Patent Review
For reprint orders, please contact [email protected]
Novel serine protease inhibitors Anilkumar R Kore* & Muthian Shanmugasundaram The pursuit of serine protease inhibitors as anticoagulants and anti-hepatitis C virus drugs continues to be an active area of research. Compounds such as P1–P3 macrocyclic peptides and linear peptides have been registered as potent hepatitis C virus protease inhibitors and compounds such as phenylglycinamide derivatives, substituted biaryls, tetrahydroquinoline derivatives, arylpropionamides, arylacrylamides, arylpropynamides, arylmethylurea analogs and peptides as factor XIa inhibitors. Given the recent US FDA approval of telaprevir and boceprevir for the treatment of hepatitis C virus, the development of new serine protease inhibitors is likely to be one of the hottest areas in the pharmaceutical industry. This review covers the patent literature on serine protease inhibitors during the period between 2009 and 2010.
Proteases, otherwise known as proteolytic enzymes, constitute an important class of enzymes that catalyze selectively the hydrolysis of peptide bonds in proteins [1–3] . Depending on their catalytic sites, proteases have been split into six major groups such as serine, aspartic, cysteine, gluatamic acid, threonine and metalloproteases. In particular, the target serine proteases display a diverse role in both health and disease. For example, they have been involved in a broad spectrum of physiological processes, such as embryogenesis, food digestion, blood clotting, defense mechanisms, immune responses and tissue reorganization. It is to be noted that many disease states such as Alzheimer’s disease, emphysema, arthritis, viral infections, stroke, cancer and inflammation are caused due to undesired, uncontrolled or unregulated proteolysis [1] . Therefore, it is essential to selectively regulate the proteases that continue to be a challenging clinical problem. A series of patent reviews reveal that the design and development of new serine protease inhibitors is demanded in view of their therapeutic intervention in a variety of disease states [4–8] . In particular, progress in the development of factor XIa inhibitors (until early 2009) has been recently reviewed in the patent literature [9] . The present review covers the patent literature on serine protease inhibitors during the period between 2009 and 2010. The scope of this review is to present the published patent literature reflecting the development of serine protease inhibitors on therapeutic and molecular biology applications and can logically be divided into three groups (Table 1) [101–124] . First, it involves the use of P1–P3 macrocyclic peptides and linear peptides as inhibitors for the treatment of hepatitis C virus (HCV). Second, compounds of the type phenylglycinamide derivatives, substituted biaryls, tetrahydroquinoline derivatives, arylpropionamides, arylacrylamides, arylpropynamides, arylmethylurea analogs and peptides as inhibitors for the treatment of thromboembolic diseases. Finally, a third group that contains peptidyl chloromethyl ketones useful in molecular biology applications is presented.
Life Technologies Corporation, Bioorganic Chemistry Division, 2130 Woodward Street, Austin, TX 78744-1832, USA *Author for correspondence: Tel.: +1 512 721 3589 Fax: +1 512 651 0201 E-mail: [email protected]
Hepatitis C serine protease inhibitors
It has been estimated that a chronic infection of HCV has affected over one hundred and 70 million people that accounts to 3% of the world’s population [10,201] . In particular, HCV has infected 1.8% of Americans (3.2 million people) and of those infected, the disease results in approximately 10,000 deaths every
10.4155/PPA.12.49 © 2012 Future Science Ltd
Pharm. Pat. Analyst (2012) 1(4), 457–468
ISSN 2046-8954
457
Patent Review
Kore & Shanmugasundaram
Table 1. Highlighted serine protease inhibitors. Lead compound
Applications
Company
Year
Ref.
Isoquinolines containing tripeptides
Hepatitis C virus
Bristol-Myers Squibb
2009
[101]
P1–P3 macrocyclic peptides
Hepatitis C virus
Enanta Pharmaceuticals 2009, 2010
Linear tripeptides
Hepatitis C virus
Enanta Pharmaceuticals 2009
[107]
Tripeptides containing acylsulfonamide P1´ moiety
Hepatitis C virus
Merck
2009
[108]
Tripeptides containing a-ketoamide P1´ moiety
Hepatitis C virus
Merck
2009
[109–112]
P1–P3 macrocyclic peptides
Hepatitis C virus
Merck
2009
Peptidomimetic compounds
Hepatitis C virus
Medivir AB
2009, 2010
[114,115]
Linear tetrapeptides containing a-ketoamide P1´ moiety
Hepatitis C virus
Vertex Pharmaceutical
2009, 2010
[116–118]
Peptidyl macrocycles
Thromboembolic disease
Bristol-Myers Squibb
2009
[119]
Phenylglycinamide derivatives
Thromboembolic disease
Bristol-Myers Squibb
2009
[120]
Substituted biaryls
Thromboembolic disease
Bristol-Myers Squibb
2009
[121]
Tetrahydroquinoline derivatives
Thromboembolic disease
Bristol-Myers Squibb
2009
[122]
Arylpropionamide, arylacrylamide, arylpropynamide and arylmethylurea
Thromboembolic disease
Bristol-Myers Squibb
2009
[123]
Peptidyl chloromethylketones
Molecular biology
Life Technologies
2010
[124]
year in the USA [201] . It is to be noted that the human problems including hepatocellular carcinoma, liver failure, and liver cirrhosis, are associated with HCV infection. HCV (Flaviviridae family) has a positive single-stranded RNA that contains a single open frame of approximately 9600 nucleotides [11,12] . The NS3 protease, a member of trypsin or chymotrypsin serine protease superfamily is one of the most intensively studied targets for anti-HCV therapy due to its crucial role in HCV replication. The NS3 protease is shown to be responsible for the proteolytic cleavage events at four junctions of precursor HCV polyprotein [12] . The N-terminal portion of the NS protein has the virally encoded protease that is responsible for processing the nonstructural portion of the polyprotein. Similar to the known digestive enzymes chymotrypsin and trypsin, the x ray data reveal that NS3 protease adopts two b-barrel conformations with a canonical Asp-HisSer catalytic triad at the active site [13,14] . Given the fact that NS3 protease displays a shallow and solventexposed substrate-binding groove, weak lipophilic and electrostatic interactions governs the inhibitor-binding energy for structure-based drug design [15] . There are two approved therapies for HCV infection. First, the use of long term intravenous administration of pegylated interferon-a [16] . Second, the improved second-generation treatment involves combination of IFN-a and oral ribavirin [17] . However,
458
[102–106]
[113]
these approved methods have considerable adverse side effects and are also effective in up to 50% of the patients. Therefore, the development of new novel antiviral agents for the treatment of HCV infection is demanded. Although Boehringer Ingelheim reported the first small-molecule hepatitis C serine protease inhibitor, BILN 2061, it failed to progress further in clinical development due to associated cardiac toxicity [18] . Subsequently, many hepatitis C serine protease inhibitors have entered into clinical trials. Small-molecule serine protease inhibitors, such as MK-7009 (Merck), R7227 (Intermune and Roche Pharmaceuticals), VX985 (Vertex Pharmaceuticals) and ABT-333 (Abbott Pharmaceuticals), are currently in Phase II clinical trials, whereas inhibitors such as BI 201335 (Boehringer Ingelheim Pharmaceuticals) [19] and TMC 435 (Medivir AB) [20] have now progressed to Phase III of clinical development [202] . It is noteworthy that recently US FDA-approved Boceprevir (trade name Victrelis™), a small-molecule protease inhibitor from Merck as the first generation of HCV drug on 13 May 2011 (Figure 1) [203] . Subsequently, the FDA-approved Incivek® (telaprevir) from Vertex Pharmaceuticals as the second direct acting antiviral drug against the HCV on 23 May 2011 [204] . These two new drugs have to be used in combination with PEG-IFN-a and ribavirin to treat patients with chronic HCV infection they promise to increase the
www.future-science.com
future science group
Patent Review
Novel serine protease inhibitors
cure rate while shortening the time required for treatment. The total treatment duration for Victrelis™ is 28 weeks for the patients without cirrhosis who have been previously untreated instead of the usual 48 weeks for the standard treatment [21,22] . The total duration of treatment with Incivek is 24 weeks for the treatment-naive and prior relapse patients [23,24] . The main pharmaceutical companies engaged in the area of HCV inhibitors during 2009 and 2010 were Bristol-Myers Squibb, Enanta Pharmaceuticals, Merck, Medivir AB and Vertex Pharmaceuticals.
O H N H N
N
H N
NH2
N
O O
N
N
N H
O
O
O
O
O Merck – boceprevir (FD A) Ki* = 14 nM EC90 = 350 nM Rat bioavailability 26% Monkey bioavailability 4–11% Dog bioavailability 30%
S Br
O
NH O
Ki* = 7 nM EC50 = 350 nM
H N
S N
N H
Vertex – telaprevir (FD A)
N
■■ Bristol-Myers Squibb Company
O
H N
O
O
N N
MeO
O Bristol-Myers Squibb reported a MeO O series of noncovalent inhibitors, O O O N namely linear tripeptides containO N NH O OH S H ing isoquinoline derivative 1 and 2 N O O N N H O that are highly specific for the NS3 H O protease and many of these memMedivir AB –TMC 435 (Phase III) Boehringer Ingelheim – BI 201335 (Phase III) bers inhibit HCV replicon replication [101] . Representative compounds were found to have an IC50 Figure 1. Hepatitis C serine protease inhibitors approved by the US FDA and in Phase III value in the range of 0.02–0.2 µM clinical trials. against the NS3/4A Bristol-Myers Squibb strain in the enzyme assay and the EC50 value clinical trials. In addition to the macrocyclic peptides, in the range of 0.1–1 µM using replicon Luciferase they reported a series of tripeptides 11 & 12, displayassay (Figure 2) . ing viral response by binding to HCV NS3/NS4A protease [107] . ■■ Enanta Pharmaceuticals An internally quenched fluorogenic substrate was Enanta Pharmaceuticals reported a series of P1–P3 used in a HCV cell-based assay (Huh 11-7 cell line) macrocyclic noncovalent NS3/NS4A protease inhibi- to determine HCV protease inhibitory activity of the tors such as quinoxalinyl macrocycles 3 & 4, phospho- compounds via quantifying HCV replicon RNA. In rous-containing macrocycle 5, tetrazolyl macrocycle 6, order to extract and purify cellular RNA, Ambion® triazolyl marcrocycle 7, and pyridazinonyl macrocycles 8–10 (Figure 3) O O [102–106] . It is claimed that the comCl Cl pounds of this invention interfere N with the life cycle of the HCV and O N S O O O O are also useful as antiviral agents. S O O NH H O N In this class of macrocyclic pepNH H N O N tides, a variety of modifications has H O O N N been achieved by the introduction H N O O N N of heterocyclic moiety attached to O H 1 2 the P2 proline group. It is interestIC50 = >0.2 µM IC50 = 0.02-0.2 µM ing that the structural framework EC50 = 0.1-1 µM EC50 = 0.1-1 µM for these compounds is common to the structure of TMS 435 (Medivir Figure 2. Hepatitis C serine protease inhibitors from Bristol-Myers Squibb. AB) that is currently in Phase III
future science group
Pharm. Pat. Analyst (2012) 1(4)
459
Patent Review
Kore & Shanmugasundaram
S S
N O
O
N
O S
N HN
O
O
N
NH
O H N
O O O
N
S
N
O
N
O
HN
O
N HN
Ph
O
O 3
H O N P Ph
O H N
O O
O
O H N
O O
N
4
5 O
O
N O
N N
N
N
N O
O O
O
H N
N
HN O
6
7
MeO
MeO N
O
N
O
MeO N
O
N
O
O H N
N
O
NH
O H N
O O
NH
N HN
IC50 EC50
N O H N
N S
9 = 0.4 nM =
For reprint orders, please contact [email protected]
Novel serine protease inhibitors Anilkumar R Kore* & Muthian Shanmugasundaram The pursuit of serine protease inhibitors as anticoagulants and anti-hepatitis C virus drugs continues to be an active area of research. Compounds such as P1–P3 macrocyclic peptides and linear peptides have been registered as potent hepatitis C virus protease inhibitors and compounds such as phenylglycinamide derivatives, substituted biaryls, tetrahydroquinoline derivatives, arylpropionamides, arylacrylamides, arylpropynamides, arylmethylurea analogs and peptides as factor XIa inhibitors. Given the recent US FDA approval of telaprevir and boceprevir for the treatment of hepatitis C virus, the development of new serine protease inhibitors is likely to be one of the hottest areas in the pharmaceutical industry. This review covers the patent literature on serine protease inhibitors during the period between 2009 and 2010.
Proteases, otherwise known as proteolytic enzymes, constitute an important class of enzymes that catalyze selectively the hydrolysis of peptide bonds in proteins [1–3] . Depending on their catalytic sites, proteases have been split into six major groups such as serine, aspartic, cysteine, gluatamic acid, threonine and metalloproteases. In particular, the target serine proteases display a diverse role in both health and disease. For example, they have been involved in a broad spectrum of physiological processes, such as embryogenesis, food digestion, blood clotting, defense mechanisms, immune responses and tissue reorganization. It is to be noted that many disease states such as Alzheimer’s disease, emphysema, arthritis, viral infections, stroke, cancer and inflammation are caused due to undesired, uncontrolled or unregulated proteolysis [1] . Therefore, it is essential to selectively regulate the proteases that continue to be a challenging clinical problem. A series of patent reviews reveal that the design and development of new serine protease inhibitors is demanded in view of their therapeutic intervention in a variety of disease states [4–8] . In particular, progress in the development of factor XIa inhibitors (until early 2009) has been recently reviewed in the patent literature [9] . The present review covers the patent literature on serine protease inhibitors during the period between 2009 and 2010. The scope of this review is to present the published patent literature reflecting the development of serine protease inhibitors on therapeutic and molecular biology applications and can logically be divided into three groups (Table 1) [101–124] . First, it involves the use of P1–P3 macrocyclic peptides and linear peptides as inhibitors for the treatment of hepatitis C virus (HCV). Second, compounds of the type phenylglycinamide derivatives, substituted biaryls, tetrahydroquinoline derivatives, arylpropionamides, arylacrylamides, arylpropynamides, arylmethylurea analogs and peptides as inhibitors for the treatment of thromboembolic diseases. Finally, a third group that contains peptidyl chloromethyl ketones useful in molecular biology applications is presented.
Life Technologies Corporation, Bioorganic Chemistry Division, 2130 Woodward Street, Austin, TX 78744-1832, USA *Author for correspondence: Tel.: +1 512 721 3589 Fax: +1 512 651 0201 E-mail: [email protected]
Hepatitis C serine protease inhibitors
It has been estimated that a chronic infection of HCV has affected over one hundred and 70 million people that accounts to 3% of the world’s population [10,201] . In particular, HCV has infected 1.8% of Americans (3.2 million people) and of those infected, the disease results in approximately 10,000 deaths every
10.4155/PPA.12.49 © 2012 Future Science Ltd
Pharm. Pat. Analyst (2012) 1(4), 457–468
ISSN 2046-8954
457
Patent Review
Kore & Shanmugasundaram
Table 1. Highlighted serine protease inhibitors. Lead compound
Applications
Company
Year
Ref.
Isoquinolines containing tripeptides
Hepatitis C virus
Bristol-Myers Squibb
2009
[101]
P1–P3 macrocyclic peptides
Hepatitis C virus
Enanta Pharmaceuticals 2009, 2010
Linear tripeptides
Hepatitis C virus
Enanta Pharmaceuticals 2009
[107]
Tripeptides containing acylsulfonamide P1´ moiety
Hepatitis C virus
Merck
2009
[108]
Tripeptides containing a-ketoamide P1´ moiety
Hepatitis C virus
Merck
2009
[109–112]
P1–P3 macrocyclic peptides
Hepatitis C virus
Merck
2009
Peptidomimetic compounds
Hepatitis C virus
Medivir AB
2009, 2010
[114,115]
Linear tetrapeptides containing a-ketoamide P1´ moiety
Hepatitis C virus
Vertex Pharmaceutical
2009, 2010
[116–118]
Peptidyl macrocycles
Thromboembolic disease
Bristol-Myers Squibb
2009
[119]
Phenylglycinamide derivatives
Thromboembolic disease
Bristol-Myers Squibb
2009
[120]
Substituted biaryls
Thromboembolic disease
Bristol-Myers Squibb
2009
[121]
Tetrahydroquinoline derivatives
Thromboembolic disease
Bristol-Myers Squibb
2009
[122]
Arylpropionamide, arylacrylamide, arylpropynamide and arylmethylurea
Thromboembolic disease
Bristol-Myers Squibb
2009
[123]
Peptidyl chloromethylketones
Molecular biology
Life Technologies
2010
[124]
year in the USA [201] . It is to be noted that the human problems including hepatocellular carcinoma, liver failure, and liver cirrhosis, are associated with HCV infection. HCV (Flaviviridae family) has a positive single-stranded RNA that contains a single open frame of approximately 9600 nucleotides [11,12] . The NS3 protease, a member of trypsin or chymotrypsin serine protease superfamily is one of the most intensively studied targets for anti-HCV therapy due to its crucial role in HCV replication. The NS3 protease is shown to be responsible for the proteolytic cleavage events at four junctions of precursor HCV polyprotein [12] . The N-terminal portion of the NS protein has the virally encoded protease that is responsible for processing the nonstructural portion of the polyprotein. Similar to the known digestive enzymes chymotrypsin and trypsin, the x ray data reveal that NS3 protease adopts two b-barrel conformations with a canonical Asp-HisSer catalytic triad at the active site [13,14] . Given the fact that NS3 protease displays a shallow and solventexposed substrate-binding groove, weak lipophilic and electrostatic interactions governs the inhibitor-binding energy for structure-based drug design [15] . There are two approved therapies for HCV infection. First, the use of long term intravenous administration of pegylated interferon-a [16] . Second, the improved second-generation treatment involves combination of IFN-a and oral ribavirin [17] . However,
458
[102–106]
[113]
these approved methods have considerable adverse side effects and are also effective in up to 50% of the patients. Therefore, the development of new novel antiviral agents for the treatment of HCV infection is demanded. Although Boehringer Ingelheim reported the first small-molecule hepatitis C serine protease inhibitor, BILN 2061, it failed to progress further in clinical development due to associated cardiac toxicity [18] . Subsequently, many hepatitis C serine protease inhibitors have entered into clinical trials. Small-molecule serine protease inhibitors, such as MK-7009 (Merck), R7227 (Intermune and Roche Pharmaceuticals), VX985 (Vertex Pharmaceuticals) and ABT-333 (Abbott Pharmaceuticals), are currently in Phase II clinical trials, whereas inhibitors such as BI 201335 (Boehringer Ingelheim Pharmaceuticals) [19] and TMC 435 (Medivir AB) [20] have now progressed to Phase III of clinical development [202] . It is noteworthy that recently US FDA-approved Boceprevir (trade name Victrelis™), a small-molecule protease inhibitor from Merck as the first generation of HCV drug on 13 May 2011 (Figure 1) [203] . Subsequently, the FDA-approved Incivek® (telaprevir) from Vertex Pharmaceuticals as the second direct acting antiviral drug against the HCV on 23 May 2011 [204] . These two new drugs have to be used in combination with PEG-IFN-a and ribavirin to treat patients with chronic HCV infection they promise to increase the
www.future-science.com
future science group
Patent Review
Novel serine protease inhibitors
cure rate while shortening the time required for treatment. The total treatment duration for Victrelis™ is 28 weeks for the patients without cirrhosis who have been previously untreated instead of the usual 48 weeks for the standard treatment [21,22] . The total duration of treatment with Incivek is 24 weeks for the treatment-naive and prior relapse patients [23,24] . The main pharmaceutical companies engaged in the area of HCV inhibitors during 2009 and 2010 were Bristol-Myers Squibb, Enanta Pharmaceuticals, Merck, Medivir AB and Vertex Pharmaceuticals.
O H N H N
N
H N
NH2
N
O O
N
N
N H
O
O
O
O
O Merck – boceprevir (FD A) Ki* = 14 nM EC90 = 350 nM Rat bioavailability 26% Monkey bioavailability 4–11% Dog bioavailability 30%
S Br
O
NH O
Ki* = 7 nM EC50 = 350 nM
H N
S N
N H
Vertex – telaprevir (FD A)
N
■■ Bristol-Myers Squibb Company
O
H N
O
O
N N
MeO
O Bristol-Myers Squibb reported a MeO O series of noncovalent inhibitors, O O O N namely linear tripeptides containO N NH O OH S H ing isoquinoline derivative 1 and 2 N O O N N H O that are highly specific for the NS3 H O protease and many of these memMedivir AB –TMC 435 (Phase III) Boehringer Ingelheim – BI 201335 (Phase III) bers inhibit HCV replicon replication [101] . Representative compounds were found to have an IC50 Figure 1. Hepatitis C serine protease inhibitors approved by the US FDA and in Phase III value in the range of 0.02–0.2 µM clinical trials. against the NS3/4A Bristol-Myers Squibb strain in the enzyme assay and the EC50 value clinical trials. In addition to the macrocyclic peptides, in the range of 0.1–1 µM using replicon Luciferase they reported a series of tripeptides 11 & 12, displayassay (Figure 2) . ing viral response by binding to HCV NS3/NS4A protease [107] . ■■ Enanta Pharmaceuticals An internally quenched fluorogenic substrate was Enanta Pharmaceuticals reported a series of P1–P3 used in a HCV cell-based assay (Huh 11-7 cell line) macrocyclic noncovalent NS3/NS4A protease inhibi- to determine HCV protease inhibitory activity of the tors such as quinoxalinyl macrocycles 3 & 4, phospho- compounds via quantifying HCV replicon RNA. In rous-containing macrocycle 5, tetrazolyl macrocycle 6, order to extract and purify cellular RNA, Ambion® triazolyl marcrocycle 7, and pyridazinonyl macrocycles 8–10 (Figure 3) O O [102–106] . It is claimed that the comCl Cl pounds of this invention interfere N with the life cycle of the HCV and O N S O O O O are also useful as antiviral agents. S O O NH H O N In this class of macrocyclic pepNH H N O N tides, a variety of modifications has H O O N N been achieved by the introduction H N O O N N of heterocyclic moiety attached to O H 1 2 the P2 proline group. It is interestIC50 = >0.2 µM IC50 = 0.02-0.2 µM ing that the structural framework EC50 = 0.1-1 µM EC50 = 0.1-1 µM for these compounds is common to the structure of TMS 435 (Medivir Figure 2. Hepatitis C serine protease inhibitors from Bristol-Myers Squibb. AB) that is currently in Phase III
future science group
Pharm. Pat. Analyst (2012) 1(4)
459
Patent Review
Kore & Shanmugasundaram
S S
N O
O
N
O S
N HN
O
O
N
NH
O H N
O O O
N
S
N
O
N
O
HN
O
N HN
Ph
O
O 3
H O N P Ph
O H N
O O
O
O H N
O O
N
4
5 O
O
N O
N N
N
N
N O
O O
O
H N
N
HN O
6
7
MeO
MeO N
O
N
O
MeO N
O
N
O
O H N
N
O
NH
O H N
O O
NH
N HN
IC50 EC50
N O H N
N S
9 = 0.4 nM =
