ABSTRACT
Structure-based design was utilized to optimize 6,6-diaryl substituted dihydropyrone and hydroxylactam to obtain inhibitors of lactate dehydrogenase (LDH) with low nanomolar biochemical and single-digit micromolar cellular potencies. Surprisingly the replacement of a phenyl with a pyridyl moiety in the chemical structure revealed a new binding mode for the inhibitors with subtle conformational change of the LDHA active site. This led to the identification of a potent, cell-active hydroxylactam inhibitor exhibiting an in vivo pharmacokinetic profile suitable for mouse tumor xenograft study.
Subject(s)
Enzyme Inhibitors/pharmacology , L-Lactate Dehydrogenase/antagonists & inhibitors , Lactams/pharmacology , Animals , Cell Line , Dogs , Dose-Response Relationship, Drug , Enzyme Inhibitors/chemistry , Humans , L-Lactate Dehydrogenase/metabolism , Lactams/chemistry , Mice , Microsomes, Liver/chemistry , Microsomes, Liver/metabolism , Molecular Structure , Structure-Activity RelationshipABSTRACT
Estrogen receptor alpha (ERα) is a well-validated drug target for ER-positive (ER+) breast cancer. Fulvestrant is FDA-approved to treat ER+ breast cancer and works through two mechanisms-as a full antagonist and selective estrogen receptor degrader (SERD)-but lacks oral bioavailability. Thus, we envisioned a "best-in-class" molecule with the same dual mechanisms as fulvestrant, but with significant oral exposure. Through lead optimization, we discovered a tool molecule 12 (GNE-149) with improved degradation and antiproliferative activity in both MCF7 and T47D cells. To illustrate the binding mode and key interactions of this scaffold with ERα, we obtained a cocrystal structure of 6 that showed ionic interaction of azetidine with Asp351 residue. Importantly, 12 showed favorable metabolic stability and good oral exposure. 12 exhibited antagonist effect in the uterus and demonstrated robust dose-dependent efficacy in xenograft models.
ABSTRACT
CDK4 and CDK6 are kinases with similar sequences that regulate cell cycle progression and are validated targets in the treatment of cancer. Glioblastoma is characterized by a high frequency of CDKN2A/CCND2/CDK4/CDK6 pathway dysregulation, making dual inhibition of CDK4 and CDK6 an attractive therapeutic approach for this disease. Abemaciclib, ribociclib, and palbociclib are approved CDK4/6 inhibitors for the treatment of HR+/HER2- breast cancer, but these drugs are not expected to show strong activity in brain tumors due to poor blood brain barrier penetration. Herein, we report the identification of a brain-penetrant CDK4/6 inhibitor derived from a literature molecule with low molecular weight and topological polar surface area (MWâ¯=â¯285 and TPSAâ¯=â¯66â¯Å2), but lacking the CDK2/1 selectivity profile due to the absence of a basic amine. Removal of a hydrogen bond donor via cyclization of the pyrazole allowed for the introduction of basic and semi-basic amines, while maintaining in many cases efflux ratios reasonable for a CNS program. Ultimately, a basic spiroazetidine (cpKaâ¯=â¯8.8) was identified that afforded acceptable selectivity over anti-target CDK1 while maintaining brain-penetration in vivo (mouse Kp,uuâ¯=â¯0.20-0.59). To probe the potency and selectivity, our lead compound was evaluated in a panel of glioblastoma cell lines. Potency comparable to abemaciclib was observed in Rb-wild type lines U87MG, DBTRG-05MG, A172, and T98G, while Rb-deficient cell lines SF539 and M059J exhibited a lack of sensitivity.
Subject(s)
Antineoplastic Agents/pharmacology , Brain Neoplasms/drug therapy , Cyclin-Dependent Kinase 4/antagonists & inhibitors , Cyclin-Dependent Kinase 6/antagonists & inhibitors , Drug Design , Glioblastoma/drug therapy , Protein Kinase Inhibitors/pharmacology , Antineoplastic Agents/chemical synthesis , Antineoplastic Agents/chemistry , Brain Neoplasms/metabolism , Brain Neoplasms/pathology , Cell Line, Tumor , Cell Proliferation/drug effects , Cyclin-Dependent Kinase 4/metabolism , Cyclin-Dependent Kinase 6/metabolism , Dose-Response Relationship, Drug , Drug Screening Assays, Antitumor , Glioblastoma/metabolism , Glioblastoma/pathology , Humans , MCF-7 Cells , Molecular Structure , Protein Kinase Inhibitors/chemical synthesis , Protein Kinase Inhibitors/chemistry , Structure-Activity RelationshipABSTRACT
Using structure- and ligand-based design principles, a novel series of piperidyl chromane arylsulfonamide Nav1.7 inhibitors was discovered. Early optimization focused on improvement of potency through refinement of the low energy ligand conformation and mitigation of high in vivo clearance. An in vitro hepatotoxicity hazard was identified and resolved through optimization of lipophilicity and lipophilic ligand efficiency to arrive at GNE-616 (24), a highly potent, metabolically stable, subtype selective inhibitor of Nav1.7. Compound 24 showed a robust PK/PD response in a Nav1.7-dependent mouse model, and site-directed mutagenesis was used to identify residues critical for the isoform selectivity profile of 24.
Subject(s)
NAV1.7 Voltage-Gated Sodium Channel/chemistry , Sulfonamides/chemistry , Voltage-Gated Sodium Channel Blockers/chemistry , Analgesics/chemistry , Analgesics/metabolism , Analgesics/pharmacology , Analgesics/therapeutic use , Animals , Binding Sites , Cell Line , Cell Survival/drug effects , Chronic Pain/drug therapy , Chronic Pain/pathology , Dogs , Half-Life , Humans , Ligands , Male , Mice , Molecular Docking Simulation , Mutagenesis, Site-Directed , NAV1.7 Voltage-Gated Sodium Channel/genetics , NAV1.7 Voltage-Gated Sodium Channel/metabolism , Protein Isoforms/antagonists & inhibitors , Protein Isoforms/metabolism , Rats , Structure-Activity Relationship , Sulfonamides/metabolism , Sulfonamides/pharmacology , Sulfonamides/therapeutic use , Voltage-Gated Sodium Channel Blockers/metabolism , Voltage-Gated Sodium Channel Blockers/pharmacology , Voltage-Gated Sodium Channel Blockers/therapeutic useABSTRACT
A novel, potent, and orally bioavailable inhibitor of the bromodomain of CBP, compound 35 (GNE-207), has been identified through SAR investigations focused on optimizing al bicyclic heteroarene to replace the aniline present in the published GNE-272 series. Compound 35â¯has excellent CBP potency (CBP IC50â¯=â¯1â¯nM, MYC EC50â¯=â¯18â¯nM), a selectively index ofâ¯>2500-fold against BRD4(1), and exhibits a good pharmacokinetic profile.
Subject(s)
Biphenyl Compounds/chemistry , Drug Design , p300-CBP Transcription Factors/antagonists & inhibitors , Animals , Binding Sites , Biphenyl Compounds/chemical synthesis , Biphenyl Compounds/metabolism , Cell Cycle Proteins , Crystallography, X-Ray , Half-Life , Humans , Hydrogen Bonding , Inhibitory Concentration 50 , Mice , Microsomes, Liver/metabolism , Molecular Dynamics Simulation , Nuclear Proteins/antagonists & inhibitors , Nuclear Proteins/metabolism , Protein Structure, Tertiary , Rats , Structure-Activity Relationship , Transcription Factors/antagonists & inhibitors , Transcription Factors/metabolism , p300-CBP Transcription Factors/metabolismABSTRACT
The epigenetic regulator CBP/P300 presents a novel therapeutic target for oncology. Previously, we disclosed the development of potent and selective CBP bromodomain inhibitors by first identifying pharmacophores that bind the KAc region and then building into the LPF shelf. Herein, we report the "hybridization" of a variety of KAc-binding fragments with a tetrahydroquinoline scaffold that makes optimal interactions with the LPF shelf, imparting enhanced potency and selectivity to the hybridized ligand. To demonstrate the utility of our hybridization approach, two analogues containing unique Asn binders and the optimized tetrahydroquinoline moiety were rapidly optimized to yield single-digit nanomolar inhibitors of CBP with exquisite selectivity over BRD4(1) and the broader bromodomain family.
Subject(s)
High-Throughput Screening Assays/methods , p300-CBP Transcription Factors/antagonists & inhibitors , Animals , Asparagine/chemistry , Asparagine/metabolism , Binding Sites , Cell Cycle Proteins , Crystallography, X-Ray , Female , Fluorescence Resonance Energy Transfer/methods , Mice, Inbred Strains , Molecular Docking Simulation , Nuclear Proteins/antagonists & inhibitors , Protein Domains , Pyrazoles/chemistry , Pyridines/chemistry , Quinolines/chemistry , Transcription Factors/antagonists & inhibitors , p300-CBP Transcription Factors/chemistry , p300-CBP Transcription Factors/metabolismABSTRACT
Inhibition of the bromodomain of the transcriptional regulator CBP/P300 is an especially interesting new therapeutic approach in oncology. We recently disclosed in vivo chemical tool 1 (GNE-272) for the bromodomain of CBP that was moderately potent and selective over BRD4(1). In pursuit of a more potent and selective CBP inhibitor, we used structure-based design. Constraining the aniline of 1 into a tetrahydroquinoline motif maintained potency and increased selectivity 2-fold. Structure-activity relationship studies coupled with further structure-based design targeting the LPF shelf, BC loop, and KAc regions allowed us to significantly increase potency and selectivity, resulting in the identification of non-CNS penetrant 19 (GNE-781, TR-FRET IC50 = 0.94 nM, BRET IC50 = 6.2 nM; BRD4(1) IC50 = 5100 nΜ) that maintained good in vivo PK properties in multiple species. Compound 19 displays antitumor activity in an AML tumor model and was also shown to decrease Foxp3 transcript levels in a dose dependent manner.
Subject(s)
Antineoplastic Agents/pharmacology , CREB-Binding Protein/antagonists & inhibitors , Pyrazoles/pharmacology , Pyridines/pharmacology , Animals , Antineoplastic Agents/chemical synthesis , Antineoplastic Agents/chemistry , Antineoplastic Agents/pharmacokinetics , CREB-Binding Protein/chemistry , Dogs , Female , Forkhead Transcription Factors/genetics , Forkhead Transcription Factors/metabolism , HEK293 Cells , Humans , Macaca fascicularis , Male , Mice , Protein Domains , Proto-Oncogene Proteins c-myc/genetics , Proto-Oncogene Proteins c-myc/metabolism , Pyrazoles/chemical synthesis , Pyrazoles/chemistry , Pyrazoles/pharmacokinetics , Pyridines/chemical synthesis , Pyridines/chemistry , Pyridines/pharmacokinetics , RNA/genetics , Rats, Sprague-Dawley , Structure-Activity Relationship , Xenograft Model Antitumor AssaysABSTRACT
A series of trisubstituted hydroxylactams was identified as potent enzymatic and cellular inhibitors of human lactate dehydrogenase A. Utilizing structure-based design and physical property optimization, multiple inhibitors were discovered with <10 µM lactate IC50 in a MiaPaca2 cell line. Optimization of the series led to 29, a potent cell active molecule (MiaPaca2 IC50 = 0.67 µM) that also possessed good exposure when dosed orally to mice.
ABSTRACT
The single bromodomain of the closely related transcriptional regulators CBP/EP300 is a target of much recent interest in cancer and immune system regulation. A co-crystal structure of a ligand-efficient screening hit and the CBP bromodomain guided initial design targeting the LPF shelf, ZA loop, and acetylated lysine binding regions. Structure-activity relationship studies allowed us to identify a more potent analogue. Optimization of permeability and microsomal stability and subsequent improvement of mouse hepatocyte stability afforded 59 (GNE-272, TR-FRET IC50 = 0.02 µM, BRET IC50 = 0.41 µM, BRD4(1) IC50 = 13 µM) that retained the best balance of cell potency, selectivity, and in vivo PK. Compound 59 showed a marked antiproliferative effect in hematologic cancer cell lines and modulates MYC expression in vivo that corresponds with antitumor activity in an AML tumor model.
Subject(s)
Antineoplastic Agents/pharmacology , Drug Discovery , Pyrazoles/pharmacology , Pyridones/pharmacology , p300-CBP Transcription Factors/antagonists & inhibitors , Animals , Antineoplastic Agents/chemical synthesis , Antineoplastic Agents/chemistry , Cell Line, Tumor , Cell Proliferation/drug effects , Dogs , Dose-Response Relationship, Drug , Drug Screening Assays, Antitumor , Female , Humans , Madin Darby Canine Kidney Cells , Mice , Mice, Nude , Models, Molecular , Molecular Structure , Pyrazoles/chemical synthesis , Pyrazoles/chemistry , Pyridones/chemical synthesis , Pyridones/chemistry , Structure-Activity RelationshipABSTRACT
Optimization of 5-(2,6-dichlorophenyl)-3-hydroxy-2-mercaptocyclohex-2-enone using structure-based design strategies resulted in inhibitors with considerable improvement in biochemical potency against human lactate dehydrogenase A (LDHA). These potent inhibitors were typically selective for LDHA over LDHB isoform (410 fold) and other structurally related malate dehydrogenases, MDH1 and MDH2 (>500 fold). An X-ray crystal structure of enzymatically most potent molecule bound to LDHA revealed two additional interactions associated with enhanced biochemical potency.
Subject(s)
Enzyme Inhibitors/chemical synthesis , L-Lactate Dehydrogenase/antagonists & inhibitors , Animals , Crystallography, X-Ray , Dogs , Enzyme Inhibitors/metabolism , Enzyme Inhibitors/pharmacology , Humans , L-Lactate Dehydrogenase/metabolism , Madin Darby Canine Kidney CellsABSTRACT
An efficient entry to the ABC network of lancifodilactone G is outlined. The C ring is constructed by way of enyne ring-closing metathesis. The AB component is established via a base-mediated biomimetic oxy-Michael addition--lactonization sequence.
Subject(s)
Triterpenes/chemical synthesis , Hydroxyl Radical/chemistry , Plants, Medicinal/chemistry , Schisandra/chemistry , Triterpenes/chemistryABSTRACT
A stereochemically linear strategy has been developed to prepare the heavily congested F-ring sector of lancifodilactone G (1) from commercially inexpensive (R)-carvone. Prominent operations in our synthesis include Negishi-type sp2-sp3 cross-coupling and intramolecular free-radical cyclization for the purpose of appending the sidearm links of the D and H rings onto the F platform.
ABSTRACT
The regioselective and enantiospecific rhodium-catalyzed allylic amination of secondary allylic carbonates with N3-benzoyl thymine in conjunction with a stereoselective free radical cyclization provides a convenient method for the construction of a new conformationally rigid nucleoside.
Subject(s)
Nucleosides/chemical synthesis , Rhodium/chemistry , Thymine/chemistry , Amination , Catalysis , Nucleic Acid Conformation , StereoisomerismABSTRACT
Transition metal-catalyzed [m+n+o] carbocyclization reactions provide powerful methods for the construction of complex polycyclic systems that are generally not accessible through classical pericyclic reactions. We have developed the first regio- and enantioselective crossed intermolecular rhodium-catalyzed [2+2+2] carbocyclization of carbon- and heteroatom-tethered 1,6-enynes with unsymmetrical 1,2-disubstituted alkynes. This study clearly delineates the ligand requirements for obtaining excellent regio- and enantioselectivity. Furthermore, the ability to utilize various electron-withdrawing groups, and to introduce quaternary carbon stereogenic centers, provides the level of versatility necessary for its application to target-directed synthesis. Additional studies on the development and application of this novel methodology to the total synthesis of natural products are currently underway.
Subject(s)
Alkynes/chemistry , Heterocyclic Compounds, 2-Ring/chemical synthesis , Organometallic Compounds/chemistry , Propionates/chemistry , Rhodium/chemistry , Catalysis , Cyclization , Molecular Structure , StereoisomerismABSTRACT
The crossed intermolecular rhodium-catalyzed [2+2+2] carbocyclization of carbon and heteroatom tethered 1,6-enynes can be accomplished with symmetrical and unsymmetrical alkynes, to afford the corresponding bicyclohexadienes in an efficient and highly selective manner.
Subject(s)
Alkynes/chemistry , Rhodium/chemistry , Catalysis , CyclizationABSTRACT
We have discovered a novel class of (E)-3-acyloxy-4-(arylmethylidene)cyclodeca-1,5-diynes which exhibit promising enediyne-like DNA cleavage and cytotoxic activities. LC-MS analysis of the incubation mixture (pH 8.5, 37 degrees C) confirmed formation of 10-membered ring enediyne presumably via an allylic cation and suggested that the 1,4-benzenoid diradical might be one of the active species for DNA damage and cytotoxicity.