U.S. patent application number 16/462779 was filed with the patent office on 2019-12-05 for cyclic phosphate substituted nucleoside derivatives for the treatment of liver diseases.
This patent application is currently assigned to IDENIX PHARMACEUTICALS LLC. The applicant listed for this patent is IDENIX PHARMACEUTICALS LLC. Invention is credited to Stephane L. Bogen, Cyril Dousson, David Dukhan, Christophe C. Parsy.
Application Number | 20190365788 16/462779 |
Document ID | / |
Family ID | 57442623 |
Filed Date | 2019-12-05 |
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United States Patent
Application |
20190365788 |
Kind Code |
A1 |
Dousson; Cyril ; et
al. |
December 5, 2019 |
CYCLIC PHOSPHATE SUBSTITUTED NUCLEOSIDE DERIVATIVES FOR THE
TREATMENT OF LIVER DISEASES
Abstract
The present invention relates to Compounds of Formula (I) or
Formula (II) or a pharmaceutically acceptable salt, solvate or
enantiomer thereof, wherein A, B, R.sup.1, R.sup.2, R.sup.3,
R.sup.4, Q and V are as defined herein. The present invention also
relates to pharmaceutical compositions comprising a Compound of
Formula (I) or Formula (II) and to their use in therapy.
##STR00001##
Inventors: |
Dousson; Cyril; (Canet,
FR) ; Dukhan; David; (Montpellier, FR) ;
Parsy; Christophe C.; (Jacou, FR) ; Bogen; Stephane
L.; (Somerset, NJ) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
IDENIX PHARMACEUTICALS LLC |
Cambridge |
MA |
US |
|
|
Assignee: |
IDENIX PHARMACEUTICALS LLC
Cambridge
MA
|
Family ID: |
57442623 |
Appl. No.: |
16/462779 |
Filed: |
November 15, 2017 |
PCT Filed: |
November 15, 2017 |
PCT NO: |
PCT/EP2017/079346 |
371 Date: |
May 21, 2019 |
Current U.S.
Class: |
1/1 |
Current CPC
Class: |
A61K 31/675 20130101;
A61P 1/16 20180101; C07F 9/65586 20130101; C07F 9/65616 20130101;
C07F 9/65744 20130101; C07H 19/11 20130101; C07H 19/213 20130101;
A61P 31/12 20180101 |
International
Class: |
A61K 31/675 20060101
A61K031/675; C07F 9/6558 20060101 C07F009/6558; C07F 9/6561
20060101 C07F009/6561; A61P 1/16 20060101 A61P001/16 |
Foreign Application Data
Date |
Code |
Application Number |
Nov 21, 2016 |
EP |
16306526.1 |
Claims
1. A compound according to Formula I or Formula II: ##STR00106## or
a pharmaceutically acceptable salt or stereoisomer thereof wherein:
A is selected from O, S, CH.sub.2, CF and C.dbd.CH.sub.2, such that
if R.sup.2 is OH and R.sup.3, R.sup.4 and V are hydrogen, then A is
other than S; and if A is CF or C.dbd.CH.sub.2, then V is absent; B
is selected from the following groups: ##STR00107## each Q is
independently O or S; V is hydrogen, halogen, --N(R.sup.13).sub.2,
--OR.sup.13, alkyl, alkenyl, alkynyl, haloalkyl, N.sub.3 or CN; W
is N, CH or CF; R.sup.1 is --CH.sub.2--X--Y--R.sup.16; X is
--C(R.sup.14).sub.2; Y is --C(R.sup.15).sub.2, or
C.sub.3-C.sub.6cycloalkylene; R.sup.2 is hydrogen, fluoro, chloro,
--OR.sup.13, --CN, --N(R.sup.13).sub.2, N.sub.3,
C.sub.1-C.sub.3alkyl, C.sub.1-C.sub.3haloalkyl,
C.sub.2-C.sub.6alkenyl or C.sub.2-C.sub.3alkynyl; R.sup.3 is
hydrogen, C.sub.1-C.sub.6alkyl, C.sub.2-C.sub.6alkenyl,
C.sub.2-C.sub.6alkynyl, C.sub.1-C.sub.6haloalkyl, --OR.sup.13,
fluoro, chloro, N.sub.3, --CN or --N(R.sup.13).sub.2 such that if
R.sup.2 is fluoro or chloro, then R.sup.3 is other than fluoro or
chloro; R.sup.4 is hydrogen, C.sub.1-C.sub.6alkyl,
C.sub.2-C.sub.6alkenyl, C.sub.2-C.sub.6alkynyl,
C.sub.1-C.sub.6haloalkyl, --OR.sup.13, fluoro, chloro, N.sub.3,
--CN or --N(R.sup.13).sub.2; R.sup.5, R.sup.6, R.sup.8 and R.sup.9
are each independently selected from hydrogen,
C.sub.1-C.sub.6alkyl, C.sub.1-C.sub.6haloalkyl,
C.sub.1-C.sub.6hydroxyalkyl, halogen, --OR.sup.18, --SR.sup.18 and
--N(R.sup.18).sub.2; R.sup.7, R.sup.10, R.sup.11 and R.sup.12 are
each independently selected from hydrogen, C.sub.1-C.sub.6alkyl,
C.sub.2-C.sub.6 alkenyl, C.sub.2-C.sub.6alkynyl,
C.sub.3-C.sub.7cycloalkyl, C.sub.5-C.sub.6heteroaryl,
C.sub.9-C.sub.10heteroaryl, halogen, --OR.sup.18, --SR.sup.18,
--S(O)R.sup.18, --S(O).sub.2R.sup.18,
--S(O).sub.2N(R.sup.18).sub.2, --NHC(O)OR.sup.18,
--NHC(O)N(R.sup.18).sub.2, C.sub.1-C.sub.6haloalkyl,
C.sub.1-C.sub.6hydroxyalkyl, --O(C.sub.1-C.sub.6haloalkyl), --CN,
--NO.sub.2, --N(R.sup.18).sub.2,
--NH(C.sub.1-C.sub.6alkylene)-(C.sub.5-C.sub.6heteroaryl),
--NH(C.sub.1-C.sub.6 alkylene)-(C.sub.9-C.sub.10heteroaryl),
--C(O)R.sup.18, --C(O)OR.sup.18, --C(O)N(R.sup.18).sub.2 and
--NHC(O)R.sup.18, wherein said C.sub.2-C.sub.6alkenyl group and
said C.sub.2-C.sub.6alkynyl group are optionally substituted with
one or more halogen; each occurrence of R.sup.13 is independently
selected from hydrogen, C.sub.1-C.sub.6alkyl --C(O)R.sup.18, or
--C(O)OR.sup.18; R.sup.14 is hydrogen, halogen,
C.sub.1-C.sub.6alkyl, C.sub.1-C.sub.6haloalkyl,
C.sub.1-C.sub.6hydroxyalkyl, C.sub.6-C.sub.10aryl-, OR.sup.17,
--OC(O)R.sup.17, --N(R.sup.12)C(O)OR.sup.17 or --C(O)OR.sup.17;
each occurrence of R.sup.15 is independently selected from
hydrogen, halogen, C.sub.1-C.sub.6alkyl, C.sub.1-C.sub.6haloalkyl,
C.sub.1-C.sub.6hydroxyalkyl, C.sub.6-C.sub.10aryl-, OR.sup.17,
--OC(O)R.sup.17, --N(R.sup.12)C(O)OR.sup.17 and --C(O)OR.sup.17 or
both R.sup.15 groups together with the carbon atom to which they
are attached can join to form a 3- to 6-membered spirocyclic
cycloalkyl group; R.sup.16 is --C(O)OR.sup.17; each occurrence of
R.sup.17 is independently selected from hydrogen, halogen,
C.sub.1-C.sub.6alkyl, C.sub.3-C.sub.7cycloalkyl and
C.sub.6-C.sub.10aryl; each occurrence of R.sup.18 is independently
selected from hydrogen, C.sub.1-C.sub.15alkyl,
C.sub.1-C.sub.6haloalkyl, C.sub.1-C.sub.6hydroxyalkyl,
--(C.sub.1-C.sub.3alkylene).sub.m-(C.sub.3-C.sub.7cycloalkyl),
--(C.sub.1-C.sub.3alkylene).sub.m-(C.sub.6-C.sub.10aryl),
--(C.sub.1-C.sub.3alkylene).sub.m-(C.sub.4-C.sub.7heterocycloalkyl),
--(C.sub.1-C.sub.3alkylene).sub.m-(C.sub.5-C.sub.6heteroaryl) and
--(C.sub.1-C.sub.3alkylene).sub.m-(C.sub.9-C.sub.10heteroaryl) and
each occurrence of m is independently 0 or 1; or the compound
##STR00108## or a pharmaceutically acceptable salt or solvate
thereof.
2. The compound of claim 1 wherein A is O or S.
3. The compound of claim 1, wherein R.sup.1 is ##STR00109## and
wherein R.sup.15 and R.sup.17 are each independently selected from
C.sub.1-C.sub.6alkyl.
4. The compound of claim 1, wherein R.sup.2 is hydrogen, fluoro or
hydroxyl.
5. The compound of claim 1, wherein R.sup.3 is hydrogen.
6. The compound of claim 1, wherein R.sup.4 is hydrogen.
7. The compound of claim 1, wherein V is hydrogen.
8. The compound of claim 1, wherein B is ##STR00110## wherein Q is
O; W is N; R.sup.5 and R.sup.6 are hydrogen; R.sup.7 is NH.sub.2 or
NHC(O)aryl; R.sup.8 is hydrogen; R.sup.9 is hydrogen or
trifluoromethyl; R.sup.10 is --NH.sub.2 or
--O--C.sub.1-C.sub.6alkyl and R.sup.11 is --NH.sub.2 or
halogen.
9. A compound of claim 1 selected from the following: ##STR00111##
##STR00112## ##STR00113## ##STR00114## or a pharmaceutically
acceptable salt thereof.
10. A pharmaceutical composition comprising a compound of claim 1,
or a pharmaceutically acceptable salt or stereoisomer thereof, and
a pharmaceutically acceptable carrier or diluent.
11. (canceled)
12. A method for treating a liver disease in a patient, the method
comprising administering to the patient a compound of claim 1, or a
pharmaceutically acceptable salt or stereoisomer thereof.
13. The method of claim 12, where the liver disease being treated
is liver cancer.
14. The method of claim 12, where the liver disease being treated
is HBV infection.
15. A method for treating Ebola virus infection in a patient, the
method comprising administering to the patient a compound of claim
1, or a pharmaceutically acceptable salt or stereoisomer
thereof.
16. (canceled)
17. The composition of claim 10 further comprising one or more
additional therapeutic agents.
Description
FIELD OF THE INVENTION
[0001] The present invention relates to compounds of Formula (I) or
Formula (II), compositions comprising the compounds of Formula (I)
or Formula (II) and the compounds of Formula (I) or Formula (II)
for use in treating or preventing liver diseases, such as cancer,
Hepatitis B virus infection or Ebolavirus infection in a
patient.
BACKGROUND OF THE INVENTION
[0002] Cancer is a disease characterised primarily by an
uncontrolled division of abnormal cells derived from a given normal
tissue and the invasion of adjacent tissues by these malignant
cells. Blood or lymphatic transportation can spread cancer cells to
other parts of the body leading to regional lymph nodes and to
distant sites (metastasis). Cancer is a complex, multistep process
that begins with minor preneoplastic changes, which may under
certain conditions progress to neoplasia. There are more than 100
different types of cancer, which can be grouped into broader
categories. The main categories include: carcinoma, sarcoma,
leukemia, lymphoma and myeloma, and central nervous system cancers.
The incidence of cancer continues to climb as the general
population ages, as new cancers develop, and as susceptible
populations (e.g., people infected with AIDS or excessively exposed
to sunlight) grow.
[0003] Primary liver cancer is one of the most common forms of
cancer in the world. Hepatocellular carcinoma, also known as
malignant hepatoma, is the most common form of primary liver
cancer, and develops within the hepatocyte. Hepatocellular
carcinoma occurs mostly in men and patients that suffer from
cirrhosis. It has been the third leading cause of cancer deaths
worldwide (Block T. M. et al., 2003, Oncogene 22:5093-5107). Many
patients with hepatocellular carcinoma remain asymptomatic until
the disease is in its advanced stages, resulting in ineffective
treatment and poor prognosis; the majority of unresectable
hepatocellular carcinoma patients die within one year.
[0004] Treatment options for hepatocellular carcinoma have been
limited, especially in the case of advanced or recurrent
hepatocellular carcinoma. Surgery and radiation therapy are options
for early stage liver cancer, but not very effective for advanced
or recurrent hepatocellular carcinoma. Systematic chemotherapies
have not been particularly effective, and there are a very limited
number of drugs available for use. The recently approved kinase
inhibitor sorafenib has been shown to be effective in treating
hepatocellular carcinoma. However, it can slow or stop advanced
liver cancer from progressing for only a few months longer than
without treatment.
[0005] Liver cancers which can be treated include primary and
secondary liver cancers. In particular cases, the liver cancer is
hepatocellular carcinoma, cholangiocarcinoma, or biliary tract
cancer. In particular cases, the liver cancer is a metastasis in
the liver originated from other cancers (such as colon cancer,
pancreatic cancer, etc.). In certain embodiments, the compounds
described herein can be used to treat cancers such as breast
cancer, ovarian cancer, lung cancer, pancreatic cancer and leukemic
cancer
[0006] Hepatitis B virus (HBV) infection is a major health problem
that can lead to chronic liver disease, such as cirrhosis and
hepatocellular carcinoma, in a substantial number of infected
individuals. Although most individuals seem to resolve the
infection following acute symptoms, approximately 30% of cases
become chronic. According to recent estimates around five percent
of the world's population have chronic hepatitis B, leading to half
a million to one million deaths per year.
[0007] HBV is a small DNA virus that is considered to be the
prototypical member of the hepadnaviridae. HBV is an enveloped
virus with an unusual mode of replication, centering on the
establishment of a covalently closed circular DNA (cccDNA) copy of
its genome in the host cell nucleus. The episomal form is
established from conversion of the partially double stranded
circular DNA (relaxed circular, or rcDNA) genome upon initial
infection and functions as the template for all HBV mRNAs. Unlike
the mechanisms of most other DNA viruses, HBV cccDNA replicates
through retrotranscription of a 1.1 genome unit-length RNA copy
(pregenomic, or pgDNA) which is originally transcribed from the
cccDNA template and which is acted upon by a virus encoded
polymerase to yield progeny rcDNA. HBV DNA synthesis is coupled to
assembly of its capsid and most copies of the encapsidated genome
then efficiently associate with the envelope proteins for viron
assembly and secretion; a minority of these genomes are shunted to
the nucleus where they are converted to cccDNA, thus amplifying
levels of the episome. HBV is classified into eight genotypes
(A-H)
[0008] Although the viral polymerase and surface antigen (HBsAg)
perform very different functions, both are essential proteins for
the virus to complete its life cycle and be infectious. That is HBV
lacking HBsAg is completely defective and cannot infect or cause
infection. HBsAg is needed to protect the virus nucleocapsid, to
begin the infectious cycle and to mediate morphogenesis and
secretion of newly forming virus from the infected cells.
[0009] HBV is transmitted parenterally by contaminated blood and
blood products, contaminated needles, or sexually and vertically
from infected mothers or carrier mothers to their off-spring.
People who are chronically affected with HBV are usually
characterised by readily detectable levels of circulating antibody
specific to the viral capsid (HBc), with little, if any, detectable
levels of antibody to HBsAg. There is some evidence that chronic
carriers do produce antibodies to HBsAg, but these antibodies are
complexed with the circulating HBsAg, which can be present in
milligram per milliliter amounts in a chronic carrier's
circulation.
[0010] As the only enzyme encoded by HBV, the polymerase has been
well exploited as a target for antiviral drug development, with
several nucleoside polymerase inhibitors approved for drug use and
others in development. Mutations in the primary sequence of the
polymerase that confer resistance to lamivudine and adefovir have
been identified clinically, and underly a rebound of serum virus
titers that 70% of treated patients experience within three years
of starting lamivudine therapy. Although resistance to telbivudine,
adefovir and entecavir occurs more rarely, it has been recorded.
.alpha.-interferon is the other major therapy available for
treatment of hepatitis B, but is limited by poor long-term response
and debilitating side-effects. Despite the availability of an
effective vaccine, immunoglobulin therapy, interferon and antiviral
drugs, there is a need for improved therapeutic agents that
effectively combat chronic HBV infection.
[0011] The viruses of the Filoviridae family are enveloped negative
sense, single-stranded, linear RNA viruses. Three genera within the
Filoviridae family are Ebolavirus, Marburgvirus and `Cuevavirus`
(tentative). The five recognized species of Ebolavirus are Ebola
virus (EBOV), Reston ebolavirus (REBOV), Sudan ebolavirus (SEBOV),
Tai Forest ebolavirus (TAFV) and Bundibugyo ebolavirus (BEBOV).
Ebolavirus and Marburgvirus are both highly infectious and
contagious. Both viruses are transmitted by direct contact with the
blood, body fluids and/or tissues of the infected persons.
Ebolavirus and Marburgvirus can also be transmitted by handling
sick or dead infected wild animals. Ebola hemorrhagic fever (EHF)
is caused by an Ebolavirus infection. Marburg virus disease (MVD)
is a human disease caused by Marburgvirus and causes Marburgvirus
hemorrhagic fever (MHF). The primary organs infected by Ebolavirus
have been shown to be the liver and the spleen as reported in Rev.
Fr. Histotechnol., 2012, vol. 25 no. 1, pages 65-80.
[0012] There remains a need for further therapies for the treatment
of liver diseases, such as cancer or HBV infection or Ebolavirus
infection which are safe and effective.
SUMMARY OF THE INVENTION
[0013] In one aspect, the present invention provides a compound
according to Formula (I) or Formula (II):
##STR00002##
or a pharmaceutically acceptable salt, solvate or enantiomer
thereof, wherein:
[0014] A is selected from O, S, CH.sub.2, CF and C.dbd.CH.sub.2,
with the proviso that if R.sup.2 is OH and R.sup.3, R.sup.4 and V
are hydrogen, then A is other than S; and if A is CF or
C.dbd.CH.sub.2, then V is absent;
[0015] B is selected from the following groups:
##STR00003##
[0016] Q is O or S;
[0017] V is hydrogen, halogen, --N(R.sup.13).sub.2, --OR.sup.13,
alkyl, alkenyl, alkynyl, haloalkyl, N.sub.3 or CN;
[0018] W is N, CH or CF;
[0019] R.sup.1 is --CH.sub.2--X--Y--R.sup.16;
[0020] X is --C(R.sup.14).sub.2;
[0021] Y is --C(R.sup.15).sub.2 or
C.sub.3-C.sub.6cycloalkylene;
[0022] R.sup.2 is hydrogen, fluoro, chloro, --OR.sup.13, --CN,
--N(R.sup.13).sub.2, N.sub.3, C.sub.1-C.sub.3alkyl,
C.sub.1-C.sub.3haloalkyl, C.sub.2-C.sub.6alkenyl or
C.sub.2-C.sub.3alkynyl;
[0023] R.sup.3 is hydrogen, C.sub.1-C.sub.6alkyl,
C.sub.2-C.sub.6alkenyl, C.sub.2-C.sub.6alkynyl,
C.sub.1-C.sub.6haloalkyl, --OR.sup.13, fluoro, chloro, N.sub.3,
--CN or --N(R.sup.13).sub.2 with the proviso that if R.sup.2 is
fluoro or chloro, then R.sup.3 is other than fluoro or chloro;
[0024] R.sup.4 is hydrogen, C.sub.1-C.sub.6alkyl,
C.sub.2-C.sub.6alkenyl, C.sub.2-C.sub.6alkynyl,
C.sub.1-C.sub.6haloalkyl, --OR.sup.13, fluoro, chloro, N.sub.3,
--CN or --N(R.sup.13).sub.2;
[0025] R.sup.5, R.sup.6, R.sup.8 and R.sup.9 are each independently
selected from hydrogen, C.sub.1-C.sub.6alkyl,
C.sub.1-C.sub.6haloalkyl, C.sub.1-C.sub.6hydroxyalkyl, halogen,
--OR.sup.18, --SR.sup.18 and --N(R.sup.18).sub.2; C.sub.1-C.sub.10
alkyl or --COOR.sup.7;
[0026] R.sup.7, R.sup.10, R.sup.11 and R.sup.12 are each
independently selected from hydrogen, C.sub.1-C.sub.6alkyl,
C.sub.2-C.sub.6 alkenyl, C.sub.2-C.sub.6alkynyl,
C.sub.3-C.sub.7cycloalkyl, C.sub.5-C.sub.6heteroary,
C.sub.9-C.sub.10heteroaryl, halogen, --OR.sup.18, --SR.sup.18,
--S(O)R.sup.18, --S(O).sub.2R.sup.18,
--S(O).sub.2N(R.sup.18).sub.2, --NHC(O)OR.sup.18,
--NHC(O)N(R.sup.18).sub.2, C.sub.1-C.sub.6haloalkyl,
C.sub.1-C.sub.6hydroxyalkyl, --O(C.sub.1-C.sub.6haloalkyl), --CN,
--NO.sub.2, --N(R.sup.18).sub.2,
--NH(C.sub.1-C.sub.6alkylene)-(C.sub.5-C.sub.6heteroaryl),
--NH(C.sub.1-C.sub.6 alkylene)-(C.sub.9-C.sub.10heteroaryl),
--C(O)R.sup.18, --C(O)OR.sup.18, --C(O)N(R.sup.18).sub.2 and
--NHC(O)R.sup.18, wherein said C.sub.2-C.sub.6alkenyl group and
said C.sub.2-C.sub.6alkynyl group are optionally substituted with
one or more halogen;
[0027] each occurrence of R.sup.13 is independently selected from
hydrogen, C.sub.1-C.sub.6 alkyl, --C(O)R.sup.18 or
--C(O)OR.sup.18;
[0028] R.sup.14 is hydrogen, halogen, C.sub.1-C.sub.6alkyl,
C.sub.1-C.sub.6haloalkyl, C.sub.1-C.sub.6hydroxyalkyl,
C.sub.6-C.sub.10aryl-, OR.sup.7, --OC(O)R.sup.17,
--N(R.sup.12)C(O)OR.sup.17 or --C(O)OR.sup.17;
[0029] each occurrence of R.sup.15 is independently selected from
hydrogen, halogen, C.sub.1-C.sub.6 alkyl, C.sub.1-C.sub.6
haloalkyl, C.sub.1-C.sub.6 hydroxyalkyl, C.sub.6-C.sub.10aryl-,
OR.sup.17, --OC(O)R.sup.17, --N(R.sup.12)C(O)OR.sup.17 and
--C(O)OR.sup.17 or both R.sup.15 groups together with the carbon
atom to which they are attached can join to form a 3- to 6-membered
spirocyclic cycloalkyl group;
[0030] R.sup.16 is --C(O)OR.sup.17;
[0031] each occurrence of R.sup.17 is independently selected from
hydrogen, halogen, C.sub.1-C.sub.6alkyl, C.sub.3-C.sub.7cycloalkyl
and C.sub.6-C.sub.10aryl;
[0032] each occurrence of R.sup.18 is independently selected from
hydrogen, C.sub.1-C.sub.15alkyl, C.sub.1-C.sub.6haloalkyl,
C.sub.1-C.sub.6hydroxyalkyl,
--(C.sub.1-C.sub.3alkylene).sub.m-(C.sub.3-C.sub.7cycloalkyl),
--(C.sub.1-C.sub.3alkylene).sub.m-(C.sub.6-C.sub.10aryl),
--(C.sub.1-C.sub.3alkylene).sub.m-(C.sub.4-C.sub.7heterocycloalkyl),
--(C.sub.1-C.sub.3alkylene).sub.m-(C.sub.5-C.sub.6heteroaryl) and
--(C.sub.1-C.sub.3alkylene).sub.m-(C.sub.9-C.sub.10heteroaryl) and
[0033] each occurrence of m is independently 0 or 1; [0034] or the
compound
##STR00004##
[0035] or a pharmaceutically acceptable salt or solvate
thereof.
[0036] In a further aspect, the present invention provides a
pharmaceutical composition comprising a compound according to
Formula (I) or Formula (II) or a pharmaceutically acceptable salt,
solvate or enantiomer thereof, and a pharmaceutically acceptable
carrier or diluent.
[0037] The compounds of Formula (I) or Formula (II) or
pharmaceutically acceptable salts, solvates or enantiomers thereof
are useful in therapy. In particular, the compounds of Formula (I)
or Formula (II) or pharmaceutically acceptable salts, solvates or
enantiomers thereof are useful in the treatment or prevention of
liver diseases in a patient. As such, compounds of Formula (I) or
Formula (II) or pharmaceutically acceptable salts, solvates or
enantiomers thereof can be useful, for example, for treating or
preventing cancer in a patient.
[0038] Compounds of Formula (I) or Formula (II) or pharmaceutically
acceptable salts, solvates or enantiomers thereof can also be
useful for inhibiting HBV replication or replicon activity and for
treating or preventing HBV infection in a patient. Compounds of
Formula (I) or Formula (II) or pharmaceutically acceptable salts,
solvates or enantiomers thereof can also be useful for treating or
preventing Ebolavirus infection in a patient.
[0039] Accordingly, the present invention provides a compound of
Formula (I) or Formula (II) or a pharmaceutically acceptable salt,
solvate or enantiomer thereof for use in the treatment or
prevention of liver diseases in a patient. The present invention
further provides a compound of Formula (I) or Formula (II) or a
pharmaceutically acceptable salt, solvate or enantiomer thereof for
use in treating or preventing cancer in a patient. The present
invention further provides a compound of Formula (I) or Formula
(II) or a pharmaceutically acceptable salt, solvate or enantiomer
thereof for use in treating or preventing HBV infection in a
patient. The present invention further provides a compound of
Formula (I) or Formula (II) or a pharmaceutically acceptable salt,
solvate or enantiomer thereof for use in treating or preventing
Ebolavirus infection in a patient.
[0040] The present invention further provides a combination
comprising a compound of Formula (I) or Formula (II), or a
pharmaceutically acceptable salt, solvate or enantiomer thereof and
one, two, three or more other therapeutic agents.
[0041] The details of the invention are set forth in the
accompanying detailed description below.
DETAILED DESCRIPTION OF THE INVENTION
Definitions and Abbreviations
[0042] The terms used herein have their ordinary meaning and the
meaning of such terms is independent at each occurrence thereof.
That notwithstanding and except where stated otherwise, the
following definitions apply throughout the specification and
claims. Chemical names, common names, and chemical structures may
be used interchangeably to describe the same structure. If a
chemical compound is referred to using both a chemical structure
and a chemical name and an ambiguity exists between the structure
and the name, the structure predominates. These definitions apply
regardless of whether a term is used by itself or in combination
with other terms, unless otherwise indicated. Hence, the definition
of "alkyl" applies to "alkyl" as well as the "alkyl" portions of
"hydroxyalkyl," "haloalkyl," "--O-alkyl," etc.
[0043] As used herein, and throughout this disclosure, the
following terms, unless otherwise indicated, shall be understood to
have the following meanings:
[0044] The term "alkyl," as used herein, refers to an aliphatic
hydrocarbon group having one of its hydrogen atoms replaced with a
bond. An alkyl group may be straight or branched and contain from
about 1 to about 20 carbon atoms. In one embodiment, an alkyl group
contains from about 1 to about 12 carbon atoms. In different
embodiments, an alkyl group contains from 1 to 6 carbon atoms
(C.sub.1-C.sub.6alkyl), or from 1 to 3 carbon atoms
(C.sub.1-C.sub.3alkyl). Examples of alkyl groups include methyl,
ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl,
tert-butyl, n-pentyl, neopentyl, isopentyl, n-hexyl, isohexyl and
neohexyl. In one embodiment, an alkyl group is linear. In another
embodiment, an alkyl group is branched. Unless otherwise indicated,
an alkyl group is unsubstituted.
[0045] The term "C.sub.1-C.sub.6 alkoxy" as used herein, refers to
a group having the formula --O--(C.sub.1-C.sub.6alkyl), where the
term "C.sub.1-C.sub.6 alkyl" is defined above herein.
[0046] The term "alkenyl," as used herein, refers to an aliphatic
hydrocarbon group containing at least one carbon-carbon double bond
and having one of its hydrogen atoms replaced with a bond. An
alkenyl group may be straight or branched and contain from about 2
to about 15 carbon atoms. In one embodiment, an alkenyl group
contains from 2 to 12 carbon atoms. In another embodiment, an
alkenyl group contains from 2 to 6 carbon atoms
(C.sub.2-C.sub.6alkenyl). Examples of alkenyl groups include
ethenyl, propenyl, n-butenyl, 3-methylbut-2-enyl, n-pentenyl,
octenyl and decenyl. Unless otherwise indicated, an alkenyl group
is unsubstituted.
[0047] The term "alkynyl," as used herein, refers to an aliphatic
hydrocarbon group containing at least one carbon-carbon triple bond
and having one of its hydrogen atoms replaced with a bond. An
alkynyl group may be straight or branched and contain from about 2
to about 15 carbon atoms. In one embodiment, an alkynyl group
contains from about 2 to about 12 carbon atoms. In another
embodiment, an alkynyl group contains from 2 to 6 carbon atoms
(C.sub.2-C.sub.6alkynyl). In another embodiment, an alkynyl group
contains from 2 to 3 carbon atoms (C.sub.2-C.sub.3alkynyl).
Examples of alkynyl groups include ethynyl, propynyl, 2-butynyl and
3-methylbutynyl. Unless otherwise indicated, an alkynyl group is
unsubstituted.
[0048] The term "alkylene," as used herein, refers to an alkyl
group, as defined above, wherein one of the alkyl group's hydrogen
atoms has been replaced with a bond. Examples of alkylene groups
include --CH.sub.2--, --CH.sub.2CH.sub.2--,
--CH.sub.2CH.sub.2CH.sub.2--, --CH.sub.2CH.sub.2CH.sub.2CH.sub.2--,
--CH(CH.sub.3)CH.sub.2CH.sub.2--, --CH(CH.sub.3)-- and
--CH.sub.2CH(CH.sub.3)CH.sub.2--. In one embodiment, an alkylene
group has from 1 to 6 carbon atoms (C.sub.1-C.sub.6alkylene). In
another embodiment, an alkylene group has from 1 to 3 carbon atoms
(C.sub.1-C.sub.3alkylene). In another embodiment, an alkylene group
is branched. In another embodiment, an alkylene group is linear. In
one embodiment, an alkylene group is --CH.sub.2--.
[0049] The term "aryl," as used herein, refers to an aromatic
monocyclic or multicyclic ring system comprising from about 6 to
about 14 carbon atoms. In one embodiment, an aryl group contains
from 6 to 10 carbon atoms (C.sub.6-C.sub.10aryl). In one
embodiment, an aryl group can be optionally fused to a cycloalkyl
or cycloalkanoyl group. Examples of aryl groups include phenyl and
naphthyl. In one embodiment, an aryl group is phenyl. The term
"aryloxy" as used herein, refers to a group having the formula
--O-aryl, where the term "aryl" is defined above herein.
[0050] The term "cycloalkyl," as used herein, refers to a
non-aromatic mono- or multicyclic ring system comprising from 3 to
about 10 ring carbon atoms. In one embodiment, a cycloalkyl
contains from 3 to 7 ring carbon atoms (C.sub.3-C.sub.7cycloalkyl).
In another embodiment, a cycloalkyl contains from 5 to 6 ring
atoms. Examples of monocyclic cycloalkyls include cyclopropyl,
cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl and cyclooctyl.
Examples of multicyclic cycloalkyls include 1-decalinyl, norbomyl
and adamantly. In one embodiment, a cycloalkyl group is
unsubstituted. The term "3 to 6-membered cycloalkyl" refers to a
cycloalkyl group having from 3 to 6 ring carbon atoms. A ring
carbon atom of a cycloalkyl group may be functionalized as a
carbonyl group. An illustrative example of such a cycloalkyl group
(also referred to herein as a "cycloalkanoyl" group) includes
cyclobutanoyl:
##STR00005##
[0051] The term "cycloalkylene," as used herein, refers to a
cycloalkyl group, as defined above, wherein one of the cycloalkyl
group's hydrogen atoms has been replaced with a bond. Examples of
cycloalkylene groups include cyclopropylene, cyclobutylene,
cyclopentylene and cyclohexylene. In one embodiment, a
cycloalkylene group has from 3 to 6 carbon atoms
(C.sub.3-C.sub.6cycloalkylene).
[0052] The term "halogen," as used herein, means --F, --Cl, --Br or
--I.
[0053] The term "haloalkyl," as used herein, refers to an alkyl
group as defined above, wherein one or more of the alkyl group's
hydrogen atoms have been replaced with a halogen. In one
embodiment, a haloalkyl group has from 1 to 6 carbon atoms
(C.sub.1-C.sub.6haloalkyl). In another embodiment, a haloalkyl
group has from 1 to 3 carbon atoms (C.sub.1-C.sub.3haloalkyl). In
another embodiment, a haloalkyl group is substituted with from 1 to
3 F atoms. Examples of haloalkyl groups include --CH.sub.2F,
--CHF.sub.2, --CF.sub.3, --CH.sub.2C.sub.1 and --CCl.sub.3.
[0054] The term "hydroxyalkyl," as used herein, refers to an alkyl
group as defined above, wherein one or more of the alkyl group's
hydrogen atoms have been replaced with an --OH group. In one
embodiment, a hydroxyalkyl group has from 1 to 6 carbon atoms
(C.sub.1-C.sub.6hydroxyalkyl). Non-limiting examples of
hydroxyalkyl groups include --CH.sub.2OH, --CH.sub.2CH.sub.2OH,
--CH.sub.2CH.sub.2CH.sub.2OH and --CH.sub.2CH(OH)CH.sub.3.
[0055] The term "5 or 6-membered monocyclic heteroaryl," or
C.sub.5-C.sub.6heteroaryl, as used herein, refers to an aromatic
monocyclic ring system comprising 5 to 6 ring atoms, wherein from 1
to 4 of the ring atoms is independently O, N or S and the remaining
ring atoms are carbon atoms. A 5 or 6-membered monocyclic
heteroaryl group is joined via a ring carbon atom, and any nitrogen
atom of a heteroaryl can be optionally oxidized to the
corresponding N-oxide. Examples of 5 or 6-membered monocyclic
heteroaryls include pyridyl, pyrazinyl, furanyl, thienyl,
pyrimidinyl, pyridone (including N-substituted pyridones),
isoxazolyl, isothiazolyl, oxazolyl, oxadiazolyl, thiazolyl,
pyrazolyl, furazanyl, pyrrolyl, triazolyl, 1,2,4-thiadiazolyl,
pyrazinyl, pyridazinyl, imidazolyl, 1,2,4-triazinyl and the like,
and all isomeric forms thereof.
[0056] The term "9 or 10-membered bicyclic heteroaryl," or
C.sub.9-C.sub.10heteroaryl as used herein, refers to an aromatic
bicyclic ring system comprising 9 to 10 ring atoms, wherein from 1
to 4 of the ring atoms is independently O, N or S and the remaining
ring atoms are carbon atoms. A 9 or 10-membered bicyclic heteroaryl
group is joined via a ring carbon atom, and any nitrogen atom of a
heteroaryl can be optionally oxidized to the corresponding N-oxide.
Examples of 9 or 10-membered bicyclic heteroaryls include
imidazo[1,2-a]pyridinyl, imidazo[2,1-b]thiazolyl, benzofurazanyl,
indolyl, azaindolyl, benzimidazolyl, benzothienyl, quinolinyl,
benzimidazolyl, quinazolinyl, pyrrolopyridyl, imidazopyridyl,
isoquinolinyl, benzoazaindolyl, benzothiazolyl, and the like, and
all isomeric forms thereof.
[0057] The term "heterocycloalkyl," as used herein, refers to a
non-aromatic monocyclic or multicyclic ring system comprising 3 to
about 11 ring atoms, wherein from 1 to 4 of the ring atoms are
independently O, S, N or Si, and the remainder of the ring atoms
are carbon atoms. A heterocycloalkyl group can be joined via a ring
carbon, ring silicon atom or ring nitrogen atom. In one embodiment,
a heterocycloalkyl group is monocyclic and has from 3 to 7 ring
atoms (C.sub.3-C.sub.7heterocycloalkyl). In another embodiment, a
heterocycloalkyl group is monocyclic has from 4 to 7 ring atoms
(C.sub.4-C.sub.7heterocycloalkyl). In another embodiment, a
heterocycloalkyl group is bicyclic and has from 7 to 11 ring atoms.
In still another embodiment, a heterocycloalkyl group is monocyclic
and has 5 or 6 ring atoms. In one embodiment, a heterocycloalkyl
group is monocyclic. In another embodiment, a heterocycloalkyl
group is bicyclic. There are no adjacent oxygen and/or sulfur atoms
present in the ring system. Any --NH group in a heterocycloalkyl
ring may exist protected such as, for example, as an --N(BOC),
--N(Cbz), --N(Tos) group and the like; such protected
heterocycloalkyl groups are considered part of this invention. The
term "heterocycloalkyl" also encompasses a heterocycloalkyl group,
as defined above, which is fused to an aryl (e.g., benzene) or
heteroaryl ring. The nitrogen or sulfur atom of the
heterocycloalkyl can be optionally oxidized to the corresponding
N-oxide, S-oxide or S,S-dioxide. Examples of monocyclic
heterocycloalkyl rings include oxetanyl, piperidyl, pyrrolidinyl,
piperazinyl, morpholinyl, thiomorpholinyl, thiazolidinyl,
1,4-dioxanyl, tetrahydrofuranyl, tetrahydrothiophenyl,
delta-lactam, delta-lactone, silacyclopentane, silapyrrolidine and
the like, and all isomers thereof. Illustrative examples of a
silyl-containing heterocycloalkyl group include:
##STR00006##
[0058] A ring carbon atom of a heterocycloalkyl group may be
functionalized as a carbonyl group. An illustrative example of such
a heterocycloalkyl group is:
##STR00007##
[0059] In one embodiment, a heterocycloalkyl group is a 5-membered
monocyclic heterocycloalkyl. In another embodiment, a
heterocycloalkyl group is a 6-membered monocyclic heterocycloalkyl.
The term "3 to 6-membered monocyclic cycloalkyl" refers to a
monocyclic heterocycloalkyl group having from 3 to 6 ring atoms.
The term "4 to 6-membered monocyclic cycloalkyl" refers to a
monocyclic heterocycloalkyl group having from 4 to 6 ring atoms.
The term "7 to 11-membered bicyclic heterocycloalkyl" refers to a
bicyclic heterocycloalkyl group having from 7 to 11 ring atoms.
Unless otherwise indicated, an heterocycloalkyl group is
unsubstituted.
[0060] The term "substituted", as used herein, means that one or
more hydrogens on the designated atom is replaced with a selection
from the indicated group, provided that the designated atom's
normal valency under the existing circumstances is not exceeded,
and that the substitution results in a stable compound.
Combinations of substituents and/or variables are permissible only
if such combinations result in stable compounds. By "stable
compound` or "stable structure" is meant a compound that is
sufficiently robust to survive isolation to a useful degree of
purity from a reaction mixture, and formulation into an efficacious
therapeutic agent.
[0061] When referring to a substituent on a sugar ring of a
nucleoside, the term "beta" refers to a substituent on the same
side of the plane of the sugar ring as the 5' carbon and the term
"alpha" refers to a substituent on the opposite side of the plane
of the sugar ring from the 5' carbon. As shown below, substituent
"A" is in the "alpha" position, and substituent "B" is in the
"beta" position with respect to the 5' carbon.
##STR00008##
[0062] The term "in substantially purified form," as used herein,
refers to the physical state of a compound after the compound is
isolated from a synthetic process (e.g., from a reaction mixture),
a natural source, or a combination thereof. The term "in
substantially purified form," also refers to the physical state of
a compound after the compound is obtained from a purification
process or processes described herein or well-known to the skilled
artisan (e.g., chromatography, recrystallization and the like), in
sufficient purity to be characterizable by standard analytical
techniques described herein or well-known to the skilled
artisan.
[0063] It should also be noted that any carbon as well as
heteroatom with unsatisfied valences in the text, schemes, examples
and tables herein is assumed to have the sufficient number of
hydrogen atom(s) to satisfy the valences.
[0064] When a functional group in a compound is termed "protected",
this means that the group is in modified form to preclude undesired
side reactions at the protected site when the compound is subjected
to a reaction. Suitable protecting groups will be recognized by
those with ordinary skill in the art as well as by reference to
standard textbooks such as, for example, T. W. Greene et al,
Protective Groups in Organic Synthesis (1991), Wiley, New York.
[0065] When any substituent or variable (e.g., alkyl, R.sup.6,
R.sup.a, etc.) occurs more than one time in any constituent or in
Formula (I) or Formula (II), its definition on each occurrence is
independent of its definition at every other occurrence, unless
otherwise indicated.
[0066] The term "composition," as used herein, is intended to
encompass a product comprising the specified ingredients in the
specified amounts, as well as any product which results directly
from combination of the specified ingredients in the specified
amounts.
[0067] IC.sub.50, as used herein, refers to an amount,
concentration or dosage of a particular test compound that achieves
a 50% inhibition of a maximal response in an assay that measures
such response.
[0068] The term "host," as used herein, refers to any unicellular
or multicellular organism, including cell lines and animals, and in
certain embodiments, a human. Alternatively, the host can be
carrying a part of the Flaviviridae or Filovaridae viral genome,
whose replication or function can be altered by the compounds of
the present invention. The term "host" specifically includes
infected cells, cells transfected with all or part of the
Flaviviridae or Filovaridae genome and animals, in particular,
primates (including chimpanzees) and humans. In most animal
applications of the present invention, the host is a human patient.
Veterinary applications, in certain indications, however, are
clearly anticipated by the present invention (such as
chimpanzees).
[0069] The terms "subject" and "patient," as used herein, are used
interchangeably. The terms "subject" and "subjects" refer to a
mammal including a non-primate (e.g., a cow, pig, horse, cat, dog,
rat, and mouse) and a primate (e.g., a monkey such as a cynomolgous
monkey, a chimpanzee and a human), and for example, a human. The
subject may also be a farm animal (e.g., a horse, a cow, a pig,
etc.) or a pet (e.g., a dog or a cat).
[0070] The terms "therapeutic agent" and "therapeutic agents," as
used herein, refer to any agent(s) which can be used in the
treatment or prevention of a disorder or one or more symptoms
thereof. The term "therapeutic agent" includes a compound provided
herein. In some cases, a therapeutic agent can be an agent which is
known to be useful for, or has been or is currently being used for
the treatment or prevention of a disorder or one or more symptoms
thereof.
[0071] The term "effective amount," as used herein, refers to an
amount of a compound of Formula (I) or Formula (II) and/or an
additional therapeutic agent, or a composition thereof that is
effective in producing the desired therapeutic, ameliorative,
inhibitory or preventative effect when administered to a patient
suffering from a viral infection or virus-related disorder. In the
combination therapies of the present invention, an effective amount
can refer to each individual agent or to the combination as a
whole, wherein the amounts of all agents administered are together
effective, but wherein the component agent of the combination may
not be present individually in an effective amount. A
"therapeutically effective amount" can vary depending on, inter
alia, the compound, the disease and its severity, and the age,
weight, etc., of the subject to be treated.
[0072] The term "treating" or "treatment" of any disease or
disorder, as used herein, refers to ameliorating a disease or
disorder that exists in a subject. In some cases, "treating" or
"treatment" includes ameliorating at least one physical parameter,
which may be indiscernible by the subject. In other cases,
"treating" or "treatment" includes modulating the disease or
disorder, either physically (e.g., stabilization of a discernible
symptom) or physiologically (e.g., stabilization of a physical
parameter) or both. In other cases, "treating" or "treatment"
includes delaying the onset of the disease or disorder.
[0073] The term "preventing," as used herein with respect to a
liver disease or disorder, refers to reducing the likelihood or
severity of the liver disease or disorder.
The Compounds of Formula (I)
[0074] The present invention provides compounds of Formula (I),
having the formula:
##STR00009##
or a pharmaceutically acceptable salt, solvate or enantiomer
thereof wherein A, B, R.sup.1, R.sup.2, R.sup.3, R.sup.4, Q and V
are defined above for the compounds of Formula (I).
[0075] In one embodiment, A is O, S or CH.sub.2, with the proviso
that if R.sup.2 is OH and R.sup.3, R.sup.4 and V are hydrogen, then
A is other than S.
[0076] In another embodiment, A is O.
[0077] In another embodiment, A is S, with the proviso that if
R.sup.2 is OH, then R.sup.3, R.sup.4 and V cannot be hydrogen.
[0078] In another embodiment A, is CF or C.dbd.CH.sub.2 and V is
absent.
[0079] In another embodiment, A is CF and V is absent.
[0080] In another embodiment, A is C.dbd.CH.sub.2 and V is
absent.
[0081] In one embodiment, Q is O.
[0082] In another embodiment, Q is S.
[0083] In one embodiment, R.sup.1 is
--CH.sub.2--C(R.sup.14).sub.2--C(R.sup.15).sub.2--C(O)OR.sup.17,
wherein R.sup.14, R.sup.15 and R.sup.17 are as previously
defined.
[0084] In one embodiment, R.sup.1 is
--CH.sub.2--C(R.sup.14).sub.2--C(R.sup.15).sub.2--C(O)OR.sup.17,
wherein R.sup.14, R.sup.15 and R.sup.17 are each independently
selected from hydrogen and C.sub.1-C.sub.6alkyl.
[0085] In another embodiment, R.sup.1 is
--CH.sub.2--CH.sub.2--C(R.sup.15).sub.2--C(O)OR.sup.17, wherein
R.sup.15 and R.sup.17 are as previously defined.
[0086] In another embodiment, R.sup.1 is
##STR00010##
wherein R.sup.15 and R.sup.17 are as previously defined.
[0087] In another embodiment, R.sup.1 is
##STR00011##
wherein R.sup.15 and R.sup.17 are each independently selected from
C.sub.1-C.sub.6alkyl.
[0088] In another embodiment, R.sup.1 is
##STR00012##
wherein R.sup.15 and R.sup.17 are each independently selected from
C.sub.1-C.sub.6alkyl.
[0089] In another embodiment, R.sup.1 is
##STR00013##
wherein R.sup.15 and R.sup.17 are each independently selected from
C.sub.1-C.sub.6alkyl.
[0090] In another embodiment, R.sup.1 is
##STR00014##
wherein R.sup.15 is methyl and R.sup.17 is selected from ethyl or
propyl.
[0091] In another embodiment, R.sup.1 is
##STR00015##
wherein R.sup.15 is methyl and R.sup.17 is selected from ethyl or
propyl.
[0092] In another embodiment, R.sup.1 is
##STR00016##
wherein R.sup.15 is methyl and R.sup.17 is selected from ethyl or
propyl.
[0093] In another embodiment, R.sup.1 is
##STR00017##
[0094] In another embodiment, R.sup.1 is
##STR00018##
[0095] In another embodiment, R.sup.1 is
##STR00019##
[0096] In another embodiment, R.sup.1 is
##STR00020##
[0097] In one embodiment, R.sup.2 is hydrogen, fluoro, chloro,
--OR.sup.13, --CN, --N(R.sup.13).sub.2 or N.sub.3 with the proviso
that if R.sup.2 is OH and R.sup.3, R.sup.4 and V are hydrogen, then
A is other than S and if R.sup.2 is fluoro or chloro, then R.sup.3
is other than fluoro or chloro.
[0098] In another embodiment, R.sup.2 is C.sub.1-C.sub.3alkyl,
C.sub.1-C.sub.3haloalkyl, C.sub.2-C.sub.6alkenyl or
C.sub.2-C.sub.3alkynyl.
[0099] In another embodiment, R.sup.2 is hydrogen, fluoro, chloro,
methyl, amino or hydroxyl with the proviso that if R.sup.2 is OH
and R.sup.3, R.sup.4 and V are hydrogen, then A is other than S and
if R.sup.2 is fluoro or chloro, then R.sup.3 is other than fluoro
or chloro.
[0100] In another embodiment, R.sup.2 is hydrogen, fluoro or --OH
with the proviso that if R.sup.2 is OH and R.sup.3, R.sup.4 and V
are hydrogen, then A is other than S and if R.sup.2 is fluoro or
chloro, then R.sup.3 is other than fluoro or chloro.
[0101] In another embodiment, R.sup.2 is hydrogen.
[0102] In another embodiment, R.sup.2 is fluoro and R.sup.3 is
other than fluoro or chloro.
[0103] In another embodiment, R.sup.2 is OH and if R.sup.3, R.sup.4
and V are hydrogen, then A is other than S.
[0104] In one embodiment, R.sup.3 is hydrogen, hydroxyl, fluoro,
chloro, N.sub.3, CN or C.sub.1-C.sub.6alkyl, with the proviso that
if R.sup.2 is fluoro or chloro, then R.sup.3 is other than fluoro
or chloro.
[0105] In another embodiment, R.sup.3 is hydrogen, hydroxyl,
fluoro, chloro or methyl with the proviso that if R.sup.2 is fluoro
or chloro, then R.sup.3 is other than fluoro or chloro.
[0106] In another embodiment, R.sup.3 is hydrogen.
[0107] In another embodiment, R.sup.4 is hydrogen,
C.sub.1-C.sub.6alkyl or C.sub.2-C.sub.6alkynyl.
[0108] In another embodiment, R.sup.4 is hydrogen or ethynyl.
[0109] In another embodiment, R.sup.4 is hydrogen.
[0110] In one embodiment, V is hydrogen or fluoro.
[0111] In another embodiment, V is hydrogen.
[0112] In one embodiment, V is hydrogen and each of R.sup.2,
R.sup.3 and R.sup.4 is hydrogen.
[0113] In another embodiment, V is fluoro and each of R.sup.2,
R.sup.3 and R.sup.4 is hydrogen.
[0114] In another embodiment, V is hydrogen, R.sup.2 is fluoro and
each of R.sup.3 and R.sup.4 is hydrogen.
[0115] In another embodiment, V is hydrogen, R.sup.2 is hydroxyl
and each of R.sup.3 and R.sup.4 is hydrogen.
[0116] In one embodiment, A is O, V is hydrogen and each of
R.sup.2, R.sup.3 and R.sup.4 is hydrogen.
[0117] In another embodiment, A is O, V is hydrogen, R.sup.2 is
fluoro and each of R.sup.3 and R.sup.4 is hydrogen.
[0118] In another embodiment, A is O, V is hydrogen, R.sup.2 is
hydroxyl and each of R.sup.3 and R.sup.4 is hydrogen.
[0119] In one embodiment, A is S, V is hydrogen and each of
R.sup.2, R.sup.3 and R.sup.4 is hydrogen.
[0120] In another embodiment, A is S, V is hydrogen, R.sup.2 is
fluoro and each of R.sup.3 and R.sup.4 is hydrogen.
[0121] In one embodiment, B is selected from the following
groups:
##STR00021##
wherein Q, W and R.sup.5 to R.sup.12 are as defined previously.
[0122] In another embodiment, B is selected from the following
groups:
##STR00022##
wherein Q, W and R.sup.5 to R.sup.11 are as defined previously.
[0123] In another embodiment, B is selected from the following
groups:
##STR00023##
wherein Q is O; W is N or CH; R.sup.5, R.sup.6 and R.sup.8 are each
hydrogen; R.sup.7 is amino or NHC(O)aryl; R.sup.9 is hydrogen or
trifluoromethyl; R.sup.10 is --NH.sub.2 or
--O--C.sub.1-C.sub.6alkyl, R.sup.11 is --NH.sub.2 or halogen and
R.sup.12 is --NH.sub.2 or halogen.
[0124] In another embodiment, B is selected from the following
groups:
##STR00024##
wherein Q is O; W is N; R.sup.5, R.sup.6 and R.sup.8 are each
hydrogen; R.sup.7 is amino or NHC(O)aryl; R.sup.9 is hydrogen or
trifluoromethyl; R.sup.10 is --NH.sub.2 or
--O--C.sub.1-C.sub.6alkyl and R.sup.11 is --NH.sub.2 or
halogen.
[0125] In another embodiment, B is
##STR00025##
wherein, Q is O, R.sup.5 and R.sup.6 are each hydrogen and R.sup.7
is amino or NHC(O)R.sup.18, wherein R.sup.18 is
C.sub.1-C.sub.6alkyl or C.sub.6-C.sub.10aryl.
[0126] In another embodiment, B is
##STR00026##
wherein, Q is O, R.sup.5 and R.sup.6 are each hydrogen and R.sup.7
is amino.
[0127] In another embodiment, B is
##STR00027##
wherein, Q is O, R.sup.5 and R.sup.6 are each hydrogen and R.sup.7
is NHC(O)R.sup.18, wherein R.sup.18 is C.sub.1-C.sub.6alkyl or
C.sub.6-C.sub.10aryl.
[0128] In another embodiment, B is:
##STR00028##
wherein Q is O and R.sup.8 is hydrogen and R.sup.9 is hydrogen or
trifluoromethyl.
[0129] In another embodiment, B is:
##STR00029##
wherein W is N, R.sup.10 is amino and R.sup.11 is chloro.
[0130] In another embodiment, B is:
##STR00030##
wherein W is N, R.sup.10 is --OR.sup.18 and R.sup.11 is
--N(R.sup.18).sub.2, wherein R.sup.18 is as previously defined and
each R.sup.18 is selected independently of each other.
[0131] In another embodiment, B is:
##STR00031##
wherein W is N, R.sup.10 is --O-- C.sub.1-C.sub.6alkyl and R.sup.11
is amino. In another embodiment, B is:
##STR00032##
wherein W is N, R.sup.10 is --O-ethyl and R.sup.11 is amino.
[0132] In another embodiment, B is:
##STR00033##
wherein W is N, R.sup.10 is --O-methyl and R.sup.11 is amino.
[0133] In another embodiment, B is:
##STR00034##
wherein W is N, R.sup.10 is amino and R.sup.11 is hydrogen.
[0134] In one embodiment is a compound according to Formula
(I):
##STR00035##
or a pharmaceutically acceptable salt, solvate or enantiomer
thereof, wherein:
[0135] Q is O;
[0136] A is O or S;
[0137] R.sup.1 is
##STR00036##
wherein R.sup.15 is methyl and R.sup.17 is selected from ethyl or
propyl.
[0138] R.sup.2 is F;
[0139] R.sup.3, R.sup.4 and V are H and
[0140] B is
##STR00037##
wherein, R.sup.5 and R.sup.6 are each hydrogen and R.sup.7 is amino
or NHC(O)R.sup.18, wherein R.sup.18 is C.sub.1-C.sub.6alkyl or
C.sub.6-C.sub.10aryl.
[0141] In another embodiment is a compound according to Formula
(I):
##STR00038##
or a pharmaceutically acceptable salt, solvate or enantiomer
thereof, wherein:
[0142] Q is O;
[0143] A is O or S;
[0144] R.sup.1 is
##STR00039##
wherein R.sup.15 is methyl and R.sup.17 is selected from ethyl or
propyl.
[0145] R.sup.2, R.sup.3, R.sup.4 and V are hydrogen and
[0146] B is
##STR00040##
wherein, R.sup.8 is hydrogen and R.sup.9 is hydrogen or
trifluoromethyl.
[0147] In another embodiment is a compound according to Formula
(I):
##STR00041##
or a pharmaceutically acceptable salt, solvate or enantiomer
thereof, wherein:
[0148] Q is O;
[0149] A is O or S;
[0150] R.sup.1 is
##STR00042##
wherein R.sup.15 is methyl and R.sup.17 is selected from ethyl or
propyl.
[0151] R.sup.2 is hydrogen or fluorine;
[0152] R.sup.3, R.sup.4 and V are hydrogen and
[0153] B is
##STR00043##
wherein W is N, R.sup.10 is amino and R.sup.11 is chloro.
[0154] In another embodiment is a compound according to Formula
(I):
##STR00044##
or a pharmaceutically acceptable salt, solvate or enantiomer
thereof, wherein:
[0155] Q is O;
[0156] A is O;
[0157] R.sup.1 is
##STR00045##
wherein R.sup.15 is methyl and R.sup.17 is selected from ethyl or
propyl.
[0158] R.sup.2 is hydroxy;
[0159] R.sup.3, R.sup.4 and V are hydrogen and
[0160] B is
##STR00046##
wherein W is N, R.sup.10 is methoxy and R.sup.11 is amino.
[0161] In another embodiment is a compound of Formula I having the
structure:
##STR00047## ##STR00048##
[0162] or a pharmaceutically acceptable salt or solvate thereof,
wherein B and R.sup.17 are as defined previously.
[0163] In another embodiment is a compound of Formula I having the
structure:
##STR00049##
[0164] or a pharmaceutically acceptable salt or solvate thereof,
wherein R.sup.17 is as defined previously.
[0165] In another embodiment is a compound of formula (I) which is
selected from:
##STR00050## ##STR00051## ##STR00052## ##STR00053##
or a pharmaceutically acceptable salt or solvate thereof.
[0166] In one embodiment, variables A, B, R.sup.1, R.sup.2,
R.sup.3, Q and V for the Compounds of Formula (I) are selected
independently of each other.
[0167] In another embodiment, the Compounds of Formula (I) are in
substantially purified form.
The Compounds of Formula (II)
[0168] The present invention provides compounds of Formula (II),
having the formula:
##STR00054##
or a pharmaceutically acceptable salt or solvate thereof, wherein
B, R.sup.1 and Q are defined above for the compounds of Formula
(II).
[0169] In one embodiment, Q is O.
[0170] In another embodiment, Q is S.
[0171] In one embodiment, R.sup.1 is
--CH.sub.2--C(R.sup.14).sub.2--C(R.sup.15).sub.2--C(O)OR.sup.17,
wherein R.sup.14, R.sup.15 and R.sup.17 are as previously
defined.
[0172] In one embodiment, R.sup.1 is
--CH.sub.2--C(R.sup.14).sub.2--C(R.sup.15).sub.2--C(O)OR.sup.17,
wherein R.sup.14, R.sup.15 and R.sup.17 are each independently
selected from hydrogen and C.sub.1-C.sub.6alkyl.
[0173] In another embodiment, R.sup.1 is
--CH.sub.2--CH.sub.2--C(R.sup.15).sub.2--C(O)OR.sup.17, wherein
R.sup.15 and R.sup.17 are as previously defined.
[0174] In another embodiment, R.sup.1 is
##STR00055##
wherein R.sup.15 and R.sup.17 are as previously defined.
[0175] In another embodiment, R.sup.1 is
##STR00056##
wherein R.sup.15 and R.sup.17 are each independently selected from
C.sub.1-C.sub.6alkyl.
[0176] In another embodiment, R.sup.1 is
##STR00057##
wherein R.sup.15 and R.sup.17 are each independently selected from
C.sub.1-C.sub.6alkyl.
[0177] In another embodiment, R.sup.1 is
##STR00058##
wherein R.sup.15 and R.sup.17 are each independently selected from
C.sub.1-C.sub.6alkyl.
[0178] In another embodiment, R.sup.1 is
##STR00059##
wherein R.sup.15 is methyl and R.sup.17 is selected from ethyl or
propyl.
[0179] In another embodiment, R.sup.1 is
##STR00060##
wherein R.sup.15 is methyl and R.sup.17 is selected from ethyl or
propyl.
[0180] In another embodiment, R.sup.1 is
##STR00061##
wherein R.sup.15 is methyl and R.sup.17 is selected from ethyl or
propyl.
[0181] In another embodiment, R.sup.1 is
##STR00062##
[0182] In another embodiment, R.sup.1 is
##STR00063##
[0183] In another embodiment, R.sup.1 is
##STR00064##
[0184] In another embodiment, R.sup.1 is
##STR00065##
[0185] In one embodiment, B is selected from the following
groups:
##STR00066##
wherein Q, W and R.sup.5 to R.sup.12 are as defined previously.
[0186] In another embodiment, B is selected from the following
groups:
##STR00067##
wherein Q, W and R.sup.5 to R.sup.11 are as defined previously.
[0187] In another embodiment, B is selected from the following
groups:
##STR00068##
wherein Q is O; W is N or CH; R.sup.5, R.sup.6 and R.sup.8 are each
hydrogen; R.sup.7 is amino; R.sup.9 is hydrogen or trifluoromethyl;
R.sup.10 is --NH.sub.2 or --O--C.sub.1-C.sub.6alkyl, R.sup.11 is
--NH.sub.2 or halogen and R.sup.12 is --NH.sub.2 or halogen.
[0188] In another embodiment, B is selected from the following
groups:
##STR00069##
wherein Q is O; W is N; R.sup.5, R.sup.6 and R.sup.8 are each
hydrogen; R.sup.7 is amino; R.sup.9 is hydrogen or trifluoromethyl;
R.sup.10 is --NH.sub.2 or --O--C.sub.1-C.sub.6alkyl and R.sup.11 is
--NH.sub.2 or halogen.
[0189] In another embodiment, B is
##STR00070##
wherein, Q is O, R.sup.5 and R.sup.6 are each hydrogen and R.sup.7
is amino or NHC(O)R.sup.18, wherein R.sup.18 is
C.sub.1-C.sub.6alkyl or is C.sub.6-C.sub.10aryl
[0190] In another embodiment, B is
##STR00071##
wherein, Q is O, R.sup.5 and R.sup.6 are each hydrogen and R.sup.7
is amino.
[0191] In another embodiment, B is
##STR00072##
wherein, Q is O, R.sup.5 and R.sup.6 are each hydrogen and R.sup.7
is NHC(O)R.sup.18, wherein R.sup.18 is C.sub.1-C.sub.6alkyl or is
C.sub.6-C.sub.10aryl
[0192] In another embodiment, B is:
##STR00073##
wherein Q is O and R.sup.8 is hydrogen and R.sup.9 is hydrogen or
trifluoromethyl.
[0193] In another embodiment, B is:
##STR00074##
wherein W is N, R.sup.10 is amino and R.sup.11 is chloro.
[0194] In another embodiment, B is:
##STR00075##
wherein W is N, R.sup.10 is --OR.sup.18 and R.sup.11 is
--N(R.sup.18).sub.2, wherein R.sup.18 is as previously defined and
each R.sup.18 is selected independently of each other.
[0195] In another embodiment, B is:
##STR00076##
wherein W is N, R.sup.10 is --O-- C.sub.1-C.sub.6alkyl and R.sup.11
is amino, wherein R.sup.18 is as previously defined and each
R.sup.18 is selected independently of each other.
[0196] In another embodiment, B is:
##STR00077##
wherein W is N, R.sup.10 is --O-ethyl and R.sup.11 is amino,
wherein R.sup.18 is as previously defined and each R.sup.18 is
selected independently of each other.
[0197] In one embodiment is a compound according to Formula
(II):
##STR00078##
or a pharmaceutically acceptable salt or solvate thereof,
wherein:
[0198] Q is O;
[0199] R.sup.1 is
##STR00079##
wherein R.sup.15 is methyl and R.sup.17 is selected from ethyl or
propyl and
[0200] B is
##STR00080##
wherein, R.sup.10 is --OR.sup.18 and R.sup.11 is NHR.sup.18,
wherein each R.sup.18 is independently hydrogen or
C.sub.1-C.sub.6alkyl.
[0201] In another embodiment is a compound according to Formula
(II):
##STR00081##
or a pharmaceutically acceptable salt or solvate thereof,
wherein:
[0202] Q is O;
##STR00082##
wherein R.sup.15 is methyl and R.sup.17 is selected from ethyl or
propyl and
[0203] B is
##STR00083##
wherein, R.sup.10 is --O-ethyl and R.sup.11 is amino.
[0204] In another embodiment is a compound of formula (II) which is
selected from:
##STR00084##
or a pharmaceutically acceptable salt or solvate thereof.
[0205] In one embodiment, variables B, R.sup.1 and Q for the
compounds of Formula (II) are selected independently of each
other.
[0206] In another embodiment, the compounds of Formula (II) are in
substantially purified form.
General Methods for Making the Compounds of Formula (I)
[0207] The compounds of Formula (I) or Formula (II) may be prepared
from known or readily prepared starting materials, following
methods known to one skilled in the art of organic synthesis.
Methods useful for making the compounds of Formula (I) or Formula
(II) are set forth in the Examples below and generalized in Schemes
A, B and C below. Alternative synthetic pathways and analogous
structures will be apparent to those skilled in the art of organic
synthesis.
[0208] The following abbreviations are used herein:
[0209] CH.sub.3CN or ACN acetonitrile
[0210] DBU 1,8-diazabicyclo(5.4.0)undec-7-ene
[0211] DCM dichloromethane
[0212] DMF N,N-dimethylformamide
[0213] Et.sub.3N triethylamine
[0214] Et.sub.2O diethylether
[0215] NMI N-methylimidazole
[0216] t-bu tert-butyl
[0217] THF tetrahydrofuran
[0218] Scheme A shows a method useful for making nucleoside
compounds of formula A4, which correspond to the compounds of
Formula (I).
##STR00085##
[0219] Phosphorus (V) oxychloride can be reacted with triethylamine
and an alcohol of formula R.sup.1OH as shown to provide a compound
of formula A2. The compound of formula A2 is then reacted as shown
with a nucleoside of formula A3 to provide a cyclic phosphate
nucleoside prodrug of formula A4.
[0220] In Scheme A, the variables are as described in the context
of Formula I. Nucleosides A3 can be prepared or obtained according
to the knowledge in the art. Additional steps and reagents not
provided in Scheme A would be known to those of skill in the art.
Exemplary methods of preparation are described in detail in the
Examples herein.
[0221] Scheme B shows an alternative method useful for making
nucleoside compounds of formula A4, which correspond to the
compounds of Formula (I).
##STR00086##
[0222] 1-Chloro-N,N,N',N'-tetraisopropylphosphinediamine (B1) can
be reacted as shown with triethylamine and an alcohol of formula
R.sup.1OH to provide a compound of formula B2. The compound of
formula B2 is then reacted as shown with a nucleoside of formula A3
to provide a cyclic phosphate nucleoside prodrug of formula A4.
[0223] In Scheme B, the variables are as described in the context
of Formula I. Nucleosides A3 can be prepared or obtained according
to the knowledge in the art. Additional steps and reagents not
provided in Scheme A would be known to those of skill in the art.
Exemplary methods of preparation are described in detail in the
Examples herein.
[0224] Scheme C shows an alternative method useful for making
nucleoside compounds of formula A4, which correspond to the
compounds of Formula (I).
##STR00087##
2,3,4,5,6-Pentafluorophenol (C2) can be reacted with 4-chlorophenyl
phosphodichloridate or 4-nitrophenyl phosphodichloridate (C1),
triethylamine and an alcohol of formula R.sup.1OH as shown to
provide a compound of formula C3. The compound of formula C3 is
then reacted with a nucleoside of formula A3 as shown to provide a
cyclic phosphate nucleoside prodrug of formula A4.
[0225] In Scheme C, the variables are as described in the context
of Formula I. Nucleosides A3 can be prepared or obtained according
to the knowledge in the art. Additional steps and reagents not
provided in Scheme A would be known to those of skill in the art.
Exemplary methods of preparation are described in detail in the
Examples herein.
[0226] Compounds of Formula II can be prepared by analogous
processes to those shown in Schemes A to C using the nucleoside A5
in place of nucleoside A3.
##STR00088##
Further Embodiments of the Compounds of Formula (I)
[0227] Prodrugs and solvates of the compounds of the invention are
also contemplated herein. A discussion of prodrugs is provided in
T. Higuchi and V. Stella, Pro-drugs as Novel Delivery Systems
(1987) 14 of the A.C.S. Symposium Series, and in Bioreversible
Carriers in Drug Design, (1987) Edward B. Roche, ed., American
Pharmaceutical Association and Pergamon Press. The term "prodrug"
means a compound (e.g., a drug precursor) that is transformed in
vivo to provide a Compound of Formula (I) or Formula (II), or a
pharmaceutically acceptable salt of the compound. The
transformation may occur by various mechanisms (e.g., by metabolic
or chemical processes), such as, for example, through hydrolysis in
blood.
[0228] For example, if a Compound of Formula (I) or Formula (II)
contains a carboxylic acid functional group, a prodrug can comprise
an ester formed by the replacement of the hydrogen atom of the acid
group with a group such as, for example, (C.sub.1-C.sub.8)alkyl,
(C.sub.2-C.sub.12)alkanoyloxymethyl, 1-(alkanoyloxy)ethyl having
from 4 to 9 carbon atoms, 1-methyl-1-(alkanoyloxy)-ethyl having
from 5 to 10 carbon atoms, alkoxycarbonyloxymethyl having from 3 to
6 carbon atoms, 1-(alkoxycarbonyloxy)ethyl having from 4 to 7
carbon atoms, 1-methyl-1-(alkoxycarbonyloxy)ethyl having from 5 to
8 carbon atoms, N-(alkoxycarbonyl)aminomethyl having from 3 to 9
carbon atoms, 1-(N-(alkoxycarbonyl)amino)ethyl having from 4 to 10
carbon atoms, 3-phthalidyl, 4-crotonolactonyl,
gamma-butyrolacton-4-yl,
di-N,N--(C.sub.1-C.sub.2)alkylamino(C.sub.2-C.sub.3)alkyl (such as
.beta.-dimethylaminoethyl), carbamoyl-(C.sub.1-C.sub.2)alkyl,
N,N-di(C.sub.1-C.sub.2)alkylcarbamoyl-(C.sub.1-C.sub.2)alkyl and
piperidino-, pyrrolidino- or morpholino(C.sub.2-C.sub.3)alkyl, and
the like.
[0229] Similarly, if a Compound of Formula (I) or Formula (II)
contains an alcohol functional group, a prodrug can be formed by
the replacement of one or more of the hydrogen atoms of the alcohol
groups with a group such as, for example,
(C.sub.1-C.sub.6)alkanoyloxymethyl,
1-((C.sub.1-C.sub.6)alkanoyloxy)ethyl,
1-methyl-1-((C.sub.1-C.sub.6)alkanoyloxy)ethyl,
(C.sub.1-C.sub.6)alkoxycarbonyloxymethyl,
N--(C.sub.1-C.sub.6)alkoxycarbonylaminomethyl, succinoyl,
(C.sub.1-C.sub.6)alkanoyl, .alpha.-amino(C.sub.1-C.sub.4)alkyl,
.alpha.-amino(C.sub.1-C.sub.4)alkylene-aryl, arylacyl and
.alpha.-aminoacyl, or .alpha.-aminoacyl-.alpha.-aminoacyl, where
each .alpha.-aminoacyl group is independently selected from the
naturally occurring L-amino acids, or glycosyl (the radical
resulting from the removal of a hydroxyl group of the hemiacetal
form of a carbohydrate). Other example of alcohol-derived prodrugs
include --P(O)(OH).sub.2; --P(O)(--O--C.sub.1-C.sub.6alkyl).sub.2;
--P(O)(--NH-(.alpha.-aminoacyl group))(-O-aryl);
--P(O)(--O--(C.sub.1-C.sub.6 alkylene)-S-acyl)(-NH-arylalkyl); and
those described in U.S. Pat. No. 7,879,815; International
Publication Nos. WO2005/003047, WO2008/082602, WO2010/0081628,
WO2010/075517 and WO2010/075549; Mehellou, Chem. Med. Chem.,
5:1841-1842 (2005); Bobeck et al., Antiviral Therapy 15:935-950
(2010); Furman et al., Future Medicinal Chemistry, 1:1429-1452
(2009); and Erion, Microsomes and Drug Oxidations, Proceedings of
the International Symposium, 17th, Saratoga Springs, N.Y., United
States, Jul. 6-10, 2008, 7-12 (2008).
[0230] If a Compound of Formula (I) or Formula (II) incorporates an
amine functional group, a prodrug can be formed by the replacement
of a hydrogen atom in the amine group with a group such as, for
example, R-carbonyl-, RO-carbonyl-, NRR'-carbonyl- wherein R and R'
are each independently (C.sub.1-C.sub.10)alkyl, (C.sub.3-C.sub.7)
cycloalkyl, benzyl, a natural .alpha.-aminoacyl,
--C(OH)C(O)OY.sup.1 wherein Y.sup.1 is H, (C.sub.1-C.sub.6)alkyl or
benzyl, --C(OY.sup.2)Y.sup.3 wherein Y.sup.2 is (C.sub.1-C.sub.4)
alkyl and Y.sup.3 is (C.sub.1-C.sub.6)alkyl; carboxy
(C.sub.1-C.sub.6)alkyl; amino(C.sub.1-C.sub.4)alkyl or mono-N- or
di-N,N--(C.sub.1-C.sub.6)alkylaminoalkyl; --C(Y.sup.4)Y.sup.5
wherein Y.sup.4 is H or methyl and Y.sup.5 is mono-N- or
di-N,N--(C.sub.1-C.sub.6)alkylamino morpholino; piperidin-1-yl or
pyrrolidin-1-yl, and the like.
[0231] Pharmaceutically acceptable esters of the present compounds
include the following groups: (1) carboxylic acid esters obtained
by esterification of the hydroxy group of a hydroxyl compound, in
which the non-carbonyl moiety of the carboxylic acid portion of the
ester grouping is selected from straight or branched chain alkyl
(e.g., methyl, ethyl, n-propyl, isopropyl, t-butyl, sec-butyl or
n-butyl), alkoxyalkyl (e.g., methoxymethyl), aralkyl (e.g.,
benzyl), aryloxyalkyl (for example, phenoxymethyl), aryl (e.g.,
phenyl optionally substituted with, for example, halogen,
C.sub.1-4alkyl, --O--(C.sub.1-4alkyl) or amino); (2) sulfonate
esters, such as alkyl- or aralkylsulfonyl (for example,
methanesulfonyl); (3) amino acid esters (e.g., L-valyl or
L-isoleucyl); (4) phosphonate esters and (5) mono-, di- or
triphosphate esters. The phosphate esters may be further esterified
by, for example, a C.sub.1-20 alcohol or reactive derivative
thereof, or by a 2,3-di (C.sub.6-24)acyl glycerol.
[0232] One or more compounds of the invention may exist in
unsolvated as well as solvated forms with pharmaceutically
acceptable solvents such as water, ethanol, and the like, and it is
intended that the invention embrace both solvated and unsolvated
forms. "Solvate" means a physical association of a compound of this
invention with one or more solvent molecules. This physical
association involves varying degrees of ionic and covalent bonding,
including hydrogen bonding. In certain instances the solvate will
be capable of isolation, for example when one or more solvent
molecules are incorporated in the crystal lattice of the
crystalline solid. "Solvate" encompasses both solution-phase and
isolatable solvates. Non-limiting examples of solvates include
ethanolates, methanolates, and the like. A "hydrate" is a solvate
wherein the solvent molecule is water.
[0233] One or more compounds of the invention may optionally be
converted to a solvate. Preparation of solvates is generally known.
Thus, for example, M. Caira et al, J. Pharmaceutical Sci., 93(3),
601-611 (2004) describe the preparation of the solvates of the
antifungal fluconazole in ethyl acetate as well as from water.
Similar preparations of solvates, hemisolvate, hydrates and the
like are described by E. C. van Tonder et al, AAPS Pharm Sci
Techours., 501), article 12 (2004); and A. L. Bingham et al, Chem.
Commun., 603-604 (2001). A typical process involves dissolving the
inventive compound in desired amounts of the desired solvent
(organic or water or mixtures thereof) at a higher than room
temperature, and cooling the solution at a rate sufficient to form
crystals which are then isolated by standard methods. Analytical
techniques such as, for example IR spectroscopy, show the presence
of the solvent (or water) in the crystals as a solvate (or
hydrate).
[0234] The compounds of Formula (I) or Formula (II) can form salts
which are also within the scope of this invention. The term
"salt(s)", as employed herein, denotes acidic salts formed with
inorganic and/or organic acids, as well as basic salts formed with
inorganic and/or organic bases. In addition, when a compound of
Formula (I) or Formula (II) contains both a basic moiety, such as,
a pyridine or imidazole, and an acidic moiety, such as, a
carboxylic acid, zwitterions ("inner salts") may be formed and are
included within the term "salt(s)" as used herein. In one
embodiment, the salt is a pharmaceutically acceptable (i.e.,
non-toxic, physiologically acceptable) salt. In another embodiment,
the salt is other than a pharmaceutically acceptable salt. Salts of
the compounds of Formula (I) or Formula (II) may be formed, for
example, by reacting a compound of Formula (I) or Formula (II) with
an amount of acid or base, such as an equivalent amount, in a
medium such as one in which the salt precipitates or in an aqueous
medium followed by lyophilization.
[0235] Exemplary acid addition salts include acetates, ascorbates,
benzoates, benzenesulfonates, bisulfates, borates, butyrates,
citrates, camphorates, camphorsulfonates, fumarates,
hydrochlorides, hydrobromides, hydroiodides, lactates, maleates,
methanesulfonates, naphthalenesulfonates, nitrates, oxalates,
phosphates, propionates, salicylates, succinates, sulfates,
tartarates, thiocyanates, toluenesulfonates (also known as
tosylates) and the like. Additionally, acids which are generally
considered suitable for the formation of pharmaceutically useful
salts from basic pharmaceutical compounds are discussed, for
example, by P. Stahl et al, Camille G. (eds.) Handbook of
Pharmaceutical Salts. Properties, Selection and Use. (2002) Zurich:
Wiley-VCH; S. Berge et al, Journal of Pharmaceutical Sciences
(1977) 66(1) 1-19; P. Gould, International J. of Pharmaceutics
(1986) 33 201-217; Anderson et al, The Practice of Medicinal
Chemistry (1996), Academic Press, New York; and in The Orange Book
(Food & Drug Administration, Washington, D.C. on their
website). These disclosures are incorporated herein by reference
thereto.
[0236] Exemplary basic salts include ammonium salts, alkali metal
salts such as sodium, lithium, and potassium salts, alkaline earth
metal salts such as calcium and magnesium salts, salts with organic
bases (for example, organic amines) such as dicyclohexylamine,
t-butyl amine, choline, and salts with amino acids such as
arginine, lysine and the like. Basic nitrogen-containing groups may
be quarternized with agents such as lower alkyl halides (e.g.,
methyl, ethyl, and butyl chlorides, bromides and iodides), dialkyl
sulfates (e.g., dimethyl, diethyl, and dibutyl sulfates), long
chain halides (e.g., decyl, lauryl, and stearyl chlorides, bromides
and iodides), aralkyl halides (e.g., benzyl and phenethyl
bromides), and others.
[0237] All such acid salts and base salts are intended to be
pharmaceutically acceptable salts within the scope of the invention
and all acid and base salts are considered equivalent to the free
forms of the corresponding compounds for purposes of the
invention.
[0238] All stereoisomers (for example, geometric isomers, optical
isomers and the like) of the present compounds (including those of
the salts and solvates of the compounds), such as those which may
exist due to asymmetric carbons on various substituents, including
enantiomeric forms (which may exist even in the absence of
asymmetric carbons), rotameric forms, atropisomers (e.g.,
substituted biaryls) and diastereomeric forms, are contemplated
within the scope of this invention. If a compound of Formula (I) or
Formula (II) incorporates a double bond or a fused ring, both the
cis- and trans-forms, as well as mixtures, are embraced within the
scope of the invention.
[0239] It is also possible that the compounds of Formula (I) or
Formula (II) may exist in different tautomeric forms, and all such
forms are embraced within the scope of the invention. For example,
all keto-enol and imine-enamine forms of the compounds are included
in the invention.
[0240] Individual stereoisomers of the compounds of the invention
may, for example, be substantially free of other isomers, or may be
admixed, for example, as racemates or with all other, or other
selected, stereoisomers. The chiral centers of the present
invention can have the S or R configuration as defined by the IUPAC
1974 Recommendations. The use of the terms "salt", "solvate" and
the like, is intended to apply equally to the salt and solvate of
enantiomers, stereoisomers, rotamers, tautomers, positional
isomers, racemates or prodrugs of the inventive compounds.
[0241] Diastereomeric mixtures may be separated into their
individual diastereomers on the basis of their physical chemical
differences by methods well-known to those skilled in the art, such
as, for example, by chromatography and/or fractional
crystallization. Enantiomers may be separated by converting the
enantiomeric mixture into a diastereomeric mixture by reaction with
an appropriate optically active compound (e.g., chiral auxiliary
such as a chiral alcohol or Mosher's acid chloride), separating the
diastereomers and converting (e.g., hydrolyzing) the individual
diastereomers to the corresponding pure enantiomers.
Sterochemically pure compounds may also be prepared by using chiral
starting materials or by employing salt resolution techniques.
Enantiomers can also be directly separated using chiral
chromatographic techniques.
[0242] In the compounds of Formula (I) or Formula (II), the atoms
may exhibit their natural isotopic abundances, or one or more of
the atoms may be artificially enriched in a particular isotope
having the same atomic number, but an atomic mass or mass number
different from the atomic mass or mass number predominantly found
in nature. The present invention is meant to include all suitable
isotopic variations of the compounds of generic Formula (I) or
Formula (II). For example, different isotopic forms of hydrogen (H)
include protium (.sup.1H) and deuterium (.sup.2H). Protium is the
predominant hydrogen isotope found in nature. Enriching for
deuterium may afford certain therapeutic advantages, such as
increasing in vivo half-life or reducing dosage requirements, or
may provide a compound useful as a standard for characterization of
biological samples. Isotopically-enriched compounds of Formula (I)
or Formula (II) may be prepared without undue experimentation by
conventional techniques well known to those skilled in the art or
by processes analogous to those described in the Schemes and
Examples herein using appropriate isotopically-enriched reagents
and/or intermediates. In one embodiment, a Compound of Formula (I)
or Formula (II) has one or more of its hydrogen atoms replaced with
deuterium.
[0243] Polymorphic forms of the compounds of Formula (I) or Formula
(II), and of the salts or solvates of the Compounds of Formula (I)
or Formula (II), are intended to be included in the present
invention.
[0244] The compounds provided herein are based, at least in part,
on the discovery that the cyclic phosphate prodrugs can provide
superior human pharmacokinetics including superior accumulation of
active nucleoside and nucleotide analogs in target cells, such as
liver cells. Any compound provided herein is preferably in the form
of a composition that is substantially free of other stereoisomers
of the compound, as described herein.
[0245] Other embodiments of the present invention include the
following:
[0246] (a) A pharmaceutical composition comprising an effective
amount of a compound of Formula (I) or Formula (II), or a
pharmaceutically acceptable salt, solvate or enantiomer thereof,
and a pharmaceutically acceptable carrier or diluent.
[0247] (b) The pharmaceutical composition of (a), further
comprising a second therapeutic agent selected from the group
consisting of immunomodulators, anti-infective agents and
anticancer agents.
[0248] (c) The pharmaceutical composition of (b), wherein the
anti-infective agent is an antiviral, such as an HBV inhibitor.
[0249] (d) A combination comprising a compound of Formula (I) or
Formula (II), or a pharmaceutically acceptable salt, solvate or
enantiomer thereof and one, two, three or more other therapeutic
agents.
[0250] (e) The combination of (d) that is (i) a Compound of Formula
(I) or (II), or a pharmaceutically acceptable salt, solvate or
enantiomer thereof, and (ii) a second therapeutic agent selected
from the group consisting of immunomodulators, and anti-infective
agents; wherein the Compound of Formula (I) or (II) and the second
therapeutic agent are each employed in an amount that renders the
combination effective for inhibiting HBV replication, or for
treating HBV infection and/or reducing the likelihood or severity
of symptoms of HBV infection.
[0251] (f) The combination of (d), wherein the anti-infective agent
is an antiviral such as an HBV inhibitor.
[0252] (g) The combination of (d) that is (i) a Compound of Formula
(I) or (II), or a pharmaceutically acceptable salt, solvate or
enantiomer thereof, and (ii) a second therapeutic agent which is an
anticancer agent; wherein the Compound of Formula (I) or (II) and
the second therapeutic agent are each employed in an amount that
renders the combination effective for treating and/or reducing the
likelihood or severity of symptoms of cancer.
[0253] (h) A Compound of Formula (I) or (II), or a pharmaceutically
acceptable salt, solvate or enantiomer thereof for use in
therapy.
[0254] (i) The Compound for use of (h), wherein the therapy is
treating cancer and/or reducing the likelihood or severity of
symptoms of cancer, for example liver cancer, breast cancer,
ovarian cancer, lung cancer, pancreatic cancer, cholangiocarcinoma,
biliary tract cancer, hepatocellular carcinoma or leukemia, in a
subject.
[0255] (j) The Compound for use of (h), wherein the therapy is
treating HBV infection and/or reducing the likelihood or severity
of symptoms of HBV infection in a subject.
[0256] (k) The Compound for use of (h), wherein the therapy is
treating Ebolavirus infection and/or reducing the likelihood or
severity of symptoms of Ebolavirus infection in a subject.
[0257] (l) The Compound for use of (h), (i), (j) or (k), wherein
said compound is administered in combination with one or more other
therapeutic agents.
[0258] (m) A method of treating or preventing cancer and/or
reducing the likelihood or severity of symptoms of cancer, for
example liver cancer, breast cancer, ovarian cancer, lung cancer,
pancreatic cancer, cholangiocarcinoma, biliary tract cancer,
hepatocellular carcinoma or leukemia, in a subject in need thereof
which comprises administering to the subject an effective amount of
a compound of Formula (I) of Formula (II) or a pharmaceutically
acceptable salt, solvate or enantiomer thereof.
[0259] (n) A method of treating or preventing cancer and/or
reducing the likelihood or severity of symptoms of cancer, for
example liver cancer, breast cancer, ovarian cancer, lung cancer,
pancreatic cancer, cholangiocarcinoma, biliary tract cancer,
hepatocellular carcinoma or leukemia, in a subject in need thereof
which comprises administering to the subject the pharmaceutical
composition of (a), (b) or (c) or the combination of (d) or
(g).
[0260] (o) A method of inhibiting HBV replication in a subject in
need thereof which comprises administering to the subject an
effective amount of a compound of Formula (I) or Formula (II) or a
pharmaceutically acceptable salt, solvate or enantiomer
thereof.
[0261] (p) A method of treating HBV infection and/or reducing the
likelihood or severity of symptoms of HBV infection in a subject in
need thereof which comprises administering to the subject an
effective amount of a compound of Formula (I) of Formula (II) or a
pharmaceutically acceptable salt, solvate or enantiomer
thereof.
[0262] (q) A method of treating Ebolavirus infection and/or
reducing the likelihood or severity of symptoms of Ebolavirus
infection in a subject in need thereof which comprises
administering to the subject an effective amount of a compound of
Formula (I) of Formula (II) or a pharmaceutically acceptable salt,
solvate or enantiomer thereof.
[0263] (r) The method of (o), (p) or (q) wherein the compound of
Formula (I) or Formula (II) is administered in combination with an
effective amount of at least one second therapeutic agent selected
from the group consisting of immunomodulators, and anti-infective
agents.
[0264] (s) The method of (r), wherein the anti-infective agent is
an antiviral, such as an HBV inhibitor.
[0265] (t) A method of inhibiting HBV replication in a subject in
need thereof which comprises administering to the subject the
pharmaceutical composition of (a), (b) or (c) or the combination of
(d), (e) or (f).
[0266] (u) A method of treating HBV infection and/or reducing the
likelihood or severity of symptoms of HBV infection in a subject in
need thereof which comprises administering to the subject the
pharmaceutical composition of (a), (b) or (c) or the combination of
(d), (e) or (f).
[0267] (v) A method of treating Ebolavirus infection and/or
reducing the likelihood or severity of symptoms of Ebolavirus
infection in a subject in need thereof which comprises
administering to the subject the pharmaceutical composition of (a),
(b) or (c) or the combination of (d).
[0268] Additional embodiments of the invention include the
pharmaceutical compositions, combinations and methods set forth in
(a)-(v) above and the uses set forth in the discussion below,
wherein the compound of the present invention employed therein is a
compound of one of the embodiments, aspects, classes, sub-classes,
or features of the compounds described above. In all of these
embodiments, the compound may optionally be used in the form of a
pharmaceutically acceptable salt or solvate as appropriate. It is
understood that references to compounds would include the compound
in its present form as well as in different forms, such as
polymorphs and solvates as applicable.
[0269] It is further to be understood that the embodiments of
compositions and methods provided as (a) through (v) above are
understood to include all embodiments of the compounds, including
such embodiments as result from combinations of embodiments.
Treatment or Prevention of Cancer
[0270] Compounds of Formula (I) and Formula (II) are further useful
in the treatment or prevention of cancer and/or reducing the
likelihood or severity of symptoms of cancer, In one embodiment,
the cancer is selected from liver cancer, breast cancer, ovarian
cancer, lung cancer, pancreatic cancer or leukemic cancer. In a
further embodiment the cancer is liver cancer. Liver cancers which
can be treated include primary and secondary liver cancers. In
particular embodiments, the liver cancer is hepatocellular
carcinoma, cholangiocarcinoma, or biliary tract cancer. In
particular embodiments, the liver cancer is a metastasis in the
liver originated from other cancers (such as colon cancer,
pancreatic cancer, etc).
[0271] Accordingly, the invention also provides methods for
treating or preventing cancer and/or reducing the likelihood or
severity of symptoms of cancer in a patient, the methods comprising
administering to the patient an effective amount of at least one
compound of Formula (I) or Formula (II) or a pharmaceutically
acceptable salt, solvate or enantiomer thereof.
[0272] In further embodiments, the cancers which can be treated by
the compounds described herein include, but are not limited to,
Acute Lymphoblastic Leukemia; Acute Myeloid Leukemia;
Adrenocortical Carcinoma; AIDS-Related Lymphoma; AIDS-Related
Malignancies; Anal Cancer; Astrocytoma; Bile Duct Cancer; Bladder
Cancer; Bone Cancer, Osteosarcoma/Malignant Fibrous Histiocytoma;
Brain Stem Glioma; Brain Tumor, Cerebellar Astrocytoma; Brain
Tumor, Cerebral Astrocytoma/Malignant Glioma; Brain Tumor,
Ependymoma; Brain Tumor, Medulloblastoma; Brain Tumor,
Supratentorial Primitive Neuroectodermal Tumors; Brain Tumor,
Visual Pathway and Hypothalamic Glioma; Breast Cancer; Bronchial
Adenomas/Carcinoids; Carcinoid Tumor; Carcinoid Tumor,
Gastrointestinal; Carcinoma, Adrenocortical; Carcinoma, Islet Cell;
Central Nervous System Lymphoma, Primary; Cerebral
Astrocytoma/Malignant Glioma; Cervical Cancer; Cholangiocarcinoma;
Chronic Lymphocytic Leukemia; Chronic Myelogenous Leukemia; Chronic
Myeloproliferative Disorders; Clear Cell Sarcoma of Tendon Sheaths;
Colon Cancer; Colorectal Cancer; Cutaneous T-Cell Lymphoma;
Endometrial Cancer; Ependymoma; Epithelial Cancer, Ovarian;
Esophageal Cancer; Esophageal Cancer; Ewing's Family of Tumors;
Extracranial Germ Cell Tumor; Extrahepatic Bile Duct Cancer; Eye
Cancer, Intraocular Melanoma; Eye Cancer, Retinoblastoma;
Gallbladder Cancer; Gastric (Stomach) Cancer; 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; Hodgkin's
Lymphoma; Hypopharyngeal Cancer; Hypothalamic and Visual Pathway
Glioma; Intraocular Melanoma; Islet Cell Carcinoma (Endocrine
Pancreas); Kaposi's Sarcoma; Kidney Cancer; Laryngeal Cancer;
Leukemia, Acute Lymphoblastic; Leukemia, Acute Myeloid; Leukemia,
Chronic Lymphocytic; Leukemia, Chronic Myelogenous; Leukemia, Hairy
Cell; Lip and Oral Cavity Cancer; Liver Cancer; Lung Cancer,
Non-Small Cell; Lung Cancer, Small Cell; Lymphoblastic Leukemia;
Lymphoma, AIDS-Related; Lymphoma, Central Nervous System (Primary);
Lymphoma, Cutaneous T-Cell; Lymphoma, Hodgkin's; Lymphoma,
Hodgkin's During Pregnancy; Lymphoma, Non-Hodgkin's; Lymphoma,
Primary Central Nervous System; Macroglobulinemia, Waldenstrom's;
Male Breast Cancer; Malignant Mesothelioma; 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; Myeloma, Multiple; Myeloproliferative
Disorders, Chronic; Nasal Cavity and Paranasal Sinus Cancer;
Nasopharyngeal Cancer; Neuroblastoma; Non-Hodgkin's Lymphoma;
Non-Small Cell Lung Cancer; Oral Cancer; Oral Cavity and Lip
Cancer; Oropharyngeal Cancer; steosarcoma/Malignant Fibrous
Histiocytoma of Bone; Ovarian Epithelial Cancer; Ovarian Germ Cell
Tumor; Ovarian Low Malignant Potential Tumor; Pancreatic Cancer;
Paranasal Sinus and Nasal Cavity Cancer; Parathyroid Cancer; Penile
Cancer; Pheochromocytoma; Pineal and Supratentorial Primitive
Neuroectodermal Tumors; 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; Prostate Cancer; Rectal Cancer; Renal Cell
(Kidney) Cancer; Renal Pelvis and Ureter, Transitional Cell Cancer;
Retinoblastoma; Rhabdomyosarcoma; Salivary Gland Cancer; Sarcoma,
Ewing's Family of Tumors; Sarcoma, Kaposi's; Sarcoma
(Osteosarcoma)/Malignant Fibrous Histiocytoma of Bone; Sarcoma,
Soft Tissue; Sezary Syndrome; Skin Cancer; Skin Cancer (Melanoma);
Skin Carcinoma, Merkel Cell; Small Cell Lung Cancer; Small
Intestine Cancer; Soft Tissue Sarcoma; Squamous Neck Cancer with
Occult Primary, Metastatic; Stomach (Gastric) Cancer;
Supratentorial Primitive Neuroectodermal Tumors; T-Cell Lymphoma,
Cutaneous; Testicular Cancer; Thymoma, Malignant; Thyroid Cancer;
Transitional Cell Cancer of the Renal Pelvis and Ureter;
Trophoblastic Tumor, Gestational; Ureter and Renal Pelvis,
Transitional Cell Cancer; Urethral Cancer; Uterine Sarcoma; Vaginal
Cancer; Visual Pathway and Hypothalamic Glioma; Vulvar Cancer;
Waldenstrom's Macro globulinemia; and Wilms' Tumor.
[0273] The compounds of Formula (I) and Formula (II) are further
useful in inhibiting the growth of a cancer cell or inhibiting
replication of a cancer cell. In certain embodiments, the cancer is
selected from liver cancer, breast cancer, ovarian cancer, lung
cancer, pancreatic cancer or leukemic cancer. In an embodiment the
cancer is liver cancer. In certain embodiments, the liver cancer is
hepatocellular carcinoma, cholangiocarcinoma, or biliary tract
cancer. In particular embodiments, the cancer is hepatocellular
carcinoma.
Treatment or Prevention of Viral Infection
[0274] The compounds of Formula (I) and Formula (II) are useful in
the inhibition of viral infection, the treatment viral infection
and/or reduction of the likelihood or severity of symptoms of viral
infection and the inhibition of viral replication and/or viral
production in a cell-based system. For example, the compounds of
Formula (I) and Formula (II) are useful in treating viral infection
after suspected past exposure to the virus by such means as blood
transfusion, exchange of body fluids, bites, accidental needle
stick, or exposure to patient blood during surgery or other medical
procedures.
[0275] In a further embodiment, the compounds of Formula (I) and
Formula (II) are useful in the inhibition of HBV, the treatment of
HBV infection and/or reduction of the likelihood or severity of
symptoms of HBV infection and the inhibition of HBV replication
and/or HBV production in a cell-based system. For example, the
compounds of Formula (I) and Formula (II) are useful in treating
infection by HBV after suspected past exposure to HBV by such means
as blood transfusion, exchange of body fluids, bites, accidental
needle stick, or exposure to patient blood during surgery or other
medical procedures.
[0276] In one embodiment, the hepatitis B infection is acute
hepatitis B. In another embodiment, the hepatitis B infection is
chronic hepatitis B.
[0277] Accordingly, in one embodiment, the invention provides
methods for treating HBV infection in a patient, the methods
comprising administering to the patient an effective amount of at
least one compound of Formula (I) or Formula (II) or a
pharmaceutically acceptable salt, solvate or enantiomer thereof. In
a specific embodiment, the amount administered is effective to
treat or prevent infection by HBV in the patient. In another
specific embodiment, the amount administered is effective to
inhibit HBV viral replication and/or viral production in the
patient.
[0278] In a further embodiment, the compounds and compositions
provided herein are useful in methods of treatment of a liver
disorder, that comprise further administration of a second or more
agent effective for the treatment of the disorder, such as HBV
infection in a subject in need thereof. The second agent can be any
agent known to those of skill in the art to be effective for the
treatment of the disorder, including those currently approved by
the FDA.
[0279] In a further embodiment, the compounds of Formula (I) and
Formula (II) are useful in the inhibition of ebola virus, the
treatment of Ebolavirus infection and/or reduction of the
likelihood or severity of symptoms of Ebolavirus infection and the
inhibition of Ebolavirus replication and/or Ebolavirus production
in a cell-based system. For example, the compounds of Formula (I)
and Formula (II) are useful in treating infection by Ebolavirus
after suspected past exposure to ebola virus by such means as blood
transfusion, exchange of body fluids, bites, accidental needle
stick, or exposure to patient blood during surgery or other medical
procedures.
Combination Therapy
[0280] In another embodiment, the present compounds, compositions,
combinations and methods for treating or preventing liver diseases,
such as cancer or viral infection, can further comprise the
administration of one or more additional therapeutic agents which
are not compounds of Formula (I) or Formula (II).
[0281] In one embodiment, the additional therapeutic agent is an
anticancer agent.
[0282] In another embodiment, the additional therapeutic agent is
an antiviral agent.
[0283] In another embodiment, the additional therapeutic agent is
an immunomodulatory agent, such as an immunosuppressive agent.
[0284] As used herein, the term "in combination" includes the use
of more than one therapy (e.g., one or more prophylactic and/or
therapeutic agents). The use of the term "in combination" does not
restrict the order in which therapies (e.g., prophylactic and/or
therapeutic agents) are administered to a subject with a disorder.
A first therapy (e.g., a prophylactic or therapeutic agent such as
a compound provided herein) can be administered prior to (e.g., 5
minutes, 15 minutes, 30 minutes, 45 minutes, 1 hour, 2 hours, 4
hours, 6 hours, 12 hours, 24 hours, 48 hours, 72 hours, 96 hours, 1
week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 8 weeks, or 12
weeks before), concomitantly with, or subsequent to (e.g., 5
minutes, 15 minutes, 30 minutes, 45 minutes, 1 hour, 2 hours, 4
hours, 6 hours, 12 hours, 24 hours, 48 hours, 72 hours, 96 hours, 1
week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 8 weeks, or 12
weeks after) the administration of a second therapy (e.g., a
prophylactic or therapeutic agent) to a subject with a
disorder.
[0285] As used herein, the term "synergistic" includes a
combination of a compound provided herein and another therapy
(e.g., a prophylactic or therapeutic agent) which has been or is
currently being used to prevent, manage or treat a disorder, which
is more effective than the additive effects of the therapies. A
synergistic effect of a combination of therapies (e.g., a
combination of prophylactic or therapeutic agents) permits the use
of lower dosages of one or more of the therapies and/or less
frequent administration of said therapies to a subject with a
disorder. The ability to utilize lower dosages of a therapy (e.g.,
a prophylactic or therapeutic agent) and/or to administer said
therapy less frequently reduces the toxicity associated with the
administration of said therapy to a subject without reducing the
efficacy of said therapy in the prevention or treatment of a
disorder). In addition, a synergistic effect can result in improved
efficacy of agents in the prevention or treatment of a disorder.
Finally, a synergistic effect of a combination of therapies (e.g.,
a combination of prophylactic or therapeutic agents) may avoid or
reduce adverse or unwanted side effects associated with the use of
either therapy alone.
[0286] When administering a combination therapy of the invention to
a patient, therapeutic agents in the combination, or a
pharmaceutical composition or compositions comprising therapeutic
agents, may be administered in any order such as, for example,
sequentially, concurrently, together, simultaneously and the like.
The amounts of the various actives in such combination therapy may
be different amounts (different dosage amounts) or same amounts
(same dosage amounts). Thus, for illustration purposes, a Compound
of Formula (I) or Formula (II) and an additional therapeutic agent
may be present in fixed amounts (dosage amounts) in a single dosage
unit (e.g., a capsule, a tablet and the like).
[0287] In one embodiment, the at least one compound of Formula (I)
or Formula (II) is administered during a time when the additional
therapeutic agent(s) exert their prophylactic or therapeutic
effect, or vice versa.
[0288] In another embodiment, the at least one compound of Formula
(I) or Formula (II) and the additional therapeutic agent(s) are
administered in doses commonly employed when such agents are used
as monotherapy for treating a cancer or a viral infection.
[0289] In another embodiment, the at least one compound of Formula
(I) or Formula (II) and the additional therapeutic agent(s) are
administered in doses lower than the doses commonly employed when
such agents are used as monotherapy for treating a cancer or a
viral infection.
[0290] In still another embodiment, the at least one compound of
Formula (I) or Formula (II) and the additional therapeutic agent(s)
act synergistically and are administered in doses lower than the
doses commonly employed when such agents are used as monotherapy
for treating a cancer or a viral infection.
[0291] In one embodiment, the at least one compound of Formula (I)
or Formula (II) and the additional therapeutic agent(s) are present
in the same composition. In one embodiment, this composition is
suitable for oral administration. In another embodiment, this
composition is suitable for intravenous administration. In another
embodiment, this composition is suitable for subcutaneous
administration. In still another embodiment, this composition is
suitable for parenteral administration.
[0292] In another embodiment, the present invention provides
compositions comprising a Compound of Formula (I) or Formula (II),
a pharmaceutically acceptable carrier, and a second therapeutic
agent selected from the group consisting of immunomodulators,
anti-infective agents and anti-cancer agents.
[0293] In another embodiment, the present invention provides
compositions comprising a Compound of Formula (I) or Formula (II),
a pharmaceutically acceptable carrier, and two additional
therapeutic agents, each of which are independently selected from
the group consisting of immunomodulators, anti-infective agents and
anti-cancer agents.
[0294] In one embodiment, the present invention provides a compound
of Formula (I) or Formula (II) for use in treating cancer in a
patient comprising administering to the patient: (i) at least one
compound of Formula (I) or Formula (II) (which may include two or
more different 2'-Substituted Nucleoside Derivatives), or a
pharmaceutically acceptable salt or solvate thereof, and (ii) at
least one additional therapeutic agent that is other than a
compound of Formula (I) or Formula (II), wherein the amounts
administered are together effective to treat or prevent cancer.
[0295] In certain embodiments the compounds and compositions
provided herein are useful in the treatment of a liver cancer, that
comprise further administration of a second agent effective for the
treatment of the liver cancer in a subject in need thereof. The
second agent can be any agent known to those of skill in the art to
be effective for the treatment of the liver cancer, including those
currently approved by the FDA.
[0296] In certain embodiments, a compound provided herein is
administered in combination with one second agent. In further
embodiments, a compound provided herein is administered in
combination with two second agents. In still further embodiments, a
compound provided herein is administered in combination with two or
more second agents.
[0297] The active compounds provided herein can be administered in
combination or alternation with another therapeutic agent, in
particular an anti-cancer agent. In combination therapy, effective
dosages of two or more agents are administered together, whereas in
alternation or sequential-step therapy, an effective dosage of each
agent is administered serially or sequentially. The dosages given
will depend on absorption, inactivation and excretion rates of the
drug as well as other factors known to those of skill in the art.
It is to be noted that dosage values will also vary with the
severity of the condition to be alleviated. It is to be further
understood that for any particular subject, specific dosage
regimens and schedules should be adjusted over time according to
the individual need and the professional judgment of the person
administering or supervising the administration of the
compositions.
[0298] In certain embodiments, the second agent is selected from
the group consisting of sorafenib tosylate (Nexavar), radiation
therapy, selective internal radiation therapy (e.g., SIR-Spheres
and TheraSphere), ethiodized oil (Lipidol), pexastimogene
devacirepvec (Pexa-Vec, JX-594, Jennarex), Quinacrine (Clevelane
BioLabs), CC-223 (Celgene), CF 102 (Can-Fite), SGI-110 (Astex), and
G-202 (Genspera).
[0299] In one embodiment, the other anticancer agent is selected
from the group consisting of vascular endothelial growth factor
(VEGF) receptor inhibitors, topoisomerase II inhibitors, smoothen
inhibitors, alkylating agents, anti-tumor antibiotics,
anti-metabolites, retinoids, immunomodulatory agents including but
not limited to anti-cancer vaccines, CTLA-4, LAG-3, PD-1
antagonists and BET bromodomain inhibitors.
[0300] Examples of vascular endothelial growth factor (VEGF)
receptor inhibitors include, bevacizumab (sold under the trademark
AVASTIN by Genentech/Roche), axitinib,
(N-methyl-2-[[3-[([pound])-2-pyridin-2-ylethenyl]-1H-indazol-6-yl]sulfany-
l]benzamide, also known as AG013736, and described in PCT
Publication No. WO 01/002369), Brivanib Alaninate
((S)--((R)-1-(4-(4-Fluoro-2-methyl-1H-indol-5-yloxy)-5-methylpyrrolo[2,1--
f][1,2,4]triazin-6-yloxy)propan-2-yl)2-aminopropanoate, also known
as BMS-582664), motesanib
(N-(2,3-dihydro-3,3-dimethyl-1H-indoi-6-y!)-2-[(4-pyridinyimethyj)amino]--
3-pyfidinecarboxamide. and described in PCT Publication No. WO
02/068470), pasireotide (also known as SO 230, and described in PCT
Publication No. WO 02/010192), and sorafenib (sold under the
tradename NEXAVAR).
[0301] Examples of topoisomerase II inhibitors include, etoposide
(also known as VP-16 and Etoposide phosphate, sold under the
tradenames TOPOSAR, VEPESID and ETOPOPHOS), and teniposide (also
known as VM-26, sold under the tradename VUMON).
[0302] Examples of alkylating agents include, 5-azacytidine (sold
under the trade name VIDAZA), decitabine (sold under the trade name
of DECOGEN), temozolomide (sold under the trade names TEMODAR and
TEMODAL by Schering-Plough/Merck), dactinomycin (also known as
actinomycin-D and sold under the tradename COSMEGEN), melphalan
(also known as L-PAM, L-sarcolysin, and phenylalanine mustard, sold
under the tradename ALKERAN), altretamine (also known as
hexamethylmelamine (HMM), sold under the tradename HEXALEN),
carmustine (sold under the tradename BCNU), bendamustine (sold
under the tradename TREANDA), busulfan (sold under the tradenames
Busulfex.RTM. and Myleran.RTM.), carboplatin (sold under the
tradename Paraplatin.RTM.), lomustine (also known as CCNU, sold
under the tradename CeeNU.RTM.), cisplatin (also known as CDDP,
sold under the tradenames Platinol.RTM. and Platinol.RTM.-AQ),
chlorambucil (sold under the tradename Leukeran.RTM.),
cyclophosphamide (sold under the tradenames Cytoxan.RTM. and
Neosar.RTM.), dacarbazine (also known as DTIC, DIC and imidazole
carboxamide, sold under the tradename DTIC-Dome.RTM.), altretamine
(also known as hexamethylmelamine (HMM) sold under the tradename
Hexalen.RTM.), ifosfamide (sold under the tradename Ifex.RTM.),
procarbazine (sold under the tradename Matulane.RTM.),
mechlorethamine (also known as nitrogen mustard, mustine and
mechloroethamine hydrochloride, sold under the tradename
Mustargen.RTM.), streptozocin (sold under the tradename
Zanosar.RTM.), thiotepa (also known as thiophosphoamide, TESPA and
TSPA, and sold under the tradename Thioplex.RTM..
[0303] Examples of anti-tumor antibiotics include, doxorubicin
(sold under the tradenames Adriamycin.RTM. and Rubex.RTM.),
bleomycin (sold under the tradename Lenoxane.RTM.), daunorubicin
(also known as dauorubicin hydrochloride, daunomycin, and
rubidomycin hydrochloride, sold under the tradename
Cerubidine.RTM.), daunorubicin liposomal (daunorubicin citrate
liposome, sold under the tradename DaunoXome.RTM.), mitoxantrone
(also known as DHAD, sold under the tradename Novantrone.RTM.),
epirubicin (sold under the tradename Ellence.TM.), idarubicin (sold
under the tradenames Idamycin.RTM., Idamycin PFS.RTM.), and
mitomycin C (sold under the tradename Mutamycin.RTM.).
[0304] Examples of anti-metabolites include, claribine
(2-chlorodeoxyadenosine, sold under the tradename Leustatin.RTM.),
5-fluorouracil (sold under the tradename Adrucil.RTM.),
6-thioguanine (sold under the tradename Purinethol.RTM.),
pemetrexed (sold under the tradename Alimta.RTM.), cytarabine (also
known as arabinosylcytosine (Ara-C), sold under the tradename
Cytosar-U.RTM.), cytarabine liposomal (also known as Liposomal
Ara-C, sold under the tradename DepoCyt.TM.), decitabine (sold
under the tradename Dacogen.RTM.), hydroxyurea (sold under the
tradenames Hydrea.RTM., Droxia.TM. and Mylocel.TM.), fludarabine
(sold under the tradename Fludara.RTM.), floxuridine (sold under
the tradename FUDR.RTM.), cladribine (also known as
2-chlorodeoxyadenosine (2-CdA) sold under the tradename
Leustatin.TM.), methotrexate (also known as amethopterin,
methotrexate sodium (MTX), sold under the tradenames
Rheumatrex.RTM. and Trexall.TM.), and pentostatin (sold under the
tradename Nipent.RTM.).
[0305] Examples of retinoids include, alitretinoin (sold under the
tradename Panretin.RTM.), tretinoin (all-trans retinoic acid, also
known as ATRA, sold under the tradename Vesanoid.RTM.),
Isotretinoin (13-c/s-retinoic acid, sold under the tradenames
Accutane.RTM., Amnesteem.RTM., Claravis.RTM., Clarus.RTM.,
Decutan.RTM., Isotane.RTM., Izotech.RTM., Oratane.RTM.,
Isotret.RTM., and Sotret.RTM.), and bexarotene (sold under the
tradename Targretin.RTM.).
[0306] "PD-1 antagonist" means any chemical compound or biological
molecule that blocks binding of PD-L1 expressed on a cancer cell to
PD-1 expressed on an immune cell (T cell, B cell or NKT cell) and
preferably also blocks binding of PD-L2 expressed on a cancer cell
to the immune-cell expressed PD-1. Alternative names or synonyms
for PD-1 and its ligands include: PDCD1, PD1, CD279 and SLEB2 for
PD-1; PDCD1L1, PDL1, B7H1, B7-4, CD274 and B7-H for PD-L1; and
PDCD1L2, PDL2, B7-DC, Btdc and CD273 for PD-L2. In any of the
treatment method, medicaments and uses of the present invention in
which a human individual is being treated, the PD-1 antagonist
blocks binding of human PD-L1 to human PD-1, and preferably blocks
binding of both human PD-L1 and PD-L2 to human PD-1. Human PD-1
amino acid sequences can be found in NCBI Locus No.: NP_005009.
Human PD-L1 and PD-L2 amino acid sequences can be found in NCBI
Locus No.: NP_054862 and NP_079515, respectively.
[0307] PD-1 antagonists useful in any of the compositions and uses
of the present invention include a monoclonal antibody (mAb), or
antigen binding fragment thereof, which specifically binds to PD-1
or PD-L1, and preferably specifically binds to human PD-1 or human
PD-L1. The mAb may be a human antibody, a humanized antibody or a
chimeric antibody, and may include a human constant region. In some
embodiments the human constant region is selected from the group
consisting of IgG1, IgG2, IgG3 and IgG4 constant regions, and in
preferred embodiments, the human constant region is an IgG1 or IgG4
constant region. In some embodiments, the antigen binding fragment
is selected from the group consisting of Fab, Fab'-SH,
F(ab').sub.2, scFv and Fv fragments.
[0308] Examples of mAbs that bind to human PD-1, and useful in the
treatment method, medicaments and uses of the present invention,
are described in U.S. Pat. Nos. 7,488,802, 7,521,051, 8,008,449,
8,354,509, 8,168,757, WO2004/004771, WO2004/072286, WO2004/056875,
and US2011/0271358.
[0309] Examples of mAbs that bind to human PD-L1, and useful in the
treatment method, medicaments and uses of the present invention,
are described in WO2013/019906, WO2010/077634 A1 and U.S. Pat. No.
8,383,796. Specific anti-human PD-L1 mAbs useful as the PD-1
antagonist in the treatment method, medicaments and uses of the
present invention include MPDL3280A, BMS-936559, MED14736,
MSB0010718C and an antibody which comprises the heavy chain and
light chain variable regions of SEQ ID NO:24 and SEQ ID NO:21,
respectively, of WO2013/019906.
[0310] Other PD-1 antagonists useful in any of the treatment
method, medicaments and uses of the present invention include, an
immunoadhesin that specifically binds to PD-1 or PD-L1, and
preferably specifically binds to human PD-1 or human PD-L1, e.g., a
fusion protein containing the extracellular or PD-1 binding portion
of PD-L1 or PD-L2 fused to a constant region such as an Fc region
of an immunoglobulin molecule. Examples of immunoadhesion molecules
that specifically bind to PD-1 are described in WO2010/027827 and
WO2011/066342. Specific fusion proteins useful as the PD-1
antagonist in the treatment method, medicaments and uses of the
present invention include AMP-224 (also known as B7-DCIg), which is
a PD-L2-FC fusion protein and binds to human PD-1.
[0311] Examples of other cytotoxic agents include, arsenic trioxide
(sold under the tradename Trisenox.RTM.), asparaginase (also known
as L-asparaginase, and Erwinia L-asparaginase, sold under the
tradenames Elspar.RTM. and Kidrolase.RTM.).
[0312] In an embodiment, the other anticancer agent is a BET
bromodomain inhibitor. Examples of BET bromodomain inhibitor
include the compounds described in U.S. Pat. No. 5,712,274,
WO1994006802, U.S. Pat. No. 8,476,260 and WO2009/084693.
[0313] In a further embodiment, the present invention provides a
compound of Formula (I) or Formula (II) for use in treating a viral
infection in a patient comprising administering to the patient: (i)
at least one compound of Formula (I) or Formula (II) (which may
include two or more different 2'-Substituted Nucleoside
Derivatives), or a pharmaceutically acceptable salt or solvate
thereof, and (ii) at least one additional therapeutic agent that is
other than a compound of Formula (I) or Formula (II), wherein the
amounts administered are together effective to treat or prevent a
viral infection.
[0314] Accordingly, in one embodiment, the present invention
provides compositions comprising: (i) at least one Compound of
Formula (I) or Formula (II) or a pharmaceutically acceptable salt,
solvate or enantiomer thereof; (ii) one or more additional
therapeutic agents that are not a Compound of Formula (I) or
Formula (II); and (iii) a pharmaceutically acceptable carrier,
wherein the amounts in the composition are together effective to
prevent HBV replication and/or treat HBV infection.
[0315] In another embodiment, the present invention provides
compositions comprising: (i) at least one Compound of Formula (I)
or Formula (II) or a pharmaceutically acceptable salt, solvate or
enantiomer thereof; (ii) one or more additional therapeutic agents
that are not a Compound of Formula (I) or Formula (II); and (iii) a
pharmaceutically acceptable carrier, wherein the amounts in the
composition are together effective to prevent Ebolavirus
replication and/or treat Ebolavirus infection.
[0316] Viral infections and virus-related disorders that may be
treated or prevented using the combination therapy of the present
invention include those listed above.
[0317] In one embodiment, the viral infection is HBV infection.
[0318] In another embodiment, the viral infection is Ebolavirus
infection.
[0319] The at least one compound of Formula (I) or Formula (II) and
the additional therapeutic agent(s) can act additively or
synergistically. A synergistic combination may allow the use of
lower dosages of one or more agents and/or less frequent
administration of one or more agents of a combination therapy. A
lower dosage or less frequent administration of one or more agents
may lower toxicity of therapy without reducing the efficacy of
therapy.
[0320] In one embodiment, the administration of at least one
compound of Formula (I) or Formula (II) and the additional
therapeutic agent(s) may inhibit the resistance of a viral
infection to these agents.
[0321] Examples of additional therapeutic agents useful in the
present compositions and methods include an interferon, an
immunomodulator, a viral replication inhibitor, an antisense agent,
a therapeutic vaccine, a viral polymerase inhibitor, a nucleoside
inhibitor, a viral protease inhibitor, a viral helicase inhibitor,
a virion production inhibitor, a viral entry inhibitor, a viral
assembly inhibitor, an antibody therapy (monoclonal or polyclonal),
and any agent useful for treating an RNA-dependent
polymerase-related disorder.
[0322] In one embodiment, one or more compounds of the invention
are administered with one or more additional therapeutic agents,
including the therapeutic agents described, supra.
[0323] In one embodiment, the additional therapeutic agent is a
viral protease inhibitor.
[0324] In another embodiment, the additional therapeutic agent is a
viral replication inhibitor.
[0325] In another embodiment, the additional therapeutic agent is
an HBV entry inhibitor.
[0326] In another embodiment, the additional therapeutic agent is
an HBV capsid inhibitor.
[0327] In still another embodiment, the additional therapeutic
agent is an HBV polymerase inhibitor.
[0328] In another embodiment, the additional therapeutic agent is a
nucleoside inhibitor.
[0329] In another embodiment, the additional therapeutic agent is
an interferon.
[0330] In another embodiment, the additional therapeutic agent is
an antisense agent.
[0331] In another embodiment, the additional therapeutic agent is a
therapeutic vaccine.
[0332] In a further embodiment, the additional therapeutic agent is
a virion production inhibitor.
[0333] In another embodiment, the additional therapeutic agent is
an antibody therapy.
[0334] In still another embodiment, one or more compounds of the
present invention are administered with two additional therapeutic
agents.
[0335] In another embodiment, one or more compounds of the present
invention are administered with three additional therapeutic
agents.
[0336] In another embodiment, one or more compounds of the present
invention are administered with an HBV polymerase inhibitor.
[0337] In another embodiment, one or more compounds of the present
invention are administered with lamivudine.
[0338] In a further embodiment, one or more compounds of the
present invention are administered with tenofovir.
[0339] In another embodiment, one or more compounds of the present
invention are administered with pegylated-interferon alpha and
ribavirin.
[0340] In one embodiment, the additional therapeutic agents
comprise a viral protease inhibitor and a viral polymerase
inhibitor.
[0341] In still another embodiment, the additional therapeutic
agents comprise a viral protease inhibitor and an immunomodulatory
agent.
[0342] In yet another embodiment, the additional therapeutic agents
comprise a polymerase inhibitor and an immunomodulatory agent.
[0343] In another embodiment, the additional therapeutic agents
comprise a viral protease inhibitor and a nucleoside.
[0344] In another embodiment, the additional therapeutic agents
comprise an immunomodulatory agent and a nucleoside.
[0345] In another embodiment, the additional therapeutic agents
comprise a nucleoside and an HBV polymerase inhibitor.
[0346] In a further embodiment, the additional therapeutic agents
comprise a viral protease inhibitor, a viral polymerase inhibitor
and an immunomodulatory agent.
[0347] In another embodiment, the additional therapeutic agent is
ribavirin.
[0348] HBV polymerase inhibitors useful in the present compositions
and treatments include lamivudine (Epivir), adefovir (Hepsera),
tenofovir (Viread), telbivudine (Tyzeka), ganciclovir (Cytovene),
entecavir (Baraclude), interferon alpha-2a, and PEGylated
interferon alpha-2a (Pegasys).
[0349] Interferons useful in the present compositions and methods
include, but are not limited to, interferon alfa-2a, interferon
alfa-2b, interferon alfacon-1 and petroleum etherG-interferon alpha
conjugates. "PEG-interferon alpha conjugates" are interferon alpha
molecules covalently attached to a petroleum etherG molecule.
Illustrative petroleum etherG-interferon alpha conjugates include
interferon alpha-2a (Roferon.TM., Hoffman La-Roche, Nutley, N.J.)
in the form of pegylated interferon alpha-2a (e.g., as sold under
the trade name Pegasys.TM.), interferon alpha-2b (Intron.TM., from
Schering-Plough Corporation) in the form of pegylated interferon
alpha-2b (e.g., as sold under the trade name petroleum
etherG-Intron.TM. from Schering-Plough Corporation), interferon
alpha-2b-XL (e.g., as sold under the trade name petroleum
etherG-Intron.TM.), interferon alpha-2c (Berofor Alpha.TM.,
Boehringer Ingelheim, Ingelheim, Germany), petroleum
etherG-interferon lambda (Bristol-Myers Squibb and ZymoGenetics),
interferon alfa-2b alpha fusion polypeptides, interferon fused with
the human blood protein albumin (Albuferon.TM., Human Genome
Sciences), Omega Interferon (Intarcia), Locteron controlled release
interferon (Biolex/OctoPlus), Biomed-510 (omega interferon),
Peg-IL-29 (ZymoGenetics), Locteron CR (Octoplus), R-7025 (Roche),
IFN-.alpha.-2b-XL (Flamel Technologies), belerofon (Nautilus) and
consensus interferon as defined by determination of a consensus
sequence of naturally occurring interferon alphas (Infergen.TM.,
Amgen, Thousand Oaks, Calif.).
[0350] Examples of other HBV inhibitors useful in the present
compositions and treatments include TLR-3 or TLR-7 agonists, virus
entry inhibitors, cccDNA formation inhibitors, silenceor of cccDNA,
nucleocapsid formation inhibitors, virion maturation, assembly or
secretion inhibitors.
[0351] The doses and dosage regimen of the other agents used in the
combination therapies of the present invention for the treatment or
prevention of the viral infection may be determined using the
attending clinician, taking into consideration the approved doses
and dosage regimen in the package insert; the age, sex and general
health of the patient; and the type and severity of the viral
infection or related disease or disorder. When administered in
combination, the compound of Formula (I) or Formula (II) and the
other agent(s) may be administered simultaneously (i.e., in the
same composition or in separate compositions one right after the
other) or sequentially. This particularly useful when the
components of the combination are given on different dosing
schedules, e.g., one component is administered once daily and
another component is administered every six hours, or when the
preferred pharmaceutical compositions are different, e.g., one is a
tablet and one is a capsule. A kit comprising the separate dosage
forms is therefore advantageous.
[0352] Generally, a total daily dosage of the at least one Compound
of Formula (I) or Formula (II) alone, or when administered as
combination therapy, can range from about 1 to about 2500 mg per
day, although variations will necessarily occur depending on the
target of therapy, the patient and the route of administration. In
one embodiment, the dosage is from about 10 to about 1000 mg/day,
administered in a single dose or in 2-4 divided doses. In another
embodiment, the dosage is from about 1 to about 500 mg/day,
administered in a single dose or in 2-4 divided doses. In still
another embodiment, the dosage is from about 1 to about 100 mg/day,
administered in a single dose or in 2-4 divided doses. In yet
another embodiment, the dosage is from about 1 to about 50 mg/day,
administered in a single dose or in 2-4 divided doses. In another
embodiment, the dosage is from about 500 to about 1500 mg/day,
administered in a single dose or in 2-4 divided doses. In still
another embodiment, the dosage is from about 500 to about 1000
mg/day, administered in a single dose or in 2-4 divided doses. In
yet another embodiment, the dosage is from about 100 to about 500
mg/day, administered in a single dose or in 2-4 divided doses.
[0353] In a further embodiment, when the additional therapeutic
agent is Ribavirin (commercially available as REBETOL ribavirin
from Schering-Plough or COPEGUS ribavirin from Hoffmann-La Roche),
this agent is administered at a daily dosage of from about 600 to
about 1400 mg/day for at least 24 weeks.
Compositions and Administration
[0354] Due to their activity, the compounds of Formula (I) and
Formula (II) are useful in veterinary and human medicine. As
described above, the compounds of Formula (I) and Formula (II) are
useful for treating or preventing liver diseases, such as cancer,
HBV infection or Ebolavirus infection in a patient.
[0355] When administered to a patient, the compounds of Formula (I)
or Formula (II) may be administered as a component of a composition
that comprises a pharmaceutically acceptable carrier or vehicle.
The present invention provides pharmaceutical compositions
comprising an effective amount of at least one compound of Formula
(I) or Formula (II) or a pharmaceutically acceptable salt, solvate
or enantiomer thereof and a pharmaceutically acceptable carrier or
diluent. In the pharmaceutical compositions and uses of the present
invention, the active ingredients will typically be administered in
admixture with suitable carrier materials suitably selected with
respect to the intended form of administration, i.e., oral tablets,
capsules (either solid-filled, semi-solid filled or liquid filled),
powders for constitution, oral gels, elixirs, dispersible granules,
syrups, suspensions, and the like, and consistent with conventional
pharmaceutical practices. For example, for oral administration in
the form of tablets or capsules, the active drug component may be
combined with any oral non-toxic pharmaceutically acceptable inert
carrier, such as lactose, starch, sucrose, cellulose, magnesium
stearate, dicalcium phosphate, calcium sulfate, talc, mannitol,
ethyl alcohol (liquid forms) and the like. Solid form preparations
include powders, tablets, dispersible granules, capsules, cachets
and suppositories. Powders and tablets may be comprised of from
about 0.5 to about 95 percent inventive composition. Tablets,
powders, cachets and capsules may be used as solid dosage forms
suitable for oral administration.
[0356] Moreover, when desired or needed, suitable binders,
lubricants, disintegrating agents and coloring agents may also be
incorporated in the mixture. Suitable binders include starch,
gelatin, natural sugars, corn sweeteners, natural and synthetic
gums such as acacia, sodium alginate, carboxymethylcellulose,
polyethylene glycol and waxes. Among the lubricants there may be
mentioned for use in these dosage forms, boric acid, sodium
benzoate, sodium acetate, sodium chloride, and the like.
Disintegrants include starch, methylcellulose, guar gum, and the
like. Sweetening and flavoring agents and preservatives may also be
included where appropriate.
[0357] Liquid form preparations include solutions, suspensions and
emulsions and may include water or water-propylene glycol solutions
for parenteral injection.
[0358] Liquid form preparations may also include solutions for
intranasal administration.
[0359] Also included are solid form preparations which are intended
to be converted, shortly before use, to liquid form preparations
for either oral or parenteral administration. Such liquid forms
include solutions, suspensions and emulsions.
[0360] For preparing suppositories, a low melting wax such as a
mixture of fatty acid glycerides or cocoa butter is first melted,
and the active ingredient is dispersed homogeneously therein as by
stirring. The molten homogeneous mixture is then poured into
convenient sized molds, allowed to cool and thereby solidify.
[0361] Additionally, the compositions of the present invention may
be formulated in sustained release form to provide the rate
controlled release of any one or more of the components or active
ingredients to optimize therapeutic effects, i.e., antiviral
activity and the like. Suitable dosage forms for sustained release
include layered tablets containing layers of varying disintegration
rates or controlled release polymeric matrices impregnated with the
active components and shaped in tablet form or capsules containing
such impregnated or encapsulated porous polymeric matrices.
[0362] In one embodiment, the one or more compounds of Formula (I)
or Formula (II) are administered orally.
[0363] In another embodiment, the one or more compounds of Formula
(I) or Formula (II) are administered intravenously.
[0364] In one embodiment, a pharmaceutical preparation comprising a
compound of Formula (I) or Formula (II) is in unit dosage form. In
such form, the preparation is subdivided into unit doses containing
effective amounts of the active components.
[0365] Compositions may be prepared according to conventional
mixing, granulating or coating methods, respectively, and the
present compositions can contain, in one embodiment, from about
0.1% to about 99% of the compound of Formula (I) or Formula (II) by
weight or volume. In various embodiments, the present compositions
can contain, in one embodiment, from about 1% to about 70% or from
about 5% to about 60% of the compound of Formula (I) or Formula
(II) by weight or volume.
[0366] The quantity of compound of Formula (I) or Formula (II) in a
unit dose of preparation may be varied or adjusted from about 1 mg
to about 2500 mg. In various embodiments, the quantity is from
about 10 mg to about 1000 mg, 1 mg to about 500 mg, 1 mg to about
100 mg, and 1 mg to about 100 mg.
[0367] For convenience, the total daily dosage may be divided and
administered in portions during the day if desired. In one
embodiment, the daily dosage is administered in one portion. In
another embodiment, the total daily dosage is administered in two
divided doses over a 24 hour period. In another embodiment, the
total daily dosage is administered in three divided doses over a 24
hour period. In still another embodiment, the total daily dosage is
administered in four divided doses over a 24 hour period.
[0368] The amount and frequency of administration of the compounds
of Formula (I) or Formula (II) will be regulated according to the
judgment of the attending clinician considering such factors as
age, condition and size of the patient as well as severity of the
symptoms being treated. Generally, a total daily dosage of the
compounds of Formula (I) or Formula (II) range from about 0.1 to
about 2000 mg per day, although variations will necessarily occur
depending on the target of therapy, the patient and the route of
administration. In one embodiment, the dosage is from about 1 to
about 200 mg/day, administered in a single dose or in 2-4 divided
doses. In another embodiment, the dosage is from about 10 to about
2000 mg/day, administered in a single dose or in 2-4 divided doses.
In another embodiment, the dosage is from about 100 to about 2000
mg/day, administered in a single dose or in 2-4 divided doses. In
still another embodiment, the dosage is from about 500 to about
2000 mg/day, administered in a single dose or in 2-4 divided
doses.
EXAMPLES
[0369] The invention is illustrated by the following examples. For
all of the examples, standard work-up and purification methods
known to those skilled in the art can be utilized. Unless otherwise
indicated, all temperatures are expressed in .degree. C. (degrees
Centigrade). All reactions are conducted at room temperature unless
otherwise noted. Synthetic methodologies illustrated herein are
intended to exemplify the applicable chemistry through the use of
specific examples and are not indicative of the scope of the
disclosure.
[0370] The following abbreviations are used herein: [0371] Ac
acetyl [0372] Ac.sub.2O acetic anhydride [0373] ACN acetonitrile
[0374] AcOH or HOAc acetic acid [0375] APCI atmospheric-pressure
chemical ionization [0376] aq aqueous [0377] Bn benzyl [0378] Boc
or BOC tert-butoxycarbonyl [0379] Bz benzoyl [0380] Cbz
benzyloxycvarbonyl [0381] calc'd calculated [0382] Celite
diatomaceous earth [0383] DBU 1,8-diazabicyclo(5.4.0)undec-7-ene
[0384] DCM dichloromethane [0385] DIEA or DIPEA
N,N-diisopropylethylamine [0386] DMAP 4-dimethylaminopyridine
[0387] DMF N,N-dimethylformamide [0388] DMSO dimethyl sulfoxide
[0389] dMTrCl 4,4'-dimethoxytrityl chloride [0390] EDCI
1-ethyl-3-(3-dimethylaminopropyl)carbodiimide [0391] EDTA
ethylenediamine tetraacetic acid [0392] ESI electrospray ionization
[0393] Et ethyl [0394] Et.sub.2O diethyl ether [0395] EtOH ethanol
[0396] EtOAc ethyl acetate [0397] Et.sub.3N triethylamine [0398] h
hour [0399] HPLC high-performance liquid chromatography [0400] IPA
isopropanol [0401] iPr isopropyl [0402] LC liquid chromatography
[0403] LCMS liquid chromatography mass spectrometry [0404] Me
Methyl [0405] MeCN acetonitrile [0406] MeOH methanol [0407] mg
milligrams [0408] min minute [0409] .mu.L microliters [0410] mL
milliliters [0411] mmol millimoles [0412] MS mass spectrometry
[0413] MTBE methyl tert-butyl ether [0414] MTS
(3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfoph-
enyl)-2H-tetrazolium) [0415] NMR nuclear magnetic resonance
spectroscopy [0416] PDA photodiode array [0417] PE petroleum ether
[0418] Ph phenyl [0419] Pr Propyl [0420] PS Polystyrene [0421] Rac
racemic mixture [0422] Rh(COD).sub.2OTf
Bis(1,5-cyclooctadiene)rhodium(I) trifluoromethanesulfonate [0423]
Rosphos bistriflate
(+)-1,2-Bis[(2S,5S)-2,5-dimethyl-(3S,4S)-3,4-dihydroxyphospholano]benzene
bis(trifluoromethanesulfonate)salt [0424] RT room temperature
[0425] Rt retention time [0426] Sat saturated [0427] SFC
supercritical fluid chromatography [0428] TBAF tert-butyl ammonium
fluoride [0429] TBS or TBDMS tert-butyldimethylsilyl [0430] TBSCL
tert-butyldimethylsilyl chloride [0431] t-Bu tert-butyl [0432] TEA
triethylamine [0433] TFA trifluoroacetic acid [0434] THF
tetrahydrofuran [0435] TLC thin layer chromatography [0436] TMS
trimethylsilyl [0437] Tris tris(hydroxymethyl)aminomethane [0438]
UPLC ultra-performance liquid chromatography
General Procedures
[0439] Reactions sensitive to moisture or air were performed under
nitrogen or argon atmosphere using anhydrous solvents and reagents.
The progress of reactions was determined using either analytical
thin layer chromatography (TLC) usually performed with E. Merck
pre-coated TLC plates, silica gel 60F-254, layer thickness 0.25 mm
or liquid chromatography-mass spectrometry (LC-MS).
[0440] The analytical UPLC-MS system used consisted of a Waters
SQD2 platform with electrospray ionization in positive and negative
detection mode with an Acquity UPLC I-class solvent manager, column
manager, sample manager and PDA detector. The column used for
standard methods was a CORTECS UPLC C18 1.6 .mu.m, 2.1.times.30 mm,
and the column used for polars method was an ACQUITY UPLC HSST3 1.8
.mu.m, 2.1.times.30 mm, the column temperature was 40.degree. C.,
the flow rate was 0.7 mL/min, and injection volume was 1 .mu.L. UV
detection was in the range 210-400 nm. The mobile phase consisted
of solvent A (water plus 0.05% formic acid) and solvent B
(acetonitrile plus 0.05% formic acid) with different gradients for
4 different methods: 1/ Starting with 99% solvent A for 0.2 minutes
changing to 98% solvent B over 1 minutes, maintained for 0.4
minutes, then reverting to 99% solvent A over 0.1 min; 2/ Starting
with 99% solvent A for 0.5 minutes changing to 98% solvent B over
3.7 minutes, maintained for 0.4 minutes, then reverting to 99%
solvent A over 0.1 min; 3/ Starting with 100% solvent A for 0.4
minutes changing to 98% solvent B over 0.9 minutes, maintained for
0.3 minutes, then reverting to 100% solvent A over 0.1 min; 4/
Starting with 100% solvent A for 0.8 minutes changing to 98%
solvent B over 3.4 minutes, maintained for 0.4 minutes, then
reverting to 100% solvent A over 0.1 minutes.
[0441] The analytical LC-MS system used consisted of a Agilent 6140
quadrupole LC/MS platform with electrospray ionization in positive
and negative detection mode with an Agilent 1200 Series solvent
manager, column manager, sample manager and PDA detector. The
column for standard method was Purospher.RTM. STAR RP-18 endcapped
2 .mu.m, Hibar.RTM. HR 50-2.1, the column temperature was
60.degree. C., the flow rate was 0.8 mL/min, and injection volume
was 0.5-5 .mu.L. UV detection was in the range 210-400 nm. The
mobile phase consisted of solvent A (water plus 0.05% formic acid)
and solvent B (acetonitrile plus 0.05% formic acid) with different
gradients for 2 different methods: 1) Starting with 98% solvent A
changing to 100% solvent B over 1.8 minutes, maintained for 0.8
min; 2) Starting with 98% solvent A changing to 100% solvent B over
5.8 minutes, maintained for 0.3 minutes.
[0442] Preparative HPLC purifications were usually performed using
a mass spectrometry directed system. Usually they were performed on
a Waters Chromatography Workstation (MassLynx V4.1) configured with
LC-MS System Consisting of: Waters ZQ.TM. 2000 (quad MS system with
Electrospray Ionization), Waters 2545 Gradient Pump, Waters 2767
Injecto/Collector, Waters 2998 PDA Detector, the MS Conditions of:
100-1400 amu, Positive Electrospray, Collection Triggered by MS,
and a Waters SUNFIRE.RTM. C-18 5 micron, 19 mm (id).times.150 mm
column. The mobile phases consisted of mixtures of acetonitrile
(5-95%) in water containing 0.02% formic acid. Flow rates were
maintained at 20 mL/min, the injection volume was 500 to 3000
.mu.L, and the UV detection range was 210-400 nm. Mobile phase
gradients were optimized for the individual compounds. The
analytical system consisted of the same system with a Waters
SUNFIRE.RTM. C-18 5 .mu.m, 4.6.times.150 mm column, or a
XSelect.RTM. CSH.TM. C-18 5 .mu.m, 4.6.times.150 mm column. The
mobile phases consisted of mixtures of acetonitrile (5-95%) in
water containing 0.02% formic acid. Flow rates were maintained at
1.2 mL/min, the injection volume was 5 to 20 .mu.L. Preparative
HPLC were also performed on a Gilson system GX-281 (Trilution). The
column was a Waters SUNFIRE.RTM. Prep C18 5 .mu.m OBD, dimension
50.times.150 mm. The mobile phase consisted of acetonitrile (5-50%)
in water containing 0.02% HCOOH over 60 minutes. Flow rates were
maintained at 117 mL/min, the injection volume was 1000 to 7000
.mu.L, and the UV detection range was 260 nm.
[0443] Reactions performed using microwave irradiation were
normally carried out using an Emrys Optimizer manufactured by
Personal Chemistry, or an Initiator manufactured by Biotage.
Concentration of solutions was carried out on a rotary evaporator
in vacuo. Flash chromatography was usually performed using a
Biotage.RTM. Flash Chromatography apparatus (Isolera) on silica gel
(15-45.mu., 40-63, or spheric silica) in pre-packed cartridges of
the size noted. .sup.1H NMR spectra were acquired at 400 MHz or 500
MHz spectrometers in CDCl.sub.3 solutions unless otherwise noted.
Chemical shifts were reported in parts per million (ppm).
Tetramethylsilane (TMS) was used as internal reference in
CDCl.sub.3 solutions, and residual CH.sub.3OH peak or TMS was used
as internal reference in CD.sub.3OD solutions. Coupling constants
(J) were reported in hertz (Hz). Chiral analytical chromatography
was performed on one of CHIRALPAK.RTM. AS, CHIRALPAK.RTM. AD,
CHIRALCEL.RTM. OD, CHIRALCEL.RTM. IA, or CHIRALCEL.RTM. OJ columns
(250.times.4.6 mm) (Daicel Chemical Industries, Ltd.) with noted
percentage of either ethanol in hexane (% Et/Hex) or isopropanol in
heptane (% IPA/Hep) as isocratic solvent systems. Chiral
preparative chromatography was conducted on one of of CHIRALPAK AS,
of CHIRALPAK AD, CHIRALCEL.RTM. OD, CHIRALCEL.RTM. IA,
CHIRALCEL.RTM. OJ columns (20.times.250 mm) (Daicel Chemical
Industries, Ltd.) with desired isocratic solvent systems identified
on chiral analytical chromatography or by supercritical fluid (SFC)
conditions.
Preparation of Intermediate Compounds
Preparation of Intermediate Compound A
Method 1:
##STR00089##
[0444] Intermediate A: Ethyl (R)-4-hydroxy-2-methylbutanoate
[0445] Step 1: To a solution of (R)-4-(benzyloxy)-2-methylbutanoic
acid (11.43 g, 54.95 mmol) in DCM (170 mL) were added
N-(3-Dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (13.40
g, 70.16 mmol), 4-dimethylamino pyridine (0.69 g, 5.65 mmol) and
ethanol (34 mL, 0.58 mol). The reaction mixture was stirred under
nitrogen at RT overnight. The resulting reaction mixture was washed
with water (100 mL). The organic layer was washed with a 10%
solution of citric acid (100 mL) and with a 1:1 mixture of brine
and water (100 mL), dried over sodium sulfate, filtered and
concentrated under reduced pressure to afford ethyl
(R)-4-(benzyloxy)-2-methylbutanoate. Step 2: To a suspension of
palladium on carbon (1.64 g, 15.41 mmol) in isopropyl acetate (400
mL) was added a solution of ethyl
(R)-4-(benzyloxy)-2-methylbutanoate (15.34 g, 65.00 mmol) in
isopropyl acetate (50 mL). The reaction mixture was degassed few
times with nitrogen. Then the flask was filled with hydrogen and it
was stirred under an atmosphere of hydrogen for 7 h. The resulting
reaction mixture was filtered through a pad of celite and washed
with ethyl acetate (180 mL). The filtrates were concentrated under
reduced pressure to afford the expected intermediate.
Method 2:
##STR00090##
[0446] Intermediate A: Ethyl (R)-4-hydroxy-2-methylbutanoate
[0447] Step 1: a/ In a glass autoclave,
4-methoxy-2-methylene-4-oxobutanoic acid (50 g, 0.347 mol) and
methanol (70 mL) were stirred until complete dissolution. The
system was then pump-fill degassed with nitrogen (3.times.). b/ To
Rosphos bistriflate (95 mg, 0.142 mmol) under N.sub.2 was added
methanol (1.5 mL, purged with N.sub.2), followed by
diisopropylethylamine (36.5 mg, 0.283 mmol) in methanol (0.5 mL).
c/ To a solution of Rh(COD).sub.2OTf (63 mg, 0.135 mmol) purged
with N.sub.2 in methanol (1 mL, purged with N.sub.2), cooled in an
ice/methanol bath, was added dropwise over 2 min the phosphine
solution prepared above. This solution was allowed to warm to RT,
and then added to the autoclave, via syringe. The autoclave was
took to 5 bar of hydrogen. The reaction mixture was stirred at
50.degree. C. for 1 h, and then at 60.degree. C. for 50 min. The
system was allowed to cool to RT overnight. The resulting solution
was transferred to an RBF with methanol, and concentrated under
reduced pressure to afford the expected intermediate without
further purification.
[0448] Step 2: To a solution of
(R)-4-methoxy-2-methyl-4-oxobutanoic acid (52 g, 356 mmol) in THF
(2 L) was added lithium borohydride (16.5 g, 712 mmol) portionwise
over 40 min at RT. The reaction mixture was stirred at RT for 40
min, and then ethanol (400 mL) was added dropwise over 90 min. The
reaction mixture was then stirred at RT overnight, and then
concentrated under reduced pressure, azeotroping with THF to afford
the expected intermediate used directly in the next step without
further purification.
[0449] Step 3: To a suspension of previous intermediate in DMF (800
mL) was added bromoethane (270 mL, 3.6 mol). The reaction mixture
was stirred for 2 days. The reaction mixture was extracted with
MTBE and a pH 6.75 buffer (NaH.sub.2PO.sub.4.H.sub.2O, 55.7 g;
Na.sub.2HPO.sub.4.2H.sub.2O, 61.6 g in 1 L of water), and then with
brine. The organic layers were dried, filtered and concentrated
under reduced pressure at 30.degree. C. to afford the crude
expected compound. This material was purified by column
chromatography (DCM/EtOAc: 0 to 30%)
Preparation of Intermediate Compound B
##STR00091##
[0450] Intermediate B: Isopropyl
(R)-4-hydroxy-2-methylbutanoate
[0451] Step 1: To a stirred solution of
(R)-4-(benzyloxy)-2-methylbutanoic acid (15.00 g, 72.00 mmol) in
DCM (220 mL), was added isopropanol (55.2 mL, 0.72 mol) followed by
the addition of a solution of
N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (16.60
g, 86.44 mmol) in DCM (100 mL). The reaction mixture was stirred,
and then, 4-dimethylamino pyridine (0.88 g, 7.20 mmol) was added to
the mixture. The reaction mixture was stirred under nitrogen at RT
overnight. The resulting reaction mixture was washed with water
(250 mL). The organic layer was washed with a 10% solution of
citric acid (.times.2) and with brine, then it was dried over
sodium sulfate, filtered and concentrated under reduced pressure to
afford isopropyl (R)-4-(benzyloxy)-2-methylbutanoate. Step 2: To a
suspension of palladium on carbon (3.30 g, 31.00 mmol) in isopropyl
acetate (50 mL) was added a solution of isopropyl
(R)-4-(benzyloxy)-2-methylbutanoate (16.50 g, 66.00 mmol) in
isopropyl acetate (500 mL). The reaction mixture was degassed few
times with nitrogen. Then the flask was filled with hydrogen and it
was stirred under an atmosphere of hydrogen overnight. The reaction
mixture was filtered through a pad of celite. The filtrates were
concentrated under reduced pressure (water bath: 30.degree. C.) and
the resulting product was dried in vacuum oven for 4 h to afford
the expected intermediate.
Preparation of Intermediate Compound C
##STR00092##
[0452] Intermediate C: Isopropyl
(S)-4-hydroxy-2-methylbutanoate
[0453] Step 1: To a stirred solution of
(S)-4-(benzyloxy)-2-methylbutanoic acid (21.0 g, 101 mmol) in DCM
(420 mL), was added isopropanol (38.5 mL, 504 mmol),
N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (23.20
g, 121 mmol) and 4-dimethylamino pyridine (1.23 g, 10.08 mmol). The
reaction mixture was stirred under nitrogen at RT overnight. The
resulting reaction mixture was washed with water (250 mL). The
organic layer was washed with a 10% solution of citric acid
(.times.2) and with brine, then it was dried over sodium sulfate,
filtered and concentrated under reduced pressure to afford the
expected intermediate; MS (ESI) m/z=251.3 (MH.sup.+). Step 2: To
isopropyl (S)-4-(benzyloxy)-2-methylbutanoate (25.3 g, 101 mmol)
was added propan-2-ol (500 mL) and dihydroxypalladium (2.13 g,
15.15 mmol). The reaction mixture was degassed few times with
nitrogen. Then the flask was filled with hydrogen and the reaction
mixture was stirred under an atmosphere of hydrogen for 9 h. The
reaction mixture was filtered through a pad of celite. The
filtrates were concentrated under reduced pressure (water bath:
30.degree. C.) to afford the expected intermediate; .sup.1H NMR
(CDCl.sub.3, 400 MHz) .delta. (ppm) 4.99 (heptuplet, J=6.24 Hz,
1H), 3.70-3.61 (m, 2H), 2.60-2.51 (m, 1H), 1.94-1.85 (m, 1H),
1.70-1.62 (m, 1H), 1.22-1.20 (m, 6H), 1.15 (d, J=7.00 Hz, 3H); MS
(ESI) m/z=183.0 (MNa.sup.+).
Preparation of Intermediate Compound D
##STR00093##
[0454] Intermediate D: Ethyl (S)-4-hydroxy-2-methylbutanoate
[0455] Step 1: To a solution of (S)-4-(benzyloxy)-2-methylbutanoic
acid (30.0 g, 144 mmol) in DCM (600 mL) were added
N-(3-Dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (33.1
g, 173 mmol), 4-dimethylamino pyridine (1.76 g, 14.41 mmol) and
ethanol (42.1 mL, 720 mmol). The reaction mixture was stirred under
nitrogen at RT overnight. The resulting reaction mixture was washed
with water (250 mL). The organic layer was washed with a 10%
solution of citric acid (250 mL) and with brine, and then filtered
through a phase separator and concentrated under reduced pressure
to afford the expected intermediate; MS (ESI) m/z=237.1 (MH.sup.+).
Step 2: To a solution of ethyl (S)-4-(benzyloxy)-2-methylbutanoate
(29.5 g, 125 mmol) in ethanol (624 mL) was added dihydroxypalladium
(2.63 g, 18.73 mmol). The reaction mixture was degassed few times
with nitrogen. Then the flask was filled with hydrogen and the
reaction mixture was stirred under an atmosphere of hydrogen for 9
h. The reaction mixture was filtered through a pad of celite. The
filtrates were concentrated under reduced pressure to afford the
expected intermediate; .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta.
(ppm) 4.04 (q, J=7.09 Hz, 2H), 3.56 (t, J=6.40 Hz, 2H), 2.57-2.48
(m, 1H), 1.81-1.79 (m, 1H), 1.61-1.53 (m, 1H), 1.18 (t, J=7.09 Hz,
3H), 1.09 (d, J=6.99 Hz, 3H).
Preparation of Intermediate Compounds E and F
##STR00094##
[0456] Intermediate E: Isopropyl 4-hydroxy-2-methylbutanoate
[0457] Step 1: A solution of .alpha.-methyl-.gamma.-butyrolactone
(50.0 g, 499 mmol) in 1M aqueous potassium hydroxide solution (499
mL, 499 mmol) was stirred under reflux for 3 hours, then cooled to
RT and concentrated under reduced pressure. The crude solid was
triturated in diethyl ether, filtered off and washed with diethyl
ether. The solid was then dried in vacuo over P.sub.2O.sub.5 at
45.degree. C. Step 2: To a solution of the product of step 1 (25.0
g, 160.0 mmol) in DMF (200 mL) was added dropwise at RT under
nitrogen 2-iodopropane (31.9 mL, 320 mmol). The reaction mixture
was stirred at RT for 5 h. 2-iodopropane (8.0 mL, 80 mmol) was
added, and the reaction mixture was stirred at RT overnight. The
mixture was diluted with EtOAc, and the organic layer was washed
with a metabisulfite solution and brine. The organic layer was
dried, filtered and concentrated under reduced pressure at
20-30.degree. C. to provide the expected intermediate. .sup.1H NMR
(400 MHz, CDCl.sub.3) .delta. 4.97 (heptuplet, J=6.23 Hz, 1H),
3.66-3.60 (m, 2H), 2.59-2.50 (m, 1H), 1.92-1.84 (m, 1H), 1.67-1.59
(m, 1H), 1.195 (d, J=6.23 Hz, 3H), 1.19 (d, J=6.23 Hz, 3H), 1.14
(d, J=7.09 Hz, 3H).
Intermediate F
[0458] Intermediate F was synthesized using the method described
for the synthesis of intermediate compound E starting for step 2
from iodoethane (1.2 eq.). .sup.1H NMR (400 MHz, CDCl.sub.3)
.delta. 4.14 (q, J=7.11 Hz, 2H), 3.69-3.63 (m, 2H), 2.67-2.58 (m,
1H), 1.98-1.89 (m, 1H), 1.71-1.63 (m, 1H), 1.26 (t, J=7.11 Hz, 3H),
1.19 (d, J=7.10 Hz, 3H).
Example 1
Preparation of Compounds 1A and 1B
##STR00095##
[0459] Compounds 1A/1B: Ethyl
(R)-4-(((2S,4aR,6R,7S,7aS)-6-(4-amino-2-oxopyrimidin-1(2H)-yl)-7-fluoro-2-
-oxidotetrahydro-4H-thieno[3,2-d][1,3,2]dioxaphosphinin-2-yl)oxy)-2-methyl-
butanoate and Ethyl
(R)-4-(((2R,4aR,6R,7S,7aS)-6-(4-amino-2-oxopyrimidin-1(2H)-yl)-7-fluoro-2-
-oxidotetrahydro-4H-thieno[3,2-d][1,3,2]dioxaphosphinin-2-yl)oxy)-2-methyl-
butanoate
[0460] Step 1: A solution of phosphorus (V) oxychloride (0.12 mL,
1.29 mmol) in anhydrous DCM (4 mL) was cooled to -20.degree. C. A
solution of intermediate A (171 mg, 1.17 mmol) and triethylamine
(0.18 mL, 1.29 mmol) in anhydrous DCM (3 mL) was added to the
previous solution dropwise at -20.degree. C. The reaction mixture
was stirred at -25.degree. C. for 64 h. The resulting reaction
mixture was warmed to RT and solvent was removed in vacuo at
30.degree. C. Diethyl ether was added to the residue, filtered and
concentrated under reduced pressure to afford the expected
intermediate, which was used in the next step without further
purification.
[0461] Step 2: To a solution of previous intermediate of step 1 in
DCM (10 mL) was added dropwise a solution of
1-((2R,3S,4S,5R)-3-fluoro-4-hydroxy-5-(hydroxymethyl)tetrahydrothiophen-2-
-yl)-4-(((4-methoxyphenyl)diphenylmethyl)amino)pyrimidin-2(1H)-one
(500 mg, 0.94 mmol) and triethylamine (0.45 mL, 3.23 mmol) in DCM
(3 mL). The reaction mixture was stirred for 30 min at RT, and
then, a solution of 1-methylimidazole (77 mg, 0.94 mmol) in DCM
(0.5 mL) was added. The reaction mixture was stirred at RT
overnight, and then, concentrated under reduced pressure. The crude
residue was dissolved in ethyl acetate (100 mL) and the resulting
solution was washed with brine. The organic layer was dried over
sodium sulfate, filtered and concentrated under reduced pressure.
The crude compound was purified via silica gel flash column
chromatography (DCM/EtOAc 50%) to afford the 2 expected isomers Sp
and Rp. Diastereoisomer 1:MS (ESI) m/z=723.8 (MH.sup.+);
Diastereoisomer 2:MS (ESI) m/z=723.6 (MH.sup.+).
[0462] Step 3: To a solution of diastereoisomer 1 of ethyl
(2R)-4-(((4aR,6R,7S,7aS)-7-fluoro-6-(4-(((4-methoxyphenyl)diphenylmethyl)-
amino)-2-oxopyrimidin-1
(2H)-yl)-2-oxidotetrahydro-4H-thieno[3,2-d][1,3,2]dioxaphosphinin-2-yl)ox-
y)-2-methylbutanoate (40 mg, 55 .mu.mol) in DCM (5 mL) was added
triethylsilane (0.5 mL). A solution of trifluoroacetic acid (0.5
mL) in DCM (5 mL) was added dropwise to the reaction mixture. The
reaction mixture was stirred at RT for 90 min. Solvent was removed
under reduced pressure and the residue was azeotroped with toluene
(.times.2) (30.degree. C. bath). The crude residue was purified via
silica gel flash column chromatography (DCM/EtOH 0-10%) to afford
the expected isomer.
[0463] Compound 1 Diastereoisomer 1: .sup.1H NMR (DMSO-d.sub.6, 400
MHz) .delta. (ppm) 8.13 (d, J=7.61 Hz, 1H), 7.47-7.41 (m, 2H),
6.62-6.61 (m, 1H), 5.81 (d, J=7.61 Hz, 1H), 5.51-5.33 (m, 1H),
5.29-5.20 (m, 1H), 4.69-4.58 (m, 2H), 4.15-4.04 (m, 2H), 4.07 (q,
J=7.03 Hz, 2H), 3.72-3.66 (m, 1H), 2.59-2.53 (m, 1H), 1.99-1.90 (m,
1H), 1.75-1.67 (m, 1H), 1.19 (t, J=7.03 Hz, 3H), 1.11 (d, J=7.17
Hz, 3H); .sup.31P NMR (DMSO-d.sub.6, 162 MHz) .delta. (ppm) -6.59
(s, 1P); 19F NMR (DMSO-d.sub.6, 376 MHz) .delta. (ppm) -198.26 (s,
1F); MS (ESI) m/z=452.4 (MH.sup.+).
[0464] Step 3 bis: To a solution of diastereoisomer 2 of ethyl
(2R)-4-(((4aR,6R,7S,7aS)-7-fluoro-6-(4-(((4-methoxyphenyl)diphenylmethyl)-
amino)-2-oxopyrimidin-1
(2H)-yl)-2-oxidotetrahydro-4H-thieno[3,2-d][1,3,2]dioxaphosphinin-2-yl)ox-
y)-2-methylbutanoate (35 mg, 48 .mu.mol) in DCM (5 mL) was added
triethylsilane (0.5 mL). A solution of trifluoroacetic acid (0.5
mL) in DCM (5 mL) was added dropwise to the reaction mixture. The
reaction mixture was stirred at RT for 2 h. Solvent was removed
under reduced pressure and the residue was azeotroped with toluene
(.times.2) (30.degree. C. bath). The crude residue was purified via
silica gel flash column chromatography (DCM/EtOH 0-10%) to afford
the expected isomer.
[0465] Compound 1 Diastereoisomer 2: .sup.1H NMR (DMSO-d.sub.6, 400
MHz) .delta. (ppm) 8.03 (d, J=7.41 Hz, 1H), 7.35 (brs, 2H),
6.66-6.63 (m, 1H), 5.81 (d, J=7.41 Hz, 1H), 5.52-5.35 (m, 1H),
4.98-4.89 (m, 1H), 4.70-4.60 (m, 1H), 4.47-4.40 (m, 1H), 4.12-4.05
(m, 4H), 3.69-3.63 (m, 1H), 2.64-2.61 (m, 1H), 2.06-1.97 (m, 1H),
1.86-1.78 (m, 1H), 1.18 (t, J=7.06 Hz, 3H), 1.13 (d, J=7.08 Hz,
3H); .sup.31P NMR (DMSO-d.sub.6, 162 MHz) .delta. (ppm) -7.99 (s,
1P); 19F NMR (DMSO-d.sub.6, 376 MHz) .delta. (ppm) -197.94 (s, 1F);
MS (ESI) m/z=452.3 (MH.sup.+).
Example 2
Preparation of Compounds 2A and 2B
##STR00096##
[0466] Compounds 2A/2B: Isopropyl
(R)-4-(((2S,4aR,6R,7S,7aS)-6-(4-amino-2-oxopyrimidin-1(2H)-yl)-7-fluoro-2-
-oxidotetrahydro-4H-thieno[32-d][1,3,2]dioxaphosphinin-2-yl)oxy)-2-methylb-
utanoate and Isopropyl
(R)-4-(((2R,4aR,6R,7S,7aS)-6-(4-amino-2-oxopyrimidin-1(2H)-yl)-7-fluoro-2-
-oxidotetrahydro-4H-thieno[3,2-d][1,3,2]dioxaphosphinin-2-yl)oxy)-2-methyl-
butanoate
[0467] Compounds 2A/2B were synthesized according to a similar
procedure than the procedure described for Compounds 1A/1B using
Intermediate B as starting material.
[0468] Compound 2 Diastereoisomer 1: .sup.1H NMR (DMSO-d.sub.6, 400
MHz) .delta. (ppm) 9.01 (brs, 1H), 8.45-8.42 (m, 2H), 6.49 (d,
J=7.96 Hz, 1H), 6.03 (d, J=7.83 Hz, 1H), 5.58-5.41 (m, 1H),
5.25-5.17 (m, 1H), 4.89 (heptuplet, J=6.26 Hz, 1H), 4.72-4.60 (m,
2H), 4.13-4.06 (m, 2H), 3.72-3.66 (m, 1H), 2.53-2.52 (m, 1H),
1.98-1.89 (m, 1H), 1.74-1.66 (m, 1H), 1.185 (d, J=6.26 Hz, 3H),
1.18 (d, J=6.26 Hz, 3H), 1.10 (d, J=7.05 Hz, 3H); .sup.31P NMR
(DMSO-d.sub.6, 162 MHz) .delta. (ppm) -6.53 (s, 1P); .sup.19F NMR
(DMSO-d.sub.6, 376 MHz) .delta. (ppm) -198.18 (s, 1F); MS (ESI)
m/z=466.5 (MH.sup.+).
[0469] Compound 2 Diastereoisomer 2: .sup.1H NMR (DMSO-d.sub.6, 400
MHz) .delta. (ppm) 8.48 (brs, 1H), 8.26 (d, J=7.65 Hz, 1H), 8.07
(brs, 1H), 6.55 (d, J=8.09 Hz, 1H), 5.97 (d, J=7.65 Hz, 1H),
5.58-5.41 (m, 1H), 4.96-4.87 (m, 2H), 4.72-4.62 (m, 1H), 4.44 (t,
J=10.50 Hz, 1H), 4.13-4.05 (m, 2H), 3.70-3.63 (m, 1H), 2.62-2.55
(m, 1H), 2.05-1.97 (m, 1H), 1.84-1.76 (m, 1H), 1.18 (d, J=6.28 Hz,
3H), 1.175 (d, J=6.28 Hz, 3H), 1.11 (d, J=7.10 Hz, 3H); .sup.31P
NMR (DMSO-d.sub.6, 162 MHz) .delta. (ppm) -8.04 (s, 1P); .sup.19F
NMR (DMSO-d.sub.6, 376 MHz) .delta. (ppm) -197.83 (s, 1F); MS (ESI)
m/z=466.5 (MH.sup.+).
Example 3
Preparation of Compounds 3A and 3B
##STR00097##
[0470] Compounds 3A/3B: Isopropyl
(S)-4-(((2S,4aR,6R,7aS)-6-(6-amino-2-chloro-9H-purin-9-yl)-2-oxidotetrahy-
dro-4H-furo[3,2-d][1,3,2]dioxaphosphinin-2-yl)oxy)-2-methylbutanoate
and Isopropyl
(S)-4-(((2R,4aR,6R,7aS)-6-(6-amino-2-chloro-9H-purin-9-yl)-2-ox-
idotetrahydro-4H-furo[3,2-d][1,3,2]dioxaphosphinin-2-yl)oxy)-2-methylbutan-
oate
[0471] Step 1: To a -15.degree. C. solution of
1-chloro-N,N,N',N'-tetraisopropylphosphinediamine (15 g, 56.2 mmol)
and triethylamine (7.84 mL, 56.2 mmol) in diethyl ether (187 mL),
under nitrogen, was added dropwise a solution of isopropyl
(S)-4-hydroxy-2-methylbutanoate (9.01 g, 56.2 mmol) in diethyl
ether (94 mL). The reaction mixture was stirred at -15.degree. C.
for 1 h and then at RT for 2 h. The resulting suspension was
filtered under nitrogen and washed with diethyl ether. The filtrate
was concentrated in vacuo at RT under nitrogen to provide a crude
intermediate compound, which was stored at -20.degree. C. under
nitrogen and was directly used in the next step without further
purification: .sup.31P NMR (162 MHz, CDCl.sub.3) .delta. 124.0 (s,
1P).
[0472] Step 2: To a solution of THF (3.2 mL) under nitrogen at RT
were simultaneously slowly added a solution of
(2R,3S,5R)-5-(6-amino-2-chloro-9H-purin-9-yl)-2-(hydroxymethyl)tetrahydro-
furan-3-ol (400 mg, 1.4 mmol) and 1H-imidazole-4,5-dicarbonitrile
(413 mg, 3.5 mmol) (coevaporated 3 times with CH.sub.3CN and THF)
in THF (10.2 mL) and CH.sub.3CN (5.1 mL), and a solution of the
intermediate compound of step 1 (729 mg, 1.68 mmol) in THF (3.2
mL). The reaction mixture was stirred at RT overnight. Hydrogen
peroxide (0.21 mL, 7.0 mmol) was then added dropwise at RT for 15
min. The reaction mixture was stirred at RT for 3 hours, and then,
diluted with EtOAc and water. The aqueous layer was washed twice
with EtOAc. The combined organic layers were washed with brine,
dried, and concentrated under reduced pressure. The crude residue
was purified by flash chromatography on silica gel (DCM/MeOH: 0 to
10%) followed by preparative MS/HPLC to afford the 2 title
compounds (isolated diasteromers at P).
[0473] Compound 3 Diastereoisomer 1: .sup.1H NMR (DMSO-d.sub.6, 400
MHz) .delta. (ppm) 8.38 (s, 1H), 7.90 (brs, 2H), 6.46-6.43 (m, 1H),
5.32-5.26 (m, 1H), 4.89 (heptuplet, J=6.25 Hz, 1H), 4.61 (ddd,
J=21.97 Hz, 9.26 Hz, 4.63 Hz, 1H), 4.34-4.29 (m, 1H), 4.18-4.09 (m,
2H), 4.05-3.99 (m, 1H), 2.80-2.76 (m, 2H), 2.68-2.59 (m, 1H),
2.09-2.00 (m, 1H), 1.86-1.78 (m, 1H), 1.16 (d, J=6.25 Hz, 6H), 1.13
(d, J=7.00 Hz, 3H); .sup.31P NMR (DMSO-d.sub.6, 162 MHz) .delta.
(ppm) -6.61 (s, 1P); MS (ESI) m/z=490.3 (MH.sup.+).
[0474] Compound 3 Diastereoisomer 2: .sup.1H NMR (DMSO-d.sub.6, 400
MHz) .delta. (ppm) 8.46 (s, 1H), 7.89 (brs, 2H), 6.48-6.45 (m, 1H),
5.33-5.26 (m, 1H), 4.91 (heptuplet, J=6.33 Hz, 1H), 4.66-4.59 (m,
1H), 4.47-4.40 (m, 1H), 4.16-4.06 (m, 3H), 2.83-2.72 (m, 2H),
2.57-2.53 (m, 1H), 2.01-1.92 (m, 1H), 1.76-1.68 (m, 1H), 1.21 (d,
J=6.33 Hz, 3H), 1.20 (d, J=6.33 Hz, 3H), 1.12 (d, J=7.06 Hz, 3H);
.sup.31P NMR (DMSO-d.sub.6, 162 MHz) .delta. (ppm) -4.93 (s, 1P);
MS (ESI) m/z=490.3 (MH.sup.+).
Example 4
Preparation of Compounds 4A and 4B
##STR00098##
[0475] Compounds 4A/4B: Isopropyl
(S)-4-(((2S,4aR,6R,7S,7aR)-6-(6-amino-2-chloro-9H-purin-9-yl)-7-fluoro-2--
oxidotetrahydro-4H-furo[3,2-d][1,3,2]dioxaphosphinin-2-yl)oxy)-2-methylbut-
anoate and Isopropyl
(S)-4-(((2R,4aR,6R,7S,7aR)-6-(6-amino-2-chloro-9H-purin-9-yl)-7-fluoro-2--
oxidotetrahydro-4H-furo[3,2-d][1,3,2]dioxaphosphinin-2-yl)oxy)-2-methylbut-
anoate
[0476] Compounds 4A/4B were synthesized according to a similar
procedure than the procedure described for Compounds 3A/3B using
(2R,3R,4S,5R)-5-(6-amino-2-chloro-9H-purin-9-yl)-4-fluoro-2-(hydroxymethy-
l)tetrahydrofuran-3-ol as starting material.
[0477] Compound 4 Diastereoisomer 1: .sup.1H NMR (DMSO-d.sub.6, 400
MHz) .delta. (ppm) 8.43 (s, 1H), 7.98 (brs, 2H), 6.63-6.60 (m, 1H),
5.96-5.79 (m, 1H), 5.60-5.50 (m, 1H), 4.90 (heptuplet, J=6.29 Hz,
1H), 4.75-4.66 (m, 1H), 4.50-4.46 (m, 1H), 4.24-4.13 (m, 3H),
2.65-2.58 (m, 1H), 2.11-2.03 (m, 1H), 1.87-1.79 (m, 1H), 1.18 (d,
J=6.29 Hz, 6H), 1.12 (d, J=7.06 Hz, 3H); .sup.31P NMR
(DMSO-d.sub.6, 162 MHz) .delta. (ppm) -6.92 (s, 1P); .sup.19F NMR
(DMSO-d.sub.6, 376 MHz) .delta. (ppm) -200.16 (s, 1F); MS (ESI)
m/z=508.3 (MH.sup.+).
[0478] Compound 4 Diastereoisomer 2: .sup.1H NMR (DMSO-d.sub.6, 400
MHz) .delta. (ppm) 8.54-8.53 (m, 1H), 7.98 (brs, 2H), 6.65-6.62 (m,
1H), 5.92-5.74 (m, 1H), 5.67-5.57 (m, 1H), 4.91 (heptuplet, J=6.33
Hz, 1H), 4.74-4.67 (m, 1H), 4.61-4.54 (m, 1H), 4.32-4.26 (m, 1H),
4.18-4.09 (m, 2H), 2.57-2.54 (m, 1H), 2.02-1.93 (m, 1H), 1.78-1.69
(m, 1H), 1.205 (d, J=6.33 Hz, 3H), 1.20 (d, J=6.33 Hz, 3H), 1.12
(d, J=7.03 Hz, 3H); .sup.31P NMR (DMSO-d.sub.6, 162 MHz) .delta.
(ppm) -5.20 (s, 1P); .sup.19F NMR (DMSO-d.sub.6, 376 MHz) .delta.
(ppm) -199.57 (s, 1F); MS (ESI) m/z=508.3 (MH.sup.+).
Example 5
Preparation of Compounds 5A and 5B
##STR00099##
[0479] Compounds 5A/5B: Isopropyl
(R)-4-(((2S,4aR,6R,7S,7aS)-6-(2-amino-6-methoxy-9H-purin-9-yl)-7-hydroxy--
2-oxidotetrahydro-4H-furo[3,2-d][1,3,2]dioxaphosphinin-2-yl)oxy)-2-methylb-
utanoate and Isopropyl
(R)-4-(((2R,4aR,6R,7S,7aS)-6-(2-amino-6-methoxy-9H-purin-9-yl)-7-hydroxy--
2-oxidotetrahydro-4H-furo[3,2-d][1,3,2]dioxaphosphinin-2-yl)oxy)-2-methylb-
utanoate
[0480] Step 1: was similar than the procedure described for Step 1
of Compounds 3A/3B using Intermediate B as starting material.
.sup.31P NMR (162 MHz, CDCl.sub.3) .delta. 123.89 (s, 1P).
[0481] Step 2: To a solution of
(2R,3S,4R,5R)-2-(2-amino-6-methoxy-9H-purin-9-yl)-4-hydroxy-5-(hydroxymet-
hyl)tetrahydrofuran-3-yl benzyl carbonate (500 mg, 1.16 mmol) in
anhydrous pyridine (11 mL, 9.5 mL/mmol) was added under nitrogen at
RT 1H-tetrazole (11 mL, 9.5 mL/mmol). The reaction mixture was
cooled to -5.degree. C. and a solution of the intermediate compound
of step 1 (453 mg, 1.16 mmol) in CH.sub.3CN (5 mL) was added
dropwise. The reaction mixture was stirred at 0.degree. C. for 1 h
and then at RT for 2 h. The reaction was monitored by LC/MS. A
solution of tert-butylhydroperoxide, 5M in decane (0.6 mL, 0.5
mL/mmol) was then added dropwise and the resulting reaction mixture
was allowed to stir for 1 h at RT. The crude reaction mixture was
concentrated under reduced pressure. The crude residue was purified
using flash chromatography on silica gel (DCM/MeOH: 0 to 10%) to
afford the intermediate compound as a mixture of diastereomers.
[0482] Step 3: To a solution of isopropyl
(2R)-4-(((4aR,6R,7S,7aR)-6-(2-amino-6-methoxy-9H-purin-9-yl)-7-(((benzylo-
xy)carbonyl)oxy)-2-oxidotetrahydro-4H-furo[3,2-d][1,3,2]dioxaphosphinin-2--
yl)oxy)-2-methylbutanoate (468 mg, 0.74 mmol) in propan-2-ol (5 mL)
was added palladium on carbon (78 mg, 0.74 mmol). The reaction
mixture was degassed few times with nitrogen. Then the flask was
filled with hydrogen and the reaction mixture was stirred under
hydrogen for 3 h. The reaction mixture was filtered through a pad
of celite. The filtrate was concentrated under reduced pressure and
purified by RP-18 chromatography (H.sub.2O/CH.sub.3CN) to afford
the expected compound as a mixture of diastereoisomers.
[0483] Compound 5: Mixture of Diastereoisomers 5A/5B: .sup.1H NMR
(DMSO-d.sub.6, 400 MHz) .delta. (ppm) 8.07 (s, 0.5H), 8.05 (s,
0.5H), 6.47 (brs, 2H), 6.31-6.29 (m, 1H), 6.07-6.04 (m, 1H),
5.04-4.99 (m, 0.5H), 4.95-4.87 (m, 1H), 4.74-4.66 (m, 1.5H),
4.62-4.44 (m, 2H), 4.20-4.03 (m, 3H), 3.96 (s, 3H), 2.63-2.55 (m,
1H), 2.09-1.91 (m, 1H), 1.86-1.68 (m, 1H), 1.21-1.18 (m, 6H),
1.15-1.10 (m, 3H); .sup.31P NMR (DMSO-d.sub.6, 162 MHz) .delta.
(ppm) -4.87 (s, 0.5P), -6.40 (s, 0.5P); MS (ESI) m/z=502.2
(MH.sup.+).
Example 6
Preparation of Compounds 6A and 6B
##STR00100##
[0484] Compounds 6A/6B: Isopropyl
4-(((2S,4aR,6R,7aS)-6-(2,4-dioxo-5-(trifluoromethyl)-3,4-dihydropyrimidin-
-1
(2H)-yl)-2-oxidotetrahydro-4H-furo[3,2-d][1,3,2]dioxaphosphinin-2-yl)ox-
y)-2-methylbutanoate and Isopropyl
4-(((2R,4aR,6R,7aS)-6-(2,4-dioxo-5-(trifluoromethyl)-3,4-dihydropyrimidin-
-1
(2H)-yl)-2-oxidotetrahydro-4H-furo[3,2-d][1,3,2]dioxaphosphinin-2-yl)ox-
y)-2-methylbutanoate
[0485] Compounds 6A/6B were synthesized according to a similar
procedure than the procedure described for Compounds 3A/3B using
1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-5-(trifluo-
romethyl)pyrimidine-2,4(1H,3H)-dione and Intermediate E as starting
materials.
[0486] Compound 6: Mixture of Diastereoisomers 6A/6B: .sup.1H NMR
(DMSO-d.sub.6, 400 MHz) .delta. (ppm) 12.00 (brs, 1H), 8.16 (s,
0.1H), 8.13 (s, 0.9H), 6.23-6.17 (m, 1H), 4.90 (heptuplet, J=6.28
Hz, 1H), 4.85-4.77 (m, 1H), 4.67-4.59 (m, 1H), 4.41-4.36 (m, 1H),
4.13-4.03 (m, 2H), 3.98-3.92 (m, 1H), 2.65-2.55 (m, 2H), 2.04-1.92
(m, 1H), 1.82-1.71 (m, 1H), 1.19 (d, J=6.28 Hz, 6H), 1.12 (d,
J=7.00 Hz, 3H); .sup.31P NMR (DMSO-d.sub.6, 162 MHz) .delta. (ppm)
-4.84 (s, 0.1P), -6.69 (s, 0.9P); .sup.19F NMR (DMSO-d.sub.6, 376
MHz) .delta. (ppm) -61.43 (s, 0.1F), -61.58-(-61.59) (s, 0.9F); MS
(ESI) m/z=523.2 (MNa.sup.+).
Example 7
Preparation of Compounds 7A and 7B
##STR00101##
[0488] Step 1: To a solution of 2,3,4,5,6-pentafluorophenol (2.77
g, 15.05 mmol) in DCM (25 mL) were added at -75.degree. C. a
solution of 4-chlorophenyl phosphodichloridate (3.69 g, 15.05 mmol)
in DCM (25 mL), followed by triethylamine (2.01 mL, 15.05 mmol)
dropwise. The reaction mixture was stirred at -75.degree. C. for 30
min, then allowed to warm up to 0.degree. C. A solution of
intermediate D (2.5 g, 15.05 mmol) in DCM (25 mL) was then added
followed by triethylamine (4.20 mL, 30.10 mmol) dropwise. The
reaction mixture was stirred at RT for 1 h and then concentrated
under reduced pressure. .sup.31P NMR (DMSO-d.sub.6, 162 MHz)
.delta. (ppm) -11.57 (s, 1P).
[0489] Step 2: To a solution of
2-((2-amino-6-ethoxy-9H-purin-9-yl)methoxy)propane-1,3-diol (50 mg,
0.18 mmol) in DMF (880 .mu.L) under nitrogen was added DBU (80
.mu.L, 0.53 mmol). After stirring for 10 min, a solution of
intermediate of step 1 (89 mg, 0.18 mmol) in DMF (880 .mu.L) was
then added dropwise at -10.degree. C. The reaction mixture was
stirred at 140.degree. C. under microwaves irradiation for 20 min.
The reaction mixture was then diluted with EtOAc and washed with
saturated NaHCO.sub.3 solution. The aqueous layer was extracted
with EtOAc. The organic layers were combined, dried, filtered and
concentrated under reduced pressure. The crude compound was
purified by preparative MS/HPLC to afford a mixture of
diastereoisomers.
[0490] Compound 7: Mixture of Diastereoisomers 7A/7B: .sup.31P NMR
(DMSO-d.sub.6, 162 MHz) .delta. (ppm) -7.12 (s, 0.5P), -9.09 (s,
0.5P); MS (ESI) m/z=474.6 (MH.sup.+).
[0491] The mixture of diastereoisomers was separated by preparative
chiral SFC (CHIRALPAK-AD-H, 20*150 mm; Mobile Phase A: 20% IPA;
Mobile Phase B CO.sub.2) to afford the 2 expected separated
diastereoisomers:
[0492] Compound 7 Diastereoisomer 1: .sup.1H NMR (DMSO-d.sub.6, 400
MHz) .delta. (ppm) 8.01 (s, 1H), 6.49 (s, 2H), 5.56 (s, 2H),
4.47-4.34 (m, 5H), 4.08-3.96 (m, 4H), 3.81 (m, 1H), 2.61-2.54 (m,
1H), 1.98-1.92 (m, 1H), 1.75-1.69 (m, 1H), 1.37-1.34 (m, 3H),
1.19-1.16 (m, 3H), 1.11-1.03 (m, 3H); .sup.31P NMR (DMSO-d.sub.6,
162 MHz) .delta. (ppm) -9.08 (s, 1P).
[0493] Compound 7 Diastereoisomer 2: .sup.1H NMR (DMSO-d.sub.6, 400
MHz) .delta. (ppm) 8.02 (s, 1H), 6.50 (s, 2H), 5.54 (s, 2H),
4.47-4.38 (m, 4H), 4.23-4.17 (m, 2H), 4.06-3.95 (m, 4H), 1.93-1.86
(m, 1H), 1.69-1.62 (m, 1H), 1.37-1.34 (m, 3H), 1.17-1.14 (m, 3H),
1.07-1.03 (m, 3H); .sup.31P NMR (DMSO-d.sub.6, 162 MHz) .delta.
(ppm) -7.11 (s, 1P).
Example 8
Preparation of Compounds 8A and 8B
##STR00102##
[0494] Compounds 8A/8B: Ethyl
4-(((2R,4aS,6S,7aR)-6-(6-amino-9H-purin-9-yl)-2-oxidotetrahydro-4H-furo[3-
,2-d][1,3,2]dioxaphosphinin-2-yl)oxy)-2-methylbutanoate and Ethyl
4-(((2S,4aS,6S,7aR)-6-(6-amino-9H-purin-9-yl)-2-oxidotetrahydro-4H-furo[3-
,2-d][1,3,2]dioxaphosphinin-2-yl)oxy)-2-methylbutanoate
[0495] Compounds 8A/8B were synthesized according to a similar
procedure than the procedure described for Compounds 3A/3B using
(2S,3R,5
S)-5-(6-amino-9H-purin-9-yl)-2-(hydroxymethyl)tetrahydrofuran-3-ol
and Intermediate F as starting materials.
[0496] Compound 8: Mixture of Diastereoisomers 8A/8B: .sup.1H NMR
(DMSO-d.sub.6, 400 MHz) .delta. (ppm) 8.40 (s, 0.2H), 8.36-8.35 (m,
0.8H), 8.19 (s, 0.2H), 8.16 (m, 0.8H), 7.36 (brs, 2H), 6.52-6.47
(m, 1H), 5.47-5.36 (m, 1H), 4.65-4.56 (m, 1H), 4.43-4.27 (m, 1H),
4.18-3.99 (m, 5H), 2.83-2.66 (m, 3H), 2.13-1.93 (m, 1H), 1.87-1.69
(m, 1H), 1.22-1.12 (m, 6H); .sup.31P NMR (DMSO-d.sub.6, 162 MHz)
.delta. (ppm) -4.86 (s, 0.1P), -4.88 (s, 0.1P), -6.60 (s, 0.8P); MS
(ESI) m/z=442.3 (MH.sup.+).
Example 9
Preparation of Compounds 9A1, 9A2, 9B1 and 9B2
##STR00103##
[0497] Compounds 9A1/9A2/9B1/9B2: Ethyl
(S)-2-methyl-4-(((2R,4aS,6S,7aR)-6-(5-methyl-2,4-dioxo-3,4-dihydropyrimid-
in-1
(2H)-yl)-2-oxidotetrahydro-4H-furo[3,2-d][1,3,2]dioxaphosphinin-2-yl)-
oxy)butanoate, Ethyl
(R)-2-methyl-4-(((2R,4aS,6S,7aR)-6-(5-methyl-2,4-dioxo-3,4-dihydropyrimid-
in-1
(2H)-yl)-2-oxidotetrahydro-4H-furo[3,2-d][1,3,2]dioxaphosphinin-2-yl)-
oxy)butanoate, Ethyl
(S)-2-methyl-4-(((2S,4aS,6S,7aR)-6-(5-methyl-2,4-dioxo-3,4-dihydropyrimid-
in-1
(2H)-yl)-2-oxidotetrahydro-4H-furo[3,2-d][1,3,2]dioxaphosphinin-2-yl)-
oxy)butanoate and Ethyl
(R)-2-methyl-4-(((2S,4aS,6S,7aR)-6-(5-methyl-2,4-dioxo-3,4-dihydropyrimid-
in-1
(2H)-yl)-2-oxidotetrahydro-4H-furo[3,2-d][1,3,2]dioxaphosphinin-2-yl)-
oxy)butanoate
[0498] Compounds 9A1/9A2/9B1/9B2 were synthesized according to a
similar procedure than the procedure described for Compounds 3A/3B
using 1-((2S,4R,5
S)-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-5-methylpyrimidine-2,-
4(1H,3H)-dione and Intermediate F as starting materials.
[0499] Compound 9: Mixture of Diastereoisomers 9A1/9A2/9B1/9B2:
.sup.1H NMR (DMSO-d.sub.6, 400 MHz) .delta. (ppm) 11.42 (s, 0.8H),
11.39 (s, 0.2H), 7.58 (s, 0.2H), 7.51 (s, 0.8H), 6.38-6.29 (m, 1H),
4.77 (q, J=9.06 Hz, 1H), 4.64-4.55 (m, 1H), 4.50-4.33 (m, 1H),
4.13-4.06 (m, 4H), 3.93-3.87 (m, 1H), 2.68-2.54 (m, 2H), 2.49-2.41
(m, 1H), 2.08-1.93 (m, 1H), 1.85-1.70 (m, 4H), 1.21-1.11 (m, 6H);
.sup.31P NMR (DMSO-d.sub.6, 162 MHz) .delta. (ppm) -4.92 (s, 0.1P),
-4.95 (s, 0.1P), -6.66 (s, 0.8P); MS (ESI) m/z=455.2
(MNa.sup.+).
[0500] The mixture of diastereoisomers was separated by preparative
chiral SFC to afford the 4 expected separated diastereoisomers:
[0501] Compound 9 Diastereoisomer 1: .sup.31P NMR (DMSO-d.sub.6,
162 MHz) .delta. (ppm) -4.94 (s, 1P).
[0502] Compound 9 Diastereoisomer 2: .sup.31P NMR (DMSO-d.sub.6,
162 MHz) (ppm) -4.91 (s, 1P).
[0503] Compound 9 Diastereoisomer 3: .sup.31P NMR (DMSO-d.sub.6,
162 MHz) .delta. (ppm) -6.66 (s, 1P).
[0504] Compound 9 Diastereoisomer 4: .sup.31P NMR (DMSO-d.sub.6,
162 MHz) .delta. (ppm) -6.66 (s, 1P).
Example 10
Preparation of Compound 10
##STR00104##
[0505] Compound 10: Isopropyl
(2R)-4-(((4aR,6R,7aR)-6-(4-benzamido-2-oxopyrimidin-1(2H)-yl)-7,7-difluor-
o-2-oxidotetrahydro-4H-furo[3,2-d][1,3,2]dioxaphosphinin-2-yl)oxy)-2-methy-
lbutanoate
[0506] To a solution of isopropyl
(2R)-4-(((4aR,6R,7aR)-6-(4-amino-2-oxopyrimidin-1(2H)-yl)-7,7-difluoro-2--
oxidotetrahydro-4H-furo[3,2-d][1,3,2]dioxaphosphinin-2-yl)oxy)-2-methylbut-
anoate (650 mg, 1.39 mmol), in DMF (4.6 mL), were added
N-ethyl-N-isopropylpropan-2-amine (630 .mu.L, 3.62 mmol), benzoic
anhydride (440 mg, 1.95 mmol) and DMAP (51 mg, 0.42 mmol). The
reaction mixture was stirred at RT for 3 hours. The crude compound
was purified by preparative MS/HPLC to afford the title compound.
.sup.1H NMR (DMSO-d.sub.6, 400 MHz) .delta. (ppm) 11.51 (brs, 1H),
8.31 (d, J=7.53 Hz, 1H), 8.03-8.01 (m, 2H), 7.67-7.63 (m, 1H),
7.56-7.52 (m, 2H), 7.44-7.43 (m, 1H), 6.48 (brs, 1H), 5.06 (brs,
1H), 4.90 (heptuplet, J=6.33 Hz, 1H), 4.83-4.74 (m, 1H), 4.65-4.60
(m, 1H), 4.44-4.38 (m, 1H), 4.21-4.13 (m, 2H), 2.63-2.55 (m, 1H),
2.09-2.01 (m, 1H), 1.85-1.77 (m, 1H), 1.19 (d, J=6.33 Hz, 3H),
1.185 (d, J=6.33 Hz, 3H), 1.13 (d, J=7.06 Hz, 3H); .sup.31P NMR
(DMSO-d.sub.6, 162 MHz) .delta. (ppm) -7.50 (s, 1P); .sup.19F NMR
(DMSO-d.sub.6, 376 MHz) .delta. (ppm) -115.93-(-117.39) (m, 2F); MS
(ESI) m/z=572.5 (MH.sup.+).
Example 11
Preparation of Compounds 11A and 11B
##STR00105##
[0507] Compounds 11A/11B: Ethyl
4-(((2R,4aS,6S,7aR)-6-(4-amino-2-oxopyrimidin-1(2H)-yl)-2-oxidotetrahydro-
-4H-furo[3,2-d][1,3,2]dioxaphosphinin-2-yl)oxy)-2-methylbutanoate
and ethyl
4-(((2S,4aS,6S,7aR)-6-(4-amino-2-oxopyrimidin-1(2H)-yl)-2-oxidotetr-
ahydro-4H-furo[3,2-d][1,3,2]dioxaphosphinin-2-yl)oxy)-2-methylbutanoate
(TFA salts)
[0508] Compounds 11A/11B were synthesized according to a similar
procedure than the procedure described for Compounds 3A/3B using
1-((2S,4R,5
S)-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-4-(((4-methoxyphenyl)-
diphenylmethyl)amino)pyrimidin-2(1H)-one and Intermediate F as
starting materials.
[0509] Compound 11: Mixture of Diastereoisomers 11A/11B: .sup.1H
NMR (DMSO-d.sub.6, 400 MHz) .delta. (ppm) 8.52 (s, 1H), 8.07 (s,
1H), 7.85 (d, J=7.59 Hz, 1H), 6.27-6.23 (m, 1H), 5.93 (d, J=7.59
Hz, 1H), 4.72-4.58 (m, 2H), 4.40 (t, J=10.05 Hz, 1H), 4.12-4.05 (m,
4H), 3.98-3.92 (m, 1H), 2.65-2.54 (m, 2H), 2.46-2.41 (m, 1H),
2.07-1.98 (m, 1H), 1.83-1.74 (m, 1H), 1.21-1.17 (m, 3H), 1.14 (d,
J=7.10 Hz, 3H); .sup.31P NMR (DMSO-d.sub.6, 162 MHz) .delta. (ppm)
-4.83 (s, 0.1P), -6.71 (s, 0.9P); MS (ESI) m/z=418.3
(MH.sup.+).
Example 12
In Vitro Inhibition
[0510] The compounds of the invention were tested in cell based in
vitro assays comprising HuH-1 cells and HepG2 cells, as in vitro
models for potential hepatocellular carcinoma activity.
HuH-1 cells were purchased from JCRB, HepG2 cells from ATCC, and
both tested with the following method:
Day 1:
[0511] Cells are suspended at a specific density in a specific
culture medium. Then, 100 .mu.l of cell suspension are plated per
well in 96-well plates.
Day 2:
[0512] Compounds or negative controls (DMSO) are added to the cells
(100 .mu.l of a compound solution in its specific medium with a
final concentration of 0.15% DMSO). Each compound concentration is
tested in duplicate (6 concentrations per compound). Cells are
incubated at 37.degree. C. for 72 h without treating again.
Day 4:
[0513] SDS is added to control wells (final concentration of 1%)
and kept for 15 min at 37.degree. C. SDS 1% represents the positive
control of proliferation inhibition. Cell supernatant is removed
and 100 .mu.l of a MTT
(3-[4.5-dimethylthiazol-2-yl]-2.5-diphenyltetrazolium bromide)
solution in fresh medium are added per well (MTT final
concentration of 0.5 mg/ml) and incubated for 4 h at 37.degree. C.
MTT reaction is stopped and homogenized by adding 100 .mu.l/well of
SDS 10%, 0.01M HCl and incubated for 2 h at 37.degree. C.
Absorbance is measured at 570 nm. The results for selected
compounds are shown in Table 1.
TABLE-US-00001 TABLE 1 In vitro inhibition of HuH-1 and HepG2:
HuH-1: HepG2: Compound Relative IC50 Relative IC50 1
Diastereoisomer 1 A ND 2 Diastereoisomer 1 A ND 2 Diastereoisomer 2
A B 3 Diastereoisomer 1 B A Mixture 5A/5B C C Mixture 6A/6B C C
[0514] The IC.sub.50 values in Table 1 are as follows:
A=<0.5 .mu.M
B=.gtoreq.0.5 .mu.M and <5 .mu.M
C=.gtoreq.5 .mu.M
Example 13
In Vitro Conversion of Prodrug to Nucleoside Triphosphate
[0515] The degree of conversion of a prodrug compound of the
present invention to its corresponding nucleoside triphosphate
(NTP) can be measured in vitro using the procedure described
below.
[0516] A 2 mM stock solution of the prodrug test compound is
prepared in 5% DMSO/95% MeOH to provide a final sample
concentration of 10M. A 5 .mu.L aliquot is removed from this stock
solution and added to 1 mL of either a rat or human cryopreserved
hepatocyte sample to provide a control sample at concentration of 1
million cells/mL. This sample is assayed in triplicate and used as
a test sample.
[0517] A 2 mM stock solution of
(2R,3R,4R,5R)-2-(4-amino-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-5-(hydroxymethy-
l)-3-methyltetrahydrofuran-3,4-diol is prepared in 5% DMSO/95% MeOH
to provide a final sample concentration of 10 .mu.M.
[0518] A 5 .mu.L aliquot is removed from this stock solution and
added to 1 mL of either a rat or human cryopreserved hepatocyte
sample to provide a control sample at a concentration of 1 million
cells/mL. This sample is assayed in triplicate and used as a
control standard.
[0519] Human and rat hepatocytes are removed from liquid nitrogen
storage and thawed by submerging the hepatocyte tube into a
pre-heated 37.degree. C. waterbath and gently shaking the tube back
& forth until thawed. The thawed hepatocytes are then gently
poured into a container of Hepatocyte Basal Medium (50 mL,
pre-warmed to 37.degree. C.) and washed. The hepatocyte tube is
then rinsed out with pre-warmed Hepatocyte Basal Medium and the
washed hepatocytes and rinse are combined and centrifuged at 500
rpm for 4 minutes at room temperature. The supernatant is then
discarded and the resulting hepatocyte pellet is resuspended with
Hepatocyte Basal Medium (pre-warmed to 37.degree. C.) and the final
hepatocyte concentration is adjusted to 1 million cells/mL to
provide the final hepatocyte suspension.
[0520] A 1 mL aliquot is removed from the 1 million cells/mL final
hepatocyte suspension, analyzed in triplicate and placed into 20 mL
scintillation vial without a cap. 2 mM of the prodrug test sample
is then added into the hepatocyte suspension to provide a 10 .mu.M
final concentration in the 1 mL hepatocyte sample. The sample is
then incubated at 37.degree. C./5% CO.sub.2 for 4 hours. A blank
hepatocyte sample as well as the control standard are also
incubated in this fashion.
[0521] The incubated hepatocyte suspension samples are transferred
to a microcentrifuge tube using a transfer pipette and centrifuged
at 500 rpm for 4 minutes at room temperature. The supernatant is
discarded and the resulting hepatocyte pellet was resuspended and
the cells are extracted with 0.25 mL of a 4.degree. C. solution of
70% methanol/30% (20 mM EDTA/20 mM EGTA) that has been adjusted to
pH 8 using sodium hydroxide. The resulting extract solution is then
stored in a refrigerator at 4.degree. C. until ready for use, at
which point the sample is first subjected to vortexing/sonication
to ensure that all hepatocyte cells have burst. The sample is then
centrifuged at 4000 rpm for 10 minutes at 4.degree. C. and a 100
.mu.L aliquot of the resulting supernatant is added into a
bioanalytical plate (2 mL Square 96 well plate w/100 uL Tapered
Reservoir), with the remaining supernatant immediately stored at
-80.degree. C. for re-assay if necessary. The blank control
supernatant is transferred to a new tube for use as a control
matrix in standard curves.
[0522] Alternatively, cryopreserved plateable hepatocytes are
obtained from Celsius-In Vitro Technologies (Baltimore, Md.) and
plated according to manufacturer's protocol at 0.7.times.10.sup.6
cells/mL in In Vitro GRO CP Medium (1.75.times.10.sup.6 cells/well
in 6-well plates) three hours prior to inhibitor treatment. An
inhibitor in DMSO at the indicated concentration in In Vitro GRO CP
Medium is added to the hepatocytes at t=0. At indicated times up to
48 hours post dosing, cells are washed in ice-cold PBS, extracted
with ice-cold 1 mL 70% methanol: 30% 20 mM EDTA/EGTA and
centrifuged. The supernatant is stored at -80.degree. C. until
analysis. For intracellular NTP analysis, an NTP calibration curve
is first generated by spiking a blank extraction buffer with known
concentrations of the NTP standard. LC/ESI-MS analysis is performed
on a QTRAP 5500 LC/MS/MS system (Applied Biosystems, Foster City,
Calif.) coupled to a Shimazu UFLC system, operated in the
positive-ion mode. The HPLC system is consisted of solvent delivery
module (LC20-AD XR), auto injector (SIL-20ACXR), and photodiode
array detector (SPD-M20A PDA) (Shimazu Corporation, Tokyo, Japan).
All HPLC separations are performed at 40.degree. C. The test
samples are analyzed on a BioBasic AX column (5 .mu.m particle
size, 100.times.2.1 mm I.D., Thermo Scientific) using A
(Acetonitrile: 10 mM NH.sub.4Ac=30:70, v:v, pH=6) and B
(Acetonitrile: 1 mM NH.sub.4Ac=30:70, v:v, pH=10) as mobile phases
at a flow rate of 1.0 mL/min. The injection volume is 50 .mu.L. The
mobile phase gradient starts at 0% B, and linearly increases to
100% B over 6 min. The MS analysis of all NTPs is performed on the
same QTRAP 5500 MS instrument in the multiple ion monitoring mode
(MRM), with Turbo-Ion-Spray ionization. The collision energy is 40
eV for all the analytes and standards. The quadrupole mass analyzer
is set to unit resolution.
The results for selected compounds are shown in Table 2:
TABLE-US-00002 TABLE 2 In vitro NTP in Human Hepatocytes: Human
Hepatocyte Compound 1 hour at 10 .mu.M, NTP 2 Diastereoisomer 2 A 1
Diastereoisomer 2 A 10 C
[0523] The NTP values in Table 2 are as follows:
A=<0.5 .mu.M
B=.gtoreq.0.5 .mu.M and <1 .mu.M
C=.gtoreq.1 .mu.M
* * * * *