Preferentially different mechanisms of inactivation of 9p21 gene cluster in liver fluke-related cholangiocarcinoma

Abstract

Cholangiocarcinoma in northeast Thailand is associated with liver fluke infection. Mechanisms of inactivation of the p15INK4b, p16INK4a, and p14ARFhave been reported in many human cancers but have not hitherto been studied in liver fluke-related cholangiocarcinoma, particularly genetic and epigenetic effects on protein expression. We investigated loss of heterozygosity and microsatellite instability and performed fine mapping of the chromosomal region 9p21-pter in 94 microdissected cholangiocarcinoma samples using polymerase chain reaction based-microsatellite markers. Methylation and protein expression of p14ARF, p15INK4b, and p16INK4awas determined using methylation-specific polymerase chain reaction and immunohistochemistry, respectively. Genetic and epigenetic alterations, including loss of protein expression, were correlated with clinicopathological data. Fine mapping at 9p21-pter showed a distinctive region between D9S286 and D9S1752 of common loss. Methylation frequency was 40.2% for p14ARF, 48.9% for p15INK4b, and 28.3% for p16INK4a. Loss of protein expression of p14ARF, p15INK4b, and p16INK4awas 30.9%, 58%, and 81.5%, respectively. Both p14ARFmethylation and allelic loss at 9p21 were associated with loss of p14ARFexpression. Poor prognosis was associated with loss of p16INK4aexpression. In conclusion, mechanisms of inactivation of p14ARF, p15INK4b, and p16INK4ain liver fluke-related cholangiocarcinoma are preferentially different, by which epigenetic event being the main mechanism of p14ARF, whereas p16INK4aand p15INK4binactivation occurs through genetic and both genetic and epigenetic events, respectively. © 2009 Elsevier Inc. All rights reserved.