ABSTRACT
A series of 2'-fluorinated C-nucleosides were prepared and tested for anti-HCV activity. Among them, the triphosphate of 2'-fluoro-2'-C-methyl adenosine C-nucleoside (15) was a potent and selective inhibitor of the NS5B polymerase and maintained activity against the S282T resistance mutant. A number of phosphoramidate prodrugs were then prepared and evaluated leading to the identification of the 1-aminocyclobutane-1-carboxylic acid isopropyl ester variant (53) with favorable pharmacokinetic properties including efficient liver delivery in animals.
Subject(s)
Antiviral Agents/chemistry , Antiviral Agents/pharmacology , Hepacivirus/drug effects , Nucleosides/chemistry , Nucleosides/pharmacology , Viral Nonstructural Proteins/antagonists & inhibitors , Amides/chemistry , Amides/pharmacokinetics , Amides/pharmacology , Animals , Antiviral Agents/pharmacokinetics , Caco-2 Cells , Cell Line , Cricetinae , Drug Discovery , Drug Resistance, Viral , Halogenation , Hepacivirus/genetics , Hepacivirus/physiology , Hepatitis C/drug therapy , Humans , Methylation , Molecular Docking Simulation , Nucleosides/pharmacokinetics , Phosphoric Acids/chemistry , Phosphoric Acids/pharmacokinetics , Phosphoric Acids/pharmacology , Point Mutation , Prodrugs/chemistry , Prodrugs/pharmacokinetics , Prodrugs/pharmacology , Viral Nonstructural Proteins/genetics , Viral Nonstructural Proteins/metabolism , Virus Replication/drug effectsABSTRACT
The recent Ebola virus (EBOV) outbreak in West Africa was the largest recorded in history with over 28,000 cases, resulting in >11,000 deaths including >500 healthcare workers. A focused screening and lead optimization effort identified 4b (GS-5734) with anti-EBOV EC50 = 86 nM in macrophages as the clinical candidate. Structure activity relationships established that the 1'-CN group and C-linked nucleobase were critical for optimal anti-EBOV potency and selectivity against host polymerases. A robust diastereoselective synthesis provided sufficient quantities of 4b to enable preclinical efficacy in a non-human-primate EBOV challenge model. Once-daily 10 mg/kg iv treatment on days 3-14 postinfection had a significant effect on viremia and mortality, resulting in 100% survival of infected treated animals [ Nature 2016 , 531 , 381 - 385 ]. A phase 2 study (PREVAIL IV) is currently enrolling and will evaluate the effect of 4b on viral shedding from sanctuary sites in EBOV survivors.
Subject(s)
Alanine/analogs & derivatives , Amides/chemistry , Hemorrhagic Fever, Ebola/drug therapy , Phosphoric Acids/chemistry , Prodrugs/chemistry , Prodrugs/pharmacology , Ribonucleotides/chemistry , Virus Diseases/drug therapy , Adenosine Monophosphate/analogs & derivatives , Alanine/chemistry , Cell Line , Drug Discovery , Humans , Microbial Sensitivity Tests , Prodrugs/chemical synthesis , Structure-Activity RelationshipABSTRACT
The most recent Ebola virus outbreak in West Africa, which was unprecedented in the number of cases and fatalities, geographic distribution, and number of nations affected, highlights the need for safe, effective, and readily available antiviral agents for treatment and prevention of acute Ebola virus (EBOV) disease (EVD) or sequelae. No antiviral therapeutics have yet received regulatory approval or demonstrated clinical efficacy. Here we report the discovery of a novel small molecule GS-5734, a monophosphoramidate prodrug of an adenosine analogue, with antiviral activity against EBOV. GS-5734 exhibits antiviral activity against multiple variants of EBOV and other filoviruses in cell-based assays. The pharmacologically active nucleoside triphosphate (NTP) is efficiently formed in multiple human cell types incubated with GS-5734 in vitro, and the NTP acts as an alternative substrate and RNA-chain terminator in primer-extension assays using a surrogate respiratory syncytial virus RNA polymerase. Intravenous administration of GS-5734 to nonhuman primates resulted in persistent NTP levels in peripheral blood mononuclear cells (half-life, 14 h) and distribution to sanctuary sites for viral replication including testes, eyes, and brain. In a rhesus monkey model of EVD, once-daily intravenous administration of 10 mg kg(-1) GS-5734 for 12 days resulted in profound suppression of EBOV replication and protected 100% of EBOV-infected animals against lethal disease, ameliorating clinical disease signs and pathophysiological markers, even when treatments were initiated three days after virus exposure when systemic viral RNA was detected in two out of six treated animals. These results show the first substantive post-exposure protection by a small-molecule antiviral compound against EBOV in nonhuman primates. The broad-spectrum antiviral activity of GS-5734 in vitro against other pathogenic RNA viruses, including filoviruses, arenaviruses, and coronaviruses, suggests the potential for wider medical use. GS-5734 is amenable to large-scale manufacturing, and clinical studies investigating the drug safety and pharmacokinetics are ongoing.
Subject(s)
Alanine/analogs & derivatives , Antiviral Agents/therapeutic use , Hemorrhagic Fever, Ebola/drug therapy , Macaca mulatta/virology , Ribonucleotides/therapeutic use , Adenosine Monophosphate/analogs & derivatives , Alanine/pharmacokinetics , Alanine/pharmacology , Alanine/therapeutic use , Amino Acid Sequence , Animals , Antiviral Agents/pharmacokinetics , Antiviral Agents/pharmacology , Cell Line, Tumor , Ebolavirus/drug effects , Female , HeLa Cells , Hemorrhagic Fever, Ebola/prevention & control , Humans , Male , Molecular Sequence Data , Organ Specificity , Prodrugs/pharmacokinetics , Prodrugs/pharmacology , Prodrugs/therapeutic use , Ribonucleotides/pharmacokinetics , Ribonucleotides/pharmacologyABSTRACT
Hepatitis C virus (HCV) infection presents an unmet medical need requiring more effective treatment options. Nucleoside inhibitors (NI) of HCV polymerase (NS5B) have demonstrated pan-genotypic activity and durable antiviral response in the clinic, and they are likely to become a key component of future treatment regimens. NI candidates that have entered clinical development thus far have all been N-nucleoside derivatives. Herein, we report the discovery of a C-nucleoside class of NS5B inhibitors. Exploration of adenosine analogs in this class identified 1'-cyano-2'-C-methyl 4-aza-7,9-dideaza adenosine as a potent and selective inhibitor of NS5B. A monophosphate prodrug approach afforded a series of compounds showing submicromolar activity in HCV replicon assays. Further pharmacokinetic optimization for sufficient oral absorption and liver triphosphate loading led to identification of a clinical development candidate GS-6620. In a phase I clinical study, the potential for potent activity was demonstrated but with high intra- and interpatient pharmacokinetic and pharmacodynamic variability.
Subject(s)
Antiviral Agents/therapeutic use , Enzyme Inhibitors/therapeutic use , Hepacivirus/enzymology , Hepatitis C/drug therapy , Nucleosides/pharmacology , Organophosphorus Compounds/pharmacology , Viral Nonstructural Proteins/antagonists & inhibitors , Animals , Antiviral Agents/pharmacokinetics , Antiviral Agents/pharmacology , Dogs , Enzyme Inhibitors/pharmacokinetics , Enzyme Inhibitors/pharmacology , Hepatitis C/enzymology , Hepatitis C/virology , Humans , Nucleosides/chemistry , Organophosphorus Compounds/chemistry , Rats , Viral LoadABSTRACT
Investigation of thiophene-2-carboxylic acid HCV NS5B site II inhibitors, guided by measurement of cell culture medium binding, revealed the structure-activity relationships for intrinsic cellular potency. The pharmacokinetic profile was enhanced through incorporation of heterocyclic ethers on the N-alkyl substituent. Hydroxyl groups were incorporated to modulate protein binding. Intrinsic potency was further improved through enantiospecific introduction of an olefin in the N-acyl motif, resulting in the discovery of the phase 2 clinical candidate GS-9669. The unexpected activity of this compound against the clinically relevant NS5B M423T mutant, relative to the wild type, was shown to arise from both the N-alkyl substituent and the N-acyl group.
Subject(s)
Antiviral Agents/pharmacology , Enzyme Inhibitors/pharmacology , Furans/pharmacology , Thiophenes/pharmacology , Viral Nonstructural Proteins/pharmacology , Antiviral Agents/chemistry , Enzyme Inhibitors/chemistry , Furans/chemistry , Magnetic Resonance Spectroscopy , Models, Molecular , Thiophenes/chemistry , Viral Nonstructural Proteins/chemistryABSTRACT
GS-9669 is a highly optimized thumb site II nonnucleoside inhibitor of the hepatitis C virus (HCV) RNA polymerase, with a binding affinity of 1.35 nM for the genotype (GT) 1b protein. It is a selective inhibitor of HCV RNA replication, with a mean 50% effective concentration (EC(50)) of ≤ 11 nM in genotype 1 and 5 replicon assays, but lacks useful activity against genotypes 2 to 4. The M423T mutation is readily generated clinically upon monotherapy with the thumb site II inhibitors filibuvir and lomibuvir, and it is notable that GS-9669 exhibited only a 3-fold loss in potency against this variant in the genotype 1b replicon. Rather than M423T, resistance predominantly tracks to residues R422K and L419M and residue I482L in GT 1b and 1a replicons, respectively. GS-9669 exhibited at least additive activity in combination with agents encompassing four other direct modes of action (NS3 protease, NS5A, NS5B via an alternative allosteric binding site, and NS5B nucleotide) as well as with alpha interferon or ribavirin in replicon assays. It exhibited high metabolic stability in in vitro human liver microsomal assays, which, in combination with its pharmacokinetic profiles in rat, dog, and two monkey species, is predictive of good human pharmacokinetics. GS-9669 is well suited for combination with other orally active, direct-acting antiviral agents in the treatment of genotype 1 chronic HCV infection. (This study has been registered at ClinicalTrials.gov under registration number NCT01431898.).
Subject(s)
Antiviral Agents/pharmacology , Furans/pharmacology , Hepacivirus/drug effects , RNA-Dependent RNA Polymerase/antagonists & inhibitors , Thiophenes/pharmacology , Viral Nonstructural Proteins/antagonists & inhibitors , Animals , Antiviral Agents/chemistry , Cell Line, Tumor , Dogs , Drug Resistance, Viral , Furans/chemistry , Humans , Interferon-alpha/pharmacology , Male , Mutation , Polymorphism, Single Nucleotide , Pyrones/pharmacology , Rats , Rats, Sprague-Dawley , Ribavirin/pharmacology , Thiophenes/chemistry , Triazoles/pharmacologyABSTRACT
This Letter describes the discovery and key structure-activity relationship (SAR) of a series of 2-aminobenzimidazoles as potent Aurora kinase inhibitors. 2-Aminobenzimidazole serves as a bioisostere of the biaryl urea residue of SNS-314 (1c), which is a potent Aurora kinase inhibitor and entered clinical testing in patients with solid tumors. Compared to SNS-314, this series of compounds offers better aqueous solubility while retaining comparable in vitro potency in biochemical and cell-based assays; in particular, 6m has also demonstrated a comparable mouse iv PK profile to SNS-314.
Subject(s)
Antineoplastic Agents/chemistry , Benzimidazoles/chemistry , Protein Kinase Inhibitors/chemistry , Protein Serine-Threonine Kinases/antagonists & inhibitors , Animals , Antineoplastic Agents/chemical synthesis , Antineoplastic Agents/pharmacokinetics , Aurora Kinases , Benzimidazoles/chemical synthesis , Benzimidazoles/pharmacokinetics , Cell Line, Tumor , Humans , Mice , Protein Kinase Inhibitors/chemical synthesis , Protein Kinase Inhibitors/pharmacokinetics , Protein Serine-Threonine Kinases/metabolism , Structure-Activity RelationshipABSTRACT
Compound 1 (SNS-314) is a potent and selective Aurora kinase inhibitor that is currently in clinical trials in patients with advanced solid tumors. This communication describes the synthesis of prodrug derivatives of 1 with improved aqueous solubility profiles. In particular, phosphonooxymethyl-derived prodrug 2g has significantly enhanced solubility and is converted to the biologically active parent (1) following iv as well as po administration to rodents.
Subject(s)
Phenylurea Compounds/chemistry , Prodrugs/chemistry , Protein Serine-Threonine Kinases/antagonists & inhibitors , Thiazoles/chemistry , Water/chemistry , Animals , Antineoplastic Agents/chemistry , Antineoplastic Agents/pharmacokinetics , Antineoplastic Agents/pharmacology , Aurora Kinases , Male , Mice , Phenylurea Compounds/pharmacokinetics , Phenylurea Compounds/pharmacology , Prodrugs/pharmacokinetics , Prodrugs/pharmacology , Protein Serine-Threonine Kinases/metabolism , Rats , Rats, Sprague-Dawley , Solubility , Thiazoles/pharmacokinetics , Thiazoles/pharmacologyABSTRACT
A series of 2-amino-pyrazolopyridines was designed and synthesized as Polo-like kinase (Plk) inhibitors based on a low micromolar hit. The SAR was developed to provide compounds exhibiting low nanomolar inhibitory activity of Plk1; the phenotype of treated cells is consistent with Plk1 inhibition. A co-crystal structure of one of these compounds with zPlk1 confirms an ATP-competitive binding mode.
Subject(s)
Cell Cycle Proteins/antagonists & inhibitors , Chemistry, Pharmaceutical/methods , Protein Serine-Threonine Kinases/antagonists & inhibitors , Proto-Oncogene Proteins/antagonists & inhibitors , Pyrazoles/chemical synthesis , Pyridines/chemical synthesis , Adenosine Triphosphate/chemistry , Amino Acid Motifs , Cell Cycle , Crystallography, X-Ray , Drug Design , Humans , Inhibitory Concentration 50 , Models, Chemical , Molecular Conformation , Phenotype , Pyrazoles/chemistry , Pyrazoles/pharmacology , Pyridines/chemistry , Pyridines/pharmacology , Structure-Activity Relationship , Polo-Like Kinase 1ABSTRACT
A series of 2-amino-isoxazolopyridines was designed and synthesized as Polo-like kinase (Plk) inhibitors. Key SAR and crystallographic data are discussed. More advanced analogues inhibit Plk1 with good enzymatic activity and modest cell-based activity. Differential selectivity among the three Plk isoforms is observed.
Subject(s)
Cell Cycle Proteins/antagonists & inhibitors , Drug Design , Isoxazoles/chemical synthesis , Isoxazoles/pharmacology , Protein Serine-Threonine Kinases/antagonists & inhibitors , Proto-Oncogene Proteins/antagonists & inhibitors , Pyridines/chemical synthesis , Pyridines/pharmacology , Combinatorial Chemistry Techniques , Crystallography, X-Ray , Drug Screening Assays, Antitumor , Humans , Isoenzymes/antagonists & inhibitors , Isoxazoles/chemistry , Microsomes, Liver/drug effects , Molecular Conformation , Molecular Structure , Pyridines/chemistry , Structure-Activity Relationship , Polo-Like Kinase 1ABSTRACT
This communication describes the discovery of a novel series of Aurora kinase inhibitors. Key SAR and critical binding elements are discussed. Some of the more advanced analogues potently inhibit cellular proliferation and induce phenotypes consistent with Aurora kinase inhibition. In particular, compound 21 (SNS-314) is a potent and selective Aurora kinase inhibitor that exhibits significant activity in pre-clinical in vivo tumor models.
Subject(s)
Neoplasms, Experimental/drug therapy , Phenylurea Compounds/chemistry , Phenylurea Compounds/pharmacology , Protein Kinase Inhibitors/chemistry , Protein Kinase Inhibitors/pharmacology , Protein Serine-Threonine Kinases/antagonists & inhibitors , Quinazolines/pharmacology , Thiazoles/chemistry , Thiazoles/pharmacology , Animals , Antineoplastic Agents/chemistry , Antineoplastic Agents/pharmacology , Aurora Kinases , Drug Screening Assays, Antitumor , HCT116 Cells , Humans , Mice , Neoplasm Transplantation , Neoplasms, Experimental/enzymology , Quinazolines/chemistry , Structure-Activity RelationshipABSTRACT
The design and synthesis of a series of novel, reversible, small molecule inhibitors of caspase-3 are described.
Subject(s)
Caspase Inhibitors , Enzyme Inhibitors/chemical synthesis , Enzyme Inhibitors/pharmacology , Aldehydes/chemistry , Caspase 3 , Indicators and Reagents , Ketones/chemistry , Kinetics , Magnetic Resonance Spectroscopy , Models, Molecular , Molecular Conformation , Stereoisomerism , Structure-Activity RelationshipABSTRACT
The design, synthesis, and in vitro activities of a series of potent and selective small-molecule inhibitors of caspase-3 are described. From extended tethering, a salicylic acid fragment was identified as having binding affinity for the S(4) pocket of caspase-3. X-ray crystallography and molecular modeling of the initial tethering hit resulted in the synthesis of 4, which reversibly inhibited caspase-3 with a K(i) = 40 nM. Further optimization led to the identification of a series of potent and selective inhibitors with K(i) values in the 20-50 nM range. One of the most potent compounds in this series, 66b, inhibited caspase-3 with a K(i) = 20 nM and selectivity of 8-500-fold for caspase-3 vs a panel of seven caspases (1, 2, and 4-8). A high-resolution X-ray cocrystal structure of 4 and 66b supports the predicted binding modes of our compounds with caspase-3.