Characterization of seven human hepatocellular carcinoma cell lines

Abstract

Hepatocellular carcinoma (HCC) continues to secure its spot among top ranks in cancer-related deaths worldwide. Known for its difficulty in management, studies to find effective treatments for HCC are on-going. To assist further in vitro study, seven HCC cell lines, SNU-1517, SNU-2655, SNU-2658, SNU-2663, SNU-3058, SNU-3059, and SNU-3160, derived from Korean patients were characterized in this paper. Growth rate varied between HCC cell lines where the slowest growing cell line had doubling time of 74.59 hours and the fastest growing cell line had its doubling time of 42.93 hours. DNA finger printing analysis was carried out for authentication of cell lines. Targeted sequencing was performed for detection of mutations of genes within comprehensive cancer panel. Frame shift deletions, insertions, in frame deletions and insertions, missense, nonsense mutations, start codon deletion and splice site mutations of various genes were found and classified based on their pathogenicity reports. Also, some of these mutations were validated by Sanger sequencing. Also this study confirms that only three (SNU-1517, SNU-2655 and SNU-3058) out of 7 HCC cell lines were shown to have integrated HBV DNA in their genome and were identified via Sanger sequencing. In addition, cell viability assay was carried out to test the sensitivity to three anti-cancer drugs widely used for HCC, 5-fluorouracil (5-FU), cisplatin, and sorafenib. Among these three chemotherapeutic agents, sorafenib is the most prominent drug known to be used in HCC regimen. Therefore, proteins involved in this pathway were tested for change in gene expression. Phospho-ERK (p-ERK) expression was examined to confirm that sorafenib successfully inhibited Ras-Raf-MEK-ERK signaling pathway. For SNU-1517, SNU-2663, and SNU-3160 cell lines, sorafenib clearly made difference in p-ERK level. However, on the other HCC cell lines, effect of sorafenib was difficult to determine since SNU-2655, 2658, and 3058 had very low basal p-ERK level to verify sorafenib inhibition, and SNU-3059 barely showed any change in expression level after sorafenib treatment. In addition to sorafenib, four phytochemicals, baicalein, curcumin, genistein and resveratrol were tested to evaluate both their individual and synergetic anti-cancer effect when partnered with sorafenib. All phytochemicals were shown to have effect on all HCC cell lines at certain concentrations, although their synergetic effect with sorafenib was ambiguous and demands further study.