RESUMO
Analysis across a growing number of single-cell perturbation datasets is hampered by poor data interoperability. To facilitate development and benchmarking of computational methods, we collect a set of 44 publicly available single-cell perturbation-response datasets with molecular readouts, including transcriptomics, proteomics and epigenomics. We apply uniform quality control pipelines and harmonize feature annotations. The resulting information resource, scPerturb, enables development and testing of computational methods, and facilitates comparison and integration across datasets. We describe energy statistics (E-statistics) for quantification of perturbation effects and significance testing, and demonstrate E-distance as a general distance measure between sets of single-cell expression profiles. We illustrate the application of E-statistics for quantifying similarity and efficacy of perturbations. The perturbation-response datasets and E-statistics computation software are publicly available at scperturb.org. This work provides an information resource for researchers working with single-cell perturbation data and recommendations for experimental design, including optimal cell counts and read depth.
Assuntos
Proteômica , Software , Perfilação da Expressão Gênica/métodos , Epigenômica , Análise de Célula ÚnicaRESUMO
Pancreatic ductal adenocarcinoma (PDAC) is one of the deadliest cancers in western countries, with a median survival of 6 months and an extremely low percentage of long-term surviving patients. KRAS mutations are known to be a driver event of PDAC, but targeting mutant KRAS has proved challenging. Targeting oncogene-driven signalling pathways is a clinically validated approach for several devastating diseases. Still, despite marked tumour shrinkage, the frequency of relapse indicates that a fraction of tumour cells survives shut down of oncogenic signalling. Here we explore the role of mutant KRAS in PDAC maintenance using a recently developed inducible mouse model of mutated Kras (Kras(G12D), herein KRas) in a p53(LoxP/WT) background. We demonstrate that a subpopulation of dormant tumour cells surviving oncogene ablation (surviving cells) and responsible for tumour relapse has features of cancer stem cells and relies on oxidative phosphorylation for survival. Transcriptomic and metabolic analyses of surviving cells reveal prominent expression of genes governing mitochondrial function, autophagy and lysosome activity, as well as a strong reliance on mitochondrial respiration and a decreased dependence on glycolysis for cellular energetics. Accordingly, surviving cells show high sensitivity to oxidative phosphorylation inhibitors, which can inhibit tumour recurrence. Our integrated analyses illuminate a therapeutic strategy of combined targeting of the KRAS pathway and mitochondrial respiration to manage pancreatic cancer.
Assuntos
Carcinoma Ductal Pancreático/metabolismo , Carcinoma Ductal Pancreático/patologia , Mitocôndrias/metabolismo , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patologia , Proteínas Proto-Oncogênicas p21(ras)/genética , Animais , Autofagia , Carcinoma Ductal Pancreático/tratamento farmacológico , Carcinoma Ductal Pancreático/genética , Respiração Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Modelos Animais de Doenças , Feminino , Regulação Neoplásica da Expressão Gênica , Genes p53/genética , Glicólise , Lisossomos/metabolismo , Camundongos , Mitocôndrias/efeitos dos fármacos , Mutação/genética , Recidiva Local de Neoplasia/prevenção & controle , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Fosforilação Oxidativa/efeitos dos fármacos , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/genética , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Recidiva , Transdução de Sinais , Neoplasias PancreáticasRESUMO
Inflammatory breast cancer (IBC) is the most metastatic variant of locally advanced breast cancer. IBC has distinctive characteristics including invasion of tumor emboli into the skin and rapid disease progression. Given our previous studies suggesting that HDAC inhibitors have promise in targeting IBC, the present study revealed that the class I HDAC inhibitor Romidepsin (FK-288, Istodax; Celgene Corporation, Summit, NJ) potently induced destruction of IBC tumor emboli and lymphatic vascular architecture. associated with inhibition of vascular endothelial growth factor (VEGF) and hypoxia-inducible factor 1alpha, (HIF1alpha) proteins in the Mary-X pre-clinical model of IBC. Romidepsin treatment induced clinically relevant biomarkers in including induction of acetylated Histone 3 (Ac-H3) proteins, apoptosis, and increased p21WAF1/CIP1. Romidepsin, alone and synergistically when combined with Paclitaxel, effectively eliminated both primary tumors and metastatic lesions at multiple sites formed by the SUM149 IBC cell line. This is the first report of the ability of an HDAC inhibitor to eradicate IBC tumor emboli, to destroy the integrity of lymphatic vessel architecture and to target metastasis. Furthermore, Romidepsin, in combination with a taxane, warrants evaluation as a therapeutic strategy that may effectively target the skin involvement and rapid metastasis that are hallmarks of IBC.
Assuntos
Depsipeptídeos/uso terapêutico , Inibidores de Histona Desacetilases/uso terapêutico , Neoplasias Inflamatórias Mamárias/tratamento farmacológico , Células Neoplásicas Circulantes/efeitos dos fármacos , Paclitaxel/administração & dosagem , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Depsipeptídeos/administração & dosagem , Sinergismo Farmacológico , Feminino , Humanos , Neoplasias Inflamatórias Mamárias/patologia , Camundongos , Metástase Neoplásica/prevenção & controle , Esferoides Celulares , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Current treatment regimens for pancreatic ductal adenocarcinoma (PDAC) yield poor 5-year survival, emphasizing the critical need to identify druggable targets essential for PDAC maintenance. We developed an unbiased and in vivo target discovery approach to identify molecular vulnerabilities in low-passage and patient-derived PDAC xenografts or genetically engineered mouse model-derived allografts. Focusing on epigenetic regulators, we identified WDR5, a core member of the COMPASS histone H3 Lys4 (H3K4) MLL (1-4) methyltransferase complex, as a top tumor maintenance hit required across multiple human and mouse tumors. Mechanistically, WDR5 functions to sustain proper execution of DNA replication in PDAC cells, as previously suggested by replication stress studies involving MLL1, and c-Myc, also found to interact with WDR5. We indeed demonstrate that interaction with c-Myc is critical for this function. By showing that ATR inhibition mimicked the effects of WDR5 suppression, these data provide rationale to test ATR and WDR5 inhibitors for activity in this disease.
Assuntos
Histona-Lisina N-Metiltransferase/metabolismo , Neoplasias Pancreáticas/metabolismo , Proteínas Proto-Oncogênicas c-myc/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto , Adenocarcinoma/genética , Adenocarcinoma/metabolismo , Adenocarcinoma/patologia , Animais , Carcinogênese/metabolismo , Carcinogênese/patologia , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/metabolismo , Carcinoma Ductal Pancreático/patologia , Linhagem Celular Tumoral , Proliferação de Células , Dano ao DNA , Progressão da Doença , Epigênese Genética , Humanos , Peptídeos e Proteínas de Sinalização Intracelular , Lentivirus/metabolismo , Camundongos , Modelos Biológicos , Complexos Multiproteicos/metabolismo , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patologia , Ligação Proteica , Subunidades Proteicas/metabolismo , RNA Interferente Pequeno/metabolismo , Estresse FisiológicoRESUMO
Endosialin/TEM1/CD248 is a cell surface protein expressed at high levels by the malignant cells of about 50% of sarcomas and neuroblastomas. The antibody-drug conjugate (ADC) anti-endosialin-MC-VC-PABC-MMAE was selectively cytotoxic to endosialin-positive cells in vitro and achieved profound and durable antitumor efficacy in preclinical human tumor xenograft models of endosialin-positive disease. MC-VC-PABC-MMAE was conjugated with anti-endosialin with 3-4 MMAE molecules per ADC. The anti-endosialin-MC-VC-PABC-MMAE conjugate was tested for activity in four human cell lines with varied endosialin levels. The HT-1080 fibrosarcoma cells do not express endosialin, A-673 Ewing sarcoma cells and SK-N-AS neuroblastoma cells are moderate expressers of endosialin, and SJSA-1 osteosarcoma cells express very high levels of endosialin. To determine whether endosialin expression was maintained in vivo, A-673 Ewing sarcoma, SK-N-AS neuroblastoma, and SJSA-1 osteosarcoma cells were grown as xenograft tumors in nude mice. The SK-N-AS neuroblastoma and the A-673 Ewing sarcoma lines were selected for in vivo efficacy testing of the anti-endosialin-MC-VC-PABC-MMAE conjugate. The treatment groups included a vehicle control, unconjugated anti-endosialin, an admix control consisting of anti-endosialin and a dose of free MMAE equivalent to the dose administered as the ADC, and the anti-endosialin-MC-VC-PABC-MMAE conjugate. The unconjugated anti-endosialin had no antitumor activity and resulted in similar tumor growth as the vehicle control. The admix control produced a modest tumor growth delay. Administration of the anti-endosialin-MC-VC-PABC-MMAE conjugate resulted in a marked prolonged tumor response of both xenograts. These proof-of-concept results break new ground and open a promising drug discovery approach to these rare and neglected tumors.
Assuntos
Anticorpos Monoclonais Humanizados/farmacologia , Antígenos CD , Antígenos de Neoplasias , Antineoplásicos/farmacologia , Imunoconjugados , Animais , Anticorpos Monoclonais Humanizados/administração & dosagem , Anticorpos Monoclonais Humanizados/química , Antígenos CD/genética , Antígenos CD/metabolismo , Antígenos de Neoplasias/genética , Antígenos de Neoplasias/metabolismo , Antineoplásicos/administração & dosagem , Antineoplásicos/química , Linhagem Celular Tumoral , Modelos Animais de Doenças , Feminino , Citometria de Fluxo , Expressão Gênica , Humanos , Imuno-Histoquímica , Camundongos , Estrutura Molecular , Sarcoma/tratamento farmacológico , Sarcoma/genética , Sarcoma/metabolismo , Sarcoma/mortalidade , Sarcoma/patologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Mutated KRAS (KRAS*) is a fundamental driver in the majority of pancreatic ductal adenocarcinomas (PDAC). Using an inducible mouse model of KRAS*-driven PDAC, we compared KRAS* genetic extinction with pharmacologic inhibition of MEK1 in tumor spheres and in vivo. KRAS* ablation blocked proliferation and induced apoptosis, whereas MEK1 inhibition exerted cytostatic effects. Proteomic analysis evidenced that MEK1 inhibition was accompanied by a sustained activation of the PI3K-AKT-MTOR pathway and by the activation of AXL, PDGFRa, and HER1-2 receptor tyrosine kinases (RTK) expressed in a large proportion of human PDAC samples analyzed. Although single inhibition of each RTK alone or plus MEK1 inhibitors was ineffective, a combination of inhibitors targeting all three coactivated RTKs and MEK1 was needed to inhibit proliferation and induce apoptosis in both mouse and human low-passage PDAC cultures. Importantly, constitutive AKT activation, which may mimic the fraction of AKT2-amplified PDAC, was able to bypass the induction of apoptosis caused by KRAS* ablation, highlighting a potential inherent resistance mechanism that may inform the clinical application of MEK inhibitor therapy. This study suggests that combinatorial-targeted therapies for pancreatic cancer must be informed by the activation state of each putative driver in a given treatment context. In addition, our work may offer explanative and predictive power in understanding why inhibitors of EGFR signaling fail in PDAC treatment and how drug resistance mechanisms may arise in strategies to directly target KRAS.