P14 ARF and p16 INK4A, two products of the same gene, are differently expressed in cervical intraepithelial neoplasia

Abstract

Objective. To study the expression patterns of two different tumor suppressor proteins p16 INK4A and p14 ARF in cervical lesions. Both proteins are encoded by the same INK4A/ARF gene on chromosome 9p21. The expression patterns of these two proteins, both playing a central role in the cell cycle, were analyzed in detail in CIN, carcinomas, and normal epithelium to test the hypothesis that p16 INK4A positive cells also demonstrate p14 ARF expression. Methods. Serial tissue sections of 9 CIN1 lesions, 10 CIN2 lesions, 12 CIN3 lesions, and 7 carcinomas were stained with monoclonal antibodies against p16 INK4A and p14 ARF. The short fragment polymerase chain reaction hybridization line probe assay was used to detect HPV. Results. Normal epithelium was negative for both proteins. Marked immunoreactivity (++) for p16 INK4A and p14 ARF was observed in 5/7 carcinomas, 10/12 CIN3, and 1/10 CIN2 lesions and 0/9 CIN1 lesions. Simultaneous expression (+ or ++) was found in 19/22 CIN2/3 and not in CIN1 lesions. The fraction of p16 INK4A-stained cells increased with CIN-grade. Overexpression of p14 ARF was observed in a subpopulation of p16 INK4A positive cells, and exclusively found in lesions infected with high-risk HPV. In two CIN3 lesions with early stromal invasion, p14 ARF positivity was mainly found in the invasive cells. In carcinomas, all cells showed p16 INK4A expression, whereas p14 ARF was limited to the peripheral cells of the invasive tumor nests and individual migrating tumor cells. Conclusions. Overexpression of p14 ARF is limited to a fraction of the p16 INK4A-expressing cells and therefore it is likely that p14 ARF− and p16 INK4A expression are not induced by the same mechanisms. Before expression of p14 ARF can be linked to invasion or invasive phenotype, larger series of (micro-) invasive squamous lesions need to be studied.