Predictive value of p16INK4a, Ki-67 and ProExC immuno-qualitative features in LSIL progression into HSIL

Abstract

The current nested case-control study was conducted to explore the prognostic value of cyclin-dependent kinase inhibitor 2A (p16^INK4a), marker of proliferation Ki-67 (Ki-67) and immunohistochemical cocktail containing antibodies directed against topoisomerase IIα (TOP2A) and minichromosome maintenance 2 (MCM2) proteins (ProExC) immuno-qualitative features to predict low-grade squamous intraepithelial lesion (LSIL) progression. A total of 92 LSIL patients were followed-up for 2 years, where those with high-grade squamous intraepithelial lesion (HSIL) or persistent LSIL were designated as the case group and those who spontaneously regressed were designated as the control group. The infection status of human papillomavirus (HPV) was evaluated using flow-through hybridization and gene chip, whilst the expression of p16^INK4a, Ki-67 and ProExC were tested in LSIL patient biopsies by immunohistochemistry. All data were collected at the beginning of the follow-up and patient outcomes were diagnosed by histopathological examination. To analyze the risk factors for LSIL progression, sensitivity, specificity, positive-negative predictive value (PPV-NPV), positive-negative likelihood ratio (PLR-NLR), Youden's index (YI) and multinomial logistic regression analysis was performed. The expression rates of p16^INK4a, Ki-67, and ProExC were found to be higher in the progression group compared with those in the persistence and regression groups. Only p16^INK4a expression significantly associated with high-risk HPV infection. With respect to predicting HSIL, p16^INK4a staining was the most sensitive but Ki-67 staining was found to be the most specific. YI was the highest (42.1%) for p16^INK4a expression in the present study, followed by ProExC (39.5%) and Ki-67 (28.3%). However, the expression of ProExC was found to be an independent risk factor for LSIL progression into HSIL. In conclusion, whilst immunohistochemical staining for p16^INK4a, Ki-67, and ProExC can be used to predict HSIL progression, only ProExC expression can be applied an independent risk factor for LSIL progression.

Keywords: DNA topoisomerase IIα; and minichromosome maintenance complex component 2 antibody cocktail; cyclin-dependent kinase inhibitor 2A; marker of proliferation Ki-67; prognostic factor; squamous intraepithelial lesions of the cervix.

Figure 1.

Schematic representation of the study design, from the recruitment of subjects to final analysis. NC, normal cervix; LSIL, low-grade cervical intraepithelial neoplasia; ASC-US, atypical squamous cells of undetermined significance; HSIL, high-grade cervical intraepithelial neoplasia; SCC, squamous cell carcinoma.

Figure 2.

Positive and negative controls. (A) Positive controls for p16^INK4a, (B) Ki67, (C) ProExC in prostate cancer tissues. (D) Negative control for p16^INK4a, Ki67 and ProExC in cervical cancer tissues. Magnification, x200.

Figure 3.

Expression of p16INK4a in low-grade cervical intraepithelial neoplasia tissues. (A) Representative image of a specimen exhibiting negative staining. (B) Representative image of a specimen exhibiting focal staining patterns, which were regarded as negative. (C) Representative image of a specimen exhibiting continuous, diffuse staining, which was regarded as positive. Magnification, x200. p16^INK4a, cyclin-dependent kinase inhibitor 2A.

Figure 4.

Expression of Ki67 in low-grade cervical intraepithelial neoplasia tissues. (A) Representative image of a specimen exhibiting staining representing <50%, interpreted as negative. (B) Representative image of a specimen exhibiting staining representing >50%, considered as positive. Magnification, x200. Ki-67, marker of proliferation Ki-67.

Figure 5.

Expression of ProExC in in low-grade cervical intraepithelial neoplasia tissues. (A) Cell staining occupying <33% of squamous epithelium was regarded as negative. (B) Cell staining occupying more thana third of squamous epithelium was regarded as positive. Magnification, x200. ProExC, DNA topoisomerase IIα and minichromosome maintenance complex component 2 antibody cocktail.