Methods: A high efficient retroviral gene transfer method was used to deliver FOXM1 and/or EGFP transgenes in human keratinocytes. Real-time quantitative PCR was used to quantify gene expression in a series of normal, oral premalignant and HNSCC derived keratinocytes. A high-resolution Affymetrix single nucleotide polymorphism (SNP) mapping technique was used to measure genomic instability in the form of loss of heterozygosity (LOH) and copy number abnormalities (CNA) in human keratinocytes.
Results: FOXM1 is upregulated in oral pre-malignant and HNSCC derived primary keratinocytes. FOXM1B was found to be the main isoform driving cell cycle progression, expressed mainly at the G2 phase of human keratinocytes. We provide the first evidence that FOXM1B upregulation was sufficient to induce genomic instability in human keratinocytes. FOXM1B-induced genomic instability was enhanced and accumulated with increasing passage number, and keratinocytes with upregulated FOXM1B showed enhanced sensitivity to subsequent genotoxic insult causing an increased accumulation of genomic instability.
Conclusions: We hypothesise that aberrant upregulation of FOXM1B is important in oral carcinogenesis whereby it serves as a ‘first hit' where cells acquire genomic instability. This predisposes cells to a ‘second hit' whereby subsequent exposure to genotoxic insults may expedite the selection of transformed cells to proliferate and accumulate further genetic aberrations required for cancer initiation.