RESUMEN
Single-cell multi-omics can provide a unique perspective on tumor cellular heterogeneity. Most previous single-cell whole-genome RNA sequencing (scWGS-RNA-seq) methods demonstrate utility with intact cells from fresh samples. Among them, many are not applicable to frozen samples that cannot produce intact single-cell suspensions. We have developed scONE-seq, a versatile scWGS-RNA-seq method that amplifies single-cell DNA and RNA without separating them from each other and hence is compatible with frozen biobanked samples. We benchmarked scONE-seq against existing methods using fresh and frozen samples to demonstrate its performance in various aspects. We identified a unique transcriptionally normal-like tumor clone by analyzing a 2-year frozen astrocytoma sample, demonstrating that performing single-cell multi-omics interrogation on biobanked tissue by scONE-seq could enable previously unidentified discoveries in tumor biology.
Asunto(s)
Multiómica , Neoplasias , Humanos , Neoplasias/genética , RNA-Seq/métodos , Genotipo , FenotipoAsunto(s)
Transformación Celular Neoplásica/metabolismo , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Serina-Treonina Quinasa 3/metabolismo , Neoplasias Gástricas/etiología , Neoplasias Gástricas/metabolismo , Proteínas ras/metabolismo , Biomarcadores de Tumor , Ciclo Celular/genética , Transformación Celular Neoplásica/genética , Susceptibilidad a Enfermedades , Regulación Neoplásica de la Expresión Génica , Humanos , Oncogenes , Pronóstico , Serina-Treonina Quinasa 3/genética , Transducción de Señal , Neoplasias Gástricas/diagnóstico , Neoplasias Gástricas/mortalidadAsunto(s)
Neoplasias Encefálicas/genética , Neoplasias Encefálicas/patología , Oligodendroglioma/genética , Oligodendroglioma/patología , Adolescente , Niño , Receptores ErbB/genética , Femenino , Amplificación de Genes , Humanos , Masculino , Mutación , Proteínas Proto-Oncogénicas B-raf/genética , Proteínas Proto-Oncogénicas c-met/genéticaRESUMEN
Despite significant medical advances, glioblastoma multiforme (GBM) remains a formidable therapeutic challenge. Advent of targeted capture sequencing and patients-derived organoid cultures may hold the key to scientifically sound individualized treatment approaches. We report on our initial experience of using the combination of these two technologies to create a tailored approach of systemic therapies for a patient with GBM, which challenges the conventional treatment paradigm, as well as specifically highlighting the complexities of such an approach and the potential for a more favorable treatment outcome.
Asunto(s)
Glioblastoma/tratamiento farmacológico , Glioblastoma/patología , Terapia Molecular Dirigida/métodos , Medicina de Precisión/métodos , Neoplasias Encefálicas/patología , Humanos , Organoides/efectos de los fármacos , Organoides/patología , Resultado del Tratamiento , Células Tumorales Cultivadas/efectos de los fármacosRESUMEN
Fibroblast growth factors (FGFs) and their receptors are significant components during fundamental cellular processes. FGF18 plays a distinctive role in modulating the activity of both tumor cells and tumor microenvironment. This study aims to comprehensively investigate the expression and functional role of FGF18 in gastric cancer (GC) and elucidate its regulatory mechanisms. The upregulation of FGF18 was detected in seven out of eleven (63.6%) GC cell lines. In primary GC samples, FGF18 was overexpressed in genomically stable and chromosomal instability subtypes of GC and its overexpression was associated with poor survival. Knocking down FGF18 inhibited tumor formation abilities, induced G1 phase cell cycle arrest and enhanced anti-cancer drug sensitivity. Expression microarray profiling revealed that silencing of FGF18 activated ATM pathway but quenched TGF-ß pathway. The key factors that altered in the related signaling were validated by western blot and immunofluorescence. Meanwhile, treating GC cells with human recombinant FGF18 or FGF18-conditioned medium accelerated tumor growth through activation of ERK-MAPK signaling. FGF18 was further confirmed to be a direct target of tumor suppressor, miR-590-5p. Their expressions showed a negative correlation in primary GC samples and more importantly, re-overexpression of FGF18 partly abolished the tumor-suppressive effect of miR-590-5p. Our study not only identified that FGF18 serves as a novel prognostic marker and a therapeutic target in GC but also enriched the knowledge of FGF-FGFR signaling during gastric tumorigenesis.
Asunto(s)
Factores de Crecimiento de Fibroblastos/fisiología , MicroARNs/genética , Proteínas de Neoplasias/fisiología , ARN Neoplásico/genética , Neoplasias Gástricas/metabolismo , Animales , Comunicación Autocrina , Línea Celular Tumoral , Transformación Celular Neoplásica , Inestabilidad Cromosómica , Factores de Crecimiento de Fibroblastos/antagonistas & inhibidores , Factores de Crecimiento de Fibroblastos/genética , Factores de Crecimiento de Fibroblastos/farmacología , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Técnicas de Silenciamiento del Gen , Xenoinjertos , Humanos , Ratones , MicroARNs/antagonistas & inhibidores , Proteínas de Neoplasias/antagonistas & inhibidores , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/farmacología , Pronóstico , Interferencia de ARN , ARN Neoplásico/antagonistas & inhibidores , Proteínas Recombinantes/farmacología , Transducción de Señal/efectos de los fármacos , Transducción de Señal/genética , Neoplasias Gástricas/etiología , Neoplasias Gástricas/genética , Regulación hacia ArribaRESUMEN
Slit-Robo GTPase-activating protein 1 (SRGAP1) functions as a GAP for Rho-family GTPases and downstream of Slit-Robo signaling. We aim to investigate the biological function of SRGAP1 and reveal its regulation by deregulated microRNAs (miRNAs) in gastric cancer (GC). mRNA and protein expression of SRGAP1 were examined by quantitative reverse transcription PCR (qRT-PCR) and western blot. The biological role of SRGAP1 was demonstrated through siRNA-mediated knockdown experiments. The regulation of SRGAP1 by miR-340 and miR-124 was confirmed by western blot, dual luciferase activity assays and rescue experiments. SRGAP1 is overexpressed in 9 out of 12 (75.0%) GC cell lines. In primary GC samples from TCGA cohort, SRGAP1 shows gene amplification in 5/258 (1.9%) of cases and its mRNA expression demonstrates a positive correlation with copy number gain. Knockdown of SRGAP1 in GC cells suppressed cell proliferation, reduced colony formation, and significantly inhibited cell invasion and migration. Luciferase reporter assays revealed that SRGAP1 knockdown significantly inhibited Wnt/ß-catenin pathway. In addition, SRGAP1 was found to be a direct target of two tumor-suppressive miRNAs, miR-340 and miR-124. Concordantly, these two miRNAs were downregulated in primary gastric tumors and these decreasing levels w5ere associated with poor outcomes. Expression of miR-340 and SRGAP1 displayed a reverse relationship in primary samples and re-expressed SRGAP1, rescued the anti-cancer effects of miR-340. Taken together, these data strongly suggest that, apart from gene amplification and mutation, the activation of SRGAP1 in GC is partly due to the downregulation of tumor-suppressive miRNAs, miR-340 and miR-124. Thus SRGAP1 is overexpressed in gastric carcinogenesis and plays an oncogenic role through activating Wnt/ß-catenin pathway.