METH-2 silencing and promoter hypermethylation in NSCLC

Abstract

The antiangiogenic factor METH-2 (ADAMTS-8) was identified in a previous dual-channel cDNA microarray analysis to be at least two-fold under-represented in 85% (28 out of 33) of primary non-small-cell lung carcinomas (NSCLCs). This observation has been validated in an independent series of NSCLCs and adjacent normal tissues by comparative multiplex RT-PCR, and METH-2 mRNA expression was dramatically reduced in all 23 tumour samples analysed. Immunohistochemical analysis of the same sample set demonstrated that METH-2 was strongly expressed in 14 out of 19 normal epithelial sites examined but only one out of 20 NSCLCs. DNA methylation analysis of the proximal promoter region of this gene revealed abnormal hypermethylation in 67% of the adenocarcinomas and 50% of squamous cell carcinomas, indicating that epigenetic mechanisms are involved in silencing this gene in NSCLC. No homozygous deletions of METH-2 were found in lung cancer cell lines. Allelic imbalance in METH-2 was assessed by an intronic single nucleotide polymorphism (SNP) assay and observed in 44% of informative primary samples. In conclusion, the downregulation of METH-2 expression in primary NSCLC, often associated with promoter hypermethylation, is a frequent event, which may be related to the development of the disease.

Expression of a disintegrin and metalloprotease (ADAM and ADAMTS) enzymes in human non-small-cell lung carcinomas (NSCLC)

“…To the best of our knowledge, the present study is the first report of a negative association between a member of ADAMTS-1 and lung cancers. Dunn et al (2004) have previously reported such a negative association for ADAMTS-8 which was in that case associated with a promoter hypermethylation in cancers.…”

Emerging roles of ADAM and ADAMTS metalloproteinases in cancer

“…Interestingly, ADAMTS-8, a potent anti-angiogenic ADAMTS is downregulated in most primary NSCLC [115] due to abnormal promoter hypermethylation [115].…”

Expression of ADAMTS-8, a secreted protease with antiangiogenic properties, is downregulated in brain tumours

“…Immunohistochemistry (IHC) was performed as described previously (Dunn et al, 2004) using an ADAMTS-8 N-terminal antibody (1 : 500 dilution) (ADAMTS-8 AB-1, Oncogene Research Products; cat#PC508) and 5 mm sections cut from formalin fixed, paraffin-embedded blocks from the same tumour resections. Each run included tissue sections from brain tumours, normal cerebellum and a positive control (normal human stomach) incubated with primary antibody, and a negative control with no primary antibody for each section.…”

“…The modified, cleaned DNA was resuspended in 50 ml TE buffer, aliquotted and stored at À801C. A cMSP assay was used to assess the methylation status of the ADAMTS-8 promoter region as described previously (Dunn et al, 2004). Polymerase chain reaction products (control: 299 bp, methylation specific: 169 bp) were analysed on 4% agarose gels containing ethidium bromide.…”

“…There is evidence of modulation of expression of several of the ADAMTS genes in cancer, with significant downregulation (two-fold or lower) of ADAMTS-8 in non-small-cell lung cancer (NSCLC) (Heighway et al, 2002;Dunn et al, 2004) and hypermethylation of the promoter region being a possible mechanism of gene silencing (Dunn et al, 2004). In comparison to non-neoplastic mammary tissue ADAMTS-8 mRNA is also significantly downregulated in breast carcinomas (Porter et al,be of therapeutic use in prolonging survival.…”