RESUMEN
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.
Asunto(s)
Antineoplásicos/química , Proteínas Nucleares/antagonistas & inhibidores , Pirroles/química , Factores de Transcripción/antagonistas & inhibidores , Animales , Antineoplásicos/síntesis química , Antineoplásicos/farmacocinética , Antineoplásicos/uso terapéutico , Sitios de Unión , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Cristalografía por Rayos X , Diseño de Fármacos , Semivida , Humanos , Ratones , Simulación de Dinámica Molecular , Mieloma Múltiple/tratamiento farmacológico , Proteínas Nucleares/metabolismo , Pirroles/síntesis química , Pirroles/farmacocinética , Pirroles/uso terapéutico , Relación Estructura-Actividad , Factores de Transcripción/metabolismo , Trasplante HeterólogoRESUMEN
The Bcl-2 family of proteins plays a critical role in controlling immune responses by regulating the expansion and contraction of activated lymphocyte clones by apoptosis. ABT-737, which was originally developed for oncology, is a potent inhibitor of Bcl-2, Bcl-x(L), and Bcl-w protein function. There is evidence that Bcl-2-associated dysregulation of lymphocyte apoptosis may contribute to the pathogenesis of autoimmunity and lead to the development of autoimmune diseases. In this study, we report that ABT-737 treatment resulted in potent inhibition of lymphocyte proliferation as measured by in vitro mitogenic or ex vivo Ag-specific stimulation. More importantly, ABT-737 significantly reduced disease severity in tissue-specific and systemic animal models of autoimmunity. Bcl-2 family antagonism by ABT-737 was efficacious in treating animal models of arthritis and lupus. Our results suggest that treatment with a Bcl-2 family antagonist represents a novel and potentially attractive therapeutic approach for the clinical treatment of autoimmunity.
Asunto(s)
Autoinmunidad/efectos de los fármacos , Compuestos de Bifenilo/farmacología , Nitrofenoles/farmacología , Proteínas Proto-Oncogénicas c-bcl-2/antagonistas & inhibidores , Sulfonamidas/farmacología , Animales , Presentación de Antígeno/efectos de los fármacos , Artritis Experimental/inducido químicamente , Artritis Experimental/inmunología , Artritis Experimental/patología , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Hemocianinas/inmunología , Humanos , Hipersensibilidad Tardía/inmunología , Interferón-alfa/farmacología , Nefritis Lúpica/inducido químicamente , Nefritis Lúpica/inmunología , Nefritis Lúpica/patología , Linfocitos/citología , Linfocitos/efectos de los fármacos , Linfocitos/inmunología , Ratones , Piperazinas/farmacología , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Especificidad por SustratoRESUMEN
Proteins in the Bcl-2 family are central regulators of programmed cell death, and members that inhibit apoptosis, such as Bcl-X(L) and Bcl-2, are overexpressed in many cancers and contribute to tumour initiation, progression and resistance to therapy. Bcl-X(L) expression correlates with chemo-resistance of tumour cell lines, and reductions in Bcl-2 increase sensitivity to anticancer drugs and enhance in vivo survival. The development of inhibitors of these proteins as potential anti-cancer therapeutics has been previously explored, but obtaining potent small-molecule inhibitors has proved difficult owing to the necessity of targeting a protein-protein interaction. Here, using nuclear magnetic resonance (NMR)-based screening, parallel synthesis and structure-based design, we have discovered ABT-737, a small-molecule inhibitor of the anti-apoptotic proteins Bcl-2, Bcl-X(L) and Bcl-w, with an affinity two to three orders of magnitude more potent than previously reported compounds. Mechanistic studies reveal that ABT-737 does not directly initiate the apoptotic process, but enhances the effects of death signals, displaying synergistic cytotoxicity with chemotherapeutics and radiation. ABT-737 exhibits single-agent-mechanism-based killing of cells from lymphoma and small-cell lung carcinoma lines, as well as primary patient-derived cells, and in animal models, ABT-737 improves survival, causes regression of established tumours, and produces cures in a high percentage of the mice.
Asunto(s)
Antineoplásicos/uso terapéutico , Compuestos de Bifenilo/farmacología , Compuestos de Bifenilo/uso terapéutico , Neoplasias/tratamiento farmacológico , Neoplasias/patología , Proteínas Proto-Oncogénicas c-bcl-2/antagonistas & inhibidores , Proteínas Proto-Oncogénicas c-bcl-2/clasificación , Animales , Antineoplásicos/síntesis química , Antineoplásicos/química , Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Compuestos de Bifenilo/síntesis química , Compuestos de Bifenilo/química , Carcinoma de Células Pequeñas/tratamiento farmacológico , Carcinoma de Células Pequeñas/patología , Línea Celular Tumoral , Citocromos c/metabolismo , Modelos Animales de Enfermedad , Sinergismo Farmacológico , Humanos , Linfoma/tratamiento farmacológico , Linfoma/patología , Espectroscopía de Resonancia Magnética , Ratones , Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , Modelos Moleculares , Nitrofenoles , Paclitaxel/farmacología , Piperazinas , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Relación Estructura-Actividad , Sulfonamidas , Tasa de SupervivenciaRESUMEN
BACKGROUND: Proteins of the BCL-2 family regulate clonal selection and survival of lymphocytes, and are frequently overexpressed in lymphomas. Navitoclax is a targeted high-affinity small molecule that inhibits the anti-apoptotic activity of BCL-2 and BCL-XL. We aimed to assess the safety and antitumour activity of navitoclax in patients with lymphoid tumours, and establish the drug's pharmacokinetic and pharmacodynamic profiles. METHODS: In this phase 1 dose-escalation study, patients (aged ≥18 years) with relapsed or refractory lymphoid malignancies were enrolled and treated at seven sites in the USA between November, 2006, and November, 2009. A modified Fibonacci 3+3 design was used to assign patients to receive oral navitoclax once daily by one of two dosing schedules: intermittently for the first 14 days of a 21-day cycle (14/21) at doses of 10, 20, 40, 80, 110, 160, 225, 315, or 440 mg/day; or continuously for 21 days of a 21-day cycle (21/21) at doses of 200, 275, 325, or 425 mg/day. Study endpoints were safety, maximum tolerated dose, pharmacokinetic profile, pharmacodynamic effects on platelets and T cells, and antitumour activity. This trial is registered with ClinicalTrials.gov, number NCT00406809. FINDINGS: 55 patients were enrolled (median age 59 years, IQR 51-67), 38 to receive the 14/21 dosing schedule, and 17 to receive the 21/21 dosing schedule. Common toxic effects included grade 1 or 2 anaemia (41 patients), infection (39), diarrhoea (31), nausea (29), and fatigue (21); and grade 3 or 4 thrombocytopenia (29), lymphocytopenia (18), and neutropenia (18). On the intermittent 14/21 schedule, dose-limiting toxic effects were hospital admissions for bronchitis (one) and pleural effusion (one), grade 3 increase in aminotransferases (one), grade 4 thrombocytopenia (one), and grade 3 cardiac arrhythmia (one). To reduce platelet nadir associated with intermittent 14/21 dosing, we assessed a 150 mg/day lead-in dose followed by a continuous 21/21 dosing schedule. On the 21/21 dosing schedule, two patients did not complete the first cycle and were excluded from assessment of dose-limiting toxic effects; dose-limiting toxic effects were grade 4 thrombocytopenia (one), grade 3 increase in aminotransferases (one), and grade 3 gastrointestinal bleeding (one). Navitoclax showed a pharmacodynamic effect on circulating platelets and T cells. Clinical responses occurred across the range of doses and in several tumour types. Ten of 46 patients with assessable disease had a partial response, and these responders had median progression-free survival of 455 days (IQR 40-218). INTERPRETATION: Navitoclax has a novel mechanism of peripheral thrombocytopenia and T-cell lymphopenia, attributable to high-affinity inhibition of BCL-XL and BCL-2, respectively. On the basis of these findings, a 150 mg 7-day lead-in dose followed by a 325 mg dose administered on a continuous 21/21 dosing schedule was selected for phase 2 study. FUNDING: Abbott Laboratories, Genentech, and National Cancer Institute, National Institutes of Health.
Asunto(s)
Compuestos de Anilina/farmacocinética , Compuestos de Anilina/uso terapéutico , Antineoplásicos/farmacocinética , Antineoplásicos/uso terapéutico , Sistemas de Liberación de Medicamentos , Linfoma/tratamiento farmacológico , Proteínas Proto-Oncogénicas c-bcl-2/antagonistas & inhibidores , Sulfonamidas/farmacocinética , Sulfonamidas/uso terapéutico , Anciano , Compuestos de Anilina/efectos adversos , Compuestos de Anilina/farmacología , Antineoplásicos/efectos adversos , Antineoplásicos/farmacología , Plaquetas/efectos de los fármacos , Formas de Dosificación , Humanos , Dosis Máxima Tolerada , Persona de Mediana Edad , Sulfonamidas/efectos adversos , Sulfonamidas/farmacología , Análisis de Supervivencia , Linfocitos T/efectos de los fármacosRESUMEN
BCL-XL, an antiapoptotic member of the BCL-2 family of proteins, drives tumor survival and maintenance and thus represents a key target for cancer treatment. Herein we report the rational design of a novel series of selective BCL-XL inhibitors exemplified by A-1293102. This molecule contains structural elements of selective BCL-XL inhibitor A-1155463 and the dual BCL-XL/BCL-2 inhibitors ABT-737 and navitoclax, while representing a distinct pharmacophore as assessed by an objective cheminformatic evaluation. A-1293102 exhibited picomolar binding affinity to BCL-XL and both efficiently and selectively killed BCL-XL-dependent tumor cells. X-ray crystallographic analysis demonstrated a key hydrogen bonding network in the P2 binding pocket of BCL-XL, while the bent-back moiety achieved efficient occupancy of the P4 pocket in a manner similar to that of navitoclax. A-1293102 represents one of the few distinct structural series of selective BCL-XL inhibitors, and thus serves as a useful tool for biological studies as well as a lead compound for further optimization.
RESUMEN
Since gaining approval for the treatment of chronic lymphocytic leukemia (CLL), the BCL-2 inhibitor venetoclax has transformed the treatment of this and other blood-related cancers. Reflecting the large and hydrophobic BH3-binding groove within BCL-2, venetoclax has significantly higher molecular weight and lipophilicity than most orally administered drugs, along with negligible water solubility. Although a technology-enabled formulation successfully achieves oral absorption in humans, venetoclax tablets have limited drug loading and therefore can present a substantial pill burden for patients in high-dose indications. We therefore generated a phosphate prodrug (3, ABBV-167) that confers significantly increased water solubility to venetoclax and, upon oral administration to healthy volunteers either as a solution or high drug-load immediate release tablet, extensively converts to the parent drug. Additionally, ABBV-167 demonstrated a lower food effect with respect to venetoclax tablets. These data indicate that beyond-rule-of-5 molecules can be successfully delivered to humans via a solubility-enhancing prodrug moiety to afford robust exposures of the parent drug following oral dosing.
Asunto(s)
Compuestos Bicíclicos Heterocíclicos con Puentes/uso terapéutico , Profármacos/uso terapéutico , Sulfonamidas/uso terapéutico , Compuestos Bicíclicos Heterocíclicos con Puentes/farmacología , Línea Celular Tumoral , Estudios Cruzados , Femenino , Voluntarios Sanos , Humanos , Profármacos/farmacología , Sulfonamidas/farmacologíaRESUMEN
We describe the development of a novel series of N-aryl-benzimidazolone HSP90 inhibitors (9) targeting the N-terminal ATP-ase site. SAR development was influenced by structure-based design based around X-ray structures of ligand bound HSP90 complexes. Lead compounds exhibited high binding affinities, ATP-ase inhibition and cellular client protein degradation.
Asunto(s)
Bencimidazoles/química , Proteínas HSP90 de Choque Térmico/antagonistas & inhibidores , Bencimidazoles/síntesis química , Bencimidazoles/farmacología , Sitios de Unión , Línea Celular Tumoral , Cristalografía por Rayos X , Proteínas HSP90 de Choque Térmico/metabolismo , Humanos , Estructura Terciaria de Proteína , Relación Estructura-ActividadRESUMEN
The Bcl-2 family of proteins plays a major role in the regulation of apoptosis, or programmed cell death. Overexpression of the anti-apoptotic members of this family (Bcl-2, Bcl-x(L), and Mcl-1) can render cancer cells resistant to chemotherapeutic agents and therefore these proteins are important targets for the development of new anti-cancer agents. Here we describe the discovery of a potent, highly selective, Bcl-2 inhibitor using SAR by NMR and structure-based drug design which could serve as a starting point for the development of a Bcl-2 selective anti-cancer agent. Such an agent would potentially overcome the Bcl-x(L) mediated thrombocytopenia observed with ABT-263.
Asunto(s)
Espectroscopía de Resonancia Magnética/métodos , Proteínas Proto-Oncogénicas c-bcl-2/antagonistas & inhibidores , Modelos Moleculares , Relación Estructura-ActividadRESUMEN
Herein we describe the discovery of A-1331852, a first-in-class orally active BCL-XL inhibitor that selectively and potently induces apoptosis in BCL-XL-dependent tumor cells. This molecule was generated by re-engineering our previously reported BCL-XL inhibitor A-1155463 using structure-based drug design. Key design elements included rigidification of the A-1155463 pharmacophore and introduction of sp3-rich moieties capable of generating highly productive interactions within the key P4 pocket of BCL-XL. A-1331852 has since been used as a critical tool molecule for further exploring BCL-2 family protein biology, while also representing an attractive entry into a drug discovery program.
RESUMEN
PURPOSE: The purpose of this study was to characterize the activity of the Bcl-2 protein family inhibitor ABT-263 in a panel of small cell lung cancer (SCLC) xenograft models. EXPERIMENTAL DESIGN: A panel of 11 SCLC xenograft models was established to evaluate the efficacy of ABT-263. Single agent activity was examined on a continuous dosing schedule in each of these models. The H146 model was used to further evaluate dose and schedule, comparison to standard cytotoxic agents, and induction of apoptosis. RESULTS: ABT-263 exhibited a range of antitumor activity, leading to complete tumor regression in several models. Significant regressions of tumors as large as 1 cc were also observed. The efficacy of ABT-263 was also quite durable; in several cases, minimal tumor regrowth was noted several weeks after the cessation of treatment. Antitumor effects were equal or superior to that of several clinically approved cytotoxic agents. Regression of large established tumors was observed through several cycles of therapy and efficacy was retained in a Pgp-1 overexpressing line. Significant efficacy was observed on several dose and therapeutic schedules and was associated with significant induction of apoptosis. CONCLUSIONS: ABT-263 is a potent, orally bioavailable inhibitor of Bcl-2 family proteins that has recently entered clinical trials. The efficacy data reported here suggest that SCLC is a promising area of clinical investigation with this agent.
Asunto(s)
Compuestos de Anilina/administración & dosificación , Antineoplásicos/administración & dosificación , Carcinoma de Células Pequeñas/tratamiento farmacológico , Neoplasias Pulmonares/tratamiento farmacológico , Sulfonamidas/administración & dosificación , Animales , Relación Dosis-Respuesta a Droga , Humanos , Ratones , Ratones Desnudos , Proteínas Proto-Oncogénicas c-bcl-2/antagonistas & inhibidores , Proteínas Proto-Oncogénicas c-bcl-2/efectos de los fármacos , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
ABT-263 is a potent, orally bioavailable inhibitor of the antiapoptotic Bcl-2 family members Bcl-2, Bcl-x(L), and Bcl-w, which is currently in phase I clinical trials. Previous work has shown that this compound has low nanomolar cell-killing activity in a variety of lymphoma and leukemia cell lines, many of which overexpress Bcl-2 through a variety of mechanisms. Rapamycin is a macrolide antibiotic that inhibits the mammalian target of rapamycin complex, leading to cell cycle arrest and inhibition of protein translation. Rapamycin (and its analogues) has shown activity in a variety of tumor cell lines primarily through induction of cell cycle arrest. Activity has also been shown clinically in mantle cell lymphoma and advanced renal cell carcinoma. Here, we show that treatment of the follicular lymphoma lines DoHH-2 and SuDHL-4 with 100 nmol/L rapamycin induces substantial G(0)-G(1) arrest. Addition of as little as 39 nmol/L ABT-263 to the rapamycin regimen induced a 3-fold increase in sub-G(0) cells. Combination of these agents also led to a significant increase in Annexin V staining over ABT-263 alone. In xenograft models of these tumors, rapamycin induced a largely cytostatic response in the DoHH-2 and SuDHL-4 models. Coadministration with ABT-263 induced significant tumor regression, with DoHH-2 and SuDHL-4 tumors showing 100% overall response rates. Apoptosis in these tumors was significantly enhanced by combination therapy as measured by staining with an antibody specific for cleaved caspase-3. These data suggest that combination of ABT-263 and rapamycin or its analogues represents a promising therapeutic strategy for the treatment of lymphoma.
Asunto(s)
Compuestos de Anilina/farmacología , Antineoplásicos/farmacología , Linfoma de Células B Grandes Difuso/patología , Sirolimus/farmacología , Sulfonamidas/farmacología , Animales , Ciclo Celular/efectos de los fármacos , Muerte Celular/efectos de los fármacos , Línea Celular Tumoral , Sinergismo Farmacológico , Humanos , Inmunohistoquímica , Linfoma de Células B Grandes Difuso/terapia , Ratones , Ratones SCID , Inducción de Remisión , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Overexpression of the antiapototic proteins Bcl-2 and Bcl-xL provides a common mechanism through which cancer cells gain a survival advantage and become resistant to conventional chemotherapy. Inhibition of these prosurvival proteins is an attractive strategy for cancer therapy. We recently described the discovery of a selective Bcl-xL antagonist that potentiates the antitumor activity of chemotherapy and radiation. Here we describe the use of structure-guided design to exploit a deep hydrophobic binding pocket on the surface of these proteins to develop the first dual, subnanomolar inhibitors of Bcl-xL and Bcl-2. This study culminated in the identification of 2, which exhibited EC50 values of 8 nM and 30 nM in Bcl-2 and Bcl-xL dependent cells, respectively. Compound 2 demonstrated single agent efficacy against human follicular lymphoma cell lines that overexpress Bcl-2, and efficacy in a murine xenograft model of lymphoma when given both as a single agent and in combination with etoposide.
Asunto(s)
Antineoplásicos/síntesis química , Compuestos de Bifenilo/síntesis química , Nitrofenoles/síntesis química , Proteínas Proto-Oncogénicas c-bcl-2/antagonistas & inhibidores , Sulfonamidas/síntesis química , Animales , Antineoplásicos/química , Antineoplásicos/farmacología , Compuestos de Bifenilo/química , Compuestos de Bifenilo/farmacología , Línea Celular Tumoral , Ensayos de Selección de Medicamentos Antitumorales , Humanos , Linfoma , Ratones , Ratones SCID , Modelos Moleculares , Nitrofenoles/química , Nitrofenoles/farmacología , Piperazinas/síntesis química , Piperazinas/química , Piperazinas/farmacología , Proteínas Proto-Oncogénicas c-bcl-2/química , Relación Estructura-Actividad , Sulfonamidas/química , Sulfonamidas/farmacología , Trasplante Heterólogo , Proteína bcl-X/antagonistas & inhibidores , Proteína bcl-X/químicaRESUMEN
ABBV-075 is a potent and selective BET family bromodomain inhibitor that recently entered phase I clinical trials. Comprehensive preclinical characterization of ABBV-075 demonstrated broad activity across cell lines and tumor models, representing a variety of hematologic malignancies and solid tumor indications. In most cancer cell lines derived from solid tumors, ABBV-075 triggers prominent G1 cell-cycle arrest without extensive apoptosis. In this study, we show that ABBV-075 efficiently triggers apoptosis in acute myeloid leukemia (AML), non-Hodgkin lymphoma, and multiple myeloma cells. Apoptosis induced by ABBV-075 was mediated in part by modulation of the intrinsic apoptotic pathway, exhibiting synergy with the BCL-2 inhibitor venetoclax in preclinical models of AML. In germinal center diffuse large B-cell lymphoma, BCL-2 levels or venetoclax sensitivity predicted the apoptotic response to ABBV-075 treatment. In vivo combination studies uncovered surprising benefits of low doses of ABBV-075 coupled with bortezomib and azacitidine treatment, despite the lack of in vitro synergy between ABBV-075 and these agents. The in vitro/in vivo activities of ABBV-075 described here may serve as a useful reference to guide the development of ABBV-075 and other BET family inhibitors for cancer therapy. Cancer Res; 77(11); 2976-89. ©2017 AACR.
Asunto(s)
Antagonistas de Andrógenos/uso terapéutico , Piridonas/uso terapéutico , Sulfonamidas/uso terapéutico , Antagonistas de Andrógenos/farmacología , Apoptosis , Línea Celular Tumoral , Sinergismo Farmacológico , Humanos , Piridonas/farmacología , Sulfonamidas/farmacología , TransfecciónRESUMEN
Members of the BET family of bromodomain containing proteins have been identified as potential targets for blocking proliferation in a variety of cancer cell lines. A two-dimensional NMR fragment screen for binders to the bromodomains of BRD4 identified a phenylpyridazinone fragment with a weak binding affinity (1, Ki = 160 µM). SAR investigation of fragment 1, aided by X-ray structure-based design, enabled the synthesis of potent pyridone and macrocyclic pyridone inhibitors exhibiting single digit nanomolar potency in both biochemical and cell based assays. Advanced analogs in these series exhibited high oral exposures in rodent PK studies and demonstrated significant tumor growth inhibition efficacy in mouse flank xenograft models.
Asunto(s)
Compuestos Macrocíclicos/química , Compuestos Macrocíclicos/farmacología , Piridonas/química , Piridonas/farmacología , Animales , Cristalografía por Rayos X , Descubrimiento de Drogas , Compuestos Macrocíclicos/farmacocinética , Estructura Molecular , Piridonas/farmacocinética , Ratas , Relación Estructura-ActividadRESUMEN
The development of bromodomain and extraterminal domain (BET) bromodomain inhibitors and their examination in clinical studies, particularly in oncology settings, has garnered substantial recent interest. An effort to generate novel BET bromodomain inhibitors with excellent potency and drug metabolism and pharmacokinetics (DMPK) properties was initiated based upon elaboration of a simple pyridone core. Efforts to develop a bidentate interaction with a critical asparagine residue resulted in the incorporation of a pyrrolopyridone core, which improved potency by 9-19-fold. Additional structure-activity relationship (SAR) efforts aimed both at increasing potency and improving pharmacokinetic properties led to the discovery of the clinical candidate 63 (ABBV-075/mivebresib), which demonstrates excellent potency in biochemical and cellular assays, advantageous exposures and half-life both in animal models and in humans, and in vivo efficacy in mouse models of cancer progression and inflammation.
Asunto(s)
Descubrimiento de Drogas , Proteínas/antagonistas & inhibidores , Piridonas/farmacología , Sulfonamidas/farmacología , Animales , Línea Celular Tumoral , Cromatografía Líquida de Alta Presión , Transferencia Resonante de Energía de Fluorescencia , Semivida , Humanos , Espectrometría de Masas , Ratones , Espectroscopía de Protones por Resonancia Magnética , Piridonas/química , Piridonas/farmacocinética , Relación Estructura-Actividad , Sulfonamidas/química , Sulfonamidas/farmacocinéticaRESUMEN
The antiapoptotic proteins Bcl-x(L) and Bcl-2 play key roles in the maintenance of normal cellular homeostasis. However, their overexpression can lead to oncogenic transformation and is responsible for drug resistance in certain types of cancer. This makes Bcl-x(L) and Bcl-2 attractive targets for the development of potential anticancer agents. Here we describe the structure-based discovery of a potent Bcl-x(L) inhibitor directed at a hydrophobic groove on the surface of the protein. This groove represents the binding site for BH3 peptides from proapoptotic Bcl-2 family members such as Bak and Bad. Application of NMR-based screening yielded an initial biaryl acid with an affinity (K(d)) of approximately 300 microM for the protein. Following the classical "SAR by NMR" approach, a second-site ligand was identified that bound proximal to the first-site ligand in the hydrophobic groove. From NMR-based structural studies and parallel synthesis, a potent ligand was obtained, which binds to Bcl-x(L) with an inhibition constant (K(i)) of 36 +/- 2 nM.
Asunto(s)
Compuestos de Anilina/síntesis química , Modelos Moleculares , Sulfonamidas/síntesis química , Proteína bcl-X/antagonistas & inhibidores , Compuestos de Anilina/química , Sitios de Unión , Interacciones Hidrofóbicas e Hidrofílicas , Ligandos , Espectroscopía de Resonancia Magnética , Unión Proteica , Solubilidad , Relación Estructura-Actividad , Sulfonamidas/química , Proteína bcl-X/químicaRESUMEN
Development of a rationally designed potentiator of cancer chemotherapy, via inhibition of Bcl-X(L) function, is described. Lead compounds generated by NMR screening and directed parallel synthesis displayed sub-microM binding but were strongly deactivated in the presence of serum. The dominant component of serum deactivation was identified as domain III of human serum albumin (HSA); NMR solution structures of inhibitors bound to both Bcl-X(L) and HSA domain III indicated two potential optimization sites for separation of affinities. Modifications at both sites resulted in compounds with improved Bcl-X(L) binding and greatly increased activity in the presence of human serum, culminating in 73R, which bound to Bcl-X(L) with a K(i) of 0.8 nM. In a cellular assay 73R reversed the protection afforded by Bcl-X(L) overexpression against cytokine deprivation in FL5.12 cells with an EC(50) of 0.47 microM. 73R showed little effect on the viability of the human non small cell lung cancer cell line A549. However, consistent with the proposed mechanism, 73R potentiated the activity of paclitaxel and UV irradiation in vitro and potentiated the antitumor efficacy of paclitaxel in a mouse xenograft model.
Asunto(s)
Compuestos de Anilina/síntesis química , Antineoplásicos/síntesis química , Piperidinas/síntesis química , Sulfonamidas/síntesis química , Proteína bcl-X/antagonistas & inhibidores , Compuestos de Anilina/química , Compuestos de Anilina/farmacología , Animales , Antineoplásicos/química , Antineoplásicos/farmacología , Disponibilidad Biológica , Línea Celular Tumoral , Ensayos de Selección de Medicamentos Antitumorales , Sinergismo Farmacológico , Polarización de Fluorescencia , Humanos , Espectroscopía de Resonancia Magnética , Ratones , Ratones SCID , Paclitaxel/farmacología , Piperidinas/química , Piperidinas/farmacología , Unión Proteica , Estructura Terciaria de Proteína , Suero , Albúmina Sérica/química , Estereoisomerismo , Sulfonamidas/química , Sulfonamidas/farmacología , Trasplante Heterólogo , Rayos UltravioletaRESUMEN
Glucocorticoids (GCs) are commonly used to treat inflammatory disease; unfortunately, the long-term use of these steroids leads to a large number of debilitating side effects. The antiinflammatory effects of GCs are a result of GC receptor (GR)-mediated inhibition of expression of proinflammatory genes as well as GR-mediated activation of antiinflammatory genes. Similarly, side effects are most likely due to both activated and repressed GR target genes in affected tissues. An as yet unachieved pharmaceutical goal is the development of a compound capable of separating detrimental side effects from antiinflammatory activity. We describe the discovery and characterization of AL-438, a GR ligand that exhibits an altered gene regulation profile, able to repress and activate only a subset of the genes normally regulated by GCs. When tested in vivo, AL-438 retains full antiinflammatory efficacy and potency comparable to steroids but its negative effects on bone metabolism and glucose control are reduced at equivalently antiinflammatory doses. The mechanism underlying this selective in vitro and in vivo activity may be the result of differential cofactor recruitment in response to ligand. AL-438 reduces the interaction between GR and peroxisomal proliferator-activated receptor gamma coactivator-1, a cofactor critical for steroid-mediated glucose up-regulation, while maintaining normal interactions with GR-interacting protein 1. This compound serves as a prototype for a unique, nonsteroidal alternative to conventional GCs in treating inflammatory disease.
Asunto(s)
Antiinflamatorios no Esteroideos/farmacología , Benzopiranos/farmacología , Quinolinas/farmacología , Animales , Antiinflamatorios no Esteroideos/efectos adversos , Antiinflamatorios no Esteroideos/metabolismo , Artritis Experimental/tratamiento farmacológico , Benzopiranos/efectos adversos , Benzopiranos/metabolismo , Huesos/efectos de los fármacos , Células Cultivadas , Evaluación Preclínica de Medicamentos/métodos , Edema/tratamiento farmacológico , Regulación de la Expresión Génica/efectos de los fármacos , Glucocorticoides/farmacología , Humanos , Inflamación/tratamiento farmacológico , Masculino , Coactivador 2 del Receptor Nuclear , Prednisolona/metabolismo , Prednisolona/farmacología , Quinolinas/efectos adversos , Quinolinas/metabolismo , Ratas , Ratas Sprague-Dawley , Tibia/anatomía & histología , Tibia/efectos de los fármacos , Factores de Transcripción/efectos de los fármacos , Factores de Transcripción/genética , Factores de Transcripción/metabolismoRESUMEN
Hyperexpression of antiapoptotic BCL-2 family proteins allows cells to survive despite the receipt of signals that would ordinarily induce their deletion, a facet frequently exploited by tumors. Tumors addicted to the BCL-2 family proteins for survival are now being targeted therapeutically. For example, navitoclax, a BCL-2/BCL-XL/BCL-W inhibitor, is currently in phase I/II clinical trials in numerous malignancies. However, the related family member, MCL-1, limits the efficacy of navitoclax and other chemotherapeutic agents. In the present study, we identify breast cancer cell lines that depend upon MCL-1 for survival and subsequently determine the mechanism of apoptosis mediated by the MCL-1 selective inhibitor A-1210477. We demonstrate that apoptosis resulting from a loss in MCL-1 function requires expression of the proapoptotic protein BAK. However, expression of BCL-XL can limit apoptosis resulting from loss in MCL-1 function through sequestration of free BIM. Finally, we demonstrate substantial synergy between navitoclax and MCL-1 siRNA, the direct MCL-1 inhibitor A-1210477, or the indirect MCL-1 inhibitor flavopiridol, highlighting the therapeutic potential for inhibiting BCL-XL and MCL-1 in breast cancer.
Asunto(s)
Neoplasias de la Mama/metabolismo , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/metabolismo , Compuestos de Anilina/farmacología , Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Apoptosis/genética , Neoplasias de la Mama/genética , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Femenino , Expresión Génica , Humanos , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/antagonistas & inhibidores , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/genética , Proteínas Proto-Oncogénicas c-bcl-2/genética , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Interferencia de ARN , ARN Interferente Pequeño/genética , Sulfonamidas/farmacología , Proteína Destructora del Antagonista Homólogo bcl-2/genética , Proteína Destructora del Antagonista Homólogo bcl-2/metabolismo , Proteína bcl-X/genética , Proteína bcl-X/metabolismoRESUMEN
The BCL-2/BCL-XL/BCL-W inhibitor ABT-263 (navitoclax) has shown promising clinical activity in lymphoid malignancies such as chronic lymphocytic leukemia. However, its efficacy in these settings is limited by thrombocytopenia caused by BCL-XL inhibition. This prompted the generation of the BCL-2-selective inhibitor venetoclax (ABT-199/GDC-0199), which demonstrates robust activity in these cancers but spares platelets. Navitoclax has also been shown to enhance the efficacy of docetaxel in preclinical models of solid tumors, but clinical use of this combination has been limited by neutropenia. We used venetoclax and the BCL-XL-selective inhibitors A-1155463 and A-1331852 to assess the relative contributions of inhibiting BCL-2 or BCL-XL to the efficacy and toxicity of the navitoclax-docetaxel combination. Selective BCL-2 inhibition suppressed granulopoiesis in vitro and in vivo, potentially accounting for the exacerbated neutropenia observed when navitoclax was combined with docetaxel clinically. By contrast, selectively inhibiting BCL-XL did not suppress granulopoiesis but was highly efficacious in combination with docetaxel when tested against a range of solid tumors. Therefore, BCL-XL-selective inhibitors have the potential to enhance the efficacy of docetaxel in solid tumors and avoid the exacerbation of neutropenia observed with navitoclax. These studies demonstrate the translational utility of this toolkit of selective BCL-2 family inhibitors and highlight their potential as improved cancer therapeutics.