RESUMEN
Current KRASG12C (OFF) inhibitors that target inactive GDP-bound KRASG12C cause responses in less than half of patients and these responses are not durable. A class of RASG12C (ON) inhibitors that targets active GTP-bound KRASG12C blocks ERK signaling more potently than the inactive-state inhibitors. Sensitivity to either class of agents is strongly correlated with inhibition of mTORC1 activity. We have previously shown that PI3K/mTOR and ERK-signaling pathways converge on key cellular processes and that inhibition of both pathways is required for inhibition of these processes and for significant antitumor activity. We find here that the combination of a KRASG12C inhibitor with a selective mTORC1 kinase inhibitor causes synergistic inhibition of Cyclin D1 expression and cap-dependent translation. Moreover, BIM upregulation by KRASG12C inhibition and inhibition of MCL-1 expression by the mTORC1 inhibitor are both required to induce significant cell death. In vivo, this combination causes deep, durable tumor regressions and is well tolerated. This study suggests that the ERK and PI3K/mTOR pathways each mitigate the effects of inhibition of the other and that combinatorial inhibition is a potential strategy for treating KRASG12C-dependent lung cancer.
Asunto(s)
Carcinoma de Pulmón de Células no Pequeñas , Sinergismo Farmacológico , Neoplasias Pulmonares , Diana Mecanicista del Complejo 1 de la Rapamicina , Proteínas Proto-Oncogénicas p21(ras) , Diana Mecanicista del Complejo 1 de la Rapamicina/metabolismo , Diana Mecanicista del Complejo 1 de la Rapamicina/antagonistas & inhibidores , Humanos , Proteínas Proto-Oncogénicas p21(ras)/genética , Proteínas Proto-Oncogénicas p21(ras)/metabolismo , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Carcinoma de Pulmón de Células no Pequeñas/metabolismo , Carcinoma de Pulmón de Células no Pequeñas/patología , Carcinoma de Pulmón de Células no Pequeñas/genética , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patología , Neoplasias Pulmonares/genética , Animales , Línea Celular Tumoral , Ratones , Serina-Treonina Quinasas TOR/metabolismo , Serina-Treonina Quinasas TOR/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/farmacología , Ensayos Antitumor por Modelo de Xenoinjerto , Transducción de Señal/efectos de los fármacos , Ciclina D1/metabolismo , Ciclina D1/genética , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/metabolismo , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/genética , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/antagonistas & inhibidores , Femenino , Proteína 11 Similar a Bcl2/metabolismo , Proteína 11 Similar a Bcl2/genéticaRESUMEN
Acute myeloid leukemia (AML) is a hematologic malignancy characterized by infiltration of the blood and bone marrow, exhibiting a low remission rate and high recurrence rate. Current research has demonstrated that class I HDAC inhibitors can downregulate anti-apoptotic proteins, leading to apoptosis of AML cells. In the present investigation, we conducted structural modifications of marine cytotoxin Santacruzamate A (SCA), a compound known for its inhibitory activity towards HDACs, resulting in the development of a novel series of potent class I HDACs hydrazide inhibitors. Representative hydrazide-based compound 25c exhibited concentration-dependent induction of apoptosis in AML cells as a single agent. Moreover, 25c exhibited a synergistic anti-AML effect when combined with Venetoclax, a clinical Bcl-2 inhibitor employed in AML therapy. This combination resulted in a more pronounced downregulation of anti-apoptotic proteins Mcl-1 and Bcl-xL, along with a significant upregulation of the pro-apoptotic protein cleaved-caspase3 and the DNA double-strand break biomarker γ-H2AX compared to monotherapy. These results highlighted the potential of 25c as a promising lead compound for AML treatment, particularly when used in combination with Venetoclax.
Asunto(s)
Antineoplásicos , Apoptosis , Compuestos Bicíclicos Heterocíclicos con Puentes , Sinergismo Farmacológico , Inhibidores de Histona Desacetilasas , Leucemia Mieloide Aguda , Sulfonamidas , Humanos , Sulfonamidas/farmacología , Sulfonamidas/química , Leucemia Mieloide Aguda/tratamiento farmacológico , Inhibidores de Histona Desacetilasas/farmacología , Inhibidores de Histona Desacetilasas/química , Compuestos Bicíclicos Heterocíclicos con Puentes/farmacología , Compuestos Bicíclicos Heterocíclicos con Puentes/química , Apoptosis/efectos de los fármacos , Línea Celular Tumoral , Antineoplásicos/farmacología , Antineoplásicos/química , Histona Desacetilasa 1/antagonistas & inhibidores , Histona Desacetilasas/metabolismo , Animales , Caspasa 3/metabolismo , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/antagonistas & inhibidoresRESUMEN
Myeloid cell leukemia 1 (Mcl1), a critical protein that regulates apoptosis, has been considered as a promising target for antitumor drugs. The conventional pharmacophore screening approach has limitations in conformation sampling and data mining. Here, we offered an innovative solution to identify Mcl1 inhibitors with molecular dynamics-refined pharmacophore and machine learning methods. Considering the safety and druggability of FDA-approved drugs, virtual screening of the database was performed to discover Mcl1 inhibitors, and the hit was subsequently validated via TR-FRET, cytotoxicity, and flow cytometry assays. To reveal the binding characteristics shared by the hit and a typical Mcl1 selective inhibitor, we employed quantum mechanics and molecular mechanics (QM/MM) calculations, umbrella sampling, and metadynamics in this work. The combined studies suggested that fluvastatin had promising cell inhibitory potency and was suitable for further investigation. We believe that this research will shed light on the discovery of novel Mcl1 inhibitors that can be used as a supplemental treatment against leukemia and provide a possible method to improve the accuracy of drug repurposing with limited computational resources while balancing the costs of experimentation well.
Asunto(s)
Antineoplásicos , Reposicionamiento de Medicamentos , Aprendizaje Automático , Simulación de Dinámica Molecular , Proteína 1 de la Secuencia de Leucemia de Células Mieloides , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/antagonistas & inhibidores , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/metabolismo , Humanos , Antineoplásicos/farmacología , Antineoplásicos/química , Teoría Cuántica , Línea Celular Tumoral , Fluvastatina/farmacología , Fluvastatina/química , FarmacóforoRESUMEN
RAS pathway mutations, which are present in 30% of patients with chronic myelomonocytic leukemia (CMML) at diagnosis, confer a high risk of resistance to and progression after hypomethylating agent (HMA) therapy, the current standard of care for the disease. Here, using single-cell, multi-omics technologies, we seek to dissect the biological mechanisms underlying the initiation and progression of RAS pathway-mutated CMML. We identify that RAS pathway mutations induce transcriptional reprogramming of hematopoietic stem and progenitor cells (HSPCs) and downstream monocytic populations in response to cell-intrinsic and -extrinsic inflammatory signaling that also impair the functions of immune cells. HSPCs expand at disease progression after therapy with HMA or the BCL2 inhibitor venetoclax and rely on the NF-κB pathway effector MCL1 to maintain survival. Our study has implications for the development of therapies to improve the survival of patients with RAS pathway-mutated CMML.
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Apoptosis , Leucemia Mielomonocítica Crónica , Mutación , Proteína 1 de la Secuencia de Leucemia de Células Mieloides , Leucemia Mielomonocítica Crónica/tratamiento farmacológico , Leucemia Mielomonocítica Crónica/patología , Leucemia Mielomonocítica Crónica/genética , Leucemia Mielomonocítica Crónica/metabolismo , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/metabolismo , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/genética , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/antagonistas & inhibidores , Humanos , Apoptosis/efectos de los fármacos , Animales , Mutación/genética , Ratones , Transducción de Señal/efectos de los fármacos , Células Madre Hematopoyéticas/metabolismo , Células Madre Hematopoyéticas/efectos de los fármacos , Progresión de la Enfermedad , Sulfonamidas/farmacología , Sulfonamidas/uso terapéutico , FN-kappa B/metabolismo , Metilación de ADN/efectos de los fármacos , Metilación de ADN/genética , Compuestos Bicíclicos Heterocíclicos con Puentes/farmacología , Compuestos Bicíclicos Heterocíclicos con Puentes/uso terapéutico , Crisis Blástica/patología , Crisis Blástica/tratamiento farmacológico , Crisis Blástica/genética , Crisis Blástica/metabolismoRESUMEN
Triple-negative breast cancer (TNBC) is an aggressive breast cancer sub-type with limited treatment options and poor prognosis. Currently, standard treatments for TNBC include surgery, chemotherapy, and anti-PDL1 therapy. These therapies have limited efficacy in advanced stages. Myeloid-cell leukemia 1 (MCL1) is an anti-apoptotic BCL2 family protein. High expression of MCL1 contributes to chemotherapy resistance and is associated with a worse prognosis in TNBC. MCL1 inhibitors are in clinical trials for TNBC, but response rates to these inhibitors can vary and predictive markers are lacking. Currently, we identified a 4-member (AXL, ETS1, IL6, EFEMP1) gene signature (GS) that predicts MCL1 inhibitor sensitivity in TNBC cells. Factors encoded by these genes regulate signaling pathways to promote MCL1 inhibitor resistance. Small molecule inhibitors of the GS factors can overcome resistance and sensitize otherwise resistant TNBC cells to MCL1 inhibitor treatment. These findings offer insights into potential therapeutic strategies and tumor stratification for MCL1 inhibitor use in TNBC.
Asunto(s)
Resistencia a Antineoplásicos , Proteína 1 de la Secuencia de Leucemia de Células Mieloides , Neoplasias de la Mama Triple Negativas , Humanos , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/metabolismo , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/genética , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/antagonistas & inhibidores , Neoplasias de la Mama Triple Negativas/metabolismo , Neoplasias de la Mama Triple Negativas/patología , Neoplasias de la Mama Triple Negativas/tratamiento farmacológico , Neoplasias de la Mama Triple Negativas/genética , Femenino , Línea Celular Tumoral , Resistencia a Antineoplásicos/efectos de los fármacos , Biomarcadores de Tumor/metabolismo , Biomarcadores de Tumor/genética , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Antineoplásicos/farmacología , Interleucina-6/metabolismo , Interleucina-6/genética , Proteína Proto-Oncogénica c-ets-1RESUMEN
Considering the established pharmacokinetics and toxicity profiles, drug repurposing has emerged as an alternative therapeutic approach for treating cancer. Mefloquine has previously demonstrated inhibitory effects on multiple cancer types. This study aims to explore the impact of mefloquine on nasopharyngeal carcinoma (NPC). We found that mefloquine, at pharmacologically achievable concentrations, displayed anti-NPC activity while sparing normal counterparts. Mefloquine inhibits proliferation and induces death by reducing the levels of Cyclin A2, Bcl-2, and Bcl-xL. Intriguingly, we observed an increase in the levels of the anti-apoptotic protein Mcl-1. Mefloquine exerts its effects on NPC cells by inducing lysosomal-mediated ROS production, and the heightened expression of Mcl-1 is a consequence of ROS generation in mefloquine-treated NPC cells. The combination of an Mcl-1 inhibitor with mefloquine synergistically inhibits NPC growth in mice without causing substantial toxicity. These findings demonstrate the effectiveness and limited toxicity of mefloquine as a monotherapy and in combination with an Mcl-1 inhibitor. Our research underscores the promise of the mefloquine and Mcl-1 inhibitor combination as a potential treatment for NPC. Additionally, the elevation of Mcl-1 is a compensatory response in cells exposed to oxidative stress, offering a potential target to overcome resistance induced by pro-oxidant therapies.
Asunto(s)
Proliferación Celular , Mefloquina , Proteína 1 de la Secuencia de Leucemia de Células Mieloides , Carcinoma Nasofaríngeo , Neoplasias Nasofaríngeas , Especies Reactivas de Oxígeno , Animales , Humanos , Ratones , Apoptosis/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Reposicionamiento de Medicamentos , Sinergismo Farmacológico , Mefloquina/farmacología , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/metabolismo , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/antagonistas & inhibidores , Carcinoma Nasofaríngeo/tratamiento farmacológico , Carcinoma Nasofaríngeo/patología , Neoplasias Nasofaríngeas/tratamiento farmacológico , Neoplasias Nasofaríngeas/patología , Especies Reactivas de Oxígeno/metabolismo , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
As a tripartite cell death switch, B-cell lymphoma protein 2 (Bcl-2) family members precisely regulate the endogenous apoptosis pathway in response to various cell signal stresses through protein-protein interactions. Myeloid leukemia-1 (Mcl-1), a key anti-apoptotic Bcl-2 family member, is positioned downstream in the endogenous apoptotic pathway and plays a central role in regulating mitochondrial function. Mcl-1 is highly expressed in a variety of hematological malignancies and solid tumors, contributing to tumorigenesis, poor prognosis, and chemoresistance, making it an attractive target for cancer treatment. This Perspective aims to discuss the mechanism by which Mcl-1 regulates apoptosis and non-apoptotic functions in cancer cells and to outline the discovery and optimization process of potent Mcl-1 modulators. In addition, we summarize the structural characteristics of potent inhibitors that bind to Mcl-1 through multiple co-crystal structures and analyze the cardiotoxicity caused by current Mcl-1 inhibitors, providing prospects for rational targeting of Mcl-1.
Asunto(s)
Antineoplásicos , Proteína 1 de la Secuencia de Leucemia de Células Mieloides , Neoplasias , Humanos , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/antagonistas & inhibidores , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/metabolismo , Antineoplásicos/farmacología , Antineoplásicos/química , Antineoplásicos/uso terapéutico , Neoplasias/tratamiento farmacológico , Neoplasias/patología , Neoplasias/metabolismo , Apoptosis/efectos de los fármacos , AnimalesRESUMEN
BAX and BAK are pro-apoptotic members of the BCL2 family that are required to permeabilize the mitochondrial outer membrane. The proteins can adopt a non-activated monomeric conformation, or an activated conformation in which the exposed BH3 domain facilitates binding either to a prosurvival protein or to another activated BAK or BAX protein to promote pore formation. Certain cancer cells are proposed to have high levels of activated BAK sequestered by MCL1 or BCLXL, thus priming these cells to undergo apoptosis in response to BH3 mimetic compounds that target MCL1 or BCLXL. Here we report the first antibody, 14G6, that is specific for the non-activated BAK conformer. A crystal structure of 14G6 Fab bound to BAK revealed a binding site encompassing both the α1 helix and α5-α6 hinge regions of BAK, two sites involved in the unfolding of BAK during its activation. In mitochondrial experiments, 14G6 inhibited BAK unfolding triggered by three diverse BAK activators, supporting crucial roles for both α1 dissociation and separation of the core (α2-α5) and latch (α6-α9) regions in BAK activation. 14G6 bound the majority of BAK in several leukaemia cell lines, and binding decreased following treatment with BH3 mimetics, indicating only minor levels of constitutively activated BAK in those cells. In summary, 14G6 provides a new means of assessing BAK status in response to anti-cancer treatments.
Asunto(s)
Proteína Destructora del Antagonista Homólogo bcl-2 , Proteína Destructora del Antagonista Homólogo bcl-2/metabolismo , Humanos , Apoptosis/efectos de los fármacos , Línea Celular Tumoral , Neoplasias/patología , Neoplasias/metabolismo , Neoplasias/tratamiento farmacológico , Mitocondrias/metabolismo , Mitocondrias/efectos de los fármacos , Animales , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/metabolismo , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/antagonistas & inhibidoresRESUMEN
BH3 mimetics, including the BCL2/BCLXL/BCLw inhibitor navitoclax and MCL1 inhibitors S64315 and tapotoclax, have undergone clinical testing for a variety of neoplasms. Because of toxicities, including thrombocytopenia after BCLXL inhibition as well as hematopoietic, hepatic and possible cardiac toxicities after MCL1 inhibition, there is substantial interest in finding agents that can safely sensitize neoplastic cells to these BH3 mimetics. Building on the observation that BH3 mimetic monotherapy induces AMP kinase (AMPK) activation in multiple acute leukemia cell lines, we report that the AMPK inhibitors (AMPKis) dorsomorphin and BAY-3827 sensitize these cells to navitoclax or MCL1 inhibitors. Cell fractionation and phosphoproteomic analyses suggest that sensitization by dorsomorphin involves dephosphorylation of the proapoptotic BCL2 family member BAD at Ser75 and Ser99, leading BAD to translocate to mitochondria and inhibit BCLXL. Consistent with these results, BAD knockout or mutation to BAD S75E/S99E abolishes the sensitizing effects of dorsomorphin. Conversely, dorsomorphin synergizes with navitoclax or the MCL1 inhibitor S63845 to induce cell death in primary acute leukemia samples ex vivo and increases the antitumor effects of navitoclax or S63845 in several xenograft models in vivo with little or no increase in toxicity in normal tissues. These results suggest that AMPK inhibition can sensitize acute leukemia to multiple BH3 mimetics, potentially allowing administration of lower doses while inducing similar antineoplastic effects.
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Proteínas Quinasas Activadas por AMP , Compuestos de Anilina , Proteína 1 de la Secuencia de Leucemia de Células Mieloides , Pirimidinas , Sulfonamidas , Proteína bcl-X , Humanos , Animales , Compuestos de Anilina/farmacología , Sulfonamidas/farmacología , Proteínas Quinasas Activadas por AMP/metabolismo , Ratones , Proteína bcl-X/metabolismo , Proteína bcl-X/antagonistas & inhibidores , Línea Celular Tumoral , Pirimidinas/farmacología , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/metabolismo , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/antagonistas & inhibidores , Pirazoles/farmacología , Proteína Letal Asociada a bcl/metabolismo , Apoptosis/efectos de los fármacos , Muerte Celular/efectos de los fármacos , Leucemia/tratamiento farmacológico , Leucemia/patología , Leucemia/metabolismo , Fosforilación/efectos de los fármacos , Fragmentos de Péptidos/farmacología , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Proto-Oncogénicas/antagonistas & inhibidores , Sinergismo FarmacológicoRESUMEN
After an initial positive response to chemotherapy, cancer patients often become resistant and experience relapse. Our previous research identified eukaryotic translation initiation factor 4E (eIF4E) as a crucial target to overcome chemoresistance. In this study, we delved further into the role and therapeutic potential of myeloid cell leukemia 1 (Mcl-1), an eIF4E-mediated target, in chemoresistance. We showed that the levels of phosphor and total eIF4E, as well as Mcl-1, were elevated in chemoresistant cervical but not colon cancer cells. Mcl-1 inhibitor S64315 decreased Mcl-1 levels in chemoresistant cancer cells, regardless of Mcl-1 upregulation, decreased viability in chemoresistant cancer cells and acted synergistically with chemotherapy drugs. The combined inhibition of Mcl-1 and B-cell lymphoma 2 (Bcl-2), employing both genetic and pharmacological approaches, led to a markedly more substantial decrease in viability compared with the inhibition of either target individually. The combination of S64315 and Bcl-2 inhibitors reduced tumor growth in chemoresistant cervical and colon cancer models without causing general toxicity in mice. This combination also prolonged overall survival compared with using S64315 or venetoclax alone. Our research highlights the therapeutic potential of inhibiting Mcl-1 and Bcl-2 simultaneously in chemoresistant cancers and provides a rationale for initiating clinical trials to investigate the combination of S64315 and venetoclax for the treatment of advanced colon and cervical cancer.
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Neoplasias del Colon , Resistencia a Antineoplásicos , Sulfonamidas , Animales , Humanos , Ratones , Apoptosis , Compuestos Bicíclicos Heterocíclicos con Puentes , Línea Celular Tumoral , Neoplasias del Colon/tratamiento farmacológico , Factor 4E Eucariótico de Iniciación , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/antagonistas & inhibidores , Proteínas Proto-Oncogénicas c-bcl-2/antagonistas & inhibidoresRESUMEN
ABBV-467 is a highly potent and selective MCL-1 inhibitor that was advanced to a phase I clinical trial for the treatment of multiple myeloma. Due to its large size and structural complexity, ABBV-467 is a challenging synthetic target. Herein, we describe the synthesis of ABBV-467 on a decagram scale, which enabled preclinical characterization. The strategy is convergent and stereoselective, featuring a hindered biaryl cross coupling, enantioselective hydrogenation, and conformationally preorganized macrocyclization by C-O bond formation as key steps.
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Antineoplásicos , Proteína 1 de la Secuencia de Leucemia de Células Mieloides , Antineoplásicos/farmacología , Hidrogenación , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/antagonistas & inhibidoresRESUMEN
Tuberculosis (TB) accounts for 1.6 million deaths annually and over 25% of deaths due to antimicrobial resistance. Mycobacterium tuberculosis (M.tb) drives MCL-1 expression (family member of anti-apoptotic BCL-2 proteins) to limit apoptosis and grow intracellularly in human macrophages. The feasibility of re-purposing specific MCL-1 and BCL-2 inhibitors to limit M.tb growth, using inhibitors that are in clinical trials and FDA-approved for cancer treatment has not be tested previously. We show that specifically inhibiting MCL-1 and BCL-2 induces apoptosis of M.tb-infected macrophages, and markedly reduces M.tb growth in human and murine macrophages, and in a pre-clinical model of human granulomas. MCL-1 and BCL-2 inhibitors limit growth of drug resistant and susceptible M.tb in macrophages and act in additive fashion with the antibiotics isoniazid and rifampicin. This exciting work uncovers targeting the intrinsic apoptosis pathway as a promising approach for TB host-directed therapy. Since safety and activity studies are underway in cancer clinics for MCL-1 and BCL-2 inhibitors, we expect that re-purposing them for TB treatment should translate more readily and rapidly to the clinic. Thus, the work supports further development of this host-directed therapy approach to augment current TB treatment.
Asunto(s)
Antineoplásicos , Antituberculosos , Reposicionamiento de Medicamentos , Mycobacterium tuberculosis , Proteína 1 de la Secuencia de Leucemia de Células Mieloides , Proteínas Proto-Oncogénicas c-bcl-2 , Tuberculosis , Animales , Humanos , Ratones , Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Antituberculosos/farmacología , Antituberculosos/uso terapéutico , Antituberculosos/metabolismo , Macrófagos/efectos de los fármacos , Mycobacterium tuberculosis/efectos de los fármacos , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/antagonistas & inhibidores , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/antagonistas & inhibidores , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Tuberculosis/tratamiento farmacológico , Tuberculosis/microbiologíaRESUMEN
PURPOSE: AZD5991 is a potent and selective macrocyclic inhibitor of Mcl-1 in clinical development. Developing an intravenous solution formulation for AZD5991 proved to be challenging primarily due to the poor intrinsic solubility of AZD5991. In this article are described studies performed to select a suitable crystalline form and to assess physicochemical properties of AZD5991 to aid in the design of a solution formulation for preclinical studies. METHODS: It is preferable that the preclinical formulation has a line of sight for clinical formulation. For AZD5991, a concentration of at least 20 mg/ml was required for toxicology studies. Toward this goal, extensive pre-formulation characterization of AZD5991 including solid form analysis, pH-solubility profiling and solubility determination in cosolvents and other solubilizing media were carried out. RESULTS & DISCUSSION: Crystalline Form A, which is more stable in aqueous solution and possesses acceptable thermal stability, was selected for preclinical and clinical development of AZD5991. Extensive solubility evaluation revealed an interesting pH-solubility profile that significantly enhances solubilization at pH > 8.5 to allow solution concentrations of at least 30 mg/ml by in situ meglumine salt formation. CONCLUSION: Developing pre-clinical formulations to support in vivo studies requires a good understanding of the physicochemical properties of the drug candidates. Candidates with challenging pharmaceutic properties like the novel macrocycle molecule AZD5991, demand extensive characterization in its polymorph landscape, solubility profile and suitability evaluation of the excipients. Meglumine, a pH-adjusting and solubilizing agent, was found to be the best choice for formulating AZD5991 into an intravenous product to support preclinical studies.
Asunto(s)
Antineoplásicos , Meglumina , Fenómenos Químicos , Excipientes/química , Cloruro de Sodio , Solubilidad , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/antagonistas & inhibidoresRESUMEN
Myelodysplastic syndromes (MDSs) belong to a group of clonal bone marrow malignancies. In light of the emergence of new molecules, a significant contribution to the understanding of the pathogenesis of the disease is the study of the B-cell CLL/lymphoma 2 (BCL-2) and the programmed cell death receptor 1 (PD-1) protein and its ligands. BCL-2-family proteins are involved in the regulation of the intrinsic apoptosis pathway. Disruptions in their interactions promote the progression and resistance of MDSs. They have become an important target for specific drugs. Bone marrow cytoarchitecture may prove to be a predictor of response to its use. The challenge is the observed resistance to venetoclax, for which the MCL-1 protein may be largely responsible. Molecules with the potential to break the associated resistance include S63845, S64315, chidamide and arsenic trioxide (ATO). Despite promising in vitro studies, the role of PD-1/PD-L1 pathway inhibitors has not yet been established. Knockdown of the PD-L1 gene in preclinical studies was associated with increased levels of BCL-2 and MCL-1 in lymphocytes T, which could increase their survival and promote tumor apoptosis. A trial (NCT03969446) is currently underway to combine inhibitors from both groups.
Asunto(s)
Antígeno B7-H1 , Síndromes Mielodisplásicos , Receptor de Muerte Celular Programada 1 , Proteínas Proto-Oncogénicas c-bcl-2 , Humanos , Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Apoptosis , Antígeno B7-H1/metabolismo , Línea Celular Tumoral , Síndromes Mielodisplásicos/tratamiento farmacológico , Síndromes Mielodisplásicos/metabolismo , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/antagonistas & inhibidores , Proteínas Proto-Oncogénicas c-bcl-2/genética , Proteínas Proto-Oncogénicas c-bcl-2/metabolismoRESUMEN
Aurora kinases (AURKs) are mitotic kinases important for regulating cell cycle progression. Small-molecule inhibitors of AURK have shown promising antitumor effects in multiple cancers; however, the utility of these inhibitors as inducers of cancer cell death has thus far been limited. Here, we examined the role of the Bcl-2 family proteins in AURK inhibition-induced apoptosis in colon cancer cells. We found that alisertib and danusertib, two small-molecule inhibitors of AURK, are inefficient inducers of apoptosis in HCT116 and DLD-1 colon cancer cells, the survival of which requires at least one of the two antiapoptotic Bcl-2 family proteins, Bcl-xL and Mcl-1. We further identified Bcl-xL as a major suppressor of alisertib- or danusertib-induced apoptosis in HCT116 cells. We demonstrate that combination of a Bcl-2 homology (BH)3-mimetic inhibitor (ABT-737), a selective inhibitor of Bcl-xL, Bcl-2, and Bcl-w, with alisertib or danusertib potently induces apoptosis through the Bcl-2 family effector protein Bax. In addition, we identified Bid, Puma, and Noxa, three BH3-only proteins of the Bcl-2 family, as mediators of alisertib-ABT-737-induced apoptosis. We show while Noxa promotes apoptosis by constitutively sequestering Mcl-1, Puma becomes associated with Mcl-1 upon alisertib treatment. On the other hand, we found that alisertib treatment causes activation of caspase-2, which promotes apoptosis by cleaving Bid into truncated Bid, a suppressor of both Bcl-xL and Mcl-1. Together, these results define the Bcl-2 protein network critically involved in AURK inhibitor-induced apoptosis and suggest that BH3-mimetics targeting Bcl-xL may help overcome resistance to AURK inhibitors in cancer cells.
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Antineoplásicos , Apoptosis , Aurora Quinasas , Proteína bcl-X , Humanos , Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Apoptosis/genética , Proteínas Reguladoras de la Apoptosis/antagonistas & inhibidores , Proteínas Reguladoras de la Apoptosis/metabolismo , Aurora Quinasas/antagonistas & inhibidores , Proteína X Asociada a bcl-2/metabolismo , Proteína bcl-X/antagonistas & inhibidores , Proteína bcl-X/metabolismo , Línea Celular Tumoral , Neoplasias del Colon/tratamiento farmacológico , Neoplasias del Colon/fisiopatología , Activación Enzimática/efectos de los fármacos , Células HCT116 , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/antagonistas & inhibidores , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/antagonistas & inhibidores , Proteínas Proto-Oncogénicas c-bcl-2/metabolismoRESUMEN
MCL-1 is a member of the BCL-2 family of proteins that regulates the mitochondrial pathway of apoptosis. Overexpression of MCL-1 is associated with the development and progression of a range of human cancers, and is also responsible for the onset of resistance to conventional chemotherapies. Although several MCL-1 inhibitors have now advanced to clinical trials, recent suspensions and terminations reveal the urgency with which new inhibitor chemotypes must be discovered. Building on our previous studies of a chiral, isomeric lead, we report the discovery of a new chemotype to inhibit MCL-1: 1-sulfonylated 1,2,3,4-tetrahydroquinoline-6-carboxylic acid. The nature of the sulfonyl moiety contributed significantly to the resulting inhibitory ability. For example, transforming a phenylsulfonyl group into a 4-chloro-3,5-dimethylphenoxy)phenyl)sulfonyl moiety elicited more than a 73-fold enhancement in inhibiton of MCL-1, possibly through targeting the p2 pocket in the BH3-binding groove, and so it is anticipated that further structure-activity studies here will lead to continued improvements in binding. It should be underscored that this class of MCL-1 inhibitors is readily accessible in four simple steps, is achiral and offers many avenues for optimization, all factors that are welcomed in the search for safe and effective inhibitors of this driver of cancer cell survival.
Asunto(s)
Antineoplásicos , Ácidos Carboxílicos , Proteína 1 de la Secuencia de Leucemia de Células Mieloides , Quinolinas , Humanos , Antineoplásicos/farmacología , Apoptosis , Ácidos Carboxílicos/farmacología , Línea Celular Tumoral , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/antagonistas & inhibidores , Neoplasias , Quinolinas/farmacologíaRESUMEN
FMS-like tyrosine kinase 3 (FLT3)-internal tandem duplication (FLT3-ITD) mutations occur in about 25% of all acute myeloid leukemia (AML) patients and confer a poor prognosis. FLT3 inhibitors have been developed to treat patients with FLT3-mutated AML and have shown promise, though the acquisition of resistance occurs, highlighting the need for combination therapies to prolong the response to FLT3 inhibitors. In this study, we investigated the selective Mcl-1 inhibitor AZD5991 in combination with the FLT3 inhibitors gilteritinib and MRX-2843. The combinations synergistically induce apoptosis in AML cell lines and primary patient samples. The FLT3 inhibitors downregulate c-Myc transcripts through the suppression of the MEK/ERK and JAK2/STAT5 pathways, resulting in the decrease in c-Myc protein. This suppression of c-Myc plays an important role in the antileukemic activity of AZD5991. Interestingly, the suppression of c-Myc enhances AZD5991-inudced cytochrome c release and the subsequent induction of apoptosis. AZD5991 enhances the antileukemic activity of the FLT3 inhibitors gilteritinib and MRX-2843 against FLT3-mutated AML in vitro, warranting further development.
Asunto(s)
Leucemia Mieloide Aguda , Proteína 1 de la Secuencia de Leucemia de Células Mieloides , Inhibidores de Proteínas Quinasas , Tirosina Quinasa 3 Similar a fms , Humanos , Compuestos de Anilina , Tirosina Quinasa 3 Similar a fms/genética , Leucemia Mieloide Aguda/tratamiento farmacológico , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/metabolismo , Compuestos Macrocíclicos , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/farmacología , Inhibidores de Proteínas Quinasas/uso terapéutico , Pirazinas/farmacologíaRESUMEN
Diabetic cardiomyopathy (DCM) is initially characterized by early diastolic dysfunction, left ventricular remodeling, hypertrophy, and myocardial fibrosis, and it is eventually characterized by clinical heart failure. MicroRNAs (miRNAs), endogenous small noncoding RNAs, play significant roles in diabetes mellitus (DM). However, it is still largely unknown about the mechanism that links miRNAs and the development of DCM. Here, we aimed to elucidate the mechanism underlying the potential role of microRNA-340-5p in DCM in db/db mouse, which is a commonly used model of type 2 DM and diabetic complications that lead to heart failure. We first demonstrated that miR-340-5p expression was dramatically increased in heart tissues of mice and cardiomyocytes under diabetic conditions. Overexpression of miR-340-5p exacerbated DCM, which was reflected by extensive myocardial fibrosis and more serious dysfunction in db/db mice as represented by increased apoptotic cardiomyocytes, elevated ROS production, and impaired mitochondrial function. Inhibition of miR-340-5p by a tough decoy (TUD) vector was beneficial for preventing ROS production and apoptosis, thus rescuing diabetic cardiomyopathy. We identified myeloid cell leukemia 1 (Mcl-1) as a major target gene for miR-340-5p and showed that the inhibition of Mcl-1 was responsible for increased functional loss of mitochondria, oxidative stress, and cardiomyocyte apoptosis, thereby caused cardiac dysfunction in diabetic mice. In conclusion, our results showed that miR-340-5p plays a crucial role in the development of DCM and can be targeted for therapeutic intervention.
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MicroARNs/metabolismo , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/metabolismo , Estrés Oxidativo/genética , Animales , Antagomirs/metabolismo , Apoptosis , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Experimental/patología , Cardiomiopatías Diabéticas/etiología , Cardiomiopatías Diabéticas/patología , Modelos Animales de Enfermedad , Humanos , Ratones , Ratones Endogámicos C57BL , Ratones Obesos , MicroARNs/antagonistas & inhibidores , MicroARNs/genética , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/antagonistas & inhibidores , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/genética , Miocitos Cardíacos/citología , Miocitos Cardíacos/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/genética , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Interferencia de ARN , ARN Interferente Pequeño/metabolismo , Especies Reactivas de Oxígeno/metabolismoRESUMEN
Overexpression of antiapoptotic BCL2 family proteins occurs in various hematologic malignancies and contributes to tumorigenesis by inhibiting the apoptotic machinery of the cells. Antagonizing BH3 mimetics provide an option for medication, with venetoclax as the first drug applied for chronic lymphocytic leukemia and for acute myeloid leukemia. To find additional hematologic entities with ectopic expression of BCL2 family members, we performed expression screening of cell lines applying the LL-100 panel. Anaplastic large cell lymphoma (ALCL) and primary effusion lymphoma (PEL), 2/22 entities covered by this panel, stood out by high expression of MCL1 and low expression of BCL2. The MCL1 inhibitor AZD-5991 induced apoptosis in cell lines from both malignancies, suggesting that this BH3 mimetic might be efficient as drug for these diseases. The ALCL cell lines also expressed BCLXL and BCL2A1, both contributing to survival of the cells. The combination of specific BH3 mimetics yielded synergistic effects, pointing to a novel strategy for the treatment of ALCL. The PI3K/mTOR inhibitor BEZ-235 could also efficiently be applied in combination with AZD-5991, offering an alternative to avoid thrombocytopenia which is associated with the use of BCLXL inhibitors.
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Linfoma Anaplásico de Células Grandes/metabolismo , Linfoma de Efusión Primaria/metabolismo , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/metabolismo , Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Proteínas Reguladoras de la Apoptosis/metabolismo , Línea Celular Tumoral , Expresión Génica/genética , Regulación Neoplásica de la Expresión Génica/genética , Humanos , Imidazoles/farmacología , Linfoma Anaplásico de Células Grandes/tratamiento farmacológico , Linfoma Anaplásico de Células Grandes/genética , Linfoma de Efusión Primaria/tratamiento farmacológico , Linfoma de Efusión Primaria/genética , Compuestos Macrocíclicos/farmacología , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/antagonistas & inhibidores , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/genética , Fragmentos de Péptidos/metabolismo , Fragmentos de Péptidos/farmacología , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Proto-Oncogénicas/farmacología , Proteínas Proto-Oncogénicas c-bcl-2/efectos de los fármacos , Proteínas Proto-Oncogénicas c-bcl-2/genética , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Quinolinas/farmacologíaRESUMEN
Myeloid cell leukemia sequence 1 (MCL1), an antiapoptotic Bcell lymphoma 2 (BCL2) family molecule frequently amplified in various human cancer cells, is known to be critical for cancer cell survival. MCL1 has been recognized as a target molecule for cancer treatment. While various agents have emerged as potential MCL1 blockers, the present study presented acriflavine (ACF) as a novel MCL1 inhibitor in triplenegative breast cancer (TNBC). Further evaluation of its treatment potential on lung adenocarcinoma and glioblastoma multiforme (GBM) was also investigated. The anticancer effect of ACF on TNBC cells was demonstrated when MDAMB231 and HS578T cells were treated with ACF. ACF significantly induced typical intrinsic apoptosis in TNBCs in a dose and timedependent manner via MCL1 downregulation. MCL1 downregulation by ACF treatment was revealed at each phase of protein expression. Initially, transcriptional regulation via reverse transcriptionquantitative PCR was validated. Then, posttranslational regulation was explained by utilizing an inhibitor against protein biosynthesis and proteasome. Lastly, immunoprecipitation of ubiquitinated MCL1 confirmed the posttranslational downregulation of MCL1. In addition, the synergistic treatment efficacy of ACF with the wellknown MCL1 inhibitor ABT263 against the TNBC cells was explored [combination index (CI)<1]. Conjointly, the anticancer effect of ACF was assessed in GBM (U87, U251 and U343), and lung cancer (A549 and NCIH69) cell lines as well, using immunoblotting, cytotoxicity assay and FACS. The effect of the combination treatment using ACF and ABT263 was estimated in GBM (U87, U343 and U251), and nonsmall cell lung cancer (A549) cells likewise. The present study suggested a novel MCL1 inhibitory function of ACF and the synergistic antitumor effect with ABT263.