferon-.alpha.
(Human Genome Sciences Inc., Rockville, Md.); Levovirin (ICN
Pharmaceuticals, Costa Mesa, Calif.); IDN-6556 (Idun
Pharmaceuticals Inc., San Diego, Calif.); IP-501 (Indevus
Pharmaceuticals Inc., Lexington, Mass.); Actimmune (InterMune Inc.,
Brisbane, Calif.); Infergen A (InterMune Pharmaceuticals Inc.,
Brisbane, Calif.); ISIS 14803 (ISIS Pharmaceuticals Inc, Carlsbad,
Calif./Elan Phamaceuticals Inc., New York, N.Y.); JTK-003 (Japan
Tobacco Inc., Tokyo, Japan); Ceplene, Pegasys and Ceplene (Maxim
Pharmaceuticals Inc., San Diego, Calif.); Civacir
(Biopharmaceuticals Inc., Boca Raton, Fla.); Intron A and Zadaxin
(RegeneRx Biopharmiceuticals Inc., Bethesda, Md./SciClone
Pharmaceuticals Inc, San Mateo, Calif.); Levovirin, Viramidine
(Ribapharm Inc., Costa Mesa, Calif.); Heptazyme (Ribozyme
Pharmaceuticals Inc., Boulder, Colo.); Intron A, PEG-Intron,
Rebetron, Ribavirin, PEG-Intron/Ribavirin (Schering-Plough
Corporation, Kenilworth, N.J.); Zadazim (SciClone Pharmaceuticals
Inc., San Mateo, Calif.); Rebif (Serono, Geneva, Switzerland);
IFN-.beta. and EMZ701 (Transition Therapeutics Inc., Ontario,
Canada); T67 (Tularik Inc., South San Francisco, Calif.); VX-497
(Vertex Pharmaceuticals Inc., Cambridge, Mass.); VX-950/LY-570310
(Vertex Pharmaceuticals Inc., Cambridge, Mass./Eli Lilly and Co.
Inc., Indianapolis, Ind.); Omniferon (Viragen Inc., Plantation,
Fla.); and XTL-002 (Biopharmaceuticals Ltd., Rehovot, Isreal).
In addition to or instead of anti-hepatitis agents, pharmaceutical
compositions and methods described herein may comprise one or more
additional active agent as appropriate. Additional active agents
include those effective in treating disorders of the endocrine
system such as diabetes and hyper-insulinemia. Examples of
anti-diabetes agents include insulin, pramlintide, exenatide,
sulfonylureas (e.g., chlorpropamide, glipizide, glyburide,
glimepiride), meglitinides (e.g., repaglinide, nateglinide),
biguanides (e.g., metformin), thiazolidinediones (e.g.,
rosiglitazone, troglitazone, pioglitazone), and .alpha.-glucosidase
inhibitors (e.g., acarbose, meglitol). Such active agents may be
administered either prior to or concurrently with administration of
the compounds disclosed herein in order to regulate plasma levels
of insulin. When administered concurrently, such additional active
agents may be administered as part of the same formulation with the
compounds disclosed herein, or they may be administered in a
separate formulation. Similarly, other active agents such as those
effective in treating diseases of the liver may also be used with
the compounds disclosed herein.
Pharmaceutical compositions comprising the compounds of the
disclosure that are suitable for the uses described herein may also
comprise a pharmaceutically acceptable carrier. Appropriate
pharmaceutical carriers may depend, for example, on the method of
administration of the compositions, as will be appreciated by one
of skill in the art.
Pharmaceutically acceptable carriers may be solid or liquid, or
mixtures thereof. Pharmaceutically acceptable carriers are
materials such as binders, lubricants, disintegrants, fillers,
surfactants, emulsifiers, coloring agents, and the like. Binders
are used to impart cohesive qualities, and thus ensure that the
composition remains intact (e.g., as an implant or tablet).
Suitable binder materials include, but are not limited to, polymer
matrices, hydrogels, starch (including corn starch and
pregelatinized starch), gelatin, sugars (including sucrose,
glucose, dextrose, and lactose), polyethylene glycol, waxes, and
natural and synthetic gums, e.g., acacia sodium alginate,
polyvinylpyrrolidone, cellulosic polymers (including hydroxypropyl
cellulose, hydroxypropyl methylcellulose, methyl cellulose,
microcrystalline cellulose, ethyl cellulose, hydroxyethyl
cellulose, and the like), and Veegum. Lubricants are used to
facilitate manufacture, promoting powder flow and preventing
particle capping (i.e., particle breakage) when pressure is
relieved. Useful lubricants are magnesium stearate, calcium
stearate, and stearic acid. Disintegrants are used to facilitate
disintegration of the composition, and are generally starches,
clays, celluloses, algins, gums, or crosslinked polymers. Fillers
include, for example, materials such as silicon dioxide, titanium
dioxide, alumina, talc, kaolin, powdered cellulose, and
microcrystalline cellulose, as well as soluble materials such as
mannitol, urea, sucrose, lactose, dextrose, sodium chloride, and
sorbitol. Surfactants are wetting agents, and may include ionic
materials such as fatty acid salts and non-ionic materials such as
PLURONICS.TM. (such as F-127, L-122, L-101, L-92, L-81, and
L-61).
For example, the pharmaceutically acceptable carrier for the
compositions disclosed herein may comprise one or more
biocompatible polymer. By "biocompatible" is meant a material that
does not illicit an adverse response when subjected to a biological
environment such as by implantation or injection in vivo.
Furthermore, in one embodiment, biocompatible materials do not
illicit an immune response when administered in vivo. Unless
otherwise stated, biocompatible materials include materials that
are bioerodible, biodegradable and bioresorbable.
Polymer carriers such as biocompatible polymers may be homopolymers
or copolymers of any of the monomer units described herein.
Furthermore, copolymers are not limited to any specific
architecture, and may consist of random, alternating, block
(including multiblock), star, comb, graft, and dendrimer-type
copolymers, as well as combinations thereof. Blends of more than
one bioerodible polymer are also within the scope of this
disclosure. It will be appreciated that crosslinked and
crosslinkable polymers may be used as long as such crosslinking
does not adversely affect the material's ability to form the
compositions described herein (e.g., the material's ability to
bioerode). For example, reversible crosslinks (wherein the
crosslinks comprise non-covalent and/or weakly covalent
intermolecular bonds) may be present prior to administration of the
compositions, or such bonds may form in vivo.
Suitable bioerodible polymers may comprisepoly(orthoester)s,
poly(lactone)s such as poly( -caprolactone) and
poly(.gamma.-caprolactone), poly(lactide)s, poly(lactic acid),
poly(glycolide)s, poly(glycolic acid), poly(ethylene
terephthalate), poly(butyric acid), poly(valeric acid), polymers of
anhydrides, poly(vinyl alcohol), poly(ethylene vinyl acetate),
polymers of .alpha.-hydroxycarboxylic acid and derivatives thereof,
albumin, collagen, gelatin, hyaluronic acid, starch, cellulose and
cellulose derivatives (e.g., methylcellulose,
hydroxypropylcellulose, hydroxypropylmethylcellulose,
carboxymethylcellulose, cellulose acetate phthalate, cellulose
acetate succinate, hydroxypropylmethylcellulose phthalate), casein,
dextrans, polysaccharides, fibrinogen, poly(ether ester) multiblock
copolymers, poly(ether)s such as poly(ethylene glycol), and
poly(butylene terephthalate), tyrosine-derived polycarbonates,
poly(hydroxyl acids), poly(hydroxybutyrate), polydioxanone,
poly(alkylcarbonate), poly(hydroxyvaleric acid), polydioxanone,
degradable polyesters, poly(malic acid), poly(tartronic acid),
poly(acrylamides), polyphosphazenes, poly(amino acids),
poly(alkylene oxide)-poly(ester) block copolymers,
poly(hydroxybutyric acid), poly(beta-butyrolactone),
poly(gamma-butyrolactone), poly(gamma-valerolactone),
poly(d-decanolactone), poly(trimethylene carbonate),
poly(1,4-dioxane-2-one) or poly(1,5-dioxepan-2-one), or
combinations thereof (i.e., copolymers of the constituent monomer
units, blends, etc.).
Examples of biodegradable polymers include synthetic polymers such
as polymers of lactic acid and glycolic acid, polyanhydrides,
poly(ortho)esters, polyurethanes, poly(butyric acid), poly(valeric
acid), and poly(lactide-co-caprolactone), and natural polymers such
as alginate and other polysaccharides including dextran and
cellulose, collagen, chemical derivatives thereof (substitutions,
additions of chemical groups, for example, alkyl, alkylene,
hydroxylations, oxidations, and other modifications routinely made
by those skilled in the art), albumin and other hydrophilic
proteins, zein and other prolamines and hydrophobic proteins,
copolymers and mixtures thereof. In general, these materials
degrade either by enzymatic hydrolysis or exposure to water in
vivo, by surface or bulk erosion
The components of a composition may be distributed homogeneously
throughout the pharmaceutically acceptable carrier, or localized
regions of concentration gradients may exist. By "homogeneous
distribution" is meant to included instances of molecular
homogeneity as well as bulk or macroscopic homogeneity. For
example, the active agent may be homogeneously distributed on a
molecular level (as for a solute homogeneously distributed within a
solvent) or on a macroscopic level (as for discrete particles of
active agent homogeneously distributed throughout the carrier).
Components of a composition may be attached (covalently or
otherwise, including physisorbed, ionically associated, and the
like) to the pharmaceutically acceptable carrier.
For compositions administered as aqueous or other solvent-based
dosage forms (e.g., for parenteral administration), a variety of
liquid carriers may be used. Aqueous solutions may include salts,
buffers, and the like. Non aqueous liquid carriers include, for
example, fatty oils, such as olive oil and corn oil, synthetic
fatty acid esters, such as ethyl oleate or triglycerides, low
molecular weight alcohols such as propylene glycol, synthetic
hydrophilic polymers such as polyethylene glycol, liposomes, and
the like
In addition to one or more pharmaceutically acceptable carrier,
pharmaceutical compositions comprising one or more of the compounds
disclosed herein and suitable for the uses described herein may
also comprise one or more additional components. Additional
components include, for example, salts, buffers, penetration
enhancers, absorption accelerants, gel forming materials such as
polymers, visualization aids, dispersing agents, stabilizers,
excipients, and plasticizers.
Buffers are compounds or solutions that are employed to aid in
maintaining the concentration of an analyte within a desired range.
For example, pharmaceutically acceptable pH buffers are used to
maintain the acidity or basicity of a solution within a
pharmaceutically acceptable range. Buffers for use in the
compositions disclosed herein may be any known or hereafter
discovered buffer.
Penetration enhancers include compounds that enable or enhance
permeation of compositions across boundaries such as membranes.
Examples of penetration enhancers may be found in the relevant
literature (e.g., Percutaneous Penetration Enhancers, Smith and
Maibach, eds., CRC Press, New York N.Y., 2005) and include
cyclohexanone derivatives, cyclic monoterpenes, pyrrolidones,
dioxolanes, 1-dodecylazacycloheptan-2-one (Azone),
dimethylsulfoxide (DMSO), and limonene.
Gel forming materials may be polymers or non-polymers, and are
generally able to form a gelatinous network. In one embodiment, gel
forming materials are able to form gels in vivo, whereas in other
embodiments, gel formation takes place ex vivo. Examples of gel
forming materials include collagen, chitosan, pectins, hyaluronic
acid, and the like.
Dispersing agents are surfactants (for example, as described
herein) in combination with a solvent such as water.
Plasticizers are compounds used to plasticize (i.e., soften)
plastic and other materials. Examples include propylene glycol,
acetyl tributyl citrate, acetyl triethyl citrate,
p-tert-butylphenyl salicylate, butyl stearate, butylphthalyl butyl
glycolate, dibutyl sebacate, di-(2-ethylhexyl)phthalate, diethyl
phthalate, diisobutyl adipate, diisooctyl phthalate,
diphenyl-2-ethylhexyl phosphate, epoxidized soybean oil,
ethylphthalyl ethyl glycolate, glycerol monooleate, monoisopropyl
citrate, mono, di-, and tristearyl citrate, triacetin (glycerol
triacetate), triethyl citrate, and
3-(2-Xenolyl)-1,2-epoxypropane.
Excipients are inactive ingredients that may be employed in the
compositions described herein for a variety of reasons. A wide
range of excipients are described in the literature (e.g., Rowe et
al., Handbook of Pharmaceutical Excipients, McGraw Hill, 2006).
Visualization aids are compounds that aid visualization of the drug
delivery composition or any of the components thereof via a
visualization method such as fluoroscopy, magnetic resonance
imaging (MRI), visible light, ultrasound, or radiography. Any
visualization aids known in the art may be used in the compositions
disclosed herein.
In one aspect, the compositions of the present disclosure include
one or more preservatives or bacteriostatic agents, present in an
effective amount to preserve the composition and/or inhibit
bacterial growth in the composition. Examples include bismuth
tribromophenate, methyl hydroxybenzoate, bacitracin, ethyl
hydroxybenzoate, propyl hydroxybenzoate, erythromycin,
5-fluorouracil, methotrexate, doxorubicin, mitoxantrone, rifamycin,
chlorocresol, benzalkonium chlorides, paraoxybenzoic acid esters,
chlorobutanol, benzylalcohol, phenethyl alcohol, dehydroacetic
acid, sorbic acid, and the like.
Stabilizers include compounds such as antioxidants, and are used to
inhibit or retard decomposition reactions that include, by way of
example, oxidative reactions. Examples of stabilizer include
butylated hydroxytoluene (BHT), butylated hydroxyanisole (BHA),
ascorbic acid, ethylene diamine tetraacetic acid (EDTA),
tocopherol-derived compounds such as alpha-tocopherol, sulfites,
tert-butylhydroquinone, citric acid, acetic acid, and pectin.
The compositions disclosed herein or the precursors thereof may
further contain porosifying agents that achieve greater surface
area of, for example, an implant or tablet. Examples of porosifying
agents include inorganic salts, sucrose, surfactants, small
molecular weight polymers, fast degrading polymers,
thermoreversible polymer precipitates, gas bubbles, and cavitation
bubbles.
The amount of active agent (as well as other active ingredients,
when present) in the compositions disclosed herein will depend on a
number of factors and will vary from subject to subject. Such
factors will be apparent to one of ordinary skill in the art, and
may include the particular disorder or condition being treated, the
mode of administration, the severity of the symptoms, the patient's
age, weight and general condition, and the judgment of the
prescribing physician.
In one embodiment, a composition comprises a compound of formula I
as an active agent and a pharmaceutically acceptable carrier. The
carrier may be used in any convenient amount relative to the active
agent, and the weight ratio of the carrier to active agent can vary
from about 0.1 to 1 to about 100,000 to 1 depending upon the
application. In one example of this embodiment, the composition
consists only of the active agent and a pharmaceutically acceptable
carrier. In another example, the composition comprises the active
agent, a carrier, and one or more additional components such as
those described herein. In a still further example, the composition
comprises the active agent, a second active agent, one or more
carriers, and one or more additional components.
Compounds having the structure of formula I as disclosed herein are
useful as medicaments and as active agents in pharmaceutical
compositions. In one embodiment, such compounds and compositions
are useful in the treatment of viral hepatitis. In particular, the
compounds and compositions are useful in the treatment of patients
suffering from hepatitis B virus (HBV) and hepatitis C virus
(HCV).
In another embodiment, the compounds described herein are useful in
an improved method of treating hepatitis C with an interferon,
wherein the improvement comprises administering an effective amount
of nitazoxanide, tizoxanide, or mixtures thereof to a subject in
need thereof. By way of this improvement, the percentage of
subjects exhibiting reduced serum HCV RNA is increased in
comparison to a method of treating hepatitis C with the interferon
or with a combination of ribavirin and the interferon. In addition,
the amount of interferon required to achieve a sustained virologic
response in the patient may be reduced compared to the amount of
interferon required to achieve a sustained virologic response in
the patient without administration of nitazoxanide, tizoxanide, or
mixtures thereof. Furthermore, the amount of interferon required to
achieve a sustained virologic response in the patient may be
reduced compared to the amount of interferon required to achieve a
sustained virologic response in the patient when treated with a
combination of ribavirin and the interferon. In one embodiment, a
method of treatment is provided wherein a patient suffering from
hepatitis C is pre-treated using nitazoxanide and/or tizoxanide
prior to being treated with an interferon (such as any of the
interferons described herein). Specific examples of this and other
embodiments are described in more detail hereinbelow.
Nitazoxanide, tizoxanide, and mixtures thereof are particularly
effective in the treatment of hepatitis C. By treating hepatitis C
patients with nitazoxanide, tizoxanide, or a mixture thereof, it
may be possible to reduce the amount of interferon needed for
effective treatment, although such reduction is not necessary. It
may also be possible to avoid the use of ribavirin completely,
although this too is not necessary. These benefits may be obtained
while simultaneously increasing the percentage of subjects who
respond favorably in terms of a reduction of serum HCV RNA. Thus,
the present disclosure describes a method of treating hepatitis C
comprising administering to a subject in need thereof an effective
amount of nitazoxanide, tizoxanide, or a mixture thereof.
Similarly, the present invention includes any of the foregoing
embodiments in which any compound of formula I or combination of
such compounds is used in place of nitazoxanide and tizoxanide.
Administration of the compositions described herein may be carried
out using any appropriate mode of administration and dosage form.
Thus, administration can be, for example, oral, ocular, buccal,
rectal, topical, parenteral, transdermal, transmucosal, sublingual,
by inhalation (using either solid or liquid compositions), or via
an implanted reservoir in a dosage form. It will be appreciated
that the most suitable route in any given case will depend on the
nature and severity of the condition being treated and on the
nature of the particular form of compound of formula I which is
being used. The term "parenteral" as used herein is intended to
include, for example, subcutaneous, intravenous, intradermal, and
intramuscular injection. The term "transmuco sal" as used herein is
intended to include, for example, rectal, vaginal, buccal,
sublingual, and penile administration. The term "inhalation" as
used herein is intended to include inhalation via the nose or the
mouth, and includes instances wherein absorption of the composition
occurs in the lungs as well as, for example, the mucosal membranes
of the mouth, nose, and throat. Administration via implants is
meant to include implants affixed anywhere on or positioned
anywhere inside the body, including within body cavities (e.g.,
intraperitoneal implants, intraocular implants, implants in joints,
etc.), within organs, and subcutaneously.
Depending on the intended mode of administration, the
pharmaceutical composition may be a solid, semi-solid, or liquid
such as, for example, a tablet, a capsule, a caplet, an aerosol, a
liquid, a suspension, an emulsion, a cream, a gel, a suppository,
granules, pellets, beads, a film, a powder, a sponge, or the
like.
In one embodiment, the composition comprises a unit dosage form
suitable for single administration of a precise dosage. In another
embodiment, the composition comprises a reservoir such as in an
implant capable of controlled delivery of the composition over
time.
Suitable pharmaceutical compositions and dosage forms may be
prepared using conventional methods known to those in the field of
pharmaceutical formulation and described in the pertinent texts and
literature, e.g., in Remington: The Science and Practice of
Pharmacy (Easton, Pa.: Mack Publishing Co., 1995). A description of
some, but not all, of the suitable dosage forms is provided
infra.
Formulations suitable for oral administration may be presented as
discrete units, such as capsules, cachets, lozenges, or tablets,
each containing a predetermined amount of a compound of formula I;
as a powder or granules; as a solution or a suspension in an
aqueous or non-aqueous liquid; or as an oil-in-water or
water-in-oil emulsion. Such formulations may be prepared by any
suitable method of pharmacy which includes the step of bringing
into association the active compound and a suitable carrier (which
may contain one or more accessory ingredients).
Tablets may be manufactured using standard tablet processing
procedures and equipment. In addition to reversine, tablets will
generally contain inactive, pharmaceutically acceptable carrier
materials as described herein. Suitable capsules may be either hard
or soft, and are generally made of gelatin, starch, or a cellulosic
material, with gelatin capsules preferred. Two-piece hard gelatin
capsules are preferably sealed, such as with gelatin bands or the
like. See, for example, Remington: The Science and Practice of
Pharmacy, cited supra, which describes materials and methods for
preparing encapsulated pharmaceuticals. Oral dosage forms, whether
tablets, capsules, caplets, or particulates, may, if desired, be
formulated so as to provide for gradual, sustained release of the
active agent over an extended time period. For example, as will be
appreciated by those of ordinary skill in the art, dosage forms may
be formulated by dispersing the active agent within a matrix of a
gradually hydrolyzable material such as a hydrophilic polymer, or
by coating a solid, drug-containing dosage form with such a
material.
One example of a preferred dosage form is Alinia.RTM. (see
Alinia.RTM. package insert and/or U.S. Pat. Nos. 5,387,598,
5,578,621, 5,968,961, 5,856,348, 5,859,138, 5,886,013, 5,965,590,
6,020,353, and 6,117,894). It is to be understood that, unless
otherwise specified, in the present disclosure (including the
examples and claims) any references made to Alinia.RTM. are
providing only as examples, and are not meant to be limiting. Thus,
such references are intended to apply equally to other formulations
comprising nitazoxanide, tizoxanide, and/or compounds having the
structure of formula I.
Formulations suitable for buccal (e.g., sub-lingual) administration
include lozenges comprising a compound of formula I, in a flavored
base, usually sucrose and acacia or tragacanth; and pastilles
comprising the compound in an inert base such as gelatin and
glycerin or sucrose and acacia.
Preparations according to this disclosure suitable for parenteral
administration include sterile aqueous and non-aqueous solutions,
suspensions, and emulsions. Such preparations are preferably
isotonic with the blood of the intended recipient. Injectable
aqueous solutions may contain the active agent in water-soluble
form, or may contain a suspension or emulsion of the active agent.
Examples of nonaqueous solvents or vehicles are described herein.
Parenteral formulations may also contain adjuvants such as
solubilizers, preservatives, wetting agents, emulsifiers,
dispersants, and stabilizers, and aqueous suspensions may contain
substances that increase the viscosity of the suspension, such as
sodium carboxymethyl cellulose, sorbitol, and dextran. Injectable
compositions may be rendered sterile via, for example,
incorporation of a sterilizing agent, filtration through a
bacteria-retaining filter, irradiation, or heat. They can also be
manufactured using a sterile injectable medium. Any active agents
present in the compositions may also be in dried, e.g.,
lyophilized, form that may be rehydrated with a suitable vehicle
immediately prior to administration via injection. Parenteral
preparations are preferably administered intravenously, although
administration may also be effected by means of subcutaneous,
intramuscular, or intradermal injection. In one embodiment, such
preparations are prepared by admixing the compound with water or a
glycine buffer and rendering the resulting solution sterile and
isotonic with the blood.
The compositions disclosed herein may also be administered through
the skin using conventional transdermal drug delivery systems,
wherein the active agent is contained within a laminated structure
that serves as a drug delivery device to be affixed to the skin. In
such a structure, the active agent composition is contained in a
layer, or "reservoir," underlying an upper backing layer. The
laminated structure may contain a single reservoir, or it may
contain multiple reservoirs. In one embodiment, the reservoir
comprises a polymeric matrix of a pharmaceutically acceptable
contact adhesive material that serves to affix the system to the
skin during drug delivery. Alternatively, the active
agent-containing reservoir and skin contact adhesive are present as
separate and distinct layers, with the adhesive underlying the
reservoir which, in this case, may be either a polymeric matrix as
described above, or it may be a liquid or hydrogel reservoir, or
may take some other form. Transdermal drug delivery systems may in
addition contain a skin permeation enhancer. Formulations for
transdermal administration may also be delivered by iontophoresis
(see, for example, Pharmaceutical Research 3(6), 318, (1986)) and
suitable formulations typically take the form of an optionally
buffered aqueous solution of a Compound of formula I. Suitable
formulations comprise, for example, citrate or bis/tris buffer (pH
6) or ethanol/water and contain from 0.1 to 0.2M active
ingredient.
The compositions disclosed herein may also be administered
topically using conventional topical dosage forms, wherein the
active agent is contained within a carrier. Dosage forms suitable
for topical application include, by way of example, creams, pastes,
jellies, gels, ointments, liquids, aerosols, oils, lotions, foams,
suspensions, and emulsions. Carriers which may be used include
vaseline, lanoline, polyethylene glycols, alcohols, and
combinations of two or more thereof.
In addition to the formulations described previously, the compounds
may also be formulated as a depot preparation for controlled
release of the active agent, preferably sustained release over an
extended time period. These sustained release dosage forms may be
administered by implantation (e.g., subcutaneously,
intraperitoneal, intramuscularly or by intramuscular
injection).
Formulations suitable for rectal administration are preferably
presented as unit dose suppositories. These may be prepared by
admixing a Compound of formula I with one or more conventional
solid carriers, for example, cocoa butter, and then shaping the
resulting mixture.
Although the compositions disclosed herein will generally be
administered orally, parenterally, transdermally, or via an
implanted depot, other modes of administration are suitable as
well. For example, administration may be rectal or vaginal,
preferably using a suppository that contains, in addition to an
active agent, excipients such as a suppository wax. Formulations
for nasal or sublingual administration are also prepared with
standard excipients well known in the art. The pharmaceutical
compositions of the invention may also be formulated for
inhalation, e.g., as a solution in saline, as a dry powder, or as
an aerosol.
It will be appreciated that the compositions disclosed herein may
be prepared and packaged as single dosage units, such as for oral
administration (e.g., tablets). The formulations may also be
prepared and packaged as multiple dose formulations, or as dosages
suitable for long-term administration, such as for topical
administration (e.g., creams), transmembrane administration (e.g.,
patches), or implantation.
The compounds disclosed herein may be administered for any length
of time suitable for the intended use. Administration of the
compounds disclosed herein will typically be carried out over a
period of about 3 days to about 104 weeks, but may be carried out
over a period longer than 104 weeks and may even be carried out
indefinitely. For example, treatment of hepatitis C using the
compounds disclosed herein will typically involve administration of
the compounds over a period of 12, 24, or 48 weeks.
Any appropriate dosage and regimen may be used for the compounds
disclosed herein and the pharmaceutical compositions comprising
such compounds. In one embodiment, a compound having the structure
of formula I is administered in conjunction with an additional
active agent such as, for example, an interferon such as any of the
interferons described herein. The compound having the structure of
formula I and the additional active agent (e.g., an interferon) may
be administered as part of the same composition, or they may be
administered in separate compositions (including in separate
compositions that vary in dosage form, release profiles, and the
like).
In one embodiment, a patient suffering from hepatitis C is first
pretreated with nitazoxanide, tizoxanide, or any of the compounds
disclosed herein having the structure of formula I. The duration of
the pretreatment period may be between about 3 days and about 6
months, for example between about 1 week and about 12 weeks, and as
a further example between about 1 week and about 4 weeks. The
pretreatment period is followed subsequently by a treatment period
wherein the pretreated patient is treated with either an interferon
alone or an interferon plus nitazoxanide, tizoxanide, or any of the
compounds having the structure of formula I. Any of the interferons
described herein may be used during the treatment period. The
duration of the treatment period will be any duration that is
required to obtain the desired response, and will typically be
between about 1 day and about 12 months or longer. For example, the
treatment period may comprise weekly injections of an interferon,
and may involve a single week of treatment, 2-4 weeks of treatment,
4-12 weeks of treatment, or more (such as 6 months, 1 year, 2
years, or indefinitely).
Examples of regimens that are suitable for administration of the
compounds disclosed herein include the following: 24 weeks of
administration of nitazoxanide followed by 12 weeks of
administration of a composition comprising nitazoxanide and
interferon .alpha.-2b or pegylated interferon .alpha.-2b; 2-4 weeks
of administration of nitazoxanide followed by 12 weeks of
administration of a composition comprising nitazoxanide and
pegylated interferon .alpha.-2b; administration of a composition
comprising nitazoxanide+pegylated interferon .alpha.-2b for 12, 24,
or 48 weeks; and 12, 24, or 48 weeks of administration of
nitazoxanide, tizoxanide, or combinations thereof. It will be
appreciated that such regimens are provided only as examples, as
suitable durations, dosages, and orders of administration will
vary. Appropriate regimens will typically be determined by a
physician.
It will be appreciated that dosages may vary, and will typically be
selected to provide a therapeutically effective amount of the
active agent to the patient. In one example, a dosage may be in the
range of about 100 mg to about 2000 mg, or in the range of about
250 mg to about 1000 mg, or preferably about 500 mg. In another
specific example, an appropriate dosage is chosen to achieve and
maintain a blood level of active agent (e.g., nitazoxanide) in the
patient that is between about 0.1 .mu.g/ml and about 10 .mu.g/ml,
preferably about 1 .mu.g/ml.
Methods of preparation for the compositions disclosed herein will
be apparent to one of ordinary skill. In one embodiment, the
formulations of the disclosure may be prepared by uniformly and
intimately admixing the active compound with a liquid or finely
divided solid carrier, or both, and then, if necessary, shaping the
resulting mixture. For example, a tablet may be prepared by
compressing or molding a coated or uncoated powder or coated or
uncoated granules containing the active compound, optionally with
one or more accessory ingredients. Compressed tablets may be
prepared by compressing, in a suitable machine, the compound in a
free-flowing form, such as a powder or granules optionally mixed
with a binder, lubricant, inert diluent, and/or surface
active/dispersing agent(s). Molded tablets may be made by molding,
in a suitable machine, the powdered compound moistened with an
inert liquid binder.
The present disclosure also provides kits for accomplishing such
treatment as described herein. The kits comprise: (i) an effective
amount of a compound of formula I; (ii) one or more
pharmaceutically acceptable carriers and/or additives; and (iii)
instructions for use (e.g., in treating hepatitis).
As used herein, the phrase "instructions for use" shall mean any
FDA-mandated labelling, instructions, or package inserts that
relate to the administration of a compound of Formula I for the
purpose of treating viral hepatitis. For example, instructions for
use may include, but are not limited to, indications for the
particular disease, identification of specific symptoms of the
specific disease that can be ameliorated by the claimed compounds,
and recommended dosage amounts for subjects suffering from the
disease. The kit of the present invention further comprises a unit
dosage amount of the compound effective for treating viral
hepatitis.
It is to be understood that while the invention has been described
in conjunction with the preferred specific embodiments thereof, the
description above as well as the examples that follow are intended
to illustrate and not limit the scope of the invention. Other
aspects, advantages and modifications within the scope of the
invention will be apparent to those skilled in the art to which the
invention pertains.
All patents, patent applications, and publications mentioned herein
are hereby incorporated by reference in their entireties. However,
where a patent, patent application, or publication containing
express definitions is incorporated by reference, those express
definitions should be understood to apply to the incorporated
patent, patent application, or publication in which they are found,
and not to the remainder of the text of this application, in
particular the claims of this application.
EXAMPLES
Example 1
Activity Against HCV Replication
Antiviral activity of nitazoxanide, tizoxanide, interferon .alpha.,
ribavirin and 2'-C-methyl cytidine was assessed in five different
HCV replicon cell lines: (1) AVA5, a subgenomic construct of
genotype 1b (Blight et al., 2000, Science 290:1972-1974); (2)
H/FL-Neo, a genotype 1a full length construct (Blight et al., 2003,
Journal of Virology 77:3181-3190); (3) JWT, a subgenomic construct
of genotype 1b (Pfeiffer and Kirkegaard, 2005, Journal of Virology,
79:2346-2355); (4) 4-3-10, a subgenomic construct of genotype 1b,
developed by a protocol that involved serial passage of JWT cells
in 100 .mu.M for one month followed by 400 .mu.M ribavirin for two
weeks (Pfeiffer and Kirkegaard, 2005, Journal of Virology,
79:2346-2355); and (5) RP7, a subgenomic construct of genotype 1b
(Elazar et al., 2003, Journal of Virology 77:6055-6061).
Antiviral activity for each test compound was determined as
previously described (Okuse et al., 2005, Antiviral Research
65:23-34). Briefly, replicon cell lines were maintained as
sub-confluent cultures on 96-well plates. Compounds were added
daily for three days in fresh medium. Twenty-four hours after the
last dose of compound, antiviral activity was determined by blot
hybridization analysis of intracellular HCV RNA, and cytotoxicity
was assessed by neutral red dye uptake. EC.sub.50, EC.sub.90,
CC.sub.50 and selectivity index were calculated for each compound
tested in a replicon cell line. EC.sub.50=drug concentration
producing a 50% reduction of intracellular HCV RNA relative to the
average levels in untreated cultures. EC.sub.90=drug concentration
producing a 90% reduction of intracellular HCV RNA relative to the
average levels in untreated cultures. CC.sub.50=drug concentration
producing a 50% reduction of neutral red dye uptake relative to the
average levels in untreated cultures. Selectivity index=CC.sub.50
divided by EC.sub.50. EC.sub.50, EC.sub.90 and CC.sub.50 values
(.+-.standard deviations [S.D.]) were calculated by linear
regression analysis using data combined from all treated cultures.
Median EC.sub.50 and EC.sub.90 values were calculated for each
compound based on the results for determined for the five different
replicon cell lines.
Nitazoxanide and tizoxanide were provided by Romark Laboratories,
L.C. (Tampa, Fla. USA). Recombinant interferon .alpha.-2b was
purchased from PBL Biomedical Laboratories (Piscataway, N.J. USA).
Ribavirin was purchased from Sigma-Aldrich (St. Louis, Mo. USA).
2'-C-methyl cytidine (Pierra, et al. 2005, Nucleosides Nucleotides
Nucleic Acids, 24:767-770) was purchased from Moraveck
Biochemicals, Inc. (La Brea, Calif. USA). Interferon .alpha.-2b was
solubilized and/or diluted in sterile phosphate-buffered saline
(PBS)/1% BSA as instructed by the manufacturer. Ribavirin,
nitazoxanide, tizoxanide and 2'-C-methyl cytidine were solubilized
in 100% tissue culture grade DMSO (Sigma). Stock solutions were
stored (-70.degree. C. for interferon .alpha.-2b, -20.degree. C.
for nitazoxanide, tizoxanide, ribavirin and 2'C-methyl cytidine) in
quantities sufficient for a single experiment and used only once.
Daily aliquots of test compounds were made from the stock solutions
in individual tubes and stored at the appropriate temperatures. On
each day of treatment, daily aliquots of the test compounds were
suspended into culture medium at room temperature, and immediately
added to the cell cultures, thereby subjecting each aliquot of test
compound to the same, limited, number of freeze-thaw cycles.
Nitazoxanide and tizoxanide selectively reduced intracellular HCV
replication in each of the five HCV genotype 1-derived replicon
cell lines (Table 1). Median EC.sub.50s were 0.13 .mu.M and 0.15
.mu.M for nitazoxanide and tizoxanide, respectively, compared to
0.86 IU/mL for interferon .alpha.-2b, 69 .mu.M for ribavirin and
2.1 .mu.M for 2'-C-methyl cytidine.
TABLE-US-00001 TABLE 1 Relative potency of test compounds against
HCV replication. Drug Cell line EC.sub.50 (.mu.M) EC.sub.90 (.mu.M)
CC.sub.50 (.mu.M) S.I..sup.1 Nitazoxanide AVA5 0.13 .+-. 0.02 1.0
.+-. 0.2 39 .+-. 3.9 300 H/FL-Neo 0.33 .+-. 0.05 1.1 .+-. 0.1 49
.+-. 1.5 149 JWT 0.11 .+-. 0.01 1.0 .+-. 0.2 39 .+-. 1.0 354 4-3-10
0.10 .+-. 0.03 0.87 .+-. 0.16 34 .+-. 0.4 340 RP7 0.16 .+-. 0.01
1.2 .+-. 0.1 38 .+-. 0.8 238 Median 0.13 1.0 Tizoxanide AVA5 0.12
.+-. 0.01 0.77 .+-. 0.10 25 .+-. 2.8 208 H/FL-Neo 0.25 .+-. 0.03
1.0 .+-. 0.1 4.2 .+-. 0.2 17 JWT 0.16 .+-. 0.02 0.76 .+-. 0.03 24
.+-. 2.9 150 4-3-10 0.11 .+-. 0.05 0.55 .+-. 0.08 21 .+-. 1.1 191
RP7 0.15 .+-. 0.01 0.94 .+-. 0.10 25 .+-. 1.0 167 Median 0.15 0.77
Interferon .alpha.-2b (IU/ml) AVA5 1.5 .+-. 0.2 8.2 .+-. 0.8
>10000 >6667 H/FL-Neo 2.1 .+-. 0.2 9.4 .+-. 0.9 >10000
>4762 JWT 0.77 .+-. 0.03 2.6 .+-. 0.2 >10000 >12987 4-3-10
0.86 .+-. 0.06 5.7 .+-. 0.4 >10000 >11627 RP7 0.41 .+-. 0.01
3.6 .+-. 0.2 >10000 >24390 Median 0.86 5.7 Ribavirin AVA5 70
.+-. 0.5 220 .+-. 34 84 .+-. 4.7 1.2 H/FL-Neo JWT 23 .+-. 2.2 62
.+-. 1.7 89 .+-. 7.5 3.9 4-3-10 RP7 69 .+-. 4.4 122 .+-. 13 77 .+-.
4.9 1.2 Median 69 122 2'-C-methyl cytidine AVA5 2.1 .+-. 0.2 8.1
.+-. 0.7 >300 >143 H/FL-Neo 1.8 .+-. 0.2 8.1 .+-. 0.8
>1000 >556 JWT 2.2 .+-. 0.1 8.2 .+-. 0.7 >300 >136
4-3-10 2.1 .+-. 0.1 8.0 .+-. 0.9 >300 >143 RP7 2.0 .+-. 0.1
9.0 .+-. 0.6 >300 >150 Median 2.1 8.1 .sup.1S.I. (Selectivity
Index) = CC.sub.50/EC.sub.50
Example 2
Synergistic Activity of Nitazoxanide and Tizoxanide with Other
Anti-HCV Drugs
Activity of combination treatments with nitazoxanide plus
interferon .alpha.-2b, tizoxanide plus interferon .alpha.-2b,
nitazoxanide plus 2'-C-methyl cytidine and tizoxanide plus
2'-C-methyl cytidine against HCV replication were evaluated in the
AVA5 replicon cell line using the methods previously described
(Okuse et al., 2005, Antiviral Research 65:23-34). Analyses of
interactions between compounds used in combination treatments were
performed using Calcusyn.TM. software (Biosoft, Cambridge, UK).
Combinations of nitazoxanide with either interferon .alpha.-2b or
2'-C-methyl cytidine and tizoxanide with either interferon
.alpha.-2b or 2'-C-methyl cytidine exhibited synergistic
interactions against HCV replication (Table 2, FIGS. 1a and 1b). In
FIGS. 1a and 1b analyses of interactions between compounds in
combination treatments are shown.
TABLE-US-00002 TABLE 2 Relative potency of combination treatments
against HCV replication in AVA5 cell cultures. Treatment EC.sub.50
(.mu.M) EC.sub.90 (.mu.M) CC.sub.50 (.mu.M) S.I..sup.1 Nitazoxanide
0.21 .+-. 0.03 0.93 .+-. 0.11 38 .+-. 1.8 181 (NTZ) Tizoxanide 0.15
.+-. 0.02 0.81 .+-. 0.92 15 .+-. 1.2 100 (TIZ) IFN.alpha.-2b 1.9
.+-. 0.22 8.9 .+-. 0.92 >100002 >5263 2'-C-methyl 1.6 .+-.
0.2 8.3 .+-. 0.7 >300 >188 cytidine (2'CMeC) 2'CMeC + 0.67
.+-. 0.007 2.3 .+-. 0.3 >300 >448 IFN.alpha.-2b, 1:1 NTZ +
0.06 .+-. 0.008 0.25 .+-. 0.03 33 .+-. 1.3 550 IFN.alpha.-2b, 1:10
NTZ + 0.07 .+-. 0.005 0.28 .+-. 0.02 35 .+-. 1.5 500 2'CMeC, 1:10
TIZ + 0.07 .+-. 0.01 0.22 .+-. 0.03 17 .+-. 1.3 245 IFN.alpha.-2b,
1:10 TIZ + 0.06 .+-. 0.004 0.19 .+-. 0.02 18 .+-. 1.1 300 2'CMeC,
1:10 .sup.1SI = CC.sub.50/EC.sub.50. .sup.2Values for IFN.alpha.-2b
expressed in IU/mL
FIG. 1a presents CI-Fa (Combination Index-Fraction (of virus)
affected) plots (Belen'kii and Schinazi, 1994, Antiviral Research
25:11-18). For these plots, a combination index [CI] greater than
1.0 indicates antagonism and a CI less than 1.0 indicates
synergism. Evaluations of synergy, additivity (summation), or
antagonism at different levels of virus inhibition (e.g. 5%, or
Fa=0.05 to 99%, or Fa=0.99) are provided by the plotted lines and
points. FIG. 1b shows conservative isobolograms. For these plots,
EC.sub.50, EC.sub.75, and EC.sub.90 (50%, 75%, and 90% effective
antiviral concentrations) values for the combination treatments are
displayed as single points. Three lines radiating out from the axes
denote the expected (e.g. additive) EC.sub.50, EC.sub.75, and
EC.sub.90 values for drug combinations as calculated from the
monotherapies. EC.sub.50, EC.sub.75, and EC.sub.90 values for the
combinations that plot to the left (e.g. less than) of the
corresponding lines indicate synergy, and values plotting to the
right (e.g. greater than) of the corresponding lines indicate
antagonism.
Example 3
Enhanced Activity of Interferon Alpha+Nitazoxanide after
Pre-Treatment with Nitazoxanide
To evaluate the effect of pre-treating with nitazoxanide prior to
treatment with combination treatments, cultures were treated for
either 3 or 6 days with nitazoxanide, interferon .alpha.-2b, or
2'-C-methyl cytidine or combinations of nitazoxanide and either
interferon .alpha.-2b or 2'-C-methyl cytidine. Alternatively,
cultures were treated with nitazoxanide for 3 days, followed by an
additional 3 days of treatment with a combination of nitazoxanide
and either interferon .alpha.-2b or 2'-C-methyl cytidine. Antiviral
activity and cytotoxicity was determined 24 hours after the end of
each respective treatment as described previously.
Pre-treatment with nitazoxanide improved the potency of combination
treatment with nitazoxanide plus interferon .alpha.-2b by
approximately 3-fold (Table 3 and FIGS. 2a and 2b). Pre-treatment
did not, however, affect the potency of combination treatment with
2'-C-methyl cytidine (Table 4). FIGS. 2a and 2b show analyses of
the effect in cultures pre-treated with nitazoxanide before
treatment with nitazoxanide plus interferon .alpha.-2b. Analyses
were performed using Calcusyn.TM. software (Biosoft, Cambridge,
UK). Two types of evaluations are presented. FIG. 2a presents CI-Fa
(Combination Index-Fraction (of virus) affected) plots (Belen'kii
and Schinazi, 1994). For these plots, a combination index [CI]
greater than 1.0 indicates antagonism and a CI less than 1.0
indicates synergism. Evaluations of synergy, additivity
(summation), or antagonism at different levels of virus inhibition
(e.g. 5%, or Fa=0.05 to 99%, or Fa=0.99) are provided by the
plotted lines and points. Dotted lines indicate 1.96 standard
deviations (not shown in FIG. 1a for clarity). FIG. 2b presents
conservative isobolograms. For these plots, EC.sub.50, EC.sub.75,
and EC.sub.90 (50%, 75%, and 90% effective antiviral
concentrations) values for the combination treatments are displayed
as single points. Three lines radiating out from the axes denote
the expected (e.g. additive) EC.sub.50, EC.sub.75, and EC.sub.90
values for drug combinations as calculated from the monotherapies.
EC.sub.50, EC.sub.75, and EC.sub.90 values for the combinations
that plot to the left (e.g. less than) of the corresponding lines
indicate synergy, and values plotting to the right (e.g. greater
than) of the corresponding lines indicate antagonism.
TABLE-US-00003 TABLE 3 Effect of NTZ Pretreatment on Activity of
NTZ + IFN.alpha. Combination Treatment Dur- ation NTZ (.mu.M)
IFN.alpha.-2b (IU/mL) Treatment (days) EC.sub.50 (.mu.M) EC.sub.90
(.mu.M) EC.sub.50 (.mu.M) EC.sub.90 (.mu.M) IFN.alpha. 3 1.9 .+-.
0.3 8.3 .+-. 0.9 IFN.alpha. 6 1.7 .+-. 0.2 7.8 .+-. 0.8 NTZ 3 0.22
.+-. 0.03 1.0 .+-. 0.1 NTZ 6 0.20 .+-. 0.02 0.92 .+-. 0.10 NTZ + 3
0.08 .+-. 0.010 0.27 .+-. 0.03 0.82 .+-. 0.07 2.7 .+-. 0.3
IFN.alpha., 1:10 NTZ + 6 0.09 .+-. 0.010 0.24 .+-. 0.04 0.75 .+-.
0.09 2.4 .+-. 0.2 IFN.alpha., 1:10 NTZ, then 6 0.03 .+-. 0.004 0.09
.+-. 0.011 0.31 .+-. 0.04 0.96 .+-. 0.12 NTZ + IFN.alpha.
TABLE-US-00004 TABLE 4 Effect of NTZ Pretreatment on Activity of
NTZ + 2'CMeC Combination Treatment Dur- ation NTZ (.mu.M) 2'CMeC
(.mu.M) Treatment (days) EC.sub.50 (.mu.M) EC.sub.90 (.mu.M)
EC.sub.50 (.mu.M) EC.sub.90 (.mu.M) 2'CMeC 3 1.7 .+-. 0.2 6.2 .+-.
0.5 2'CMeC 6 1.3 .+-. 0.2 5.8 .+-. 0.9 NTZ 3 0.22 .+-. 0.03 1.0
.+-. 0.1 NTZ 6 0.20 .+-. 0.02 0.92 .+-. 0.10 NTZ + 3 0.05 .+-.
0.006 0.16 .+-. 0.02 0.57 .+-. 0.07 1.8 .+-. 0.2 2'CMeC, 1:10 NTZ +
6 0.05 .+-. 0.007 0.17 .+-. 0.03 0.54 .+-. 0.06 1.9 .+-. 0.2
2'CMeC, 1:10 NTZ, then 6 0.06 .+-. 0.005 0.15 .+-. 0.02 0.58 .+-.
0.08 1.7 .+-. 0.3 NTZ + 2'CMeC
Example 4
Enhanced Activity of Interferon Alpha after Pre-Treatment with
Nitazoxanide or Tizoxanide
To evaluate the effect of interferon alpha following pre-treatment
with nitazoxanide or tizoxanide, a parental replicon-containing
cell line (RP-7) was serially passaged in increasing concentrations
of nitazoxanide or tizoxanide. Anti-HCV activity of interferon
alpha-2b was determined using the parental cell line and using the
cell lines obtained after passage in nitazoxanide or tizoxanide.
Anti-HCV activity was determined by the methods described
above.
The parental replicon-containing cell line was established by
electroporation of RNA transcribed in vitro off of the
Sca-I-linearized Bart 79I plasmid into Huh-7 cells (Elazar et al.,
2003). Bart79I encodes for a second-generation high-efficiency
bi-cistronic sub-genomic replicon of genotype 1b containing a
single adaptive mutation (S1179I) in the NS5A gene, and the
neomycinphosphotransferase gene in the first cistron. The
electroporated cells were plated along with naive Huh-7 feeder
cells and grown in medium--DMEM (4.5 g/l glucose, L-glutamine and
sodium pyruvate--Mediatech 10-013-CV), 10% fetal bovine serum, 1%
Penicillin-streptomycin, 1% L-glutamine (final concentration 2 mM),
1.times.MEM Non-Essential Amino Acids (100.times.)
(Invitrogen)--and 1 mg/ml G418. After 3 weeks, G418-resistant
colonies appeared. One of the resulting colonies was isolated,
expanded, passaged in 700 .mu.g/ml G418, and termed RP-7.
RP-7 cells were subjected to a resistance-promoting regimen as
follows. The cells were grown in the medium described above
containing 700 .mu.g/ml G418 (Invitrogen), 1% tissue culture grade
DMSO (Sigma), and an initial low concentration of nitazoxanide or
tizoxanide which was then steadily increased every week, with an
intervening 2-day drug holiday in between each dose increase. On
days 1 through 5 of each dose of drug, the media was changed daily
to provide a source of fresh drug. No media changes were performed
on days 6 and 7 (the drug holiday). The initial concentration of
nitazoxanide or tizoxanide was 0.02 .mu.M, followed by 0.05 .mu.M.
0.1 .mu.M, 0.5 .mu.M, 1 .mu.M, and subsequent weekly increases of 1
.mu.M. A final concentration of 11 .mu.M was used for the cells
passaged in nitazoxanide while a final concentration of 8 .mu.M was
used for cells passaged in tizoxanide. The resulting cells were
subsequently passaged at this final concentration for at least 2
months prior to being used to test the anti-HCV activity of
interferon alpha-2b.
Results are presented in Table 5. Serial passage of the parental
cell line in increasing concentrations of nitazoxanide or
tizoxanide did not induce resistance to interferon alpha-2b. The
cell lines passaged in nitazoxanide or tizoxanide were actually 2.5
to 7.6-fold more susceptible to interferon alpha-2b than the
parental replicon-containing cell line, which was not passaged in
nitazoxanide or tizoxanide.
TABLE-US-00005 TABLE 5 Potency of Interferon .alpha.-2b Against HCV
Replication in RP7 Cells Before and After Serial Passage in
Increasing Concentrations of Nitazoxanide and Tizoxanide Cell line
EC.sub.50 (.mu.M) EC.sub.90 (.mu.M) CC.sub.50 (.mu.M) SI Parental
cell 0.41 .+-. 0.01 3.6 .+-. 0.2 >10000 >24390 line (RP-7)
RP7 cells 0.11 .+-. 0.02 0.47 .+-. 0.04 >10000 >90909
passaged in nitazoxanide RP7 cells 0.16 .+-. 0.01 0.42 .+-. 0.04
>10000 >62500 passaged in tizoxanide
Example 5
Treatment of Chronic Hepatitis C with a Combination of Nitazoxanide
and Tizoxanide
Fifty (50) patients were enrolled in a double-blind study of
Alinia.RTM. (pharmaceutical composition comprising 99% nitazoxanide
and 1% tizoxanide as active agents) administered orally as a 500 mg
tablet twice daily for 24 weeks compared to a placebo in treating
patients with chronic hepatitis C genotype 4. The 50 patients were
enrolled at three study sites in Egypt: 32 at Cairo, 12 at
Alexandria and 6 at Tanta. Three patients dropped out of the study
immediately after enrollment and did not return for any
post-treatment follow-up. One patient did not return for follow-up
after week 12. Each of the remaining 46 patients completed the
study. See FIG. 3 for a Patient Disposition Flowchart. One patient
was co-infected with hepatitis B virus. The patient was
HBeAg-negative, and an exception was made to allow enrollment of
this patient. The protocol called for use of an intent-to-treat
population (all patients randomized) for the primary efficacy
analysis. The three patients that dropped out before receiving any
medication were excluded from the efficacy analysis. The patient
who dropped out after week 12 was included in the efficacy analysis
and analyzed on the basis of last observation carried forward.
Demographic and disease-related characteristics for the 47 patients
included in the efficacy analysis is summarized by treatment group
in Table 6.
At each study visit, the patients were questioned regarding
treatment compliance. With one exception, each of the patients
completing the study reported that they had been compliant with
taking the medication. One patient completed the study but reported
sporadic noncompliance with taking medication due to abdominal
pain.
TABLE-US-00006 TABLE 6 Demographic and Disease-Related
Characteristics All Subjects Active Placebo P.sup.1 Race: Caucasian
47 23 24 1.0 Gender: Male/Female 39/8 19/4 20/4 1.0 Age (years):
Mean .+-. SD 47.3 .+-. 9.3 49.7 .+-. 8.4 45.0 .+-. 9.6 .08 Median
(Range) 48 (27-67) 51 (35-67) 46 (27-64) Weight (kgs): Mean .+-. SD
86.2 .+-. 18.8 84.8 .+-. 16.7 87.5 .+-. 21.0 .62 Median (Range) 84
(64-143) 84 (64-130) 82 (65-143) Body Mass Index: Mean .+-. SD 29.4
.+-. 5.5 29.0 .+-. 5.1 29.8 .+-. 6.0 .62 Median (Range) 28.2
(21-47) 27.3 (22-47) 28.3 (21-46) Viral load (log10 IU/mL): Mean
.+-. SD 5.2 .+-. 0.7 5.3 .+-. 0.7 5.2 .+-. 0.8 .43 Median (Range)
5.3 (3.5-6.5) 5.4 (4.0-6.3) 5.3 (3.5-6.5) Viral load >800,000 10
6 4 .49 IU/mL Elevated ALT 31 13 18 .23 Necroinflammatory score:
Mean .+-. SD 6.0 .+-. 3.2 6.3 .+-. 3.3 5.7 .+-. 2.7 .51 Median
(Range) 5 (2-17) 5 (3-17) 5.5 (2-11) Liver disease: No fibrosis 8 4
4 .95 Fibrous portal 18 8 10 expansion Bridging fibrosis 14 7 7
Cirrhosis (com- 3 1 2 pensated) Cirrhosis (decom- 4 3 1 pensated)
Previously treated with 5 3 2 .67 peginterferon/ribavirin Diabetes
mellitus Controlled 7 4 3 .70 Uncontrolled 3 1 2 1.0 .sup.1Fisher's
exact test or chi-square test used for comparing proportions,
t-test for means.
Virologic responses are summarized by treatment group in Table 7.
The proportion of virologic responders in the active treatment
group was significantly higher than in the placebo treatment group
(P=0.0039). Virologic responses (undetectable serum HCV RNA) were
observed at weeks 4 (n=3), week 8 (n=3) and week 20 (n=1). Each of
these responses were maintained throughout the treatment
period.
TABLE-US-00007 TABLE 7 Virologic Responses by Treatment Group
Active Placebo P.sup.1 Responders/Total (%) 7/23 (30.4%) 0/24 (0%)
0.0039 .sup.1two-sided Fisher's exact test
Demographic characteristics, baseline laboratory data, data from
liver biopsies and medical histories were evaluated to identify
independent predictors of virologic response within the active
treatment group. Predictors of response are listed in Table 8. The
most significant predictor of response was lower viral load at
baseline. All responders had baseline viral loads.ltoreq.384,615
IU/mL. Laboratory values at baseline (platelet counts, prothrombin
time and alfa fetoprotein) also suggested that the responders had
less severe liver disease.
TABLE-US-00008 TABLE 8 Independent Predictors of Response
Predictors of Response P Lower viral load at baseline .0086
Indicators of less serious liver disease Higher platelet counts
.0385 Lower prothrombin time .0579 Lower alfa fetoprotein .0696
Further analysis of patients with complicating disease-related
factors such as high viral loads, cirrhosis, uncontrolled diabetes
mellitus or hepatitis B co-infection showed very poor response
rates in these subsets of patients (see Table 9). Fifteen (15) of
the 16 Alinia.RTM. treatment failures had high viral load, advanced
liver disease, uncontrolled diabetes mellitus or hepatitis B virus
co-infection. The Alinia.RTM. responders can, therefore, be
described as patients with low viral loads (<800,000 IU/mL)
whose disease had not advanced to cirrhosis and who did not have
uncontrolled diabetes mellitus or hepatitis B virus co-infection.
Two (2) virologic responders in the active treatment group had a
prior history of treatment with peginterferon/ribavirin. One was
unable to tolerate peginteferon/ribavirin and discontinued therapy
after 5 weeks. The other relapsed following completion of 48 weeks
of peginterferon/ribavirin.
TABLE-US-00009 TABLE 9 Response Rates in Patients with Complicating
Disease-Related Factors Responders/ Complicating disease-related
factors Total High viral load (>800,000 IU/mL) 0/3 Advanced
liver disease: cirrhosis 0/3 Advanced liver disease: bridging
fibrosis 3/5 Uncontrolled diabetes mellitus 0/3 Hepatitis B virus
co-infection 0/1 High viral load and cirrhosis 0/1 High viral load
and bridging fibrosis 0/1 High viral load, uncontrolled diabetes
and 0/1 bridging fibrosis
Sustained Virologic Response: The 7 virologic responders were
followed up at least 24 weeks after the end of treatment, and 5 of
these patients had a sustained virologic response (undetectable
serum HCV RNA) at follow-up. Sustained virologic response rates are
presented by treatment group in Table 10. Two patients failed to
maintain their virologic responses off-treatment. One patient only
completed 8 weeks of treatment. One patient completed the study,
but reported sporadic noncompliance with taking medication due to
abdominal pain. Each of these two patients had advanced liver
disease (bridging fibrosis).
TABLE-US-00010 TABLE 10 Sustained Virologic Responses by Treatment
Group Active Placebo P* Responders/Total (%) 5/23 (21.7%) 0/24 (0%)
0.0219 *two-sided Fisher's exact test
Changes in Quantitative Serum HCV RNA (Viral Load): Mean
quantitative viral loads for the active treatment group, the
placebo treatment group, active treatment group virologic
responders, and active treatment group virologic failures are
presented in Table 11 and FIG. 4. Reduction of the mean
quantitative viral load from baseline to end of treatment was
significantly greater for the active treatment group (reduction of
1.55.+-.2.34 log.sub.10 IU/mL) than for the placebo group
(reduction of 0.21.+-.0.98 log.sub.10 IU/mL) observed for the
placebo treatment group (P=0.0166, t-test). The reduction in mean
viral load observed for the active treatment group was entirely
attributed to the virologic responders. Changes in viral loads of
nonresponders were not significantly different than changes noted
for the placebo treatment group. Actual quantitative viral loads
for the 7 virologic responders over time are presented in Table 12
and FIG. 5.
TABLE-US-00011 TABLE 11 Mean Quantitative Serum HCV RNA over Time
by Treatment Group and Virologic Response (Log.sub.10 IU/mL) Week
Week Week Week Baseline Week 4 Week 8 12 16 20 24 Alinia Non- 5.5
5.21 5.21 5.23 5.54 5.61 5.42 responsders Placebo 5.16 5.17 4.73
4.96 5.15 5.13 4.94 Alinia 5.33 4.53 3.87 3.9 4.02 3.9 3.77 Alinia
4.92 2.98 0.80 0.85 0.53 * * Responders * All values below lower
limit of detection (10 IU/mL)
TABLE-US-00012 TABLE 12 Quantitative Serum HCV RNA over Time for
Virologic Responders (Log.sub.10 IU/mL) Week Week Week Patient
Baseline Week 4 Week 8 Week 12 16 20 24 #1 4.37 * * * * * * #6 5.59
5.64 * * * * * #15 5.22 5.43 5.57 5.98 3.74 * * #17 5.30 * * * * *
* #21 5.00 4.23 * * * * * #37 4.70 * * * * * * #40 4.25 5.56 * * *
* * * Below limit of detection (10 IU/mL)
Changes in ALT: Mean changes in ALT from baseline to end of
treatment were not significantly different for the two treatment
groups (-3.9.+-.32 for the active treatment group and -1.3.+-.42
for the placebo group, P=0.82, t test). Categorical changes in ALT
from baseline to end of treatment are summarized by treatment group
in Table 13. Three of the virologic responders in the active
treatment group had normal ALT values at baseline, which remained
normal at the end of treatment. One of the four virologic
responders with elevated ALT at baseline had normal ALT at the end
of treatment while the ALT for the other 3 remained elevated. Four
of the five patients with sustained virologic responses also had
normal ALT after 24 weeks off-treatment.
TABLE-US-00013 TABLE 13 Change in ALT from Baseline to End of
Treatment Active Placebo Normalized 3 2 Remained Normal 7 4
Remained Elevated 10 16 Normal to Elevated 3 2
Quantitative HCV RNA values were missing for one patient at week 24
and for one patient at weeks 12, 16, 20 and 24. End of treatment
data for these patients was analyzed using the last data point
available (last observation carried forward). An interim analysis
of end of treatment virologic response was conducted for the first
21 patients enrolled in the study. For purposes of this report, no
adjustments have been made to account for multiple analyses.
Virologic response rates are presented by treatment group by study
center in Table 14. The higher response rate observed in the active
treatment group for the Cairo study center is attributed to
disease-related characteristics of patients enrolled at the
different sites. Each of the 9 patients enrolled in the active
treatment group at the Alexandria and Tanta centers had high viral
loads (>800,000 IU/mL), advanced liver disease, uncontrolled
diabetes mellitus or hepatitis B virus co-infection.
TABLE-US-00014 TABLE 14 Virologic Response by Treatment Group and
Study Center Cairo Alexandria Tanta Active 7/14 (50%) 0/6 (0%) 0/3
(0%) Placebo 0/15 (0%) 0/6 (0%) 0/3 (0%) P = 0.0453,
Cochran-Mantel-Haenszel test
A summary of response rates for the active treatment group by
disease-related complications and study center is presented in
Table 15.
TABLE-US-00015 TABLE 15 Response Rates for the Active Treatment
Group by Complicating Disease-Related Factors and Study Center
Study Center Complicating factors Cairo Alexandria Tanta High viral
load 0/1 0/2 -- Cirrhosis 0/2 -- 0/1 Bridging fibrosis 3/4 -- 0/1
Uncontrolled diabetes mellitus 0/1 0/2 -- Hepatitis B virus
co-infection -- 0/1 -- High viral load and cirrhosis -- 0/1 -- High
viral load + bridging fibrosis 0/1 -- -- High viral load, diabetes,
bridging -- -- 0/1 fibrosis Patients without complicating factors
4/5 -- -- Totals 7/14 0/6 0/3
There were no significant protocol deviations that would warrant an
efficacy subset analysis. An analysis of the subset of patients
with low viral loads and no cirrhosis, uncontrolled diabetes or
hepatitis B virus co-infection is presented in Table 16.
TABLE-US-00016 TABLE 16 Virologic Responses by Treatment Group,
Subset of Patients with Low Viral Loads and No Cirrhosis,
Uncontrolled Diabetes or Hepatitis B Co-infection Active Placebo P*
Responders/Total (%) 7/10 (70%) 0/15 (0%) 0.0002 *two-sided
Fisher's exact test
The Alinia.RTM. tablets administered 500 mg twice daily with food
for 24 weeks produced virologic responses (undetectable serum HCV
RNA) in 7 of 23 patients (30.4%) compared to zero of 25 patients
(0%) from the placebo group (P=0.0039).
The virologic responses occurred between 4 and 20 weeks of
treatment (3 at week 4, 3 at week 8, 1 at week 20) and were
maintained through the end of treatment with no virological
breakthroughs.
Virologic response was sustained in 5 of 23 patients in the
Alinia.RTM. treatment group at least 24 weeks after the end of
treatment (P=0.0219). Each of the two patients that relapsed
following the end of treatment visit had advanced liver disease
(bridging fibrosis). One dropped out of the study after 8 weeks of
treatment, and the other reported sporadic noncompliance with
taking the study medication.
Low viral load was the most significant independent predictor of
virologic response (P=0.0086). None of the patients with cirrhosis,
uncontrolled diabetes mellitus or hepatitis B virus co-infection
responded to treatment.
When patients with high viral loads, cirrhosis, uncontrolled
diabetes or hepatitis B co-infection were excluded from the
efficacy analysis, virologic response rates were 7/10 (70%) for the
active treatment group and 0/15 for the placebo group (P=0.0002).
Two of the three Alinia.RTM.-treated failures included in this
analysis had advanced liver disease with bridging fibrosis.
These results indicate that 24 weeks of Alinia.RTM. monotherapy is
effective in achieving a sustained virologic response in patients
with chronic hepatitis C genotype 4 when the patients have low
viral loads and no other complicating factors such as cirrhosis,
uncontrolled diabetes or hepatitis B co-infection.
Safety measures were examined in patients receiving Alinia.RTM.
compared to patients receiving placebo tablets. The extent of
exposure is summarized in Table 17. Three patients (2 randomized to
the Alinia.RTM. treatment group, 1 randomized to the placebo group)
dropped out of the study before returning for any follow-up visits.
These patients did not report taking any medication or experiencing
any adverse events, and they were excluded from the safety
analyses.
TABLE-US-00017 TABLE 17 Extent of Exposure Treatment/Exposure No.
of Patients Alinia 500 mg twice daily .times. 24 weeks 22 Alinia
500 mg twice daily .times. 12 weeks 1 Placebo twice daily .times.
24 weeks 24
Sixteen patients (11 from Alinia.RTM. group, 5 from placebo group)
reported a total of 33 adverse events. There were two serious
adverse events. One patient in the placebo group experienced severe
hematemesis and a patient in the Alinia.RTM. treatment group
experienced moderate melena. Both events required hospitalization
but resolved without discontinuing treatment. The remaining adverse
events were mild to moderate and transient in nature, none
requiring modification or discontinuation of treatment. Adverse
events are displayed by body system, standard term, severity and
causality in Table 18 for the active treatment group and in Table
19 for the placebo treatment group. The proportions of patients
reporting each adverse event were compared by treatment group.
There were no significant differences in the frequency or nature of
adverse events reported by the two treatment groups.
TABLE-US-00018 TABLE 18 Adverse Events: Patients Exposed to Alinia
.RTM. (N = 23) Patients Severity and Relationship to Use of the
Drug.sup.2 Adverse event Reporting AEs Mild Moderate Severe
(Affected system).sup.1 Number % N U P PR N U P PR N U P PR
Jaundice (DIG) 2 8.7 -- 2 -- -- -- -- -- -- -- -- -- -- Anorexia
(DIG) 1 4.3 -- -- 1 -- -- -- -- -- -- -- -- -- Constipation (DIG) 1
4.3 -- 1 -- -- -- -- -- -- -- -- -- -- Diarrhea (DIG) 1 4.3 -- 1 --
-- -- -- -- -- -- -- -- -- Flatulence (DIG) 1 4.3 -- 1 -- -- -- --
-- -- -- -- -- -- GI Disorder (DIG) 1 4.3 -- -- -- -- 1 -- -- -- --
-- -- -- Melena (DIG) 1 4.3 -- -- -- -- -- 1 -- -- -- -- -- --
Nausia (DIG) 1 4.3 -- 1 -- -- -- -- -- -- -- -- -- -- Asthenia
(BODY) 4 17.4 -- 4 -- -- -- -- -- -- -- -- -- -- Pain Abdo (BODY) 1
4.3 -- 1 -- -- -- -- -- -- -- -- -- -- Dysuria (UG) 2 4.3 -- 1 --
-- -- 1 -- -- -- -- -- -- Epistaxis (RES) 1 4.3 1 -- -- -- -- -- --
-- -- -- -- -- Palpitation (CV) 1 4.3 -- 1 -- -- -- -- -- -- -- --
-- -- Myalgia (MS) 1 4.3 -- -- -- -- -- 1 -- -- -- -- -- --
Somnolence (NER) 1 4.3 -- 1 -- -- -- -- -- -- -- -- -- -- Skin
Discolor (SKIN) 1 4.3 -- 1 -- -- -- -- -- -- -- -- -- -- .sup.1DIG
= Digestive; BODY = Body as a whole or Nonspecific system; UG =
Urogenital; RES = respiratory; CV = Cardiovascular; MS =
Musculoskeletal; NER = Nervous; SKIN = Skin. .sup.2Relationship to
use of the drug: N = not related, U = unlikely related, P =
possibly related, PR = probably related
TABLE-US-00019 TABLE 19 Adverse Events: Patients Exposed to Placebo
(N = 24) Patients Severity and Relationship to Use of the
Drug.sup.2 Adverse event Reporting AEs Mild Moderate Severe
(Affected system).sup.1 Number % N U P PR N U P PR N U P PR
Jaundice (DIG) 1 4.2 -- 1 -- -- -- -- -- -- -- -- -- -- Hematemesis
(DIG) 1 4.2 -- -- -- -- -- -- -- -- 1 -- -- -- Vomit (DIG) 1 4.2 --
1 -- -- -- -- -- -- -- -- -- -- Asthenia (BODY) 2 8.3 -- 2 -- -- --
-- -- -- -- -- -- -- Pain Abdo (BODY) 2 8.3 -- 2 -- -- -- -- -- --
-- -- -- -- Headache (BODY) 1 4.2 -- 1 -- -- -- -- -- -- -- -- --
-- Fever (BODY) 1 4.2 -- -- -- -- -- 1 -- -- -- -- -- -- Urine
Abnorm (UG) 1 4.2 -- 1 -- -- -- -- -- -- -- -- -- -- Hemoptysis
(RES) 1 4.2 -- 1 -- -- -- -- -- -- -- -- -- -- Diabetes Mell (MAN)
1 4.2 -- 1 -- -- -- -- -- -- -- -- -- -- .sup.1DIG = Digestive;
BODY = Body as a whole or Nonspecific system; UG = Urogenital; RES
= respiratory; MAN = Metabolic and Nutritional. .sup.2Relationship
to use of the drug: N = not related, U = unlikely related, P =
possibly related, PR = probably related
Changes in laboratory safety parameters over time were analyzed by
treatment group using repeated measures analysis of variance for
continuous data and Fisher's Exact tests for categorical data. No
significant changes in laboratory safety parameters were
observed.
No safety concerns were identified during the course of this study.
The Alinia.RTM. tablets administered 500 mg twice daily with food
in patients with chronic hepatitis C were safe and well tolerated.
Adverse events reported for patients treated with Alinia.RTM.
tablets were similar to those reported by patients treated with
placebo.
In this study, Alinia.RTM. tablets administered 500 mg twice daily
with food for 24 weeks produced virologic responses (undetectable
serum HCV RNA) in 7 of 23 patients (30.4%) compared to zero of 25
patients (0%) from the placebo group (P=0.0039). The virologic
responses occurred between 4 and 20 weeks of treatment (3 at week
4, 3 at week 8, 1 at week 20) and were maintained through the end
of treatment with no virological breakthroughs. Virologic response
was sustained in 5 patients at least 24 weeks after the end of
treatment.
Low viral load was the most significant independent predictor of
virologic response (P=0.0086). None of the patients with cirrhosis,
uncontrolled diabetes mellitus or hepatitis B virus co-infection
responded to treatment.
When patients with high viral loads, cirrhosis, uncontrolled
diabetes or hepatitis B co-infection were excluded from the
efficacy analysis, virologic response rates were 7/10 (70%) for the
active treatment group and 0/15 for the placebo group (P=0.0002).
Two of the three Alinia.RTM.-treated failures included in this
analysis had advanced liver disease with bridging fibrosis.
These results indicate that 24 weeks of Alinia monotherapy is
effective in achieving a sustained virologic response in patients
with chronic hepatitis C genotype 4 when the patients have low
viral loads and no other complicating factors such as cirrhosis,
uncontrolled diabetes or hepatitis B co-infection.
No safety concerns were identified during the course of the study.
Adverse events reported for patients in the Alinia.RTM. treatment
group were similar to those reported for the placebo group. There
were no significant changes in clinical laboratory values over the
24-week course of treatment for the Alinia.RTM. treatment group
compared to the placebo group.
Example 6
Treatment of Viral Hepatitis with Alinia and Pegylated Interferon
Alpha-2B
Thirty-six (36) patients were enrolled in a clinical study to
evaluate the effectiveness and safety of combination therapy with
Alinia.RTM. plus pegylated interferon alpha-2b (PegIFN .alpha.-2b)
compared to a placebo plus PegIFN .alpha.-2b in treating chronic
hepatitis C. The patients were recruited as follows: Upon
completing the 24-week treatment phase of study RM01-3027 (see
Example 4), a randomized double-blind placebo-controlled study of
Alinia.RTM., eighteen (18) non-responders were offered the
opportunity to participate in this clinical trial. Two patients
declined enrollment due to the advanced stage of their disease and
unwillingness to be treated with pegylated interferon. Sixteen (16)
patients were enrolled in the study. These patients continued their
blinded oral study medication along with 12 weekly injections of
PegIFN .alpha.-2b. Twenty (20) treatment-naive patients were
recruited for the study to initiate blinded study medication plus
PegIFN .alpha.-2b at the same time (first PegIFN injection and
first dose of oral blinded medication on the same day). See FIG. 6
for a Patient Disposition Flowchart. One patient was enrolled with
HCV genotype 2 (randomized to the pre-treated active group). One
patient dropped out of the study immediately after receiving his
first dose of PegIFN and did not return for any post-treatment
follow-up. One patient did not return for follow-up after week 8.
Each of the remaining 34 patients completed the study. An
intent-to-treat population (all patients randomized) was used for
the primary efficacy analysis with drop-outs being treated as
failures. Demographic data and disease-related characteristics are
summarized by treatment group in Table 20.
TABLE-US-00020 TABLE 20 Demographic and Disease-Related
Characteristics Pre-Treated Not Pre-Treated Active Placebo Active
Placebo P.sup.1 Race: Caucasian 8 8 10 10 1.0 Gender: Male/Female
8/0 7/1 8/2 10/0 .20 Age (years): Mean .+-. SD 45.1 .+-. 5.5 41.3
.+-. 10.1 46.0 .+-. 9.1 39.1 .+-. 8.9 .28 Median (Range) 46.5
(38-52) 42.5 (27-55) 48 (26-56) 40 (21-49) Weight (kgs): Mean .+-.
SD 77.8 .+-. 6.6 84.0 .+-. 13.0 77.1 .+-. 11.8 77.7 .+-. 9.7 .51
Median (Range) 79.5 (68-86) 86 (67-105) 79.5 (56-100) 75 (64-94)
Body Mass Index Mean .+-. SD 26.0 .+-. 2.3 28.3 .+-. 4.5 26.1 .+-.
3.4 26.8 .+-. 3.8 .54 Median (Range) 26.3 (21-29) 29.0 (22-36) 27.0
(20-31) 25.7 (21-36) Viral load (log.sub.10 IU/mL).sup.2 Mean .+-.
SD 5.5 .+-. 0.6 5.6 .+-. 0.5 5.9 .+-. 0.5 5.6 .+-. 0 .4 .34 Median
(Range) 5.6 (4.3-6.1) 5.6 (4.9-6.5) 5.9 (4.9-6.6) 5.7 (4.5-6.1)
Viral load .gtoreq.800,000 3 (38%) 2 (25%) 4 (40%) 1 (10%) .39
IU/mL Elevated ALT 7 (88%) 7 (88%) 9 (90%) 8 (80%) .95 Advanced
liver disease Cirrhosis 1 (13%) -- -- 1 (10%) .34 Bridging fibrosis
2 (25%) 1 (13%) -- -- Diabetes mellitus 3 (38%) 1 (13%) 1 (10%) 1
(10%) .42 .sup.1Chi-square test used for comparing proportions,
analysis of variance for means. .sup.2For pre-treated patients,
viral loads are presented as determined before the pre-treatment
period.
Each of the weekly peginterferon injections were administered by
the physicians. At each study visit, patients were questioned
regarding compliance with administration of the oral study
medication (Alinia or placebo). With the exception of one patient
who dropped out of the study during the first week and another
patient who did not return for evaluation at week 12 and was
treated as a nonresponder, each of the patients reported that they
had been compliant with taking the medication. None of the patients
returned unused medication.
Virologic responses are summarized by treatment group in Table 21.
The response rate for the pre-treated active group (5/8, 63%) was
higher than that of the pre-treated placebo group (P=0.15734),
non-pretreated active group (P=0.08824), the non-pretreated placebo
group (P=0.31859), the two placebo groups combined (P=0.16888) and
the three other groups combined (P=0.09102).
TABLE-US-00021 TABLE 21 Virologic Responses by Treatment Group
Pre-treated Not Pre-treated Active Placebo Active Placebo
Responders/ 5/8 (63%) 2/8 (25%) 2/10 (20%) 4/10 (40%) Total (%) P =
0.26, chi-square test
Logistic regression analyses identified lower fasted blood glucose
as a significant independent predictor of virologic response
(P=0.0101) for the entire population of patients studied (n=36).
The relationship between fasted blood glucose and virologic
response was most significant (P=0.0011) in the pre-treated active
group where there were three patients with uncontrolled diabetes
mellitus.
Given the relationships observed between virologic response and
fasted blood glucose, the efficacy analysis was repeated for a
subset of patients which excluded patients with uncontrolled
diabetes mellitus. The results of this analysis are presented in
Table 5. In this subset of non-diabetic patients, the response rate
for the pre-treated active group (5/5, 100%) was higher than that
of the pre-treated placebo group (P=0.02652), non-pretreated active
group (P=0.01049), the non-pretreated placebo group (P=0.06294),
the two placebo groups combined (P=0.02270) and the three other
groups combined (P=0.00903). Demographic and disease-related
characteristics of the subset of non-diabetic patients analyzed in
Table 22 were compared by treatment group, and there were no
significant differences between groups.
TABLE-US-00022 TABLE 22 Virologic Responses by Treatment Group,
Excluding Patients with Uncontrolled Diabetes Mellitus Pre-Treated
Not Pre-Treated Active Placebo Active Placebo Responders/ 5/5
(100%) 2/7 (29%) 2/9 (22%) 4/9 (44%) Total (%) P = 0.01, chi-square
test
Each of the virologic responders in the pre-treated
Alinia.RTM.+pegIFN group had complicating disease-related factors
that might ordinarily reduce the probability of treatment success
with pegIFN-ribavirin. Response rates for subsets of patients with
high viral loads, advanced liver disease, and uncontrolled diabetes
are presented by treatment group in Table 23.
TABLE-US-00023 TABLE 23 Response Rates in Patients with
Complicating Disease-Related Factors No. Responders/Total
Pre-Treated Not Pre-Treated Active Placebo Active Placebo Viral
load >800,000 IU/mL 2/2 0/1 1/3 0/1 Advanced liver disease:
Cirrhosis 1/1 -- -- -- Bridging fibrosis 1/1 1/1 -- -- HBV
co-infection 1/1 -- -- -- Uncontrolled diabetes 0/2 -- -- -- with
high viral load (HVL) -- 0/1 0/1 -- with HVL and bridging fibrosis
0/1 -- -- -- with cirrhosis -- -- -- 0/1 None of the above -- 1/5
1/6 4/8
Two-log drop in serum HCV RNA. All patients with a 2-log drop in
serum HCV RNA at the end of treatment also had undetectable serum
HCV RNA. The results are, therefore, the same as presented in
Tables 21, 22, and 23.
Changes in ALT from baseline to week 12 are summarized by treatment
group in Table 24.
TABLE-US-00024 TABLE 24 Changes in ALT by Treatment Group
Pre-Treated Not Pre-Treated Active Placebo Active Placebo
Normalized 3 1 2 2 Remained Elevated 4 6 6 4 Remained Normal 1 1 1
1 Normal to Elevated -- -- -- 1 Note: 3 patients not evaluable due
to missing ALT data at either baseline or end of treatment.
Virologic responses by treatment group are presented for each of
two study centers in Table 25. The same data is presented for the
subset of patients without uncontrolled diabetes in Table 26. In
the overall analysis, there was no significant difference between
the response rates observed for the two study centers. In the
subset analysis, the response rates were significantly different
because the second study center had two patients that responded on
placebo+pegIFN. These two patients were 27 and 30 year-old males
with low viral loads and no complicating disease-related
conditions. The patient enrolled in the non-pretreated active group
with genotype 2 was a nonresponder. There were no other significant
protocol deviations.
TABLE-US-00025 TABLE 25 Virologic Responses by Study Site and
Treatment Group No. Responders/Total Pre-treated Not Pre-treated
Active Placebo Active Placebo First study center 3/5 0/5 2/10 4/10
Second study center 2/3 2/3 -- -- P = 0.35, Cochran-Mantel-Haenszel
test
TABLE-US-00026 TABLE 26 Virologic Responses by Study Site and
Treatment Group, Patients without Uncontrolled Diabetes Mellitus
No. Responders/Total Pre-treated Not Pre-treated Active Placebo
Active Placebo First study center 3/3 0/4 2/9 4/9 Second study
center 2/2 2/3 -- -- P = 0.0465, Cochran-Mantel-Haenszel test
Administration of 24 weeks of Alinia.RTM. followed by 12 weeks of
Alinia plus pegIFN alfa-2b produced higher virologic response rates
(5/8, 63%) than either pegIFN alfa-2b plus placebo for 12 weeks
(6/18, 33%) or Alinia.RTM. plus pegIFN alfa-2b for 12 weeks without
pre-treatment (2/10, 20%).
When patients with uncontrolled diabetes mellitus were excluded,
the response rate for the pre-treated active group (5/5, 100%) was
higher than that of the pre-treated placebo group (2/7, 29%,
P=0.02652), non-pretreated active group (2/9, 22%, P=0.01049), the
non-pretreated placebo group (4/9, 44%, P=0.06294), the two placebo
groups combined (6/16, 38%, P=0.02270) and the three other groups
combined (8/25, 32%, P=0.00903).
Each of the 5 virologic responders in the pre-treated active
treatment group had disease-related complications that might
typically reduce the probability of success with pegIFN-ribavirin
therapy: 2 with viral load>800,000 IU/mL, 2 with advanced liver
disease (1 cirrhosis, 1 bridging fibrosis) and 1 with hepatitis B
virus co-infection.
These results indicate that pre-treatment of patients with
Alinia.RTM. before adding pegIFN potentiates the effect of pegIFN,
producing response rates that are significantly higher than those
for pegIFN alone or Alinia plus pegIFN without a pre-treatment
period.
Drug safety measures were examined for patients treated with
Alinia.RTM. plus pegIFN and for those receiving placebo plus
pegIFN. The extent of exposure is summarized in Table 27.
TABLE-US-00027 TABLE 27 Extent of Exposure No. of
Treatment/Exposure Patients Alinia 500 mg twice daily .times. 12
weeks + 18 weekly pegIFN injections Placebo twice daily .times. 24
weeks + 17 weekly pegIFN injections One peginterferon injection
(dropped out) 1
Four mild adverse events (AEs) were reported, three for patients in
the placebo treatment group and one for a patient in the active
treatment group. There were no serious adverse events. None of the
adverse events required modification or discontinuation of
treatment. Adverse events are displayed by body system, standard
term, severity and causality in Table 28 for the active treatment
group and in Table 29 for the placebo treatment group. The
proportions of patients reporting each adverse event were compared
by treatment group. There were no significant differences in the
frequency or nature of adverse events reported by the two treatment
groups. No deaths, serious AEs, or other significant AEs were
reported. No laboratory adverse events were reported during the
study.
TABLE-US-00028 TABLE 28 Adverse Events: Patients Exposed to Alinia
(N = 18) Patients Severity and Relationship to Use of the
Drug.sup.2 Adverse event Reporting AEs Mild Moderate Severe
(Affected system).sup.1 Number % N U P PR N U P PR N U P PR
Depression (NER) 1 5.6 -- 1 -- -- -- -- -- -- -- -- -- -- .sup.1NER
= Nervous system .sup.2Relationship to use of the drug: N = not
related, U = unlikely related, P = possibly related, PR = probably
related
TABLE-US-00029 TABLE 29 Adverse Events: Patients Exposed to Placebo
(N = 17) Patients Severity and Relationship to Use of the
Drug.sup.2 Adverse event Reporting AEs Mild Moderate Severe
(Affected system).sup.1 Number % N U P PR N U P PR N U P PR
Petechia (HAL) 1 5.8 -- 1 -- -- -- -- -- -- -- -- -- -- Depression
(NER) 1 5.8 -- 1 -- -- -- -- -- -- -- -- -- -- Photosensitivity 1
5.8 -- 1 -- -- -- -- -- -- -- -- -- -- (BODY) .sup.1HAL = Heme and
Lymphatic System. NER = Nervous system. BODY = Body as a whole or
Nonspecific system .sup.2Relationship to use of the drug: N = not
related, U = unlikely related, P = possibly related, PR = probably
related
Changes in laboratory safety parameters over time were analyzed by
treatment group using repeated measures analysis of variance for
continuous data and Fisher's Exact tests for categorical data.
Significant differences were observed for two parameters: platelet
counts over time were higher for the patients treated with
Alinia+pegIFN than for patients treated with pegIFN+placebo
(P=0.0138), as shown in FIG. 7; and absolute neutrophil counts over
time were higher for patients treated with Alinia+pegIFN than for
patients treated with pegIFN+placebo (P=0.0205), as shown for FIG.
8.
Values recorded for platelet counts and neutrophil counts increased
from week 8 to week 12. A number of patients had their week 12
serum sample collected 3 to 7 days late (10 to 14 days after the
last injection of pegIFN), and their platelet and neutrophil counts
had begun to recover. To eliminate the effect of data collected
late at week 12, data from baseline to week 8 was analyzed
separately. When the week 12 data point was eliminated the
differences in platelet counts and absolute neutrophil counts over
time remained significant (P=0.0044 for platelets, P=0.0101 for
neutrophils).
Analyses were conducted to evaluate the effect of virologic
response or pre-treatment with Alinia on the change in platelet
counts or neutrophil counts over time. The differences were not
related to virologic response or pre-treatment with Alinia.
Vital signs, physical findings, and other observations related to
safety provided no significant findings.
The administration of Alinia tablets administered 500 mg twice
daily with food along with weekly injections of pegylated
interferon alfa-2b for 12 weeks in patients with chronic hepatitis
C was safe and well tolerated.
Reductions of platelet counts and neutrophil counts typically
associated with administration of pegIFN were significantly smaller
in patients treated with Alinia (P=0.0044 and 0.0101,
respectively).
Administration of 24 weeks of Alinia followed by 12 weeks of Alinia
plus pegIFN alfa-2b produced higher virologic response rates (5/8,
63%) than either pegIFN alfa-2b plus placebo for 12 weeks (6/18,
33%) or Alinia plus pegIFN alfa-2b for 12 weeks without
pre-treatment (2/10, 20%).
When patients with uncontrolled diabetes mellitus were excluded,
the response rate for the pre-treated active group (5/5, 100%) was
higher than that of the pre-treated placebo group (2/7, 29%,
P=0.02652), non-pretreated active group (2/9, 22%, P=0.01049), the
non-pretreated placebo group (4/9, 44%, P=0.06294), the two placebo
groups combined (6/16, 38%, P=0.02270) and the three other groups
combined (8/25, 32%, P=0.00903).
Each of the 5 virologic responders in the pre-treated active
treatment group had disease-related complications that might
typically reduce the probability of success with pegIFN-ribavirin
therapy: 2 with viral load>800,000 IU/mL, 2 with advanced liver
disease (1 cirrhosis, 1 bridging fibrosis) and 1 with hepatitis B
virus co-infection.
The administration of Alinia along with pegIFN alfa-2b in patients
with chronic hepatitis C was safe and well tolerated. No safety
concerns were identified.
Reductions of platelet counts and neutrophil counts typically
associated with administration of pegIFN were significantly smaller
in patients treated with Alinia (P=0.0044 and 0.0101,
respectively).
These results indicate that pre-treatment of patients with Alinia
before adding pegIFN potentiates the effect of pegIFN, producing
response rates significantly higher than those for pegIFN alone or
Alinia plus pegIFN without a pre-treatment period. Concomitant
administration of Alinia may furthermore reduce the hematologic
toxicity of pegIFN.
* * * * *
References