Marcadores genéticos e bioquímicos no diagnóstico e prognóstico do carcinoma hepatocelular

Abstract

Hepatocellular carcinoma (HCC) stands out as the most aggressive malignant liver tumor. The identification of genetic and biochemical markers may help to understand the physiopathology of HCC and to make an early diagnosis of the disease with new therapeutic interventions. Objectives - To evaluate the association of genetic variants (VEGF-C936T e VEGF-A1154G), serum levels and gene expression involved in the angiogenesis process, protection, communication and cell signaling with HCC. Patients and Methods - A total of 393 subjects were studied, distributed into Group 1 (G1) - 102 patients with HCC; Group 2 (G2) - 117 patients with cirrhosis of any etiology; Group 3 (G3) - 33 patients with hepatitis C virus (HCV); Group 4 (G4) - 141 individuals without clinical or biochemical signs of any liver disease (controls). For gene expression analysis of vascular endothelial growth factor (VEGF), β-catenina (CTNNB1), Alpha-fetoprotein (AFP), Cystatin-B (CSTB) and Glypican-3 (GPC3), 26 patients with HCC, 16 with cirrhosis, 9 with HCV and 14 controls undergoing bileoenteric surgery were selected. The polymorphisms were analyzed by PCR/RFLP (polymerase chain reaction/ restriction fragments length polymorphism), gene expression (liver tissue) by qPCR (quantitative / polymerase chain reaction), and serum levels by ELISA (Enzyme Linked Immunosorbent Assay). α -error was set at 5%. Results - Wild genotype both VEGF polymorphisms prevailed in all the groups (P> 0.05). Increased levels of VEGF were observed in G1 and G3, related to the mutant allele of VEGF-C936T and VEGF-A1154G, respectively, compared to the wild-type genotype (P=0.028; P=0.028), in G1 vs. G2 and G3 for VEGF-C936T, and in G1 compared to G2 for VEGF-A1154G (P<0.05 for both). Overexpression of VEGF in G1 (3.85) was observed, compared to G2 (0.76) and G3 (1.95; P=0.02 for both). Higher levels of expression of VEGF and AFP could be observed in patients classified into stage D (19.97 and 8.86), compared to those in stage B (1.96 and 0.03;P=0.008 and P=0.001, respectively). Patients in stage C showed higher expression of CTNNB1, compared to those in stage B (18.40 vs. 0.90; P=0.008). There was an increase of GPC3 (2.8ng/mL), CSTB (0.99ng/mL), VEGF (264.8pg/mL) and AFP (20.3ng/mL) in patients with HCC, compared to controls (2.9ng/mL, 0.6ng/mL, 185.2pg/mL, 2.6ng/mL; P<0.05 for all comparisons). GPC3 was relevant when discriminating G1 and G4 (area under the curve = 0.82). For VEGF, discriminatory power between HCC patients and controls (area under the curve = 0.71) was observed. CSTB showed discriminatory power when comparing groups with HCV and controls (area under the curve = 0.74). A positive correlation was observed for expression among all genes, except CSTB, which correlated only with CTNNB1 (r=0.43; P=0.01), and GPC3, which correlated only with VEGF (r= 0.49; P=0.008). Conclusion – VEGF-A polymorphisms (C936T and A1154G) are not associated with cirrhosis, HCC and survival, however, the presence of the mutant alleles may influence serum levels of VEGF. Emphasis should be given to the sensitivity of serum CSTB (cell protection) for the diagnosis of HCV, VEGF (angiogenesis) and serum GPC3 (cell proliferation) demonstrate sensitivity and specificity for the diagnosis of HCC. Increased expression, particularly of VEGF in patients with end-stage HCC (BCLC-D) confirms VEGF as a prognostic indicator in liver carcinogenesis. The correlation between VEGF expression with AFP and GPC3 CTNNB1 suggests integration of markers of angiogenesis, signaling and cell proliferation in HCC and hepatitis C.