U.S. patent application number 15/454844 was filed with the patent office on 2017-08-24 for deuterated derivatives of ruxolitinib.
The applicant listed for this patent is Concert Pharmaceuticals, Inc.. Invention is credited to Scott L. Harbeson, Julie F. Liu, Adam J. Morgan, Bhaumik Pandya, I. Robert Silverman.
Application Number | 20170239254 15/454844 |
Document ID | / |
Family ID | 53520766 |
Filed Date | 2017-08-24 |
United States Patent
Application |
20170239254 |
Kind Code |
A1 |
Silverman; I. Robert ; et
al. |
August 24, 2017 |
DEUTERATED DERIVATIVES OF RUXOLITINIB
Abstract
The present invention in one embodiment provides a compound of
Formula A: ##STR00001## or a pharmaceutically acceptable salt
thereof; pharmaceutical compositions comprising the compound; and
methods of treating the indications disclosed herein.
Inventors: |
Silverman; I. Robert;
(Arlington, MA) ; Liu; Julie F.; (Lexington,
MA) ; Morgan; Adam J.; (Ashland, MA) ; Pandya;
Bhaumik; (Arlington, MA) ; Harbeson; Scott L.;
(Cambridge, MA) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
Concert Pharmaceuticals, Inc. |
Lexington |
MA |
US |
|
|
Family ID: |
53520766 |
Appl. No.: |
15/454844 |
Filed: |
March 9, 2017 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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14570954 |
Dec 15, 2014 |
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15454844 |
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PCT/US2013/045919 |
Jun 14, 2013 |
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14570954 |
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61660428 |
Jun 15, 2012 |
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61678795 |
Aug 2, 2012 |
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61917589 |
Dec 18, 2013 |
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Current U.S.
Class: |
1/1 |
Current CPC
Class: |
C07B 2200/05 20130101;
A61K 31/454 20130101; A61K 31/4045 20130101; A61K 31/7068 20130101;
A61K 45/06 20130101; A61K 31/5685 20130101; A61K 31/7068 20130101;
A61K 31/4045 20130101; C07D 487/04 20130101; C07D 487/04 20130101;
A61P 35/00 20180101; A61K 31/454 20130101; A61K 31/5685 20130101;
A61K 31/519 20130101; A61K 31/519 20130101; A61K 2300/00 20130101;
A61K 2300/00 20130101; A61K 2300/00 20130101; A61K 2300/00
20130101; A61K 2300/00 20130101; C07B 2200/05 20130101 |
International
Class: |
A61K 31/519 20060101
A61K031/519 |
Claims
1-11. (canceled)
12. A method of inhibiting one or more of JAK1 and JAK2 in a cell,
comprising contacting the cell with a compound represented by the
following structural formula: ##STR00019## or a pharmaceutically
acceptable salt thereof.
13. The method of claim 12, wherein each position in the compound
designated specifically as deuterium has at least 90% incorporation
of deuterium, and wherein any atom not designated as deuterium is
present at its natural isotopic abundance.
14. The method of claim 12, wherein each position in the compound
designated specifically as deuterium has at least 95% incorporation
of deuterium, and wherein any atom not designated as deuterium is
present at its natural isotopic abundance.
15. The method of claim 12, wherein, the pharmaceutically
acceptable salt of the compound is the phosphate salt.
Description
RELATED APPLICATIONS
[0001] This application is a Continuation of U.S. application Ser.
No. 14/570,954, filed Dec. 15, 2014, which is a
continuation-in-part of International Application No.
PCT/US2013/045919, which designated the United States and was filed
on Jun. 14, 2013, published in English, which claims the benefit of
U.S. Provisional Application Ser. No. 61/660,428, filed Jun. 15,
2012, and U.S. Provisional Application Ser. No. 61/678,795, filed
Aug. 2, 2012. This application also claims the benefit of U.S.
Provisional Application Ser. No. 61/917,589, filed Dec. 18, 2013.
The entire teachings of the above applications are incorporated
herein by reference.
BACKGROUND OF THE INVENTION
[0002] Many current medicines suffer from poor absorption,
distribution, metabolism and/or excretion (ADME) properties that
prevent their wider use or limit their use in certain indications.
Poor ADME properties are also a major reason for the failure of
drug candidates in clinical trials. While formulation technologies
and prodrug strategies can be employed in some cases to improve
certain ADME properties, these approaches often fail to address the
underlying ADME problems that exist for many drugs and drug
candidates. One such problem is rapid metabolism that causes a
number of drugs, which otherwise would be highly effective in
treating a disease, to be cleared too rapidly from the body. A
possible solution to rapid drug clearance is frequent or high
dosing to attain a sufficiently high plasma level of drug. This,
however, introduces a number of potential treatment problems such
as poor patient compliance with the dosing regimen, side effects
that become more acute with higher doses, and increased cost of
treatment. A rapidly metabolized drug may also expose patients to
undesirable toxic or reactive metabolites.
[0003] Another ADME limitation that affects many medicines is the
formation of toxic or biologically reactive metabolites. As a
result, some patients receiving the drug may experience toxicities,
or the safe dosing of such drugs may be limited such that patients
receive a suboptimal amount of the active agent. In certain cases,
modifying dosing intervals or formulation approaches can help to
reduce clinical adverse effects, but often the formation of such
undesirable metabolites is intrinsic to the metabolism of the
compound.
[0004] In some select cases, a metabolic inhibitor will be
co-administered with a drug that is cleared too rapidly. Such is
the case with the protease inhibitor class of drugs that are used
to treat HIV infection. The FDA recommends that these drugs be
co-dosed with ritonavir, an inhibitor of cytochrome P450 enzyme 3A4
(CYP3A4), the enzyme typically responsible for their metabolism
(see Kempf, D. J. et al., Antimicrobial agents and chemotherapy,
1997, 41(3): 654-60). Ritonavir, however, causes adverse effects
and adds to the pill burden for HIV patients who must already take
a combination of different drugs. Similarly, the CYP2D6 inhibitor
quinidine has been added to dextromethorphan for the purpose of
reducing rapid CYP2D6 metabolism of dextromethorphan in a treatment
of pseudobulbar affect. Quinidine, however, has unwanted side
effects that greatly limit its use in potential combination therapy
(see Wang, L et al., Clinical Pharmacology and Therapeutics, 1994,
56(6 Pt 1): 659-67; and FDA label for quinidine at
www.accessdata.fda.gov).
[0005] In general, combining drugs with cytochrome P450 inhibitors
is not a satisfactory strategy for decreasing drug clearance. The
inhibition of a CYP enzyme's activity can affect the metabolism and
clearance of other drugs metabolized by that same enzyme. CYP
inhibition can cause other drugs to accumulate in the body to toxic
levels.
[0006] A potentially attractive strategy for improving a drug's
metabolic properties is deuterium modification. In this approach,
one attempts to slow the CYP-mediated metabolism of a drug or to
reduce the formation of undesirable metabolites by replacing one or
more hydrogen atoms with deuterium atoms. Deuterium is a safe,
stable, non-radioactive isotope of hydrogen. Compared to hydrogen,
deuterium forms stronger bonds with carbon. In select cases, the
increased bond strength imparted by deuterium can positively impact
the ADME properties of a drug, creating the potential for improved
drug efficacy, safety, and/or tolerability. At the same time,
because the size and shape of deuterium are essentially identical
to those of hydrogen, replacement of hydrogen by deuterium would
not be expected to affect the biochemical potency and selectivity
of the drug as compared to the original chemical entity that
contains only hydrogen.
[0007] Over the past 35 years, the effects of deuterium
substitution on the rate of metabolism have been reported for a
very small percentage of approved drugs (see, e.g., Blake, M I et
al, J Pharm Sci, 1975, 64:367-91; Foster, A B, Adv Drug Res 1985,
14:1-40 ("Foster"); Kushner, D J et al, Can J Physiol Pharmacol
1999, 79-88; Fisher, M B et al, Curr Opin Drug Discov Devel, 2006,
9:101-09 ("Fisher")). Many of the examples in these references
report a local deuterium isotope effect (an effect on the rate of
metabolism at a specific site of deuteration in the substrate)
rather than the effect of deuteration on the overall metabolic
stability of the drug, i.e., the overall substrate consumption via
metabolism. The reported results of those studies measuring
deuterium substitution's effect on overall metabolic stability are
variable and unpredictable. For some compounds deuteration caused
decreased metabolic clearance in vivo. For others, there was no
change in metabolism. Still others demonstrated increased metabolic
clearance. The variability in deuterium effects has also led
experts to question or dismiss deuterium modification as a viable
drug design strategy for inhibiting adverse metabolism (see Foster
at p. 35 and Fisher at p. 101).
[0008] The effects of deuterium modification on a drug's metabolic
properties are not predictable even when deuterium atoms are
incorporated at known sites of metabolism. Only by actually
preparing and testing a deuterated drug can one determine if and
how the rate of metabolism will differ from that of its
non-deuterated counterpart. See, for example, Fukuto et al. (J.
Med. Chem. 1991, 34, 2871-76). Many drugs have multiple sites where
metabolism is possible. The site(s) where deuterium substitution is
required and the extent of deuteration necessary to see an effect
on metabolism, if any, will be different for each drug.
[0009] Ruxolitinib phosphate, is a heteroaryl-substituted
pyrrolo[2,3-d]pyrimidines also known as
3(R)-cyclopentyl-3-[4-(7H-pyrrolo[2,3-d]pyrimidin-4-yl)-1H-pyrazol-1-yl]p-
ropanenitrile phosphate and as
(R)-3-(4-(7H-pyrrolo[2,3-d]pyrimidin-4-yl)-1H-pyrazol-1-yl)-3-cyclopentyl-
propanenitrile phosphate, inhibits Janus Associated Kinases (JAKs)
JAK1 and JAK2. These kinases mediate the signaling of a number of
cytokines and growth factors important for hematopoiesis and immune
function. JAK signaling involves recruitment of STATs (signal
transducers and activators of transcription) to cytokine receptors,
activation and subsequent localization of STATs to the nucleus
leading to modulation of gene expression.
[0010] Ruxolitinib phosphate is currently approved for the
treatment of patients with intermediate or high-risk myelofibrosis,
including primary myelofibrosis, post-polycythemia vera
myelofibrosis and post-essential thrombocythemia myelofibrosis.
Ruxolitinib phosphate is also currently in clinical trials for the
treatment of essential thrombocythemia, pancreatic cancer, prostate
cancer, breast cancer, leukemia, non-Hodgkin's lymphoma, multiple
myeloma and psoriasis.
[0011] Three metabolites in humans have been identified as active,
that resulting from hydroxylation at the 2-position on the
cyclopentyl moiety, that resulting from hydroxylation at the
3-position on the cyclopentyl moiety and the ketone resulting from
further oxidation at the 3-position on the cyclopentyl moiety. (See
Shilling, A. D. et al., Drug Metabolism and Disposition, 2010,
38(11): 2023-2031; FDA Prescribing Information and
US20080312258).
[0012] The most common hematologic adverse reactions associated
with the dosing of ruxolitinib are thrombocytopenia and anemia. The
most common non-hematologic adverse reactions are bruising,
dizziness and headache.
[0013] Despite the beneficial activities of ruxolitinib, there is a
continuing need for new compounds to treat the aforementioned
diseases and conditions.
SUMMARY OF THE INVENTION
[0014] This invention relates to novel heteroaryl-substituted
pyrrolo[2,3-d]pyrimidines, and pharmaceutically acceptable salts
thereof. This invention also provides compositions comprising a
compound of this invention and the use of such compositions in
methods of treating diseases and conditions that are beneficially
treated by administering an inhibitor of Janus-associated kinase
with selectivity for subtypes 1 and 2 (JAK1/JAK2).
BRIEF DESCRIPTION OF THE DRAWINGS
[0015] The FIGURE shows the results of metabolic stability testing
of the referenced compounds.
DETAILED DESCRIPTION OF THE INVENTION
Definitions
[0016] The term "treat" means decrease, suppress, attenuate,
diminish, arrest, or stabilize the development or progression of a
disease (e.g., a disease or disorder delineated herein), lessen the
severity of the disease or improve the symptoms associated with the
disease.
[0017] "Disease" means any condition or disorder that damages or
interferes with the normal function of a cell, tissue, or
organ.
[0018] It will be recognized that some variation of natural
isotopic abundance occurs in a synthesized compound depending upon
the origin of chemical materials used in the synthesis. Thus, a
preparation of ruxolitinib will inherently contain small amounts of
deuterated isotopologues. The concentration of naturally abundant
stable hydrogen and carbon isotopes, notwithstanding this
variation, is small and immaterial as compared to the degree of
stable isotopic substitution of compounds of this invention. See,
for instance, Wada, E et al., Seikagaku, 1994, 66:15; Gannes, L Z
et al., Comp Biochem Physiol Mol Integr Physiol, 1998, 119:725.
[0019] In the compounds of this invention any atom not specifically
designated as a particular isotope is meant to represent any stable
isotope of that atom. Unless otherwise stated, when a position is
designated specifically as "H" or "hydrogen", the position is
understood to have hydrogen at its natural abundance isotopic
composition. Also unless otherwise stated, when a position is
designated specifically as "D" or "deuterium", the position is
understood to have deuterium at an abundance that is at least 3000
times greater than the natural abundance of deuterium, which is
0.015% (i.e., at least 45% incorporation of deuterium).
[0020] The term "isotopic enrichment factor" as used herein means
the ratio between the isotopic abundance and the natural abundance
of a specified isotope.
[0021] In other embodiments, a compound of this invention has an
isotopic enrichment factor for each designated deuterium atom of at
least 3500 (52.5% deuterium incorporation at each designated
deuterium atom), at least 4000 (60% deuterium incorporation), at
least 4500 (67.5% deuterium incorporation), at least 5000 (75%
deuterium), at least 5500 (82.5% deuterium incorporation), at least
6000 (90% deuterium incorporation), at least 6333.3 (95% deuterium
incorporation), at least 6466.7 (97% deuterium incorporation), at
least 6600 (99% deuterium incorporation), or at least 6633.3 (99.5%
deuterium incorporation).
[0022] The term "isotopologue" refers to a species in which the
chemical structure differs from a specific compound of this
invention only in the isotopic composition thereof.
[0023] The term "compound," when referring to a compound of this
invention, refers to a collection of molecules having an identical
chemical structure, except that there may be isotopic variation
among the constituent atoms of the molecules. Thus, it will be
clear to those of skill in the art that a compound represented by a
particular chemical structure containing indicated deuterium atoms,
will also contain lesser amounts of isotopologues having hydrogen
atoms at one or more of the designated deuterium positions in that
structure. The relative amount of such isotopologues in a compound
of this invention will depend upon a number of factors including
the isotopic purity of deuterated reagents used to make the
compound and the efficiency of incorporation of deuterium in the
various synthesis steps used to prepare the compound. However, as
set forth above the relative amount of such isotopologues in toto
will be less than 49.9% of the compound. In other embodiments, the
relative amount of such isotopologues in toto will be less than
47.5%, less than 40%, less than 32.5%, less than 25%, less than
17.5%, less than 10%, less than 5%, less than 3%, less than 1%, or
less than 0.5% of the compound.
[0024] The invention also provides salts of the compounds of the
invention.
[0025] A salt of a compound of this invention is formed between an
acid and a basic group of the compound, such as an amino functional
group, or a base and an acidic group of the compound, such as a
carboxyl functional group. According to another embodiment, the
compound is a pharmaceutically acceptable acid addition salt.
[0026] The term "pharmaceutically acceptable," as used herein,
refers to a component that is, within the scope of sound medical
judgment, suitable for use in contact with the tissues of humans
and other mammals without undue toxicity, irritation, allergic
response and the like, and are commensurate with a reasonable
benefit/risk ratio. A "pharmaceutically acceptable salt" means any
non-toxic salt that, upon administration to a recipient, is capable
of providing, either directly or indirectly, a compound of this
invention. A "pharmaceutically acceptable counterion" is an ionic
portion of a salt that is not toxic when released from the salt
upon administration to a recipient.
[0027] Acids commonly employed to form pharmaceutically acceptable
salts include inorganic acids such as hydrogen bisulfide,
hydrochloric acid, hydrobromic acid, hydroiodic acid, sulfuric acid
and phosphoric acid, as well as organic acids such as
para-toluenesulfonic acid, salicylic acid, tartaric acid,
bitartaric acid, ascorbic acid, maleic acid, besylic acid, fumaric
acid, gluconic acid, glucuronic acid, formic acid, glutamic acid,
methanesulfonic acid, ethanesulfonic acid, benzenesulfonic acid,
lactic acid, oxalic acid, para-bromophenylsulfonic acid, carbonic
acid, succinic acid, citric acid, benzoic acid and acetic acid, as
well as related inorganic and organic acids. Such pharmaceutically
acceptable salts thus include sulfate, pyrosulfate, bisulfate,
sulfite, bisulfite, phosphate, monohydrogenphosphate,
dihydrogenphosphate, metaphosphate, pyrophosphate, chloride,
bromide, iodide, acetate, propionate, decanoate, caprylate,
acrylate, formate, isobutyrate, caprate, heptanoate, propiolate,
oxalate, malonate, succinate, suberate, sebacate, fumarate,
maleate, butyne-1,4-dioate, hexyne-1,6-dioate, benzoate,
chlorobenzoate, methylbenzoate, dinitrobenzoate, hydroxybenzoate,
methoxybenzoate, phthalate, terephthalate, sulfonate, xylene
sulfonate, phenylacetate, phenylpropionate, phenylbutyrate,
citrate, lactate, .beta.-hydroxybutyrate, glycolate, maleate,
tartrate, methanesulfonate, propanesulfonate,
naphthalene-1-sulfonate, naphthalene-2-sulfonate, mandelate and
other salts. In one embodiment, pharmaceutically acceptable acid
addition salts include those formed with mineral acids such as
hydrochloric acid and hydrobromic acid, and especially those formed
with organic acids such as maleic acid.
[0028] The compounds of the present invention (e.g., compounds of
Formula I or Formula A), may contain an asymmetric carbon atom, for
example, as the result of deuterium substitution or otherwise. As
such, compounds of this invention can exist as either individual
enantiomers, or mixtures of the two enantiomers. Accordingly, a
compound of the present invention may exist as either a racemic
mixture or a scalemic mixture, or as individual respective
stereoisomers that are substantially free from another possible
stereoisomer. The term "substantially free of other stereoisomers"
as used herein means less than 25% of other stereoisomers,
preferably less than 10% of other stereoisomers, more preferably
less than 5% of other stereoisomers and most preferably less than
2% of other stereoisomers are present. Methods of obtaining or
synthesizing an individual enantiomer for a given compound are
known in the art and may be applied as practicable to final
compounds or to starting material or intermediates.
[0029] Unless otherwise indicated, when a disclosed compound is
named or depicted by a structure without specifying the
stereochemistry and has one or more chiral centers, it is
understood to represent all possible stereoisomers of the
compound.
[0030] The term "mammal" as used herein includes a human or a
non-human animal, such as mouse, rat, guinea pig, dog, cat, horse,
cow, pig, monkey, chimpanzee, baboon, or rhesus. In one embodiment,
the mammal is a non-human animal. In another embodiment, the mammal
is a human.
[0031] The term "stable compounds," as used herein, refers to
compounds which possess stability sufficient to allow for their
manufacture and which maintain the integrity of the compound for a
sufficient period of time to be useful for the purposes detailed
herein (e.g., formulation into therapeutic products, intermediates
for use in production of therapeutic compounds, isolatable or
storable intermediate compounds, treating a disease or condition
responsive to therapeutic agents).
[0032] "D" and "d" both refer to deuterium. "Stereoisomer" refers
to both enantiomers and diastereomers. "Tert" and "" each refer to
tertiary. "US" refers to the United States of America.
[0033] "Substituted with deuterium" refers to the replacement of
one or more hydrogen atoms with a corresponding number of deuterium
atoms.
[0034] Throughout this specification, a variable may be referred to
generally (e.g.,"each R") or may be referred to specifically (e.g.,
R.sup.1, R.sup.2, R.sup.3, etc.). Unless otherwise indicated, when
a variable is referred to generally, it is meant to include all
specific embodiments of that particular variable.
Therapeutic Compounds
[0035] The present invention in one embodiment provides a compound
of Formula A:
##STR00002##
or a pharmaceutically acceptable salt thereof, wherein: [0036]
Y.sup.1 is selected from hydrogen and deuterium; [0037] each
Y.sup.2 is independently selected from hydrogen and deuterium,
provided that each Y.sup.2 attached to a common carbon is the same;
[0038] each Y.sup.3 is independently selected from hydrogen and
deuterium, provided that each Y.sup.3 attached to a common carbon
is the same; [0039] Y.sup.4 is selected from hydrogen and
deuterium; [0040] each Y.sup.5 is the same and is selected from
hydrogen and deuterium; and [0041] Y.sup.6, Y.sup.7, Y.sup.8,
Y.sup.9, and Y.sup.10 are each independently selected from hydrogen
and deuterium; provided that when Y.sup.1 is hydrogen, each Y.sup.2
and each Y.sup.3 are hydrogen, Y.sup.4 is hydrogen, and each of
Y.sup.6, Y.sup.7, Y.sup.8, Y.sup.9, and Y.sup.m is hydrogen, then
each Y.sup.5 is deuterium.
[0042] In one embodiment of Formula A each Y.sup.2 is the same,
each Y.sup.3 is the same and each Y.sup.5 is the same. In one
aspect of this embodiment, each Y.sup.2 is deuterium. In a further
aspect each Y.sup.3 is deuterium. In another further aspect each
Y.sup.3 is hydrogen. In another aspect of this embodiment, each
Y.sup.2 is hydrogen. In a further aspect each Y.sup.3 is deuterium.
In another further aspect each Y.sup.3 is hydrogen. In one example
of any of the foregoing aspects, Y.sup.1 is deuterium. In another
example of any of the foregoing aspects, Y.sup.1 is hydrogen. In a
more particular example of any of the foregoing aspects, Y.sup.1 is
deuterium, Y.sup.4 is deuterium, and each Y.sup.5 is deuterium. In
another more particular example of any of the foregoing aspects,
Y.sup.1 is deuterium, Y.sup.4 is deuterium, and each Y.sup.5 is
hydrogen. In another more particular example of any of the
foregoing aspects, Y.sup.1 is deuterium, Y.sup.4 is hydrogen, and
each Y.sup.5 is hydrogen. In another more particular example of any
of the foregoing aspects, Y.sup.1 is hydrogen, Y.sup.4 is hydrogen,
and each Y.sup.5 is hydrogen. In another more particular example of
any of the foregoing aspects, Y.sup.1 is hydrogen, Y.sup.4 is
hydrogen, and each Y.sup.5 is deuterium. In another more particular
example of any of the foregoing aspects, Y.sup.1 is hydrogen,
Y.sup.4 is deuterium, and each Y.sup.5 is deuterium. In another
more particular example of any of the foregoing aspects, Y.sup.1 is
hydrogen, Y.sup.4 is deuterium, and each Y.sup.5 is hydrogen.
[0043] In one embodiment, Y.sup.6 is deuterium. In one aspect of
this embodiment, each of Y.sup.7 and Y.sup.8 is deuterium. In
another aspect of this embodiment, each of Y.sup.7 and Y.sup.8 is
hydrogen.
[0044] In one embodiment, Y.sup.6 is hydrogen. In one aspect of
this embodiment, each of Y.sup.7 and Y.sup.8 is deuterium. In
another aspect of this embodiment, each of Y.sup.7 and Y.sup.8 is
hydrogen.
[0045] The present invention in one embodiment provides a compound
of Formula I:
##STR00003##
or a pharmaceutically acceptable salt thereof, wherein: [0046]
Y.sup.1 is selected from hydrogen and deuterium; [0047] each
Y.sup.2 is independently selected from hydrogen and deuterium,
provided that each Y.sup.2 attached to a common carbon is the same;
[0048] each Y.sup.3 is independently selected from hydrogen and
deuterium, provided that each Y.sup.3 attached to a common carbon
is the same; [0049] Y.sup.4 is selected from hydrogen and
deuterium; [0050] each Y.sup.5 is the same and is selected from
hydrogen and deuterium; and [0051] Y.sup.6, Y.sup.7, and Y.sup.8
are each independently selected from hydrogen and deuterium;
provided that when Y.sup.1 is hydrogen, each Y.sup.2 and each
Y.sup.3 are hydrogen, Y.sup.4 is hydrogen, and each of Y.sup.6,
Y.sup.7 and Y.sup.8 is hydrogen, then each Y.sup.5 is
deuterium.
[0052] In one embodiment each Y.sup.2 is the same, each Y.sup.3 is
the same and each Y.sup.5 is the same. In one aspect of this
embodiment, each Y.sup.2 is deuterium. In a further aspect each
Y.sup.3 is deuterium. In another further aspect each Y.sup.3 is
hydrogen. In another aspect of this embodiment, each Y.sup.2 is
hydrogen. In a further aspect each Y.sup.3 is deuterium. In another
further aspect each Y.sup.3 is hydrogen. In one example of any of
the foregoing aspects, Y.sup.1 is deuterium. In another example of
any of the foregoing aspects, Y.sup.1 is hydrogen. In a more
particular example of any of the foregoing aspects, Y.sup.1 is
deuterium, Y.sup.4 is deuterium, and each Y.sup.5 is deuterium. In
another more particular example of any of the foregoing aspects,
Y.sup.1 is deuterium, Y.sup.4 is deuterium, and each Y.sup.5 is
hydrogen. In another more particular example of any of the
foregoing aspects, Y.sup.1 is deuterium, Y.sup.4 is hydrogen, and
each Y.sup.5 is hydrogen. In another more particular example of any
of the foregoing aspects, Y.sup.1 is hydrogen, Y.sup.4 is hydrogen,
and each Y.sup.5 is hydrogen. In another more particular example of
any of the foregoing aspects, Y.sup.1 is hydrogen, Y.sup.4 is
hydrogen, and each Y.sup.5 is deuterium. In another more particular
example of any of the foregoing aspects, Y.sup.1 is hydrogen,
Y.sup.4 is deuterium, and each Y.sup.5 is deuterium. In another
more particular example of any of the foregoing aspects, Y.sup.1 is
hydrogen, Y.sup.4 is deuterium, and each Y.sup.5 is hydrogen.
[0053] In one embodiment, Y.sup.6 is deuterium. In one aspect of
this embodiment, each of Y.sup.7 and Y.sup.8 is deuterium. In
another aspect of this embodiment, each of Y.sup.7 and Y.sup.8 is
hydrogen.
[0054] In one embodiment, Y.sup.6 is hydrogen. In one aspect of
this embodiment, each of Y.sup.7 and Y.sup.8 is deuterium. In
another aspect of this embodiment, each of Y.sup.7 and Y.sup.8 is
hydrogen.
[0055] In one embodiment, the compound is a compound of Formula I
wherein Y.sup.6, Y.sup.7 and Y.sup.8 are each hydrogen and the
compound is selected from any one of the compounds (Cmpd) set forth
in Table 1 (below):
TABLE-US-00001 TABLE 1 Exemplary Embodiments of Formula I Cmpd
Y.sup.1 Each Y.sup.2 Each Y.sup.3 Y.sup.4 each Y.sup.5 100 H H H D
H 101 H H H H D 102 H H H D D 103 H H D H H 104 H H D D H 105 H H D
H D 106 H H D D D 107 H D H H H 108 H D H D H 109 H D H H D 110 H D
H D D 111 H D D H H 112 H D D D H 113 H D D H D 114 H D D D D 115 D
H H H H 116 D H H D H 117 D H H H D 118 D H H D D 119 D H D H H 120
D H D D H 121 D H D H D 122 D H D D D 123 D D H H H 124 D D H D H
125 D D H H D 126 D D H D D 127 D D D H H 128 D D D D H 129 D D D H
D 130 D D D D D
or a pharmaceutically acceptable salt thereof, wherein any atom not
designated as deuterium is present at its natural isotopic
abundance.
[0056] In one embodiment, the compound is a compound of Formula I
wherein Y.sup.6, Y.sup.7 and Y.sup.8 are each D and the compound is
selected from any one of the compounds (Cmpd) set forth in Table 2
(below):
TABLE-US-00002 TABLE 2 Exemplary Embodiments of Formula I Cmpd
Y.sup.1 Each Y.sup.2 Each Y.sup.3 Y.sup.4 each Y.sup.5 200 H H H D
H 201 H H H H D 202 H H H D D 203 H H D H H 204 H H D D H 205 H H D
H D 206 H H D D D 207 H D H H H 208 H D H D H 209 H D H H D 210 H D
H D D 211 H D D H H 212 H D D D H 213 H D D H D 214 H D D D D 215 D
H H H H 216 D H H D H 217 D H H H D 218 D H H D D 219 D H D H H 220
D H D D H 221 D H D H D 222 D H D D D 223 D D H H H 224 D D H D H
225 D D H H D 226 D D H D D 227 D D D H H 228 D D D D H 229 D D D H
D 230 D D D D D 231 H H H H H
or a pharmaceutically acceptable salt thereof, wherein any atom not
designated as deuterium is present at its natural isotopic
abundance.
[0057] In another set of embodiments, any atom not designated as
deuterium in any of the embodiments set forth above is present at
its natural isotopic abundance.
[0058] The following compounds are useful for making various
compounds of this invention:
##STR00004## ##STR00005## ##STR00006## ##STR00007##
or a salt thereof, wherein any atom not designated as deuterium is
present at its natural isotopic abundance.
[0059] The synthesis of compounds of Formula I or Formula A may be
readily achieved by synthetic chemists of ordinary skill by
reference to the Exemplary Synthesis and Examples disclosed herein.
Relevant procedures analogous to those of use for the preparation
of compounds of Formula I or Formula A and intermediates thereof
are disclosed, for instance, in U.S. Pat. No. 7,598,257 and in
Organic Letters, 2009, 11(9): 1999-2009.
[0060] Such methods can be carried out utilizing corresponding
deuterated and optionally, other isotope-containing reagents and/or
intermediates to synthesize the compounds delineated herein, or
invoking standard synthetic protocols known in the art for
introducing isotopic atoms to a chemical structure.
Exemplary Synthesis
[0061] Compounds of Formula I or Formula A may be prepared in a
manner analogous to those syntheses presented in US Patent
7,598,257 and in Organic Letters, 2009, 11(9): 1999-2009 using
appropriately deuterated starting materials.
[0062] Compounds of Formula I or Formula A may also be prepared as
shown in the schemes below.
##STR00008## ##STR00009##
[0063] Scheme 1 discloses an exemplary preparation of the compound
of formula I wherein Y.sup.1, each Y.sup.2 and each Y.sup.3 are
deuterium and Y.sup.4, each Y.sup.5, Y.sup.6, Y.sup.7 and Y.sup.8
are hydrogen. In a manner analogous to that described in WO
2010/083283, commercially available,
4-chloro-7H-pyrrolo[2,3-d]pyrimidine 11 (Aldrich) is treated with
sodium hydride and SEM chloride to afford 12, which is reacted with
commercially available 13 to provide 14. Instead of 11,
4-bromo-7H-pyrrolo[2,3-d]pyrimidine may also be used in the first
step to provide the SEM-protected
4-bromo-7H-pyrrolo[2,3-d]pyrimidine (analogous to 12) which can be
reacted with 13 to provide 14. Reaction of 14 with 15, prepared as
disclosed in Scheme 2a below, is performed in a manner analogous to
that described in Lin, Q. et al. Org. Lett. 2009, 11, 1999, to give
16. The reaction is performed in the presence of chiral ligand 27,
prepared as described in Lin, Q. et al. 16 is converted to 17 by
treatment with NH4OH and I.sub.2. The SEM protecting group of 17 is
then deprotected with LiBF.sub.4 and NH.sub.4OH to give a compound
of Formula I.
##STR00010##
[0064] As shown in Scheme 2a, commercially available 18 is treated
with phosphonium ylide 20 and DCl/D.sub.2O to provide 19, which is
treated with 20 and DiBAl-H to afford 15.
##STR00011##
##STR00012##
[0065] Compounds analogous to 15 may also be prepared. For example,
as shown in Scheme 2b, commercially available 21 may be converted
to 23 in a manner analogous to that disclosed in Scheme 2a. As
another example, as shown in Scheme 2c, commercially available 24
may be converted to 26 in a manner analogous to that disclosed in
Scheme 2a and Scheme 2b. 23 may be converted, in a manner similar
to that disclosed in Scheme 1, to a compound of formula I wherein
Y.sup.1 and each Y.sup.3 are deuterium and Y.sup.4, each Y.sup.2,
each Y.sup.5, Y.sup.6, Y.sup.7 and Y.sup.8 are hydrogen. Likewise,
26 may be converted, in a manner similar to that disclosed in
Scheme 1, to a compound of formula I wherein Y.sup.1 and each
Y.sup.2 are deuterium and Y.sup.4, each Y.sup.3, each Y.sup.5,
Y.sup.6, Y.sup.7 and Y.sup.8 are hydrogen.
[0066] The specific approaches and compounds shown above are not
intended to be limiting. The chemical structures in the schemes
herein depict variables that are hereby defined commensurately with
chemical group definitions (moieties, atoms, etc.) of the
corresponding position in the compound formulae herein, whether
identified by the same variable name (i.e., R.sup.1, R.sup.2,
R.sup.3, etc.) or not. The suitability of a chemical group in a
compound structure for use in the synthesis of another compound is
within the knowledge of one of ordinary skill in the art.
[0067] Additional methods of synthesizing compounds of Formula I or
Formula A and their synthetic precursors, including those within
routes not explicitly shown in schemes herein, are within the means
of chemists of ordinary skill in the art. Synthetic chemistry
transformations and protecting group methodologies (protection and
deprotection) useful in synthesizing the applicable compounds are
known in the art and include, for example, those described in
Larock R, Comprehensive Organic Transformations, VCH Publishers
(1989); Greene, T W et al., Protective Groups in Organic Synthesis,
3.sup.rd Ed., John Wiley and Sons (1999); Fieser, L et al., Fieser
and Fieser's Reagents for Organic Synthesis, John Wiley and Sons
(1994); and Paquette, L, ed., Encyclopedia of Reagents for Organic
Synthesis, John Wiley and Sons (1995) and subsequent editions
thereof.
[0068] Combinations of substituents and variables envisioned by
this invention are only those that result in the formation of
stable compounds.
Compositions
[0069] The invention also provides pyrogen-free pharmaceutical
compositions comprising an effective amount of a compound of
Formula I or Formula A (e.g., including any of the formulae
herein), or a pharmaceutically acceptable salt of said compound;
and a pharmaceutically acceptable carrier. The carrier(s) are
"acceptable" in the sense of being compatible with the other
ingredients of the formulation and, in the case of a
pharmaceutically acceptable carrier, not deleterious to the
recipient thereof in an amount used in the medicament.
[0070] Pharmaceutically acceptable carriers, adjuvants and vehicles
that may be used in the pharmaceutical compositions of this
invention include, but are not limited to, ion exchangers, alumina,
aluminum stearate, lecithin, serum proteins, such as human serum
albumin, buffer substances such as phosphates, glycine, sorbic
acid, potassium sorbate, partial glyceride mixtures of saturated
vegetable fatty acids, water, salts or electrolytes, such as
protamine sulfate, disodium hydrogen phosphate, potassium hydrogen
phosphate, sodium chloride, zinc salts, colloidal silica, magnesium
trisilicate, polyvinyl pyrrolidone, cellulose-based substances,
polyethylene glycol, sodium carboxymethylcellulose, polyacrylates,
waxes, polyethylene-polyoxypropylene-block polymers, polyethylene
glycol and wool fat.
[0071] If required, the solubility and bioavailability of the
compounds of the present invention in pharmaceutical compositions
may be enhanced by methods well-known in the art. One method
includes the use of lipid excipients in the formulation. See "Oral
Lipid-Based Formulations: Enhancing the Bioavailability of Poorly
Water-Soluble Drugs (Drugs and the Pharmaceutical Sciences)," David
J. Hauss, ed. Informa Healthcare, 2007; and "Role of Lipid
Excipients in Modifying Oral and Parenteral Drug Delivery: Basic
Principles and Biological Examples," Kishor M. Wasan, ed.
Wiley-Interscience, 2006.
[0072] Another known method of enhancing bioavailability is the use
of an amorphous form of a compound of this invention optionally
formulated with a poloxamer, such as LUTROL.TM. and PLURONIC.TM.
(BASF Corporation), or block copolymers of ethylene oxide and
propylene oxide. See U.S. Pat. No. 7,014,866; and United States
patent publications 20060094744 and 20060079502.
[0073] The pharmaceutical compositions of the invention include
those suitable for oral, rectal, nasal, topical (including buccal
and sublingual), vaginal or parenteral (including subcutaneous,
intramuscular, intravenous and intradermal) administration. In
certain embodiments, the compound of the formulae herein is
administered transdermally (e.g., using a transdermal patch or
iontophoretic techniques). Other formulations may conveniently be
presented in unit dosage form, e.g., tablets, sustained release
capsules, and in liposomes, and may be prepared by any methods well
known in the art of pharmacy. See, for example, Remington: The
Science and Practice of Pharmacy, Lippincott Williams &
Wilkins, Baltimore, Md. (20th ed. 2000).
[0074] Such preparative methods include the step of bringing into
association with the molecule to be administered ingredients such
as the carrier that constitutes one or more accessory ingredients.
In general, the compositions are prepared by uniformly and
intimately bringing into association the active ingredients with
liquid carriers, liposomes or finely divided solid carriers, or
both, and then, if necessary, shaping the product.
[0075] In certain embodiments, the compound is administered orally.
Compositions of the present invention suitable for oral
administration may be presented as discrete units such as capsules,
sachets, or tablets each containing a predetermined amount of the
active ingredient; a powder or granules; a solution or a suspension
in an aqueous liquid or a non-aqueous liquid; an oil-in-water
liquid emulsion; a water-in-oil liquid emulsion; packed in
liposomes; or as a bolus, etc. Soft gelatin capsules can be useful
for containing such suspensions, which may beneficially increase
the rate of compound absorption.
[0076] In the case of tablets for oral use, carriers that are
commonly used include lactose and corn starch. Lubricating agents,
such as magnesium stearate, are also typically added. For oral
administration in a capsule form, useful diluents include lactose
and dried cornstarch. When aqueous suspensions are administered
orally, the active ingredient is combined with emulsifying and
suspending agents. If desired, certain sweetening and/or flavoring
and/or coloring agents may be added.
[0077] Compositions suitable for oral administration include
lozenges comprising the ingredients in a flavored basis, usually
sucrose and acacia or tragacanth; and pastilles comprising the
active ingredient in an inert basis such as gelatin and glycerin,
or sucrose and acacia.
[0078] Compositions suitable for parenteral administration include
aqueous and non-aqueous sterile injection solutions which may
contain anti-oxidants, buffers, bacteriostats and solutes which
render the formulation isotonic with the blood of the intended
recipient; and aqueous and non-aqueous sterile suspensions which
may include suspending agents and thickening agents. The
formulations may be presented in unit-dose or multi-dose
containers, for example, sealed ampules and vials, and may be
stored in a freeze dried (lyophilized) condition requiring only the
addition of the sterile liquid carrier, for example water for
injections, immediately prior to use. Extemporaneous injection
solutions and suspensions may be prepared from sterile powders,
granules and tablets.
[0079] Such injection solutions may be in the form, for example, of
a sterile injectable aqueous or oleaginous suspension. This
suspension may be formulated according to techniques known in the
art using suitable dispersing or wetting agents (such as, for
example, Tween 80) and suspending agents. The sterile injectable
preparation may also be a sterile injectable solution or suspension
in a non-toxic parenterally-acceptable diluent or solvent, for
example, as a solution in 1,3-butanediol. Among the acceptable
vehicles and solvents that may be employed are mannitol, water,
Ringer's solution and isotonic sodium chloride solution. In
addition, sterile, fixed oils are conventionally employed as a
solvent or suspending medium. For this purpose, any bland fixed oil
may be employed including synthetic mono- or diglycerides. Fatty
acids, such as oleic acid and its glyceride derivatives are useful
in the preparation of injectables, as are natural
pharmaceutically-acceptable oils, such as olive oil or castor oil,
especially in their polyoxyethylated versions. These oil solutions
or suspensions may also contain a long-chain alcohol diluent or
dispersant.
[0080] The pharmaceutical compositions of this invention may be
administered in the form of suppositories for rectal
administration. These compositions can be prepared by mixing a
compound of this invention with a suitable non-irritating excipient
which is solid at room temperature but liquid at the rectal
temperature and therefore will melt in the rectum to release the
active components. Such materials include, but are not limited to,
cocoa butter, beeswax and polyethylene glycols.
[0081] The pharmaceutical compositions of this invention may be
administered by nasal aerosol or inhalation. Such compositions are
prepared according to techniques well-known in the art of
pharmaceutical formulation and may be prepared as solutions in
saline, employing benzyl alcohol or other suitable preservatives,
absorption promoters to enhance bioavailability, fluorocarbons,
and/or other solubilizing or dispersing agents known in the art.
See, e.g.: Rabinowitz J D and Zaffaroni A C, U.S. Pat. No.
6,803,031, assigned to Alexza Molecular Delivery Corporation.
[0082] Topical administration of the pharmaceutical compositions of
this invention is especially useful when the desired treatment
involves areas or organs readily accessible by topical application.
For topical application topically to the skin, the pharmaceutical
composition should be formulated with a suitable ointment
containing the active components suspended or dissolved in a
carrier. Carriers for topical administration of the compounds of
this invention include, but are not limited to, mineral oil, liquid
petroleum, white petroleum, propylene glycol, polyoxyethylene
polyoxypropylene compound, emulsifying wax, and water.
Alternatively, the pharmaceutical composition can be formulated
with a suitable lotion or cream containing the active compound
suspended or dissolved in a carrier. Suitable carriers include, but
are not limited to, mineral oil, sorbitan monostearate, polysorbate
60, cetyl esters wax, cetearyl alcohol, 2-octyldodecanol, benzyl
alcohol, and water. The pharmaceutical compositions of this
invention may also be topically applied to the lower intestinal
tract by rectal suppository formulation or in a suitable enema
formulation. Topically-transdermal patches and iontophoretic
administration are also included in this invention.
[0083] Application of the subject therapeutics may be local, so as
to be administered at the site of interest. Various techniques can
be used for providing the subject compositions at the site of
interest, such as injection, use of catheters, trocars,
projectiles, pluronic gel, stents, sustained drug release polymers
or other device which provides for internal access.
[0084] Thus, according to yet another embodiment, the compounds of
this invention may be incorporated into compositions for coating an
implantable medical device, such as prostheses, artificial valves,
vascular grafts, stents, or catheters. Suitable coatings and the
general preparation of coated implantable devices are known in the
art and are exemplified in U.S. Pat. Nos. 6,099,562; 5,886,026; and
5,304,121. The coatings are typically biocompatible polymeric
materials such as a hydrogel polymer, polymethyldisiloxane,
polycaprolactone, polyethylene glycol, polylactic acid, ethylene
vinyl acetate, and mixtures thereof. The coatings may optionally be
further covered by a suitable topcoat of fluorosilicone,
polysaccharides, polyethylene glycol, phospholipids or combinations
thereof to impart controlled release characteristics in the
composition. Coatings for invasive devices are to be included
within the definition of pharmaceutically acceptable carrier,
adjuvant or vehicle, as those terms are used herein.
[0085] According to another embodiment, the invention provides a
method of coating an implantable medical device comprising the step
of contacting said device with the coating composition described
above. It will be obvious to those skilled in the art that the
coating of the device will occur prior to implantation into a
mammal.
[0086] According to another embodiment, the invention provides a
method of impregnating an implantable drug release device
comprising the step of contacting said drug release device with a
compound or composition of this invention. Implantable drug release
devices include, but are not limited to, biodegradable polymer
capsules or bullets, non-degradable, diffusible polymer capsules
and biodegradable polymer wafers.
[0087] According to another embodiment, the invention provides an
implantable medical device coated with a compound or a composition
comprising a compound of this invention, such that said compound is
therapeutically active.
[0088] According to another embodiment, the invention provides an
implantable drug release device impregnated with or containing a
compound or a composition comprising a compound of this invention,
such that said compound is released from said device and is
therapeutically active.
[0089] Where an organ or tissue is accessible because of removal
from the subject, such organ or tissue may be bathed in a medium
containing a composition of this invention, a composition of this
invention may be painted onto the organ, or a composition of this
invention may be applied in any other convenient way.
[0090] In another embodiment, a composition of this invention
further comprises a second therapeutic agent. The second
therapeutic agent may be selected from any compound or therapeutic
agent known to have or that demonstrates advantageous properties
when administered with a compound having the same mechanism of
action as ruxolitinib. Such agents include those indicated as being
useful in combination with ruxolitinib.
[0091] Preferably, the second therapeutic agent is an agent useful
in the treatment or prevention of a disease or condition selected
from myelofibrosis, including primary myelofibrosis, polycythemia
vera, post-polycythemia vera myelofibrosis, chronic idiopathic
myelofibrosis, post-essential thrombocythemia myelofibrosis, and
essential thrombocythemia, pancreatic cancer, prostate cancer,
breast cancer, leukemia, non-Hodgkin's lymphoma, multiple myeloma,
psoriasis and alopecia areata.
[0092] In one embodiment, the second therapeutic agent is selected
from lenalidomide, panobinostat, capecitabine, exemestane, and
combinations thereof.
[0093] In another embodiment, the invention provides separate
dosage forms of a compound of this invention and one or more of any
of the above-described second therapeutic agents, wherein the
compound and second therapeutic agent are associated with one
another. The term "associated with one another" as used herein
means that the separate dosage forms are packaged together or
otherwise attached to one another such that it is readily apparent
that the separate dosage forms are intended to be sold and
administered together (within less than 24 hours of one another,
consecutively or simultaneously).
[0094] In the pharmaceutical compositions of the invention, the
compound of the present invention is present in an effective
amount. As used herein, the term "effective amount" refers to an
amount which, when administered in a proper dosing regimen, is
sufficient to treat the target disorder.
[0095] The interrelationship of dosages for animals and humans
(based on milligrams per meter squared of body surface) is
described in Freireich et al., Cancer Chemother. Rep, 1966, 50:
219. Body surface area may be approximately determined from height
and weight of the subject. See, e.g., Scientific Tables, Geigy
Pharmaceuticals, Ardsley, N.Y., 1970, 537.
[0096] In one embodiment, an effective amount of a compound of this
invention can range from 1 mg to 500 mg, such as 5 mg to 100 mg,
such as 5 mg to 50 mg. Examples of ranges are from 40 mg to 50 mg,
from 25 mg to 40 mg, from 25 mg to 50 mg, from 20 mg to 40 mg, from
20 mg to 50 mg, from 10 mg to 25 mg, from 10 mg to 20 mg, from 5 mg
to 25 mg, from 5 mg to 20 mg, and from 5 mg to 10 mg. In one
embodiment, a dose of 10 mg, 20 mg, 40 mg, and 50 mg is
administered once a day. In one embodiment a dose of 5 mg, 10 mg,
20 mg, 40 mg, and 50 mg is administered twice a day.
[0097] Effective doses will also vary, as recognized by those
skilled in the art, depending on the diseases treated, the severity
of the disease, the route of administration, the sex, age and
general health condition of the subject, excipient usage, the
possibility of co-usage with other therapeutic treatments such as
use of other agents and the judgment of the treating physician. For
example, guidance for selecting an effective dose can be determined
by reference to the prescribing information for ruxolitinib.
[0098] For pharmaceutical compositions that comprise a second
therapeutic agent, an effective amount of the second therapeutic
agent is between about 20% and 100% of the dosage normally utilized
in a monotherapy regime using just that agent. Preferably, an
effective amount is between about 70% and 100% of the normal
monotherapeutic dose. The normal monotherapeutic dosages of these
second therapeutic agents are well known in the art. See, e.g.,
Wells et al., eds., Pharmacotherapy Handbook, 2nd Edition, Appleton
and Lange, Stamford, Conn. (2000); PDR Pharmacopoeia, Tarascon
Pocket Pharmacopoeia 2000, Deluxe Edition, Tarascon Publishing,
Loma Linda, Calif (2000), each of which references are incorporated
herein by reference in their entirety.
[0099] It is expected that some of the second therapeutic agents
referenced above will act synergistically with the compounds of
this invention. When this occurs, it will allow the effective
dosage of the second therapeutic agent and/or the compound of this
invention to be reduced from that required in a monotherapy. This
has the advantage of minimizing toxic side effects of either the
second therapeutic agent of a compound of this invention,
synergistic improvements in efficacy, improved ease of
administration or use and/or reduced overall expense of compound
preparation or formulation.
Methods of Treatment
[0100] In another embodiment, the invention provides a method of
inhibiting one or more of Janus Associated Kinases (JAKs) JAK1 and
JAK2 in a cell, comprising contacting a cell with one or more
compounds of Formula I or Formula A herein, or a pharmaceutically
acceptable salt thereof.
[0101] According to another embodiment, the invention provides a
method of treating a disease that is beneficially treated by
ruxolitinib in a subject in need thereof, comprising the step of
administering to the subject an effective amount of a compound or a
composition of this invention. In one embodiment the subject is a
patient in need of such treatment. Such diseases are well known in
the art and are disclosed in, but not limited to the following
patent: U.S. Pat. No. 7,598,257. Such diseases include, but are not
limited to, diseases involving the immune system including, for
example, organ transplant rejection (e.g., allograft refection and
graft versus host disease); autoimmune diseases such as multiple
sclerosis, rheumatoid arthritis, juvenile arthritis, type I
diabetes, lupus, psoriasis, inflammatory bowel disease, ulcerative
colitis, Crohn's disease, myasthenia gravis, immunoglobulin
nephropathies, autoimmune thyroid disorders; allergic conditions
such as asthma, food allergies, atopic dermatitis and rhinitis;
viral diseases such as Epstein Barr virus (EBV), hepatitis B,
hepatitis C, HIV, HTLV 1, varicella-zoster virus (VZV) and human
papilloma virus (HPV); skin disorders such as psoriasis (for
example, psoriasis vulgaris), atopic dermatitis, skin rash, skin
irritation, skin sensitization (e.g., contact dermatitis or
allergic contact dermatitis; cancer, including those characterized
by solid tumors (e.g., prostate cancer, renal cancer, hepatic
cancer, pancreatic cancer, gastric cancer, breast cancer, lung
cancer, cancers of the head and neck, thyroid cancer, glioblastoma,
Kaposi's sarcoma, Castleman's disease, melanoma), hematological
cancers (e.g., lymphoma, leukemia such as acute lymphoblastic
leukemia, or multiple myeloma), and skin cancer such as cutaneous
T-cell lymphoma (CTCL) and cutaneous B-cell lymphoma (examples of
which include Sezary syndrome and mycosis fungoides;
myeloproliferative disorders (MPDs) such as polycythemia vera (PV),
essential thrombocythemia (ET), myeloid metaplasia with
myelofibrosis (MMM), chronic myelomonocytic leukemia (CMML),
hypereosinophilic syndrome (HES), systemic mast cell disease
(SMCD); inflammation and inflammatory diseases, such as
inflammatory diseases of the eye (e.g., iritis, uveitis, scleritis,
conjunctivitis, or related disease), inflammatory diseases of the
respiratory tract (e.g., the upper respiratory tract including the
nose and sinuses such as rhinitis or sinusitis or the lower
respiratory tract including bronchitis, chronic obstructive
pulmonary disease, and the like), inflammatory myopathy such as
myocarditis; systemic inflammatory response syndrome (SIRS) and
septic shock; ischemia reperfusion injuries or a disease or
condition related to an inflammatory ischemic event such as stroke
or cardiac arrest; anorexia; cachexia; fatigue such as that
resulting from or associated with cancer; restenosis;
sclerodermitis; fibrosis; conditions associated with hypoxia or
astrogliosis such as, for example diabetic retinopathy, cancer or
neurodegeneration; gout; increased prostate size due to, e.g.,
benign prostatic hypertrophy or benign prostatic hyperplasia.
[0102] In one particular embodiment, the method of this invention
is used to treat a disease or condition selected from
myelofibrosis, including primary myelofibrosis, post-polycythemia
vera myelofibrosis, post-essential thrombocythemia myelofibrosis,
essential thrombocythemia or a combination thereof; pancreatic
cancer; prostate cancer; breast cancer; leukemia, non-Hodgkin's
lymphoma; multiple myeloma;psoriasis and a combination thereof in a
subject in need thereof.
[0103] In another particular embodiment, the method of this
invention is used to treat a disease or condition selected from
myelofibrosis, including primary myelofibrosis, post-polycythemia
vera myelofibrosis and post-essential thrombocythemia myelofibrosis
in a subject in need thereof.
[0104] Identifying a subject in need of such treatment can be in
the judgment of a subject or a health care professional and can be
subjective (e.g. opinion) or objective (e.g. measurable by a test
or diagnostic method).
[0105] In another embodiment, any of the above methods of treatment
comprises the further step of co-administering to the subject in
need thereof one or more second therapeutic agents. The choice of
second therapeutic agent may be made from any second therapeutic
agent known to be useful for co-administration with ruxolitinib.
The choice of second therapeutic agent is also dependent upon the
particular disease or condition to be treated. Examples of second
therapeutic agents that may be employed in the methods of this
invention are those set forth above for use in combination
compositions comprising a compound of this invention and a second
therapeutic agent.
[0106] In particular, the combination therapies of this invention
include co-administering a compound of Formula I or Formula A and a
second therapeutic agent to a subject in need thereof for treatment
of the following conditions (with the particular second therapeutic
agent indicated in parentheses following the indication:
myelofibrosis (lenalidomide or panobinostat); pancreatic cancer
(capecitabine); and breast cancer (exemestane).
[0107] The term "co-administered" as used herein means that the
second therapeutic agent may be administered together with a
compound of this invention as part of a single dosage form (such as
a composition of this invention comprising a compound of the
invention and an second therapeutic agent as described above) or as
separate, multiple dosage forms. Alternatively, the additional
agent may be administered prior to, consecutively with, or
following the administration of a compound of this invention. In
such combination therapy treatment, both the compounds of this
invention and the second therapeutic agent(s) are administered by
conventional methods. The administration of a composition of this
invention, comprising both a compound of the invention and a second
therapeutic agent, to a subject does not preclude the separate
administration of that same therapeutic agent, any other second
therapeutic agent or any compound of this invention to said subject
at another time during a course of treatment.
[0108] Effective amounts of these second therapeutic agents are
well known to those skilled in the art and guidance for dosing may
be found in patents and published patent applications referenced
herein, as well as in Wells et al., eds., Pharmacotherapy Handbook,
2nd Edition, Appleton and Lange, Stamford, Conn. (2000); PDR
Pharmacopoeia, Tarascon Pocket Pharmacopoeia 2000, Deluxe Edition,
Tarascon Publishing, Loma Linda, Calif. (2000), and other medical
texts. However, it is well within the skilled artisan's purview to
determine the second therapeutic agent's optimal effective-amount
range.
[0109] In one embodiment of the invention, where a second
therapeutic agent is administered to a subject, the effective
amount of the compound of this invention is less than its effective
amount would be where the second therapeutic agent is not
administered. In another embodiment, the effective amount of the
second therapeutic agent is less than its effective amount would be
where the compound of this invention is not administered. In this
way, undesired side effects associated with high doses of either
agent may be minimized. Other potential advantages (including
without limitation improved dosing regimens and/or reduced drug
cost) will be apparent to those of skill in the art.
[0110] In yet another aspect, the invention provides the use of a
compound of Formula I or Formula A alone or together with one or
more of the above-described second therapeutic agents in the
manufacture of a medicament, either as a single composition or as
separate dosage forms, for treatment or prevention in a subject of
a disease, disorder or symptom set forth above. Another aspect of
the invention is a compound of Formula I or Formula A for use in
the treatment or prevention in a subject of a disease, disorder or
symptom thereof delineated herein.
EXAMPLES
Example 1
Synthesis of
(R)-3-(4-(7H-Pyrrolo[2,3-d]pyrimidin-4-yl)-1H-pyrazol-1-yl)-3
-(2,2,5,5-d.sub.4-cyclopentyl)propanenitrile (Compound 107)
##STR00013## ##STR00014##
[0112] Step 1. Diethyl
2,2,5,5-d.sub.4-cyclopentane-1,1-dicarboxylate (32). To a solution
of diethyl malonate (6.57 mL, 43.3 mmol) in ethanol (40 mL) was
added a 21 wt % solution of sodium ethoxide in ethanol (32.3 mL,
86.6 mmol) followed by 1,1,4,4-tetradeutero-1,4-dibromobutane (31,
5.53 mL, 45.5 mmol, CDN Isotopes, 98 atom % D). The resulting
solution was stirred at reflux for two hours then cooled to room
temperature and diluted with excess water. The majority of the
ethanol was then removed via distillation and the resulting aqueous
solution was extracted with ethyl acetate (3.times.75 mL). The
organic layers were combined, washed with brine, dried
(Na.sub.2SO.sub.4), filtered and concentrated under reduced
pressure to afford 32 as a yellow oil which was carried forward
without purification. (9.45 g, 100%).
[0113] Step 2. 2,2,5,5-d.sub.4-Cyclopentane-1-carboxylic acid (33).
To a solution of 32 (9.45 g, 43.3 mmol) in ethanol (20 mL) was
added a 5M solution of sodium hydroxide (20 mL). Additional water
(15 mL) was then added and the reaction stirred at reflux for three
hours. Upon cooling to room temperature, the reaction was diluted
with excess water and the majority of ethanol was removed via
distillation. The aqueous solution was rendered acidic (pH<2)
with 1N HCl and subsequently extracted with diethyl ether
(3.times.50 mL). The organic layers were combined, dried
(Na.sub.2SO.sub.4), filtered and concentrated under reduced
pressure. The resulting light orange solid was transfered to a
pressure flask and water (140 mL) was added. The pressure flask was
sealed and the reaction stirred at 160.degree. C. for 15 hours then
was cooled to room temperature. The reaction was diluted with 1N
HCl and extracted with diethyl ether (3.times.50 mL). The organic
layers were combined, dried (Na.sub.2SO.sub.4), filtered and
concentrated under reduced pressure to afford 33 (4.37 g, 86%) as
an amber oil which was used without purification.
[0114] Step 3.
2,2,5,5-d.sub.4-N-Methoxy-N-methylcyclopentanecarboxamide (34). To
a solution of 33 (4.37 g, 37.0 mmol) in acetonitrile (60 mL) at
0.degree. C. was added N,O-dimethylhydroxylamine hydrochloride
(4.33 g, 44.4 mmol), TBTU (12.5 g, 38.9 mmol) and
N,N-diisopropylethylamine (19.0 mL, 111 mmol). The reaction stirred
at room temperature for 15 hours, then was diluted with 1N HCl and
extracted with ethyl acetate (3.times.50 mL). The organic layers
were combined, washed with sat. NaHCO.sub.3, dried
(Na.sub.2SO.sub.4), filtered and concentrated under reduced
pressure. The reulting product was purified by column
chromatography (SiO2, 0-50% ethyl acetate/hexanes) to afford 34
(2.22 g, 37%) as a clear oil. MS (ESI) 162.3 [(M+H).sup.+].
[0115] Step 4. 2,2,5,5-d.sub.4-Cyclopentane-1-carboxaldehyde (35).
To a solution of 34 (2.22 g, 13.8 mmol) in THF (50 mL) at 0.degree.
C. was added dropwise a 1M solution of LiAlH.sub.4 in THF (24.8 mL,
24.8 mmol). The reaction stirred at 0.degree. C. for one hour then
was quenched by sequential dropwise addition of water (940 .mu.L),
15% NaOH (940 .mu.L) and water (2.82 mL). The quenched reaction
stirred at room temperature for 30 minutes then was filtered
through Celite.RTM. and concentrated under reduced pressure. The
resulting oil was diluted with 1N HCl and extracted with diethyl
ether (3.times.50 mL). The organic layers were combined, dried
(Na.sub.2SO.sub.4), filtered and concentrated under reduced
pressure to afford 35 (850 mg, 60%) as a clear oil which was used
without purification.
[0116] Step 5. 3-(2,2,5,5-d.sub.4-cyclopentyl)acrylonitrile (36).
To a 1M solution of potassium tert-butoxide in THF (8.74 mL, 8.74
mmol) at 0.degree. C. was added dropwise a solution of diethyl
cyanomethylphosphonate (1.48 mL, 9.15 mmol) in THF (12 mL). The
reaction was warmed to room temperature, stirred for 15 minutes,
then cooled to 0.degree. C. Aldehyde 35 (850 mg, 8.32 mmol) was
then added dropwise as a solution in THF (3 mL). The reaction was
stirred at room temperature for 48 hours then diluted with excess
water and extracted with diethyl ether (1.times.50 mL) and ethyl
acetate (3.times.50 mL). The organic layers were combined, dried
(Na.sub.2SO.sub.4), filtered and concentrated under reduced
pressure to afford 36 (1.17 g, >100%) as a light orange oil
which was used without purification.
[0117] Step 6.
(+/-)-(4-(1-(2-Cyano-1-(2,2,5,5-d.sub.4-cyclopentyl)ethyl)-1H-pyrazol-4-y-
l)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)methyl pivalate ((+/-)38). To a
solution of 37 (400 mg, 1.34 mmol, preparation described in Lin, Q.
et al. Org. Lett., 2009, 11, 1999-2002) in acetonitrile (10 mL) was
added 36 (418 mg, 3.34 mmol) followed by DBU (421 .mu.L, 2.81
mmol). The reaction stirred at room temperature for 15 hours then
was concentrated under reduced vacuum. The resulting crude mixture
was diluted with water and extracted with ethyl acetate (3.times.50
mL). The organic layers were combined, washed with 1N HCl, dried
(Na.sub.2SO.sub.4), filtered and concentrated underreduced
pressure. Purification via normal phase column chromatography
(SiO.sub.2, 0-60% ethyl acetate/hexanes) followed by reverse phase
column chromatography (C18, 5-70% acetonitrile/water containing
0.1% formic acid) afforded (+/-)38 (68 mg, 12%) as a white foam. 1H
NMR (DMSO-d.sub.6, 400 MHz) .delta. 8.84 (s, 1H), 8.79 (s, 1H),
8.40 (s, 1H), 7.74 (d, J=3.8 Hz, 1H), 7.12 (d, J=3.8 Hz, 1H), 6.24
(s, 2H), 4.54 (td, J=9.7, 4.3 Hz, 1H), 3.30-3.15 (m, 2H), 2.39 (d,
J=9.8 Hz, 1H), 1.68-1.36 (m, 4H), 1.08 (s, 9H); MS (ESI) 425.3
[(M+H).sup.+].
[0118] Step 7.
(R)-(4-(1-(2-cyano-1-(2,2,5,5-tetradeuterocyclopentyl)ethyl)-1H-pyrazol-4-
-yl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)methyl pivalate ((R)-38).
Racemic compound (+/-)38 (62 mg) was dissolved in acetonitrile at a
concentration of 30 mg/mL and subjected to chiral separation by
preparative HPLC on a Daicel ChiralPak AD column (20.times.250 mm,
10 .mu.m) with 500 .mu.L of (+/-)38 solution per injection using an
isocratic method: 30% isopropanol (+0.1% diethylamine)/70% hexane
(+0.1% diethylamine) at a flow rate of 17 mL/min. Under these
conditions baseline separation was achieved with (S)-38 eluting at
15.0 minutes and (R)-38 eluting at 20.2 minutes.
[0119] Fractions containing each enantiomer were pooled and
concentrated yielding 28 mg of (S)-38 as a colorless film and 29 mg
of (R)-38 as a colorless film.
[0120] Step 8.
(R)-3-(4-(7H-Pyrrolo[2,3-d]pyrimidin-4-yl)-1H-pyrazol-1-yl)-3-(2,2,5,5-te-
tradeuterocyclopentyl)propanenitrile (Compound 107). Compound
(R)-38 (28 mg, 0.066 mmol, 1 equiv) was dissolved in methanol (1
mL) in a 20 mL scintillation vial. Sodium hydroxide (0.13 mL of a 1
M solution, 0.13 mmol, 2 equiv) was added and the reaction was
stirred at room temperature for 18 hours. The reaction was diluted
with water (10 mL) and brine (20 mL). The aqueous mixture was
extracted with ethyl acetate (2.times.20 mL). The combined organic
layers were washed with brine (20 mL), dried over sodium sulfate,
filtered, and evaporated. The crude material was purified using an
Analogix automated chromatography system eluting with 0 to 6%
methanol in dichloromethane. Product fractions were pooled and
evaporated yielding compound 107 as a white foam. The chiral purity
was found to be >99% ee (Chiralpak OD 4.6.times.250 mm, 10 um,
70% (hexane+0.1% diethylamine)+30% (isopropanol+0.1% diethylamine),
1 mL/min, 254 nm retention time=8.85 min).
Example 2
Synthesis of
(R)-3-(4-(7H-Pyrrolo[2,3-d]pyrimidin-4-yl)-1H-pyrazol-1-yl)-3-(3,3,4,4-d.-
sub.4-cyclopentyl)propanenitrile (Compound 103).
##STR00015## ##STR00016##
[0122] Step 1. Diethyl
3,3,4,4-d.sub.4-cyclopentane-1,1-dicarboxylate (40). To a solution
of diethyl malonate (3.25 mL, 21.4 mmol) in ethanol (20 mL) was
added a 21 wt % solution of sodium ethoxide in ethanol (16.0 mL,
42.8 mmol) followed by 2,2,3,3-tetradeutero-1,4-dibromobutane (39,
4.95 g, 22.5 mmol, CDN Isotopes, 98 atom % D). The resulting
solution was stirred at reflux for two hours then cooled to room
temperature and diluted with excess water. The majority of the
ethanol was then removed via distillation and the resulting aqueous
solution was extracted with ethyl acetate (3.times.75 mL). The
organic layers were combined, washed with brine, dried
(Na.sub.2SO.sub.4), filtered and concentrated under reduced
pressure to afford 40 as a yellow oil which was carried forward
without purification. (4.67 g, 100%).
[0123] Step 2. 3,3,4,4-d.sub.4-Cyclopentane-1-carboxylic acid (41).
To a solution of 40 (4.67 g, 21.4 mmol) in ethanol (10 mL) was
added a 5M solution of sodium hydroxide (10 mL). Additional water
(10 mL) was then added and the reaction stirred at reflux for three
hours. Upon cooling to room temperature, the reaction was diluted
with excess water and the majority of ethanol was removed via
distillation. The aqueous solution was rendered acidic (pH<2)
with 1N HCl and subsequently extracted with diethyl ether
(3.times.50 mL). The organic layers were combined, dried
(Na.sub.2SO.sub.4), filtered and concentrated under reduced
pressure. The resulting light orange solid was transfered to a
pressure flask and water (70 mL) was added. The pressure flask was
sealed and the reaction stirred at 160.degree. C. for 15 hours then
was cooled to room temperature. The reaction was diluted with 1N
HCl and extracted with diethyl ether (3.times.50 mL). The organic
layers were combined, dried (Na.sub.2SO.sub.4), filtered and
concentrated under reduced pressure to afford 41 (1.93 g, 76%) as
an amber oil which was used without purification.
[0124] Step 3.
3,3,4,4-d.sub.4-N-Methoxy-N-methylcyclopentanecarboxamide (42). To
a solution of 41 (1.93 g, 16.3 mmol) in acetonitrile (30 mL) at
0.degree. C. was added N,O-dimethylhydroxylamine hydrochloride
(1.91 g, 19.6 mmol), TBTU (5.50 g, 17.1 mmol) and
N,N-diisopropylethylamine (8.52 mL, 48.9 mmol). The reaction
stirred at room temperature for 15 hours, then was diluted with 1N
HCl and extracted with ethyl acetate (3.times.50 mL). The organic
layers were combined, washed with sat. NaHCO.sub.3, dried
(Na.sub.2SO.sub.4), filtered and concentrated under reduced
pressure. The resulting product was purified by column
chromatography (SiO.sub.2, 0-40% acetone/hexanes) to afford 42
(1.47 g, 56%) as a clear oil. MS (ESI) 162.3 [(M+H).sup.+].
[0125] Step 4. 3,3,4,4-d.sub.4-Cyclopentane-1-carboxaldehyde (43).
To a solution of 42 (1.47 g, 9.12 mmol) in THF (35 mL) at 0.degree.
C. was added dropwise a 1M solution of LiAlH.sub.4 in THF (16.4 mL,
16.4 mmol). The reaction stirred at room temperature for one hour
then was quenched at 0.degree. C. by sequential dropwise addition
of water (623 .mu.L), 15% NaOH (623 .mu.L) and water (1.87 mL). The
quenched reaction stirred at room temperature for 30 minutes then
was filtered through Celite.RTM. and concentrated under reduced
pressure. The resulting oil was diluted with 1N HCl and extracted
with diethyl ether (3.times.50 mL). The organic layers were
combined, dried (Na.sub.2SO.sub.4), filtered and concentrated under
reduced pressure to afford 43 (767 mg, 82%) as a clear oil which
was used without purification.
[0126] Step 5. 3-(3,3,4,4-d.sub.4-Cyclopentyl)acrylonitrile (44).
To a solution of diethyl cyanomethylphosphonate (0.607 mL, 3.75
mmol) in THF (10 mL) at 0.degree. C. was added dropwise a 1M
solution of potassium tert-butoxide in THF (3.75 mL, 3.75 mmol).
The reaction stirred at 0.degree. C. for 1 hour. Aldehyde 43 (767
mg, 7.51 mmol) was then added dropwise as a solution in THF (3 mL).
The reaction was stirred at room temperature for 15 hours then
diluted with excess 1:1 water/brine and extracted with MTBE
(3.times.50 mL). The organic layers were combined, dried
(Na.sub.2SO.sub.4), filtered and concentrated under reduced
pressure. The resulting oil was dissolved in CH.sub.2Cl.sub.2 (100
ml) and washed with NaHSO.sub.3 (3.times.25 mL). The organic layer
was dried (Na.sub.2SO.sub.4), filtered and concentrated under
reduced pressure to afford 44 (537 mg, 57%) as a light orange oil
which was used without purification.
[0127] Step 6.
(+/-)-(4-(1-(2-Cyano-1-(3,3,4,4-d.sub.4-cyclopentyl)ethyl)-1H-pyrazol-4-y-
l)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)methyl pivalate ((+/-)45). To a
solution of 37 (514 mg, 1.72 mmol, preparation described in Lin, Q.
et al. Org. Lett., 2009, 11, 1999-2002) in acetonitrile (15 mL) was
added 44 (537 mg, 4.29 mmol) followed by DBU (540 .mu.L, 3.61
mmol). The reaction stirred at room temperature for 15 hours then
was concentrated under reduced vacuum. The resulting crude mixture
was diluted with water and extracted with ethyl acetate (3.times.50
mL). The organic layers were combined, washed with 1N HCl, dried
(Na.sub.2SO.sub.4), filtered and concentrated under reduced
pressure. Purification via normal phase column chromatography
(SiO.sub.2, 0-60% ethyl acetate/hexanes) afforded (+/-) 45 (368 mg,
50%) as a white foam. 1H NMR (DMSO-d.sub.6, 400 MHz) .delta. 8.84
(s, 1H), 8.79 (s, 1H), 8.40 (s, 1H), 7.75 (d, J=3.7 Hz, 1H), 7.12
(d, J=3.7 Hz, 1H), 6.24 (s, 2H), 4.53 (td, J=9.7, 4.2 Hz, 1H),
3.32-3.14 (m, 2H), 2.41 (q, J=8.7 Hz, 1H), 1.79 (dd, J=12.6, 7.6
Hz, 1H), 1.36-1.11 (m, 3H), 1.08 (s, 9H).; MS (ESI) 425.2
[(M+H).sup.+].
[0128] Step 7.
(R)-(4-(1-(2-Cyano-1-(3,3,4,4-d.sub.4-cyclopentyl)ethyl)-1H-pyrazol-4-yl)-
-7H-pyrrolo[2,3-d]pyrimidin-7-yl)methyl pivalate ((R)-45). Racemic
compound (+/-)45 (151 mg) was dissolved in acetonitrile at a
concentration of 30 mg/mL and subjected to chiral separation by
preparative HPLC on a Daicel ChiralPak AD column (20.times.250 mm,
10 .mu.m) with 1000 .mu.L of (+/-)45 solution per injection using
an isocratic method: 30% isopropanol (+0.1% diethylamine)/70%
hexane (+0.1% diethylamine) at a flow rate of 17 mL/min. Under
these conditions baseline separation was achieved with (S)-45
eluting at 15.5 minutes and (R)-45 eluting at 20.7 minutes.
[0129] Fractions containing each enantiomer were pooled separately
and concentrated to give g 51 mg of (S)-45 as a colorless film and
53 mg of (R)-45 as a colorless film.
[0130] Step 8.
(R)-3-(4-(7H-Pyrrolo[2,3-d]pyrimidin-4-yl)-1H-pyrazol-1-yl)-3-(3,3,4,4-d.-
sub.4-cyclopentyl)propanenitrile (Compound 103). (R)-45 (53 mg,
0.13 mmol, 1 equiv) was dissolved in methanol (2 mL) in a 20 mL
scintillation vial. Sodium hydroxide (0.25 mL of a 1 M solution,
0.25 mmol, 2 equiv) was added and the reaction mixture was stirred
at room temperature for 18 hours. The reaction mixture was diluted
with water (10 mL) and brine (20 mL). The aqueous mixture was
extracted with ethyl acetate (2.times.20 mL). The combined organic
layers were washed with brine (20 mL), dried over sodium sulfate,
filtered, and concentrated. The crude material was purified using
an Analogix automated chromatography system eluting with 0 to 6%
methanol in dichloromethane. Product fractions were pooled and
evaporated to give Compound 103 as a white foam in .about.90%
purity with the incompletely deprotected hydroxymethyl intermediate
as the main impurity. Further chromatography failed to further
improve the purity. The 90% pure material was dissolved in THF (2
mL) and treated with several drops of 10% aqueous sodium hydroxide
at 40.degree. C. for 8 hours resulting in complete conversion to
Compound 103. The reaction mixture was diluted with water (10 mL)
and extracted with ethyl acetate (2.times.10 mL). The combined
organic layers were dried over sodium sulfate, filtered, and
concentrated to a white foam. The foam was dissolved in minimal
acetonitrile, diluted with water, and lyophilized to give Compound
103 (14 mg, 35% yield) as a white solid. The chiral purity was
found to be >99% ee (Chiralpak OD 4.6.times.250 mm, 10 um, 70%
(hexane+0.1% diethylamine)+30% (isopropanol+0.1% diethylamine), 1
mL/min, 254 nm retention time=7.56 min).
Example 3
Synthesis of
(R)-3-(4-(7H-Pyrrolo[2,3-d]pyrimidin-4-yl)-1H-byrazol-1-yl)-3-(cyclopenty-
l-d.sub.9)propanenitrile (Compound 127).
##STR00017## ##STR00018##
[0132] Step 1. Diethyl
2,2,3,3,4,4,5,5-d.sub.8-Cyclopentane-1,1-dicarboxylate (47). To a
solution of diethyl malonate (6.24 mL, 41.1 mmol) in ethanol (40
mL) was added a 21 wt % solution of sodium ethoxide in ethanol
(30.7 mL, 82.2 mmol) followed by
1,1,2,2,3,3,4,4-octadeutero-1,4-dibromobutane (46, 9.67 g, 43.2
mmol, CDN Isotopes, 98 atom % D). The resulting solution was
stirred at reflux for two hours then cooled to room temperature and
diluted with excess water. The majority of the ethanol was then
removed via distillation and the resulting aqueous solution was
extracted with ethyl acetate (3.times.75 mL). The organic layers
were combined, washed with brine, dried (Na.sub.2SO.sub.4),
filtered and concentrated under reduced pressure to afford 47 as a
yellow oil (9.12 g, 100%) which was carried forward without
purification.
[0133] Step 2. Perdeuterocyclopentane-1-carboxylic acid (48). To a
solution of 47 (9.12 g, 41.1 mmol) in ethanol (20 mL) was added a
5M solution of sodium hydroxide (20 mL). Additional water (15 mL)
was then added and the reaction stirred at reflux for three hours.
Upon cooling to room temperature, the reaction was diluted with
excess water and the majority of ethanol was removed via
distillation. The aqueous solution was rendered acidic (pH<2)
with 1N HCl and subsequently extracted with diethyl ether
(3.times.50 mL). The organic layers were combined, dried
(Na.sub.2SO.sub.4), filtered and concentrated under reduced
pressure. The resulting light orange solid was transfered to a
pressure flask and D.sub.2O (120 mL) was added. The pressure flask
was sealed and the reaction stirred at 160.degree. C. for 15 hours
then was cooled to room temperature. The reaction was diluted with
1N HCl and extracted with diethyl ether (3.times.50 mL). The
organic layers were combined, dried (Na.sub.2SO.sub.4), filtered
and concentrated under reduced pressure to afford 48 (4.58 g, 90%)
as a yellow oil which was used without purification.
[0134] Step 3. N-Methoxy-N-methyl(cyclopentane-d.sub.9)carboxamide
(49). To a solution of 48 (4.58 g, 37.2 mmol) in acetonitrile (60
mL) at 0.degree. C. was added N,O-dimethylhydroxylamine
hydrochloride (4.35 g, 44.6 mmol), TBTU (12.5 g, 39.1 mmol) and
N,N-diisopropylethylamine (19.4 mL, 112 mmol). The reaction stirred
at room temperature for 15 hours, then was diluted with 1N HCl and
extracted with ethyl acetate (3.times.50 mL). The organic layers
were combined, washed with sat. NaHCO.sub.3, dried
(Na.sub.2SO.sub.4), filtered and concentrated under reduced
pressure. The resulting product was purified by column
chromatography (SiO.sub.2, 0-50% ethyl acetate/hexanes) to afford
49 (3.41 g, 55%) as a clear oil. MS (ESI) 167.2 [(M+H).sup.+].
[0135] Step 4. Perdeuterocyclopentane-1-carboxaldehyde (50). To a
solution of 49 (3.41 g, 20.5 mmol) in THF (80 mL) at 0.degree. C.
was added dropwise a 1M solution of LiAlH.sub.4 in THF (37.0 mL,
37.0 mmol). The reaction stirred at room temperature for one hour
then was quenched at 0.degree. C. by sequential dropwise addition
of D.sub.2O (1.41 mL), 15% NaOD/D.sub.2O (1.41 mL) and D.sub.2O
(4.23 mL). The quenched reaction stirred at room temperature for 30
minutes then was filtered through Celite.RTM. and concentrated
under reduced pressure. The resulting oil was diluted with 1N
DCl/D.sub.2O and extracted with diethyl ether (3.times.50 mL). The
organic layers were combined, dried (MgSO.sub.4), filtered and
concentrated under reduced pressure to afford 50 (1.79 g, 82%) as a
clear oil which was used without purification.
[0136] Step 5. 3-(Perdeuterocyclopentyl)acrylonitrile (51). To a
solution of diethyl cyanomethylphosphonate (1.35 mL, 8.34 mmol) in
THF (25 mL) at 0.degree. C. was added dropwise a 1M solution of
potassium tert-butoxide in THF (8.34 mL, 8.34 mmol). The reaction
stirred at 0.degree. C. for 1 hour. Aldehyde 50 (1.79 g, 16.7 mmol)
was then added dropwise as a solution in THF (5 mL). The reaction
was stirred at room temperature for 15 hours then diluted with
excess 1:1 water/brine and extracted with MTBE (3.times.50 mL). The
organic layers were combined, dried (Na.sub.2SO.sub.4), filtered
and concentrated underreduced pressure. The organic layers were
combined, dried (Na.sub.2SO.sub.4), filtered and concentrated
underreduced pressure to afford 51 (1.61 g, 74%) as a light orange
oil which was used without purification.
[0137] Step 6.
(+/-)-(4-(1-(2-Cyano-1-(cyclopentyl-d.sub.9)ethyl)-1H-pyrazol-4-yl)-7H-py-
rrolo[2,3-d]pyrimidin-7-yl)methyl pivalate ((+/-)52). To a solution
of 37 (619 mg, 2.07 mmol, preparation described in Lin, Q. et al.
Org. Lett., 2009, 11, 1999-2002) in acetonitrile (15 mL) was added
51 (673 mg, 5.17 mmol) followed by DBU (650 .mu.L, 4.35 mmol). The
reaction stirred at room temperature for 15 hours then was
concentrated under reduced vacuum. The resulting crude mixture was
diluted with water and extracted with ethyl acetate (3.times.50
mL). The organic layers were combined, washed with 1N HCl, dried
(Na.sub.2SO.sub.4), filtered and concentrated underreduced
pressure. Purification via normal phase column chromatography
(SiO.sub.2, 0-60% ethyl acetate/hexanes) afforded (+/-)52 (447 mg,
50%) as a white foam. 1H NMR (DMSO-d.sub.6, 400 MHz) .delta. 8.84
(s, 1H), 8.79 (s, 1H), 8.39 (s, 1H), 7.75 (d, J=3.7 Hz, 1H), 7.12
(d, J=3.7 Hz, 1H), 6.24 (s, 2H), 4.53 (dd, J=9.6, 4.2 Hz, 1H),
3.32-3.13 (m, 2H), 1.08 (s, 9H).; MS (ESI) 430.3[(M+H).sup.+].
[0138] Step 7.
(R)-(4-(1-(2-Cyano-1-(cyclopentyl-d)ethyl)-1H-pyrazol-4-yl)-7H-pyrrolo[2,-
3-d]pyrimidin-7-yl)methyl pivalate ((R)-52). Racemic compound
(+/-)52 (162 mg) was dissolved in acetonitrile at a concentration
of 30 mg/mL and subjected to chiral separation by preparative HPLC
on a Daicel ChiralPak AD column (20.times.250 mm, 10 .mu.m) with
1000 .mu.L, of (+/-)52 solution per injection using an isocratic
method: 30% isopropanol (+0.1% diethylamine)/70% hexane (+0.1%
diethylamine) at a flow rate of 17 mL/min. Under these conditions
baseline separation was achieved with (S)-52 eluting at 15.4
minutes and (R)-52 eluting at 20.5 minutes.
[0139] Fractions containing each enantiomer were pooled separately
and concentrated to give 61 mg of (S)-52 as a colorless film and 63
mg of (R)-52 as a colorless film.
[0140] Step 8.
(R)-3-(4-(7H-Pyrrolo[2,3-d]pyrimidin-4-yl)-1H-pyrazol-1-yl)-3-(cyclopenty-
l-d.sub.9)propanenitrile (Compound 127). (R)-52 (60 mg, 0.14 mmol,
1 equiv) was dissolved in methanol (2 mL) in a 20 mL scintillation
vial. Sodium hydroxide (0.28 mL of a 1 M solution, 0.28 mmol, 2
equiv) was added and the reaction mixture was stirred at room
temperature for 18 hours. The reaction mixture was diluted with
water (10 mL) and brine (20 mL). The aqueous mixture was extracted
with ethyl acetate (2.times.20 mL). The combined organic layers
were washed with brine (20 mL), dried over sodium sulfate,
filtered, and concentrated. The crude material was purified using
an Analogix automated chromatography system eluting with 0 to 6%
methanol in dichloromethane. Product fractions were pooled and
evaporated to give Compound 127 (34 mg) as a white foam in
.about.90% purity with the incompletely deprotected hydroxymethyl
intermediate as the main impurity. Further chromatography failed to
further improve the purity. The 90% pure material was dissolved in
THF (2 mL) and treated with several drops of 10% aqueous sodium
hydroxide at 40.degree. C. for 8 hours resulting in complete
conversion to Compound 127. The reaction mixture was diluted with
water (10 mL) and extracted with ethyl acetate (2.times.10 mL). The
combined organic layers were dried over sodium sulfate, filtered,
and concentrated to a white foam. The foam was dissolved in minimal
acetonitrile, diluted with water, and lyophilized to give Compound
127 (19 mg, 42% yield) as a white solid. The chiral purity was
found to be >99% ee (Chiralpak OD 4.6.times.250 mm, 10 um, 70%
(hexane+0.1% diethylamine)+30% (isopropanol+0.1% diethylamine), 1
mL/min, 254 nm retention time=7.55 min).
Example 4
Evaluation of Metabolic Stability in CYP3A4 Supersomes.TM.
[0141] Evaluation of Metabolic Stability of Compounds 103, 107 and
127 in Human CYP3A4 Supersomes.TM..
[0142] SUPERSOMES.TM. Assay. 10 mM stock solutions of test
compounds, Compounds 103, 107, 127 and ruxolitinib, were prepared
in DMSO. The 10 mM stock solutions were diluted to 15.6 .mu.M in
acetonitrile (ACN). Human CYP3A4 supersomes.TM. (1000 pmol/mL,
purchased from BD Gentest.TM. Products and Services) were diluted
to 62.5 pmol/mL in 0.1 M potassium phosphate buffer, pH 7.4,
containing 3 mM MgCl.sub.2. The diluted supersomes were added to
wells of a 96-well polypropylene plate in triplicate. A 10 .mu.L
aliquot of the 15.6 .mu.M test compound was added to the supersomes
and the mixture was pre-warmed for 10 minutes. Reactions were
initiated by addition of pre-warmed NADPH solution. The final
reaction volume was 0.5 mL and contained 50 pmol/mL CYP3A4
supersomes.TM., 0.25 .mu.M test compound, and 2 mM NADPH in 0.1 M
potassium phosphate buffer, pH 7.4, and 3 mM MgCl.sub.2. The
reaction mixtures were incubated at 37.degree. C., and 50 .mu.L
aliquots were removed at 0, 5, 10, 20, and 30 minutes and added to
96-well plates which contained 50 .mu.L of ice-cold ACN with
internal standard to stop the reactions. The plates were stored at
4.degree. C. for 20 minutes after which 100 mL of water was added
to the wells of the plate before centrifugation to pellet
precipitated proteins. Supernatants were transferred to another
96-well plate and analyzed for amounts of parent remaining by
LC-MS/MS using an Applied Bio-systems API 4000 mass
spectrometer.
[0143] Data analysis: The in vitro half-lives (t.sub.1/2 values)
for test compounds were calculated from the slopes of the linear
regression of LN (% parent remaining) vs incubation time
relationship:
in vitro t.sub.1/2=0.693/k, where k=-[slope of linear regression of
% parent remaining (ln) vs incubation time].
[0144] The results of this experiment are shown in Table 3 and FIG.
1. As shown in Table 3, the half-life of ruxolitinib was calculated
to be 14.5 minutes. In contrast, each of Compounds 103, 107 and 127
were more stable in the supersomes with calculated half-lives of
16.9, 17.9 and 32.0 minutes respectively. This respresents a 17%
increase in t.sub.1/2 for compound 103, a 23% increase in t.sub.1/2
for compound 107, and a 121% increase in t.sub.1/2 for compound
127.
TABLE-US-00003 TABLE 3 Metabolic Stability of Compounds 103, 107
and 127 versus Ruxolitinib in Human CYP3A4 Supersomes .TM.
t.sub.1/2 (minutes) Compound Experiment 1 Experiment 2 Ave .+-. SD
Ruxolitinib 14.5 14.5 14.5 .+-. 00 Compound 103 17.5 16.3 16.9 .+-.
0.9 (17%*) Compound 107 18.4 17.0 17.9 .+-. 1.0 (23%*) Compound 127
31.4 32.1 32.0 .+-. 0.5 (121%*) *% .DELTA. = [(deuterated species)
- (nondeuterated species)](100)/(nondeuterated species)
Example 5
Evaluation of Metabolic Stability in Human Liver Microsomes
[0145] Microsomal Assay: Human liver microsomes (20 mg/mL) are
obtained from Xenotech, LLC (Lenexa, Kans.). .beta.-nicotinamide
adenine dinucleotide phosphate, reduced form (NADPH), magnesium
chloride (MgCl.sub.2), and dimethyl sulfoxide (DMSO) are purchased
from Sigma-Aldrich.
[0146] Determination of Metabolic Stability: 7.5 mM stock solutions
of test compounds are prepared in DMSO. The 7.5 mM stock solutions
are diluted to 12.5-50 82 M in acetonitrile (ACN). The 20 mg/mL
human liver microsomes are diluted to 0.625 mg/mL in 0.1 M
potassium phosphate buffer, pH 7.4, containing 3 mM MgCl.sub.2. The
diluted microsomes are added to wells of a 96-well deep-well
polypropylene plate in triplicate. A 10 .mu.L aliquot of the
12.5-50 .mu.M test compound is added to the microsomes and the
mixture is pre-warmed for 10 minutes. Reactions are initiated by
addition of pre-warmed NADPH solution. The final reaction volume is
0.5 mL and contains 0.5 mg/mL human liver microsomes, 0.25-1.0
.mu.M test compound, and 2 mM NADPH in 0.1 M potassium phosphate
buffer, pH 7.4, and 3 mM MgCl.sub.2. The reaction mixtures are
incubated at 37.degree. C., and 50 .mu.L aliquots are removed at 0,
5, 10, 20, and 30 minutes and added to shallow-well 96-well plates
which contain 50 .mu.L of ice-cold ACN with internal standard to
stop the reactions. The plates are stored at 4.degree. C. for 20
minutes after which 100 .mu.L of water is added to the wells of the
plate before centrifugation to pellet precipitated proteins.
Supernatants are transferred to another 96-well plate and analyzed
for amounts of parent remaining by LC-MS/MS using an Applied
Bio-systems API 4000 mass spectrometer. The same procedure is
followed for the non-deuterated counterpart of the compound of
Formula I or Formula A and the positive control, 7-ethoxycoumarin
(1 .mu.M). Testing is done in triplicate.
[0147] Data analysis: The in vitro t.sub.1/2s for test compounds
are calculated from the slopes of the linear regression of % parent
remaining (ln) vs incubation time relationship.
in vitro t.sub.1/2=0.693/k [0148] k=-[slope of linear regression of
% parent remaining(ln) vs incubation time]
[0149] Data analysis is performed using Microsoft Excel
Software.
[0150] Without further description, it is believed that one of
ordinary skill in the art can, using the preceding description and
the illustrative examples, make and utilize the compounds of the
present invention and practice the claimed methods. It should be
understood that the foregoing discussion and examples merely
present a detailed description of certain preferred embodiments. It
will be apparent to those of ordinary skill in the art that various
modifications and equivalents can be made without departing from
the spirit and scope of the invention.
* * * * *
References