Methylation-mediated silencing of the LIM homeobox 6 (LHX6) gene promotes cell proliferation in human pancreatic cancer

概要

Epigenetic gene silencing by aberrant DNA methylation leads to loss of key cellular pathways in tumorigenesis. DNA methylation-mediated silenced genes in pancreatic cancer were searched for using the methyl-CpG targeted transcriptional activation (MeTA) method, and LHX6 (LIM homeobox 6), a transcription factor involved in embryogenesis as well as CNS and craniofacial development, was selected as a strong candidate gene. LHX6 was downregulated in most of the pancreatic cancer cell lines (83%, 10/12), mainly through promoter hypermethylation and histone deacetylation. Furthermore, LHX6 was methylated in primary pancreatic cancer specimens (57%, 16/28) in a tumor-specific manner. In order to assess the biological significance of LHX6 in pancreatic tumorigenesis, I first performed colony formation assay and found that LHX6 re-expression inhibited colony formation in LHX6 low-expressing pancreatic cancer cell lines, PK-1 and PK-9. Similarly, inducible expression of LHX6 inhibited cell proliferation and migration in PK-1 and PK-9 cells. In contrast, knockdown of LHX6 accelerated cell proliferation in LHX6 high-expressing pancreatic cancer cell lines, PCI-35 and MIA PaCa-2. Although LHX6 has been reported to be involved in the Wnt/ -catenin pathway in breast and lung cancers and my microarray analysis indicated that LHX6 induction in both PK-1 and PK-9 cells upregulated DKK4, which functions as a Wnt antagonist, LHX6 induction did not affect any CTNNB1 protein expressions. On the other hand, I also found that TFPI2, Tissue factor pathway inhibitor-2, was upregulated in accordance with LHX6 induction in both PK-1 and PK-9 cells. TFPI2 may function as a tumor suppressor in a LHX6-dependent manner, which differs from a well-known promoter hypermethylation. My present results suggest that epigenetic inactivation of LHX6 plays an important role in pancreatic tumorigenesis by promoting cell proliferation through the regulation of several downstream genes.