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1.
Cell ; 184(25): 6119-6137.e26, 2021 12 09.
Artigo em Inglês | MEDLINE | ID: mdl-34890551

RESUMO

Prognostically relevant RNA expression states exist in pancreatic ductal adenocarcinoma (PDAC), but our understanding of their drivers, stability, and relationship to therapeutic response is limited. To examine these attributes systematically, we profiled metastatic biopsies and matched organoid models at single-cell resolution. In vivo, we identify a new intermediate PDAC transcriptional cell state and uncover distinct site- and state-specific tumor microenvironments (TMEs). Benchmarking models against this reference map, we reveal strong culture-specific biases in cancer cell transcriptional state representation driven by altered TME signals. We restore expression state heterogeneity by adding back in vivo-relevant factors and show plasticity in culture models. Further, we prove that non-genetic modulation of cell state can strongly influence drug responses, uncovering state-specific vulnerabilities. This work provides a broadly applicable framework for aligning cell states across in vivo and ex vivo settings, identifying drivers of transcriptional plasticity and manipulating cell state to target associated vulnerabilities.


Assuntos
Biomarcadores Tumorais/metabolismo , Carcinoma Ductal Pancreático/metabolismo , Neoplasias Pancreáticas/metabolismo , Microambiente Tumoral , Adulto , Idoso , Linhagem Celular Tumoral , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Pessoa de Meia-Idade , Análise de Célula Única
2.
Nat Chem Biol ; 16(6): 635-643, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32251410

RESUMO

Doublecortin like kinase 1 (DCLK1) is an understudied kinase that is upregulated in a wide range of cancers, including pancreatic ductal adenocarcinoma (PDAC). However, little is known about its potential as a therapeutic target. We used chemoproteomic profiling and structure-based design to develop a selective, in vivo-compatible chemical probe of the DCLK1 kinase domain, DCLK1-IN-1. We demonstrate activity of DCLK1-IN-1 against clinically relevant patient-derived PDAC organoid models and use a combination of RNA-sequencing, proteomics and phosphoproteomics analysis to reveal that DCLK1 inhibition modulates proteins and pathways associated with cell motility in this context. DCLK1-IN-1 will serve as a versatile tool to investigate DCLK1 biology and establish its role in cancer.


Assuntos
Carcinoma Ductal Pancreático/tratamento farmacológico , Peptídeos e Proteínas de Sinalização Intracelular/antagonistas & inibidores , Neoplasias Pancreáticas/tratamento farmacológico , Inibidores de Proteínas Quinases/química , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Animais , Linhagem Celular Tumoral , Movimento Celular , Proteína Duplacortina , Quinases Semelhantes a Duplacortina , Ensaios de Seleção de Medicamentos Antitumorais , Regulação da Expressão Gênica , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Masculino , Camundongos , Simulação de Acoplamento Molecular , Estrutura Molecular , Inibidores de Proteínas Quinases/farmacocinética , Proteômica , Ratos , Relação Estrutura-Atividade , Peixe-Zebra , Neoplasias Pancreáticas
3.
Cancer Res ; 83(3): 441-455, 2023 02 03.
Artigo em Inglês | MEDLINE | ID: mdl-36459568

RESUMO

Pancreatic ductal adenocarcinoma (PDAC) has been classified into classical and basal-like transcriptional subtypes by bulk RNA measurements. However, recent work has uncovered greater complexity to transcriptional subtypes than was initially appreciated using bulk RNA expression profiling. To provide a deeper understanding of PDAC subtypes, we developed a multiplex immunofluorescence (mIF) pipeline that quantifies protein expression of six PDAC subtype markers (CLDN18.2, TFF1, GATA6, KRT17, KRT5, and S100A2) and permits spatially resolved, single-cell interrogation of pancreatic tumors from resection specimens and core needle biopsies. Both primary and metastatic tumors displayed striking intratumoral subtype heterogeneity that was associated with patient outcomes, existed at the scale of individual glands, and was significantly reduced in patient-derived organoid cultures. Tumor cells co-expressing classical and basal markers were present in > 90% of tumors, existed on a basal-classical polarization continuum, and were enriched in tumors containing a greater admixture of basal and classical cell populations. Cell-cell neighbor analyses within tumor glands further suggested that co-expressor cells may represent an intermediate state between expression subtype poles. The extensive intratumoral heterogeneity identified through this clinically applicable mIF pipeline may inform prognosis and treatment selection for patients with PDAC. SIGNIFICANCE: A high-throughput pipeline using multiplex immunofluorescence in pancreatic cancer reveals striking expression subtype intratumoral heterogeneity with implications for therapy selection and identifies co-expressor cells that may serve as intermediates during subtype switching.


Assuntos
Carcinoma Ductal Pancreático , Neoplasias Pancreáticas , Humanos , Neoplasias Pancreáticas/patologia , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/patologia , Prognóstico , Fenótipo , RNA , Regulação Neoplásica da Expressão Gênica , Claudinas
4.
Cell Rep ; 33(11): 108493, 2020 12 15.
Artigo em Inglês | MEDLINE | ID: mdl-33326793

RESUMO

Few therapies target the loss of tumor suppressor genes in cancer. We examine CRISPR-SpCas9 and RNA-interference loss-of-function screens to identify new therapeutic targets associated with genomic loss of tumor suppressor genes. The endosomal sorting complexes required for transport (ESCRT) ATPases VPS4A and VPS4B score as strong synthetic lethal dependencies. VPS4A is essential in cancers harboring loss of VPS4B adjacent to SMAD4 on chromosome 18q and VPS4B is required in tumors with co-deletion of VPS4A and CDH1 (E-cadherin) on chromosome 16q. We demonstrate that more than 30% of cancers selectively require VPS4A or VPS4B. VPS4A suppression in VPS4B-deficient cells selectively leads to ESCRT-III filament accumulation, cytokinesis defects, nuclear deformation, G2/M arrest, apoptosis, and potent tumor regression. CRISPR-SpCas9 screening and integrative genomic analysis reveal other ESCRT members, regulators of abscission, and interferon signaling as modifiers of VPS4A dependency. We describe a compendium of synthetic lethal vulnerabilities and nominate VPS4A and VPS4B as high-priority therapeutic targets for cancers with 18q or 16q loss.


Assuntos
ATPases Associadas a Diversas Atividades Celulares/metabolismo , Complexos Endossomais de Distribuição Requeridos para Transporte/metabolismo , Neoplasias/genética , ATPases Vacuolares Próton-Translocadoras/metabolismo , Linhagem Celular Tumoral , Humanos
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