Background: KRT16 codifies for acidic-intermedium cytokeratin constitutively expressed in squamous epithelia. Its role in carcinogenesis may be related to its expression in the hyperproliferative phase on epidermal cultures, but yet need to be clarified. In a previous cDNA microarray analysis, the KRT16 gene was differentially expressed between healthy cervical tissue and a high grade squamous intraepithelial lesion (HSIL) with confirmed HPV-18 transforming infection. Gene expression profile differences between uterine cervix healthy tissue, low grade, and high grade squamous intraepithelial lesion (LSIL, HSIL) could contribute to identify potential biomarkers for cervical premalignant lesions progression.
Design: A chromogenic in situ hybridization (CISH) assay, (RNAscope), was performed to evaluate the KRT16 mRNA in situ expression in a tissue microarray (TMA) constructed from excision specimens (conization and hysterotomy) from 169 patients, with a mean age of 35.37 years (SD ±10.4), with representative areas from normal cervix tissue, LSIL and HSIL. The RNAscope 2.5HD Red KRT16 assay, and the RNAscope 2.5HD Red E6/E7 HPV-HR18 assay, designed to detect E6/E7 mRNA for eighteen different high-risk (HR) HPV genotypes, were performed. Stainings were visually scored from 0 to 4 by two pathologist based on the average number of dots per cell according to a semi-quantitative analysis; score values were compared using GraphPad Prism 7.0 software. A p< 0.05 was considered statistically significant.
Results: The KRT16 mRNA in situ expression in healthy cervical tissue, LSIL and HSIL was determined by CISH in a cervical TMA including, normal cervix: 84, LSIL: 77, and HSIL: 157, tissue cores. The mean KRT16 mRNA expression scores in squamous epithelium lining normal cervix (0.95, 95% CI: 0.70-1.20), LSIL (0.88, 95% CI:0.60-1.15), and HSIL (1.11, 95% CI:0.89-1.33), were not significantly different (p=0.573). Mean HPV E6/E7 mRNA expression score in HSIL was significantly higher than mean HPV E6/E7 mRNA expression in LSIL (p<0.0001). There was a positive-low correlation between KRT16 mRNA expression and E6/E7 HR-HPV mRNA expression (Spearman r =0.27).
Conclusions: Although previous cDNA microarray analysis identified differential expression of KRT16 between healthy cervical tissue and a HSIL with confirmed HPV-18 transforming infection, this was not confirmed when comparing its in situ mRNA expression between squamous epithelium lining normal cervix, LSIL and HSIL. Also, there is not a strong correlation with the expression of E6/E7 mRNA for eighteen different HR-HPV. The fact that KRT16 is constitutively expressed in basal cells may help to elucidate this phenomenon. The role of KRT16 in proliferative phase of basal cells and cervical carcinogenesis remains unclear, further studies are required to define it.
Citation Format: Ines Benedetti, Reinhard Rodríguez, Lia Barrios. KRT16 and E6-E7 HR-HPV mRNA in situ expression in normal cervical tissue, and low-grade and high-grade squamous intraepithelial lesions [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2153.