Hepatocarcinoma in chronic viral hepatitis: from epidemiological data, through new pathophysiological findings, up to new possible clinical tools

Background and aim: Hepatocellular carcinoma (HCC) is the first cause of liver tumor and the leading cause of death among patients with liver cirrhosis. HCC is a major public health concern, with an incidence and mortality of HCC in increasing in North America and in some European regions1. In this contest, is strategically relevant to determine HCC epidemiology, physiopathological alterations and its potential early biomarkers. The aim of the three studies presented in this thesis is to define HCC incidence in HCV patients treated with direct acting antiviral drugs (DAAs), and investigate the role of novel targets and circulating biomarker in HCC related to chronic viral hepatitis. The main aim of study 1 is to evaluate incidence of newly diagnosed HCC in patients with advanced hepatitis C treated with DAAs. Aim of study 2 is to evaluate the expression of mi-RNAs in patients with HCC, compared with patients without HCC. The main aims of study 3 is to better define the role of OPN in HCC and elucidate the immunoregulatory functions of OPN in Hepatocellular Carcinoma Methods: Study1. The study is based on the NAVIGATORE platform, a prospectively recording database of all patients with hepatitis C receiving DAAs in Veneto region (Italy). Inclusion criteria: fibrosis stage ≥ F3. Exclusion criteria: Child-Pugh C, liver transplantation before DAAs, history or presence of HCC, follow-up <4 weeks after starting DAAs. Study 2. A total of 68 patients with cirrhosis HCV related who achieved SVR after therapy with DAAs have bee enrolled. 20 patients with HCC within the 24 weeks after the end of therapy with DAAs and 48 patients without HCC after DAAs. In the two groups selected micro-RNAs have been analyzed at the start and at the end of DAAs therapy. Study 3. This study includes: measurement of Osteopontin (OPN) in the plasma of patients with HCC, analysis of OPN on supernatant of Peripheral blood mononuclear cells (PBMCs) culture in HCC patients and evaluation of the impact of recombinant OPN on modulating the phenotype of PBMCs in patients with HCC. Results: Study 1. During the first year, HCC incidence was 0.46% (95% CI: 0.12-1.17) in F3, 1.49% (1.03-2.08) in Child-Pugh-A and 3.61% (1.86-6.31) in Child-Pugh-B cirrhotics. In the second year HCC incidences were: 0%, 0.2%, and 0.69%, respectively. Study 2. When we have compared mi-RNAs expression between the two groups (patients with HCC or without HCC after DAAs) at baseline, miR28-5p is resulted significantly increased (p=0.029) in HCC group compared to not HCC group. Study 3. In patients with HCC hepatitis B (HCC-CHB) related and with HCC non-viral related, the levels of OPN are higher than patients with chronic hepatitis without HCC (p=0.0016 and p=0.0063 respectively). In the analysis of OPN on supernatant of PBMCs cultured with Glypican 3, following PD1 blockade, OPN production was reduced (p=0.023) in HCC-non viral but not HCC-CHB or Healthy controls. We also observed a reduction in OPN in both HCC-non viral (p=0.0078) and HCC-CHB (p=0.0010) in PBMCs cultured with p53. Conclusions: Study 1 These data, obtained in a large, prospective, population-based study, indicate that in patients with advanced hepatitis C receiving DAAs, the risk of "de novo" hepatocarcinoma during the first year is not higher, and might be lower, than that of untreated patients, and further declines thereafter. Study 2. In conclusion the expression of miR 28-5p is altered in patients treated with DAAs who develop HCC, suggesting its potential prognostic role in HCC related to HCV. Study 3. These results suggest its use as biomarker of HCC and reveal an immunomodulatory role for Osteopontin in HCC and warrant its further investigation as an immunotherapeutic target for HCC.