Studies were conducted to investigate the biochemical reactions of TNM with nucleosides and DNA in vitro. No reaction products were detected when analyzed by HPLC. These data indicate that TNM either interacts with DNA indirectly (e.g. by binding to nucleoproteins) or that TNM must be metabolized to a reactive intermediate before binding to DNA and other macromolecules. Initial studies to evaluate the potential metabolism of TNM by lung microsomes revealed no metabolic products; however, these studies are continuing. Methods are being developed to identify and isolate differentially expressed genes in cells treated with TNM in vitro. The differential display method involves amplification of partial sequences from subsets of mRNA by reverse transcription and PCR followed by visualization of differences in their mRNA patterns on a sequencing gel. These studies will provide the basis for subsequent mechanistic studies performed in cultured lung cells exposed to TNM.
