RESUMO
Pancreatic ductal adenocarcinoma (PDAC) is a highly lethal and treatment-refractory cancer. Molecular stratification in pancreatic cancer remains rudimentary and does not yet inform clinical management or therapeutic development. Here, we construct a high-resolution molecular landscape of the cellular subtypes and spatial communities that compose PDAC using single-nucleus RNA sequencing and whole-transcriptome digital spatial profiling (DSP) of 43 primary PDAC tumor specimens that either received neoadjuvant therapy or were treatment naive. We uncovered recurrent expression programs across malignant cells and fibroblasts, including a newly identified neural-like progenitor malignant cell program that was enriched after chemotherapy and radiotherapy and associated with poor prognosis in independent cohorts. Integrating spatial and cellular profiles revealed three multicellular communities with distinct contributions from malignant, fibroblast and immune subtypes: classical, squamoid-basaloid and treatment enriched. Our refined molecular and cellular taxonomy can provide a framework for stratification in clinical trials and serve as a roadmap for therapeutic targeting of specific cellular phenotypes and multicellular interactions.
Assuntos
Carcinoma Ductal Pancreático , Neoplasias Pancreáticas , Biomarcadores Tumorais/genética , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/patologia , Carcinoma Ductal Pancreático/terapia , Perfilação da Expressão Gênica , Humanos , Terapia Neoadjuvante , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/genética , Prognóstico , Transcriptoma/genética , Neoplasias PancreáticasRESUMO
Establishing causal relationships between genetic alterations of human cancers and specific phenotypes of malignancy remains a challenge. We sequentially introduced mutations into healthy human melanocytes in up to five genes spanning six commonly disrupted melanoma pathways, forming nine genetically distinct cellular models of melanoma. We connected mutant melanocyte genotypes to malignant cell expression programs in vitro and in vivo, replicative immortality, malignancy, rapid tumor growth, pigmentation, metastasis, and histopathology. Mutations in malignant cells also affected tumor microenvironment composition and cell states. Our melanoma models shared genotype-associated expression programs with patient melanomas, and a deep learning model showed that these models partially recapitulated genotype-associated histopathological features as well. Thus, a progressive series of genome-edited human cancer models can causally connect genotypes carrying multiple mutations to phenotype.
Assuntos
Melanoma , Neoplasias Cutâneas , Humanos , Melanócitos/metabolismo , Melanoma/patologia , Mutação , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/patologia , Microambiente Tumoral/genéticaRESUMO
Identifying gene-regulatory targets of nuclear proteins in tissues is a challenge. Here we describe intranuclear cellular indexing of transcriptomes and epitopes (inCITE-seq), a scalable method that measures multiplexed intranuclear protein levels and the transcriptome in parallel across thousands of nuclei, enabling joint analysis of transcription factor (TF) levels and gene expression in vivo. We apply inCITE-seq to characterize cell state-related changes upon pharmacological induction of neuronal activity in the mouse brain. Modeling gene expression as a linear combination of quantitative protein levels revealed genome-wide associations of each TF and recovered known gene targets. TF-associated genes were coexpressed as distinct modules that each reflected positive or negative TF levels, showing that our approach can disentangle relative putative contributions of TFs to gene expression and add interpretability to inferred gene networks. inCITE-seq can illuminate how combinations of nuclear proteins shape gene expression in native tissue contexts, with direct applications to solid or frozen tissues and clinical specimens.
Assuntos
Biologia Computacional/métodos , Proteínas Nucleares/metabolismo , Análise de Célula Única/métodos , Animais , Anticorpos , Encéfalo/metabolismo , Regulação da Expressão Gênica/fisiologia , Estudo de Associação Genômica Ampla , Ácido Caínico/toxicidade , Camundongos , Proteínas Nucleares/genética , RNA , Reprodutibilidade dos Testes , Convulsões/induzido quimicamente , Fator de Transcrição RelA/genética , Fator de Transcrição RelA/metabolismoRESUMO
The CD155/TIGIT axis can be co-opted during immune evasion in chronic viral infections and cancer. Pancreatic adenocarcinoma (PDAC) is a highly lethal malignancy, and immune-based strategies to combat this disease have been largely unsuccessful to date. We corroborate prior reports that a substantial portion of PDAC harbors predicted high-affinity MHC class I-restricted neoepitopes and extend these findings to advanced/metastatic disease. Using multiple preclinical models of neoantigen-expressing PDAC, we demonstrate that intratumoral neoantigen-specific CD8+ T cells adopt multiple states of dysfunction, resembling those in tumor-infiltrating lymphocytes of PDAC patients. Mechanistically, genetic and/or pharmacologic modulation of the CD155/TIGIT axis was sufficient to promote immune evasion in autochthonous neoantigen-expressing PDAC. Finally, we demonstrate that the CD155/TIGIT axis is critical in maintaining immune evasion in PDAC and uncover a combination immunotherapy (TIGIT/PD-1 co-blockade plus CD40 agonism) that elicits profound anti-tumor responses in preclinical models, now poised for clinical evaluation.
Assuntos
Evasão da Resposta Imune/imunologia , Imunoterapia/métodos , Linfócitos do Interstício Tumoral/metabolismo , Neoplasias Pancreáticas/imunologia , Receptores Virais/imunologia , Animais , Humanos , Camundongos , Neoplasias PancreáticasRESUMO
COVID-19, which is caused by SARS-CoV-2, can result in acute respiratory distress syndrome and multiple organ failure1-4, but little is known about its pathophysiology. Here we generated single-cell atlases of 24 lung, 16 kidney, 16 liver and 19 heart autopsy tissue samples and spatial atlases of 14 lung samples from donors who died of COVID-19. Integrated computational analysis uncovered substantial remodelling in the lung epithelial, immune and stromal compartments, with evidence of multiple paths of failed tissue regeneration, including defective alveolar type 2 differentiation and expansion of fibroblasts and putative TP63+ intrapulmonary basal-like progenitor cells. Viral RNAs were enriched in mononuclear phagocytic and endothelial lung cells, which induced specific host programs. Spatial analysis in lung distinguished inflammatory host responses in lung regions with and without viral RNA. Analysis of the other tissue atlases showed transcriptional alterations in multiple cell types in heart tissue from donors with COVID-19, and mapped cell types and genes implicated with disease severity based on COVID-19 genome-wide association studies. Our foundational dataset elucidates the biological effect of severe SARS-CoV-2 infection across the body, a key step towards new treatments.
Assuntos
COVID-19/patologia , COVID-19/virologia , Rim/patologia , Fígado/patologia , Pulmão/patologia , Miocárdio/patologia , SARS-CoV-2/patogenicidade , Adulto , Idoso , Idoso de 80 Anos ou mais , Atlas como Assunto , Autopsia , Bancos de Espécimes Biológicos , COVID-19/genética , COVID-19/imunologia , Células Endoteliais , Células Epiteliais/patologia , Células Epiteliais/virologia , Feminino , Fibroblastos , Estudo de Associação Genômica Ampla , Coração/virologia , Humanos , Inflamação/patologia , Inflamação/virologia , Rim/virologia , Fígado/virologia , Pulmão/virologia , Masculino , Pessoa de Meia-Idade , Especificidade de Órgãos , Fagócitos , Alvéolos Pulmonares/patologia , Alvéolos Pulmonares/virologia , RNA Viral/análise , Regeneração , SARS-CoV-2/imunologia , Análise de Célula Única , Carga ViralRESUMO
Tumor evolution from a single cell into a malignant, heterogeneous tissue remains poorly understood. Here, we profile single-cell transcriptomes of genetically engineered mouse lung tumors at seven stages, from pre-neoplastic hyperplasia to adenocarcinoma. The diversity of transcriptional states increases over time and is reproducible across tumors and mice. Cancer cells progressively adopt alternate lineage identities, computationally predicted to be mediated through a common transitional, high-plasticity cell state (HPCS). Accordingly, HPCS cells prospectively isolated from mouse tumors and human patient-derived xenografts display high capacity for differentiation and proliferation. The HPCS program is associated with poor survival across human cancers and demonstrates chemoresistance in mice. Our study reveals a central principle underpinning intra-tumoral heterogeneity and motivates therapeutic targeting of the HPCS.