RESUMEN
OBJECTIVE: We investigated the mutational landscape of mammalian target of rapamycin (mTOR) signalling cascade in hepatocellular carcinomas (HCCs) with chronic HBV background, aiming to evaluate and delineate mutation-dependent mechanism of mTOR hyperactivation in hepatocarcinogenesis. DESIGN: We performed next-generation sequencing on human HCC samples and cell line panel. Systematic mutational screening of mTOR pathway-related genes was undertaken and mutant genes were evaluated based on their recurrence. Protein expressions of tuberous sclerosis complex (TSC)1, TSC2 and pRPS6 were assessed by immunohistochemistry in human HCC samples. Rapamycin sensitivity was estimated by colony-formation assay in HCC cell lines and the treatment was further tested using our patient-derived tumour xenograft (PDTX) models. RESULTS: We identified and confirmed multiple mTOR components as recurrently mutated in HBV-associated HCCs. Of significance, we detected frequent (16.2%, n=18/111) mutations of TSC1 and TSC2 genes in the HCC samples. The spectrum of TSC1/2 mutations likely disrupts the endogenous gene functions in suppressing the downstream mTOR activity through different mechanisms and leads to more aggressive tumour behaviour. Mutational disruption of TSC1 and TSC2 was also observed in HCC cell lines and our PDTX models. TSC-mutant cells exhibited reduced colony-forming ability on rapamycin treatment. With the use of biologically relevant TSC2-mutant PDTXs, we demonstrated the therapeutic benefits of the hypersensitivity towards rapamycin treatment. CONCLUSIONS: Taken together, our findings suggest the significance of previously undocumented mutation-dependent mTOR hyperactivation and frequent TSC1/2 mutations in HBV-associated HCCs. They define a molecular subset of HCC having genetic aberrations in mTOR signalling, with potential significance of effective specific drug therapy.
Asunto(s)
Antibióticos Antineoplásicos/uso terapéutico , Carcinoma Hepatocelular/genética , Neoplasias Hepáticas/genética , Sirolimus/uso terapéutico , Serina-Treonina Quinasas TOR/metabolismo , Proteínas Supresoras de Tumor/genética , Adulto , Anciano , Animales , Antibióticos Antineoplásicos/farmacología , Proteína Axina/genética , Carcinoma Hepatocelular/química , Carcinoma Hepatocelular/metabolismo , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Proliferación Celular/genética , Análisis Mutacional de ADN , Proteínas de Unión al ADN , Femenino , Humanos , Neoplasias Hepáticas/química , Neoplasias Hepáticas/metabolismo , Masculino , Ratones , Persona de Mediana Edad , Tasa de Mutación , Trasplante de Neoplasias , Proteínas Nucleares/genética , Transducción de Señal , Sirolimus/farmacología , Factores de Transcripción/genética , Proteína 1 del Complejo de la Esclerosis Tuberosa , Proteína 2 del Complejo de la Esclerosis Tuberosa , Ensayo de Tumor de Célula Madre , Proteína p53 Supresora de Tumor/genética , Proteínas Supresoras de Tumor/análisis , Adulto Joven , beta Catenina/genéticaRESUMEN
Spinal muscular atrophy (SMA) is the most common genetic motoneuron degenerative disorder, but the mechanism(s) of motoneuron degeneration is unclear. We previously generated SMA model mice, which genotypically and phenotypically mimicked human SMA patients, by a combination of knockout and transgenic techniques. Here, we used these SMA model mice to decipher the apoptotic mechanism(s) involved in SMA motoneuron degeneration. We found a significant increase in proapoptotic Bax expression in the spinal cords of SMA mice in comparison with their wild-type littermates. After crossing SMA mice with Bax knockout mice, we produced in vivo evidence indicating that Bax protein plays an important role in the degeneration of SMA spinal motoneurons. Progeny Bax-deficient SMA mice showed milder disease severity, longer life spans, and significant increases in spinal motoneuron densities compared to SMA littermates with wild-type Bax genes. Our results strongly suggest that suppression of Bax-involved apoptosis has the potential for amelioration of SMA.