RESUMEN
The L265P somatic mutation in the Myeloid Differentiation Primary Response 88 (MYD88) gene is a recurrent mutation in chronic lymphocytic leukaemia (CLL). This mutation has functional effects in various haematological malignancies but its role in CLL remains to be fully elucidated. Here, we report that MYD88 L265P mutations are associated with mutated immunoglobulin heavy-chain gene (IGHV-M) status and that among IGHV-M patients, the presence of MYD88 L265P is associated with younger age at diagnosis. Using microarray and RNA-Seq gene expression analysis, we further observe that the MYD88 L265P mutation is associated with a distinctive gene expression signature that predicts both failure-free survival and overall survival. This association was validated in an independent cohort of patients. To determine whether MYD88 L265P mutations can be therapeutically exploited in CLL, we treated primary cells with an inhibitor of interleukin 1 receptor-associated kinase 4 (IRAK4), a critical effector of the MYD88 pathway. IRAK4 inhibition decreased downstream nuclear factor-κB signalling and cell viability in CLL cells, indicating the potential of the MYD88 pathway as a therapeutic target in CLL.
Asunto(s)
Leucemia Linfocítica Crónica de Células B/genética , Factor 88 de Diferenciación Mieloide/genética , Adulto , Anciano , Estudios de Cohortes , Citocinas/biosíntesis , Femenino , Genes de las Cadenas Pesadas de las Inmunoglobulinas , Humanos , Quinasas Asociadas a Receptores de Interleucina-1/antagonistas & inhibidores , Leucemia Linfocítica Crónica de Células B/tratamiento farmacológico , Leucemia Linfocítica Crónica de Células B/metabolismo , Masculino , Persona de Mediana Edad , Terapia Molecular Dirigida , Mutación , Factor 88 de Diferenciación Mieloide/metabolismo , Pronóstico , Transducción de Señal , TranscriptomaRESUMEN
Enhancer profiling is a powerful approach for discovering cis-regulatory elements that define the core transcriptional regulatory circuits of normal and malignant cells. Gene control through enhancer activity is often dominated by a subset of lineage-specific transcription factors. By integrating measures of chromatin accessibility and enrichment for H3K27 acetylation, we have generated regulatory landscapes of chronic lymphocytic leukemia (CLL) samples and representative cell lines. With super enhancer-based modeling of regulatory circuits and assessments of transcription factor dependencies, we discover that the essential super enhancer factor PAX5 dominates CLL regulatory nodes and is essential for CLL cell survival. Targeting enhancer signaling via BET bromodomain inhibition disrupts super enhancer-dependent gene expression with selective effects on CLL core regulatory circuitry, conferring potent anti-tumor activity.
Asunto(s)
Cromatina/genética , Elementos de Facilitación Genéticos/genética , Regulación Leucémica de la Expresión Génica/genética , Leucemia Linfocítica Crónica de Células B/genética , Acetilación , Animales , Azepinas/farmacología , Línea Celular Tumoral , Cromatina/efectos de los fármacos , Cromatina/metabolismo , Regulación Leucémica de la Expresión Génica/efectos de los fármacos , Histonas/metabolismo , Humanos , Leucemia Linfocítica Crónica de Células B/tratamiento farmacológico , Leucemia Linfocítica Crónica de Células B/metabolismo , Ratones Noqueados , Factor de Transcripción PAX5/genética , Factor de Transcripción PAX5/metabolismo , Unión Proteica , Proteínas/antagonistas & inhibidores , Proteínas/genética , Proteínas/metabolismo , Triazoles/farmacología , Ensayos Antitumor por Modelo de Xenoinjerto/métodosRESUMEN
Diagnostic and prognostic evaluation of chronic lymphocytic leukemia (CLL) involves blood cell counts, immunophenotyping, IgVH mutation status, and cytogenetic analyses. We generated B-cell associated gene-signatures (BAGS) based on six naturally occurring B-cell subsets within normal bone marrow. Our hypothesis is that by segregating CLL according to BAGS, we can identify subtypes with prognostic implications in support of pathogenetic value of BAGS. Microarray-based gene-expression samples from eight independent CLL cohorts (1,024 untreated patients) were BAGS-stratified into pre-BI, pre-BII, immature, naïve, memory, or plasma cell subtypes; the majority falling within the memory (24.5-45.8%) or naïve (14.5-32.3%) categories. For a subset of CLL patients (n = 296), time to treatment (TTT) was shorter amongst early differentiation subtypes (pre-BI/pre-BII/immature) compared to late subtypes (memory/plasma cell, HR: 0.53 [0.35-0.78]). Particularly, pre-BII subtype patients had the shortest TTT among all subtypes. Correlates derived for BAGS subtype and IgVH mutation (n = 405) revealed an elevated mutation frequency in late vs. early subtypes (71% vs. 45%, P < .001). Predictions for BAGS subtype resistance towards rituximab and cyclophosphamide varied for rituximab, whereas all subtypes were sensitive to cyclophosphamide. This study supports our hypothesis that BAGS-subtyping may be of tangible prognostic and pathogenetic value for CLL patients.
Asunto(s)
Subgrupos de Linfocitos B/metabolismo , Médula Ósea/metabolismo , Leucemia Linfocítica Crónica de Células B/clasificación , Leucemia Linfocítica Crónica de Células B/metabolismo , Adulto , Anciano , Anciano de 80 o más Años , Antineoplásicos Alquilantes/uso terapéutico , Antineoplásicos Inmunológicos/uso terapéutico , Ciclofosfamida/uso terapéutico , Resistencia a Antineoplásicos/fisiología , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Leucemia Linfocítica Crónica de Células B/terapia , Masculino , Análisis por Micromatrices , Persona de Mediana Edad , Pronóstico , Prueba de Estudio Conceptual , Estudios Retrospectivos , Rituximab/uso terapéutico , Análisis de Supervivencia , Tiempo de TratamientoRESUMEN
The treatment of chronic lymphocytic leukaemia (CLL) has been improved by introduction of monoclonal antibodies (mAbs) that exert their effect through secondary effector mechanisms. CLL cells are characterized by expression of CD5 and CD23 along with CD19 and CD20, hence anti-CD5 Abs that engage secondary effector functions represent an attractive opportunity for CLL treatment. Here, a repertoire of mAbs against human CD5 was generated and tested for ability to induce complement-dependent cytotoxicity (CDC) and antibody-dependent cell-mediated cytotoxicity (ADCC) both as single mAbs and combinations of two mAbs against non-overlapping epitopes on human CD5. The results demonstrated that combinations of two mAbs significantly increased the level of CDC compared to the single mAbs, while no enhancement of ADCC was seen with anti-CD5 mAb combinations. High levels of CDC and ADCC correlated with low levels of Ab-induced CD5 internalization and degradation. Importantly, an anti-CD5 mAb combination enhanced CDC of CLL cells when combined with the anti-CD20 mAbs rituximab and ofatumumab as well as with the anti-CD52 mAb alemtuzumab. These results suggest that an anti-CD5 mAb combination inducing CDC and ADCC may be effective alone, in combination with mAbs against other targets or combined with chemotherapy for CLL and other CD5-expressing haematological or lymphoid malignancies.
Asunto(s)
Anticuerpos Monoclonales/farmacología , Antígenos CD5/metabolismo , Proteínas del Sistema Complemento/inmunología , Citotoxicidad Inmunológica/efectos de los fármacos , Leucemia Linfocítica Crónica de Células B/inmunología , Leucemia Linfocítica Crónica de Células B/metabolismo , Adulto , Anciano , Anciano de 80 o más Años , Alemtuzumab , Anticuerpos Monoclonales/inmunología , Anticuerpos Monoclonales Humanizados/farmacología , Citotoxicidad Celular Dependiente de Anticuerpos/efectos de los fármacos , Citotoxicidad Celular Dependiente de Anticuerpos/inmunología , Antígenos CD20/inmunología , Antígenos CD20/metabolismo , Antígenos CD5/inmunología , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Aberraciones Cromosómicas , Sinergismo Farmacológico , Femenino , Humanos , Leucemia Linfocítica Crónica de Células B/genética , Masculino , Persona de Mediana EdadRESUMEN
The transcription factor STAT5 is an essential mediator of the pathogenesis of chronic myelogenous leukemia (CML). In CML, the BCR/ABL fusion kinase causes the constitutive activation of STAT5, thereby driving the expression of genes promoting survival. BCR/ABL kinase inhibitors have become the mainstay of therapy for CML, although CML cells can develop resistance through mutations in BCR/ABL. To overcome this problem, we used a cell-based screen to identify drugs that inhibit STAT-dependent gene expression. Using this approach, we identified the psychotropic drug pimozide as a STAT5 inhibitor. Pimozide decreases STAT5 tyrosine phosphorylation, although it does not inhibit BCR/ABL or other tyrosine kinases. Furthermore, pimozide decreases the expression of STAT5 target genes and induces cell cycle arrest and apoptosis in CML cell lines. Pimozide also selectively inhibits colony formation of CD34(+) bone marrow cells from CML patients. Importantly, pimozide induces similar effects in the presence of the T315I BCR/ABL mutation that renders the kinase resistant to presently available inhibitors. Simultaneously inhibiting STAT5 with pimozide and the kinase inhibitors imatinib or nilotinib shows enhanced effects in inhibiting STAT5 phosphorylation and in inducing apoptosis. Thus, targeting STAT5 may be an effective strategy for the treatment of CML and other myeloproliferative diseases.
Asunto(s)
Resistencia a Antineoplásicos/efectos de los fármacos , Leucemia Mielógena Crónica BCR-ABL Positiva/patología , Pimozida/farmacología , Inhibidores de Proteínas Quinasas/uso terapéutico , Factor de Transcripción STAT5/antagonistas & inhibidores , Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Supervivencia Celular/efectos de los fármacos , Regulación hacia Abajo/efectos de los fármacos , Evaluación Preclínica de Medicamentos , Proteínas de Fusión bcr-abl/antagonistas & inhibidores , Humanos , Células K562 , Leucemia Mielógena Crónica BCR-ABL Positiva/tratamiento farmacológico , Modelos Biológicos , Terapia Molecular Dirigida , Pimozida/uso terapéutico , Insuficiencia del Tratamiento , Células Tumorales CultivadasRESUMEN
The immunogenicity of therapeutic Abs is a concern as anti-drug Abs may impact negatively on the pharmacodynamics and safety profile of Ab drugs. The factors governing induction of anti-drug Abs are not fully understood. In this study, we describe a model based on mouse-human chimeric Abs for the study of Ab immunogenicity in vivo. Six chimeric Abs containing human V regions and mouse C regions were generated from six human anti-Rhesus D Abs and the Ag-binding characteristics of the parental human Abs were retained. Analysis of the immune response toward the individual chimeric Abs revealed the induction of anti-variable domain Abs including anti-idiotypic Abs against some of these, thereby demonstrating the applicability of the model for studying anti-drug Ab responses in vivo. Immunization of BALB/c, C57, and outbred NMRI mice with a polyclonal composition consisting of all six chimeric Abs demonstrated that the immunogenicity of the individual Abs was haplotype dependent. Chimeric Abs, which were nonimmunogenic when administered individually, did not become immunogenic as part of the polyclonal composition, implying the absence of epitope spreading. Ex vivo Ab-binding studies established a clear correlation between the level of immunogenicity of the Abs comprised in the composition and the impact on the pharmacology of the Abs. These analyses demonstrate that under these conditions this polyclonal Ab composition was generally less susceptible to blocking Abs than the respective mAbs.