RESUMO
Recent discoveries suggest that the critical events leading to the anti-proliferative activity of the IMiD immunomodulatory agents lenalidomide and pomalidomide in multiple myeloma (MM) cells are initiated by Cereblon-dependent ubiquitination and proteasomal degradation of substrate proteins Ikaros (IKZF1) and Aiolos (IKZF3). By performing kinetic analyses, we found that the downregulation or proteasomal degradation of Ikaros and Aiolos led to specific and sequential downregulation of c-Myc followed by IRF4 and subsequent growth inhibition and apoptosis. Notably, to ensure growth inhibition and cell death, sustained downregulation of Ikaros and Aiolos, c-Myc or IRF4 expression was required. In addition, we found that the half-maximal rate, rather than the final extent of Ikaros and Aiolos degradation, correlated to the relative efficacy of growth inhibition by lenalidomide or pomalidomide. Finally, we observed that all four transcription factors were elevated in primary MM samples compared with normal plasma cells. Taken together, our results suggest a functional link between Ikaros and Aiolos, and the pathological dysregulation of c-Myc and IRF4, and provide a new mechanistic understanding of the relative efficacy of lenalidomide and pomalidomide based on the kinetics of substrate degradation and downregulation of their downstream targets.
Assuntos
Antineoplásicos/farmacologia , Fator de Transcrição Ikaros/metabolismo , Fatores Reguladores de Interferon/metabolismo , Mieloma Múltiplo/metabolismo , Proteínas Proto-Oncogênicas c-myc/metabolismo , Apoptose/efeitos dos fármacos , Apoptose/fisiologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/fisiologia , Regulação para Baixo , Citometria de Fluxo , Técnicas de Silenciamento de Genes , Humanos , Immunoblotting , Imuno-Histoquímica , Lenalidomida , Mieloma Múltiplo/patologia , RNA Interferente Pequeno , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Talidomida/análogos & derivados , Talidomida/farmacologiaRESUMO
The development of resistance to therapy is unavoidable in the history of multiple myeloma patients. Therefore, the study of its characteristics and mechanisms is critical in the search for novel therapeutic approaches to overcome it. This effort is hampered by the absence of appropriate preclinical models, especially those mimicking acquired resistance. Here we present an in vivo model of acquired resistance based on the continuous treatment of mice bearing subcutaneous MM1S plasmacytomas. Xenografts acquired resistance to two generations of immunomodulatory drugs (IMiDs; lenalidomide and pomalidomide) in combination with dexamethasone, that was reversible after a wash-out period. Furthermore, lenalidomide-dexamethasone (LD) or pomalidomide-dexamethasone (PD) did not display cross-resistance, which could be due to the differential requirements of the key target Cereblon and its substrates Aiolos and Ikaros observed in cells resistant to each combination. Differential gene expression profiles of LD and PD could also explain the absence of cross-resistance. Onset of resistance to both combinations was accompanied by upregulation of the mitogen-activated protein kinase/extracellular signal-regulated kinase (ERK) kinase (MEK)/ERK pathway and addition of selumetinib, a small-molecule MEK inhibitor, could resensitize resistant cells. Our results provide insights into the mechanisms of acquired resistance to LD and PD combinations and offer possible therapeutic approaches to addressing IMiD resistance in the clinic.
Assuntos
Antineoplásicos/farmacologia , Dexametasona/farmacologia , Regulação Neoplásica da Expressão Gênica , Plasmocitoma/tratamento farmacológico , Talidomida/análogos & derivados , Proteínas Adaptadoras de Transdução de Sinal , Animais , Apoptose/efeitos dos fármacos , Benzimidazóis/farmacologia , Linhagem Celular Tumoral , Modelos Animais de Doenças , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/genética , Quimioterapia Combinada , MAP Quinases Reguladas por Sinal Extracelular/genética , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Humanos , Fator de Transcrição Ikaros/genética , Fator de Transcrição Ikaros/metabolismo , Lenalidomida , Camundongos , Mieloma Múltiplo/tratamento farmacológico , Mieloma Múltiplo/genética , Mieloma Múltiplo/metabolismo , Mieloma Múltiplo/patologia , Transplante de Neoplasias , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Plasmocitoma/genética , Plasmocitoma/metabolismo , Plasmocitoma/patologia , Transdução de Sinais , Talidomida/farmacologia , Transativadores/genética , Transativadores/metabolismoRESUMO
Resistance of myeloma to lenalidomide is an emerging clinical problem, and though it has been associated in part with activation of Wnt/ß-catenin signaling, the mediators of this phenotype remained undefined. Lenalidomide-resistant models were found to overexpress the hyaluronan (HA)-binding protein CD44, a downstream Wnt/ß-catenin transcriptional target. Consistent with a role of CD44 in cell adhesion-mediated drug resistance (CAM-DR), lenalidomide-resistant myeloma cells were more adhesive to bone marrow stroma and HA-coated plates. Blockade of CD44 with monoclonal antibodies, free HA or CD44 knockdown reduced adhesion and sensitized to lenalidomide. Wnt/ß-catenin suppression by FH535 enhanced the activity of lenalidomide, as did interleukin-6 neutralization with siltuximab. Notably, all-trans retinoic acid (ATRA) downregulated total ß-catenin, cell-surface and total CD44, reduced adhesion of lenalidomide-resistant myeloma cells and enhanced the activity of lenalidomide in a lenalidomide-resistant in vivo murine xenograft model. Finally, ATRA sensitized primary myeloma samples from patients that had relapsed and/or refractory disease after lenalidomide therapy to this immunomodulatory agent ex vivo. Taken together, our findings support the hypotheses that CD44 and CAM-DR contribute to lenalidomide resistance in multiple myeloma, that CD44 should be evaluated as a putative biomarker of sensitivity to lenalidomide, and that ATRA or other approaches that target CD44 may overcome clinical lenalidomide resistance.
Assuntos
Antineoplásicos/uso terapêutico , Resistencia a Medicamentos Antineoplásicos/genética , Receptores de Hialuronatos/genética , Fatores Imunológicos/uso terapêutico , Mieloma Múltiplo/tratamento farmacológico , Mieloma Múltiplo/genética , Talidomida/análogos & derivados , Animais , Anticorpos Monoclonais/farmacologia , Citotoxicidade Celular Dependente de Anticorpos , Adesão Celular/genética , Linhagem Celular Tumoral , Modelos Animais de Doenças , Expressão Gênica , Humanos , Receptores de Hialuronatos/metabolismo , Ácido Hialurônico/metabolismo , Lenalidomida , Camundongos , Ligação Proteica/efeitos dos fármacos , Talidomida/uso terapêutico , Tretinoína/farmacologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
The ubiquitin-proteasome pathway (UPP) is an attractive chemotherapeutic target due to its intrinsically stringent regulation of cell cycle, pro-survival, and anti-apoptotic regulators that disproportionately favor survival and proliferation in malignant cells. A reversible first-in-class proteasome inhibitor, bortezomib, is Food and Drug Administration approved for multiple myeloma and relapsed/refractory mantle cell lymphoma and has proven to be extremely effective, both as a single agent and in combination. An irreversible second generation proteasome inhibitor, carfilzomib, has shown preclinical effectiveness against hematological and solid malignancies both in vitro and in vivo. Carfilzomib, a peptidyl-epoxyketone functions similarly to bortezomib through primary inhibition of chymotrypsin-like (ChT-L) activity at the b5 subunits of the core 20S proteasome. Carfilzomib is also currently achieving successful response rates within the clinical setting. In addition to conventional proteasome inhibitors, a novel approach may be to specifically target the hematological-specific immunoproteasome, thereby increasing overall effectiveness and reducing negative off-target effects. The immunoproteasome-specific inhibitor, IPSI-001, was shown to have inhibitory preference over the constitutive proteasome, and display enhanced efficiency of apoptotic induction of tumor cells from a hematologic origin. Herein, we discuss the preclinical and clinical development of carfilzomib and explore the potential of immunoproteasome-specific inhibitors, like IPSI-001, as a rational approach to exclusively target hematological malignancies.