RESUMO
Despite the success of PD-1 blockade in melanoma and other cancers, effective treatment strategies to overcome resistance to cancer immunotherapy are lacking1,2. Here we identify the innate immune kinase TANK-binding kinase 1 (TBK1)3 as a candidate immune-evasion gene in a pooled genetic screen4. Using a suite of genetic and pharmacological tools across multiple experimental model systems, we confirm a role for TBK1 as an immune-evasion gene. Targeting TBK1 enhances responses to PD-1 blockade by decreasing the cytotoxicity threshold to effector cytokines (TNF and IFNγ). TBK1 inhibition in combination with PD-1 blockade also demonstrated efficacy using patient-derived tumour models, with concordant findings in matched patient-derived organotypic tumour spheroids and matched patient-derived organoids. Tumour cells lacking TBK1 are primed to undergo RIPK- and caspase-dependent cell death in response to TNF and IFNγ in a JAK-STAT-dependent manner. Taken together, our results demonstrate that targeting TBK1 is an effective strategy to overcome resistance to cancer immunotherapy.
Assuntos
Resistencia a Medicamentos Antineoplásicos , Evasão da Resposta Imune , Imunoterapia , Proteínas Serina-Treonina Quinases , Humanos , Evasão da Resposta Imune/genética , Evasão da Resposta Imune/imunologia , Imunoterapia/métodos , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Serina-Treonina Quinases/genética , Organoides , Fatores de Necrose Tumoral/imunologia , Interferon gama/imunologia , Esferoides Celulares , Caspases , Janus Quinases , Fatores de Transcrição STATAssuntos
Líquen Plano , Alopecia , Humanos , Líquen Plano/radioterapia , Projetos Piloto , Estudos Prospectivos , Couro CabeludoRESUMO
Giant congenital melanocytic nevi are NRAS-driven proliferations that may cover up to 80% of the body surface. Their most dangerous consequence is progression to melanoma. This risk often triggers preemptive extensive surgical excisions in childhood, producing severe lifelong challenges. We have presented preclinical models, including multiple genetically engineered mice and xenografted human lesions, which enabled testing locally applied pharmacologic agents to avoid surgery. The murine models permitted the identification of proliferative versus senescent nevus phases and treatments targeting both. These nevi recapitulated the histologic and molecular features of human giant congenital nevi, including the risk of melanoma transformation. Cutaneously delivered MEK, PI3K, and c-KIT inhibitors or proinflammatory squaric acid dibutylester (SADBE) achieved major regressions. SADBE triggered innate immunity that ablated detectable nevocytes, fully prevented melanoma, and regressed human giant nevus xenografts. These findings reveal nevus mechanistic vulnerabilities and suggest opportunities for topical interventions that may alter the therapeutic options for children with congenital giant nevi.
Assuntos
Melanoma , Nevo Pigmentado , Neoplasias Cutâneas , Animais , Xenoenxertos , Humanos , Melanoma/tratamento farmacológico , Melanoma/patologia , Camundongos , Transplante de Neoplasias , Nevo Pigmentado/congênito , Nevo Pigmentado/tratamento farmacológico , Nevo Pigmentado/patologia , Neoplasias Cutâneas/tratamento farmacológico , Neoplasias Cutâneas/patologia , Neoplasias Cutâneas/prevenção & controleRESUMO
Although immune checkpoint inhibitors (ICIs), such as anti-programmed cell death protein-1 (PD-1), can deliver durable antitumor effects, most patients with cancer fail to respond. Recent studies suggest that ICI efficacy correlates with a higher load of tumor-specific neoantigens and development of vitiligo in patients with melanoma. Here, we report that patients with low melanoma neoantigen burdens who responded to ICI had tumors with higher expression of pigmentation-related genes. Moreover, expansion of peripheral blood CD8+ T cell populations specific for melanocyte antigens was observed only in patients who responded to anti-PD-1 therapy, suggesting that ICI can promote breakdown of tolerance toward tumor-lineage self-antigens. In a mouse model of poorly immunogenic melanomas, spreading of epitope recognition toward wild-type melanocyte antigens was associated with markedly improved anti-PD-1 efficacy in two independent approaches: introduction of neoantigens by ultraviolet (UV) B radiation mutagenesis or the therapeutic combination of ablative fractional photothermolysis plus imiquimod. Complete responses against UV mutation-bearing tumors after anti-PD-1 resulted in protection from subsequent engraftment of melanomas lacking any shared neoantigens, as well as pancreatic adenocarcinomas forcibly overexpressing melanocyte-lineage antigens. Our data demonstrate that somatic mutations are sufficient to provoke strong antitumor responses after checkpoint blockade, but long-term responses are not restricted to these putative neoantigens. Epitope spreading toward T cell recognition of wild-type tumor-lineage self-antigens represents a common pathway for successful response to ICI, which can be evoked in neoantigen-deficient tumors by combination therapy with ablative fractional photothermolysis and imiquimod.
Assuntos
Inibidores de Checkpoint Imunológico , Melanoma , Animais , Antígenos de Neoplasias , Epitopos , Humanos , Melanócitos , Melanoma/terapia , CamundongosRESUMO
In a Perspective, Mack Su and David Fisher discuss the development of immunotherapies for treatment of melanoma and other cancer types.
Assuntos
Imunoterapia/tendências , Melanoma/terapia , Medicina de Precisão/métodos , Humanos , Medicina de Precisão/tendênciasRESUMO
The retinoblastoma gene (RB1) has been implicated as a tumor suppressor in multiple myeloma (MM), yet its role remains unclear because in the majority of cases with 13q14 deletions, un-mutated RB1 remains expressed from the retained allele. To explore the role of Rb1 in MM, we examined the functional consequences of single- and double-copy Rb1 loss in germinal center B cells, the cells of origin of MM. We generated mice without Rb1 function in germinal center B cells by crossing Rb1(Flox/Flox) with C-γ-1-Cre (Cγ1) mice expressing the Cre recombinase in class-switched B cells in a p107(-/-) background to prevent p107 from compensating for Rb1 loss (Cγ1-Rb1(F/F)-p107(-/-)). All mice developed normally, but B cells with two copies of Rb1 deleted (Cγ1-Rb1(F/F)-p107(-/-)) exhibited increased proliferation and cell death compared with Cγ1-Rb1(+/+)-p107(-/-) controls ex vivo. In vivo, Cγ1-Rb1(F/F)-p107(-/-) mice had a lower percentage of splenic B220+ cells and reduced numbers of bone marrow antigen-specific secreting cells compared with control mice. Our data indicate that Rb1 loss induces both cell proliferation and death in germinal center B cells. Because no B-cell malignancies developed after 1 year of observation, our data also suggest that Rb1 loss is not sufficient to transform post-germinal center B cells and that additional, specific mutations are likely required to cooperate with Rb1 loss to induce malignant transformation.
Assuntos
Linfócitos B/metabolismo , Proliferação de Células , Transformação Celular Neoplásica/metabolismo , Centro Germinativo/metabolismo , Mutação , Proteína do Retinoblastoma/deficiência , Animais , Linfócitos B/patologia , Morte Celular , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/patologia , Centro Germinativo/patologia , Camundongos , Camundongos KnockoutRESUMO
T follicular helper (Tfh) cells are a subset of CD4(+) T helper cells that migrate into germinal centers and promote B-cell maturation into memory B and plasma cells. Tfh cells are necessary for promotion of protective humoral immunity following pathogen challenge, but when aberrantly regulated, drive pathogenic antibody formation in autoimmunity and undergo neoplastic transformation in angioimmunoblastic T-cell lymphoma and other primary cutaneous T-cell lymphomas. Limited information is available on the expression and regulation of genes in human Tfh cells. Using a fluorescence-activated cell sorting-based strategy, we obtained primary Tfh and non-Tfh T effector cells from tonsils and prepared genome-wide maps of active, intermediate, and poised enhancers determined by chromatin immunoprecipitation-sequencing, with parallel transcriptome analyses determined by RNA sequencing. Tfh cell enhancers were enriched near genes highly expressed in lymphoid cells or involved in lymphoid cell function, with many mapping to sites previously associated with autoimmune disease in genome-wide association studies. A group of active enhancers unique to Tfh cells associated with differentially expressed genes was identified. Fragments from these regions directed expression in reporter gene assays. These data provide a significant resource for studies of T lymphocyte development and differentiation and normal and perturbed Tfh cell function.
Assuntos
Elementos Facilitadores Genéticos/genética , Subpopulações de Linfócitos T/metabolismo , Linfócitos T Auxiliares-Indutores/metabolismo , Transcriptoma/genética , Células Cultivadas , Citometria de Fluxo , Genoma Humano/genética , Humanos , Análise de Sequência com Séries de Oligonucleotídeos , Tonsila Palatina/citologia , Cultura Primária de Células , Análise de Sequência de RNA , Subpopulações de Linfócitos T/citologia , Linfócitos T Auxiliares-Indutores/citologiaRESUMO
Activating mutations in Ras (N- and K-) are the most common point mutations found in patients with multiple myeloma (MM) and are associated with poor clinical outcome. We sought to directly examine the role of Ras activation in MM pathogenesis and used two different tissue-specific Cre recombinase mouse lines (Cγ1-Cre and AID-Cre), to generate mice with mutant Kras (Kras(G12D) ) activated specifically in germinal center B-cells. We also generated mice with activation of the Kras(G12D) allele in a tumor-prone Arf-null genetic background. Surprisingly, we observed no significant disruption in B-cell homeostasis in any of these models by serum immunoglobulin ELISA, SPEP, flow cytometry and histological examination. We observed development of non-overlapping tumor types due to off-target Cre expression, but despite successful recombination in germinal center and later B-cell populations, we observed no B-cell phenotype. Together, these data demonstrate that Ras activation is not sufficient to transform primary germinal center B-cells, even in an Arf-null context, and that the temporal order of mutation acquisition may be critical for myeloma development. Specific pathways, yet to be identified, are required before Kras can contribute to the development of MM.
Assuntos
Linfócitos B/patologia , Transformação Celular Neoplásica , Inibidor p16 de Quinase Dependente de Ciclina/deficiência , Deleção de Genes , Centro Germinativo/imunologia , Proteínas Proto-Oncogênicas/genética , Proteínas Supressoras de Tumor/deficiência , Proteínas ras/genética , Animais , Inibidor p16 de Quinase Dependente de Ciclina/genética , Linfoma/patologia , Camundongos , Camundongos Transgênicos , Papiloma/patologia , Proteínas Proto-Oncogênicas p21(ras) , Neoplasias Cutâneas/patologia , Timoma/patologia , Proteínas Supressoras de Tumor/genéticaRESUMO
Identification of cell type-specific enhancers is important for understanding the regulation of programs controlling cellular development and differentiation. Enhancers are typically marked by the co-transcriptional activator protein p300 or by groups of cell-expressed transcription factors. We hypothesized that a unique set of enhancers regulates gene expression in human erythroid cells, a highly specialized cell type evolved to provide adequate amounts of oxygen throughout the body. Using chromatin immunoprecipitation followed by massively parallel sequencing, genome-wide maps of candidate enhancers were constructed for p300 and four transcription factors, GATA1, NF-E2, KLF1, and SCL, using primary human erythroid cells. These data were combined with gene expression analyses, and candidate enhancers were identified. Consistent with their predicted function as candidate enhancers, there was statistically significant enrichment of p300 and combinations of co-localizing erythroid transcription factors within 1-50 kb of the transcriptional start site (TSS) of genes highly expressed in erythroid cells. Candidate enhancers were also enriched near genes with known erythroid cell function or phenotype. Candidate enhancers exhibited moderate conservation with mouse and minimal conservation with nonplacental vertebrates. Candidate enhancers were mapped to a set of erythroid-associated, biologically relevant, SNPs from the genome-wide association studies (GWAS) catalogue of NHGRI, National Institutes of Health. Fourteen candidate enhancers, representing 10 genetic loci, mapped to sites associated with biologically relevant erythroid traits. Fragments from these loci directed statistically significant expression in reporter gene assays. Identification of enhancers in human erythroid cells will allow a better understanding of erythroid cell development, differentiation, structure, and function and provide insights into inherited and acquired hematologic disease.
Assuntos
Elementos Facilitadores Genéticos , Células Eritroides/metabolismo , Regulação da Expressão Gênica , Sequência de Bases , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Fatores de Transcrição Hélice-Alça-Hélice Básicos/fisiologia , Células Cultivadas , Cromatina/genética , Cromatina/metabolismo , Imunoprecipitação da Cromatina , Sequência Conservada , Proteína p300 Associada a E1A/metabolismo , Fator de Transcrição GATA1/metabolismo , Fator de Transcrição GATA1/fisiologia , Genes Reporter , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Fatores de Transcrição Kruppel-Like/metabolismo , Fatores de Transcrição Kruppel-Like/fisiologia , Luciferases de Vaga-Lume/biossíntese , Luciferases de Vaga-Lume/genética , Anotação de Sequência Molecular , Subunidade p45 do Fator de Transcrição NF-E2/metabolismo , Subunidade p45 do Fator de Transcrição NF-E2/fisiologia , Análise de Sequência com Séries de Oligonucleotídeos , Polimorfismo de Nucleotídeo Único , Regiões Promotoras Genéticas , Ligação Proteica , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas/fisiologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Análise de Sequência de DNA , Proteína 1 de Leucemia Linfocítica Aguda de Células T , TranscriptomaRESUMO
The histone methyltransferase WHSC1 (also known as MMSET) is overexpressed in multiple myeloma (MM) as a result of the t(4;14) chromosomal translocation and in a broad variety of other cancers by unclear mechanisms. Overexpression of WHSC1 did not transform wild-type or tumor-prone primary hematopoietic cells. We found that ACA11, an orphan box H/ACA class small nucleolar RNA (snoRNA) encoded within an intron of WHSC1, was highly expressed in t(4;14)-positive MM and other cancers. ACA11 localized to nucleoli and bound what we believe to be a novel small nuclear ribonucleoprotein (snRNP) complex composed of several proteins involved in postsplicing intron complexes. RNA targets of this uncharacterized snRNP included snoRNA intermediates hosted within ribosomal protein (RP) genes, and an RP gene signature was strongly associated with t(4;14) in patients with MM. Expression of ACA11 was sufficient to downregulate RP genes and other snoRNAs implicated in the control of oxidative stress. ACA11 suppressed oxidative stress, afforded resistance to chemotherapy, and increased the proliferation of MM cells, demonstrating that ACA11 is a critical target of the t(4;14) translocation in MM and suggesting an oncogenic role in other cancers as well.