Molecular profiling of liver cancer: Genetic and epigenetic variation analysis for developing potential targets for therapy

Hepatocellular carcinoma (HCC) is one of the most common cancers in the world with poor prognosis. HCC is a complex cancer, marked by vast cellular and molecular heterogeneity. This project aimed to discover genetic and epigenetic targets molecules as potential therapeutic efficacy that may encompass HCC heterogeneity. This is a translational study, combining data taken from human HCC specimens, in vitro model, in vivo mouse model and in silico analysis. For in vitro, we used models of heterogeneity composed of various HCC cell lines under two subtypes: S1/TGFβ-Wnt-activated, S2/progenitor HCC subtype compared to a normal immortalized hepatocyte. The first task was the investigation on the epigenetic role of suppressor of cytokine signaling 1 (SOCS1), a gene involved in hepatocarcinogenesis. The presence of SOCS1 methylation was significantly higher in HCC tissues compared to peri- and non-tumoral tissues. In vitro treatment with 5-Azacytidine (5-AZA) reduced DNMT1 expression and restored SOCS1 expression to different extents across cells. The second task was to discover potential targets for HCC therapy. An in silico strategy was performed to identify potential molecular targets. Protein-protein interaction analysis (PPI) using published datasets identified 16 proto-oncogenes targets. In vitro evaluation of three treatment modalities, 5-AZA, Sorafenib (SOR), and PD-L1 gene silencing revealed that SOR was effective to down-regulate targets mostly in S2/progenitor subtype while PD-L1 silencing was able to decrease the expression of targets in all HCC subtypes. This suggests that PD-L1 regulation may be a good approach to encompass HCC heterogeneity. On the third task, we performed further analysis on the interaction of those targets in task 2, through gene enrichment analysis. Data showed a unique clustering of Src family of kinases (SFKs). In vitro and in vivo assessments confirmed dysregulation of SFKs during liver damage. Two SFK inhibitors were evaluated as monotherapy and in combination with SOR. Data showed a promising potential of SFK inhibitors in combination therapy, but not as monotherapy, indicated by a dose-dependent toxicity to HCC cells. In conclusion, this study highlights that HCC response to therapy is cellular subtype dependent. This study proposes that a combination therapy may be a potential treatment modality to improve HCC therapy to encompass cellular heterogeneity.

Hepatocellular carcinoma (HCC) is one of the most common cancers in the world with poor prognosis. HCC is a complex cancer, marked by vast cellular and molecular heterogeneity. This project aimed to discover genetic and epigenetic targets molecules as potential therapeutic efficacy that may encompass HCC heterogeneity. This is a translational study, combining data taken from human HCC specimens, in vitro model, in vivo mouse model and in silico analysis. For in vitro, we used models of heterogeneity composed of various HCC cell lines under two subtypes: S1/TGFβ-Wnt-activated, S2/progenitor HCC subtype compared to a normal immortalized hepatocyte. The first task was the investigation on the epigenetic role of suppressor of cytokine signaling 1 (SOCS1), a gene involved in hepatocarcinogenesis. The presence of SOCS1 methylation was significantly higher in HCC tissues compared to peri- and non-tumoral tissues. In vitro treatment with 5-Azacytidine (5-AZA) reduced DNMT1 expression and restored SOCS1 expression to different extents across cells. The second task was to discover potential targets for HCC therapy. An in silico strategy was performed to identify potential molecular targets. Protein-protein interaction analysis (PPI) using published datasets identified 16 proto-oncogenes targets. In vitro evaluation of three treatment modalities, 5-AZA, Sorafenib (SOR), and PD-L1 gene silencing revealed that SOR was effective to down-regulate targets mostly in S2/progenitor subtype while PD-L1 silencing was able to decrease the expression of targets in all HCC subtypes. This suggests that PD-L1 regulation may be a good apprach to encompass HCC heterogeneity. On the third task, we performed further analysis on the interaction of those targets in task 2, through gene enrichment analysis. Data showed a unique clustering of Src family of kinases (SFKs). In vitro and in vivo assesments confirmed dysregulation of SFKs during liver damage. Two SFK inhibitors were evaluated as monotherapy and in combination with SOR. Data showed a promising potential of SFK inhibitors in combination therapy, but not as monotherapy, indicated by a dose-dependent toxicity to HCC cells. In conclusion, this study highlights that HCC response to therapy is cellular subtype dependent. This study proposes that a combination therapy may be a potential treatment modality to improve HCC therapy to encompass cellular heterogeneity.