High frequency of telomerase reverse-transcriptase promoter somatic mutations in hepatocellular carcinoma and preneoplastic lesions

Abstract

Somatic mutations activating telomerase reverse-trancriptase promoter were recently identified in several tumour types. Here we identify frequent similar mutations in human hepatocellular carcinomas (59%), cirrhotic preneoplastic macronodules (25%) and hepatocellular adenomas with malignant transformation in hepatocellular carcinomas (44%). In hepatocellular tumours, telomerase reverse-transcripase- and CTNNB1-activating mutations are significantly associated. Moreover, preliminary data suggest that telomerase reverse-trancriptase promoter mutations can increase the expression of telomerase transcript. In conclusion, telomerase reverse-trancriptase promoter mutation is the earliest recurrent genetic event identified in cirrhotic preneoplastic lesions so far and is also the most frequent genetic alteration in hepatocellular carcinomas, arising from both the cirrhotic or non-cirrhotic liver.

Figure 1. Somatic mutations of the TERT promoter in human liver carcinogenesis.

(a) Mutation spectrum of the TERT promoter in 179 human HCCs: substitution at two hot spots (−124 and −146 bp from the ATG start site, g.1,295,228 and g.1,295,250, respectively) that create a new ETS-/TCF-binding sequences. Distribution (%) of the 179 mutations along the TERT promoter (59% of all HCCs) is indicated. Mutations are represented on the +strand of DNA. (b) TERT promoter mutations in the multistep process of liver carcinogenesis. Percentage of TERT promoter mutations is reported at each step of liver carcinogenesis. On cirrhosis, 5/20 (25%) cirrhotic macronodules (three with dysplasia and two without dysplasia) and 71/110 (65%) HCCs were mutated for TERT. On normal liver, no mutations of the TERT promoter (0%) were identified among 60 HCAs; in contrast, 7/16 (48%) of malignant transformation of HCA and 58/106 (55%) of HCCs without signs of malignant transformation from HCA exhibited mutations of the TERT promoter. Other genes most frequently mutated in HCC and HCA are indicated.

Figure 2. TERT transcript expression according to mutation status of the TERT promoter.

(a) TERT messenger RNA (mRNA) is not expressed in 4 normal livers, 24 cirrhosis and 48 HCAs, whereas it is overexpressed in 156 HCCs mutated and 109 HCCs not mutated for the TERT promoter. (b) TERT transcript was overexpressed in all mutated cirrhotic macronodules (n=4) when compared with non-mutated macronodules (n=8). (c) TERT mRNA was not overexpressed in HCA with CTNNB1 mutations (n=9) compared with HCA without CTNNB1 mutations (n=39). (d) β-catenin target genes (GLUL and LGR5) were not overexpressed in HCC harbouring TERT promoter mutations without CTNNB1 mutations (n=82) compared with HCC lacking both TERT promoter and CTNNB1 mutations (n=80). TERT, GLUL and LGR5 mRNA levels were measured using quantitative RT–PCR. All the results were normalized with the mean of normal liver tissues (see Methods section). Results were reported in median with interquartile range and compared using non-parametric Mann–Whitney test. Differences were considered significant when the P-value was <0.05.