Mitotic Checkpoint CHK2 Upregulation is Linked to Chromosomal Instability in Human Hepatocellular Carcinoma

Background and Aims: Although the epidemiologic risk factors for hepatocellular carcinoma (HCC) are well known, the molecular mechanisms underlying hepatocarcinogenesis are not well characterized. Surgery, liver transplantation, and radiologic intervention are viable options for patients with HCC, however the prognosis still remains uncertain. Histologically, 80% of small hepatocellular carcinoma are moderately differentiated, and another 20% consist of varying mixtures of cancerous tissuesChromosomal instability (CIN), is a major hallmark of HCC progression and hampers prognosis and therapy. Methods: To identify genes involved in CIN we screened specimens obtained from patients with primary hepatocellular carcinoma for the expression of mitotic checkpoint gene CHK2 . Chk2 promotes proper mitotic spindle assembly, chromosome alignment, and segregation through largely undisclosed mechanisms. First, with the use of immunohistochemical analysis we investigated the expression levels of Chk2 in liver biopsy of patients with cirrhotic- related hepatocellular carcinoma (n = 40), focal nodular hyperplasia (n = 10), and normal liver (n = 40). Results: We observed differential expression of Chk2 in grade I and grade II HCC identifying different subtypes within a histological grade. Normal liver and focal nodular hyperplasia were negative. These findings prompted us to investigate the function of Chk2 in HCC assuming that overexpressed Chk2 may induce pathways not normally triggered by Chk2 in non- transformed hepatocytes. Chk2-loss and gain of function studies were accomplished with a panel of HCC cell lines. Most importantly, immortalized human hepatocytes with telomerase reverse transcriptase TERT , a gene frequently activated in HCC, showed overexpressed Chk2 localized with centrosome proteins γ -tubulin and Nek2 kinase to mitotic spindle poles and kinetochore protein CENP-H causing chromosome missegregation and chromosomal instability assessed by karyotype analysis. Loss of function studies confirmed the critical role of Chk2 inchromosomal instability and tumorigenicity. Conclusions: Identification of reliable biomarkers that can be used to predict patient outcomes ensures more effective clinical management, testing for the presence of Chk2 as a marker of chromosomal instability at diagnosis may be helpful not only for determining the prognosis but also for identifying patients who are likely to have a response to treatment.