Yoichi Hiasa, Nobuyuki Enomoto, Yasuhiro Asahina, Atsumasa Komori, Atsunori Kusakabe, Shintaro Ogawa, Koichi Takaguchi, Masanori Isogawa, Hitoshi Yoshiji, Kazuho Ikeo, Tatsuya Ide, Mina Nakagawa, Atsushi Suetsugu, Yosuke Kawai, Eiichi Tomita, Ken Shirabe, Takuya Genda, Shuhei Nishiguchi, Kaname Kojima, Eiji Kajiwara, Noritomo Shimada, Misako Matsubara, Akihiro Tamori, Namiki Izumi, Masayuki Kurosaki, Naoya Sakamoto, Hisayoshi Watanabe, Masao Honda, Isao Sakaida, Etsuko Iio, Yasuhito Tanaka, Masao Nagasaki, Tadashi Namisaki, Takashi Kumada, Yasuteru Kondo, Katsushi Tokunaga, Hiromi Sawai, Hidenori Toyoda, Junichi Sugihara, Kentaro Matsuura, Shuichi Kaneko, Mitsuo Shimada, Yoshito Itoh, Norifumi Kawada, Sohji Nishina, and Eiji Tanaka
Background & Aims There is still a risk for hepatocellular carcinoma (HCC) development after eradication of hepatitis C virus (HCV) infection with antiviral agents. We investigated genetic factors associated with the development of HCC in patients with a sustained virologic response (SVR) to treatment for chronic HCV infection. Methods We obtained genomic DNA from 457 patients in Japan with a SVR to interferon-based treatment for chronic HCV infection from 2007 through 2015. We conducted a genome-wide association study (GWAS), followed by a replication analysis of 79 candidate single nucleotide polymorphisms (SNPs) in an independent set of 486 patients in Japan. The study end point was HCC diagnosis or confirmation of lack of HCC (at follow-up examinations until December 2014 in the GWAS cohort, and until January 2016 in the replication cohort). We collected clinical and laboratory data from all patients. We analyzed expression levels of candidate gene variants in human hepatic stellate cells, rats with steatohepatitis caused by a choline-deficient L-amino acid-defined diet, and a mouse model of liver injury caused by administration of carbon tetrachloride. We also analyzed expression levels in liver tissues of patients with chronic HCV infection with different stages of fibrosis or tumors vs patients without HCV infection (controls). Results We found a strong association between the SNP rs17047200, located within the intron of the tolloid like 1 gene ( TLL1 ) on chromosome 4, and development of HCC; there was a genome-wide level of significance when the results of the GWAS and replication study were combined (odds ratio, 2.37; P = 2.66 × 10 −8 ). Multivariate analysis showed rs17047200 AT/TT to be an independent risk factor for HCC (hazard ratio, 1.78; P = .008), along with male sex, older age, lower level of albumin, advanced stage of hepatic fibrosis, presence of diabetes, and higher post-treatment level of α-fetoprotein. Combining the rs17047200 genotype with other factors, we developed prediction models for HCC development in patients with mild or advanced hepatic fibrosis. Levels of TLL1 messenger RNA (mRNA) in human hepatic stellate cells increased with activation. Levels of Tll1 mRNA increased in liver tissues of rodents with hepatic fibrogenesis compared with controls. Levels of TLL1 mRNA increased in liver tissues of patients with progression of fibrosis. Gene expression levels of TLL1 short variants, including isoform 2, were higher in patients with rs17047200 AT/TT. Conclusions In a GWAS, we identified the association between the SNP rs17047200, within the intron of TLL1 , and development of HCC in patients who achieved an SVR to treatment for chronic HCV infection. We found levels of Tll1/TLL1 mRNA to be increased in rodent models of liver injury and liver tissues of patients with fibrosis, compared with controls. We propose that this SNP might affect splicing of TLL1 mRNA, yielding short variants with high catalytic activity that accelerates hepatic fibrogenesis and carcinogenesis. Further studies are needed to determine how rs17047200 affects TLL1 mRNA levels, splicing, and translation, as well as the prevalence of this variant among other patients with HCC. Tests for the TLL1 SNP might be used to identify patients at risk for HCC after an SVR to treatment of HCV infection.