Epigenetic inactivation of ADAMTS9 via promoter methylation in multiple myeloma

A disintegrin-like and metalloprotease with thrombospondin type I motifs (ADAMTS) are a family of 19 secreted mammalian metalloproteases. ADAMTS9 was reported to be a novel tumor suppressor gene and is downregulated in various types of human cancer due to hypermethylation of promoter CpG islands. In the present study, the silencing mechanism of the ADAMTS9 gene was analyzed in the multiple myeloma (MM) cell lines, KM3 and RPMI-8226. Control and MM samples were obtained by conventional bone marrow (BM) biopsy of normal and MM adult BM, respectively. RT-PCR revealed a high expression of the ADAMTS9 gene in normal samples and RPMI-8226 cells while marked gene silencing of ADAMTS9 was observed in MM patients and KM3 cells. Promoter methylation of ADAMTS9 was detected in the KM3 cell line and 66% (37/56) MM patients by methylation-specific PCR. In addition, the DNA demethylating agent, 5-aza-2'-deoxycytidine and trichostatin A restored ADAMTS9 expression by suppressing promoter methylation in KM3 cells. Ectopic expression of ADAMTS9 in ADAMTS9-silenced MM cells was found to significantly suppress cell colony formation and proliferation. In the present study, DNA methylation was found to play a key role in ADAMTS9 gene silencing and the biological behavior of myeloma cells. The results demonstrate that ADAMTS9 silencing by methylation may be a novel tumor marker for MM and the applicability of demethylating agents in the treatment of MM.