Role of p16/MTS1, cyclin D1 and RB in primary oral cancer and oral cancer cell lines

One of the most important components of G1 checkpoint is the retinoblastoma protein (pRB(110)). The activity of pRB is regulated by its phosphorylatio n, which is mediated by genes such as cyclin D1 and p16/MTS1. All th ree ge nes have been shown to be commonly altered in human malignancies. We have s creened a panel of 26 oral squamous cell carcinomas (OSGC), nine premaligna nt and three normal oral tissue samples as well as eight established OSCC c ell lines for mutations in the p16/MTS1 gene. The expression of p16/MTS1, c yclin D1 and pRB(110) was also studied in the same panel. We have found p16 /MTS1 gene alterations in 5/26 (19%) primary tumours and 6/8 (75%) cell lin es. Two primary tumours and five OSCC cell lines had p16/MTS1 point mutatio ns and another three primary and one OSCC cell line contained partial gene deletions. Six of seven p16/MTS1 point mutations resulted in termination co dons and the remaining mutation caused a frameshift. Western blot analysis showed absence of p16/MTS1 expression in 18/26 (69%) OSCC, 7/9 (78%) premal ignant lesions and 8/8 cell lines. One cell line, H314, contained a framesh ift mutation possibly resulting in a truncated p16/MTS1 protein. pRB was de tected in 14/25 (56%) of OSCC but only 11/14 (78%) of these contained all o r some hypophosphorylated (active) pRB. In premalignant samples, 6/8 (75%) displayed pRB, and all three normal samples and eight cell lines analysed c ontained RE protein, p16/MTS1 protein was undetectable in 10/11 (91%) OSCCs with positive pRB. Overexpression of cyclin D1 was observed in 9/22 (41%) OSCC, 3/9 (33%) premalignant and 8/8 (100%) of OSCC cell lines. Our data su ggest p16/MTS1 mutations and loss of expression to be very common in oral c ancer cell lines and less frequent in primary OSCC tumours. A different pat tern of p16/MTS1 mutations was observed in OSCC compared to other cancers w ith all the detected p16/MTS1 mutations resulting in premature termination codons or a frameshift. The RE protein was expressed in about half (44%) of OSCCs and its expression inversely correlated with p16/MTS1 expression. In conclusion, we show that abnormalities of the RE pathway are a common mech anism of oral carcinogenesis.