RESUMO
ABSTRACT: Glucocorticoids are key components of the standard-of-care treatment regimens for B-cell malignancy. However, systemic glucocorticoid treatment is associated with several adverse events. ABBV-319 is a CD19-targeting antibody-drug conjugate engineered to reduce glucocorticoid-associated toxicities while possessing 3 distinct mechanisms of action (MOA) to increase therapeutic efficacy: (1) antibody-mediated delivery of a glucocorticoid receptor modulator (GRM) payload to activate apoptosis, (2) inhibition of CD19 signaling, and (3) enhanced fragment crystallizable (Fc)-mediated effector function via afucosylation of the antibody backbone. ABBV-319 elicited potent GRM-driven antitumor activity against multiple malignant B-cell lines in vitro, as well as in cell line-derived xenografts and patient-derived xenografts (PDXs) in vivo. Remarkably, a single dose of ABBV-319 induced sustained tumor regression and enhanced antitumor activity compared with repeated dosing of systemic prednisolone at the maximum tolerated dose in mice. The unconjugated CD19 monoclonal antibody (mAb) also displayed antiproliferative activity in a subset of B-cell lymphoma cell lines through the inhibition of phosphoinositide 3-kinase signaling. Moreover, afucosylation of CD19 mAb enhanced Fc-mediated antibody-dependent cellular cytotoxicity. Notably, ABBV-319 displayed superior efficacy compared with afucosylated CD19 mAb in human CD34+ peripheral blood mononuclear cell-engrafted NSG-Tg(Hu-IL15) transgenic mice, demonstrating enhanced antitumor activity when multiple MOAs are enabled. ABBV-319 also showed durable antitumor activity across multiple B-cell lymphoma PDX models, including nongerminal center B-cell diffuse large B-cell lymphoma and relapsed lymphoma after R-CHOP treatment. Collectively, these data support the ongoing evaluation of ABBV-319 in a phase 1 clinical trial.
Assuntos
Antígenos CD19 , Imunoconjugados , Receptores de Glucocorticoides , Ensaios Antitumorais Modelo de Xenoenxerto , Humanos , Animais , Antígenos CD19/imunologia , Camundongos , Imunoconjugados/farmacologia , Imunoconjugados/uso terapêutico , Receptores de Glucocorticoides/antagonistas & inibidores , Anticorpos Monoclonais Humanizados/farmacologia , Anticorpos Monoclonais Humanizados/uso terapêutico , Anticorpos Monoclonais Humanizados/administração & dosagem , Linfoma de Células B/tratamento farmacológico , Linfoma de Células B/patologia , Linhagem Celular Tumoral , Camundongos SCID , Feminino , Maitansina/análogos & derivadosRESUMO
We performed a high-throughput phenotypic whole cell screen of Mycobacterium tuberculosis against a diverse chemical library of approximately 100,000 compounds from the AbbVie corporate collection and identified 24 chemotypes with anti-tubercular activity. We selected two series for further exploration and conducted structure-activity relationship studies with new analogs for the 4-phenyl piperidines (4PP) and phenylcyclobutane carboxamides (PCB). Strains with mutations in MmpL3 demonstrated resistance to both compound series. We isolated resistant mutants for the two series and found mutations in MmpL3. These data suggest that MmpL3 is the target, or mechanism of resistance for both series.
Assuntos
Mycobacterium tuberculosis , Antituberculosos/química , Antituberculosos/farmacologia , Proteínas de Bactérias/metabolismo , Ensaios de Triagem em Larga Escala , Proteínas de Membrana Transportadoras/genética , Testes de Sensibilidade Microbiana , Mycobacterium tuberculosis/metabolismoRESUMO
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.
RESUMO
Antibody-drug conjugates (ADCs) are an increasingly prevalent drug class utilized as chemotherapeutic agents. The complexity of ADCs, including their large size, array of drug conjugation sites, and heterogeneous compositions containing from zero to several payloads, demands the use of advanced analytical characterization methods. Tandem mass spectrometry (MS/MS) strategies, including a variety of bottom-up, middle-down, and even top-down approaches, frequently applied for the analysis of antibodies are increasingly being adapted for antibody-drug conjugates. Middle-down tandem mass spectrometry, often focusing on the analysis of â¼25 kDa protein subunits, offers the potential for complete sequence confirmation as well as the identification of multiple conjugation states. While middle-down studies have been extensively developed for monoclonal antibodies, middle-down characterization of ADCs has been limited by the high complexity of the drug molecules. This study seeks to bridge the gap by utilizing a combination of 193 nm ultraviolet photodissociation (UVPD), electron-transfer dissociation (ETD), and electron-transfer/higher-energy collision dissociation (EThcD). The compilation of these MS/MS methods leads to high sequence coverages of 60-80% for each subunit of the ADC. Moreover, the combined fragmentation patterns provide sufficient information to allow confirmation of both the sequence of the complementarity-determining regions and the payload conjugation sites.
Assuntos
Imunoconjugados/química , Espectrometria de Massas em Tandem/métodos , Sequência de Aminoácidos , Antineoplásicos/administração & dosagem , Antineoplásicos/química , Fracionamento Químico , Subunidades ProteicasRESUMO
The physical stability of antibody drug conjugates is dictated by the properties of the antibody, linker-drug, and conjugation site. Two linker-drugs were chosen that are different in terms of hydrophobicity and polar surface area to evaluate the effect of linker-drug properties on antibody-drug conjugate (ADC) behavior. Site-specific and non-site-specific conjugation was used to investigate the role of conjugation site in conformational and colloidal stability. Finally, 2 antibodies were selected to determine if the observed results were antibody-specific. The conformational stability is affected, with the highest degree of destabilization observed when conjugation results in the removal of interchain disulfide bonds. Although conformational destabilization occurred in the domain in which conjugation occurred and domains distinct from the conjugation site, no correlation could be drawn between linker-drug properties and conformational stability. Evaluation of aggregation by size exclusion HPLC confirmed a relationship between linker-drug hydrophobicity and aggregation propensity under thermal stress in all ADCs tested. The extent of aggregation was far greater in the conjugates generated with a more hydrophobic antibody, illustrating that the properties of both the antibody and linker-drug contribute to aggregation. These studies emphasize that the distinct properties of the molecule as a whole warrant a case-by-case evaluation of each ADC.
Assuntos
Vacinas Anticâncer , Imunoconjugados , Preparações Farmacêuticas , Cromatografia Líquida de Alta Pressão , Interações Hidrofóbicas e HidrofílicasRESUMO
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.
Assuntos
Neoplasias da Mama/metabolismo , Proteína de Sequência 1 de Leucemia de Células Mieloides/metabolismo , Compostos de Anilina/farmacologia , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Apoptose/genética , Neoplasias da Mama/genética , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Feminino , Expressão Gênica , Humanos , Proteína de Sequência 1 de Leucemia de Células Mieloides/antagonistas & inibidores , Proteína de Sequência 1 de Leucemia de Células Mieloides/genética , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Interferência de RNA , RNA Interferente Pequeno/genética , Sulfonamidas/farmacologia , Proteína Killer-Antagonista Homóloga a bcl-2/genética , Proteína Killer-Antagonista Homóloga a bcl-2/metabolismo , Proteína bcl-X/genética , Proteína bcl-X/metabolismoRESUMO
Myeloid cell leukemia 1 (MCL-1) is a BCL-2 family protein that has been implicated in the progression and survival of multiple tumor types. Herein we report a series of MCL-1 inhibitors that emanated from a high throughput screening (HTS) hit and progressed via iterative cycles of structure-guided design. Advanced compounds from this series exhibited subnanomolar affinity for MCL-1 and excellent selectivity over other BCL-2 family proteins as well as multiple kinases and GPCRs. In a MCL-1 dependent human tumor cell line, administration of compound 30b rapidly induced caspase activation with associated loss in cell viability. The small molecules described herein thus comprise effective tools for studying MCL-1 biology.
Assuntos
Antineoplásicos/química , Antineoplásicos/farmacologia , Desenho de Fármacos , Mieloma Múltiplo/tratamento farmacológico , Proteína de Sequência 1 de Leucemia de Células Mieloides/química , Neoplasias Pancreáticas/tratamento farmacológico , Apoptose/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Cristalografia por Raios X , Bases de Dados Factuais , Ensaios de Triagem em Larga Escala , Humanos , Simulação de Acoplamento Molecular , Estrutura Molecular , Mieloma Múltiplo/metabolismo , Mieloma Múltiplo/patologia , Proteína de Sequência 1 de Leucemia de Células Mieloides/metabolismo , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patologia , Ligação Proteica , Relação Estrutura-Atividade , Células Tumorais CultivadasRESUMO
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.
Assuntos
Benzimidazóis/química , Proteínas de Choque Térmico HSP90/antagonistas & inibidores , Benzimidazóis/síntese química , Benzimidazóis/farmacologia , Sítios de Ligação , Linhagem Celular Tumoral , Cristalografia por Raios X , Proteínas de Choque Térmico HSP90/metabolismo , Humanos , Estrutura Terciária de Proteína , Relação Estrutura-AtividadeRESUMO
Overexpression of prosurvival proteins such as Bcl-2 and Bcl-X L has been correlated with tumorigenesis and resistance to chemotherapy, and thus, the development of antagonists of these proteins may provide a novel means for the treatment of cancer. We recently described the discovery of 1 (ABT-737), which binds Bcl-2, Bcl-X L, and Bcl-w with high affinity, shows robust antitumor activity in murine tumor xenograft models, but is not orally bioavailable. Herein, we report that targeted modifications at three key positions of 1 resulted in a 20-fold improvement in the pharmacokinetic/pharmacodynamic relationship (PK/PD) between oral exposure (AUC) and in vitro efficacy in human tumor cell lines (EC 50). The resulting compound, 2 (ABT-263), is orally efficacious in an established xenograft model of human small cell lung cancer, inducing complete tumor regressions in all animals. Compound 2 is currently in multiple phase 1 clinical trials in patients with small cell lung cancer and hematological malignancies.
Assuntos
Proteínas Proto-Oncogênicas c-bcl-2/antagonistas & inibidores , Administração Oral , Animais , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Avaliação Pré-Clínica de Medicamentos , Humanos , Camundongos , Estrutura Molecular , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Relação Estrutura-Atividade , Sulfonamidas/síntese química , Sulfonamidas/química , Sulfonamidas/farmacologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
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.
Assuntos
Antineoplásicos/síntese química , Compostos de Bifenilo/síntese química , Nitrofenóis/síntese química , Proteínas Proto-Oncogênicas c-bcl-2/antagonistas & inibidores , Sulfonamidas/síntese química , Animais , Antineoplásicos/química , Antineoplásicos/farmacologia , Compostos de Bifenilo/química , Compostos de Bifenilo/farmacologia , Linhagem Celular Tumoral , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Linfoma , Camundongos , Camundongos SCID , Modelos Moleculares , Nitrofenóis/química , Nitrofenóis/farmacologia , Piperazinas/síntese química , Piperazinas/química , Piperazinas/farmacologia , Proteínas Proto-Oncogênicas c-bcl-2/química , Relação Estrutura-Atividade , Sulfonamidas/química , Sulfonamidas/farmacologia , Transplante Heterólogo , Proteína bcl-X/antagonistas & inibidores , Proteína bcl-X/químicaRESUMO
Inhibition of the prosurvival members of the Bcl-2 family of proteins represents an attractive strategy for the treatment of cancer. We have previously reported the activity of ABT-737, a potent inhibitor of Bcl-2, Bcl-X(L), and Bcl-w, which exhibits monotherapy efficacy in xenograft models of small-cell lung cancer and lymphoma and potentiates the activity of numerous cytotoxic agents. Here we describe the biological activity of A-385358, a small molecule with relative selectivity for binding to Bcl-X(L) versus Bcl-2 (K(i)'s of 0.80 and 67 nmol/L for Bcl-X(L) and Bcl-2, respectively). This compound efficiently enters cells and co-localizes with the mitochondrial membrane. Although A-385358 shows relatively modest single-agent cytotoxic activity against most tumor cell lines, it has an EC(50) of <500 nmol/L in cells dependent on Bcl-X(L) for survival. In addition, A-385358 enhances the in vitro cytotoxic activity of numerous chemotherapeutic agents (paclitaxel, etoposide, cisplatin, and doxorubicin) in several tumor cell lines. In A549 non-small-cell lung cancer cells, A-385358 potentiates the activity of paclitaxel by as much as 25-fold. Importantly, A-385358 also potentiated the activity of paclitaxel in vivo. Significant inhibition of tumor growth was observed when A-385358 was added to maximally tolerated or half maximally tolerated doses of paclitaxel in the A549 xenograft model. In tumors, the combination therapy also resulted in a significant increase in mitotic arrest followed by apoptosis relative to paclitaxel monotherapy.
Assuntos
Compostos de Anilina/farmacologia , Antineoplásicos/uso terapêutico , Compostos de Bifenilo/uso terapêutico , Neoplasias Pulmonares/tratamento farmacológico , Nitrofenóis/uso terapêutico , Sulfonamidas/farmacologia , Sulfonamidas/uso terapêutico , Proteína bcl-X/antagonistas & inibidores , Compostos de Anilina/farmacocinética , Compostos de Anilina/uso terapêutico , Animais , Antineoplásicos/farmacocinética , Antineoplásicos/farmacologia , Compostos de Bifenilo/farmacocinética , Compostos de Bifenilo/farmacologia , Linhagem Celular Tumoral , Sinergismo Farmacológico , Humanos , Cinética , Masculino , Camundongos , Camundongos SCID , Nitrofenóis/farmacocinética , Nitrofenóis/farmacologia , Paclitaxel/farmacocinética , Piperazinas/farmacocinética , Piperazinas/farmacologia , Piperazinas/uso terapêutico , Sulfonamidas/farmacocinética , Transplante HeterólogoRESUMO
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.
Assuntos
Compostos de Anilina/síntese química , Antineoplásicos/síntese química , Piperidinas/síntese química , Sulfonamidas/síntese química , Proteína bcl-X/antagonistas & inibidores , Compostos de Anilina/química , Compostos de Anilina/farmacologia , Animais , Antineoplásicos/química , Antineoplásicos/farmacologia , Disponibilidade Biológica , Linhagem Celular Tumoral , Ensaios de Seleção de Medicamentos Antitumorais , Sinergismo Farmacológico , Polarização de Fluorescência , Humanos , Espectroscopia de Ressonância Magnética , Camundongos , Camundongos SCID , Paclitaxel/farmacologia , Piperidinas/química , Piperidinas/farmacologia , Ligação Proteica , Estrutura Terciária de Proteína , Soro , Albumina Sérica/química , Estereoisomerismo , Sulfonamidas/química , Sulfonamidas/farmacologia , Transplante Heterólogo , Raios UltravioletaRESUMO
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.
Assuntos
Antineoplásicos/uso terapêutico , Compostos de Bifenilo/farmacologia , Compostos de Bifenilo/uso terapêutico , Neoplasias/tratamento farmacológico , Neoplasias/patologia , Proteínas Proto-Oncogênicas c-bcl-2/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-bcl-2/classificação , Animais , Antineoplásicos/síntese química , Antineoplásicos/química , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Compostos de Bifenilo/síntese química , Compostos de Bifenilo/química , Carcinoma de Células Pequenas/tratamento farmacológico , Carcinoma de Células Pequenas/patologia , Linhagem Celular Tumoral , Citocromos c/metabolismo , Modelos Animais de Doenças , Sinergismo Farmacológico , Humanos , Linfoma/tratamento farmacológico , Linfoma/patologia , Espectroscopia de Ressonância Magnética , Camundongos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Modelos Moleculares , Nitrofenóis , Paclitaxel/farmacologia , Piperazinas , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Relação Estrutura-Atividade , Sulfonamidas , Taxa de SobrevidaRESUMO
A series of non-amide-linked 6-substituted-2-naphthamidine urokinase plasminogen activator (uPA) inhibitors are described. These compounds possess excellent binding activities and selectivities with significantly improved pharmacokinetic profiles versus previously described amide-linked inhibitors.