Fingerprinting Genomewide Alterations During Malignant Transformation in Human Oral Keratinocytes

Objectives: FOXM1 transcription factor is known to be upregulated in many solid human cancers. Our recent data showed that FOXM1 gene was upregulated in head and neck squamous cell carcinoma (HNSCC) and that overexpression of FOXM1 in oral keratinocytes induces malignant transformation following nicotine exposure. Genomic instability is a hallmark of malignant transformation in many cancers including HNSCC. In order to understand global genomic alterations during malignant transformation, we have used the Affymetrix 10K single nucleotide polymorphism (SNP) mapping array to fingerprint genomewide alterations during malignant transformation induced by FOXM1 overexpression. Methods: A premalignant oral keratinocyte line SVpgC2a was retrovirally transduced to overexpress either FOXM1 or a control protein EGFP. These cells were then exposed to carcinogenic doses of nicotine to induce malignant transformation. EGFP expressing cells did not give rise to transformed cells. Eight FOXM1-transformed colonies were isolated, expanded into individual cell lines and genomic DNA extracted for SNP fingerprinting. Pattern of loss of heterozygosity (LOH) and copy number alterations (CNA) were mapped by comparing DNA from transformed cell lines with the untransformed parental cells. Results: Our data showed for the first time that overexpression of an oncogene FOXM1 alone in oral keratinocytes can induce LOH by uniparental disomy (UPD), an allelic imbalance-independent mechanism. Further carcinogenic stress such as nicotine exposure caused additional genomic instability affecting discrete chromosomal regions harbouring genes with functions in cellular adhesion/migration, matrix remodelling cell cycle/apoptosis and tumour suppression. Conclusions: This study presented the first evidence that unique patterns of genomic LOH could be induced by FOXM1 and that subsequent nicotine exposure resulting in additional genomic instability producing a non-random pattern of genomic LOH regions harbouring genes that may be implicated in malignant transformation. Further functional association study is required to validate the roles of these putative genes in relation to malignant transformation.