p16/Ki-67 dual staining in cervico-vaginal cytology: correlation with histology, Human Papillomavirus detection and genotyping in women undergoing colposcopy.
ABSTRACT To evaluate the CINtec PLUS assay (mtm laboratories), a new immunocytochemical method for the simultaneous detection of p16(INK4a) and Ki-67, in liquid-based cervico-vaginal cytology, investigating the association of the dual staining with HPV infection and genotyping as well as cytological and histological abnormalities.
140 women with a cervico-vaginal sample obtained immediately before the colposcopy were enrolled. This cytological sample was used for HPV testing with the Linear Array HPV Genotyping Test, the dual staining with the CINtec PLUS kit and the morphology assessment.
Cytology results were 38 NILM, 16 ASC-US, 32L-SIL, 54H-SIL or worse. 113 patients also had a colposcopy-guided biopsy, classified as 14 negative, 35 CIN1, 24 CIN2, 37 CIN3, 3 invasive SCC. A strong association between p16/Ki-67 and HR-HPV infection was found (COR=6.86, 95% CI: 1.84-31.14). Importantly, the association between p16/Ki-67 positivity and HPV16 and/or 18 infection was 2-fold stronger compared to that with the infection by other HR-HPV types (COR=9.92, 95% CI: 2.39-47.77 vs COR=4.20, 95% CI: 0.99-20.87). In addition, p16/Ki-67 positivity rate significantly increased with the severity of the cytological and histological abnormalities (p<0.05 in both cases). p16/Ki-67 positivity resulted strongly associated with a CIN2+ diagnosis (COR=10.86 95% CI: 4.16-29.12).
This preliminary study evidenced that p16/Ki-67 immunostaining might have a relevant clinical role, since the dual staining was significantly associated with HR-HPV infection, particularly with HPV 16 and 18, and the increasing grade of the cervical lesions, the positivity for this biomarker being strongly related to the presence of a CIN2+ lesion.
- SourceAvailable from: PubMed Central[show abstract] [hide abstract]
ABSTRACT: Background: Atypical squamous cell cannot exclude high-grade squamous intraepithelial lesion (ASC-H) and low-grade intraepithelial lesion cannot exclude high-grade squamous intraepithelial lesion (LSIL-H) are ambiguous diagnostic entities for the prediction of high-grade cervical lesion. Objective and reproducible tests for predicting high-grade cervical lesions are needed to reduce unnecessary colposcopic referrals or follow-ups. Objective: We aimed to identify an adequate set of adjunctive markers to predict cervical intraepithelial neoplasia grade 2+ (CIN2+) in residual liquid-based cytology specimens (LBCS). Methods: We conducted p16 (INK4a)/Ki-67 and L1 capsid protein immunostaining and human papillomavirus (HPV) DNA typing on 56 LBCS diagnosed with ASC-H or LSIL-H, all of which were subjected to histologic confirmation or follow-up cytologic examination. Results: Positivity for p16 (INK4a)/Ki-67 was associated with a histology of CIN2+ (P=0.047) and CIN3+ (P=0.002). Negativity for L1 capsid protein was associated with CIN2+ confirmed at follow-up (P=0.02).Positivity for high-risk HPV (HR-HPV) was associated with CIN2+ confirmed at follow-up (P=0.036) and a histology of CIN2+ (P=0.037). The sensitivity, specificity, positive predictive value, and negative predictive value for predicting follow-up CIN2+ were 76.2%, 51.4%, 48.5%, and 78.3%, respectively, for p16 (INK4a)/Ki-67 immunostaining; 95.2%, 34.3%, 46.5%, and 92.3%, respectively, for L1 capsid protein; and 66.7%, 67.7%, 54.5%, and 77.8%, respectively, for HR-HPV. The classification and regression tree analysis showed that the combined results of p16 (INK4a)/Ki-67 andL1 capsid protein immunostaining and the HR-HPV test, conducted sequentially, correctly classified 81.8% of samples (27/33)in the prediction of a histology of CIN2 + in ASC-H or LSIL-H. For determination of the histology of cervical intraepithelial neoplasia grade 3+ (CIN3+)in ASC-H or LSIL-H, we found that the combined results of p16 (INK4a)/Ki-67 and L1 capsid protein immunostaining correctly classified 78.8% (26/33) of samples. Conclusions: p16(INK4a)/Ki-67 and L1 capsid protein immunostaining and HR-HPV testing of residual LBCS diagnosed with ASC-H or LSIL-H are useful objective biomarkers for predicting CIN2+. Immunostaining for p16(INK4a)/Ki-67 and L1 capsid protein are sufficient to predict CIN3+.International journal of medical sciences 01/2013; 10(12):1602-7. · 2.07 Impact Factor
- [show abstract] [hide abstract]
ABSTRACT: Pap test, and especially HPV DNA test, identify a large group of women who do not have any clinically relevant lesions, i.e., CIN2+ (Cervical Intraepithelial Neoplasia grade 2 or worse), but who are at greater risk of getting lesions in the future. The follow up of these women needs new biomarkers with prognostic value. The objective of this study is to evaluate the prognostic value of E6/E7 mRNA over-expression assay (PreTect HPV-Proofer, Norchip) for 5 HR-HPV types (16, 18, 31, 33, and 45) for progression to CIN2+ after a negative colposcopy. This prospective study, conducted at four Italian centres, enrolled 673 women with either a negative colposcopy or a negative or CIN1 histology. The clinical end-point was histological confirmation of CIN2+. Women were classified at baseline according to mRNA results and managed according to local colposcopy protocols. At least one conclusive follow-up test was obtained for 347 women (25 months average lapse since recruitment, range 5-74). Only seven CIN2+ were detected during follow up, three among the 82 women positive for mRNA at baseline, two among the 250 negative (Fisher exact test, p = 0.02), and two among the 12 with an invalid test. Absolute CIN2+ risk was 6.7/1,000 person/years in the whole cohort. The absolute CIN2+ risk was 18.4/1,000 person/years and 3.6/1,000 person/years in mRNA-positive and mRNA-negative women, respectively. In conclusion, E6/E7 mRNA over-expression appears to be a good candidate as a prognostic biomarker to manage HR-HPV DNA-positive women with negative colposcopy or histology, particularly in order to decrease follow-up intensity in those who are negative.PLoS ONE 01/2013; 8(2):e57600. · 3.73 Impact Factor