ABSTRACT
Proteins of the bromodomain and extra-terminal (BET) domain family are epigenetic readers that bind acetylated histones through their bromodomains to regulate gene transcription. Dual-bromodomain BET inhibitors (DbBi) that bind with similar affinities to the first (BD1) and second (BD2) bromodomains of BRD2, BRD3, BRD4 and BRDt have displayed modest clinical activity in monotherapy cancer trials. A reduced number of thrombocytes in the blood (thrombocytopenia) as well as symptoms of gastrointestinal toxicity are dose-limiting adverse events for some types of DbBi1-5. Given that similar haematological and gastrointestinal defects were observed after genetic silencing of Brd4 in mice6, the platelet and gastrointestinal toxicities may represent on-target activities associated with BET inhibition. The two individual bromodomains in BET family proteins may have distinct functions7-9 and different cellular phenotypes after pharmacological inhibition of one or both bromodomains have been reported10,11, suggesting that selectively targeting one of the bromodomains may result in a different efficacy and tolerability profile compared with DbBi. Available compounds that are selective to individual domains lack sufficient potency and the pharmacokinetics properties that are required for in vivo efficacy and tolerability assessment10-13. Here we carried out a medicinal chemistry campaign that led to the discovery of ABBV-744, a highly potent and selective inhibitor of the BD2 domain of BET family proteins with drug-like properties. In contrast to the broad range of cell growth inhibition induced by DbBi, the antiproliferative activity of ABBV-744 was largely, but not exclusively, restricted to cell lines of acute myeloid leukaemia and prostate cancer that expressed the full-length androgen receptor (AR). ABBV-744 retained robust activity in prostate cancer xenografts, and showed fewer platelet and gastrointestinal toxicities than the DbBi ABBV-07514. Analyses of RNA expression and chromatin immunoprecipitation followed by sequencing revealed that ABBV-744 displaced BRD4 from AR-containing super-enhancers and inhibited AR-dependent transcription, with less impact on global transcription compared with ABBV-075. These results underscore the potential value of selectively targeting the BD2 domain of BET family proteins for cancer therapy.
Subject(s)
Cell Cycle Proteins/antagonists & inhibitors , Cell Cycle Proteins/chemistry , Prostatic Neoplasms/drug therapy , Prostatic Neoplasms/metabolism , Protein Domains/drug effects , Pyridines/pharmacology , Pyrroles/pharmacology , Transcription Factors/antagonists & inhibitors , Transcription Factors/chemistry , Animals , Cell Cycle Proteins/genetics , Cell Cycle Proteins/metabolism , Cell Line, Tumor , Cell Proliferation/drug effects , Enhancer Elements, Genetic/genetics , Gene Expression Regulation, Neoplastic/drug effects , Humans , Male , Mice , Pyridines/adverse effects , Pyridines/toxicity , Pyrroles/adverse effects , Pyrroles/toxicity , Rats , Receptors, Androgen/metabolism , Transcription Factors/genetics , Transcription Factors/metabolism , Transcription, Genetic/drug effects , Xenograft Model Antitumor AssaysABSTRACT
Cancer therapies targeting the vascular endothelial growth factor (VEGF) signaling pathway can lead to renal damage by disrupting the glomerular ultrafiltration apparatus. The objective of the current study was to identify sensitive biomarkers for VEGF inhibition-induced glomerular changes in rats. Male Sprague-Dawley rats were administered an experimental VEGF receptor (VEGFR) inhibitor, ABT-123, for seven days to investigate the correlation of several biomarkers with microscopic and ultrastructural changes. Glomeruli obtained by laser capture microdissection were also subjected to gene expression analysis to investigate the underlying molecular events of VEGFR inhibition in glomerulus. ABT-123 induced characteristic glomerular ultrastructural changes in rats, including fusion of podocyte foot processes, the presence of subendothelial electron-dense deposits, and swelling and loss of fenestrations in glomerular endothelium. The subtle morphological changes cannot be detected with light microscopy or by changes in standard clinical chemistry and urinalysis. However, urinary albumin increased 44-fold as early as Day three. Urinary ß2-microglobulin levels were also increased. Other urinary biomarkers that are typically associated with tubular injury were not significantly impacted. Such patterns in urinary biomarkers can provide valuable diagnostic insight to VEGF inhibition therapy-induced glomeruli injuries.
Subject(s)
Kidney Diseases/urine , Protein Kinase Inhibitors/adverse effects , Signal Transduction/drug effects , beta 2-Microglobulin/urine , Albumins/metabolism , Animals , Biomarkers/urine , Disease Models, Animal , Kidney Diseases/chemically induced , Kidney Diseases/metabolism , Laser Capture Microdissection , Male , Rats , Rats, Sprague-Dawley , Receptors, Vascular Endothelial Growth Factor/antagonists & inhibitors , Vascular Endothelial Growth Factor A/metabolismABSTRACT
Novel conformationally constrained BET bromodomain inhibitors have been developed. These inhibitors were optimized in two similar, yet distinct chemical series, the 6-methyl-1H-pyrrolo[2,3-c]pyridin-7(6H)-ones (A) and the 1-methyl-1H-pyrrolo[2,3-c]pyridin-7(6H)-ones (B). Each series demonstrated excellent activity in binding and cellular assays, and lead compounds from each series demonstrated significant efficacy in in vivo tumor xenograft models.
Subject(s)
Nuclear Proteins/antagonists & inhibitors , Pyridones/chemistry , Transcription Factors/antagonists & inhibitors , Animals , Binding Sites , Cell Cycle Proteins , Cell Line, Tumor , Cell Proliferation/drug effects , Crystallography, X-Ray , Drug Evaluation, Preclinical , Half-Life , Humans , Mice , Microsomes/metabolism , Molecular Dynamics Simulation , Multiple Myeloma/drug therapy , Nuclear Proteins/metabolism , Protein Structure, Tertiary , Pyridones/pharmacokinetics , Pyridones/pharmacology , Pyridones/therapeutic use , Structure-Activity Relationship , Transcription Factors/metabolism , Transplantation, HeterologousABSTRACT
An NMR fragment screen for binders to the bromodomains of BRD4 identified 2-methyl-3-ketopyrroles 1 and 2. Elaboration of these fragments guided by structure-based design provided lead molecules with significant activity in a mouse tumor model. Further modifications to the methylpyrrole core provided compounds with improved properties and enhanced activity in a mouse model of multiple myeloma.
Subject(s)
Antineoplastic Agents/chemistry , Nuclear Proteins/antagonists & inhibitors , Pyrroles/chemistry , Transcription Factors/antagonists & inhibitors , Animals , Antineoplastic Agents/chemical synthesis , Antineoplastic Agents/pharmacokinetics , Antineoplastic Agents/therapeutic use , Binding Sites , Cell Line, Tumor , Cell Proliferation/drug effects , Crystallography, X-Ray , Drug Design , Half-Life , Humans , Mice , Molecular Dynamics Simulation , Multiple Myeloma/drug therapy , Nuclear Proteins/metabolism , Pyrroles/chemical synthesis , Pyrroles/pharmacokinetics , Pyrroles/therapeutic use , Structure-Activity Relationship , Transcription Factors/metabolism , Transplantation, HeterologousABSTRACT
BACKGROUND: Ilorasertib (ABT-348) is a novel inhibitor of Aurora kinase, vascular endothelial growth factor (VEGF) and platelet-derived growth factor receptors, and the Src families of tyrosine kinases. Ilorasertib alone or in combination with azacitidine demonstrated activity in preclinical models in various hematological malignancies, indicating that pan-Aurora kinase and multiple kinase inhibition may have preferential antileukemic activity. This phase 1 trial determined the safety, pharmacokinetics, and preliminary antitumor activity of ilorasertib alone or combined with azacitidine in advanced hematologic malignancies. PATIENTS AND METHODS: Fifty-two patients (median age, 67 years; 35 % with >4 prior regimens) with acute myelogenous leukaemia (AML; n = 38), myelodysplastic syndrome (n = 12), or chronic myelomonocytic leukaemia (n = 2) received 3 or 6 doses of ilorasertib per 28-day cycle and were assigned to arm A (once-weekly oral), B (twice-weekly oral), C (once-weekly oral plus azacitidine), or D (once-weekly intravenous) treatment. RESULTS: Maximum tolerated doses were not determined; the recommended phase 2 oral monotherapy doses were 540 mg once weekly and 480 mg twice weekly. The most common grade 3/4 adverse events were hypertension (28.8 %), hypokalemia (15.4 %), anemia (13.5 %), and hypophosphatemia (11.5 %). Oral ilorasertib pharmacokinetics appeared dose proportional, with a 15-hour half-life and no interaction with azacitidine. Ilorasertib inhibited biomarkers for Aurora kinase and VEGF receptors, and demonstrated clinical responses in 3 AML patients. CONCLUSIONS: Ilorasertib exhibited acceptable safety and pharmacokinetics at or below the recommended phase 2 dose, displayed evidence of dual Aurora kinase and VEGF receptor kinase inhibition, and activity in AML.
Subject(s)
Aminopyridines , Antineoplastic Agents , Hematologic Neoplasms/drug therapy , Phenylurea Compounds , Protein Kinase Inhibitors , Aged , Aminopyridines/adverse effects , Aminopyridines/pharmacokinetics , Aminopyridines/therapeutic use , Antineoplastic Agents/adverse effects , Antineoplastic Agents/pharmacokinetics , Antineoplastic Agents/therapeutic use , Antineoplastic Combined Chemotherapy Protocols , Aurora Kinases/antagonists & inhibitors , Azacitidine/adverse effects , Azacitidine/pharmacokinetics , Azacitidine/therapeutic use , Female , Hematologic Neoplasms/metabolism , Humans , Male , Maximum Tolerated Dose , Phenylurea Compounds/adverse effects , Phenylurea Compounds/pharmacokinetics , Phenylurea Compounds/therapeutic use , Protein Kinase Inhibitors/adverse effects , Protein Kinase Inhibitors/pharmacokinetics , Protein Kinase Inhibitors/therapeutic use , Receptors, Vascular Endothelial Growth FactorABSTRACT
The objective of this study was to determine the utility of (99m)Tc-3P-Arg-Gly-Asp (RGD2) single photon emission computed tomography (SPECT)/computed tomography (CT) for noninvasive monitoring of integrin αvß3-expression response to antiangiogenic treatment with linifanib. Linifanib or vehicle therapy was carried out in female athymic nu/nu mice bearing U87MG glioma (high αvß3 expression) or PC-3 prostate (low αvß3 expression) tumors at 12.5 mg/kg twice daily. The average tumor volume was 180 ± 90 mm(3) the day prior to baseline SPECT/CT. Longitudinal (99m)Tc-3P-RGD2 SPECT/CT imaging was performed at baseline (-1 day) and days 1, 4, 11, and 18. Tumors were harvested at all imaging time points for histopathological analysis with H&E and immunohistochemistry. A significant difference in tumor volumes between vehicle- and linifanib-treated groups was observed after 4 days of linifanib therapy in the U87MG model. The percent injected dose (%ID) tumor uptake of (99m)Tc-3P-RGD2 peaked in the vehicle-treated group at day 11, while the %ID/cm(3) tumor uptake decreased slowly over the whole study period. During the first 2 days of linifanib treatment, a rapid decrease in both %ID/cm(3) tumor uptake and tumor/muscle ratios of (99m)Tc-3P-RGD2 was observed, followed by a slow decrease until day 18. No decrease in tumor uptake of (99m)Tc-3P-RGD2 or tumor volume was observed for either treatment group in the PC-3 model. Changes in tumor vasculature were confirmed by histopathological H&E analysis and immunohistochemistry. Longitudinal imaging using (99m)Tc-3P-RGD2 SPECT/CT may be a useful tool for monitoring the downstream biologic effects of linifanib therapy.
Subject(s)
Angiogenesis Inhibitors/therapeutic use , Indazoles/therapeutic use , Integrin alphaVbeta3/metabolism , Oligopeptides , Phenylurea Compounds/therapeutic use , Protein-Tyrosine Kinases/antagonists & inhibitors , Radiopharmaceuticals , Animals , Biomarkers, Tumor/metabolism , Dimerization , Female , Glioma , Humans , Male , Mice , Mice, Nude , Multimodal Imaging , Organotechnetium Compounds , Peptides, Cyclic , Positron-Emission Tomography , Prostatic Neoplasms , Technetium , Tomography, X-Ray Computed , Xenograft Model Antitumor AssaysABSTRACT
ABT-348 [1-(4-(4-amino-7-(1-(2-hydroxyethyl)-1H-pyrazol-4-yl)thieno[3,2-c]pyridin-3-yl)phenyl)-3-(3-fluorophenyl)urea] is a novel ATP-competitive multitargeted kinase inhibitor with nanomolar potency (IC(50)) for inhibiting binding and cellular autophosphorylation of Aurora B (7 and 13 nM), C (1 and 13 nM), and A (120 and 189 nM). Cellular activity against Aurora B is reflected by inhibition of phosphorylation of histone H3, induction of polyploidy, and inhibition of proliferation of a variety of leukemia, lymphoma, and solid tumor cell lines (IC(50) = 0.3-21 nM). In vivo inhibition of Aurora B was confirmed in an engrafted leukemia model by observing a decrease in phosphorylation of histone H3 that persisted in a dose-dependent manner for 8 h and correlated with plasma concentration of ABT-348. Evaluation of ABT-348 across a panel of 128 kinases revealed additional potent binding activity (K(i) < 30 nM) against vascular endothelial growth factor receptor (VEGFR)/platelet-derived growth factor receptor (PDGFR) families and the Src family of cytoplasmic tyrosine kinases. VEGFR/PDGFR binding activity correlated with inhibition of autophosphorylation in cells and inhibition of vascular endothelial growth factor (VEGF)-stimulated endothelial cell proliferation (IC(50) ≤ 0.3 nM). Evidence of on-target activity in vivo was provided by the potency for blocking VEGF-mediated vascular permeability and inducing plasma placental growth factor. Activity against the Src kinase family was evident in antiproliferative activity against BCR-ABL chronic myeloid leukemia cells and cells expressing the gleevec-resistant BCR-ABL T315I mutation. On the basis of its unique spectrum of activity, ABT-348 was evaluated and found effective in representative solid tumor [HT1080 and pancreatic carcinoma (MiaPaCa), tumor stasis] and hematological malignancy (RS4;11, regression) xenografts. These results provide the rationale for clinical assessment of ABT-348 as a therapeutic agent in the treatment of cancer.
Subject(s)
Aminopyridines/pharmacology , Antineoplastic Agents/pharmacology , Phenylurea Compounds/pharmacology , Protein Serine-Threonine Kinases/antagonists & inhibitors , Receptors, Platelet-Derived Growth Factor/antagonists & inhibitors , Receptors, Vascular Endothelial Growth Factor/antagonists & inhibitors , src-Family Kinases/antagonists & inhibitors , Aminopyridines/chemistry , Aminopyridines/pharmacokinetics , Aminopyridines/therapeutic use , Animals , Antineoplastic Agents/chemistry , Antineoplastic Agents/pharmacokinetics , Antineoplastic Agents/therapeutic use , Aurora Kinase B , Aurora Kinases , Cell Line, Tumor , Dose-Response Relationship, Drug , Female , Histones/antagonists & inhibitors , Human Umbilical Vein Endothelial Cells , Humans , Leukemia, Experimental/drug therapy , Leukemia, Experimental/enzymology , Male , Mice , Mice, Inbred BALB C , Mice, SCID , Molecular Structure , NIH 3T3 Cells , Phenylurea Compounds/chemistry , Phenylurea Compounds/pharmacokinetics , Phenylurea Compounds/therapeutic use , Time Factors , Xenograft Model Antitumor AssaysABSTRACT
Four hinge-binding scaffolds have been explored for novel selective Aurora kinase inhibitors. The structure activity relationship, selectivity and pharmacokinetic profiles have been evaluated.
Subject(s)
Protein Kinase Inhibitors/chemical synthesis , Protein Serine-Threonine Kinases/antagonists & inhibitors , Animals , Aurora Kinases , Cell Line , Mice , Models, Molecular , Molecular Structure , Protein Kinase Inhibitors/pharmacology , Structure-Activity RelationshipABSTRACT
In an effort to identify kinase inhibitors with dual KDR/Aurora B activity and improved aqueous solubility compared to the Abbott dual inhibitor ABT-348, a series of novel pyrazole pyrimidines structurally related to kinase inhibitor AS703569 were prepared. SAR work provided analogs with significant cellular activity, measureable aqueous solubility and moderate antitumor activity in a mouse tumor model after weekly ip dosing. Unfortunately these compounds were pan-kinase inhibitors that suffered from narrow therapeutic indices which prohibited their use as antitumor agents.
Subject(s)
Protein Kinase Inhibitors/chemistry , Protein Serine-Threonine Kinases/antagonists & inhibitors , Pyrazoles/chemistry , Pyrimidines/chemistry , Vascular Endothelial Growth Factor Receptor-2/antagonists & inhibitors , Amination , Animals , Antineoplastic Agents/chemistry , Antineoplastic Agents/pharmacology , Aurora Kinase B , Aurora Kinases , Cell Line, Tumor , Cell Proliferation/drug effects , Humans , Mice , Microsomes, Liver/drug effects , Microsomes, Liver/enzymology , Models, Molecular , Molecular Structure , Pyrimidines/pharmacology , Structure-Activity Relationship , Xenograft Model Antitumor AssaysABSTRACT
In an effort to identify multi-targeted kinase inhibitors with a novel spectrum of kinase activity, a screen of Abbott proprietary KDR inhibitors against a broad panel of kinases was conducted and revealed a series of thienopyridine ureas with promising activity against the Aurora kinases. Modification of the diphenyl urea and C7 moiety of these compounds provided potent inhibitors with good pharmacokinetic profiles that were efficacious in mouse tumor models after oral dosing. Compound 2 (ABT-348) of this series is currently undergoing Phase I clinical trials in solid and hematological cancer populations.
Subject(s)
Protein Kinase Inhibitors/pharmacology , Pyridines/pharmacology , Urea/pharmacology , Animals , Antineoplastic Agents/chemistry , Cell Line, Tumor , Humans , Mice , Protein Kinase Inhibitors/chemistry , Vascular Endothelial Growth Factor AABSTRACT
BACKGROUND/AIM: The therapeutic potential of bromodomain and extra-terminal motif (BET) inhibitors in hematological cancers has been well established in preclinical and early-stage clinical trials, although as of yet, no BETtargeting agent has achieved approval. To add insight into potential response to mivebresib (ABBV-075), a broadspectrum BET inhibitor, co-clinical modeling of individual patient biopsies was conducted in the context of a Phase I trial in acute myeloid leukemia (AML). MATERIALS AND METHODS: Co-clinical modeling involves taking the patient's biopsy and implanting it in mice with limited passage so that it closely retains the original characteristics of the malignancy and allows comparisons of response between animal model and clinical data. Procedures were developed, initially with neonate NOD/Shi-scid-IL2rγnull (NOG) mice and then optimized with juvenile NOG-EXL as host mice, eventually resulting in a robust rate of engraftment (16 out of 26, 62%). RESULTS: Results from the co-clinical AML patient-derived xenograft (PDX) modeling (6 with >60% inhibition of bone marrow blasts) were consistent with the equivalent clinical data from patients receiving mivebresib in monotherapy, and in combination with venetoclax. The modeling system also demonstrated the activity of a novel BD2-selective BET inhibitor (ABBV-744) in the preclinical AML setting. Both agents were also highly effective in inhibiting blast counts in the spleen (10/10 and 5/6 models, respectively). CONCLUSION: These findings confirm the validity of the model system in the co-clinical setting, establish highly relevant in vivo models for the discovery of cancer therapy, and indicate the therapeutic value of BET inhibitors for AML and, potentially, myelofibrosis treatment.
Subject(s)
Leukemia, Myeloid, Acute , Pyridones , Animals , Cell Line, Tumor , Humans , Leukemia, Myeloid, Acute/drug therapy , Leukemia, Myeloid, Acute/pathology , Mice , Mice, Inbred NOD , Mice, SCID , Pyridones/pharmacology , Pyridones/therapeutic use , SulfonamidesABSTRACT
N-[4-(3-amino-1H-indazol-4-yl)phenyl]-N1-(2-fluoro-5-methyl phenyl)-urea (ABT-869) is a novel multitargeted receptor tyrosine kinase inhibitor that demonstrates single-agent activity in preclinical studies and has undergone phase I and II clinical trials. We characterized the mechanism of action of ABT-869 by examining vascular changes after treatment (25 mg/kg per day) in HT1080 fibrosarcoma and SW620 colon carcinoma cells, using immunohistochemistry, dynamic contrast enhanced-magnetic resonance imaging (DCE-MRI), and hypoxic protein detection. We observed the inhibition of vascular endothelial growth factor receptor 2 and platelet-derived growth factor receptor ß phosphorylation in both tumors and changes in tumor vasculature. Reductions in microvessel density and diameter were observed. Vascular-wall integrity was assessed by colocalization of pericytes and basement membrane. Although both microvessel density and total number of pericytes decreased with treatment, the percentage of pericyte coverage on remaining vessels significantly increased. These data suggest the selective ablation of microvessels lacking pericyte coverage. Functional vascular measures DCE-MRI and hypoxia formation were also tested. After 2 days of treatment on the HT1080 model, vascular permeability, K(trans), was reduced by >60% and hypoxic tumor fraction was significantly decreased, which was also seen in the SW620 tumors after 4 days of treatment. Taken together, decreases in vascular permeability and changes in vascular integrity observed in these studies define the mode of action of ABT-869 and may aid in optimizing the timing of therapeutic window for combination therapies.
Subject(s)
Capillary Permeability/drug effects , Endothelium, Vascular/drug effects , Indazoles/pharmacology , Indazoles/therapeutic use , Neovascularization, Pathologic/drug therapy , Phenylurea Compounds/pharmacology , Phenylurea Compounds/therapeutic use , Receptor Protein-Tyrosine Kinases/antagonists & inhibitors , Xenograft Model Antitumor Assays , Animals , Capillary Permeability/physiology , Cell Line, Tumor , Drug Evaluation, Preclinical/methods , Endothelium, Vascular/metabolism , Humans , Mice , Neovascularization, Pathologic/enzymology , Neovascularization, Pathologic/pathology , Receptor Protein-Tyrosine Kinases/metabolism , Xenograft Model Antitumor Assays/methodsABSTRACT
To further investigate potential mechanisms of resistance to FLT3 inhibitors, we developed a resistant cell line by long-term culture of MV4-11 cells with ABT-869, designated as MV4-11-R. Gene profiling reveals up-regulation of FLT3LG (FLT3 ligand) and BIRC5 (survivin), but down-regulation of SOCS1, SOCS2, and SOCS3 in MV4-11-R cells. Hypermethylation of these SOCS genes leads to their transcriptional silencing. Survivin is directly regulated by STAT3. Stimulation of the parental MV4-11 cells with FLT3 ligand increases the expression of survivin and phosphorylated protein STAT1, STAT3, STAT5. Targeting survivin by short-hairpin RNA (shRNA) in MV4-11-R cells induces apoptosis and augments ABT-869-mediated cytotoxicity. Overexpression of survivin protects MV4-11 from apoptosis. Subtoxic dose of indirubin derivative (IDR) E804 resensitizes MV4-11-R to ABT-869 treatment by inhibiting STAT signaling activity and abolishing survivin expression. Combining IDR E804 with ABT-869 shows potent in vivo efficacy in the MV4-11-R xenograft model. Taken together, these results demonstrate that enhanced activation of STAT pathways and overexpression of survivin are important mechanisms of resistance to ABT-869, suggesting that the STAT pathways and survivin could be potential targets for reducing resistance developed in patients receiving FLT3 inhibitors.
Subject(s)
Gene Expression Regulation, Neoplastic/genetics , Leukemia, Myeloid, Acute/metabolism , Leukemia, Myeloid, Acute/pathology , Microtubule-Associated Proteins/metabolism , STAT3 Transcription Factor/metabolism , Signal Transduction/drug effects , fms-Like Tyrosine Kinase 3/antagonists & inhibitors , Animals , Apoptosis/drug effects , Cell Line, Tumor , Drug Resistance, Neoplasm/drug effects , Epoxy Compounds/pharmacology , Female , Humans , Indazoles/pharmacology , Inhibitor of Apoptosis Proteins , Leukemia, Myeloid, Acute/genetics , Ligands , Mice , Mice, Inbred BALB C , Mice, Nude , Microtubule-Associated Proteins/genetics , Phenylurea Compounds/pharmacology , STAT3 Transcription Factor/genetics , Sesquiterpenes/pharmacology , Substrate Specificity , Survivin , Up-Regulation/genetics , Xenograft Model Antitumor Assays , fms-Like Tyrosine Kinase 3/metabolismABSTRACT
In an effort to discover Aurora kinase inhibitors, an HTS hit revealed an amide containing pyrrolopyrimidine compound. Replacement of the pyrrolopyrimidine residue with a thienopyrimidine moiety led to a series of potent and selective Aurora inhibitors.
Subject(s)
Drug Discovery , Protein Kinase Inhibitors/pharmacology , Protein Serine-Threonine Kinases/antagonists & inhibitors , Pyrimidines/pharmacology , Animals , Aurora Kinases , Dose-Response Relationship, Drug , High-Throughput Screening Assays , Hydrogen Bonding , Mice , Models, Molecular , Molecular Structure , Protein Kinase Inhibitors/chemical synthesis , Protein Kinase Inhibitors/chemistry , Protein Serine-Threonine Kinases/metabolism , Pyrimidines/chemical synthesis , Pyrimidines/chemistry , Stereoisomerism , Structure-Activity RelationshipABSTRACT
Cyclin-dependent kinase 9 (CDK9) is a serine/threonine kinase involved in the regulation of transcription elongation. An inhibition of CDK9 downregulates a number of short-lived proteins responsible for tumor maintenance and survival, including the antiapoptotic BCL-2 family member MCL-1. As pan-CDK inhibitors under development have faced dosing and toxicity challenges in the clinical setting, we generated selective CDK9 inhibitors that could be amenable to an oral administration. Here, we report the lead optimization of a series of azaindole-based inhibitors. To overcome early challenges with promiscuity and cardiovascular toxicity, carboxylates were introduced into the pharmacophore en route to compounds such as 14 and 16. These CDK9 inhibitors demonstrated a reduced toxicity, adequate pharmacokinetic properties, and a robust in vivo efficacy in mice upon oral dosing.
ABSTRACT
Dual bromodomain BET inhibitors that bind with similar affinities to the first and second bromodomains across BRD2, BRD3, BRD4, and BRDT have displayed modest activity as monotherapy in clinical trials. Thrombocytopenia, closely followed by symptoms characteristic of gastrointestinal toxicity, have presented as dose-limiting adverse events that may have prevented escalation to higher dose levels required for more robust efficacy. ABBV-744 is a highly selective inhibitor for the second bromodomain of the four BET family proteins. In contrast to the broad antiproliferative activities observed with dual bromodomain BET inhibitors, ABBV-744 displayed significant antiproliferative activities largely although not exclusively in cancer cell lines derived from acute myeloid leukemia and androgen receptor positive prostate cancer. Studies in acute myeloid leukemia xenograft models demonstrated antitumor efficacy for ABBV-744 that was comparable with the pan-BET inhibitor ABBV-075 but with an improved therapeutic index. Enhanced antitumor efficacy was also observed with the combination of ABBV-744 and the BCL-2 inhibitor, venetoclax compared with monotherapies of either agent alone. These results collectively support the clinical evaluation of ABBV-744 in AML (Clinical Trials.gov identifier: NCT03360006).
Subject(s)
Bridged Bicyclo Compounds, Heterocyclic/pharmacology , Leukemia, Myeloid, Acute/drug therapy , Proteins/antagonists & inhibitors , Proto-Oncogene Proteins c-bcl-2/antagonists & inhibitors , Pyridines/pharmacology , Pyrroles/pharmacology , Sulfonamides/pharmacology , Animals , Antineoplastic Agents/pharmacology , Apoptosis , Cell Proliferation , Drug Therapy, Combination , Female , Humans , Leukemia, Myeloid, Acute/metabolism , Leukemia, Myeloid, Acute/pathology , Mice , Mice, Inbred NOD , Mice, SCID , Tumor Cells, Cultured , Xenograft Model Antitumor AssaysABSTRACT
MCL-1 is one of the most frequently amplified genes in cancer, facilitating tumor initiation and maintenance and enabling resistance to anti-tumorigenic agents including the BCL-2 selective inhibitor venetoclax. The expression of MCL-1 is maintained via P-TEFb-mediated transcription, where the kinase CDK9 is a critical component. Consequently, we developed a series of potent small-molecule inhibitors of CDK9, exemplified by the orally active A-1592668, with CDK selectivity profiles that are distinct from related molecules that have been extensively studied clinically. Short-term treatment with A-1592668 rapidly downregulates RNA pol-II (Ser 2) phosphorylation resulting in the loss of MCL-1 protein and apoptosis in MCL-1-dependent hematologic tumor cell lines. This cell death could be attenuated by either inhibiting caspases or overexpressing BCL-2 protein. Synergistic cell killing was also observed between A-1592668 or the related analog A-1467729, and venetoclax in a number of hematologic cell lines and primary NHL patient samples. Importantly, the CDK9 inhibitor plus venetoclax combination was well tolerated in vivo and demonstrated efficacy superior to either agent alone in mouse models of lymphoma and AML. These data indicate that CDK9 inhibitors could be highly efficacious in tumors that depend on MCL-1 for survival or when used in combination with venetoclax in malignancies dependent on MCL-1 and BCL-2.
Subject(s)
Antineoplastic Agents/pharmacology , Cyclin-Dependent Kinase 9/antagonists & inhibitors , Hematologic Neoplasms , Protein Kinase Inhibitors/pharmacology , Animals , Apoptosis/drug effects , Bridged Bicyclo Compounds, Heterocyclic/pharmacology , Cell Line, Tumor , Cell Survival/drug effects , Drug Synergism , Humans , Mice , Sulfonamides/pharmacology , Tumor Cells, Cultured , Xenograft Model Antitumor AssaysABSTRACT
The BET family of proteins consists of BRD2, BRD3, BRD4, and BRDt. Each protein contains two distinct bromodomains (BD1 and BD2). BET family bromodomain inhibitors under clinical development for oncology bind to each of the eight bromodomains with similar affinities. We hypothesized that it may be possible to achieve an improved therapeutic index by selectively targeting subsets of the BET bromodomains. Both BD1 and BD2 are highly conserved across family members (>70% identity), whereas BD1 and BD2 from the same protein exhibit a larger degree of divergence (â¼40% identity), suggesting selectivity between BD1 and BD2 of all family members would be more straightforward to achieve. Exploiting the Asp144/His437 and Ile146/Val439 sequence differences (BRD4 BD1/BD2 numbering) allowed the identification of compound 27 demonstrating greater than 100-fold selectivity for BRD4 BD2 over BRD4 BD1. Further optimization to improve BD2 selectivity and oral bioavailability resulted in the clinical development compound 46 (ABBV-744).
Subject(s)
Cell Cycle Proteins/antagonists & inhibitors , Cell Cycle Proteins/metabolism , Drug Discovery/methods , Pyridines/chemistry , Pyridines/metabolism , Pyrroles/chemistry , Pyrroles/metabolism , Transcription Factors/antagonists & inhibitors , Transcription Factors/metabolism , Animals , Female , HeLa Cells , Humans , Mice , Mice, SCID , Protein Domains/drug effects , Protein Domains/physiology , Protein Structure, Secondary , Protein Structure, Tertiary , Pyridines/pharmacology , Pyrroles/pharmacology , Xenograft Model Antitumor Assays/methodsABSTRACT
ABT-869 [N-(4-(3-amino-1H-indazol-4-yl)phenyl)-N'-(2-fluoro-5-methylphenyl)urea] is a novel multitargeted inhibitor of the vascular endothelial growth factor (VEGF) and platelet-derived growth factor (PDGF) receptor tyrosine kinase family members. ABT-869 demonstrates tumor growth inhibition in multiple preclinical animal models and in early clinical trials. VEGF receptor inhibition is also associated with reversible hypertension that may limit its benefit clinically. To evaluate optimal therapeutic approaches to prevent hypertension with VEGF receptor inhibition, we characterized the dose-dependent effects of seven antihypertensive agents from three mechanistic classes [angiotensin-converting enzyme inhibitors (ACEis), angiotensin receptor blockers (ARBs), calcium channel blockers (CCBs)] on hypertension induced by ABT-869 in conscious telemetry rats. We report that ABT-869-induced hypertension can be prevented and reversed with subtherapeutic or therapeutic doses of antihypertensive drugs with a general rank order of ACEi > ARB > CCB. In SCID mice, the ACE inhibitor, enalapril (C(20)H(28)N(2)O(5) x C(4)H(4)O(4)) at 30 mg/kg, prevented hypertension, with no attenuation of the antitumor efficacy of ABT-869. These studies demonstrate that the adverse cardiovascular effects of the VEGF/PDGF receptor tyrosine kinase inhibitor, ABT-869, are readily controlled by conventional antihypertensive therapy without affecting antitumor efficacy.
Subject(s)
Antihypertensive Agents/pharmacology , Blood Pressure/drug effects , Indazoles/pharmacology , Neoplasms/drug therapy , Phenylurea Compounds/pharmacology , Protein Kinase Inhibitors/pharmacology , Protein-Tyrosine Kinases/antagonists & inhibitors , Acrylates/pharmacology , Amlodipine/pharmacology , Angiotensin Receptor Antagonists , Angiotensin-Converting Enzyme Inhibitors/pharmacology , Animals , Benzimidazoles/pharmacology , Benzoates/pharmacology , Calcium Channel Blockers/pharmacology , Dose-Response Relationship, Drug , Enalapril/pharmacology , Humans , Imidazoles/pharmacology , Indazoles/adverse effects , Indazoles/therapeutic use , Lisinopril/pharmacology , Male , Mice , Mice, SCID , Neoplasms/pathology , Nifedipine/pharmacology , Phenylurea Compounds/adverse effects , Phenylurea Compounds/therapeutic use , Protein Kinase Inhibitors/blood , Protein Kinase Inhibitors/therapeutic use , Ramipril/pharmacology , Rats , Rats, Sprague-Dawley , Telmisartan , Thiophenes/pharmacology , Xenograft Model Antitumor AssaysABSTRACT
ABT-869 is a novel multitargeted inhibitor of vascular endothelial growth factor and platelet-derived growth factor receptor tyrosine kinases (RTKs) with potent antiangiogenic properties that slow tumor progression. Vascular endothelial growth factor receptor blockade has been shown to produce hypertension. Atrasentan is a potent and selective endothelin (ETA) receptor antagonist that lowers blood pressure and affects tumor growth. To assess the utility of ETA receptor blockade in controlling hypertension with RTK inhibition, we evaluated the ability of atrasentan to block hypertension with ABT-869 in conscious, telemetry-instrumented rats. Changes in mean arterial pressure (MAP) and heart rate (HR) were evaluated using mean values and the area under the curve (AUC). Atrasentan (0.5, 1.5, and 5.0 mg kg(-1) d(-1) for 5 days) elicited dose-dependent decreases in MAP-AUC (-16.7 +/- 1.3, -20.94 +/- 3.68, and -30.12 +/- 3.57 mm Hg x day, respectively) compared with vehicle. ABT-869 (1, 3, 10, 30 mg kg(-1) d(-1) for 5 days) increased MAP compared with vehicle (MAP-AUC values of -5.52 +/- 3.75, 12.7 +/- 8.4, 37.5 +/- 4.4, and 63.8 +/- 3.3 mm Hg x day, respectively). Pretreatment with atrasentan (5 mg/kg for 5 days) prevented and abolished the hypertensive effects of ABT-869. Thus, ETA receptor blockade effectively alleviated hypertension with RTK inhibition and may serve a dual therapeutic role by preventing hypertension and slowing tumor progression.