RESUMO
Proteasome inhibitors (PI) are extensively used for the therapy of multiple myeloma (MM) and mantle cell lymphoma. However, patients continuously relapse or are intrinsically resistant to this class of drugs. Here, to identify targets that synergize with PI, we carried out a functional screening in MM cell lines using a short hairpin RNA library against cancer driver genes. Isocitrate dehydrogenase 2 (IDH2) was identified as a top candidate, showing a synthetic lethal activity with the PI carfilzomib (CFZ). Combinations of US Food and Drug Administration-approved PI with a pharmacological IDH2 inhibitor (AGI-6780) triggered synergistic cytotoxicity in MM, mantle cell lymphoma, and Burkitt lymphoma cell lines. CFZ/AGI-6780 treatment increased death of primary CD138+ cells from MM patients and exhibited a favorable cytotoxicity profile toward peripheral blood mononuclear cells and bone marrow-derived stromal cells. Mechanistically, the CFZ/AGI-6780 combination significantly decreased tricarboxylic acid cycle activity and adenosine triphosphate levels as a consequence of enhanced IDH2 enzymatic inhibition. Specifically, CFZ treatment reduced the expression of nicotinamide phosphoribosyltransferase (NAMPT), thus limiting IDH2 activation through the NAD+-dependent deacetylase SIRT3. Consistently, combination of CFZ with either NAMPT or SIRT3 inhibitors impaired IDH2 activity and increased MM cell death. Finally, inducible IDH2 knockdown enhanced the therapeutic efficacy of CFZ in a subcutaneous xenograft model of MM, resulting in inhibition of tumor progression and extended survival. Taken together, these findings indicate that NAMPT/SIRT3/IDH2 pathway inhibition enhances the therapeutic efficacy of PI, thus providing compelling evidence for treatments with lower and less toxic doses and broadening the application of PI to other malignancies.
Assuntos
Resistencia a Medicamentos Antineoplásicos , Neoplasias Hematológicas/tratamento farmacológico , Isocitrato Desidrogenase/antagonistas & inibidores , Oligopeptídeos/farmacologia , Inibidores de Proteassoma/farmacologia , Animais , Apoptose , Proliferação de Células , Citocinas/antagonistas & inibidores , Citocinas/genética , Neoplasias Hematológicas/genética , Neoplasias Hematológicas/patologia , Humanos , Isocitrato Desidrogenase/genética , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Nicotinamida Fosforribosiltransferase/antagonistas & inibidores , Nicotinamida Fosforribosiltransferase/genética , RNA Interferente Pequeno/genética , Sirtuína 3/antagonistas & inibidores , Sirtuína 3/genética , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Anaplastic large-cell lymphoma (ALCL) is a clinical and biological heterogeneous disease that includes systemic anaplastic lymphoma kinase (ALK)-positive and ALK-negative entities. To discover biomarkers and/or genes involved in ALK-negative ALCL pathogenesis, we applied the cancer outlier profile analysis algorithm to a gene expression profiling data set including 249 cases of T-cell non-Hodgkin lymphoma and normal T cells. Ectopic coexpression of ERBB4 and COL29A1 genes was detected in 24% of ALK-negative ALCL patients. RNA sequencing and 5' RNA ligase-mediated rapid amplification of complementary DNA ends identified 2 novel ERBB4-truncated transcripts displaying intronic transcription start sites. By luciferase assays, we defined that the expression of ERBB4-aberrant transcripts is promoted by endogenous intronic long terminal repeats. ERBB4 expression was confirmed at the protein level by western blot analysis and immunohistochemistry. Lastly, we demonstrated that ERBB4-truncated forms show oncogenic potentials and that ERBB4 pharmacologic inhibition partially controls ALCL cell growth and disease progression in an ERBB4-positive patient-derived tumorgraft model. In conclusion, we identified a new subclass of ALK-negative ALCL characterized by aberrant expression of ERBB4-truncated transcripts carrying intronic 5' untranslated regions.
Assuntos
Linfoma Anaplásico de Células Grandes/genética , Receptores Proteína Tirosina Quinases/genética , Receptor ErbB-4/genética , Regiões 5' não Traduzidas , Quinase do Linfoma Anaplásico , Animais , Códon sem Sentido , Regulação Neoplásica da Expressão Gênica , Células HEK293 , Humanos , Linfoma Anaplásico de Células Grandes/classificação , Linfoma Anaplásico de Células Grandes/patologia , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Camundongos Transgênicos , Dados de Sequência Molecular , Proteínas Mutantes/genética , Proteínas Mutantes/metabolismo , Células NIH 3T3 , RNA Mensageiro/metabolismo , Receptores Proteína Tirosina Quinases/metabolismo , Receptor ErbB-4/metabolismoRESUMO
Anaplastic large-cell lymphomas (ALCLs) are a group of clinically and biologically heterogeneous diseases including the ALK(+) and ALK(-) systemic forms. Whereas ALK(+) ALCLs are molecularly characterized and can be readily diagnosed, specific immunophenotypic or genetic features to define ALK(-) ALCL are missing, and their distinction from other T-cell non-Hodgkin lymphomas (T-NHLs) remains controversial. In the present study, we undertook a transcriptional profiling meta-analysis of 309 cases, including ALCL and other primary T-NHL samples. Pathway discovery and prediction analyses defined a minimum set of genes capable of recognizing ALK(-) ALCL. Application of quantitative RT-PCR in independent datasets from cryopreserved and formalin-fixed paraffin-embedded samples validated a 3-gene model (TNFRSF8, BATF3, and TMOD1) able to successfully separate ALK(-) ALCL from peripheral T-cell lymphoma not otherwise specified, with overall accuracy near 97%. In conclusion, our data justify the possibility of translating quantitative RT-PCR protocols to routine clinical settings as a new approach to objectively dissect T-NHL and to select more appropriate therapeutic protocols.
Assuntos
Biomarcadores Tumorais/genética , Genes Neoplásicos , Linfoma Anaplásico de Células Grandes/diagnóstico , Linfoma Anaplásico de Células Grandes/genética , Técnicas de Diagnóstico Molecular/métodos , Receptores Proteína Tirosina Quinases/genética , Adulto , Quinase do Linfoma Anaplásico , Biomarcadores Tumorais/isolamento & purificação , Biomarcadores Tumorais/fisiologia , Estudos de Casos e Controles , Diagnóstico Diferencial , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Genes Neoplásicos/fisiologia , Humanos , Análise em Microsséries , Modelos Estatísticos , Valor Preditivo dos Testes , Prognóstico , Receptores Proteína Tirosina Quinases/metabolismoRESUMO
Chimeric antigen receptor (CAR) T cell therapy targeting CD19 elicits remarkable clinical efficacy in B-cell malignancies, but many patients relapse due to failed expansion and/or progressive loss of CAR-T cells. We recently reported a strategy to potently restimulate CAR-T cells in vivo, enhancing their functionality by administration of a vaccine-like stimulus comprised of surrogate peptide ligands for a CAR linked to a lymph node-targeting amphiphilic PEG-lipid (termed CAR-T-vax). Here, we demonstrate a general strategy to generate and optimize peptide mimotopes enabling CAR-T-vax generation for any CAR. Using the clinical CD19 CAR FMC63 as a test case, we employed yeast surface display to identify peptide binders to soluble IgG versions of FMC63, which were subsequently affinity matured by directed evolution. CAR-T vaccines using these optimized mimotopes triggered marked expansion of both murine CD19 CAR-T cells in a syngeneic model and human CAR-T cells in a humanized mouse model of B cell acute lymphoblastic leukemia (B-ALL), and enhanced control of leukemia progression. This approach thus enables vaccine boosting to be applied to any clinically-relevant CAR-T cell product.
RESUMO
Selection of the best tumor antigen is critical for the therapeutic success of chimeric antigen receptor (CAR) T cells in hematologic malignancies and solid tumors. The anaplastic lymphoma kinase (ALK) receptor is expressed by most neuroblastomas while virtually absent in most normal tissues. ALK is an oncogenic driver in neuroblastoma and ALK inhibitors show promising clinical activity. Here, we describe the development of ALK.CAR-T cells that show potent efficacy in monotherapy against neuroblastoma with high ALK expression without toxicity. For neuroblastoma with low ALK expression, combination with ALK inhibitors specifically potentiates ALK.CAR-T cells but not GD2.CAR-T cells. Mechanistically, ALK inhibitors impair tumor growth and upregulate the expression of ALK, thereby facilitating the activity of ALK.CAR-T cells against neuroblastoma. Thus, while neither ALK inhibitors nor ALK.CAR-T cells will likely be sufficient as monotherapy in neuroblastoma with low ALK density, their combination specifically enhances therapeutic efficacy.
Assuntos
Neuroblastoma , Humanos , Quinase do Linfoma Anaplásico/genética , Quinase do Linfoma Anaplásico/metabolismo , Neuroblastoma/tratamento farmacológico , Neuroblastoma/genética , Neuroblastoma/metabolismo , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/uso terapêutico , Antígenos de Neoplasias , Linfócitos T , Linhagem Celular TumoralRESUMO
Anaplastic lymphoma kinase (ALK)-rearranged non-small cell lung cancer (NSCLC) is treated with ALK tyrosine kinase inhibitors (TKIs), but the lack of activity of immune checkpoint inhibitors (ICIs) is poorly understood. Here, we identified immunogenic ALK peptides to show that ICIs induced rejection of ALK+ tumors in the flank but not in the lung. A single-peptide vaccination restored priming of ALK-specific CD8+ T cells, eradicated lung tumors in combination with ALK TKIs and prevented metastatic dissemination of tumors to the brain. The poor response of ALK+ NSCLC to ICIs was due to ineffective CD8+ T cell priming against ALK antigens and is circumvented through specific vaccination. Finally, we identified human ALK peptides displayed by HLA-A*02:01 and HLA-B*07:02 molecules. These peptides were immunogenic in HLA-transgenic mice and were recognized by CD8+ T cells from individuals with NSCLC, paving the way for the development of a clinical vaccine to treat ALK+ NSCLC.
Assuntos
Vacinas Anticâncer , Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , Camundongos , Animais , Humanos , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/genética , Quinase do Linfoma Anaplásico/genética , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/patologia , Vacinas Anticâncer/uso terapêutico , Receptores Proteína Tirosina Quinases/uso terapêutico , Linfócitos T CD8-Positivos/patologia , Vacinas de Subunidades Antigênicas/uso terapêutico , Inibidores de Proteínas Quinases/farmacologia , Proteínas Tirosina Quinases/uso terapêutico , Camundongos Transgênicos , VacinaçãoRESUMO
Isocitrate dehydrogenases (IDHs) are enzymes that catalyze the oxidative decarboxylation of isocitrate, producing α-ketoglutarate (αKG) and CO2. The discovery of IDH1 and IDH2 mutations in several malignancies has brought to the approval of drugs targeting IDH1/2 mutants in cancers. Here, we summarized findings addressing the impact of IDH mutants in rare pathologies and focused on the relevance of non-mutated IDH enzymes in tumors. Several pieces of evidence suggest that the enzymatic inhibition of IDHs may have therapeutic potentials also in wild-type IDH cancers. Moreover, IDHs inhibition could enhance the efficacy of canonical cancer therapies, such as chemotherapy, target therapy, and radiotherapy. However, further studies are required to elucidate whether IDH proteins are diagnostic/prognostic markers, instrumental for tumor initiation and maintenance, and could be exploited as targets for anticancer therapy. The development of wild-type IDH inhibitors is expected to improve our understanding of a potential non-oncogenic addition to IDH1/2 activities and to fully address their applicability in combination with other therapies.
RESUMO
Recent studies reported the expression of anaplastic lymphoma kinase (ALK) in malignant melanomas. The aim of this study was to investigate whether ALK expression is associated with specific clinical and molecular characteristics of melanoma metastases, and to evaluate its correlation with survival outcomes. Seventy-one patients with metastatic melanoma were investigated. Clinical features and survival outcomes were analyzed and correlated to ALK expression, as detected by immunohistochemistry and reverse transcription-quantitative polymerase chain reaction, and to the mutational status of BRAF, KRAS, NRAS, and PIK3CA. No translocations or ALK alternative isoforms were identified. ALK expression was mainly detected in NRAS mutated metastatic lesions. Interestingly, among NRAS-mutated patients, ALK positive samples displayed a significantly more favorable outcome in terms of disease specific survival, as compared to ALK negative ones. In conclusion, we suggest that ALK positive/NRAS mutated metastases represent a specific subset of metastatic melanomas, associated with a better prognosis. Validation of these observations in larger cohorts could contribute to understand the molecular events cooperating to melanoma progression, in addition to open new perspectives in the clinical and therapeutic management of this subgroup of patients.