U.S. patent application number 14/277957 was filed with the patent office on 2014-10-30 for pyruvate kinase m2 modulators, therapeutic compositions and related methods of use.
This patent application is currently assigned to AGIOS PHARMACEUTICALS, INC. The applicant listed for this patent is Francesco G. Salituro, Jeffrey O. Saunders. Invention is credited to Francesco G. Salituro, Jeffrey O. Saunders.
Application Number | 20140323729 14/277957 |
Document ID | / |
Family ID | 42224321 |
Filed Date | 2014-10-30 |
United States Patent
Application |
20140323729 |
Kind Code |
A1 |
Salituro; Francesco G. ; et
al. |
October 30, 2014 |
PYRUVATE KINASE M2 MODULATORS, THERAPEUTIC COMPOSITIONS AND RELATED
METHODS OF USE
Abstract
Compositions comprising compounds that modulate pyruvate kinase
M2 (PKM2) are described herein. Also described herein are methods
of using the compounds that modulate PKM2 in the treatment of
cancer.
Inventors: |
Salituro; Francesco G.;
(Marlborough, MA) ; Saunders; Jeffrey O.;
(Lincoln, MA) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
Salituro; Francesco G.
Saunders; Jeffrey O. |
Marlborough
Lincoln |
MA
MA |
US
US |
|
|
Assignee: |
AGIOS PHARMACEUTICALS, INC
Cambridge
MA
|
Family ID: |
42224321 |
Appl. No.: |
14/277957 |
Filed: |
May 15, 2014 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
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13267494 |
Oct 6, 2011 |
8742119 |
|
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14277957 |
|
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PCT/US2010/030139 |
Apr 6, 2010 |
|
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13267494 |
|
|
|
|
61233470 |
Aug 12, 2009 |
|
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61167017 |
Apr 6, 2009 |
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Current U.S.
Class: |
544/327 ;
544/331; 546/274.1; 546/276.1 |
Current CPC
Class: |
A61P 3/04 20180101; A61P
13/08 20180101; C07D 403/04 20130101; C07D 405/14 20130101; A61P
37/02 20180101; C07D 409/04 20130101; A61P 43/00 20180101; C07D
401/04 20130101; A61P 3/10 20180101; C07D 417/04 20130101; A61P
35/00 20180101 |
Class at
Publication: |
544/327 ;
546/274.1; 544/331; 546/276.1 |
International
Class: |
C07D 405/14 20060101
C07D405/14; C07D 403/04 20060101 C07D403/04; C07D 401/04 20060101
C07D401/04 |
Claims
1. A compound selected from formula (II): ##STR00240## wherein A,
B, D and E are each independently selected from H,
--SO.sub.2--NR.sup.4R.sup.5 and R.sup.3; wherein at least one of A,
B, D, or E is --SO.sub.2--NR.sup.4R.sup.5; Y.sup.1, Y.sup.2,
Y.sup.3 and Y.sup.4 are each independently selected from N and
CR.sup.1, wherein at least one of Y.sup.1, Y.sup.2, Y.sup.3 and
Y.sup.4 are N; each R.sup.4 is independently selected from
C.sub.1-8 alkyl, aryl and heteroaryl, each of which is substituted
with n occurrences of R.sup.2; each R.sup.5 is independently
hydrogen or C.sub.1-8 alkyl; each R.sup.1 is independently selected
from hydrogen, C.sub.1-8 alkyl, C.sub.1-8 terminal alkynyl,
C.sub.1-8 alkoxy, halogen, haloalkyl and haloalkoxy; each R.sup.2
is independently selected from halo, haloalkyl, C.sub.1-4 alkyl,
C.sub.2-4 alkenyl, C.sub.2-4 alknynyl, aryl, heteroaryl, aralkyl,
heteroaralkyl, cyano, --OR.sup.a, --COOR.sup.b and
--CONR.sup.cR.sup.c'; wherein two R.sup.2, together with the
carbons to which they are attached, may form an optionally
substituted ring, each of which can be further substituted; each
R.sup.3 is independently selected from C.sub.1-8 alkyl, --OR.sup.a,
halogen, haloalkyl, haloalkoxy and optionally substituted
heteroaryl; each R.sup.a is independently selected from alkyl,
haloalkyl, optionally substituted heteroaryl and optionally
substituted heterocyclyl; each R.sup.b is independently alkyl; and
each R.sup.c is independently selected from hydrogen and alkyl; and
n is 0, 1, 2 or 3.
2. A compound selected from formula (VI): ##STR00241## wherein A, B
and E are each independently selected from H,
--SO.sub.2--NR.sup.4R.sup.5 and R.sup.3; wherein at least one of A,
B or E is --SO.sub.2--NR.sup.4R.sup.5; Y.sup.1, Y.sup.2, Y.sup.3
and Y.sup.4 are each independently selected from N and CR.sup.1,
wherein at least one of Y.sup.1, Y.sup.2, Y.sup.3 and Y.sup.4 are
N; each R.sup.4 is independently selected from C.sub.1-8 alkyl,
aryl and heteroaryl, each of which is substituted with n
occurrences of R.sup.2; each R.sup.5 is independently hydrogen or
C.sub.1-8 alkyl; each R.sup.1 is independently selected from
hydrogen, C.sub.1-8 alkyl, C.sub.1-8 terminal alkynyl, C.sub.1-8
alkoxy, halogen, haloalkyl and haloalkoxy; each R.sup.2 is
independently selected from halo, haloalkyl, C.sub.1-4 alkyl,
C.sub.2-4 alkenyl, C.sub.2-4 alknynyl, aryl, heteroaryl, aralkyl,
heteroaralkyl, cyano, --OR.sup.a, --COOR.sup.b and
--CONR.sup.cR.sup.c'; wherein two R.sup.2, together with the
carbons to which they are attached, may form an optionally
substituted ring, each of which can be further substituted; each
R.sup.3 is independently selected from C.sub.1-8 alkyl, --OR.sup.a,
halogen, haloalkyl, haloalkoxy and optionally substituted
heteroaryl; each R.sup.a is independently selected from alkyl,
haloalkyl, optionally substituted heteroaryl and optionally
substituted heterocyclyl; each R.sup.b is independently alkyl; and
each R.sup.c is independently selected from hydrogen and alkyl; and
n is 0, 1, 2 or 3.
Description
CLAIM OF PRIORITY
[0001] This application is a divisional of U.S. Ser. No.
13/267,494, filed Oct. 6, 2011, which is a continuation of
International Application No. PCT/US2010/030139, filed Apr. 6,
2010, which claims priority from U.S. Ser. No. 61/233,470, filed
Aug. 12, 2009, and U.S. Ser. No. 61/167,017, filed Apr. 6, 2009,
each of which is incorporated herein by reference in its
entirety.
BACKGROUND OF INVENTION
[0002] Cancer cells rely primarily on glycolysis to generate
cellular energy and biochemical intermediates for biosynthesis of
lipids and nucleotides, while the majority of "normal" cells in
adult tissues utilize aerobic respiration. This fundamental
difference in cellular metabolism between cancer cells and normal
cells, termed the Warburg Effect, has been exploited for diagnostic
purposes, but has not yet been exploited for therapeutic
benefit.
[0003] Pyruvate kinase (PK) is a metabolic enzyme that converts
phosphoenolpyruvate to pyruvate during glycolysis. Four PK isoforms
exist in mammals: the L and R isoforms are expressed in liver and
red blood cells, the M1 isoform is expressed in most adult tissues,
and the M2 isoform is a splice variant of M1 expressed during
embryonic development. All tumor cells exclusively express the
embryonic M2 isoform. A well-known difference between the M1 and M2
isoforms of PK is that M2 is a low-activity enzyme that relies on
allosteric activation by the upstream glycolytic intermediate,
fructose-1,6-bisphosphate (FBP), whereas M1 is a constitutively
active enzyme.
[0004] All tumor cells exclusively express the embryonic M2 isoform
of pyruvate kinase, suggesting PKM2 as a potential target for
cancer therapy. PKM2 is also expressed in adipose tissue and
activated T-cells. Thus, the modulation (e.g., inhibition or
activation) of PKM2 may be effective in the treatment of, e.g.,
obesity, diabetes, autoimmune conditions, and
proliferation-dependent diseases, e.g., benign prostatic
hyperplasia (BPH). Current modulators (e.g., inhibitors) of
pyruvate kinase are not selective, making it difficult to treat
disease related to pyruvate kinase function.
[0005] Furthermore, phosphotyrosine peptide binding to PKM2 leads
to a dissociation of FBP from PKM2 and conformational changes of
PKM2 from an active, tetrameric form to an inactive form. Compounds
that bind to PKM2 and lock the enzyme in the active confirmation
will lead to the loss of allosteric control of PKM2 needed for
shunting biochemical intermediates from glycolysis into
biosynthesis of nucleotides and lipids. Thus, the activation of
PKM2 can also inhibit the growth and proliferation of cancer cells,
activated immune cells, and fat cells.
[0006] There is a continuing need for novel treatments of diseases
such as cancer, diabetes, obesity, autoimmune conditions,
proliferation-dependent diseases (e.g., BPH), and other diseases
related to the function of pyruvate kinase (e.g., PKM2).
SUMMARY OF INVENTION
[0007] Described herein are compounds that modulate pyruvate kinase
M2 (PKM2) and pharmaceutically acceptable salts, solvates, and
hydrates thereof, for example, compounds that modulate PKM2. This
invention also provides compositions and pharmaceutical kits
comprising a compound of this invention and the use of such
compositions and kits in methods of treating diseases and
conditions that are related to pyruvate kinase function (e.g., PKM2
function), including, e.g., cancer, diabetes, obesity, autoimmune
disorders, and benign prostatic hyperplasia (BPH).
[0008] In one aspect, the present invention features a compound or
pharmaceutically acceptable salt thereof of formula (I):
##STR00001##
wherein
[0009] X.sup.1 is N or CE;
[0010] X.sup.2 is N or CD;
[0011] X.sup.3 is N or CB;
[0012] X.sup.4 is N or CA;
[0013] Y.sup.1, Y.sup.2, Y.sup.3 and Y.sup.4 are each independently
selected from N and CR.sup.1;
[0014] A, B, D and E are each independently selected from H,
R.sup.3 and --SO.sub.2--NR.sup.4R.sup.5;
[0015] wherein at least one of X.sup.1, X.sup.2, X.sup.3, X.sup.4,
Y.sup.1, Y.sup.2, Y.sup.3 and Y.sup.4 is N; and at least one of
X.sup.1, X.sup.2, X.sup.3, X.sup.4, is
C--SO.sub.2--NR.sup.4R.sup.5;
[0016] each R.sup.4 is independently selected from C.sub.1-8 alkyl,
aryl and heteroaryl, each of which is substituted with n
occurrences of R.sup.2;
[0017] each R.sup.5 is independently hydrogen or C.sub.1-8
alkyl;
[0018] each R.sup.1 is independently selected from hydrogen,
C.sub.1-8 alkyl, C.sub.1-8 terminal alkynyl, C.sub.1-8 alkoxy,
halogen, haloalkyl and haloalkoxy;
[0019] each R.sup.2 is independently selected from halo, haloalkyl,
C.sub.1-4 alkyl, C.sub.2-4 alkenyl, C.sub.2-4 alknynyl, aryl,
heteroaryl, aralkyl, heteroaralkyl, cyano, --OR.sup.a, --COOR.sup.b
and --CONR.sup.cR.sup.c'; wherein two R.sup.2, together with the
carbons to which they are attached, may form an optionally
substituted ring, each of which can be further substituted;
[0020] each R.sup.3 is independently selected from C.sub.1-8 alkyl,
--OR.sup.a, halogen, haloalkyl, haloalkoxy and optionally
substituted heteroaryl;
[0021] each R.sup.a is independently selected from alkyl,
haloalkyl, optionally substituted heteroaryl and optionally
substituted heterocyclyl;
[0022] each R.sup.b is independently alkyl; and
[0023] each R.sup.c is independently selected from hydrogen and
alkyl; and
[0024] n is 0, 1, 2 or 3.
[0025] In some embodiments, one of X.sup.1, X.sup.2, X.sup.3,
X.sup.4, is C--SO.sub.2--NR.sup.4R.sup.5;
[0026] In some embodiments, X.sup.4 is A and A is
--SO.sub.2--NR.sup.4R.sup.5. In some embodiments, X.sup.3 is B and
B is --SO.sub.2--NR.sup.4R.sup.5. In some embodiments, X.sup.2 is D
and D is --SO.sub.2--NR.sup.4R.sup.5. In some embodiments, X.sup.1
is CE and E is --SO.sub.2--NR.sup.4R.sup.5.
[0027] In some embodiments, the compound is a compound of formula
(Ia):
##STR00002##
wherein A, B, D, R.sup.1, R.sup.2, R.sup.3, R.sup.4, R.sup.5 and n
are as defined in formula (I).
[0028] In some embodiments, A or B is
--SO.sub.2--NR.sup.4R.sup.5.
[0029] In some embodiments, the compound is a compound of formula
(Ib):
##STR00003##
wherein A, B, R.sup.1, R.sup.2, R.sup.3, R.sup.4, R.sup.5 and n are
as defined in formula (I).
[0030] In some embodiments, A or B is
--SO.sub.2--NR.sup.4R.sup.5.
[0031] In some embodiments, the compound is a compound of formula
(Ic):
##STR00004##
wherein A, D, R.sup.1, R.sup.2, R.sup.3, R.sup.4, R.sup.5 and n are
as defined in formula (I).
[0032] In some embodiments, A or D is
--SO.sub.2--NR.sup.4R.sup.5.
[0033] In some embodiments, the compound is a compound of formula
(Id):
##STR00005##
wherein A, D, E, R.sup.1, R.sup.2, R.sup.3, R.sup.4, R.sup.5 and n
are as defined in formula (I).
[0034] In some embodiments, A is --SO.sub.2--NR.sup.4R.sup.5.
[0035] In some embodiments, the compound is a compound of formula
(Ie):
##STR00006##
wherein A, B, E, R.sup.1, R.sup.2, R.sup.3, R.sup.4, R.sup.5 and n
are as defined in formula (I).
[0036] In some embodiments, A or B is
--SO.sub.2--NR.sup.4R.sup.5.
[0037] In some embodiments, the compound is a compound of formula
(If):
##STR00007##
wherein A, B, D, R.sup.1, R.sup.2, R.sup.3, R.sup.4, R.sup.5 and n
are as defined in formula (I).
[0038] In some embodiments, A or B is
--SO.sub.2--NR.sup.4R.sup.5.
[0039] In some embodiments, the compound is a compound of formula
(Ig):
##STR00008##
wherein A, B, R.sup.1, R.sup.2, R.sup.3, R.sup.4, R and n are as
defined in formula (I).
[0040] In some embodiments, A is --SO.sub.2--NR.sup.4R.sup.5.
[0041] In some embodiments, B is --SO.sub.2--NR.sup.4R.sup.5.
[0042] In some embodiments, the compound is a compound of formula
(Ih):
##STR00009##
wherein A, D, R.sup.1, R.sup.2, R.sup.3, R.sup.4, R.sup.5 and n are
as defined in formula (I).
[0043] In some embodiments, A is --SO.sub.2--NR.sup.4R.sup.5.
[0044] In some embodiments, the compound is a compound of formula
(Ij):
##STR00010##
wherein A, D, E, R.sup.1, R.sup.2, R.sup.3, R.sup.4, R.sup.5 and n
are as defined in formula (I).
[0045] In some embodiments, A is --SO.sub.2--NR.sup.4R.sup.5.
[0046] In some embodiments, the compound is a compound of formula
(Ik):
##STR00011##
wherein A, B, E, R.sup.1, R.sup.2, R.sup.3, R.sup.4, R.sup.5 and n
are as defined in formula (I).
[0047] In some embodiments, A or B is
--SO.sub.2--NR.sup.4R.sup.5.
[0048] In some embodiments, the compound is a compound of formula
(Im):
##STR00012##
wherein A, B, D, R.sup.1, R.sup.2, R.sup.3, R.sup.4, R.sup.5 and n
are as defined in formula (I).
[0049] In some embodiments, A or B is
--SO.sub.2--NR.sup.4R.sup.5.
[0050] In some embodiments, the compound is a compound of formula
(In):
##STR00013##
wherein A, B, R.sup.1, R.sup.2, R.sup.3, R.sup.4, R.sup.5 and n are
as defined in formula (I).
[0051] In some embodiments, A is --SO.sub.2--NR.sup.4R.sup.5.
[0052] In some embodiments, B is --SO.sub.2--NR.sup.4R.sup.5.
[0053] In some embodiments, the compound is a compound of formula
(Io):
##STR00014##
wherein A, D, R.sup.1, R.sup.2, R.sup.3, R.sup.4, R.sup.5 and n are
as defined in formula (I).
[0054] In some embodiments, A is --SO.sub.2--NR.sup.4R.sup.5.
[0055] In some embodiments, the compound is a compound of formula
(Ip):
##STR00015##
wherein A, D, E, R.sup.1, R.sup.2, R.sup.3, R.sup.4, R.sup.5 and n
are as defined in formula (I).
[0056] In some embodiments, A is --SO.sub.2--NR.sup.4R.sup.5.
[0057] In some embodiments, the compound is a compound of formula
(Iq):
##STR00016##
wherein A, B, E, R.sup.1, R.sup.2, R.sup.3, R.sup.4, R.sup.5 and n
are as defined in formula (I).
[0058] In some embodiments, A or B is
--SO.sub.2--NR.sup.4R.sup.5.
[0059] In some embodiments, the compound is a compound of formula
(Ir):
##STR00017##
wherein A, B, D, R.sup.1, R.sup.2, R.sup.3, R.sup.4, R.sup.5 and n
are as defined in formula (I).
[0060] In some embodiments, A or B is
--SO.sub.2--NR.sup.4R.sup.5.
[0061] In some embodiments, the compound is a compound of formula
(Is):
##STR00018##
wherein A, B, D, E, R.sup.1, R.sup.2, R.sup.3, R.sup.4, R.sup.5, m
and n are as defined in formula (I).
[0062] In some embodiments, A is --SO.sub.2--NR.sup.4R.sup.5.
[0063] In some embodiments, B is --SO.sub.2--NR.sup.4R.sup.5.
[0064] In some embodiments, the compound is a compound of formula
(It):
##STR00019##
wherein A, D, R.sup.1, R.sup.2, R.sup.3, R.sup.4, R.sup.5 and n are
as defined in formula (I).
[0065] In some embodiments, A is --SO.sub.2--NR.sup.4R.sup.5.
[0066] In some embodiments, the compound is a compound of formula
(Iu):
##STR00020##
wherein A, D, E, R.sup.1, R.sup.2, R.sup.3, R.sup.4, R.sup.5 and n
are as defined in formula (I).
[0067] In some embodiments, A is --SO.sub.2--NR.sup.4R.sup.5.
[0068] In some embodiments, the compound is a compound of formula
(Iv):
##STR00021##
wherein A, B, E, R.sup.1, R.sup.2, R.sup.3, R, R and n are as
defined in formula (I).
[0069] In some embodiments, A or B is
--SO.sub.2--NR.sup.4R.sup.5.
[0070] In some embodiments, the compound is a compound of formula
(Iw):
##STR00022##
wherein A, B, D, R.sup.1, R.sup.2, R.sup.3, R.sup.4, R.sup.5 and n
are as defined in formula (I).
[0071] In some embodiments, A or B is
--SO.sub.2--NR.sup.4R.sup.5.
[0072] In some embodiments, the compound is a compound of formula
(Ix):
##STR00023##
wherein A, B, R.sup.1, R.sup.2, R.sup.3, R.sup.4, R.sup.5 and n are
as defined in formula (I).
[0073] In some embodiments, A is --SO.sub.2--NR.sup.4R.sup.5.
[0074] In some embodiments, B is --SO.sub.2--NR.sup.4R.sup.5.
[0075] In some embodiments, the compound is a compound of formula
(Iy):
##STR00024##
wherein A, D, R, R.sup.2, R.sup.3, R.sup.4, R.sup.5 and n are as
defined in formula (I).
[0076] In some embodiments, A is --SO.sub.2--NR.sup.4R.sup.5.
[0077] In some embodiments, the compound is a compound of formula
(Iz):
##STR00025##
wherein A, D, E, R.sup.1, R.sup.2, R.sup.3, R.sup.4, R.sup.5 and n
are as defined in formula (I).
[0078] In some embodiments, A is --SO.sub.2--NR.sup.4R.sup.5.
[0079] In some embodiments, the compound is a compound of formula
(Iaa):
##STR00026##
wherein A, B, E, R.sup.1, R.sup.2, R.sup.3, R.sup.4, R.sup.5 and n
are as defined in formula (I).
[0080] In some embodiments, A is --SO.sub.2--NR.sup.4R.sup.5.
[0081] In some embodiments, B is --SO.sub.2--NR.sup.4R.sup.5.
[0082] In some embodiments, the compound is a compound of formula
(Ibb):
##STR00027##
wherein A, B, D, R.sup.1, R.sup.2, R.sup.3, R.sup.4, R.sup.5 and n
are as defined in formula (I).
[0083] In some embodiments, A is --SO.sub.2--NR.sup.4R.sup.5.
[0084] In some embodiments, B is --SO.sub.2--NR.sup.4R.sup.5.
[0085] In some embodiments, the compound is a compound of formula
(Icc)
##STR00028##
wherein A, B, R.sup.1, R.sup.2, R.sup.3, R.sup.4, R.sup.5 and n are
as defined in formula (I).
[0086] In some embodiments, A is --SO.sub.2--NR.sup.4R.sup.5.
[0087] In some embodiments, B is --SO.sub.2--NR.sup.4R.sup.5.
[0088] In some embodiments, the compound is a compound of formula
(Idd):
##STR00029##
wherein A, D, R.sup.1, R.sup.2, R.sup.3, R.sup.4, R.sup.5 and n are
as defined in formula (I).
[0089] In some embodiments, A is --SO.sub.2--NR.sup.4R.sup.5.
[0090] In some embodiments, the compound is a compound of formula
(lee):
##STR00030##
wherein A, D, E, R.sup.1, R.sup.2, R.sup.3, R.sup.4, R.sup.5 and n
are as defined in formula (I).
[0091] In some embodiments, A is --SO.sub.2--NR.sup.4R.sup.5.
[0092] In some embodiments, the compound is a compound of formula
(Iff):
##STR00031##
wherein A, B, E, R.sup.1, R.sup.2, R.sup.3, R.sup.4, R.sup.5 and n
are as defined in formula (I).
[0093] In some embodiments, A is --SO.sub.2--NR.sup.4R.sup.5.
[0094] In some embodiments, B is --SO.sub.2--NR.sup.4R.sup.5.
[0095] In one aspect, the invention features a pharmaceutical
composition comprising a compound of formula (I) as described
herein, or a pharmaceutically acceptable salt thereof.
[0096] In one aspect, the invention features a method of treating a
disorder described herein (e.g., cancer) comprising administering
to a subject a compound of formula (I) as described herein or a
pharmaceutically acceptable salt thereof.
[0097] In one aspect, the invention features a method of preventing
(e.g., preventing the onset of at least one symptom) or delaying
the onset of a disorder as described herein (e.g., cancer)
comprising administering to a subject a compound of formula (I) as
described herein or a pharmaceutically acceptable salt thereof.
[0098] In one aspect, the present invention features a compound or
pharmaceutically acceptable salt thereof of formula (II):
##STR00032##
wherein
[0099] A, B, D and E are each independently selected from H,
--SO.sub.2--NR.sup.4R.sup.5 and R.sup.3; wherein at least one of A,
B, D, or E is --SO.sub.2--NR.sup.4R.sup.5;
[0100] Y.sup.1, Y.sup.2, Y.sup.3 and Y.sup.4 are each independently
selected from N and CR.sup.1, wherein at least one of Y.sup.1,
Y.sup.2, Y.sup.3 and Y.sup.4 are N;
[0101] each R.sup.4 is independently selected from C.sub.1-8 alkyl,
aryl and heteroaryl, each of which is substituted with n
occurrences of R.sup.2;
[0102] each R.sup.5 is independently hydrogen or C.sub.1-8
alkyl;
[0103] each R.sup.1 is independently selected from hydrogen,
C.sub.1-8 alkyl, C.sub.1-8 terminal alkynyl, C.sub.1-8 alkoxy,
halogen, haloalkyl and haloalkoxy;
[0104] each R.sup.2 is independently selected from halo, haloalkyl,
C.sub.1-4 alkyl, C.sub.2-4 alkenyl, C.sub.2-4 alknynyl, aryl,
heteroaryl, aralkyl, heteroaralkyl, cyano, --OR.sup.a, --COOR.sup.b
and --CONR.sup.cR.sup.c'; wherein two R.sup.2, together with the
carbons to which they are attached, may form an optionally
substituted ring, each of which can be further substituted;
[0105] each R.sup.3 is independently selected from C.sub.1-8 alkyl,
--OR.sup.a, halogen, haloalkyl, haloalkoxy and optionally
substituted heteroaryl;
[0106] each R.sup.a is independently selected from alkyl,
haloalkyl, optionally substituted heteroaryl and optionally
substituted heterocyclyl;
[0107] each R.sup.b is independently alkyl; and
[0108] each R.sup.c is independently selected from hydrogen and
alkyl; and
[0109] n is 0, 1, 2 or 3.
[0110] In some embodiments, at least one of Y.sup.1, Y.sup.2,
Y.sup.3 and Y.sup.4 is N. In some embodiments, at least one of
Y.sup.1, Y.sup.2, Y.sup.3 and Y.sup.4 are CH. In some embodiments,
Y.sup.1 is N. In some embodiments, Y.sup.3 is N.
[0111] In some embodiments, each R.sup.1 is independently
hydrogen.
[0112] In some embodiments, the invention features a compound of
formula (IIa):
##STR00033##
wherein n, B, D, E, R.sup.1, R.sup.2, R.sup.3, R.sup.4 and R.sup.5
are defined as above.
[0113] In some embodiments, the invention features a compound of
formula (IIb):
##STR00034##
wherein n, B, D, E, R.sup.1, R.sup.2, R.sup.3, R.sup.4 and R.sup.5
are defined as above.
[0114] In some embodiments, B, D and E are each independently
selected from H.
[0115] In some embodiments, R.sup.5 is hydrogen.
[0116] In some embodiments, each R.sup.1 is independently hydrogen.
In some embodiments, each R.sup.1 is independently selected from
C.sub.1-8 alkyl, halogen or haloalkyl. In some embodiments, each
R.sup.1 is independently selected from halogen or haloalkyl. In
some embodiments, each R.sup.1 is independently selected from
halogen (e.g., chlorine or fluorine). In some embodiments, each
R.sup.1 is independently haloalkyl (e.g., trifluoroalkyl).
[0117] In some embodiments, R.sup.4 is selected from aryl or
heteroaryl. In some embodiments, R.sup.4 is aryl substituted with n
occurrences of R.sup.2. In some embodiments, R.sup.4 is C.sub.5-8
monocyclic aryl or C.sub.8-14 bicyclic aryl. In some embodiments,
R.sup.4 is C.sub.5-8 monocyclic aryl (e.g., optionally substituted
phenyl). In some embodiments, R.sup.4 is phenyl substituted with n
occurrences of R.sup.2.
[0118] In some embodiments, n is 0. In some embodiments, n is
1.
[0119] In some embodiments, R.sup.2 is halo, C.sub.1-4 alkyl or
haloalkyl, each of which can be further substituted.
[0120] In some embodiments, R.sup.2 is C.sub.1-4 alkyl (e.g.,
ethyl). In some embodiments, R.sup.2 is halo (e.g., chloro). In
some embodiments, R.sup.2 is haloalkyl (e.g., trifluoromethyl).
[0121] In some embodiments, n is 2. In some embodiments, both
R.sup.2 are C.sub.1-4 alkyl (e.g., methyl). In some embodiments, n
is 2. In some embodiments, both R.sup.2 are halo (e.g., fluoro or
chloro). In some embodiments, n is 2. In some embodiments, one
R.sup.2 is haloalkyl (e.g., trifluoroalkyl) and the other R.sup.2
is --OR.sup.a. In some embodiments, R.sup.a is alkyl (e.g., methyl
or ethyl). In some embodiments, n is 2. In some embodiments, one
R.sup.2 is halo (e.g., fluoro or chloro) and the other R.sup.2 is
C.sub.1-4 alkyl (e.g., methyl or ethyl).
[0122] In some embodiments, n is 2. In some embodiments, two
R.sup.2, together with the carbon atoms to which they are attached,
form a 5-membered heterocyclic ring. In some embodiments, two
R.sup.2, together with the phenyl ring to which they are attached,
form the following structure:
##STR00035##
[0123] In some embodiments, n is 3. In some embodiments, all
R.sup.2 are halo (e.g., fluoro or chloro).
[0124] In another aspect, the invention features a pharmaceutical
composition comprising a compound selected from Formula (II), (IIa)
or (IIb) as described herein or a pharmaceutically acceptable salt
thereof.
[0125] In another aspect, the invention features a method of
treating a disorder described herein (e.g., cancer) comprising
administering to a subject a compound of formula (II), (IIa) or
(IIb) as described herein or a pharmaceutically acceptable salt
thereof.
[0126] In another aspect, the invention features a method of
preventing (e.g., preventing the onset of at least one symptom) or
delaying the onset of a disorder as described herein (e.g., cancer)
comprising administering to a subject a compound of formula (II),
(IIa) or (IIb) as described herein or a pharmaceutically acceptable
salt thereof.
[0127] In one aspect, the present invention features a compound or
pharmaceutically acceptable salt thereof of formula (III):
##STR00036##
wherein
[0128] X.sup.1 is N or CE;
[0129] X.sup.2 is N or CD;
[0130] X.sup.3 is N or CB;
[0131] X.sup.4 is N or CA, wherein at least one of X.sup.1,
X.sup.2, X.sup.3 and X.sup.4 is N and at least one of X.sup.1,
X.sup.2, X.sup.3, X.sup.4, is C--SO.sub.2--NR.sup.4R.sup.5;
[0132] A, B, D and E are each independently selected from H,
R.sup.3 and --SO.sub.2--NR.sup.4R.sup.5;
[0133] Y.sup.2 is selected from N and CR.sup.1;
[0134] each R.sup.4 is independently selected from C.sub.1-8 alkyl,
aryl and heteroaryl, each of which is substituted with n
occurrences of R.sup.2;
[0135] R.sup.5 is hydrogen or C.sub.1-8 alkyl;
[0136] each R.sup.1 is independently selected from hydrogen,
C.sub.1-8 alkyl, C.sub.1-8 terminal alkynyl, C.sub.1-8 alkoxy,
halogen, haloalkyl and haloalkoxy;
[0137] each R.sup.2 is independently selected from halo, haloalkyl,
C.sub.1-4 alkyl, C.sub.2-4 alkenyl, C.sub.2-4 alkynyl, aryl,
heteroaryl, aralkyl, heteroaralkyl, cyano, --OR.sup.a, --COOR.sup.b
and --CONR.sup.cR.sup.c'; wherein two R.sup.2, together with the
carbons to which they are attached, may form an optionally
substituted ring, each of which can be further substituted;
[0138] each R.sup.3 is independently selected from C.sub.1-8 alkyl,
--OR.sup.a, halogen, haloalkyl, haloalkoxy and optionally
substituted heteroaryl;
[0139] each R.sup.a is independently selected from alkyl,
haloalkyl, optionally substituted heteroaryl and optionally
substituted heterocyclyl;
[0140] each R.sup.b is independently alkyl; and
[0141] each R.sup.c is independently selected from hydrogen and
alkyl; and
[0142] n is 0, 1, 2 or 3.
[0143] In some embodiments, the invention features a compound of
formula (IIIa):
##STR00037##
wherein n, B, D, E, R.sup.1, R.sup.4, R.sup.2 and R.sup.5 are
defined as in formula (III).
[0144] In some embodiments, the invention features a compound of
formula (IIIb):
##STR00038##
wherein n, B, E, R.sup.1, R.sup.4, R.sup.2 and R.sup.5 are defined
as in formula (III).
[0145] In some embodiments, the invention features a compound of
formula (IIIc):
##STR00039##
wherein n, B, D, R.sup.1, R.sup.4, R.sup.2 and R.sup.5 are defined
as in formula (III).
[0146] In some embodiments, B and E are each independently selected
from H.
[0147] In some embodiments, R.sup.5 is hydrogen.
[0148] In some embodiments, each R.sup.1 is independently hydrogen.
In some embodiments, each R.sup.1 is independently selected from
C.sub.1-8 alkyl, halogen or haloalkyl. In some embodiments, each
R.sup.1 is independently selected from halogen or haloalkyl. In
some embodiments, each R.sup.1 is independently selected from
halogen (e.g., chlorine or fluorine). In some embodiments, each
R.sup.1 is independently haloalkyl (e.g., trifluoroalkyl).
[0149] In some embodiments, R.sup.4 is selected from aryl or
heteroaryl. In some embodiments, R.sup.4 is aryl substituted with n
occurrences of R.sup.2. In some embodiments, R.sup.4 is C.sub.5-8
monocyclic aryl or C.sub.8-14 bicyclic aryl. In some embodiments,
R.sup.4 is C.sub.5-8 monocyclic aryl (e.g., optionally substituted
phenyl). In some embodiments, R.sup.4 is phenyl substituted with n
occurrences of R.sup.2.
[0150] In some embodiments, n is 0. In some embodiments, n is
1.
[0151] In some embodiments, R.sup.2 is halo, C.sub.1-4 alkyl or
haloalkyl, each of which can be further substituted.
[0152] In some embodiments, R.sup.2 is C.sub.1-4 alkyl (e.g.,
methyl or ethyl). In some embodiments, R.sup.2 is halo (e.g.,
chloro).
[0153] In some embodiments, n is 2. In some embodiments, both
R.sup.2 are C.sub.1-4 alkyl (e.g., methyl or ethyl). In some
embodiments, n is 2. In some embodiments, both R.sup.2 are halo
(e.g., fluoro or chloro). In some embodiments, n is 2. In some
embodiments, one R.sup.2 is haloalkyl (e.g., trifluoroalkyl) and
the other R.sup.2 is --OR.sup.a. In some embodiments, R.sup.a is
alkyl (e.g., methyl or ethyl). In some embodiments, n is 2. In some
embodiments, one R.sup.2 is C.sub.1-4 alkyl (e.g., methyl or ethyl)
and the other R.sup.2 is halo (e.g., fluoro or chloro).
[0154] In some embodiments, n is 2. In some embodiments, two
R.sup.2, together with the carbon atoms to which they are attached,
form a 5-membered heterocyclic ring. In some embodiments, two
R.sup.2, together with the phenyl ring to which they are attached,
form the following structure:
##STR00040##
[0155] In some embodiments, n is 3. In some embodiments, all
R.sup.2 are halo (e.g., fluoro or chloro).
[0156] In another aspect, the invention features a pharmaceutical
composition comprising a compound selected from formula (III),
(IIIa), (IIIb) or (IIIc) as described herein or a pharmaceutically
acceptable salt thereof.
[0157] In another aspect, the invention features a method of
treating a disorder described herein (e.g., cancer) comprising
administering to a subject a compound of formula (III), (IIIa),
(IIIb) or (IIIc) as described herein or a pharmaceutically
acceptable salt thereof.
[0158] In another aspect, the invention features a method of
preventing (e.g., preventing the onset of at least one symptom) or
delaying the onset of a disorder described here (e.g., cancer)
comprising administering to a subject a compound of formula (III),
(IIIa), (IIIb) or (IIIc) as described herein or a pharmaceutically
acceptable salt thereof.
[0159] In another aspect, the invention features a compound or
pharmaceutically acceptable salt thereof selected from the
following formula:
##STR00041##
wherein
[0160] n is 0, 1, 2 or 3;
[0161] X.sup.1 is N or CE;
[0162] X.sup.2 is N or CD;
[0163] X.sup.3 is N or CB;
[0164] X.sup.4 is N or CA, wherein at least one of X.sup.1,
X.sup.2, X.sup.3 and X.sup.4 is N; and and at least one of X.sup.1,
X.sup.2, X.sup.3, X.sup.4, is C--SO.sub.2--NR.sup.4R.sup.5;
[0165] A, B, D and E are each independently selected from H and
--SO.sub.2--NR.sup.4R.sup.5;
[0166] each R.sup.4 is independently selected from C.sub.1-8 alkyl,
aryl and heteroaryl, each of which is substituted with n
occurrences of R.sup.2;
[0167] each R.sup.5 is independently hydrogen or C.sub.1-8
alkyl;
[0168] each R.sup.1 is independently selected from hydrogen,
C.sub.1-8 alkyl, C.sub.1-8 alkoxy, halogen, haloalkyl and
haloalkoxy;
[0169] each R.sup.2 is independently selected from halo, haloalkyl,
C.sub.1-4alkyl, C.sub.2-4 alkenyl, C.sub.2-4 alkynylheteroaryl,
aryl, aralkyl, heteroaralkyl, cyano, --OR.sup.a, --COOR.sup.b and
--CONR.sup.cR.sup.c'; wherein two R.sup.2, together with the
carbons to which they are attached, may form an optionally
substituted ring, each of which can be further substituted;
[0170] each R.sup.3 is independently selected from C.sub.1-8 alkyl,
--OR.sup.a, halogen, haloalkyl, haloalkoxy or optionally
substituted heteroaryl;
[0171] R.sup.a is independently selected from alkyl, haloalkyl,
optionally substituted heteroaryl and optionally substituted
heterocyclyl;
[0172] each R.sup.b is independently alkyl; and
[0173] each R.sup.c is independently selected from hydrogen and
alkyl.
[0174] In some embodiments, at least one of X.sup.3 and X.sup.4 are
CH.
[0175] In some embodiments, at least one of A, B, D and E are H. In
some embodiments, at least one of A, B, D and E are
--SO.sub.2--NH--R.sup.4. In some embodiments, A is
--SO.sub.2--NH--R.sup.4. In some embodiments, B is
--SO.sub.2--NH--R.sup.4.
[0176] In some embodiments, the invention features a compound of
formula (IVa):
##STR00042##
wherein n, B, E, R.sup.1, R.sup.4, R.sup.3, R.sup.2 and R.sup.5 are
defined as above.
[0177] In some embodiments, the invention features a compound of
formula (IVb):
##STR00043##
wherein n, A, D, E, R.sup.1, R.sup.4, R.sup.3, R.sup.2, and R.sup.5
are defined as above.
[0178] In some embodiments, A, and D are H. In some embodiments, B
and E are H.
[0179] In some embodiments, R.sup.5 is hydrogen.
[0180] In some embodiments, each R.sup.1 is independently H. In
some embodiments, each R.sup.1 is independently selected from
C.sub.1-8 alkyl, halogen or haloalkyl. In some embodiments, each
R.sup.1 is independently selected from halogen or haloalkyl. In
some embodiments, each R.sup.1 is independently selected from
halogen (e.g., chlorine or fluorine). In some embodiments, each
R.sup.1 is independently haloalkyl (e.g., trifluoroalkyl).
[0181] In some embodiments, R.sup.4 is selected from aryl or
heteroaryl. In some embodiments, R.sup.4 is aryl substituted with n
occurrences of R.sup.2. In some embodiments, R.sup.4 is C.sub.5-8
monocyclic aryl or C.sub.8-14 bicyclic aryl. In some embodiments,
R.sup.4 is C.sub.5-8 monocyclic aryl (e.g., phenyl). In some
embodiments, R.sup.4 is phenyl substituted with n occurrences of
R.sup.2. In some embodiments, R.sup.4 is C.sub.8-14 bicyclic aryl
(e.g., napthyl). In some embodiments, R.sup.4 is a 5-8 membered
heteroaryl or a 8-14 membered heteroaryl. In some embodiments,
R.sup.4 is a 8-14 membered heteroaryl (e.g., 5-quinolyl or
6-quinolyl). In some embodiments, R.sup.4 is quinolyl (e.g.,
5-quinolyl or 6-quinolyl) substituted with n occurrences of
R.sup.2
[0182] In some embodiments, n is 0. In some embodiments, n is
1.
[0183] In some embodiments, R.sup.2 is selected from halo,
C.sub.1-4 alkyl, cyano, haloalkyl, --OR.sup.a or two R.sup.2, taken
together with the carbon atoms to which they are attached form an
optionally substituted ring, each of which can be further
substituted.
[0184] In some embodiments, R.sup.2 is halo (e.g., chloro or
fluoro). In some embodiments, R.sup.2 is C.sub.1-4 alkyl (e.g.,
methyl or ethyl). In some embodiments, R.sup.2 is cyano. In some
embodiments, R.sup.2 is haloalkyl (e.g., trifluoromethyl). In some
embodiments, R.sup.2 is --OR.sup.a. In some embodiments, R.sup.a is
alkyl (e.g., methyl).
[0185] In some embodiments, n is 2. In some embodiments, both
R.sup.2 are C.sub.1-4 alkyl (e.g., methyl). In some embodiments, n
is 2. In some embodiments, both R.sup.2 are halo (e.g., fluoro or
chloro). In some embodiments, n is 2. In some embodiments, one
R.sup.2 is C.sub.1-4 alkyl and the other is halo (e.g., methyl and
chloro or methyl and fluoro). In some embodiments, n is 2. In some
embodiments, both R.sup.2 are haloalkyl (e.g., trifluoromethyl). In
some embodiments, n is 2. In some embodiments, both R.sup.2 are
--OR.sup.a. In some embodiments, both R.sup.a are alkyl (e.g,
methyl). In some embodiments, n is 2. In some embodiments, one
R.sup.2 is haloalkyl (e.g., trifluoromethyl) and one is --OR.sup.a.
In some embodiments, R.sup.a is alkyl (e.g., methyl).
[0186] In some embodiments, n is 2. In some embodiments, two
R.sup.2, taken together with the carbon atoms to which they are
attached, form a 6-membered heterocyclic ring. In some embodiments,
two R.sup.2, taken together with the phenyl ring to which they are
attached, for the following structure:
##STR00044##
[0187] In some embodiments, n is 3. In some embodiments, three
R.sup.2 are halo (e.g., fluoro).
[0188] In another aspect, the invention features a pharmaceutical
composition comprising a compound selected from formula (IV), (IVa)
or (IVb) as described herein or a pharmaceutically acceptable salt
thereof.
[0189] In another aspect, the invention features a method of
treating a disorder described herein (e.g., cancer) comprising
administering to a subject a compound of formula (IV), (IVa) or
(IVb) as described herein or a pharmaceutically acceptable salt
thereof.
[0190] In another aspect, the invention features a method of
preventing (e.g., preventing the onset of at least one symptom) or
delaying the onset of a disorder described here (e.g., cancer)
comprising administering to a subject a compound of formula (IV),
(IVa) or (IVb) as described herein or a pharmaceutically acceptable
salt thereof.
[0191] In another aspect, the invention features a pharmaceutical
composition comprising a compound of formula (V):
##STR00045##
[0192] wherein
[0193] B, D and E are each independently selected from H and
R.sup.3;
[0194] each R.sup.1 is independently selected from hydrogen, halo
and haloalkyl;
[0195] R.sup.4 is hydrogen, C.sub.1-8 alkyl, and aryl, substituted
with n occurrences of R.sup.2;
[0196] each R.sup.2 is independently selected from halo, haloalkyl,
alkyl, aryl, heteroaryl, aralkyl, heteroaralkyl, cyano, --OR.sup.a,
--COOR.sup.b and --CONR.sup.cR.sup.c'; wherein two R.sup.2 together
with the carbons to which they are attached, may form an optionally
substituted ring, each of which can be further substituted;
[0197] each R.sup.3 is independently selected from halo, haloalkyl
and --OR.sup.a;
[0198] R.sup.5 is hydrogen or C.sub.1-8 alkyl;
[0199] R.sup.a is independently selected from alkyl, haloalkyl,
optionally substituted heteroaryl and optionally substituted
heterocyclyl;
[0200] each R.sup.b is independently alkyl;
[0201] each R.sup.c is independently selected from hydrogen and
alkyl; and
[0202] n is 0, 1, 2, or 3.
[0203] In some embodiments, B, D and E are each independently
H.
[0204] In some embodiments, each R.sup.1 is independently H. In
some embodiments, each R.sup.1 is independently halo (e.g.,
chloro). In some embodiments, each R.sup.1 is independently
haloalkyl (e.g., trifluoromethyl).
[0205] In some embodiments, one R.sup.1 is halo and one R.sup.1 is
haloalkyl. In some embodiments, one R.sup.1 is chloro and one
R.sup.1 is trifluoromethyl.
[0206] In some embodiments, R.sup.5 is hydrogen.
[0207] In some embodiments, R.sup.5 is C.sub.1-8 alkyl (e.g.,
methyl).
[0208] In some embodiments, R.sup.4 is hydrogen.
[0209] In some embodiments, R.sup.4 is C.sub.1-8 alkyl or aryl
substituted with n occurrences of R.sup.2. In some embodiments,
R.sup.4 is C.sub.1-8 alkyl (e.g., methyl or ethyl) substituted with
n occurrences of R.sup.2.
[0210] In some embodiments, each R.sup.2 is independently selected
from halo, haloalkyl, alkyl, aryl, heteroaryl, heteroaralkyl,
cyano, --OR.sup.a, --COOR.sup.b and --CONR.sup.cR.sup.c'; wherein
two R.sup.2, together with the carbons to which they are attached,
may form an optionally substituted ring, each of which can be
further substituted.
[0211] In some embodiments, n is 0. In some embodiments, n is 1. In
some embodiments, R.sup.2 is --OR.sup.a. In some embodiments,
R.sup.a is alkyl (e.g., methyl). In some embodiments, R.sup.2 is
optionally substituted heteroaryl. In some embodiments, R.sup.2 is
optionally substituted monocyclic heteroaryl (e.g., 3-pyridyl). In
some embodiments, R.sup.2 is optionally substituted aryl. In some
embodiments, R.sup.2 is optionally substituted monocyclic aryl
(e.g., 4-chlorophenyl).
[0212] In some embodiments, R.sup.4 is aryl (e.g., phenyl)
substituted with n occurrences of R.sup.2. In some embodiments,
R.sup.4 is phenyl substituted with n occurrences of R.sup.2. In
some embodiments, n is 0.
[0213] In some embodiments, n is 1. In some embodiments, R.sup.2 is
halo (e.g., fluoro or chloro). In some embodiments, R.sup.2 is
haloalkyl (e.g., trifluoromethyl). In some embodiments, R.sup.2 is
alkyl (e.g., methyl or ethyl). In some embodiments, R.sup.2 is
heteoaralkyl. In some embodiments, R.sup.2 is optionally
substituted monocyclic heteroaralkyl (e.g.,
methyl-4-trifluoromethyl-2-pyridyl). In some embodiments, R.sup.2
is cyano. In some embodiments, R.sup.2 is --OR.sup.a. In some
embodiments, R.sup.a is alkyl (e.g., methyl). In some embodiments,
R.sup.2 is --COOR.sup.b. In some embodiments, R.sup.b is alkyl
(e.g., ethyl). In some embodiments, R.sup.2 is optionally
substituted monocyclic heteroaryl. In some embodiments, R.sup.2 is
optionally substituted pyridyl. In some embodiments, R.sup.2 is
pyridyl substituted with haloalkyl (e.g., trifluoromethyl).
[0214] In some embodiments, n is 2. In some embodiments, both
R.sup.2 are halo (e.g., fluoro or chloro). In some embodiments, n
is 2. In some embodiments, both R.sup.2 are alkyl (e.g., methyl).
In some embodiments, n is 2. In some embodiments, one R.sup.2 is
halo (e.g., fluoro or chloro) and one is alkyl (e.g., methyl). In
some embodiments, n is 2. In some embodiments, one R.sup.2 is halo
and one is --CONR.sup.cR.sup.c'. In some embodiments, n is 2. In
some embodiments, one R.sup.2 is chloro and one is
--CONR.sup.cR.sup.c'. In some embodiments, R.sup.c is alkyl (e.g.,
methyl or isopropyl). In some embodiments, n is 2. In some
embodiments, one R.sup.2 is alkyl and one is --CONHR.sup.c'. In
some embodiments, n is 2. In some embodiments, one R.sup.2 is
methyl and one is --CONHR.sup.c'. In some embodiments, R.sup.c is
alkyl (e.g., methyl or isopropyl).
[0215] In some embodiments, n is 2. In some embodiments, one
R.sup.2 is haloalkyl (e.g., trifluoromethyl) and the other is
--OR.sup.a. In some embodiments, R.sup.a is alkyl (e.g., methyl).
In some embodiments, n is 2. In some embodiments, one R.sup.2 is
halo and the other is --OR.sup.a. In some embodiments, n is 2. In
some embodiments, one R.sup.2 is chloro and the other is
--OR.sup.a. In some embodiments, R.sup.a is optionally substituted
heteroaryl. In some embodiments, n is 2. In some embodiments, both
R.sup.2 are --OR.sup.a. In some embodiments, R.sup.a is alkyl
(e.g., methyl). In some embodiments, R.sup.a is optionally
substituted pyridyl. In some embodiments, R.sup.a is pyridyl
substituted with haloalkyl (e.g., trifluoromethyl). In some
embodiments, R.sup.a is optionally substituted heterocyclyl. In
some embodiments, R.sup.a is an optionally substituted 5-membered
heterocyclyl. In some embodiments, R.sup.a is optionally
substituted pyrrolidinyl. In some embodiments, R.sup.a is N-methyl
pyrrolidinyl. In some embodiments, R.sup.a is:
##STR00046##
[0216] In some embodiments, n is 2. In some embodiments, two
R.sup.2, together with the carbon atoms to which they are attached,
form a 5-membered heterocyclic ring. In some embodiments, two
R.sup.2, together with the phenyl ring to which they are attached,
form the following structure:
##STR00047##
[0217] In some embodiments, n is 3. In some embodiments, each
R.sup.2 is halo (e.g., In some embodiments, n is 3. In some
embodiments, each R.sup.2 is halo (e.g., fluoro). In some
embodiments, n is 3. In some embodiments, two R.sup.2 are halo and
one R.sup.2 is --CONR.sup.cR.sup.c'. In some embodiments, two
R.sup.2 are chloro and one R.sup.2 is --CONHR.sup.c'. In some
embodiments, R.sup.c' is alkyl (e.g., methyl or isopropyl). In some
embodiments, one R.sup.2 is chloro, one R.sup.2 is bromo, and one
R.sup.2 is --CONHR.sup.c'. In some embodiments, R.sup.c' is alkyl
(e.g., methyl or isopropyl). In some embodiments, n is 3. In some
embodiments, one R.sup.2 is halo, one R.sup.2 is alkyl, and one
R.sup.2 is --CONR.sup.cR.sup.c'. In some embodiments, one R.sup.2
is chloro, one R.sup.2 is methyl, and one R.sup.2 is
--CONHR.sup.c'. In some embodiments, R.sup.c' is alkyl (e.g.,
methyl or isopropyl). In some embodiments, one R.sup.2 is bromo,
one R.sup.2 is methyl, and one R.sup.2 is --CONHR.sup.c'. In some
embodiments, R.sup.c' is alkyl (e.g., methyl or isopropyl).
[0218] In some embodiments, R.sup.3 is halo (e.g., chloro or
bromo). In some embodiments, R.sup.3 is haloalkyl (e.g.,
trifluoromethyl). In some embodiments, R.sup.3 is --OR.sup.a. In
some embodiments, R.sup.a is haloalkyl (e.g., difluoromethoxy,
trifluoromethoxy or trifluoroethoxy). In some embodiments, R.sup.a
is --CH.sub.2CF.sub.3.
[0219] In another aspect, the invention features a pharmaceutical
composition comprising a compound of formula (Va):
##STR00048##
[0220] wherein
[0221] B and D are each independently selected from H and
SO.sub.2NR.sup.4R.sup.5; wherein at least one of B or D is
--SO.sub.2--NR.sup.4R.sup.5;
[0222] A and E are each independently selected from H and
R.sup.3;
[0223] each R.sup.1 is independently selected from hydrogen, halo
and haloalkyl;
[0224] R.sup.4 is hydrogen, C.sub.1-8 alkyl, and aryl, substituted
with n occurrences of R.sup.2;
[0225] each R.sup.2 is independently selected from halo, haloalkyl,
alkyl, aryl, heteroaryl, aralkyl, heteroaralkyl, cyano, --OR.sup.a,
--COOR.sup.b and --CONR.sup.cR.sup.c'; wherein two R.sup.2,
together with the carbons to which they are attached, may form an
optionally substituted ring, each of which can be further
substituted;
[0226] each R.sup.3 is independently selected from halo, haloalkyl
and --OR.sup.a;
[0227] R.sup.5 is hydrogen or C.sub.1-8 alkyl;
[0228] R.sup.a is independently selected from alkyl, haloalkyl,
optionally substituted heteroaryl and optionally substituted
heterocyclyl;
[0229] each R.sup.b is independently alkyl;
[0230] each R.sup.c is independently selected from hydrogen and
alkyl; and
[0231] n is 0, 1, 2, or 3.
[0232] In another aspect, the invention features a method of
treating a disorder described herein (e.g., cancer) comprising
administering to a subject a compound of formula (V) or (Va) as
described herein or a pharmaceutically acceptable salt thereof.
[0233] In another aspect, the invention features a method of
preventing (e.g., preventing the onset of at least one symptom) or
delaying the onset of a disorder described here (e.g., cancer)
comprising administering to a subject a compound of formula (V) or
(Va) as described herein or a pharmaceutically acceptable salt
thereof.
[0234] In some embodiments, A and E are each H.
[0235] In some embodiments, B is SO.sub.2NR.sup.4R.sup.5 and D is
H. In some embodiments, B is H and D is
SO.sub.2NR.sup.4R.sup.5.
[0236] In one aspect, the present invention features a compound or
pharmaceutically acceptable salt thereof of formula (VI):
##STR00049##
wherein
[0237] A, B and E are each independently selected from H,
--SO.sub.2--NR.sup.4R.sup.5 and R.sup.3; wherein at least one of A,
B or E is --SO.sub.2--NR.sup.4R.sup.5;
[0238] Y.sup.1, Y.sup.2, Y.sup.3 and Y.sup.4 are each independently
selected from N and CR.sup.1, wherein at least one of Y.sup.1,
Y.sup.2, Y.sup.3 and Y.sup.4 are N;
[0239] each R.sup.4 is independently selected from C.sub.1-8 alkyl,
aryl and heteroaryl, each of which is substituted with n
occurrences of R.sup.2;
[0240] each R.sup.5 is independently hydrogen or C.sub.1-8
alkyl;
[0241] each R.sup.1 is independently selected from hydrogen,
C.sub.1-8 alkyl, C.sub.1-8 terminal alkynyl, C.sub.1-8 alkoxy,
halogen, haloalkyl and haloalkoxy;
[0242] each R.sup.2 is independently selected from halo, haloalkyl,
C.sub.1-4 alkyl, C.sub.2-4 alkenyl, C.sub.2-4 alknynyl, aryl,
heteroaryl, aralkyl, heteroaralkyl, cyano, --OR.sup.a, --COOR.sup.b
and --CONR.sup.cR.sup.c'; wherein two R.sup.2, together with the
carbons to which they are attached, may form an optionally
substituted ring, each of which can be further substituted;
[0243] each R.sup.3 is independently selected from C.sub.1-8 alkyl,
--OR.sup.a, halogen, haloalkyl, haloalkoxy and optionally
substituted heteroaryl;
[0244] each R.sup.a is independently selected from alkyl,
haloalkyl, optionally substituted heteroaryl and optionally
substituted heterocyclyl;
[0245] each R.sup.b is independently alkyl; and
[0246] each R.sup.c is independently selected from hydrogen and
alkyl; and
[0247] n is 0, 1, 2 or 3.
[0248] In some embodiments, at least one of Y.sup.1, Y.sup.2,
Y.sup.3 and Y.sup.4 is N. In some embodiments, at least one of
Y.sup.1, Y.sup.2, Y.sup.3 and Y.sup.4 are CH. In some embodiments,
Y.sup.1 is N. In some embodiments, Y.sup.3 is N.
[0249] In some embodiments, each R.sup.1 is independently
hydrogen.
[0250] In some embodiments, the invention features a compound of
formula (VIa):
##STR00050##
wherein n, B, E, R.sup.1, R.sup.2, R.sup.3, R.sup.4 and R.sup.5 are
defined as above.
[0251] In some embodiments, the invention features a compound of
formula (VIb):
##STR00051##
wherein n, B, E, R.sup.1, R.sup.2, R.sup.3, R.sup.4 and R.sup.5 are
defined as above.
[0252] In some embodiments, B and E are each independently selected
from H.
[0253] In some embodiments, R.sup.5 is hydrogen.
[0254] In some embodiments, each R.sup.1 is independently hydrogen.
In some embodiments, each R.sup.1 is independently selected from
C.sub.1-8 alkyl, halogen or haloalkyl. In some embodiments, each
R.sup.1 is independently selected from halogen or haloalkyl. In
some embodiments, each R.sup.1 is independently selected from
halogen (e.g., chlorine or fluorine). In some embodiments, each
R.sup.1 is independently haloalkyl (e.g., trifluoroalkyl).
[0255] In some embodiments, R.sup.4 is selected from aryl or
heteroaryl. In some embodiments, R.sup.4 is aryl substituted with n
occurrences of R.sup.2. In some embodiments, R.sup.4 is C.sub.5-8
monocyclic aryl or C.sub.8-14 bicyclic aryl. In some embodiments,
R.sup.4 is C.sub.5-8 monocyclic aryl (e.g., optionally substituted
phenyl). In some embodiments, R.sup.4 is phenyl substituted with n
occurrences of R.sup.2. In some embodiments, R.sup.2 is heteroaryl
substituted with n occurrences of R.sup.2. In some embodiments,
R.sup.4 is a 5-8 membered heteroaryl or 8-14 membered heteroaryl.
In some embodiments, R.sup.4 is an 8-12 membered heteroaryl (e.g.,
5-quinolyl or 6-quinolyl). In some embodiments, R.sup.4 is quinolyl
(e.g., 5-quinolyl or 6-quinolyl) substituted with n occurrences of
R.sup.2.
[0256] In some embodiments, n is 0. In some embodiments, n is
1.
[0257] In some embodiments, R.sup.2 is halo, C.sub.1-4 alkyl or
haloalkyl, each of which can be further substituted.
[0258] In some embodiments, R.sup.2 is C.sub.1-4 alkyl (e.g.,
ethyl). In some embodiments, R.sup.2 is halo (e.g., fluoro or
chloro). In some embodiments, R.sup.2 is haloalkyl (e.g.,
trifluoromethyl).
[0259] In some embodiments, n is 2. In some embodiments, both
R.sup.2 are C.sub.1-4 alkyl (e.g., methyl). In some embodiments, n
is 2. In some embodiments, both R.sup.2 are halo (e.g., fluoro or
chloro). In some embodiments, n is 2. In some embodiments, one
R.sup.2 is haloalkyl (e.g., trifluoroalkyl) and the other R.sup.2
is --OR.sup.a. In some embodiments, R.sup.a is alkyl (e.g., methyl
or ethyl). In some embodiments, n is 2. In some embodiments, on
R.sup.2 is halo (e.g., fluoro or chloro) and the other R.sup.2 is
C.sub.1-4 alkyl (e.g., methyl or ethyl).
[0260] In some embodiments, n is 2. In some embodiments, two
R.sup.2, together with the carbon atoms to which they are attached,
form a 5-membered heterocyclic ring. In some embodiments, two
R.sup.2, together with the phenyl ring to which they are attached,
form the following structure:
##STR00052##
[0261] In some embodiments, n is 3. In some embodiments, all
R.sup.2 are halo (e.g., fluoro or chloro).
[0262] In another aspect, the invention features a pharmaceutical
composition comprising a compound selected from Formula (VI), (VIa)
or (VIb) as described herein or a pharmaceutically acceptable salt
thereof.
[0263] In another aspect, the invention features a method of
treating a disorder described herein (e.g., cancer) comprising
administering to a subject a compound of formula (VI), (VIa) or
(VIb) as described herein or a pharmaceutically acceptable salt
thereof.
[0264] In another aspect, the invention features a method of
preventing (e.g., preventing the onset of at least one symptom) or
delaying the onset of a disorder as described herein (e.g., cancer)
comprising administering to a subject a compound of formula (VI),
(VIa) or (VIb) as described herein or a pharmaceutically acceptable
salt thereof.
[0265] In one aspect, the present invention features a compound or
pharmaceutically acceptable salt thereof of formula (VII):
##STR00053##
wherein
[0266] X.sup.1 is N or CE;
[0267] X.sup.2 is N or CD;
[0268] X.sup.3 is N or CB;
[0269] X.sup.4 is N or CA, wherein at least one of X.sup.1,
X.sup.2, X.sup.3 and X.sup.4 is N and at least one of X.sup.1,
X.sup.2, X.sup.3, X.sup.4, is C--SO.sub.2--NR.sup.4R.sup.5;
[0270] A, B, D and E are each independently selected from H,
R.sup.3 and --SO.sub.2--NR.sup.4R.sup.5;
[0271] Y.sup.1 is selected from N and CR.sup.1;
[0272] each R.sup.4 is independently selected from C.sub.1-8 alkyl,
aryl and heteroaryl, each of which is substituted with n
occurrences of R.sup.2;
[0273] R.sup.5 is hydrogen or C.sub.1-8 alkyl;
[0274] each R.sup.1 is independently selected from hydrogen,
C.sub.1-8 alkyl, C.sub.1-8 terminal alkynyl, C.sub.1-8 alkoxy,
halogen, haloalkyl and haloalkoxy;
[0275] each R.sup.2 is independently selected from halo, haloalkyl,
C.sub.1-4 alkyl, C.sub.2-4 alkenyl, C.sub.2-4 alkynyl, aryl,
heteroaryl, aralkyl, heteroaralkyl, cyano, --OR.sup.a, --COOR.sup.b
and --CONR.sup.cR.sup.c'; wherein two R.sup.2, together with the
carbons to which they are attached, may form an optionally
substituted ring, each of which can be further substituted;
[0276] each R.sup.3 is independently selected from C.sub.1-8 alkyl,
--OR.sup.a, halogen, haloalkyl, haloalkoxy and optionally
substituted heteroaryl;
[0277] each R.sup.a is independently selected from alkyl,
haloalkyl, optionally substituted heteroaryl and optionally
substituted heterocyclyl;
[0278] each R.sup.b is independently alkyl; and
[0279] each R.sup.c is independently selected from hydrogen and
alkyl; and
[0280] n is 0, 1, 2 or 3.
[0281] In some embodiments, the invention features a compound of
formula (VIIa):
##STR00054##
wherein n, B, D, R.sup.1, R.sup.4, R.sup.2 and R.sup.5 are defined
as in formula (VII).
[0282] In some embodiments, the invention features a compound of
formula (VIIa):
##STR00055##
wherein n, B, E, R.sup.1, R.sup.4, R.sup.2 and R.sup.5 are defined
as in formula (VII).
[0283] In some embodiments, the invention features a compound of
formula (VIIb):
##STR00056##
wherein n, B, D, R.sup.1, R.sup.4, R.sup.2 and R.sup.5 are defined
as in formula (VII).
[0284] In some embodiments, the invention features a compound of
formula (VIIa):
##STR00057##
wherein n, B, E, R.sup.1, R.sup.4, R.sup.2 and R.sup.5 are defined
as in formula (VII).
[0285] In some embodiments, B and E are each independently selected
from H.
[0286] In some embodiments, B and D are each independently selected
from H.
[0287] In some embodiments, R.sup.5 is hydrogen.
[0288] In some embodiments, each R.sup.1 is independently hydrogen.
In some embodiments, each R.sup.1 is independently selected from
C.sub.1-8 alkyl, halogen or haloalkyl. In some embodiments, each
R.sup.1 is independently selected from halogen or haloalkyl. In
some embodiments, each R.sup.1 is independently selected from
halogen (e.g., chlorine or fluorine). In some embodiments, each
R.sup.1 is independently haloalkyl (e.g., trifluoroalkyl).
[0289] In some embodiments, R.sup.4 is selected from aryl or
heteroaryl. In some embodiments, R.sup.4 is aryl substituted with n
occurrences of R.sup.2. In some embodiments, R.sup.4 is C.sub.5-8
monocyclic aryl or C.sub.8-14 bicyclic aryl. In some embodiments,
R.sup.4 is C.sub.5-8 monocyclic aryl (e.g., optionally substituted
phenyl). In some embodiments, R.sup.4 is phenyl substituted with n
occurrences of R.sup.2.
[0290] In some embodiments, n is 0. In some embodiments, n is
1.
[0291] In some embodiments, R.sup.2 is halo, C.sub.1-4 alkyl or
haloalkyl, each of which can be further substituted.
[0292] In some embodiments, R.sup.2 is C.sub.1-4 alkyl (e.g.,
methyl or ethyl). In some embodiments, R.sup.2 is halo (e.g.,
fluoro or chloro).
[0293] In some embodiments, n is 2. In some embodiments, both
R.sup.2 are C.sub.1-4 alkyl (e.g., methyl or ethyl). In some
embodiments, n is 2. In some embodiments, both R.sup.2 are halo
(e.g., fluoro or chloro). In some embodiments, n is 2. In some
embodiments, one R.sup.2 is haloalkyl (e.g., trifluoroalkyl) and
the other R.sup.2 is --OR.sup.a. In some embodiments, R.sup.a is
alkyl (e.g., methyl or ethyl). In some embodiments, n is 2. In some
embodiments, one R.sup.2 is C.sub.1-4 alkyl (e.g., methyl or ethyl)
and the other R.sup.2 is halo (e.g., fluoro or chloro).
[0294] In some embodiments, n is 2. In some embodiments, two
R.sup.2, together with the carbon atoms to which they are attached,
form a 5-membered heterocyclic ring. In some embodiments, two
R.sup.2, together with the phenyl ring to which they are attached,
form the following structure:
##STR00058##
[0295] In some embodiments, n is 3. In some embodiments, all
R.sup.2 are halo (e.g., fluoro or chloro).
[0296] In another aspect, the invention features a pharmaceutical
composition comprising a compound selected from formula (VII),
(VIIa), (VIIb), (VIIc) or (VIId) as described herein or a
pharmaceutically acceptable salt thereof.
[0297] In another aspect, the invention features a method of
treating a disorder described herein (e.g., cancer) comprising
administering to a subject a compound of formula (VII), (VIIa),
(VIIb), (VIIc) or (VIId) as described herein or a pharmaceutically
acceptable salt thereof.
[0298] In another aspect, the invention features a method of
preventing (e.g., preventing the onset of at least one symptom) or
delaying the onset of a disorder described here (e.g., cancer)
comprising administering to a subject a compound of formula (VII),
(VIIa), (VIIb), (VIIc) or (VIId) as described herein or a
pharmaceutically acceptable salt thereof.
[0299] In one aspect, the invention features a method of modulating
(e.g., increasing or decreasing) the level of PKM2 activity and/or
glycolysis (e.g., modulating the endogenous ability of a cell in
the patient to down regulate PKM2) in a patient in need thereof.
The method comprises the step of administering an effective amount
of a compound described herein to the patient in need thereof,
thereby modulating (e.g., increasing or decreasing) the level of
PKM2 activity and/or glycolysis in the patient. In some embodiments
of the invention an activator is used to maintain PKM2 in its
active conformation or activate pyruvate kinase activity in
proliferating cells as a means to divert glucose metabolites into
catabolic rather than anabolic processes in the patient.
[0300] In another aspect, the invention features a method of
regulating cell proliferation in a patient in need thereof. The
method comprises the step of administering an effective amount of a
compound described herein to the patient in need thereof, thereby
regulating cell proliferation in the patient. E.g., this method can
modulate growth of a transformed cell, e.g., a cancer cell, or
generally modulate growth in a PKM2-dependent cell that undergoes
aerobic glycolysis.
[0301] In another aspect, the invention features a method of
treating a patient suffering from or susceptible to a disease or
disorder associated with the function of PKM2 in a patient in need
thereof. The method comprises the step of administering an
effective amount of a compound described herein to the patient in
need thereof, thereby treating, preventing or ameliorating the
disease or disorder in the patient. In another embodiment the
modulator is provided in a pharmaceutical composition.
[0302] In another embodiment the method includes identifying or
selecting a patient who would benefit from modulation (e.g.,
activation or inhibition) of PKM2. E.g., the patient can be
identified on the basis of the level of PKM2 activity in a cell of
the patient (e.g., as opposed to merely being in need of treatment
of the disorder itself, e.g., cancer). In another embodiment the
selected patient is a patient suffering from or susceptible to a
disorder or disease identified herein, e.g., a disorder
characterized by unwanted cell growth or proliferation, e.g.,
cancer, obesity, diabetes, atherosclerosis, restenosis, and
autoimmune diseases.
[0303] In one aspect, the invention features a method of evaluating
a candidate compound the method comprising:
[0304] optionally supplying the candidate compound;
[0305] contacting the compound with a cell, e.g., a cell having an
intact plasma membrane;
[0306] evaluating the ability of the compound to interact
intracellularly with, e.g., to form a complex with, to bind, e.g.,
specifically to, or to modulate (e.g., activate or inhibit) the
activity of, a target kinase, e.g., pyruvate kinase, e.g.,
PKM2;
[0307] thereby evaluating the candidate compound.
[0308] In some embodiments, evaluating the candidate compound
comprises evaluating the candidate compound for use as an
anti-proliferative or anti-cancer agent and the ability of the
candidate compound to interact intracellulary with the target
compound is correlated to efficacy as an anti-proliferative or
anti-cancer agent.
[0309] In some embodiments, evaluating the candidate compound
comprises evaluating the ability of the candidate compound cross
the cell membrane and the ability of the candidate compound to
interact intracellulary with the target compound is correlated to
with ability of the candidate compound to cross the cell
membrane.
[0310] In some embodiments, evaluating the candidate compound
comprises evaluating the ability of the candidate compound to
modulate any of the following properties: a conformational state in
the target kinase, binding of the target kinase to an endogenous
modulator of target kinase activity, e.g., FBP or a phosphotyrsine
containing polypeptide, or other property of a target kinase
disclosed herein, and the ability of the candidate compound to
interact with the target compound is correlated with one or more of
said properties.
[0311] In some embodiments, the method further comprises separating
the cell from candidate compound that has not entered the cell,
e.g., by washing the cell or removing the cell from an animal to
which the candidate compound has been administered.
[0312] In some embodiments, the method further comprises lysing
(e.g., by disrupting or dissolving the cell membrane) the cell,
e.g., prior to evaluating the ability of the candidate compound to
interact with the target kinase.
[0313] In some embodiments, contacting the candidate compound with
a cell comprises contacting the candidate compound with a whole
animal, a tissue that is not part of a whole animal, an organ which
is not part of a whole animal or a cell which is not part of a
whole animal.
[0314] In some embodiments, the cell is a cultured cell, e.g., a
primary cell, a secondary cell.
[0315] In some embodiments, the cell is a mammal, primate, human,
rodent, mouse, rat, or hamster cell.
[0316] In some embodiments, the cell is a tumor or transformed
cell, e.g., a solid tumor cell.
[0317] In some embodiments, a plurality of target compounds are
evaluated, e.g., at least 10, 20, 50, 100, or 500 candidate
compounds are evaluated.
[0318] In some embodiments, a plurality of candidate compounds are
evaluated simultaneously, e.g., wherein each of a plurality of
candidate compounds is evaluated individually but simultaneously,
or wherein a plurality of candidate compounds are pooled and
contacted with the same cell or same aliquot of cells.
[0319] In some embodiments, a plurality of candidate compounds are
evaluated in an automated device.
[0320] In some embodiments, evaluating a candidate compound
comprises providing a value for the ability of the candidate
compound to interact with the target kinase and, optionally,
comparing that value to a predetermined value, e.g., a value for a
positive and or negative control.
[0321] In some embodiments, the method further comprises selecting
a candidate compound have a value for interacting with the target
kinase which has a preselected relationship with a reference value,
e.g., the value for the candidate compound exceeds a preselected
minimum value, e.g., a preselected minimum value for activation of
the target kinase.
[0322] In some embodiments, the method further comprises
evaluating, e.g., confirming, the ability of a candidate compound
(e.g., a candidate compound which meets a predetermined level of
interaction (e.g., complex formation, specific binding, or
modulation (e.g., activation or inhibition) in the evaluating step)
to interact with, e.g., to form a complex with, to bind
specifically to, or to modulate (e.g., activate or inhibit) the
activity of the target kinase in a second method.
[0323] In some embodiments, the method further comprises selecting
a candidate compound and repeating the evaluation under the same or
different conditions, e.g., at the same or a different
concentration.
[0324] In some embodiments, the method further comprises selecting
a candidate compound evaluated in a cell other than a whole animal
and confirming the activity determined in the cell-based assay by
evaluation in a whole animal.
[0325] In some embodiments, the method further comprises selecting
a candidate compound and confirming the activity determined in the
evaluation by a second, different assay.
[0326] In some embodiments, a plurality of structurally related
candidate candidates are evaluated, e.g., a plurality of candidate
candidates having a common core or scaffold.
[0327] In some embodiments, the method comprises providing a
plurality of second generation candidate candidates which are
analogs of a candidate compound.
[0328] In some embodiments, the method comprises evaluating a first
candidate compound, comparing the structure of the first candidate
compound to a second candidate compound and evaluating the second
candidate compound
[0329] In some embodiments, the candidate compound is contacted
with cultured cells, e.g., cultured cells having a preselected
level of confluency, e.g., from about 60% to about 95%, preferably
from about 70% to about 90% confluent.
[0330] In some embodiments, the candidate compound is contacted
with the cell for a preselected length of time, e.g., a time period
sufficient to allow a positive control to enter the cell and
interact with the target kinase.
[0331] In some embodiments, the contacting step comprises
contacting the compound with the cell for at least about 0.1, 0.5,
1, 2, 3, 4, 5, or 6 hours.
[0332] In some embodiments, the compound forms a complex with the
target kinase.
[0333] In some embodiments, the compound binds, e.g., specifically
to the target kinase.
[0334] In some embodiments, the target kinase is PKM2 and the
candidate compound induces a conformational change (e.g., from a
non-activated or less activated conformation to an activated or
more activated conformation or from an activated or more activated
conformation to a non-activated or less activated conformation) in
the target kinase.
[0335] In some embodiments, the target kinase is PKM2 and the
candidate compound increases the activity of PKM2.
[0336] In some embodiments, the target kinase is PKM2 and the
candidate compound decreases the activity of PKM2.
[0337] In some embodiments, the method further comprises evaluating
the presence and/or amount of lactate, e.g., in the media.
[0338] In some embodiments, the lysing step comprises snap-freezing
the cell, e.g., on dry ice.
[0339] In some embodiments, the lysing step comprises adding a
lysis buffer, e.g., a detergent-containing (e.g.,
Triton-containing) lysis buffer (e.g., a lysis buffer described in
Table 3), to the cell.
[0340] In some embodiments, the detergent is used at a
concentration that does not disrupt the interaction (e.g., binding)
between the compound and the kinase, e.g., pyruvate kinase, e.g.,
PKM2, e.g., at no more than about 0.1, 0.5, 1, 1.5, 2, or 5%.
[0341] In one aspect, the invention features a method of evaluating
a candidate compound the method comprising:
[0342] optionally supplying the candidate compound;
[0343] contacting the candidate compound with a cell, which cell is
outside an animal, e.g., a cell having an intact plasma
membrane;
[0344] separating the cell from candidate compound that has not
entered the cell;
[0345] lysing said cell under conditions that do not abolish the
binding of the candidate compound to PKM2; and
[0346] evaluating the ability of the compound to interact
intracellularly with, e.g., to form a complex with, to bind, e.g.,
specifically to, or to modulate (e.g., activate or inhibit) the
activity of PKM2;
[0347] thereby evaluating the candidate compound.
[0348] In one aspect, the invention features a method of evaluating
a candidate compound the method comprising:
[0349] optionally supplying the candidate compound;
[0350] contacting the candidate compound with a cell which is part
of a whole animal;
[0351] lysing said cell under conditions that do not abolish the
binding of the candidate compound to PKM2; and
[0352] evaluating the ability of the compound to interact
intracellularly with, e.g., to form a complex with, to bind, e.g.,
specifically to, or to modulate (e.g., activate or inhibit) the
activity of PKM2;
[0353] thereby evaluating the candidate compound.
[0354] In another embodiment the compound described herein is
administered at a dosage and frequency sufficient to increase
lactate production or oxidative phosphorylation.
[0355] The term "halo" or "halogen" refers to any radical of
fluorine, chlorine, bromine or iodine.
[0356] The term "alkyl" refers to a hydrocarbon chain that may be a
straight chain or branched chain, containing the indicated number
of carbon atoms. For example, C.sub.1-C.sub.12 alkyl indicates that
the group may have from 1 to 12 (inclusive) carbon atoms in it. The
term "haloalkyl" refers to an alkyl in which one or more hydrogen
atoms are replaced by halo, and includes alkyl moieties in which
all hydrogens have been replaced by halo (e.g., perfluoroalkyl).
The terms "arylalkyl" or "aralkyl" refer to an alkyl moiety in
which an alkyl hydrogen atom is replaced by an aryl group. Aralkyl
includes groups in which more than one hydrogen atom has been
replaced by an aryl group. Examples of "arylalkyl" or "aralkyl"
include benzyl, 2-phenylethyl, 3-phenylpropyl, 9-fluorenyl,
benzhydryl, and trityl groups.
[0357] The term "alkylene" refers to a divalent alkyl, e.g.,
--CH.sub.2--, --CH.sub.2CH.sub.2--, and
--CH.sub.2CH.sub.2CH.sub.2--.
[0358] The term "alkenyl" refers to a straight or branched
hydrocarbon chain containing 2-12 carbon atoms and having one or
more double bonds. Examples of alkenyl groups include, but are not
limited to, allyl, propenyl, 2-butenyl, 3-hexenyl and 3-octenyl
groups. One of the double bond carbons may optionally be the point
of attachment of the alkenyl substituent. The term "alkynyl" refers
to a straight or branched hydrocarbon chain containing 2-12 carbon
atoms and characterized in having one or more triple bonds.
Examples of alkynyl groups include, but are not limited to,
ethynyl, propargyl, and 3-hexynyl. One of the triple bond carbons
may optionally be the point of attachment of the alkynyl
substituent.
[0359] The terms "alkylamino" and "dialkylamino" refer to
--NH(alkyl) and --NH(alkyl).sub.2 radicals respectively. The term
"aralkylamino" refers to a --) radical. The term alkylaminoalkyl
refers to a (alkyl)NH-alkyl- radical; the term dialkylaminoalkyl
refers to a (alkyl).sub.2N-alkyl- radical The term "alkoxy" refers
to an --O-alkyl radical. The term "mercapto" refers to an SH
radical. The term "thioalkoxy" refers to an --S-alkyl radical. The
term thioaryloxy refers to an --S-aryl radical.
[0360] The term "aryl" refers to an aromatic monocyclic, bicyclic,
or tricyclic hydrocarbon ring system, wherein any ring atom capable
of substitution can be substituted (e.g., by one or more
substituents). Examples of aryl moieties include, but are not
limited to, phenyl, naphthyl, and anthracenyl.
[0361] The term "cycloalkyl" as employed herein includes saturated
cyclic, bicyclic, tricyclic, or polycyclic hydrocarbon groups
having 3 to 12 carbons. Any ring atom can be substituted (e.g., by
one or more substituents). The cycloalkyl groups can contain fused
rings. Fused rings are rings that share a common carbon atom.
Examples of cycloalkyl moieties include, but are not limited to,
cyclopropyl, cyclohexyl, methylcyclohexyl, adamantyl, and
norbornyl.
[0362] The term "heterocyclyl" refers to a nonaromatic 3-10
membered monocyclic, 8-12 membered bicyclic, or 11-14 membered
tricyclic ring system having 1-3 heteroatoms if monocyclic, 1-6
heteroatoms if bicyclic, or 1-9 heteroatoms if tricyclic, said
heteroatoms selected from O, N, or S (e.g., carbon atoms and 1-3,
1-6, or 1-9 heteroatoms of N, O, or S if monocyclic, bicyclic, or
tricyclic, respectively). The heteroatom may optionally be the
point of attachment of the heterocyclyl substituent. Any ring atom
can be substituted (e.g., by one or more substituents). The
heterocyclyl groups can contain fused rings. Fused rings are rings
that share a common carbon atom. Examples of heterocyclyl include,
but are not limited to, tetrahydrofuranyl, tetrahydropyranyl,
piperidinyl, morpholino, pyrrolinyl, pyrimidinyl, quinolinyl, and
pyrrolidinyl.
[0363] The term "cycloalkenyl" refers to partially unsaturated,
nonaromatic, cyclic, bicyclic, tricyclic, or polycyclic hydrocarbon
groups having 5 to 12 carbons, preferably 5 to 8 carbons. The
unsaturated carbon may optionally be the point of attachment of the
cycloalkenyl substituent. Any ring atom can be substituted (e.g.,
by one or more substituents). The cycloalkenyl groups can contain
fused rings. Fused rings are rings that share a common carbon atom.
Examples of cycloalkenyl moieties include, but are not limited to,
cyclohexenyl, cyclohexadienyl, or norbornenyl.
[0364] The term "heterocycloalkenyl" refers to a partially
saturated, nonaromatic 5-10 membered monocyclic, 8-12 membered
bicyclic, or 11-14 membered tricyclic ring system having 1-3
heteroatoms if monocyclic, 1-6 heteroatoms if bicyclic, or 1-9
heteroatoms if tricyclic, said heteroatoms selected from O, N, or S
(e.g., carbon atoms and 1-3, 1-6, or 1-9 heteroatoms of N, O, or S
if monocyclic, bicyclic, or tricyclic, respectively). The
unsaturated carbon or the heteroatom may optionally be the point of
attachment of the heterocycloalkenyl substituent. Any ring atom can
be substituted (e.g., by one or more substituents). The
heterocycloalkenyl groups can contain fused rings. Fused rings are
rings that share a common carbon atom. Examples of
heterocycloalkenyl include but are not limited to tetrahydropyridyl
and dihydropyranyl.
[0365] The term "heteroaryl" refers to an aromatic 5-8 membered
monocyclic, 8-12 membered bicyclic, or 11-14 membered tricyclic
ring system having 1-3 heteroatoms if monocyclic, 1-6 heteroatoms
if bicyclic, or 1-9 heteroatoms if tricyclic, said heteroatoms
selected from O, N, or S (e.g., carbon atoms and 1-3, 1-6, or 1-9
heteroatoms of N, O, or S if monocyclic, bicyclic, or tricyclic,
respectively). Any ring atom can be substituted (e.g., by one or
more substituents).
[0366] The term "oxo" refers to an oxygen atom, which forms a
carbonyl when attached to carbon, an N-oxide when attached to
nitrogen, and a sulfoxide or sulfone when attached to sulfur.
[0367] The term "acyl" refers to an alkylcarbonyl,
cycloalkylcarbonyl, arylcarbonyl, heterocyclylcarbonyl, or
heteroarylcarbonyl substituent, any of which may be further
substituted (e.g., by one or more substituents).
[0368] The term "substituents" refers to a group "substituted" on
an alkyl, cycloalkyl, alkenyl, alkynyl, heterocyclyl,
heterocycloalkenyl, cycloalkenyl, aryl, or heteroaryl group at any
atom of that group. Any atom can be substituted. Suitable
substituents include, without limitation, alkyl (e.g., C1, C2, C3,
C4, C5, C6, C7, C8, C9, C10, C11, C12 straight or branched chain
alkyl), cycloalkyl, haloalkyl (e.g., perfluoroalkyl such as
CF.sub.3), aryl, heteroaryl, aralkyl, heteroaralkyl, heterocyclyl,
alkenyl, alkynyl, cycloalkenyl, heterocycloalkenyl, alkoxy,
haloalkoxy (e.g., perfluoroalkoxy such as OCF.sub.3), halo,
hydroxy, carboxy, carboxylate, cyano, nitro, amino, alkyl amino,
SO.sub.3H, sulfate, phosphate, methylenedioxy (--O--CH.sub.2--O--
wherein oxygens are attached to vicinal atoms), ethylenedioxy, oxo,
thioxo (e.g., C.dbd.S), imino (alkyl, aryl, aralkyl),
S(O).sub.nalkyl (where n is 0-2), S(O).sub.n aryl (where n is 0-2),
S(O).sub.n heteroaryl (where n is 0-2), S(O).sub.n heterocyclyl
(where n is 0-2), amine (mono-, di-, alkyl, cycloalkyl, aralkyl,
heteroaralkyl, aryl, heteroaryl, and combinations thereof), ester
(alkyl, aralkyl, heteroaralkyl, aryl, heteroaryl), amide (mono-,
di-, alkyl, aralkyl, heteroaralkyl, aryl, heteroaryl, and
combinations thereof), sulfonamide (mono-, di-, alkyl, aralkyl,
heteroaralkyl, and combinations thereof). In one aspect, the
substituents on a group are independently any one single, or any
subset of the aforementioned substituents. In another aspect, a
substituent may itself be substituted with any one of the above
substituents.
[0369] The term "selective" is meant at least 2-fold, 3-fold,
4-fold, 5-fold, 6-fold, or 10-fold greater modulation (e.g.,
inhibition) of M2 than M1.
[0370] The term "activator" as used herein means an agent that
(measurably) increases the activity of a pyruvate kinase (e.g.,
PKM2) or causes pyruvate kinase (e.g., PKM2) activity to increase
to a level that is greater than PKM2's basal levels of activity.
For example, the activator may mimic the effect caused by a natural
ligand (e.g., FBP). The activator effect caused by the agent may be
to the same, or to a greater, or to a lesser extent than the
activating effect caused by a natural ligand, but the same type of
effect is caused. Peptides, nucleic acids, and small molecules may
be activators. An agent can be evaluated to determine if it is an
activator by measuring either directly or indirectly the activity
of the pyruvate kinase when subjected to the agent. The activity of
the agent can be measured, for example, against a control
substance. In some instances, the activity measured of the agent is
for activation of PKM2. The activity of PKM2 can be measured, for
example, by monitoring the concentration of a substrate such as ATP
or NADH.
[0371] The term "inhibitor" as used herein means an agent that
measurably slows, stops, decreases or inactivates the enzymatic
activity of pyruvate kinase (e.g., PKM2) to decrease to a level
that is less than the pyruvate kinase's (e.g., PKM2's) basal levels
of activity. Inhibitors of pyruvate kinase (e.g., PKM2) may be
peptides or nucleic acids. An agent can be evaluated to determine
if it is an inhibitor by measuring either directly or indirectly
the activity of the pyruvate kinase when subjected to the agent.
The activity of the agent can be measured, for example, against a
control substance. In some instances, the activity measured of the
agent is for inhibition of PKM2. The activity of PKM2 can be
measured, for example, by monitoring the concentration of a
substrate such as ATP or NADH.
[0372] The term "modulate" refers to an increase or decrease, e.g.,
in the activity of an enzyme in response to exposure to a compound
or composition described herein, e.g., the activation or inhibition
of PKM2, in at least a sub-population of cells in a subject such
that a desired end result is achieved (e.g., a therapeutic result).
In some embodiments, a compound as described herein inhibits a
target described herein, e.g., PKM2. In some embodiments, a
compound as described herein is activates a target described
herein, e.g., PKM2.
DETAILED DESCRIPTION
[0373] This invention is not limited in its application to the
details of construction and the arrangement of components set forth
in the following description or illustrated in the drawings. The
invention is capable of other embodiments and of being practiced or
of being carried out in various ways. Also, the phraseology and
terminology used herein is for the purpose of description and
should not be regarded as limiting. The use of "including,"
"comprising," or "having," "containing", "involving", and
variations thereof herein, is meant to encompass the items listed
thereafter and equivalents thereof as well as additional items.
Compounds
[0374] Described herein are compounds and compositions that
modulate PKM2, for example, activate or inhibit PKM2. Compounds
that modulate PKM2, e.g., activate or inhibit PKM2, can be used to
treat disorders such as neoplastic disorders (e.g., cancer) or fat
related disorders (e.g., obesity). Exemplary compounds include the
compounds of Formulas (I), (II), (III), (IV), (V), (VI) and (VII)
described herein. In some embodiments, a compound described herein
modulates PKM2 by interacting (e.g., binding) with the FBP binding
pocket. For example, a compound described herein can compete with
FBP binding in PKM2.
[0375] In some embodiments a compound described herein has one or
more properties described herein, e.g., one or more of the
following properties: it is an allosteric modulator (e.g.,
inhibitor or activator); it modulates the release of FBP (e.g.,
inhibits or promotes); it is a modulator (e.g., agonist or
antagonist) of FBP, e.g., an agonist which binds with a lower,
about the same, or higher affinity than does FBP; it modulates
(e.g., inhibits or promotes) the dissolution of tetrameric PKM2; it
modulates (e.g., promotes or inhibits) the assembly of tetrameric
PKM2; it selectively modulates (e.g., inhibits or activates) PKM2
over at least one other isoform of PK, e.g., it is selective for
PKM2 over PKR, PKM1, or PKL; is has an affinity for PKM2 which is
greater than its affinity for at least one other isoform of PK,
e.g., PKR, PKM1, or PKL.
[0376] In another embodiment the activator of PKM2 utilized in the
methods and compositions of this invention operates by or has one
or more of the following mechanisms or properties: [0377] a. it is
an allosteric activator of PKM2; [0378] b. it modulates (e.g.,
stabilizes or inhibits) the binding of FBP in a binding pocket of
PKM2; [0379] c. it modulates (e.g., inhibits or promotes) the
release of FBP from a binding pocket of PKM2; [0380] d. it is a
modulator (e.g., an agonist or antagonist), e.g., an analog, of
FBP, e.g., an agonist which binds PKM2 with a lower, about the
same, or higher affinity than does FBP; [0381] e. it modulates
(e.g., inhibits or promotes) the dissolution of tetrameric PKM2;
[0382] f. it modulates (e.g., inhibits or promotes) the assembly of
tetrameric PKM2; [0383] g. it modulates (e.g., stabilizes or
inhibits) the tetrameric conformation of PKM2; [0384] h. it
modulates (e.g., inhibits or promotes) the binding of a
phosphotyrosine containing polypeptide to PKM2; [0385] i. it
modulates (e.g., inhibits or promotes) the ability of a
phosphotyrosine containing polypeptide to induce release of FBP
from PKM2, e.g., by inducing a change in the conformation of PKM2,
e.g., in the position of Lys 433, thereby hindering the release of
FBP; [0386] k. it binds to or changes the position of Lys 433
relative to the FBP binding pocket; [0387] l. it selectively
modulates (e.g., activates or inhibits) PKM2 over at least one
other isoform of PK, e.g., it is selective for PKM2 over one or
more of PKR, PKM1, or PKL; [0388] m. it has an affinity for PKM2
which is greater than its affinity for at least one other isoform
of PK, e.g., PKR, PKM1, or PKL.
[0389] A compound described herein may be an activator of PKM2.
Exemplary compounds are shown in Table 1. As shown in Table 1, A
refers to an activator of PKM2 with an EC.sub.50<100 nM. B
refers to an activator of PKM2 with an EC.sub.50 between 100 nM and
500 nM. C refers to an activator of PKM2 with an EC.sub.50 between
500 nM and 1000 nM. D refers to an activator of PKM2 with an
EC.sub.50 between 1 .mu.M and 10 .mu.M. E refers to an activator of
PKM2 with an EC.sub.50>10 .mu.M ND means not determined.
[0390] Ex vivo data is provided as follows: + refers to a compound
having an activity of .ltoreq.1 .mu.M; ++ refers to a compound
having an activity of >1 .mu.M; ND means not determined
TABLE-US-00001 TABLE 1 Ex- Structure PKM2_AC50 Vivo_AC50
##STR00059## A + ##STR00060## D ND ##STR00061## B + ##STR00062## D
ND ##STR00063## C ND ##STR00064## B ++ ##STR00065## A +
##STR00066## D ND ##STR00067## B ND ##STR00068## C ND ##STR00069##
B ND ##STR00070## A + ##STR00071## A + ##STR00072## D ND
##STR00073## D ND ##STR00074## A + ##STR00075## D ND ##STR00076## C
ND ##STR00077## E ND ##STR00078## A ++ ##STR00079## D ND
##STR00080## C ND ##STR00081## A + ##STR00082## D ND ##STR00083## B
ND ##STR00084## A + ##STR00085## A + ##STR00086## D ND ##STR00087##
B + ##STR00088## A + ##STR00089## E ND ##STR00090## E ND
##STR00091## A + ##STR00092## C ND ##STR00093## A + ##STR00094## C
ND ##STR00095## B ND ##STR00096## B ND ##STR00097## D ND
##STR00098## A ++ ##STR00099## A + ##STR00100## B ND ##STR00101## A
ND ##STR00102## D ND ##STR00103## D ND ##STR00104## B +
##STR00105## D ND ##STR00106## D ND ##STR00107## C ND ##STR00108##
D ND ##STR00109## D ND ##STR00110## B + ##STR00111## B ND
##STR00112## B ND ##STR00113## D ND ##STR00114## D ND ##STR00115##
D ND ##STR00116## A + ##STR00117## B ND ##STR00118## E ND
##STR00119## D ND ##STR00120## E ND ##STR00121## A + ##STR00122## D
ND ##STR00123## D ND ##STR00124## D ND ##STR00125## D ND
##STR00126## A + ##STR00127## B ND ##STR00128## A + ##STR00129## D
ND ##STR00130## C ND ##STR00131## A + ##STR00132## A ND
##STR00133## B + ##STR00134## B ND ##STR00135## B ND ##STR00136## D
ND ##STR00137## B ND ##STR00138## E ND ##STR00139## ND ND
##STR00140## D ND ##STR00141## E ND ##STR00142## A + ##STR00143## D
ND ##STR00144## C ND ##STR00145## B ND ##STR00146## A +
##STR00147## A + ##STR00148## C ND ##STR00149## D ND ##STR00150## B
ND ##STR00151## B + ##STR00152## A + ##STR00153## B ND ##STR00154##
B ND ##STR00155## D ND ##STR00156## D ND ##STR00157## D ND
##STR00158## B + ##STR00159## D ND ##STR00160## B + ##STR00161## B
+ ##STR00162## A ND ##STR00163## D ND ##STR00164## D ND
##STR00165## B ND ##STR00166## B ND ##STR00167## C ND ##STR00168##
D ND ##STR00169## C ND ##STR00170## D ND ##STR00171## A ++
##STR00172## B ND ##STR00173## B ND ##STR00174## D + ##STR00175## B
ND ##STR00176## C ND ##STR00177## C ND ##STR00178## A ND
##STR00179## A ND ##STR00180## D ND ##STR00181## E ND
##STR00182## D ND ##STR00183## D ND ##STR00184## D ND ##STR00185##
B ND ##STR00186## C ND ##STR00187## B ND ##STR00188## A ND
##STR00189## ##STR00190## D ND ##STR00191## B ND ##STR00192## B ND
##STR00193## A ND ##STR00194## D ND ##STR00195## D ND ##STR00196##
D ND ##STR00197## E ND ##STR00198## D ND ##STR00199## E ND
##STR00200## C ND ##STR00201## D ND ##STR00202## D ND ##STR00203##
D ND ##STR00204## C ND ##STR00205## B ND
[0391] The compounds described herein can be made using a variety
of synthetic techniques. Scheme 1 below depicts a representative
synthesis of certain compounds described herein.
##STR00206##
[0392] As can be appreciated by the skilled artisan, methods of
synthesizing the compounds of the formulae herein will be evident
to those of ordinary skill in the art. Additionally, the various
synthetic steps may be performed in an alternate sequence or order
to give the desired compounds. Synthetic chemistry transformations
and protecting group methodologies (protection and deprotection)
useful in synthesizing the compounds described herein are known in
the art and include, for example, those such as described in R.
Larock, Comprehensive Organic Transformations, VCH Publishers
(1989); T. W. Greene and P. G. M. Wuts, Protective Groups in
Organic Synthesis, 2d. Ed., John Wiley and Sons (1991); L. Fieser
and M. Fieser, Fieser and Fieser's Reagents for Organic Synthesis,
John Wiley and Sons (1994); and L. Paquette, ed., Encyclopedia of
Reagents for Organic Synthesis, John Wiley and Sons (1995), and
subsequent editions thereof.
[0393] The compounds of this invention may contain one or more
asymmetric centers and thus occur as racemates and racemic
mixtures, single enantiomers, individual diastereomers and
diastereomeric mixtures. All such isomeric forms of these compounds
are expressly included in the present invention. The compounds of
this invention may also contain linkages (e.g., carbon-carbon
bonds) or substituents that can restrict bond rotation, e.g.
restriction resulting from the presence of a ring or double bond.
Accordingly, all cis/trans and E/Z isomers are expressly included
in the present invention.
[0394] The compounds of this invention may also be represented in
multiple tautomeric forms, in such instances, the invention
expressly includes all tautomeric forms of the compounds described
herein, even though only a single tautomeric form may be
represented (e.g., alkylation of a ring system may result in
alkylation at multiple sites, the invention expressly includes all
such reaction products). All such isomeric forms of such compounds
are expressly included in the present invention. All crystal forms
of the compounds described herein are expressly included in the
present invention including hydrates and other solvates.
[0395] The compounds of this invention include the compounds
themselves, as well as their salts and their prodrugs, if
applicable. A salt, for example, can be formed between an anion and
a positively charged substituent (e.g., amino) on a compound
described herein. Suitable anions include chloride, bromide,
iodide, sulfate, nitrate, phosphate, citrate, methanesulfonate,
trifluoroacetate, and acetate. Likewise, a salt can also be formed
between a cation and a negatively charged substituent (e.g.,
carboxylate) on a compound described herein. Suitable cations
include sodium ion, potassium ion, magnesium ion, calcium ion, and
an ammonium cation such as tetramethylammonium ion. Examples of
prodrugs include esters and other pharmaceutically acceptable
derivatives, which, upon administration to a subject, are capable
of providing active compounds.
[0396] The compounds of this invention may be modified by appending
appropriate functionalities to enhance selected biological
properties, e.g., targeting to a particular tissue. Such
modifications are known in the art and include those which increase
biological penetration into a given biological compartment (e.g.,
blood, lymphatic system, central nervous system), increase oral
availability, increase solubility to allow administration by
injection, alter metabolism and alter rate of excretion.
[0397] In an alternate embodiment, the compounds described herein
may be used as platforms or scaffolds that may be utilized in
combinatorial chemistry techniques for preparation of derivatives
and/or chemical libraries of compounds. Such derivatives and
libraries of compounds have biological activity and are useful for
identifying and designing compounds possessing a particular
activity. Combinatorial techniques suitable for utilizing the
compounds described herein are known in the art as exemplified by
Obrecht, D. and Villalgrodo, J. M., Solid-Supported Combinatorial
and Parallel Synthesis of Small-Molecular-Weight Compound
Libraries, Pergamon-Elsevier Science Limited (1998), and include
those such as the "split and pool" or "parallel" synthesis
techniques, solid-phase and solution-phase techniques, and encoding
techniques (see, for example, Czarnik, A. W., Curr. Opin. Chem.
Bio., (1997) 1, 60. Thus, one embodiment relates to a method of
using the compounds described herein for generating derivatives or
chemical libraries comprising: 1) providing a body comprising a
plurality of wells; 2) providing one or more compounds identified
by methods described herein in each well; 3) providing an
additional one or more chemicals in each well; 4) isolating the
resulting one or more products from each well. An alternate
embodiment relates to a method of using the compounds described
herein for generating derivatives or chemical libraries comprising:
1) providing one or more compounds described herein attached to a
solid support; 2) treating the one or more compounds identified by
methods described herein attached to a solid support with one or
more additional chemicals; 3) isolating the resulting one or more
products from the solid support. In the methods described above,
"tags" or identifier or labeling moieties may be attached to and/or
detached from the compounds described herein or their derivatives,
to facilitate tracking, identification or isolation of the desired
products or their intermediates. Such moieties are known in the
art. The chemicals used in the aforementioned methods may include,
for example, solvents, reagents, catalysts, protecting group and
deprotecting group reagents and the like. Examples of such
chemicals are those that appear in the various synthetic and
protecting group chemistry texts and treatises referenced
herein.
Methods of Evaluating Compounds
[0398] The compounds described herein can be evaluated for ability
to modulate PKM2 (e.g., activate or inhibit PKM2) by methods known
in the art. Exemplary methods include contacting the compound with
a cell-based assay which allows assessment of the ability to
modulate (e.g., activate or inhibit) PKM2. E.g., the candidate
compound can be contacted with a cell and measuring the consumption
of oxygen or production of lactate. A change in cellular
phosphoenolpyruvate, a change in glycerol-phosphate, a change in
ribose or deoxyribose, a change in lipid synthesis, or a change in
glucose conversion to lipid or nucleic acids or amino acids or
protein can also be used to evaluate a compound for its ability to
modulate PKM2 (e.g., activate or inhibit PKM2). The evaluation
could also include measuring a change in pyruvate or a
determination of an alteration in mitochondrial membrane potential,
e.g., as measured by fluorescent potentiometric dyes.
[0399] PKM1 and PKM2 for use in the screening method may be
produced by any method known in the art for expression of
recombinant proteins. For example, nucleic acids that encode the
desired polypeptide may be introduced into various cell types or
cell-free systems for expression. Eukaryotic (e.g., COS, HEK293T,
CHO, and NIH cell lines) and prokaryotic (e.g., E. coli) expression
systems may be generated in which a PKM sequence is introduced into
a plasmid or other vector, which is then used to transform living
cells. Constructs in which the PKM cDNA contains the entire open
reading frame, or biologically active fragment thereof, are
inserted in the correct orientation into an expression plasmid and
may be used for protein expression. Prokaryotic and eukaryotic
expression systems allow for the expression and recovery of fusion
proteins in which the PKM protein is covalently linked to a tag
molecule on either the amino terminal or carboxy terminal side,
which facilitates identification and/or purification. Examples of
tags that can be used include hexahistidine, HA, FLAG, and c-myc
epitope tags. An enzymatic or chemical cleavage site can be
engineered between the PKM protein and the tag molecule so that the
tag can be removed following purification.
[0400] The activity of the PKM enzyme measured in the screening
assay may be measured by, e.g., monitoring the concentration of a
substrate (e.g., ATP or NADH) present in the reaction mixture.
Pyruvate, produced by the enzymatic activity of pyruvate kinase, is
converted into lactate by lactate dehydrogenase, which requires the
consumption of NADH (NADH.fwdarw.NAD+). Thus, the activity of PKM2
can be indirectly measured by monitoring the consumption of NADH
through, e.g., fluorescence assays. Additionally, the activity of
the PKM2 enzyme can be directly monitored by measuring the
production of ATP, as ATP is produced when phosphoenolpyruvate is
converted to pyruvate. Methods for monitoring the amount of
substrate in a reaction mixture include, e.g., absorbance,
fluorescence, Raman scattering, phosphorescence, luminescence,
luciferase assays, and radioactivity.
[0401] The screening procedure requires the presence of specific
components in the reaction mixture. Components utilized in the
assay include, e.g., a nucleoside diphosphate (e.g., ADP),
phosphoenolpyruvate, NADH, lactate dehydrogenase, FBP, a reducing
agent (e.g., dithiothreitol), a detergent (e.g., Brij 35),
glycerol, and a solvent (e.g., DMSO). Exemplary reaction conditions
are found in Table 2.
TABLE-US-00002 TABLE 2 Component of Amount in Inhibition Amount in
Reaction Condition Assay Activation Assay ADP 0.1-5.0 mM 0.1-5.0 mM
Phosphoenolpyruvate 0.1-5.0 mM 0.1-5.0 mM NADH 10-1000 .mu.M
10-1000 .mu.M Lactate dehydrogenase 0.1-10 units 0.1-10 units
Fructose-1,6-bisphosphate 1-500 .mu.M 0 DTT 0.1-50 mM 0.1-50 mM
Brij 35 0.01-1% 0.01-1% Glycerol 0.1-10% 0.1-10% Pyruvate Kinase M2
1-100 pg 1-100 pg (used for screen) DMSO 1-10% 1-10%
[0402] Candidate inhibitory compounds are chosen if they
demonstrate specificity for PKM2 and inhibition of the PKM2 enzyme
greater than 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70,
75, 80, 85, 90, 95, 99, or 99.9%.
[0403] Candidate activator compounds are chosen if they demonstrate
specificity and activation of PKM2 enzyme in the absence of FBP to
a level greater than that of 10, 15, 20, 25, 30, 35, 40, 45, 50,
55, 60, 65, 70, 75, 80, 85, 90, 95, 99, or 100% in the presence of
FBP. Furthermore, specific candidate activators of PKM2 can be
evaluated in the presence or absence of a phosphotyrosine peptide.
Phosphotyrosine peptide binding to PKM2 leads to a dissociation of
FBP from PKM2 and conformational changes of PKM2 from an active,
tetrameric form to an inactive form. Compounds that bind to PKM2
and lock the enzyme in the active confirmation even in the presence
of a phosphotyrosine peptide will lead to the loss of allosteric
control of PKM2 needed for shunting the biochemical intermediates
from glycolysis into biosynthesis of other intermediates. This, in
turn, will lead to inhibition of growth of cancer cells, activated
immune cells and fat cells.
[0404] Exemplary screening assays also include ex vivo assays, for
example, an ex vivo assay described herein.
Methods of Treatment
[0405] The compounds and compositions described herein can be
administered to cells in culture, e.g. in vitro or ex vivo, or to a
subject, e.g., in vivo, to treat, prevent, and/or diagnose a
variety of disorders, including those described herein below.
[0406] As used herein, the term "treat" or "treatment" is defined
as the application or administration of a compound, alone or in
combination with, a second compound to a subject, e.g., a patient,
or application or administration of the compound to an isolated
tissue or cell, e.g., cell line, from a subject, e.g., a patient,
who has a disorder (e.g., a disorder as described herein), a
symptom of a disorder, or a predisposition toward a disorder, with
the purpose to cure, heal, alleviate, relieve, alter, remedy,
ameliorate, improve or affect the disorder, one or more symptoms of
the disorder or the predisposition toward the disorder (e.g., to
prevent at least one symptom of the disorder or to delay onset of
at least one symptom of the disorder).
[0407] As used herein, an amount of a compound effective to treat a
disorder, or a "therapeutically effective amount" refers to an
amount of the compound which is effective, upon single or multiple
dose administration to a subject, in treating a cell, or in curing,
alleviating, relieving or improving a subject with a disorder
beyond that expected in the absence of such treatment.
[0408] As used herein, an amount of a compound effective to prevent
a disorder, or a "a prophylactically effective amount" of the
compound refers to an amount effective, upon single- or
multiple-dose administration to the subject, in preventing or
delaying the occurrence of the onset or recurrence of a disorder or
a symptom of the disorder.
[0409] As used herein, the term "subject" is intended to include
human and non-human animals. Exemplary human subjects include a
human patient having a disorder, e.g., a disorder described herein
or a normal subject. The term "non-human animals" of the invention
includes all vertebrates, e.g., non-mammals (such as chickens,
amphibians, reptiles) and mammals, such as non-human primates,
domesticated and/or agriculturally useful animals, e.g., sheep,
dog, cat, cow, pig, etc.
Neoplastic Disorders
[0410] A compound or composition described herein can be used to
treat a neoplastic disorder. A "neoplastic disorder" is a disease
or disorder characterized by cells that have the capacity for
autonomous growth or replication, e.g., an abnormal state or
condition characterized by proliferative cell growth. Exemplary
neoplastic disorders include: carcinoma, sarcoma, metastatic
disorders (e.g., tumors arising from prostate, colon, lung, breast
and liver origin), hematopoietic neoplastic disorders, e.g.,
leukemias, metastatic tumors. Prevalent cancers include: breast,
prostate, colon, lung, liver, and pancreatic cancers. Treatment
with the compound may be in an amount effective to ameliorate at
least one symptom of the neoplastic disorder, e.g., reduced cell
proliferation, reduced tumor mass, etc.
[0411] The disclosed methods are useful in the prevention and
treatment of cancer, including for example, solid tumors, soft
tissue tumors, and metastases thereof. The disclosed methods are
also useful in treating non-solid cancers. Exemplary solid tumors
include malignancies (e.g., sarcomas, adenocarcinomas, and
carcinomas) of the various organ systems, such as those of lung,
breast, lymphoid, gastrointestinal (e.g., colon), and genitourinary
(e.g., renal, urothelial, or testicular tumors) tracts, pharynx,
prostate, and ovary. Exemplary adenocarcinomas include colorectal
cancers, renal-cell carcinoma, liver cancer, non-small cell
carcinoma of the lung, and cancer of the small intestine.
[0412] Exemplary cancers described by the national cancer institute
include: Acute Lymphoblastic Leukemia, Adult; Acute Lymphoblastic
Leukemia, Childhood; Acute Myeloid Leukemia, Adult; Adrenocortical
Carcinoma; Adrenocortical Carcinoma, Childhood; AIDS-Related
Lymphoma; AIDS-Related Malignancies; Anal Cancer; Astrocytoma,
Childhood Cerebellar; Astrocytoma, Childhood Cerebral; Bile Duct
Cancer, Extrahepatic; Bladder Cancer; Bladder Cancer, Childhood;
Bone Cancer, Osteosarcoma/Malignant Fibrous Histiocytoma; Brain
Stem Glioma, Childhood; Brain Tumor, Adult; Brain Tumor, Brain Stem
Glioma, Childhood; Brain Tumor, Cerebellar Astrocytoma, Childhood;
Brain Tumor, Cerebral Astrocytoma/Malignant Glioma, Childhood;
Brain Tumor, Ependymoma, Childhood; Brain Tumor, Medulloblastoma,
Childhood; Brain Tumor, Supratentorial Primitive Neuroectodermal
Tumors, Childhood; Brain Tumor, Visual Pathway and Hypothalamic
Glioma, Childhood; Brain Tumor, Childhood (Other); Breast Cancer;
Breast Cancer and Pregnancy; Breast Cancer, Childhood; Breast
Cancer, Male; Bronchial Adenomas/Carcinoids, Childhood; Carcinoid
Tumor, Childhood; Carcinoid Tumor, Gastrointestinal; Carcinoma,
Adrenocortical; Carcinoma, Islet Cell; Carcinoma of Unknown
Primaiy; Central Nervous System Lymphoma, Primary; Cerebellar
Astrocytoma, Childhood; Cerebral Astrocytoma/Malignant Glioma,
Childhood; Cervical Cancer; Childhood Cancers; Chronic Lymphocytic
Leukemia; Chronic Myelogenous Leukemia; Chronic Myeloproliferative
Disorders; Clear Cell Sarcoma of Tendon Sheaths; Colon Cancer;
Colorectal Cancer, Childhood; Cutaneous T-Cell Lymphoma;
Endometrial Cancer; Ependymoma, Childhood; Epithelial Cancer,
Ovarian; Esophageal Cancer; Esophageal Cancer, Childhood; Ewing's
Family of Tumors; Extracranial Germ Cell Tumor, Childhood;
Extragonadal Germ Cell Tumor; Extrahepatic Bile Duct Cancer; Eye
Cancer, Intraocular Melanoma; Eye Cancer, Retinoblastoma;
Gallbladder Cancer; Gastric (Stomach) Cancer; Gastric (Stomach)
Cancer, Childhood; Gastrointestinal Carcinoid Tumor; Germ Cell
Tumor, Extracranial, Childhood; Germ Cell Tumor, Extragonadal; Germ
Cell Tumor, Ovarian; Gestational Trophoblastic Tumor; Glioma,
Childhood Brain Stem; Glioma, Childhood Visual Pathway and
Hypothalamic; Hairy Cell Leukemia; Head and Neck Cancer;
Hepatocellular (Liver) Cancer, Adult (Primary); Hepatocellular
(Liver) Cancer, Childhood (Primary); Hodgkin's Lymphoma, Adult;
Hodgkin's Lymphoma, Childhood; Hodgkin's Lymphoma During Pregnancy;
Hypopharyngeal Cancer; Hypothalamic and Visual Pathway Glioma,
Childhood; Intraocular Melanoma; Islet Cell Carcinoma (Endocrine
Pancreas); Kaposi's Sarcoma; Kidney Cancer; Laryngeal Cancer;
Laryngeal Cancer, Childhood; Leukemia, Acute Lymphoblastic, Adult;
Leukemia, Acute Lymphoblastic, Childhood; Leukemia, Acute Myeloid,
Adult; Leukemia, Acute Myeloid, Childhood; Leukemia, Chronic
Lymphocytic; Leukemia, Chronic Myelogenous; Leukemia, Hairy Cell;
Lip and Oral Cavity Cancer; Liver Cancer, Adult (Primary); Liver
Cancer, Childhood (Primary); Lung Cancer, Non-Small Cell; Lung
Cancer, Small Cell; Lymphoblastic Leukemia, Adult Acute;
Lymphoblastic Leukemia, Childhood Acute; Lymphocytic Leukemia,
Chronic; Lymphoma, AIDS-Related; Lymphoma, Central Nervous System
(Primary); Lymphoma, Cutaneous T-Cell; Lymphoma, Hodgkin's, Adult;
Lymphoma, Hodgkin's, Childhood; Lymphoma, Hodgkin's During
Pregnancy; Lymphoma, Non-Hodgkin's, Adult; Lymphoma, Non-Hodgkin's,
Childhood; Lymphoma, Non-Hodgkin's During Pregnancy; Lymphoma,
Primary Central Nervous System; Macroglobulinemia, Waldenstrom's;
Male Breast Cancer; Malignant Mesothelioma, Adult; Malignant
Mesothelioma, Childhood; Malignant Thymoma; Medulloblastoma,
Childhood; Melanoma; Melanoma, Intraocular; Merkel Cell Carcinoma;
Mesothelioma, Malignant; Metastatic Squamous Neck Cancer with
Occult Primary; Multiple Endocrine Neoplasia Syndrome, Childhood;
Multiple Myeloma/Plasma Cell Neoplasm; Mycosis Fungoides;
Myelodysplastic Syndromes; Myelogenous Leukemia, Chronic; Myeloid
Leukemia, Childhood Acute; Myeloma, Multiple; Myeloproliferative
Disorders, Chronic; Nasal Cavity and Paranasal Sinus Cancer;
Nasopharyngeal Cancer; Nasopharyngeal Cancer, Childhood;
Neuroblastoma; Non-Hodgkin's Lymphoma, Adult; Non-Hodgkin's
Lymphoma, Childhood; Non-Hodgkin's Lymphoma During Pregnancy;
Non-Small Cell Lung Cancer; Oral Cancer, Childhood; Oral Cavity and
Lip Cancer; Oropharyngeal Cancer; Osteosarcoma/Malignant Fibrous
Histiocytoma of Bone; Ovarian Cancer, Childhood; Ovarian Epithelial
Cancer; Ovarian Germ Cell Tumor; Ovarian Low Malignant Potential
Tumor; Pancreatic Cancer; Pancreatic Cancer, Childhood; Pancreatic
Cancer, Islet Cell; Paranasal Sinus and Nasal Cavity Cancer;
Parathyroid Cancer; Penile Cancer; Pheochromocytoma; Pineal and
Supratentorial Primitive Neuroectodermal Tumors, Childhood;
Pituitary Tumor; Plasma Cell Neoplasm/Multiple Myeloma;
Pleuropulmonary Blastoma; Pregnancy and Breast Cancer; Pregnancy
and Hodgkin's Lymphoma; Pregnancy and Non-Hodgkin's Lymphoma;
Primary Central Nervous System Lymphoma; Primary Liver Cancer,
Adult; Primary Liver Cancer, Childhood; Prostate Cancer; Rectal
Cancer; Renal Cell (Kidney) Cancer; Renal Cell Cancer, Childhood;
Renal Pelvis and Ureter, Transitional Cell Cancer; Retinoblastoma;
Rhabdomyosarcoma, Childhood; Salivary Gland Cancer; Salivary Gland
Cancer, Childhood; Sarcoma, Ewing's Family of Tumors; Sarcoma,
Kaposi's; Sarcoma (Osteosarcoma)/Malignant Fibrous Histiocytoma of
Bone; Sarcoma, Rhabdomyosarcoma, Childhood; Sarcoma, Soft Tissue,
Adult; Sarcoma, Soft Tissue, Childhood; Sezary Syndrome; Skin
Cancer; Skin Cancer, Childhood; Skin Cancer (Melanoma); Skin
Carcinoma, Merkel Cell; Small Cell Lung Cancer; Small Intestine
Cancer; Soft Tissue Sarcoma, Adult; Soft Tissue Sarcoma, Childhood;
Squamous Neck Cancer with Occult Primary, Metastatic; Stomach
(Gastric) Cancer; Stomach (Gastric) Cancer, Childhood;
Supratentorial Primitive Neuroectodermal Tumors, Childhood; T-Cell
Lymphoma, Cutaneous; Testicular Cancer; Thymoma, Childhood;
Thymoma, Malignant; Thyroid Cancer; Thyroid Cancer, Childhood;
Transitional Cell Cancer of the Renal Pelvis and Ureter;
Trophoblastic Tumor, Gestational; Unknown Primary Site, Cancer of,
Childhood; Unusual Cancers of Childhood; Ureter and Renal Pelvis,
Transitional Cell Cancer; Urethral Cancer; Uterine Sarcoma; Vaginal
Cancer; Visual Pathway and Hypothalamic Glioma, Childhood; Vulvar
Cancer; Waldenstrom's Macro globulinemia; and Wilms' Tumor.
Metastases of the aforementioned cancers can also be treated or
prevented in accordance with the methods described herein.
Cancer Combination therapies
[0413] In some embodiments, a compound described herein is
administered together with an additional cancer treatment.
Exemplary cancer treatments include, for example: chemotherapy,
targeted therapies such as antibody therapies, immunotherapy, and
hormonal therapy. Examples of each of these treatments are provided
below.
[0414] Chemotherapy
[0415] In some embodiments, a compound described herein is
administered with a chemotherapy. Chemotherapy is the treatment of
cancer with drugs that can destroy cancer cells. "Chemotherapy"
usually refers to cytotoxic drugs which affect rapidly dividing
cells in general, in contrast with targeted therapy. Chemotherapy
drugs interfere with cell division in various possible ways, e.g.,
with the duplication of DNA or the separation of newly formed
chromosomes. Most forms of chemotherapy target all rapidly dividing
cells and are not specific for cancer cells, although some degree
of specificity may come from the inability of many cancer cells to
repair DNA damage, while normal cells generally can.
[0416] Examples of chemotherapeutic agents used in cancer therapy
include, for example, antimetabolites (e.g., folic acid, purine,
and pyrimidine derivatives) and alkylating agents (e.g., nitrogen
mustards, nitrosoureas, platinum, alkyl sulfonates, hydrazines,
triazenes, aziridines, spindle poison, cytotoxic agents,
toposimerase inhibitors and others). Exemplary agents include
Aclarubicin, Actinomycin, Alitretinon, Altretamine, Aminopterin,
Aminolevulinic acid, Amrubicin, Amsacrine, Anagrelide, Arsenic
trioxide, Asparaginase, Atrasentan, Belotecan, Bexarotene,
endamustine, Bleomycin, Bortezomib, Busulfan, Camptothecin,
Capecitabine, Carboplatin, Carboquone, Carmofur, Carmustine,
Celecoxib, Chlorambucil, Chlormethine, Cisplatin, Cladribine,
Clofarabine, Crisantaspase, Cyclophosphamide, Cytarabine,
Dacarbazine, Dactinomycin, Daunorubicin, Decitabine, Demecolcine,
Docetaxel, Doxorubicin, Efaproxiral, Elesclomol, Elsamitrucin,
Enocitabine, Epirubicin, Estramustine, Etoglucid, Etoposide,
Floxuridine, Fludarabine, Fluorouracil (5FU), Fotemustine,
Gemcitabine, Gliadel implants, Hydroxycarbamide, Hydroxyurea,
Idarubicin, Ifosfamide, Irinotecan, Irofulven, Ixabepilone,
Larotaxel, Leucovorin, Liposomal doxorubicin, Liposomal
daunorubicin, Lonidamine, Lomustine, Lucanthone, Mannosulfan,
Masoprocol, Melphalan, Mercaptopurine, Mesna, Methotrexate, Methyl
aminolevulinate, Mitobronitol, Mitoguazone, Mitotane, Mitomycin,
Mitoxantrone, Nedaplatin, Nimustine, Oblimersen, Omacetaxine,
Ortataxel, Oxaliplatin, Paclitaxel, Pegaspargase, Pemetrexed,
Pentostatin, Pirarubicin, Pixantrone, Plicamycin, Porfimer sodium,
Prednimustine, Procarbazine, Raltitrexed, Ranimustine, Rubitecan,
Sapacitabine, Semustine, Sitimagene ceradenovec, Strataplatin,
Streptozocin, Talaporfin, Tegafur-uracil, Temoporfin, Temozolomide,
Teniposide, Tesetaxel, Testolactone, Tetranitrate, Thiotepa,
Tiazofurine, Tioguanine, Tipifarnib, Topotecan, Trabectedin,
Triaziquone, Triethylenemelamine, Triplatin, Tretinoin, Treosulfan,
Trofosfamide, Uramustine, Valrubicin, Verteporfin, Vinblastine,
Vincristine, Vindesine, Vinflunine, Vinorelbine, Vorinostat,
Zorubicin, and other cytostatic or cytotoxic agents described
herein.
[0417] Because some drugs work better together than alone, two or
more drugs are often given at the same time. Often, two or more
chemotherapy agents are used as combination chemotherapy. In some
embodiments, the chemotherapy agents (including combination
chemotherapy) can be used in combination with a compound described
herein.
[0418] Targeted Therapy
[0419] In some embodiments, a compound described herein is
administered with a targeted therapy. Targeted therapy constitutes
the use of agents specific for the deregulated proteins of cancer
cells. Small molecule targeted therapy drugs are generally
inhibitors of enzymatic domains on mutated, overexpressed, or
otherwise critical proteins within the cancer cell. Prominent
examples are the tyrosine kinase inhibitors such as Axitinib,
Bosutinib, Cediranib, desatinib, erolotinib, imatinib, gefitinib,
lapatinib, Lestaurtinib, Nilotinib, Semaxanib, Sorafenib,
Sunitinib, and Vandetanib, and also cyclin-depdendent kinase
inhibitors such as Alvocidib and Seliciclib. Monoclonal antibody
therapy is another strategy in which the therapeutic agent is an
antibody which specifically binds to a protein on the surface of
the cancer cells. Examples include the anti-HER2/neu antibody
trastuzumab (HERCEPTIN.RTM.) typically used in breast cancer, and
the anti-CD20 antibody rituximab and Tositumomab typically used in
a variety of B-cell malignancies. Other exemplary anbitodies
include Ctuximab, Panitumumab, Trastuzumab, Alemtuzumab,
Bevacizumab, Edrecolomab, and Gemtuzumab. Exemplary fusion proteins
include Aflibercept and Denileukin diftitox. In some embodiments,
the targeted therapy can be used in combination with a compound
described herein.
[0420] Targeted therapy can also involve small peptides as "homing
devices" which can bind to cell surface receptors or affected
extracellular matrix surrounding the tumor. Radionuclides which are
attached to these peptides (e.g., RGDs) eventually kill the cancer
cell if the nuclide decays in the vicinity of the cell. An example
of such therapy includes BEXXAR.RTM..
[0421] Immunotherapy
[0422] In some embodiments, a compound described herein is
administered with an immunotherapy. Cancer immunotherapy refers to
a diverse set of therapeutic strategies designed to induce the
patient's own immune system to fight the tumor. Contemporary
methods for generating an immune response against tumors include
intravesicular BCG immunotherapy for superficial bladder cancer,
and use of interferons and other cytokines to induce an immune
response in renal cell carcinoma and melanoma patients.
[0423] Allogeneic hematopoietic stem cell transplantation can be
considered a form of immunotherapy, since the donor's immune cells
will often attack the tumor in a graft-versus-tumor effect. In some
embodiments, the immunotherapy agents can be used in combination
with a compound described herein.
[0424] Hormonal Therapy
[0425] In some embodiments, a compound described herein is
administered with a hormonal therapy. The growth of some cancers
can be inhibited by providing or blocking certain hormones. Common
examples of hormone-sensitive tumors include certain types of
breast and prostate cancers. Removing or blocking estrogen or
testosterone is often an important additional treatment. In certain
cancers, administration of hormone agonists, such as progestogens
may be therapeutically beneficial. In some embodiments, the
hormonal therapy agents can be used in combination with a compound
described herein.
Obesity and Fat Disorders
[0426] A compound or composition described herein can be used to
treat or prevent obesity, e.g., in a human subject, e.g. a child or
adult subject. "Obesity" refers to a condition in which a subject
has a body mass index of greater than or equal to 30. Many
compounds described herein can be used to treat or prevent an
over-weight condition. "Over-weight" refers to a condition in which
a subject has a body mass index of greater or equal to 25.0. The
body mass index (BMI) and other definitions are according to the
"NIH Clinical Guidelines on the Identification and Evaluation, and
Treatment of Overweight and Obesity in Adults" (1998). Treatment
with the compound may be in an amount effective to alter the weight
of the subject, e.g., by at least 2, 5, 7, 10, 12, 15, 20, 25, 30,
25, 40, 45, 50, or 55%. Treatment with a compound may be in an
amount effective to reduce the body mass index of the subject,
e.g., to less than 30, 28, 27, 25, 22, 20, or 18. The compounds can
be used to treat or prevent aberrant or inappropriate weight gain,
metabolic rate, or fat deposition, e.g., anorexia, bulimia,
obesity, diabetes, or hyperlipidemia (e.g., elevated triglycerides
and/or elevated cholesterol), as well as disorders of fat or lipid
metabolism.
[0427] A compound or composition described herein can be
administered to treat obesity associated with Prader-Willi Syndrome
(PWS). PWS is a genetic disorder associated with obesity (e.g.,
morbid obesity).
[0428] A compound or composition described herein can be used to
reduce body fat, prevent increased body fat, reduce cholesterol
(e.g., total cholesterol and/or ratios of total cholesterol to HDL
cholesterol), and/or reduce appetite in individuals having PWS
associated obesity, and/or reduce comorbidities such as diabetes,
cardiovascular disease, and stroke.
Compositions and Routes of Administration
[0429] The compositions delineated herein include the compounds
delineated herein (e.g., a compound described herein), as well as
additional therapeutic agents if present, in amounts effective for
achieving a modulation of disease or disease symptoms, including
those described herein.
[0430] The term "pharmaceutically acceptable carrier or adjuvant"
refers to a carrier or adjuvant that may be administered to a
patient, together with a compound of this invention, and which does
not destroy the pharmacological activity thereof and is nontoxic
when administered in doses sufficient to deliver a therapeutic
amount of the compound.
[0431] 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, self-emulsifying drug delivery systems
(SEDDS) such as d-.alpha.-tocopherol polyethyleneglycol 1000
succinate, surfactants used in pharmaceutical dosage forms such as
Tweens or other similar polymeric delivery matrices, 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. Cyclodextrins such as .alpha.-, .beta.-, and
.gamma.-cyclodextrin, or chemically modified derivatives such as
hydroxyalkylcyclodextrins, including 2- and
3-hydroxypropyl-.beta.-cyclodextrins, or other solubilized
derivatives may also be advantageously used to enhance delivery of
compounds of the formulae described herein.
[0432] The pharmaceutical compositions of this invention may be
administered orally, parenterally, by inhalation spray, topically,
rectally, nasally, buccally, vaginally or via an implanted
reservoir, preferably by oral administration or administration by
injection. The pharmaceutical compositions of this invention may
contain any conventional non-toxic pharmaceutically-acceptable
carriers, adjuvants or vehicles. In some cases, the pH of the
formulation may be adjusted with pharmaceutically acceptable acids,
bases or buffers to enhance the stability of the formulated
compound or its delivery form. The term parenteral as used herein
includes subcutaneous, intracutaneous, intravenous, intramuscular,
intraarticular, intraarterial, intrasynovial, intrasternal,
intrathecal, intralesional and intracranial injection or infusion
techniques.
[0433] The pharmaceutical compositions may be in the form of a
sterile injectable preparation, for example, as 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, or carboxymethyl cellulose or similar dispersing agents
which are commonly used in the formulation of pharmaceutically
acceptable dosage forms such as emulsions and or suspensions. Other
commonly used surfactants such as Tweens or Spans and/or other
similar emulsifying agents or bioavailability enhancers which are
commonly used in the manufacture of pharmaceutically acceptable
solid, liquid, or other dosage forms may also be used for the
purposes of formulation.
[0434] The pharmaceutical compositions of this invention may be
orally administered in any orally acceptable dosage form including,
but not limited to, capsules, tablets, emulsions and aqueous
suspensions, dispersions and solutions. In the case of tablets for
oral use, carriers which 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 corn starch. When aqueous
suspensions and/or emulsions are administered orally, the active
ingredient may be suspended or dissolved in an oily phase is
combined with emulsifying and/or suspending agents. If desired,
certain sweetening and/or flavoring and/or coloring agents may be
added.
[0435] The pharmaceutical compositions of this invention may also
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.
[0436] Topical administration of the pharmaceutical compositions of
this invention is useful when the desired treatment involves areas
or organs readily accessible by topical application. For
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 with suitable emulsifying agents. 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 are also
included in this invention.
[0437] 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.
[0438] When the compositions of this invention comprise a
combination of a compound of the formulae described herein and one
or more additional therapeutic or prophylactic agents, both the
compound and the additional agent should be present at dosage
levels of between about 1 to 100%, and more preferably between
about 5 to 95% of the dosage normally administered in a monotherapy
regimen. The additional agents may be administered separately, as
part of a multiple dose regimen, from the compounds of this
invention. Alternatively, those agents may be part of a single
dosage form, mixed together with the compounds of this invention in
a single composition.
[0439] The compounds described herein can, for example, be
administered by injection, intravenously, intraarterially,
subdermally, intraperitoneally, intramuscularly, or subcutaneously;
or orally, buccally, nasally, transmucosally, topically, in an
ophthalmic preparation, or by inhalation, with a dosage ranging
from about 0.5 to about 100 mg/kg of body weight, alternatively
dosages between 1 mg and 1000 mg/dose, every 4 to 120 hours, or
according to the requirements of the particular drug. The methods
herein contemplate administration of an effective amount of
compound or compound composition to achieve the desired or stated
effect. Typically, the pharmaceutical compositions of this
invention will be administered from about 1 to about 6 times per
day or alternatively, as a continuous infusion. Such administration
can be used as a chronic or acute therapy. The amount of active
ingredient that may be combined with the carrier materials to
produce a single dosage form will vary depending upon the host
treated and the particular mode of administration. A typical
preparation will contain from about 5% to about 95% active compound
(w/w). Alternatively, such preparations contain from about 20% to
about 80% active compound.
[0440] Lower or higher doses than those recited above may be
required. Specific dosage and treatment regimens for any particular
patient will depend upon a variety of factors, including the
activity of the specific compound employed, the age, body weight,
general health status, sex, diet, time of administration, rate of
excretion, drug combination, the severity and course of the
disease, condition or symptoms, the patient's disposition to the
disease, condition or symptoms, and the judgment of the treating
physician.
[0441] Upon improvement of a patient's condition, a maintenance
dose of a compound, composition or combination of this invention
may be administered, if necessary. Subsequently, the dosage or
frequency of administration, or both, may be reduced, as a function
of the symptoms, to a level at which the improved condition is
retained when the symptoms have been alleviated to the desired
level. Patients may, however, require intermittent treatment on a
long-term basis upon any recurrence of disease symptoms.
Patient Selection and Monitoring
[0442] The compounds described herein can modulate PKM2.
Accordingly, a patient and/or subject can be selected for treatment
using a compound described herein by first evaluating the patient
and/or subject to determine whether the subject is in need of
modulation of PKM2, and if the subject is determined to be in need
of modulation of PKM2, then optionally administering to the subject
a compound described herein.
[0443] A subject can be evaluated as being in need of modulation of
PKM2 using methods known in the art, e.g., by measuring the
presence and/or activity of PKM2 in the patient. In some
embodiments, the activity and/or level of PKM2 is evaluated in the
cancer.
[0444] A patient receiving a compound described herein can be
monitored, for example, for improvement in the condition and/or
adverse effects. Improvement of a patient's condition can be
evaluated, for example, by monitoring the growth, absence of
growth, or regression of the cancer (e.g., a tumor). In some
embodiments, the patient is evaluated using a radiological assay or
evaluation of hemolytic parameters.
Examples
Example 1
PKM2 Assay
Procedure:
[0445] PKM2 stock enzyme solution was diluted in Reaction Buffer
[0446] 2 .mu.L of compound was added into each well first, and then
180 .mu.L of the Reaction Mix was added. [0447] Reaction mixture
with compound (without ADP) were incubated for 30 minutes at
4.degree. C. [0448] Plates were re-equilibrated to room temperature
prior to adding 20 .mu.L ADP to initiate the reaction. [0449]
Reaction progress was measured as changes in absorbance at 340 nm
wavelength at room temperature (25.degree. C.)
Reaction Mix:
[0450] PKM2 (50 ng/well), ADP (0.7 mM), PEP (0.15 mM), NADH (180
.mu.M), LDH (2 units) in Reaction Buffer
Reaction Buffer:
[0451] 100 mM KCl, 50 mM Tris pH 7.5, 5 mM MgCl2, 1 mM DTT, 0.03%
BSA.
Example 2
PKM2 Ex-Vivo Assay
[0452] Described herein is a method to measure the activity of PKM2
activators in living cells and tissue. One of ordinary skill in the
art would recognize and understand that this method can be adapted
to high throughput format, and can accommodate a variety of cell
lines and growth conditions.
[0453] In the assay, cells are treated with a compound described
herein (i.e., a PKM2 activator). This compound is capable of
entering the cell and binding to PKM2, inducing an activated
conformation. The excess unbound compound is washed away with PBS,
and the cells are lysed by snap-freezing on dry ice, followed by
addition of a detergent-containing lysis buffer. The lysate, in
which activated PKM2 remains intact, is removed and added to a
chemical cocktail including the chemicals necessary to measure
pyruvate kinase activity, in an assay that is coupled to the LDHa
enzyme. The amount of pyruvate kinase activity that is measured is
normalized to the total protein content in the lysate, and related
to the concentration of PKM2 activator that was added to the cell.
This allows an AC.sub.50 (concentration at which PKM2 is activated
50%) value to be derived. The total fold-increase in activity over
mock-treated cells can also be calculated, and the "maximum level
of activation" can be used to distinguish between compounds that
fully activate PKM2 and compounds that can only partially activate
PKM2.
[0454] In the case of measuring PKM2 activity from tissue (for
example, in a cell tumor), animals harboring the tissue/tumor of
interest are dosed with a compound. After a specified period of
time in which exposure has been achieved in the target tissue/tumor
of interest, the tissue/tumor is harvested from the animal,
snap-frozen, and then lysed and homogenized. The amount of pyruvate
kinase activity in this lysate can then be quantitated as described
above.
Materials:
TABLE-US-00003 [0455] Lysis buffer* 20 mM Tris-HCl (pH 7.5) 150 mM
NaCl 1 mM Na.sub.2EDTA 1 mM EGTA 1% Triton 2.5 mM sodium
pyrophosphate 1 mM beta-glycerophosphate 1 mM Na.sub.3VO.sub.4 1
.mu.g/ml leupeptin 1 mM PMSF** *This lysis buffer (without PMSF) is
available from Cell Signaling Technology as a 10x stock (#9803) **1
mM PMSF is added fresh from a 100 mM stock solution made up in
isopropanol. The stock solution can be stored at 4 degrees
indefinitely.
Pyruvate Kinase Assay Master Mix (Same for PKM2 Activator
Assay):
TABLE-US-00004 [0456] TABLE 3 KCl 100 mM Tris (pH 7.5) 50 mM
MgCl.sub.2 5.0 mM PEP 0.10 mM NADH 0.18 mM DTT 1.00 mM BSA 0.3
mg/mL LDH 0.5 units H.sub.2O to 180 uL ADP solution: ADP 7.0 mM
H.sub.2O to 20 uL
Procedure:
[0457] On the first day (day 1) cells are normally cultured in
RPMI-1640 (Lonza #12-115.degree. F.) (with 25 mM Hepes,
L-glutamine)/10% FBS. The cells are subsequently trypsinized and
plated in RPMI-1640 (Lonza, #12-918F) (no phenol red, supplemented
with L-glutamine 300 mg/L (Sigma, #G8540))/10% FBS at the following
densities in 96 well plates:
[0458] A549: 40 k/well
[0459] 100 uL final volume of media per well.
[0460] On the second day (Day 2), the cells should be 70-90%
confluent. The cells are then treated with a compound described
herein dissolved in media at final assay concentrations in a
96-well assay block (500 uL) (Costar, #3956). The final DMSO
concentration is 0.1% (0.5 L into 500 uL). Compound dilutions in
DMSO are prepared so that the final DMSO concentration is constant
at all compound concentrations. The media for the assay is
RPMI-1640 (no phenol red, with L-glutamine 300 mg/L).
[0461] The media is then aspirated carefully from the cells using a
multi-channel aspirator. 100 L of media w/compounds is added onto
cells with a multichannel pipette. Each compound concentration is
then assayed in triplicate (a duplicate assay is also
sufficient).
[0462] The cells are treated for 1-4 hrs (this time is determined
empirically compared to DMSO reference treatment). During the cell
treatment, PBS (containing calcium and magnesium) and lysis buffer
is cooled on ice.
[0463] The cells are lysed and the pyruvate kinase activity is
assayed. The remaining media is aspirated and the cells are washed
2.times. with 100 uL ice-cold PBS. The PBS is removed, and the cell
plate frozen on dry ice for 5 minutes. The cells are lysed in 50 L
cold lysis buffer. Cells are subsequently kept on ice for 5
minutes, and then agitated on a plate shaker for 5 minutes (repeat
3.times.). Remove 10 L for protein quantitation (or use OD.sub.280
on entire plate).
[0464] In a fresh plate, 170 uL of pyruvate kinase assay master mix
was added to each well (see end for recipe). 10 uL of cell lysate
was then transferred into each well. The assay was initiated upon
addition of 20 uL of ADP solution. The rates were then calculated
against the initial rates to determine pyruvate kinase specific
activity.
[0465] The concentration and type of detergent in the lysis buffer
can be varied to accommodate the specific physicochemical
properties of the specific PKM2 activator. For instance, the
interaction between some PKM2 activators and PKM2 can be disrupted
by higher detergent concentrations, but preserved when cells are
lysed with lower detergent concentrations.
Example 3
Compounds and Synthesis
##STR00207##
[0467] General Procedure for Compound 2:
[0468] Chlorosulfonic acid (24 mL) was added slowly over a period
of 15 min to imidazole (4.0 g, 58.82 mmol) at 0.degree. C. The
resulting mixture was heated at 120-130.degree. C. for 3 h. After
completion of SM, the reaction mixture was quenched with ice cold
water (25 mL) and extracted with DCM (3.times.40 mL). The combined
organic layers were dried over Na.sub.2SO.sub.4 and concentrated
under reduced pressure. The desired Compound-2 was obtained as a
solid (3.0 g, 42% yield).
[0469] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 14.29 (bs, 1H),
8.90 (s, 1H), 7.64 (s, 1H); Mass (M+1): 166.9.
[0470] General Procedure for Compound 4:
[0471] To a solution of compound-3 (500 mg, 4.13 mmol) in DCM (10
mL) under a nitrogen atmosphere, pyridine (0.65 ml, 8.26 mmol) was
added and stirred at room temperature for 15 min. The reaction
mixture was then cooled to 0.degree. C. followed by the dropwise
addition of Compound-2 (820 mg, 4.96 mmol) in DCM (4 mL). The
resulting reaction mixture was stirred for 2 h at room temperature.
After completion of the reaction, 0.5N HCl solution was added and
extracted with DCM (2.times.10 mL). The combined organic layers
were washed with brine (1.times.20 mL), dried over Na.sub.2SO.sub.4
and concentrated under reduced pressure to provide the desired
compound 4 as a solid (500 mg, 50% yield).
[0472] General Procedure for Compound 6:
[0473] In a two neck RB flask, compound-4 (50 mg, 0.2 mmoL),
compound-5 (51 mg, 0.24 mmoL) and K.sub.2CO.sub.3 (54 mg, 0.4 mmoL)
were charged in DMF (6 mL) under N.sub.2 atmosphere. The resulting
reaction mixture was stirred at 90.degree. C. for 3 h. The progress
of the reaction was monitored by TLC. After completion of SM, the
reaction mixture was quenched with water (15 mL) and extracted with
ethyl acetate (2.times.20 mL). The combined organic layers were
dried over Na.sub.2SO.sub.4 and concentrated under reduced
pressure. The crude product was purified by column chromatography
(Silica gel 60-120, 3:7, ethyl acetate/hexane) to obtain the
desired compound-6 as a solid (40 mg, 47% yield).
[0474] .sup.1H NMR (500 MHz, DMSO-D6) .delta.: 10.29 (s, 1H), 9.03
(s, 1H), 8.84 (s, 1H), 8.37 (s, 1H), 8.32 (s, 1H), 7.82 (s, 2H),
6.63 (s, 1H), 2.18 (s, 6H); HPLC Purity: 93.13%; Mass (M+1):
431.1.
1-(3-chloro-5-(trifluromethyl)pyridine-2-yl)-N-(3,5-dimethylphenyl)-1H-imi-
dazole-5-sulfonamide
##STR00208##
[0476] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 10.29 (s, 1H),
9.03 (s, 1H), 8.84 (s, 1H), 8.37 (s, 1H), 8.32 (s, 1H), 7.82 (s,
2H), 6.63 (s, 1H), 2.18 (s, 6H); HPLC Purity: 93.13%; Mass (M+1):
431.1.
1-(3-chloro-5-(trifluromethyl)pyridine-2-yl)-N-(4-methoxyphenyl)-1H-imidaz-
ole-5-sulfonamide
##STR00209##
[0478] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 10.04 (bs, 1H),
9.01 (s, 1H), 8.83 (s, 1H), 8.38 (s, 1H), 8.18 (s, 1H), 7.08 (d,
2H), 6.82 (d, 2H), 3.67 (s, 3H); HPLC Purity: 91.04%; Mass (M+1):
433.
N-(4-methoxyphenyl)-1-5-(trifluromethyl)pyridine-2-yl)-1H-imidazole-5-sulf-
onamide
##STR00210##
[0480] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 10.11 (s, 1H),
8.97 (s, 1H), 8.81 (s, 1H), 8.52 (m, 2H), 8.19 (d, 1H), 7.48 (d,
2H), 6.82 (d, 2H), 3.65 (s, 3H); HPLC Purity: 96.12%; Mass (M+1):
399.
N-(2,3-dihydrobenzo[b][1,4]dioxin-6-yl)-1-5-(trifluromethyl)pyridine-2-yl)-
-1H-imidazole-5-sulfonamide
##STR00211##
[0482] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 10.53 (s, 1H),
8.96 (s, 1H), 8.79 (s, 1H), 8.58 (s, 1H), 8.56 (s, 1H), 8.18 (d,
1H), 6.66 (s, 2H), 6.63 (d, 1H) 4.17 (t, 4H); HPLC Purity: 98.48%;
Mass (M+1): 427.
1-(3-chloro-5-(trifluromethyl)pyridin-2-yl)-N-(2,3-dihydrobenzo[b][1,4]dio-
xin-6-yl)-1H-imidazole-5-sulfonamide
##STR00212##
[0484] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 10.13 (s, 1H),
9.02 (s, 1H), 8.84 (s, 1H), 8.39 (s, 1H), 8.22 (s, 1H), 6.78-6.58
(m, 3H), 4.18 (s, 4H); HPLC Purity: 96.98%; Mass (M+1): 461.
N-(4-flurophenyl)-1-(5-(trifluromethyl)pyridine-2-yl)-1H-imidazole-5-sulfo-
namide
##STR00213##
[0486] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 10.42 (s, 1H),
8.97 (s, 1H), 8.80 (s, 1H), 8.62 (s, 1H), 8.46 (d, 1H), 8.19 (d,
1H), 7.23 (d, 2H), 7.08 (d, 2H); HPLC Purity: 99.35%; Mass (M+1):
387.
1-(3-chloro-5-(trifluromethyl)pyridin-2-yl)-N-(4-flurophenyl)-1H-imidazole-
-5-sulfonamide
##STR00214##
[0488] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 10.42 (s, 1H),
9.02 (s, 1H), 8.84 (s, 1H), 8.39 (s, 1H), 8.24 (s, 1H), 7.22 (d,
2H), 7.08 (d, 2H); HPLC Purity: 92.78%; Mass (M+1): 421.
N-(3,5-dimethylphenyl)-1-(5-(trifluromethyl)pyridine-2-yl)-1H-imidazole-5--
sulfonamide
##STR00215##
[0490] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 10.28 (s, 1H),
8.96 (s, 1H), 8.78 (s, 1H), 8.63 (s, 1H), 8.33 (d, 1H), 8.19 (d,
1H), 7.82 (s, 2H), 7.62 (s, 1H), 2.18 (s, 6H); HPLC Purity: 99.23%;
Mass (M+1): 397.1.
1-(3-chloro-5-(trifluromethyl)pyridin-2-yl)-N-(3,5-dimethoxyphenyl)-1H-imi-
dazole-5-sulfonamide
##STR00216##
[0492] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 10.41 (s, 1H),
9.02 (s, 1H), 8.87 (s, 1H), 8.38 (s, 2H), 6.39 (s, 2H), 6.17 (s,
1H), 3.67 (s, 6H); HPLC Purity: 97.82%; Mass (M+1): 463.1.
N-(4-chloro-3-methylphenyl)-1-(3-chloro-(5-(trifluromethyl)pyridine-2-yl)--
1H-imidazole-5-sulfonamide
##STR00217##
[0494] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 10.53 (s, 1H),
9.03 (s, 1H), 8.84 (s, 1H), 8.37 (d, 2H), 7.28 (d, 1H), 7.16 (s,
1H), 7.03 (d, 1H), 2.22 (s, 3H); HPLC Purity: 98.76%; Mass (M+1):
451.
1-(3-chloro-5-(trifluromethyl)pyridin-2-yl)-N-(3,4-dimethylphenyl)-1H-imid-
azole-5-sulfonamide
##STR00218##
[0496] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 10.21 (s, 1H),
9.02 (s, 1H), 8.85 (s, 1H), 8.38 (s, 2H), 8.24 (s, 1H), 6.98-6.87
(m, 3H), 2.13 (s, 6H); HPLC Purity: 99.06%; Mass (M+1): 431.
1-(3-chloro-5-(trifluromethyl)pyridin-2-yl)-N-(3-chlorophenyl)-1H-imidazol-
e-5-sulfonamide
##STR00219##
[0498] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 10.78 (s, 1H),
9.03 (s, 1H), 8.84 (s, 1H), 8.38 (d, 2H), 7.31-7.24 (m, 2H), 7.16
(d, 1H), 7.08 (d, 1H); HPLC Purity: 95.26%; Mass (M+1): 436.9.
1-(3-chloro-5-(trifluromethyl)pyridin-2-yl)-N-(3-(trifluromethyl)phenyl)-1-
H-imidazole-5-sulfonamide
##STR00220##
[0500] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 10.96 (s, 1H),
9.02 (s, 1H), 8.87 (s, 1H), 8.38 (d, 2H), 7.52-7.38 (m, 4H); HPLC
Purity: 90.31%; Mass (M+1): 471.1.
1-(3-chloro-5-(trifluromethyl)pyridin-2-yl)-N-(3,5-dichlorophenyl)-1H-imid-
azole-5-sulfonamide
##STR00221##
[0502] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 11.03 (s, 1H),
9.03 (s, 1H), 8.49 (s, 1H), 8.39 (s, 1H), 7.24-7.21 (m, 4H); HPLC
Purity: 97.52%; Mass (M+1): 472.8.
1-(3-chloro-5-(trifluromethyl)pyridin-2-yl)-N-(3-ethylphenyl)-1H-imidazole-
-5-sulfonamide
##STR00222##
[0504] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 10.38 (s, 1H),
9.02 (s, 1H), 8.84 (s, 1H), 8.37 (d, 2H), 7.17 (m, 1H), 7.01 (m,
2H), 6.86 (d, 1H), 2.52 (q, 2H), 1.13 (t, 3H); HPLC Purity: 93.13%;
Mass (M+1): 431.1.
1-(3-chloro-5-(trifluromethyl)pyridin-2-yl)-N-(3-cyanophenyl)-1H-imidazole-
-5-sulfonamide
##STR00223##
[0506] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 10.99 (s, 1H),
9.04 (s, 1H), 8.87 (s, 1H), 8.44 (s, 1H), 8.38 (s, 1H), 7.51 (m,
4H); HPLC Purity: 90.19%; Mass (M+1): 428.1.
##STR00224##
[0507] General Procedure for Compound 3&3':
[0508] In a two neck RB flask, compound-2 (3.0 g, 0.02 mole),
pyrrole (2.69 g, 0.04 mole) and CS.sub.2CO.sub.3 (19.68 g, 0.06
mole) were charged in toluene (60 mL) under N.sub.2 atmosphere.
BINAP (620 mg, 0.00099 mole) and Pd(OAc).sub.2 (990 mg, 0.004 mole)
were then added to the reaction mixture under N.sub.2 atmosphere
and stirred at 80 C for 3 h. The progress of the reaction was
monitored by TLC. After completion of SM, the reaction mixture was
filtered through celite and quenched with water (5 mL). The organic
layer was separated, dried over Na.sub.2SO.sub.4 and concentrated
under reduced pressure. The crude product was subjected to silica
gel column chromatography (silica gel 60-120, 1:9, ethyl
acetate/hexane) to separate the sulfonyl chlorides 3 & 3' as
viscous oil (compound-3: 1.1 g 30.55% and compound-3': 684 mg
19.0%). Overall 49.55% yield obtained.
[0509] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 8.53 (d, 1H),
7.54 (d, 2H), 7.15 (d, 1H), 6.41 (d, 2H); Mass (M+1): 180.
[0510] General Procedure for Compound 4:
[0511] In a single neck RB flask, to compound-3 (100 mg, 0.59 mmol)
was added chlorosulfonic acid (690 mg, 5.9 mmol) slowly over a
period of 15 min at 0.degree. C. The resulting mixture was stirred
for 3 h at room temperature. After completion of SM, the reaction
mixture was quenched with water (3 mL) and extracted with DCM
(2.times.5 mL). The combined organic layers were dried over
Na.sub.2SO.sub.4 and concentrated under reduced pressure. The crude
compound was washed with pentane and dried under vacuum to provide
the desired compound-4 as a solid (60 mg, 38.7%).
[0512] General Procedure for Compound 6:
[0513] In a two neck RB flask, compound-5 (9.5 mg, 0.16 mmol) and
pyridine (0.15 mL) were taken and stirred for 10 minutes at
0.degree. C. Compound-4 (30 mg, 0.1 mmol) was then added at
0.degree. C. and the resulting mixture was stirred for 3 h at room
temperature. After completion of the reaction, HCl solution (6N, 4
mL) was added and extracted with DCM (2.times.5 mL). The combined
organic layers were washed with brine (1.times.20 mL), dried over
anhydrous Na.sub.2SO.sub.4 and concentrated under reduced pressure.
The crude product was purified by column chromatography (silica gel
60-120, 2:8, ethyl acetate/hexane) to provide the desired
compound-6 as a solid (20.4 mg, 68.1%).
[0514] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 8.81 (s, 1H),
8.19 (s, 1H), 7.97 (s, 1H), 7.81 (s, 1H), 6.81 (s, 2H), 6.69 (s,
1H), 6.58 (s, 1H), 2.19 (s, 6H); HPLC Purity: 92.07%; Mass (M+1):
362.9.
[0515] General Procedure for Compound 7:
[0516] In a single neck RB flask, to compound-3' (100 mg, 0.59
mmol) was added chlorosulfonic acid (690 mg, 5.9 mmol) over a
period of 15 min at 0.degree. C. The resulting mixture was then
stirred for 3 h at room temperature. Upon completion of SM (as
indicated by TLC), the reaction mixture was quenched with water (3
mL) and extracted with DCM (2.times.5 mL). The combined organic
layers were dried over Na.sub.2SO.sub.4 and concentrated under
reduced pressure. The crude compound was washed with pentane and
dried under vacuum to provide the desired compound-7 as a solid (35
mg, 22% yield).
[0517] General Procedure for Compound 8:
[0518] In a two neck RB flask, compound-5 (14.9 mg, 0.12 mmol) and
pyridine (0.1 mL) were transferred and stirred for 10 minutes at
0.degree. C. Compound-7 (20 mg, 0.08 mmol) was then added at
0.degree. C. The resulting mixture was stirred for 3 h at room
temperature. After completion of the reaction, HCl (6N, 4 mL) was
added and extracted with DCM (2.times.5 mL). The combined organic
layers were washed with brine (1.times.20 mL), dried over
Na.sub.2SO.sub.4 and concentrated under reduced pressure. The crude
product was purified by column chromatography (silica gel 60-120,
2:8, ethyl acetate/hexane) to provide the desired compound-8 as a
solid (18.62 mg, 71% yield).
[0519] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 10.0 (s, 1H),
8.81 (s, 1H), 8.01 (s, 1H), 7.78 (s, 1H), 7.63 (s, 1H), 6.82 (s,
2H), 6.68 (s, 1H), 6.54 (s, 1H), 2.19 (s, 6H); HPLC Purity: 98.01%;
Mass (M+1): 363.
1-(4-chloropyrimidin-2-yl)-N-(3,5-dimethylphenyl)-1H-pyrrole-2-sulfonamide
##STR00225##
[0521] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 10.0 (s, 1H),
8.81 (s, 1H), 8.01 (s, 1H), 7.78 (s, 1H), 7.63 (s, 1H), 6.82 (s,
2H), 6.68 (s, 1H), 6.54 (s, 1H), 2.19 (s, 6H); HPLC Purity: 98.01%;
Mass (M+1): 363.
1-(2-chloropyrimidin-4-yl)-N-(3,5-dimethylphenyl)-1H-pyrrole-2-sulfonamide
##STR00226##
[0523] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 8.81 (s, 1H),
8.19 (s, 1H), 7.97 (s, 1H), 7.81 (s, 1H), 6.81 (s, 2H), 6.69 (s,
1H), 6.58 (s, 1H), 2.19 (s, 6H); HPLC Purity: 92.07%; Mass (M+1):
362.9.
N-(4-chlorophenyl)-1-(2-chloropyrimidin-2-yl)-1H-pyrrole-2-sulfonamide
##STR00227##
[0525] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 10.34 (s, 1H),
8.82 (s, 1H), 8.23 (s, 1H), 8.07 (s, 1H), 7.37 (d, 2H), 7.08 (d,
2H), 6.57 (s, 1H); HPLC Purity: 96.74%; Mass (M+1): 369.
N-(4-chlorophenyl)-1-(4-chloropyrimidin-2-yl)-1H-pyrrole-2-sulfonamide
##STR00228##
[0527] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 10.29 (s, 1H),
8.90 (s, 1H), 8.02 (s, 1H), 7.77 (s, 1H), 7.64 (s, 1H), 7.33 (d,
2H), 7.18 (d, 2H), 6.54 (s, 1H); HPLC Purity: 91.17%; Mass (M+1):
368.9.
N-(4-chlorophenyl)-1-(pyrimidin-2-yl)-1H-pyrrole-2-sulfonamide
##STR00229##
[0529] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 10.27 (s, 1H),
8.83 (d, 2H), 8.07 (s, 1H), 7.79 (s, 1H), 7.48 (d, 1H), 7.32 (d,
2H), 7.18 (d, 2H), 6.53 (s, 1H); HPLC Purity: 97.05%; Mass (M+1):
334.9.
N-(3,5-dimethylphenyl)-1-(pyrimidin-2-yl)-1H-pyrrole-2-sulfonamide
##STR00230##
[0531] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 9.88 (s, 1H),
8.84 (d, 2H), 8.04 (s, 1H), 7.78 (s, 1H), 7.43 (d, 1H), 6.79 (s,
2H), 6.63 (s, 1H), 6.49 (s, 1H); HPLC Purity: 97.48%; Mass (M+1):
329.
##STR00231##
[0532] General Procedure for Compound 2:
[0533] In a single neck RB flask chlorosulfonic acid (9.7 ml, 145
mmol) was added slowly over a period of 15 minutes to pyrazole (2.0
g, 29.0 mmol) at 0.degree. C. The resulting mixture was then heated
at 100.degree. C. for 3 h. Upon completion, the reaction mixture
was quenched with ice cold water (25 mL) and extracted with DCM
(3.times.40 mL). The combined organic layers were dried over
Na.sub.2SO.sub.4 and concentrated under reduced pressure. The
desired product 2 was obtained as a solid (4.8 g, 63% yield).
[0534] .sup.1H NMR (500 MHz, CDCl3) .delta.: 8.19 (s, 2H).
[0535] General Procedure for Compound 3:
[0536] To a stirred solution of compound-2 (60 mg, 0.496 mmol) in
DCM (10 mL) under a nitrogen atmosphere, pyridine was added (0.1
mL, 0.99 mmol) and stirred at room temperature for 15 minutes. The
reaction mixture was then cooled to 0.degree. C. followed by the
dropwise addition of compound-2 solution (91.1 mg, 0.545 mmol in 4
mL of DCM). The resulting mixture was stirred for 2 h at room
temperature. After completion of the reaction, 0.5N HCl solution
was added and extracted with DCM (2.times.10 mL). The combined
organic layers were washed with brine (1.times.20 mL), dried over
Na.sub.2SO.sub.4 and concentrated under reduced pressure to furnish
the desired compound-4 as a solid (120 mg, 97% yield).
[0537] .sup.1H NMR (500 MHz, CDCl.sub.3) .delta.: 10.4 (bs, 1H),
7.81 (s, 2H), 6.79 (s, 1H), 6.72 (s, 2H), 6.47 (s, 1H), 2.23 (s,
6H); Mass (M+1): 252.
[0538] General Procedure for Compound 4:
[0539] In a two neck RB flask, compound-3 (50 mg, 0.2 mmol),
compound-C (0.03 ml, 0.29 mmol) and K.sub.2CO.sub.3 (83 mg, 0.6
mmol) were charged in DMF (5 mL) under N.sub.2 atmosphere and
stirred at 90.degree. C. for 3 h. The progress of the reaction was
monitored by TLC. After completion of SM, the reaction mixture was
quenched with water (15 mL) and extracted with ethyl acetate
(2.times.20 mL). The combined organic layers were dried over
Na.sub.2SO.sub.4 and concentrated under reduced pressure. The crude
product was purified by column chromatography (Silica gel 60-120,
3:7, ethyl acetate/hexane) to give the desired product 4 as a solid
(70 mg, 81.3% yield).
1-(3-chloro-5-(trifluromethyl)pyridin-2-yl)-N-(3,5-dimethylphenyl)-1H-pyra-
zole-4-sulfonamide
##STR00232##
[0541] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 10.10 (s, 1H),
9.01 (s, 1H), 8.82 (s, 1H), 8.75 (s, 1H), 8.15 (s, 1H), 6.81 (s,
2H), 6.77 (s, 1H), 2.19 (s, 6H); HPLC Purity: 94.73%; Mass (M+1):
431.1.
N-(4-methoxyphenyl)-1-(5-(trifluromethyl)pyridin-2-yl)-1H-pyrazole-4-sulfo-
namide
##STR00233##
[0543] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 9.98 (s, 1H),
8.97 (s, 1H), 8.81 (s, 1H), 8.45 (d, 1H), 8.10 (d, 1H), 8.05 (s,
1H), 7.07 (d, 2H), 6.85 (d, 2H), 3.68 (s, 3H); HPLC Purity:
99.35%.
N-(4-chlorophenyl)-1-(5-(trifluromethyl)pyridin-2-yl)-1H-pyrazole-4-sulfon-
amide
##STR00234##
[0545] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 10.51 (s, 1H),
8.97 (s, 2H), 8.46 (d, 1H), 8.18 (s, 1H), 8.12 (d, 1H), 7.38 (d,
2H), 7.21 (d, 2H); HPLC Purity: 99.60%; Mass (M+1): 403.
1-(3-chloro-5-(trifluromethyl)pyridin-2-yl)-N-(4-chlorophenyl)-1H-pyrazole-
-4-sulfonamide
##STR00235##
[0547] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 10.49 (s, 1H),
9.01 (s, 1H), 8.83 (s, 1H), 8.78 (s, 1H), 8.13 (s, 1H), 7.38 (d,
2H), 7.19 (d, 2H); HPLC Purity: 99.52%; Mass (M+1): 437.
1-(3-chloro-5-(trifluromethyl)pyridin-2-yl)-N-(4-methoxyphenyl)-1H-pyrazol-
e-4-sulfonamide
##STR00236##
[0549] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 9.98 (s, 1H),
9.01 (s, 1H), 8.81 (s, 1H), 8.64 (s, 1H), 8.02 (s, 1H), 7.07 (d,
2H), 6.84 (d, 2H), 3.68 (s, 3H); HPLC Purity: 99.35%; Mass (M+1):
432.9.
##STR00237##
[0550] General Procedure for Preparation of Compound a:
[0551] A solution of mixture of triethyl orthoformate (11.2 g, 75
mmol) and malononitrile (5.0 g, 75 mmol) in acetic anhydride (15
mL) was heated to 90.degree. C. for 12 h. After completion of
reaction (TLC shows absent of S.M), the solvent (excess acetic
anhydride and acetic acid) was removed under high vacuum. The
residue (6.0 g, 65.2% of yield) was used in the next reaction
without further purification.
[0552] .sup.1H NMR (500 MHz, DMSO-d6) .delta.: 8.59 (s, 1H), 4.55
(m, 2H), 1.38 (m, 3H).
[0553] General Procedure for Compound 2:
[0554] To a stirred solution of 2-bromo-5-(trifluoromethyl)pyridine
(compound-1) (1.0 g, 4.0 mmol) in ethanol (5 mL), hydrazine hydrate
(980 mg, 20 mmol) was added at room temperature and the reaction
mixture was heated to 90.degree. C. for 12 h. After completion of
reaction (TLC shows absent of S.M), the solvent was removed under
high vacuum. The residue was treated with water (30 mL) and
extracted with ethyl acetate (2.times.50 mL). The organic layer was
washed with brine (50 mL), dried over sodium sulfate, filtered and
concentrated to provide the desired compound-2 as a brown color
solid. (700 mg, 90% yield).
[0555] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 8.42 (d, 2H),
8.25 (s, 1H), 5.0 (bs, 3H); Mass (M+1): 178.
[0556] General Procedure for Compound 3:
[0557] A solution of 2-hydrazinyl-5-(trifluoromethyl)pyridine
(compound-2) (250 mg, 1.4 mmol) and
2-(ethoxymethylene)malononitrile (compound-A) (170 mg, 1.4 mmol) in
ethanol (10 mL) was heated to 90.degree. C. for 12 h. After
completion of reaction (TLC shows absent of S.M), the solvent was
removed under high vacuum. The residue was treated with water (30
mL) and extracted with ethyl acetate (2.times.50 mL). The organic
layer was washed with brine (50 mL), dried over sodium sulphate,
filtered and concentrated to provide the desired compound-3 as an
off white color solid. (280 mg, 80% yield).
[0558] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 8.91 (s, 1H),
8.45 (d, 1H), 8.32 (s, 2H), 8.15 (d, 1H), 8.0 (s, 1H); Mass (M+1):
254.
[0559] General Procedure for Compound 4:
[0560] In a two neck RB flask, compound-3 (50 mg, 0.1 mmoL) was
taken in THF (4 mL) and cooled to 0.degree. C. under nitrogen
atmosphere. NaH (60% NaH, 5 mg, 0.1 mmol) was added to the reaction
mixture and stirred for 10 min followed by the addition of solution
of 3,5-dimethylbenzene-1-sulphonyl chloride (44 mg, 0.2 mmoL) in
THF (2 mL) at 0.degree. C. The reaction mixture was then stirred
for 30 min. at room temperature. After completion of the reaction,
the reaction mixture was quenched with ice cold water and extracted
with ethyl acetate, and the organic layer was washed with brine (10
mL), dried over sodium sulphate, filtered and concentrated to
provide the desired compound-4 as a solid (60 mg, 81.2% yield).
[0561] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 8.83 (s, 1H),
8.28 (d, 1H), 8.17 (d, 1H), 7.77 (s, 1H), 7.31 (s, 2H), 7.01 (s,
1H), 2.21 (s, 6H); HPLC Purity: 98.81%; Mass (M+1): 422.
N-(4-cyano-1-(5-(trifluromethyl)pyridin-2-yl)-1H-pyrazole-3,5
dimethylbenzenesulfonamide
##STR00238##
[0563] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 8.83 (s, 1H),
8.28 (d, 1H), 8.17 (d, 1H), 7.77 (s, 1H), 7.31 (s, 2H), 7.01 (s,
1H), 2.21 (s, 6H); HPLC Purity: 98.81%; Mass (M+1): 422.
4-chloro-N-(4-cyano-1-(5-(trifluromethyl)pyridin-2-yl)-1H-pyrazol-5-yl)ben-
zenesulfonamide
##STR00239##
[0565] .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.: 8.88 (s, 1H),
8.31 (d, 1H), 8.14 (bs, 1H), 7.73 (m, 3H), 7.41 (m, 2H); HPLC
Purity: 97.80%; Mass (M+1): 428.
[0566] Having thus described several aspects of at least one
embodiment of this invention, it is to be appreciated various
alterations, modifications, and improvements will readily occur to
those skilled in the art. Such alterations, modifications, and
improvements are intended to be part of this disclosure, and are
intended to be within the spirit and scope of the invention.
Accordingly, the foregoing description and drawings are by way of
example only.
[0567] Having thus described several aspects of at least one
embodiment of this invention, it is to be appreciated various
alterations, modifications, and improvements will readily occur to
those skilled in the art. Such alterations, modifications, and
improvements are intended to be part of this disclosure, and are
intended to be within the spirit and scope of the invention.
Accordingly, the foregoing description and drawings are by way of
example only.
* * * * *