In an effort to identify all the genes that are affected by DNA methylation in lung cancer, we treated several lung cancer cell lines with the DNA methyltransferase inhibitor 5’-aza-2-deoxycitidine (5’-Aza). We found that 5’-Aza induced the expression of between 184 (H157) –1289 (H2347) genes >4 fold. There was significant heterogeneity in the gene expression profiles of the lung cancer cell lines, both before and after treatment with 5’-aza. However, we identified 291 genes that were upregulated >4 fold in at least two of the cell lines. As 5’-aza treatment is known to affect gene transcription in general and a number of the genes we found are known to be imprinted or are located on the X chromosome, we sought to further isolate genes that are specific to cancer. To this end, we treated a cell line which has been immortalized without the use of dominant oncogenes with 5’-aza. We found that 419 genes were induced >4 fold. Of these, 26 were also upregulated >4 fold in the cancer cell lines. To delineate cancer specific gene induction, we subtracted genes that were induced in both the immortalized cell line and at least two of the cancer cell lines. This resulted in a subset of 265 genes that were induced by DNA demethylation in only lung cancer cells. We then compared this set of genes to a broader panel of normal human bronchial and lung cancer cell lines and identified 63 genes that were expressed in the normal cells, but were not expressed by the majority of cancer cell lines. We confirmed gene expression and induction by real time PCR and Western Blot.We also identified 30 genes that were upregulated in the cancer cell lines and were not expressed in untreated normal cells, but were induced by 5’-Aza in the normal bronchial cells. This subset represents a novel group of genes that may act as dominant oncogenes after induction by genome-wide hypomethylation. This integrated approach has identified a large number of new candidate genes that may be involved in the pathogenesis of lung cancer and will require functional testing.

[Proc Amer Assoc Cancer Res, Volume 45, 2004]