RESUMEN
The casein kinase 1 enzymes (CK1) form a family of serine/threonine kinases with seven CK1 isoforms identified in humans. The most important substrates of CK1 kinases are proteins that act in the regulatory nodes essential for tumorigenesis of hematological malignancies. Among those, the most important are the functions of CK1s in the regulation of Wnt pathways, cell proliferation, apoptosis and autophagy. In this review we summarize the recent developments in the understanding of biology and therapeutic potential of the inhibition of CK1 isoforms in the pathogenesis of chronic lymphocytic leukemia (CLL), other non-Hodgkin lymphomas (NHL), myelodysplastic syndrome (MDS), acute myeloid leukemia (AML) and multiple myeloma (MM). CK1δ/ε inhibitors block CLL development in preclinical models via inhibition of WNT-5A/ROR1-driven non-canonical Wnt pathway. While no selective CK1 inhibitors have reached clinical stage to date, one dual PI3Kδ and CK1ε inhibitor, umbralisib, is currently in clinical trials for CLL and NHL patients. In MDS, AML and MM, inhibition of CK1α, acting via activation of p53 pathway, showed promising preclinical activities and the first CK1α inhibitor has now entered the clinical trials.
Asunto(s)
Quinasa de la Caseína I/metabolismo , Neoplasias Hematológicas/tratamiento farmacológico , Neoplasias Hematológicas/enzimología , Terapia Molecular Dirigida , Animales , Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Quinasa de la Caseína I/antagonistas & inhibidores , Quinasa de la Caseína I/química , Neoplasias Hematológicas/patología , Humanos , Células Madre Neoplásicas/efectos de los fármacos , Células Madre Neoplásicas/metabolismo , Células Madre Neoplásicas/patología , Vía de Señalización WntRESUMEN
In the activated B-cell-like (ABC) subtype of diffuse large B-cell lymphoma (DLBCL), NF-κB activity is essential for viability of the malignant cells and is sustained by constitutive activity of IκB kinase (IKK) in the cytoplasm. Here, we report an unexpected role for the bromodomain and extraterminal domain (BET) proteins BRD2 and BRD4 in maintaining oncogenic IKK activity in ABC DLBCL. IKK activity was reduced by small molecules targeting BET proteins as well as by genetic knockdown of BRD2 and BRD4 expression, thereby inhibiting downstream NF-κB-driven transcriptional programs and killing ABC DLBCL cells. Using a high-throughput platform to screen for drug-drug synergy, we observed that the BET inhibitor JQ1 combined favorably with multiple drugs targeting B-cell receptor signaling, one pathway that activates IKK in ABC DLBCL. The BTK kinase inhibitor ibrutinib, which is in clinical development for the treatment of ABC DLBCL, synergized strongly with BET inhibitors in killing ABC DLBCL cells in vitro and in a xenograft mouse model. These findings provide a mechanistic basis for the clinical development of BET protein inhibitors in ABC DLBCL, particularly in combination with other modulators of oncogenic IKK signaling.
Asunto(s)
Quinasa I-kappa B/antagonistas & inhibidores , Linfoma de Células B Grandes Difuso/enzimología , Proteínas Nucleares/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Factores de Transcripción/antagonistas & inhibidores , Adenina/análogos & derivados , Animales , Azepinas/farmacología , Azepinas/toxicidad , Proteínas de Ciclo Celular , Muerte Celular/efectos de los fármacos , Línea Celular Tumoral , Supervivencia Celular , Sinergismo Farmacológico , Humanos , Quinasa I-kappa B/química , Quinasa I-kappa B/metabolismo , Linfoma de Células B Grandes Difuso/patología , Ratones , Ratones SCID , Proteínas Nucleares/metabolismo , Piperidinas , Proteínas Serina-Treonina Quinasas/metabolismo , Estructura Terciaria de Proteína , Pirazoles/farmacología , Pirimidinas/farmacología , Transducción de Señal/efectos de los fármacos , Factores de Transcripción/metabolismo , Triazoles/farmacología , Triazoles/toxicidad , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
The discovery and optimization of a series of small molecule EZH2 inhibitors is described. Starting from dimethylpyridone HTS hit (2), a series of indole-based EZH2 inhibitors were identified. Biochemical potency and microsomal stability were optimized during these studies and afforded compound 22. This compound demonstrates nanomolar levels of biochemical potency (IC50=0.002 µM), cellular potency (EC50=0.080 µM), and afforded tumor regression when dosed (200 mpk SC BID) in an EZH2 dependent tumor xenograft model.
Asunto(s)
Antineoplásicos/química , Antineoplásicos/farmacología , Indoles/química , Complejo Represivo Polycomb 2/antagonistas & inhibidores , Animales , Antineoplásicos/síntesis química , Línea Celular Tumoral , Técnicas de Química Sintética , Diseño de Fármacos , Descubrimiento de Drogas , Ensayos de Selección de Medicamentos Antitumorales , Estabilidad de Medicamentos , Proteína Potenciadora del Homólogo Zeste 2 , Células HeLa/efectos de los fármacos , Humanos , Concentración 50 Inhibidora , Ratones , Terapia Molecular Dirigida/métodos , Bibliotecas de Moléculas Pequeñas/química , Bibliotecas de Moléculas Pequeñas/farmacología , Relación Estructura-Actividad , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Despite the promising introduction of the proteasome inhibitor bortezomib in the treatment of mantle cell lymphoma (MCL), not all patients respond, and resistance often appears after initial treatment. By analyzing a set of 18 MCL samples, including cell lines with constitutive or induced resistance to bortezomib, we found a high correlation between loss of sensitivity to the proteasome inhibitor and up-regulation of the prosurvival chaperone BiP/Grp78. BiP/Grp78 stabilization was ensured at a posttranscriptional level by an increase in the chaperoning activity of heat shock protein of 90 kDa (Hsp90). In bortezomib-resistant cells, both BiP/Grp78 knockdown and cell pretreatment with the Hsp90 inhibitor of the ansamycin class, IPI-504, led to synergistic induction of apoptotic cell death when combined with bortezomib. Cell exposure to the IPI-504-bortezomib combination provoked the dissociation of Hsp90/BiP complexes, leading to BiP/Grp78 depletion, inhibition of unfolded protein response, and promotion of NOXA-mediated mitochondrial depolarization. The IPI-504-bortezomib combination also prevented BiP/Grp78 accumulation, thereby promoting apoptosis and inhibiting the growth of bortezomib-resistant tumors in a mouse model of MCL xenotransplantation. These results suggest that targeting unfolded protein response activation by the inhibition of Hsp90 may be an attractive model for the design of a new bortezomib-based combination therapy for MCL.
Asunto(s)
Benzoquinonas/farmacología , Ácidos Borónicos/uso terapéutico , Resistencia a Antineoplásicos/efectos de los fármacos , Proteínas de Choque Térmico/genética , Lactamas Macrocíclicas/farmacología , Linfoma de Células del Manto/tratamiento farmacológico , Pirazinas/uso terapéutico , Adulto , Anciano , Anciano de 80 o más Años , Animales , Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Benzoquinonas/uso terapéutico , Bortezomib , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/genética , Regulación hacia Abajo/efectos de los fármacos , Resistencia a Antineoplásicos/genética , Chaperón BiP del Retículo Endoplásmico , Femenino , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Técnicas de Silenciamiento del Gen , Proteínas HSP90 de Choque Térmico/antagonistas & inhibidores , Proteínas de Choque Térmico/metabolismo , Humanos , Lactamas Macrocíclicas/uso terapéutico , Linfoma de Células del Manto/genética , Masculino , Ratones , Ratones SCID , Ratones Transgénicos , Persona de Mediana Edad , Chaperonas Moleculares/genética , Chaperonas Moleculares/metabolismo , Receptores de Estrógenos/metabolismo , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Background: Immunotherapy-based regimens have considerably improved the survival rate of B-cell non-Hodgkin lymphoma (B-NHL) patients in the last decades; however, most disease subtypes remain almost incurable. TG-1801, a bispecific antibody that targets CD47 selectively on CD19+ B-cells, is under clinical evaluation in relapsed/refractory (R/R) B-NHL patients either as a single-agent or in combination with ublituximab, a new generation CD20 antibody. Methods: A set of eight B-NHL cell lines and primary samples were cultured in vitro in the presence of bone marrow-derived stromal cells, M2-polarized primary macrophages, and primary circulating PBMCs as a source of effector cells. Cell response to TG-1801 alone or combined with the U2 regimen associating ublituximab to the PI3Kδ inhibitor umbralisib, was analyzed by proliferation assay, western blot, transcriptomic analysis (qPCR array and RNA sequencing followed by gene set enrichment analysis) and/or quantification of antibody-dependent cell death (ADCC) and antibody-dependent cell phagocytosis (ADCP). CRISPR-Cas9 gene edition was used to selectively abrogate GPR183 gene expression in B-NHL cells. In vivo, drug efficacy was determined in immunodeficient (NSG mice) or immune-competent (chicken embryo chorioallantoic membrane (CAM)) B-NHL xenograft models. Results: Using a panel of B-NHL co-cultures, we show that TG-1801, by disrupting the CD47-SIRPα axis, potentiates anti-CD20-mediated ADCC and ADCP. This led to a remarkable and durable antitumor effect of the triplet therapy composed by TG-1801 and U2 regimen, in vitro, as well as in mice and CAM xenograft models of B-NHL. Transcriptomic analysis also uncovered the upregulation of the G protein-coupled and inflammatory receptor, GPR183, as a crucial event associated with the efficacy of the triplet combination. Genetic depletion and pharmacological inhibition of GPR183 impaired ADCP initiation, cytoskeleton remodeling and cell migration in 2D and 3D spheroid B-NHL co-cultures, and disrupted macrophage-mediated control of tumor growth in B-NHL CAM xenografts. Conclusions: Altogether, our results support a crucial role for GPR183 in the recognition and elimination of malignant B cells upon concomitant targeting of CD20, CD47 and PI3Kδ, and warrant further clinical evaluation of this triplet regimen in B-NHL.
Asunto(s)
Anticuerpos Biespecíficos , Linfoma de Burkitt , Linfoma de Células B , Neoplasias , Embrión de Pollo , Humanos , Ratones , Animales , Inhibidores de Puntos de Control Inmunológico/uso terapéutico , Antígeno CD47 , Neoplasias/metabolismo , Linfoma de Células B/tratamiento farmacológico , Anticuerpos Biespecíficos/farmacología , Anticuerpos Biespecíficos/uso terapéutico , Modelos Animales de Enfermedad , Receptores Acoplados a Proteínas GRESUMEN
PURPOSE: PI3K inhibitors (PI3Ki) are approved for relapsed chronic lymphocytic leukemia (CLL). Although patients may show an initial response to these therapies, development of treatment intolerance or resistance remain clinical challenges. To overcome these, prediction of individual treatment responses based on actionable biomarkers is needed. Here, we characterized the activity and cellular effects of 10 PI3Ki and investigated whether functional analyses can identify treatment vulnerabilities in PI3Ki-refractory/intolerant CLL and stratify responders to PI3Ki. EXPERIMENTAL DESIGN: Peripheral blood mononuclear cell samples (n = 51 in total) from treatment-naïve and PI3Ki-treated patients with CLL were studied. Cells were profiled against 10 PI3Ki and the Bcl-2 antagonist venetoclax. Cell signaling and immune phenotypes were analyzed by flow cytometry. Cell viability was monitored by detection of cleaved caspase-3 and the CellTiter-Glo assay. RESULTS: pan-PI3Kis were most effective at inhibiting PI3K signaling and cell viability, and showed activity in CLL cells from both treatment-naïve and idelalisib-refractory/intolerant patients. CLL cells from idelalisib-refractory/intolerant patients showed overall reduced protein phosphorylation levels. The pan-PI3Ki copanlisib, but not the p110δ inhibitor idelalisib, inhibited PI3K signaling in CD4+ and CD8+ T cells in addition to CD19+ B cells, but did not significantly affect T-cell numbers. Combination treatment with a PI3Ki and venetoclax resulted in synergistic induction of apoptosis. Analysis of drug sensitivities to 73 drug combinations and profiling of 31 proteins stratified responders to idelalisib and umbralisib, respectively. CONCLUSIONS: Our findings suggest novel treatment vulnerabilities in idelalisib-refractory/intolerant CLL, and indicate that ex vivo functional profiling may stratify PI3Ki responders.
Asunto(s)
Antineoplásicos , Leucemia Linfocítica Crónica de Células B , Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Compuestos Bicíclicos Heterocíclicos con Puentes , Caspasa 3 , Humanos , Leucemia Linfocítica Crónica de Células B/tratamiento farmacológico , Leucocitos Mononucleares/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Inhibidores de las Quinasa Fosfoinosítidos-3 , Proteínas Proto-Oncogénicas c-bcl-2/genética , Quinazolinonas/farmacología , Quinazolinonas/uso terapéutico , SulfonamidasRESUMEN
Several Hsp90 (heat shock protein 90) inhibitors are currently under clinical evaluation as anticancer agents. However, the correlation between the duration and magnitude of Hsp90 inhibition and the downstream effects on client protein degradation and cancer cell growth inhibition has not been thoroughly investigated. To investigate the relationship between Hsp90 inhibition and cellular effects, we developed a method that measures drug occupancy on Hsp90 after treatment with the Hsp90 inhibitor IPI-504 in living cells and in tumor xenografts. In cells, we find the level of Hsp90 occupancy to be directly correlated with cell growth inhibition. At the molecular level, the relationship between Hsp90 occupancy and Hsp90 client protein degradation was examined for different client proteins. For sensitive Hsp90 clients (e.g. HER2 (human epidermal growth factor receptor 2), client protein levels directly mirror Hsp90 occupancy at all time points after IPI-504 administration. For insensitive client proteins, we find that protein abundance matches Hsp90 occupancy only after prolonged incubation with drug. Additionally, we investigate the correlation between plasma pharmacokinetics (PK), tumor PK, pharmacodynamics (PD) (client protein degradation), tumor growth inhibition, and Hsp90 occupancy in a xenograft model of human cancer. Our results indicate Hsp90 occupancy to be a better predictor of PD than either plasma PK or tumor PK. In the nonsmall cell lung cancer xenograft model studied, a linear correlation between Hsp90 occupancy and tumor growth inhibition was found. This novel binding assay was evaluated both in vitro and in vivo and could be used as a pharmacodynamic readout in the clinic.
Asunto(s)
Antineoplásicos/farmacocinética , Benzoquinonas/farmacocinética , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Proteínas HSP90 de Choque Térmico/antagonistas & inhibidores , Lactamas Macrocíclicas/farmacocinética , Neoplasias Pulmonares/tratamiento farmacológico , Proteínas de Neoplasias/antagonistas & inhibidores , Ensayos Antitumor por Modelo de Xenoinjerto , Animales , Antineoplásicos/farmacología , Benzoquinonas/farmacología , Carcinoma de Pulmón de Células no Pequeñas/metabolismo , Proteínas HSP90 de Choque Térmico/metabolismo , Células HeLa , Humanos , Lactamas Macrocíclicas/farmacología , Neoplasias Pulmonares/metabolismo , Ratones , Ratones DesnudosRESUMEN
PURPOSE: Despite the remarkable activity of BTK inhibitors (BTKi) in relapsed B-cell non-Hodgkin lymphoma (B-NHL), no clinically-relevant biomarker has been associated to these agents so far. The relevance of phosphoproteomic profiling for the early identification of BTKi responders remains underexplored. EXPERIMENTAL DESIGN: A set of six clinical samples from an ongoing phase I trial dosing patients with chronic lymphocytic leukemia (CLL) with TG-1701, a novel irreversible and highly specific BTKi, were characterized by phosphoproteomic and RNA sequencing (RNA-seq) analysis. The activity of TG-1701 was evaluated in a panel of 11 B-NHL cell lines and mouse xenografts, including two NF-κB- and BTKC481S-driven BTKi-resistant models. Biomarker validation and signal transduction analysis were conducted through real-time PCR, Western blot analysis, immunostaining, and gene knockout (KO) experiments. RESULTS: A nonsupervised, phosphoproteomic-based clustering did match the early clinical outcomes of patients with CLL and separated a group of "early-responders" from a group of "late-responders." This clustering was based on a selected list of 96 phosphosites with Ikaros-pSer442/445 as a potential biomarker for TG-1701 efficacy. TG-1701 treatment was further shown to blunt Ikaros gene signature, including YES1 and MYC, in early-responder patients as well as in BTKi-sensitive B-NHL cell lines and xenografts. In contrast, Ikaros nuclear activity and signaling remained unaffected by the drug in vitro and in vivo in late-responder patients and in BTKC481S, BTKKO, and noncanonical NF-κB models. CONCLUSIONS: These data validate phosphoproteomic as a valuable tool for the early detection of response to BTK inhibition in the clinic, and for the determination of drug mechanism of action.
Asunto(s)
Leucemia Linfocítica Crónica de Células B , Linfoma no Hodgkin , Agammaglobulinemia Tirosina Quinasa , Animales , Humanos , Leucemia Linfocítica Crónica de Células B/tratamiento farmacológico , Linfoma no Hodgkin/tratamiento farmacológico , Ratones , Piperidinas/uso terapéutico , Inhibidores de Proteínas Quinasas/farmacología , Inhibidores de Proteínas Quinasas/uso terapéutico , Pirazoles/farmacología , Pirimidinas/farmacología , Transducción de SeñalRESUMEN
BACKGROUND: CD47 is a broadly expressed cell surface glycoprotein associated with immune evasion. Interaction with the inhibitory receptor signal regulatory protein alpha (SIRPα), primarily expressed on myeloid cells, normally serves to restrict effector function (eg, phagocytosis and immune cell homeostasis). CD47/SIRPα antagonists, commonly referred to as 'macrophage checkpoint' inhibitors, are being developed as cancer interventions. SRF231 is an investigational fully human IgG4 anti-CD47 antibody that is currently under evaluation in a phase 1 clinical trial. The development and preclinical characterization of SRF231 are reported here. METHODS: SRF231 was characterized in assays designed to probe CD47/SIRPα blocking potential and effects on red blood cell (RBC) phagocytosis and agglutination. Additionally, SRF231-mediated phagocytosis and cell death were assessed in macrophage:tumor cell in vitro coculture systems. Further mechanistic studies were conducted within these coculture systems to ascertain the dependency of SRF231-mediated antitumor activity on Fc receptor engagement vs CD47/SIRPα blockade. In vivo, SRF231 was evaluated in a variety of hematologic xenograft models, and the mechanism of antitumor activity was assessed using cytokine and macrophage infiltration analyses following SRF231 treatment. RESULTS: SRF231 binds CD47 and disrupts the CD47/SIRPα interaction without causing hemagglutination or RBC phagocytosis. SRF231 exerts antitumor activity in vitro through both phagocytosis and cell death in a manner dependent on the activating Fc-gamma receptor (FcγR), CD32a. Through its Fc domain, SRF231 engagement with macrophage-derived CD32a serves dual purposes by eliciting FcγR-mediated phagocytosis of cancer cells and acting as a scaffold to drive CD47-mediated death signaling into tumor cells. Robust antitumor activity occurs across multiple hematologic xenograft models either as a single agent or in combination with rituximab. In tumor-bearing mice, SRF231 increases tumor macrophage infiltration and induction of the macrophage cytokines, mouse chemoattractant protein 1 and macrophage inflammatory protein 1 alpha. Macrophage depletion results in diminished SRF231 antitumor activity, underscoring a mechanistic role for macrophage engagement by SRF231. CONCLUSION: SRF231 elicits antitumor activity via apoptosis and phagocytosis involving macrophage engagement in a manner dependent on the FcγR, CD32a.
Asunto(s)
Antígeno CD47/metabolismo , Neoplasias/genética , Receptores de IgG/metabolismo , Animales , Humanos , Ratones , Neoplasias/patología , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
BACKGROUND: Inhibitors of heat shock protein (Hsp) 90 induce apoptosis in multiple myeloma (MM) cells, but the molecular mechanisms underlying this cytotoxic outcome are not clear. Here, we investigate the effect of IPI-504, a novel and highly soluble inhibitor of the Hsp90 ATPase activity, on the unfolded protein response (UPR) in MM cells. The UPR is a stress response pathway triggered by sensors located at the endoplasmic reticulum (ER) membrane whose function is to reduce an excessive accumulation of misfolded protein in the ER. During normal development of B-lymphocytes to antibody-producing plasma cells, a partial UPR has been described, where IREalpha and ATF-6 are stimulated, whereas the third sensor, PERK, is not induced. METHODS: Levels of the activated forms of the three main UPR sensors ATF-6, XBP-1 and PERK/eIF-2 were monitored in two different MM cells lines and one non-MM cell lines under various experimental conditions including incubation with increasing concentration of IPI-504. Also, MM cells were incubated with IPI-504 and several apoptosis markers were monitored. RESULTS: We show here that a partial UPR is constitutively activated in plasma cell-derived MM cells and that IPI-504 can potently inhibit this pathway. IPI-504 achieves this by inactivating the transcription factors XBP1 and ATF6. In addition, IPI-504 also blocks the tunicamycin-induced phosphorylation of eIF2 by PERK. Dose-response and time course experiments reveal that IPI-504's inhibitory effect on the UPR parallels its cytotoxic and pro-apoptotic effects on MM cells. CONCLUSION: The results presented here suggest that the IPI-504-induced apoptosis might be, in part, mediated by the inhibition of the partial UPR. Other malignancies that rely on intact and efficient UPR to survive could be considered as new indications for Hsp90 inhibitors.
Asunto(s)
Antineoplásicos/farmacología , Benzoquinonas/farmacología , Proteínas HSP90 de Choque Térmico/antagonistas & inhibidores , Lactamas Macrocíclicas/farmacología , Mieloma Múltiple/tratamiento farmacológico , Mieloma Múltiple/metabolismo , Pliegue de Proteína , Factor de Transcripción Activador 6/genética , Factor de Transcripción Activador 6/fisiología , Antivirales/farmacología , Apoptosis/efectos de los fármacos , Apoptosis/fisiología , Western Blotting , Muerte Celular/efectos de los fármacos , Línea Celular Tumoral , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/fisiología , Relación Dosis-Respuesta a Droga , Factor 2 Eucariótico de Iniciación/genética , Factor 2 Eucariótico de Iniciación/fisiología , Genes Reporteros/genética , Humanos , Luciferasas/genética , Mieloma Múltiple/patología , Proteínas Nucleares/genética , Proteínas Nucleares/fisiología , Fosforilación , Factores de Transcripción del Factor Regulador X , Factores de Transcripción , Transfección , Tunicamicina/farmacología , Proteína 1 de Unión a la X-Box , eIF-2 Quinasa/genética , eIF-2 Quinasa/fisiologíaRESUMEN
17-Allylamino-17-demethoxygeldanamycin (17-AAG)1 is a semisynthetic inhibitor of the 90 kDa heat shock protein (Hsp90) currently in clinical trials for the treatment of cancer. However, 17-AAG faces challenging formulation issues due to its poor solubility. Here we report the synthesis and evaluation of a highly soluble hydroquinone hydrochloride derivative of 17-AAG, 1a (IPI-504), and several of the physiological metabolites. These compounds show comparable binding affinity to human Hsp90 and its endoplasmic reticulum (ER) homologue, the 94 kDa glucose regulated protein (Grp94). Furthermore, the compounds inhibit the growth of the human cancer cell lines SKBR3 and SKOV3, which overexpress Hsp90 client protein Her2, and cause down-regulation of Her2 as well as induction of Hsp70 consistent with Hsp90 inhibition. There is a clear correlation between the measured binding affinity of the compounds and their cellular activities. Upon the basis of its potent activity against Hsp90 and a significant improvement in solubility, 1a is currently under evaluation in Phase I clinical trials for cancer.
Asunto(s)
Antineoplásicos/síntesis química , Benzoquinonas/síntesis química , Proteínas HSP90 de Choque Térmico/antagonistas & inhibidores , Hidroquinonas/síntesis química , Lactamas Macrocíclicas/síntesis química , Rifabutina/análogos & derivados , Animales , Antineoplásicos/química , Antineoplásicos/farmacología , Benzoquinonas/química , Benzoquinonas/farmacología , Unión Competitiva , Línea Celular Tumoral , Perros , Ensayos de Selección de Medicamentos Antitumorales , Polarización de Fluorescencia , Proteínas HSP70 de Choque Térmico/biosíntesis , Proteínas HSP70 de Choque Térmico/química , Proteínas HSP90 de Choque Térmico/química , Humanos , Hidroquinonas/química , Hidroquinonas/farmacología , Lactamas Macrocíclicas/química , Lactamas Macrocíclicas/farmacología , Proteínas de la Membrana/química , Modelos Moleculares , Isoformas de Proteínas/química , Receptor ErbB-2/antagonistas & inhibidores , Receptor ErbB-2/biosíntesis , Rifabutina/síntesis química , Rifabutina/química , Rifabutina/farmacología , Solubilidad , Relación Estructura-Actividad , AguaRESUMEN
In recent years, inhibition of the interaction between the bromodomain and extra-terminal domain (BET) family of chromatin adaptors and acetyl-lysine residues on chromatin has emerged as a promising approach to regulate the expression of important disease-relevant genes, including MYC, BCL-2, and NF-κB. Here we describe the identification and characterization of a potent and selective benzoisoxazoloazepine BET bromodomain inhibitor that attenuates BET-dependent gene expression in vivo, demonstrates antitumor efficacy in an MV-4-11 mouse xenograft model, and is currently undergoing human clinical trials for hematological malignancies (CPI-0610).
Asunto(s)
Antineoplásicos/química , Antineoplásicos/uso terapéutico , Azepinas/química , Azepinas/uso terapéutico , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Leucemia Mieloide Aguda/tratamiento farmacológico , Proteínas Nucleares/antagonistas & inhibidores , Factores de Transcripción/antagonistas & inhibidores , Animales , Antineoplásicos/farmacocinética , Antineoplásicos/farmacología , Azepinas/farmacocinética , Azepinas/farmacología , Proteínas de Ciclo Celular , Línea Celular Tumoral , Ensayos Clínicos como Asunto , Perros , Genes myc/efectos de los fármacos , Humanos , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/metabolismo , Ratones , Proteínas Nucleares/química , Proteínas Nucleares/metabolismo , Proteínas Proto-Oncogénicas c-myc/genética , Ratas , Factores de Transcripción/química , Factores de Transcripción/metabolismo , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Polycomb repressive complex 2 (PRC2) has been shown to play a major role in transcriptional silencing in part by installing methylation marks on lysine 27 of histone 3. Dysregulation of PRC2 function correlates with certain malignancies and poor prognosis. EZH2 is the catalytic engine of the PRC2 complex and thus represents a key candidate oncology target for pharmacological intervention. Here we report the optimization of our indole-based EZH2 inhibitor series that led to the identification of CPI-1205, a highly potent (biochemical IC50 = 0.002 µM, cellular EC50 = 0.032 µM) and selective inhibitor of EZH2. This compound demonstrates robust antitumor effects in a Karpas-422 xenograft model when dosed at 160 mg/kg BID and is currently in Phase I clinical trials. Additionally, we disclose the co-crystal structure of our inhibitor series bound to the human PRC2 complex.
Asunto(s)
Antineoplásicos/farmacología , Ensayos Clínicos Fase I como Asunto , Inhibidores Enzimáticos/farmacología , N-Metiltransferasa de Histona-Lisina/antagonistas & inhibidores , Indoles/farmacología , Linfoma de Células B/tratamiento farmacológico , Piperidinas/farmacología , Animales , Antineoplásicos/síntesis química , Antineoplásicos/química , Proliferación Celular/efectos de los fármacos , Perros , Relación Dosis-Respuesta a Droga , Ensayos de Selección de Medicamentos Antitumorales , Inhibidores Enzimáticos/síntesis química , Inhibidores Enzimáticos/química , Histona Metiltransferasas , N-Metiltransferasa de Histona-Lisina/metabolismo , Humanos , Indoles/síntesis química , Indoles/química , Modelos Moleculares , Estructura Molecular , Neoplasias Experimentales/tratamiento farmacológico , Neoplasias Experimentales/patología , Piperidinas/síntesis química , Piperidinas/química , Ratas , Relación Estructura-ActividadRESUMEN
The histone lysine methyltransferase (MT) Enhancer of Zeste Homolog 2 (EZH2) is considered an oncogenic driver in a subset of germinal center B-cell-like diffuse large B cell lymphoma (GCB-DLBCL) and follicular lymphoma due to the presence of recurrent, monoallelic mutations in the EZH2 catalytic domain. These genomic data suggest that targeting the EZH2 MT activity is a valid therapeutic strategy for the treatment of lymphoma patients with EZH2 mutations. Here we report the identification of highly potent and selective EZH2 small molecule inhibitors, their validation by a cellular thermal shift assay, application across a large cell panel representing various non-Hodgkin's lymphoma (NHL) subtypes, and their efficacy in EZH2mutant-containing GCB-DLBCL xenograft models. Surprisingly, our EZH2 inhibitors selectively affect the turnover of trimethylated, but not monomethylated histone H3 lysine 27 at pharmacologically relevant doses. Importantly, we find that these inhibitors are broadly efficacious also in NHL models with wild-type EZH2.
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Apoptosis/efectos de los fármacos , Inhibidores Enzimáticos/toxicidad , Histonas/metabolismo , Complejo Represivo Polycomb 2/antagonistas & inhibidores , Bibliotecas de Moléculas Pequeñas/toxicidad , Secuencia de Aminoácidos , Animales , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Modelos Animales de Enfermedad , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/uso terapéutico , Histonas/química , Humanos , Cinética , Linfoma de Células B Grandes Difuso/tratamiento farmacológico , Linfoma de Células B Grandes Difuso/patología , Linfoma no Hodgkin/metabolismo , Linfoma no Hodgkin/patología , Metilación , Ratones , Ratones Desnudos , Mutación , Péptidos/análisis , Complejo Represivo Polycomb 2/genética , Complejo Represivo Polycomb 2/metabolismo , Bibliotecas de Moléculas Pequeñas/química , Bibliotecas de Moléculas Pequeñas/uso terapéutico , Trasplante HeterólogoRESUMEN
PURPOSE: Heat shock protein 90 (HSP90) is required for the proper folding, function, and stability of various client proteins, two of which (KIT and PDGFRα) are critical in the pathogenesis and progression of gastrointestinal stromal tumors (GIST). This phase I study investigated the safety and maximum tolerated dose (MTD) of retaspimycin hydrochloride (IPI-504), a novel potent and selective HSP90 inhibitor, in patients with metastatic and/or unresectable GIST or other soft-tissue sarcomas (STS). EXPERIMENTAL DESIGN: IPI-504 was administered intravenously at doses ranging from 90 to 500 mg/m(2) twice weekly for 2 weeks on/1 week off. Safety, pharmacokinetic, and pharmacodynamic profiles were determined. Response was assessed by Response Evaluation Criteria for Solid Tumors (RECIST) 1.0 and optionally via 18-fluorodeoxyglucose positron emission tomography (18-FDG-PET) imaging. RESULTS: Fifty-four patients received IPI-504; 37 with GIST and 17 with other STS. The MTD was 400 mg/m(2) twice weekly for 2 weeks on/1 week off. Common related adverse events were fatigue (59%), headache (44%), and nausea (43%). Exposure to IPI-504, 17-AAG, and 17-AG increased with IPI-504 dose. Stable disease (SD) was observed in 70% (26 of 37) of patients with GIST and 59% (10 of 17) of patients with STS. There was one confirmed partial response (PR) in a patient with GIST and one PR in a patient with liposarcoma. Metabolic partial responses occurred in 11 of 29 (38%) patients with GIST. CONCLUSIONS: In this study of advanced GIST or other STS, IPI-504 was generally well-tolerated with some evidence of antitumor activity, serving as a clinical proof-of-concept that HSP90 inhibition remains a promising strategy.
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Antineoplásicos/uso terapéutico , Benzoquinonas/uso terapéutico , Neoplasias Gastrointestinales/tratamiento farmacológico , Tumores del Estroma Gastrointestinal/tratamiento farmacológico , Lactamas Macrocíclicas/uso terapéutico , Inhibidores de Proteínas Quinasas/uso terapéutico , Sarcoma/tratamiento farmacológico , Adulto , Anciano , Anciano de 80 o más Años , Antineoplásicos/farmacología , Benzoquinonas/farmacología , Femenino , Neoplasias Gastrointestinales/mortalidad , Tumores del Estroma Gastrointestinal/mortalidad , Proteínas HSP90 de Choque Térmico/antagonistas & inhibidores , Humanos , Lactamas Macrocíclicas/farmacología , Masculino , Persona de Mediana Edad , Inhibidores de Proteínas Quinasas/farmacología , Sarcoma/mortalidad , Resultado del Tratamiento , Adulto JovenRESUMEN
Treatment of gastroenteropancreatic neuroendocrine tumors (GEP-NET) is still unsatisfactory and innovative therapeutic approaches are urgently needed. Heat shock protein 90 (Hsp90) is overexpressed in a wide range of tumor types and is an emerging target for the treatment of cancer. However, the potential activity of Hsp90 inhibitors in GEP-NET has not yet been investigated. We studied the antineoplastic activity of the Hsp90 inhibitor IPI-504 on GEPNET cells, and characterized its mechanism of action. In human insulinoma (CM) and pancreatic carcinoid (BON) cells IPI-504 induced a dose-dependent growth inhibition by almost 70%. The antiproliferative effect of IPI-504 correlated with a reduction in protein levels of the IGF-1 receptor. Additionally, several proteins of the PI3K/AKT/mTOR pathway, downstream of IGF-1 receptor activation in GEP-NETs, were downregulated as a consequence of Hsp90 inhibition. Combination treatment of IPI-504 with mTOR- or AKT-inhibitors led to additive antiproliferative effects. In addition, effects of IGF-1 receptor tyrosine kinase inhibition were strongly enhanced by IPI-504. Cancer gene expression profiling and FACS analysis revealed that IPI-504 antiproliferative effects were due to both induction of cell cycle arrest and apoptosis. A modified chick chorioallantoic membrane (CAM) assay confirmed the antineoplastic activity of IPI-504 in GEP-NETs in vivo. In conclusion, this study showed that Hsp90 inhibition may be an attractive target for innovative GEP-NET treatment alone or in combination with either IGF-1R or mTOR inhibitors.
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Antineoplásicos/farmacología , Benzoquinonas/farmacología , Proliferación Celular/efectos de los fármacos , Neoplasias Gastrointestinales/metabolismo , Proteínas HSP90 de Choque Térmico/antagonistas & inhibidores , Lactamas Macrocíclicas/farmacología , Tumores Neuroendocrinos/metabolismo , Animales , Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Apoptosis/efectos de los fármacos , Puntos de Control del Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Movimiento Celular/efectos de los fármacos , Embrión de Pollo , Membrana Corioalantoides/irrigación sanguínea , Relación Dosis-Respuesta a Droga , Citometría de Flujo , Neoplasias Gastrointestinales/genética , Neoplasias Gastrointestinales/patología , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Proteínas HSP90 de Choque Térmico/metabolismo , Humanos , Tumores Neuroendocrinos/genética , Tumores Neuroendocrinos/patología , Fosfatidilinositol 3-Quinasa/metabolismo , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Proto-Oncogénicas c-akt/antagonistas & inhibidores , Proteínas Proto-Oncogénicas c-akt/metabolismo , Receptor IGF Tipo 1/antagonistas & inhibidores , Receptor IGF Tipo 1/metabolismo , Transducción de Señal/efectos de los fármacos , Serina-Treonina Quinasas TOR/antagonistas & inhibidores , Serina-Treonina Quinasas TOR/metabolismoRESUMEN
PURPOSE: KIT activity is crucial for gastrointestinal stromal tumors (GIST). Imatinib (IMA) and sunitinib (SUN) are very effective KIT-inhibitors in patients with advanced GIST but have no curative potential. We evaluated the efficacy of the novel HSP90 inhibitor IPI-493 alone, or in combination with IMA or SUN in GIST xenografts with KIT mutations. EXPERIMENTAL DESIGN: Nude mice (n = 98) were grafted bilaterally with human GIST carrying KIT exon 11 (GIST-PSW), KIT exon 9 (GIST-BOE), or double, KIT imatinib-sensitive exon 11 and imatinib-resistant exon 17 mutations (GIST-48). Mice were divided into six treatment groups and dosed orally for 15 days as follows: (i) control group, sterile water; (ii) IMA alone; (iii) SUN alone; (iv) IPI-493 alone; (v) IPI-493+IMA; and (vi) IPI-493+SUN. RESULTS: Treatment with IPI-493 resulted in tumor growth stabilization, variable proliferation arrest, induction of apoptosis and necrosis, and downregulation of KIT and its signaling cascade, especially in the GIST-BOE model. Significant reduction of vessel density was observed with IPI-493 treatment, and was equal to SUN treatment in GIST-PSW and GIST-BOE xenografts. IPI-493 treatment effects were enhanced in combination with TKIs, especially with IPI-493+SUN. In our hands, IPI-493 showed dose-dependent liver damages. CONCLUSIONS: When administered as a single agent in a xenograft model, the HSP90 inhibitor IPI-493 has consistent antitumor activity and induces KIT downregulation in GISTs with heterogeneous KIT mutations. IPI-493 synergizes with TKIs that are commonly used for the treatment of advanced or IMA-resistant GIST. The antitumor response of IPI-493 is particularly enhanced in combination with SUN.
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Benzoquinonas/uso terapéutico , Neoplasias Gastrointestinales/tratamiento farmacológico , Tumores del Estroma Gastrointestinal/tratamiento farmacológico , Proteínas HSP90 de Choque Térmico/antagonistas & inhibidores , Lactamas Macrocíclicas/uso terapéutico , Proteínas Proto-Oncogénicas c-kit/genética , Factor de Células Madre/efectos de los fármacos , Animales , Antineoplásicos/administración & dosificación , Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Apoptosis/efectos de los fármacos , Benzamidas , Benzoquinonas/administración & dosificación , Benzoquinonas/farmacología , Línea Celular Tumoral , Neoplasias Gastrointestinales/genética , Neoplasias Gastrointestinales/patología , Tumores del Estroma Gastrointestinal/genética , Tumores del Estroma Gastrointestinal/patología , Humanos , Mesilato de Imatinib , Indoles/administración & dosificación , Indoles/farmacología , Indoles/uso terapéutico , Lactamas Macrocíclicas/administración & dosificación , Lactamas Macrocíclicas/farmacología , Ratones , Ratones Desnudos , Piperazinas/administración & dosificación , Piperazinas/farmacología , Piperazinas/uso terapéutico , Pirimidinas/administración & dosificación , Pirimidinas/farmacología , Pirimidinas/uso terapéutico , Pirroles/administración & dosificación , Pirroles/farmacología , Pirroles/uso terapéutico , Factor de Células Madre/genética , Factor de Células Madre/metabolismo , Sunitinib , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
The activity of the receptor tyrosine kinase KIT is crucial for gastrointestinal stromal tumor (GIST) growth and survival. Imatinib and sunitinib are very effective in advanced GIST, but have no curative potential. The observation that heat shock protein 90 (HSP90) inhibition results in KIT degradation prompted us to assess the efficacy of the HSP90 inhibitor retaspimycin hydrochloride (IPI-504) alone or in combination with imatinib or sunitinib in two GIST xenografts with distinctive KIT mutations. Nude mice were grafted with human GIST carrying KIT exon 13 (GIST-882; n = 59) or exon 11 (GIST-PSW; n = 44) mutations and dosed with imatinib (50 mg/kg twice daily), sunitinib (40 mg/kg once daily), IPI-504 (100 mg/kg 3 times per week), IPI-504 + imatinib, or IPI-504 + sunitinib. We evaluated tumor volume, proliferation and apoptosis, KIT expression and activation, as well as adverse events during treatment. Treatment with IPI-504 alone resulted in tumor regression, proliferation arrest, and induction of tumor necrosis. We documented downregulation of KIT and its signaling cascade in IPI-504-treated animals. Treatment effects were enhanced by combining IPI-504 with imatinib or sunitinib. On histologic examination, liver damage was frequently observed in animals exposed to combination treatments. In conclusion, IPI-504 shows consistent antitumor activity and induces KIT downregulation in GIST, as a single agent, and is more potent in combination with imatinib or sunitinib. The sequence of drug administration in the combination arms warrants further studies.
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Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Benzoquinonas/farmacología , Tumores del Estroma Gastrointestinal/tratamiento farmacológico , Proteínas HSP90 de Choque Térmico/antagonistas & inhibidores , Lactamas Macrocíclicas/farmacología , Proteínas Proto-Oncogénicas c-kit/metabolismo , Animales , Benzamidas , Benzoquinonas/administración & dosificación , Procesos de Crecimiento Celular/efectos de los fármacos , Línea Celular Tumoral , Femenino , Tumores del Estroma Gastrointestinal/genética , Tumores del Estroma Gastrointestinal/metabolismo , Tumores del Estroma Gastrointestinal/patología , Proteínas HSP90 de Choque Térmico/metabolismo , Humanos , Mesilato de Imatinib , Indoles/administración & dosificación , Lactamas Macrocíclicas/administración & dosificación , Ratones , Ratones Desnudos , Piperazinas/administración & dosificación , Proteínas Proto-Oncogénicas c-kit/genética , Pirimidinas/administración & dosificación , Pirroles/administración & dosificación , Sunitinib , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Abstract A phase 1 study of IPI-504 (retaspimycin hydrochloride) administered intravenously twice weekly for 2 weeks at 22.5, 45, 90, 150, 225, 300 or 400 mg/m(2) followed by 10 days off-treatment was conducted to determine the safety and maximum tolerated dose (MTD) of IPI-504 in patients with relapsed or relapsed/refractory multiple myeloma (MM). Anti-tumor activity and pharmacokinetics were also evaluated. Eighteen patients (mean age 60.5 years; median 9 prior therapies) were enrolled. No dose-limiting toxicities (DLTs) were reported for IPI-504 doses up to 400 mg/m(2). The most common treatment-related adverse event was grade 1 infusion site pain (four patients). All other treatment-related events were assessed as grade 1 or 2 in severity. The area under the curve (AUC) increased with increasing dose, and the mean half-life was approximately 2-4 h for IPI-504 and its metabolites. Four patients had stable disease, demonstrating modest single-agent activity in relapsed or relapsed/refractory MM.