Expression of MAPK and PI3K/AKT/mTOR Proteins according to the Chronic Liver Disease Etiology in Hepatocellular Carcinoma

Abstract

Aims: Chronic liver disease (CLD) of different etiologies leads to hepatocellular carcinoma (HCC) by multiple mechanisms that may be translated into clinicopathological differences. We evaluated the tissue expression of the MAPK and PI3K/Akt/mTOR pathway proteins and their association with long-term outcome and other parameters, according to the etiology of the CLD, in HCC patients.

Methods: Clinicopathological data from 80 patients who underwent orthotopic liver transplantation for HCC treatment in a Brazilian referral center were compared according to CLD etiology. Event (tumor recurrence or death from any cause) occurrence and event-free survival (EFS) were analyzed. Pathway protein expression was assessed by immunohistochemistry (IHQ) in both tumor and underlying cirrhosis and by RT-PCR in tumor tissue.

Results: Strong expression (SE) of KRAS was more frequent in tumors arising from viral (26.8%) than the nonviral group of liver disease (7.7%, p=0.024) and also than cirrhotic parenchyma (0%, p=0.004). SE of PI3K was more frequent in tumor than in cirrhosis (p=0.048, p < 0.01), without differences in its tumor expression among etiologic groups (p=0.111). mRNA of ERK, PI3K, and BRAF was expressed in the tumor, without differences between CLD etiologies, and there was no association with IHQ findings. Older age and microvascular invasion (MIV) were the only parameters independently associated with the event. MIV was also associated with shorter EFS.

Conclusions: Hepatitis B and C virus can lead to HCC by different mechanisms compared with nonviral hepatopathy. KRAS and PI3K may have a role in carcinogenesis. The prognostic and therapeutic implications need to be investigated.

Figure 1

Frequency of strong expression of MAPK pathway proteins in hepatocellular carcinoma patients according to the etiology of the underlying chronic liver disease in tumor and adjacent cirrhosis.

Figure 2

Frequency of strong expression of PI3K/AKT/mTOR pathway proteins in hepatocellular carcinoma patients according to the etiology of underlying chronic liver disease in tumor and adjacent cirrhosis.

Figure 3

Relative RNA expression of some pathway proteins in tumor tissue according to the etiology of the underlying chronic liver disease. The data are normalized for the level of GAPDH and expressed as 2^−ΔCt. ΔCt refers to the difference between Ct of GAPDH and Ct of primer analyzed per sample. The results are expressed as median (interquartile range), three technical replicates per sample. (a) BRAF 0.423 (0.003–0.573), n = 8/8; 0.222 (0.052–0.722), n = 7/8; 0.183 (0.043–0.604), n = 8/10; p=0.25. (b) ERK1 0.182 (0.008–0.281), n = 4/8; median NA, n = 3/8; 1.041 (0.064–18.480), n = 4/10; p = =0.08. (c) ERK2 0.033 (0.006–0.633), n = 4/8; 0.125 (0.037–NA), n = 3/8; 0.375 (0.006–0.744), n = 4/10; p=0.258. (d) PI3K 0.007 (0.002–0.383), n = 4/8; 0.188 (0.006–0.308), n = 4/8; median NA, n = 2/10; p=0.480. All data refer to cryptogenic, ethanolic, and HCV etiologic groups, respectively. Comparisons were performed among the three groups (Kruskal–Wallis test) or between two groups (Mann–Whitney U test), and no differences were detected. It was impossible to extract RNA from the hepatitis B virus etiology and data are not shown. AKT and mTOR were also tested, but the relative RNA expression was not specifically labeled, and the data could not be shown. GAPDH, glyceraldehyde 3-phosphate dehydrogenase; HCV, hepatitis C virus, Ct, cycle threshold; CLD, chronic liver disease; NA, not available.