RESUMEN
Despite significant progress in treating chronic lymphocytic leukaemia (CLL), resistance to therapy remains challenging. NOTCH1 activation, common in CLL, confers adverse prognosis. This study explores the impact of NOTCH1 signalling on venetoclax sensitivity in vitro. Although NOTCH1 activation minimally impaired the susceptibility of CLL cells to venetoclax, ex vivo cell competition studies reveal that cells with constitutive NOTCH1 activation outgrew their wild-type counterparts in the presence of ongoing venetoclax exposure. Our findings suggest that while NOTCH1 activation is insufficient to confer venetoclax refractoriness, there is enhanced potential for cells with NOTCH1 activation to escape and thus become fully resistant to venetoclax.
RESUMEN
Venetoclax is an effective treatment for certain blood cancers, such as chronic lymphocytic leukemia (CLL) and acute myeloid leukemia (AML). However, most patients relapse while on venetoclax and further treatment options are limited. Combining venetoclax with immunotherapies is an attractive approach; however, a detailed understanding of how venetoclax treatment impacts normal immune cells in patients is lacking. In this study, we performed deep profiling of peripheral blood (PB) cells from patients with CLL and AML before and after short-term treatment with venetoclax using mass cytometry (cytometry by time of flight) and found no impact on the concentrations of key T-cell subsets or their expression of checkpoint molecules. We also analyzed PB from patients with breast cancer receiving venetoclax long-term using a single-cell multiomics approach (cellular indexing of transcriptomes and epitopes by sequencing) and functional assays. We found significant depletion of B-cell populations with low expression of MCL-1 relative to other immune cells, attended by extensive transcriptomic changes. By contrast, there was less impact on circulating T cells and natural killer (NK) cells, with no changes in their subset composition, transcriptome, or function following venetoclax treatment. Our data indicate that venetoclax has minimal impact on circulating T or NK cells, supporting the rationale of combining this BH3 mimetic drug with cancer immunotherapies for more durable antitumor responses.
Asunto(s)
Leucemia Linfocítica Crónica de Células B , Leucemia Mieloide Aguda , Humanos , Leucemia Linfocítica Crónica de Células B/tratamiento farmacológico , Células Asesinas Naturales , Leucemia Mieloide Aguda/tratamiento farmacológico , Compuestos Bicíclicos Heterocíclicos con Puentes/farmacología , Compuestos Bicíclicos Heterocíclicos con Puentes/uso terapéuticoRESUMEN
Preventing or overcoming resistance to the Bcl-2 inhibitor venetoclax is an emerging unmet clinical need in patients with chronic lymphocytic leukaemia (CLL). The upregulation of anti-apoptotic Bcl-2 members through signalling pathways within the tumor microenvironment appears as a major factor leading to resistance to venetoclax. Previously, we reported that T cells can drive resistance through CD40 and non-canonical NF-κB activation and subsequent Bcl-XL induction. Moreover, the T cell-derived cytokines IL-21 and IL-4 differentially affect Bcl-XL expression and sensitivity to venetoclax via unknown mechanisms. Here, we mechanistically dissected how Bcl-XL is regulated in the context of JAK-STAT signalling in primary CLL. First, we demonstrated a clear antagonistic role of IL-21/STAT3 signalling in the NF-κB-mediated expression of Bcl-XL, whereas IL-4/STAT6 further promoted the expression of Bcl-XL. In comparison, Bfl-1, another NF-κB target, was not differentially affected by either cytokine. Second, STAT3 and STAT6 affected Bcl-XL transcription by binding to its promoter without disrupting the DNA-binding activity of NF-κB. Third, in situ proximity ligation assays (isPLAs) indicated crosstalk between JAK-STAT signalling and NF-κB, in which STAT3 inhibited canonical NF-κB by accelerating nuclear export, and STAT6 promoted non-canonical NF-κB. Finally, NF-κB inducing kinase (NIK) inhibition interrupted the NF-κB/STAT crosstalk and resensitized CLL cells to venetoclax. In conclusion, we uncovered distinct crosstalk mechanisms that shape the NF-κB response in CLL towards venetoclax sensitivity or resistance via Bcl-XL, thereby revealing new potential therapeutic targets.
Asunto(s)
Leucemia Linfocítica Crónica de Células B , FN-kappa B , Humanos , Apoptosis , Resistencia a Antineoplásicos , Interleucina-4/uso terapéutico , Leucemia Linfocítica Crónica de Células B/tratamiento farmacológico , Leucemia Linfocítica Crónica de Células B/patología , FN-kappa B/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Microambiente TumoralRESUMEN
Venetoclax (VEN) inhibits the prosurvival protein BCL2 to induce apoptosis and is a standard therapy for chronic lymphocytic leukemia (CLL), delivering high complete remission rates and prolonged progression-free survival in relapsed CLL but with eventual loss of efficacy. A spectrum of subclonal genetic changes associated with VEN resistance has now been described. To fully understand clinical resistance to VEN, we combined single-cell short- and long-read RNA-sequencing to reveal the previously unappreciated scale of genetic and epigenetic changes underpinning acquired VEN resistance. These appear to be multilayered. One layer comprises changes in the BCL2 family of apoptosis regulators, especially the prosurvival family members. This includes previously described mutations in BCL2 and amplification of the MCL1 gene but is heterogeneous across and within individual patient leukemias. Changes in the proapoptotic genes are notably uncommon, except for single cases with subclonal losses of BAX or NOXA. Much more prominent was universal MCL1 gene upregulation. This was driven by an overlying layer of emergent NF-κB (nuclear factor kappa B) activation, which persisted in circulating cells during VEN therapy. We discovered that MCL1 could be a direct transcriptional target of NF-κB. Both the switch to alternative prosurvival factors and NF-κB activation largely dissipate following VEN discontinuation. Our studies reveal the extent of plasticity of CLL cells in their ability to evade VEN-induced apoptosis. Importantly, these findings pinpoint new approaches to circumvent VEN resistance and provide a specific biological justification for the strategy of VEN discontinuation once a maximal response is achieved rather than maintaining long-term selective pressure with the drug.
Asunto(s)
Antineoplásicos , Leucemia Linfocítica Crónica de Células B , Humanos , Leucemia Linfocítica Crónica de Células B/tratamiento farmacológico , Leucemia Linfocítica Crónica de Células B/genética , Leucemia Linfocítica Crónica de Células B/metabolismo , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , FN-kappa B , Resistencia a Antineoplásicos/genética , Compuestos Bicíclicos Heterocíclicos con Puentes/farmacología , Compuestos Bicíclicos Heterocíclicos con Puentes/uso terapéutico , Recurrencia , Antineoplásicos/uso terapéuticoRESUMEN
Normalization of single cell RNA-seq data remains a challenging task. The performance of different methods can vary greatly between datasets when unwanted factors and biology are associated. Most normalization methods also only remove the effects of unwanted variation for the cell embedding but not from gene-level data typically used for differential expression (DE) analysis to identify marker genes. We propose RUV-III-NB, a method that can be used to remove unwanted variation from both the cell embedding and gene-level counts. Using pseudo-replicates, RUV-III-NB explicitly takes into account potential association with biology when removing unwanted variation. The method can be used for both UMI or read counts and returns adjusted counts that can be used for downstream analyses such as clustering, DE and pseudotime analyses. Using published datasets with different technological platforms, kinds of biology and levels of association between biology and unwanted variation, we show that RUV-III-NB manages to remove library size and batch effects, strengthen biological signals, improve DE analyses, and lead to results exhibiting greater concordance with independent datasets of the same kind. The performance of RUV-III-NB is consistent and is not sensitive to the number of factors assumed to contribute to the unwanted variation.
Asunto(s)
Perfilación de la Expresión Génica , Perfilación de la Expresión Génica/métodos , Biblioteca de Genes , RNA-Seq , Análisis de Secuencia de ARN/métodosRESUMEN
The genomic landscape of resistance to targeted agents (TAs) used as monotherapy in chronic lymphocytic leukemia (CLL) is complex and often heterogeneous at the patient level. To gain insight into the clonal architecture of acquired genomic resistance to Bruton tyrosine kinase (BTK) inhibitors and B-cell lymphoma 2 (BCL2) inhibitors in CLL, particularly in patients carrying multiple resistance mutations, we performed targeted single-cell DNA sequencing of 8 patients who developed progressive disease (PD) on TAs (either class). In all cases, analysis of single-cell architecture revealed mutual exclusivity between multiple resistance mutations to the same TA class, variable clonal co-occurrence of multiple mutations affecting different TAs in patients exposed to both classes, and a phenomenon of multiple independent emergences of identical nucleotide changes leading to canonical resistance mutations. We also report the first observation of established BCL2 resistance mutations in a patient with mantle cell lymphoma (MCL) following PD on sequential monotherapy, implicating BCL2 as a venetoclax resistance mechanism in MCL. Taken together, these data reveal the significant clonal complexity of CLL and MCL progression on TAs at the nucleotide level and confirm the presence of multiple, clonally independent, mechanisms of TA resistance within each individual disease context.
Asunto(s)
Antineoplásicos , Leucemia Linfocítica Crónica de Células B , Linfoma de Células del Manto , Adulto , Antineoplásicos/uso terapéutico , Humanos , Leucemia Linfocítica Crónica de Células B/tratamiento farmacológico , Leucemia Linfocítica Crónica de Células B/genética , Leucemia Linfocítica Crónica de Células B/patología , Linfoma de Células del Manto/tratamiento farmacológico , Mutación , Proteínas Proto-Oncogénicas c-bcl-2/genéticaRESUMEN
A modified Chromium 10x droplet-based protocol that subsamples cells for both short-read and long-read (nanopore) sequencing together with a new computational pipeline (FLAMES) is developed to enable isoform discovery, splicing analysis, and mutation detection in single cells. We identify thousands of unannotated isoforms and find conserved functional modules that are enriched for alternative transcript usage in different cell types and species, including ribosome biogenesis and mRNA splicing. Analysis at the transcript level allows data integration with scATAC-seq on individual promoters, improved correlation with protein expression data, and linked mutations known to confer drug resistance to transcriptome heterogeneity.
Asunto(s)
Secuenciación de Nanoporos/métodos , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Empalme Alternativo , Animales , Exones , Perfilación de la Expresión Génica/métodos , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Ratones , Empalme del ARN , ARN Mensajero , TranscriptomaRESUMEN
Selective targeting of BCL-2 with the BH3-mimetic venetoclax has been a transformative treatment for patients with various leukemias. TP-53 controls apoptosis upstream of where BCL-2 and its prosurvival relatives, such as MCL-1, act. Therefore, targeting these prosurvival proteins could trigger apoptosis across diverse blood cancers, irrespective of TP53 mutation status. Indeed, targeting BCL-2 has produced clinically relevant responses in blood cancers with aberrant TP-53. However, in our study, TP53-mutated or -deficient myeloid and lymphoid leukemias outcompeted isogenic controls with intact TP-53, unless sufficient concentrations of BH3-mimetics targeting BCL-2 or MCL-1 were applied. Strikingly, tumor cells with TP-53 dysfunction escaped and thrived over time if inhibition of BCL-2 or MCL-1 was sublethal, in part because of an increased threshold for BAX/BAK activation in these cells. Our study revealed the key role of TP-53 in shaping long-term responses to BH3-mimetic drugs and reconciled the disparate pattern of initial clinical response to venetoclax, followed by subsequent treatment failure among patients with TP53-mutant chronic lymphocytic leukemia or acute myeloid leukemia. In contrast to BH3-mimetics targeting just BCL-2 or MCL-1 at doses that are individually sublethal, a combined BH3-mimetic approach targeting both prosurvival proteins enhanced lethality and durably suppressed the leukemia burden, regardless of TP53 mutation status. Our findings highlight the importance of using sufficiently lethal treatment strategies to maximize outcomes of patients with TP53-mutant disease. In addition, our findings caution against use of sublethal BH3-mimetic drug regimens that may enhance the risk of disease progression driven by emergent TP53-mutant clones.
Asunto(s)
Antineoplásicos/farmacología , Proteínas Reguladoras de la Apoptosis/antagonistas & inhibidores , Apoptosis/efectos de los fármacos , Compuestos Bicíclicos Heterocíclicos con Puentes/farmacología , Indolizinas/farmacología , Isoquinolinas/farmacología , Leucemia Linfocítica Crónica de Células B/tratamiento farmacológico , Leucemia Mieloide Aguda/tratamiento farmacológico , Morfolinas/farmacología , Proteínas de Neoplasias/fisiología , Fragmentos de Péptidos/antagonistas & inhibidores , Proteínas Proto-Oncogénicas/antagonistas & inhibidores , Sulfonamidas/farmacología , Proteína p53 Supresora de Tumor/fisiología , Animales , Antineoplásicos/administración & dosificación , Antineoplásicos/uso terapéutico , Apoptosis/fisiología , Proteínas Reguladoras de la Apoptosis/fisiología , Compuestos Bicíclicos Heterocíclicos con Puentes/administración & dosificación , Compuestos Bicíclicos Heterocíclicos con Puentes/uso terapéutico , Sistemas CRISPR-Cas , Línea Celular Tumoral , Daño del ADN , Genes p53 , Humanos , Indolizinas/uso terapéutico , Subunidad alfa del Receptor de Interleucina-2/deficiencia , Isoquinolinas/uso terapéutico , Leucemia Mieloide Aguda/patología , Leucemia Mieloide Aguda/terapia , Ratones , Ratones Endogámicos NOD , Ratones Noqueados , Ratones SCID , Morfolinas/uso terapéutico , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/antagonistas & inhibidores , Proteínas de Neoplasias/antagonistas & inhibidores , Fosforilación Oxidativa/efectos de los fármacos , Proteínas Proto-Oncogénicas c-bcl-2/antagonistas & inhibidores , Sulfonamidas/administración & dosificación , Sulfonamidas/uso terapéutico , Proteína p53 Supresora de Tumor/deficiencia , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
In chronic lymphocytic leukemia (CLL), the lymph node (LN) microenvironment delivers critical survival signals by inducing the expression of anti-apoptotic Bcl-2 members Bcl-XL, Bfl-1, and Mcl-1, resulting in apoptosis blockade. We determined previously that resistance against various drugs, among which is the clinically applied BH3 mimetic venetoclax, is dominated by upregulation of the anti-apoptotic regulator Bcl-XL. Direct clinical targeting of Bcl-XL by, e.g., Navitoclax is however not desirable due to induction of thrombocytopenia. Since the actual regulation of Bcl-XL in CLL in the context of the LN microenvironment is not well elucidated, we investigated various candidate LN signals to drive Bcl-XL expression. We found a dominance for NF-κB signaling upon CD40 stimulation, which results in activation of both the canonical and non-canonical NF-κB signaling pathways. We demonstrate that expression of Bcl-XL is first induced by the canonical NF-κB pathway, and subsequently boosted and continued via non-canonical NF-κB signaling through stabilization of NIK. NF-κB subunits p65 and p52 can both bind to the Bcl-XL promoter and activate transcription upon CD40 stimulation. Moreover, canonical NF-κB signaling was correlated with Bfl-1 expression, whereas Mcl-1 in contrast, was not transcriptionally regulated by NF-κB. Finally, we applied a novel compound targeting NIK to selectively inhibit the non-canonical NF-κB pathway and showed that venetoclax-resistant CLL cells were sensitized to venetoclax. In conclusion, protective signals from the CLL microenvironment can be tipped towards apoptosis sensitivity by interfering with non-canonical NF-κB signaling.
Asunto(s)
Leucemia Linfocítica Crónica de Células B/genética , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Proteína bcl-X/metabolismo , Animales , Línea Celular Tumoral , Resistencia a Antineoplásicos , Humanos , Ratones , FN-kappa B/metabolismo , Transducción de Señal , Transfección , Microambiente TumoralRESUMEN
The discovery of the link between defective apoptotic regulation and cancer cell survival engendered the idea of targeting aberrant components of the apoptotic machinery for cancer therapy. The intrinsic pathway of apoptosis is tightly controlled by interactions amongst members of three distinct subgroups of the B-cell lymphoma 2 (BCL2) family of proteins. The pro-survival BCL2 proteins prevent apoptosis by keeping the pro-apoptotic effector proteins BCL2-associated X protein (BAX) and BCL2 homologous antagonist/killer (BAK) in check, while the BH3-only proteins initiate apoptosis by either neutralizing the pro-survival BCL2 proteins or directly activating the pro-apoptotic effector proteins. This tripartite regulatory mechanism is commonly perturbed in B-cell malignancies facilitating cell death evasion. Over the past two decades, structure-based drug discovery has resulted in the development of a series of small molecules that mimic the function of BH3-only proteins called the BH3 mimetics. The most clinically advanced of these is venetoclax, which is a highly selective inhibitor of BCL2 that has transformed the treatment landscape for chronic lymphocytic leukemia (CLL). Other BH3 mimetics, which selectively target myeloid cell leukemia 1 (MCL1) and B-cell lymphoma extra large (BCLxL), are currently under investigation for use in diverse malignancies. Here, we review the current role of BH3 mimetics in the treatment of CLL and other B-cell malignancies and address open questions in this rapidly evolving field.
Asunto(s)
Antineoplásicos/uso terapéutico , Carcinogénesis/patología , Linfoma/tratamiento farmacológico , Linfoma/patología , Proteínas Proto-Oncogénicas c-myc/metabolismo , Proteína Serina-Treonina Quinasas de Interacción con Receptores/deficiencia , Animales , Antineoplásicos/farmacología , Carcinogénesis/metabolismo , Muerte Celular/efectos de los fármacos , Estimación de Kaplan-Meier , Linfoma/metabolismo , Ratones Transgénicos , Proteína Serina-Treonina Quinasas de Interacción con Receptores/metabolismoAsunto(s)
Compuestos Bicíclicos Heterocíclicos con Puentes/uso terapéutico , Leucemia Linfocítica Crónica de Células B/tratamiento farmacológico , Leucemia Linfocítica Crónica de Células B/genética , Tasa de Mutación , Proteínas Proto-Oncogénicas c-bcl-2/genética , Sulfonamidas/uso terapéutico , Sustitución de Aminoácidos/genética , Antineoplásicos/uso terapéutico , Estudios de Cohortes , Análisis Mutacional de ADN , Progresión de la Enfermedad , Resistencia a Antineoplásicos/genética , Frecuencia de los Genes , Glicina/genética , Humanos , Leucemia Linfocítica Crónica de Células B/patología , Modelos Moleculares , Mutación , Proteínas Proto-Oncogénicas c-bcl-2/química , Células Tumorales Cultivadas , Valina/genéticaRESUMEN
Apoptosis-regulating BCL-2 family members, which can promote malignant transformation and resistance to therapy, have become prime therapeutic targets, as illustrated by the striking efficacy in certain lymphoid malignancies of the BCL-2-specific inhibitor venetoclax. In other lymphoid malignancies, however, such as the aggressive mantle cell lymphoma (MCL), cell survival might rely instead or also on BCL-2 relative MCL-1. We have explored MCL-1 as a target for killing MCL cells by both genetic and pharmacologic approaches. In several MCL cell lines, MCL-1 knockout with an inducible CRISPR/Cas9 system triggered spontaneous apoptosis. Accordingly, most MCL cell lines proved sensitive to the specific MCL-1 inhibitor S63845, and MCL-1 inhibition also proved efficacious in an MCL xenograft model. Furthermore, its killing efficacy rose on combination with venetoclax, the BCL-XL-specific inhibitor A-1331852, or Bruton's tyrosine kinase (BTK) inhibitor ibrutinib, which reduced pro-survival signals. We also tested the MCL-1 inhibitor in primary samples from 13 MCL patients, using CD40L-expressing feeder cells to model their microenvironmental support. Notably, all unstimulated primary MCL samples were very sensitive to S63845, but the CD40L stimulation attenuated their sensitivity. Mass cytometric analysis revealed that the stimulation likely conveyed protection by elevating BCL-XL and MCL-1. Accordingly, sensitivity of the CD40L-stimulated cells to S63845 was substantially restored by co-treatment with venetoclax, the BCL-XL-specific inhibitor or ibrutinib. Overall, our findings indicate that MCL-1 is very important for survival of MCL cells and that the MCL-1 inhibitor, both alone and together with ibrutinib, venetoclax or a BCL-XL inhibitor, offers promise for novel improved MCL therapies.
Asunto(s)
Linfoma de Células del Manto/tratamiento farmacológico , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/antagonistas & inhibidores , Pirimidinas/farmacología , Tiofenos/farmacología , Proteína bcl-X/antagonistas & inhibidores , Animales , Antineoplásicos/farmacología , Apoptosis , Compuestos Bicíclicos Heterocíclicos con Puentes/farmacología , Proliferación Celular , Humanos , Linfoma de Células del Manto/metabolismo , Linfoma de Células del Manto/patología , Ratones , Ratones Endogámicos NOD , Ratones SCID , Sulfonamidas/farmacología , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
In this issue of Cancer Cell, Guièza et al. describe that overexpression of the pro-survival protein MCL1 and cellular energy metabolic reprogramming can contribute to resistance to the BCL2 inhibitor venetoclax in patients with chronic lymphocytic leukemia. This adds a new dimension to understanding of secondary clinical resistance to venetoclax.
Asunto(s)
Antineoplásicos , Leucemia Linfocítica Crónica de Células B , Compuestos Bicíclicos Heterocíclicos con Puentes , Humanos , Proteínas Proto-Oncogénicas c-bcl-2 , SulfonamidasRESUMEN
Venetoclax is a first-in-class cancer therapy that interacts with the cellular apoptotic machinery promoting apoptosis. Treatment of patients suffering chronic lymphocytic leukaemia with this BCL-2 antagonist has revealed emergence of a drug-selected BCL-2 mutation (G101V) in some patients failing therapy. To understand the molecular basis of this acquired resistance we describe the crystal structures of venetoclax bound to both BCL-2 and the G101V mutant. The pose of venetoclax in its binding site on BCL-2 reveals small but unexpected differences as compared to published structures of complexes with venetoclax analogues. The G101V mutant complex structure and mutant binding assays reveal that resistance is acquired by a knock-on effect of V101 on an adjacent residue, E152, with venetoclax binding restored by a E152A mutation. This provides a framework for considering analogues of venetoclax that might be effective in combating this mutation.
Asunto(s)
Antineoplásicos/metabolismo , Compuestos Bicíclicos Heterocíclicos con Puentes/metabolismo , Resistencia a Antineoplásicos/genética , Leucemia Linfocítica Crónica de Células B/tratamiento farmacológico , Proteínas Proto-Oncogénicas c-bcl-2/genética , Sulfonamidas/metabolismo , Antineoplásicos/uso terapéutico , Compuestos Bicíclicos Heterocíclicos con Puentes/uso terapéutico , Cristalización , Cristalografía por Rayos X , Humanos , Mutación , Unión Proteica , Proteínas Proto-Oncogénicas c-bcl-2/antagonistas & inhibidores , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Sulfonamidas/uso terapéutico , Resonancia por Plasmón de SuperficieRESUMEN
Ibrutinib plus venetoclax is a highly effective combination in mantle cell lymphoma. However, strategies to enable the evaluation of therapeutic response are required. Our prospective analyses of patients within the AIM study revealed genomic profiles that clearly dichotomized responders and nonresponders. Mutations in ATM were present in most patients who achieved a complete response, while chromosome 9p21.1-p24.3 loss and/or mutations in components of the SWI-SNF chromatin-remodeling complex were present in all patients with primary resistance and two-thirds of patients with relapsed disease. Circulating tumor DNA analysis revealed that these alterations could be dynamically monitored, providing concurrent information on treatment response and tumor evolution. Functional modeling demonstrated that compromise of the SWI-SNF complex facilitated transcriptional upregulation of BCL2L1 (Bcl-xL) providing a selective advantage against ibrutinib plus venetoclax. Together these data highlight important insights into the molecular basis of therapeutic response and provide a model for real-time assessment of innovative targeted therapies.
Asunto(s)
Compuestos Bicíclicos Heterocíclicos con Puentes/uso terapéutico , Proteínas Cromosómicas no Histona/genética , Resistencia a Antineoplásicos/genética , Linfoma de Células del Manto/tratamiento farmacológico , Linfoma de Células del Manto/genética , Mutación/genética , Pirazoles/uso terapéutico , Pirimidinas/uso terapéutico , Sulfonamidas/uso terapéutico , Factores de Transcripción/genética , Factor de Transcripción Activador 3/metabolismo , Adenina/análogos & derivados , Línea Celular Tumoral , Cromatina/metabolismo , Proteínas Cromosómicas no Histona/metabolismo , ADN Tumoral Circulante/genética , Estudios de Cohortes , ADN Helicasas/metabolismo , Genoma Humano , Humanos , Modelos Biológicos , Proteínas Nucleares/metabolismo , Piperidinas , Pronóstico , Factores de Transcripción/metabolismo , Resultado del Tratamiento , Proteína bcl-X/metabolismoRESUMEN
The BCL2 inhibitor venetoclax induces high rates of durable remission in patients with previously treated chronic lymphocytic leukemia (CLL). However, despite continuous daily treatment, leukemia recurs in most patients. To investigate the mechanisms of secondary resistance, we analyzed paired pre-venetoclax and progression samples from 15 patients with CLL progression enrolled on venetoclax clinical trials. The novel Gly101Val mutation in BCL2 was identified at progression in 7 patients, but not at study entry. It was first detectable after 19 to 42 months of therapy, and its emergence anticipated clinical disease progression by many months. Gly101Val reduces the affinity of BCL2 for venetoclax by â¼180-fold in surface plasmon resonance assays, thereby preventing the drug from displacing proapoptotic mediators from BCL2 in cells and conferring acquired resistance in cell lines and primary patient cells. This mutation provides new insights into the pathobiology of venetoclax resistance and provides a potential biomarker of impending clinical relapse. SIGNIFICANCE: Why CLL recurs in patients who achieve remission with the BCL2 inhibitor venetoclax has been unknown. We provide the first description of an acquired point mutation in BCL2 arising recurrently and exclusively in venetoclax-treated patients. The mutation reduces venetoclax binding and is sufficient to confer resistance.See related commentary by Thangavadivel and Byrd, p. 320.This article is highlighted in the In This Issue feature, p. 305.
Asunto(s)
Compuestos Bicíclicos Heterocíclicos con Puentes/farmacología , Leucemia Linfocítica Crónica de Células B/tratamiento farmacológico , Leucemia Linfocítica Crónica de Células B/genética , Mutación , Proteínas Proto-Oncogénicas c-bcl-2/genética , Sulfonamidas/farmacología , Antineoplásicos/farmacología , Proliferación Celular/efectos de los fármacos , Resistencia a Antineoplásicos , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Leucemia Linfocítica Crónica de Células B/patología , Modelos Moleculares , Recurrencia Local de Neoplasia/tratamiento farmacológico , Recurrencia Local de Neoplasia/genética , Recurrencia Local de Neoplasia/patología , Conformación Proteica , Células Tumorales CultivadasRESUMEN
Inhibition of B-cell receptor (BCR) signaling pathways in chronic lymphocytic leukemia (CLL) provides significant clinical benefit to patients, mainly by blocking adhesion of CLL cells in the lymph node microenvironment. The currently applied inhibitors ibrutinib and idelalisib have limited capacity however to induce cell death as monotherapy and are unlikely to eradicate the disease. Acquired resistance to therapy in CLL is often caused by mutations in the response network being targeted, both for DNA damage or BCR signaling pathways. Thus, drugs with dual targeting capacity could offer improved therapeutic value. Here, the potency of CC-115, a novel inhibitor of mammalian target of rapamycin kinase (TORK) and DNA-dependent protein kinase (DNA-PK), was evaluated in primary CLL cells in vitro and in CLL patients. Combined TORK and DNA-PK inhibition in vitro resulted in caspase-dependent cell killing irrespective of p53, ATM, NOTCH1, or SF3B1 status. Proliferation induced by CD40(+) interleukin-21 stimulation was completely blocked by CC-115, and CD40-mediated resistance to fludarabine and venetoclax could be reverted by CC-115. BCR-mediated signaling was inhibited by CC-115 and also in CLL samples obtained from patients with acquired resistance to idelalisib treatment. Clinical efficacy of CC-115 was demonstrated in 8 patients with relapsed/refractory CLL/small lymphocytic lymphoma harboring ATM deletions/mutations; all but 1 patient had a decrease in lymphadenopathy, resulting in 1 IWCLL partial response (PR) and 3 PRs with lymphocytosis. In conclusion, these preclinical results, along with early promising clinical activity, suggest that CC-115 may be developed further for treatment of CLL. The trial was registered at www.clinicaltrials.gov as #NCT01353625.