Identification of transcriptional changes induced by FHIT in lung cancer cells NCI-H460

The FHIT gene, at 3p14.2, encompasses the common fragile site FRA3B and is frequently inactivated in primary tumors and cell lines of lung, head and neck, stomach, esophagus, cervix and breast cancer. In particular, loss of the FHIT protein is one of the most frequent alteration in lung tumors and pre-invasive lung lesions, suggesting a role for this gene in the early stages of lung carcinogenesis. Adenoviral-mediated restoration of FHIT expression in FHIT-negative cell lines results in cell cycle alteration, induction of apoptosis (through activation of the cytoplasmic apoptotic pathway) and regression of the tumorigenic phenotype. Taken together these observations support the hypothesis of a role for FHIT in human carcinogenesis, but little is known about its mechanism of action. Identification of transcriptional targets of FHIT is therefore critical to understand the pathways by which FHIT promotes growth arrest and apoptosis. RNA was extracted from NCI-H460 lung cancer cells (FHIT-negative) transduced with AdFHIT or AdLACZ (control), 48 hours post infection at a multiplicity of infection of 10. Three independent infections were harvested and RNA preparations were pooled, transcribed into cDNA, labeled, and hybridized following the manufacturer's protocol to the oligonucleotide microarray Human Gene Chip HG-U95Av2 (Affymetrix). Two independent hybridizations were performed (rep1 and rep2) and scan images aquired before (sc1) and after (sc2) anti-streptavidin antibody enhancement step. The whole procedure was repeated twice (starting from thawing of new aliquot of NCI-H460 cells) resulting in two independent experiments (exp1 and exp2).