Z Gastroenterol 2005; 43 - 2_24
DOI: 10.1055/s-2005-861637

Promotor-Hypermethylation Is Causing Functional Relevant Downregulation of Methylthioadenosine Phosphorylase (MTAP) Expression in Hepatocellular Carcinoma

C Hellerbrand 1, M Mühlbauer 1, M Schuierer 2, S Wallner 2, I Behrmann 3, F Bataille 2, J Schölmerich 1
  • 1Klinik und Poliklinik für Innere Medizin I der Universität Regensburg, Regensburg
  • 2Institut für Pathologie der Universität Regensburg, Regensburg
  • 3Laboratoire de Biologie et Physiologie Intégrée, Université du Luxembourg

Homozygous deletions of human chromosomal region 9p21 occur frequently in several kinds of cancer and are associated with the loss of the tumor suppressor genes p16 (INK4a) and p15 (INK4b). In the same chromosomal region the methyladenosine phosphorylase (MTAP) gene is localised and therefore, may also serve as a tumor suppressor gene.

The aim of this study was to analyse MTAP mutations and expression patterns in hepatocarcinoma (HCC).

Methods: MTAP mRNA and protein expression were analysed with real time PCR, western blotting and immunohistochemistry in a) primary human hepatocytes and 3 different hepatoma cell lines (HepG2, PLC, Hep3B), and b) HCC and surrounding non-tumorous tissue samples (n=7). Furthermore, exon 1 to 8 of the MTAP gene were analysed by sequencing and MTAP promotor methylation studies were performed. MTAP expression was re-induced in hepatoma cells by stable transfection with an MTAP expression construct (MTAP-pos), and invasion assays and cell-proliferation assays with or without interferon treatment were performed in comparison to mock transfected hepatoma cells.

Results: MTAP expression was markedly downregulated in hepatoma cells. This was not due to homozygous deletion of the genomic region but to promoter hypermethylation. Reduced MTAP expression was confirmed in vivo in HCC compared to normal tissue on both

mRNA and protein levels. Interestingly, the invasive potential of MTAP re-expressing cell clones was strongly reduced, while no effects on the ability to grow in soft agar or cell proliferation were observed compared to mock transfected cells. However, MTAP re-expressing cell clones revealed a significantly stronger inhibition of cell proliferation compared to mock transfected control.

Conclusion: Our results suggest an important role of MTAP inactivation in the HCC development and invasiveness. Furthermore, our findings are in line with a recent report suggesting an association between MTAP activity and interferon sensitivity which may have clinical significance for therapeutic strategies.

Schlüsselwörter

HCC - MTAP - Promotor Methylierung