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[The p16INK4A mRNA transcripts estimation in cervical smear with different degrees of cervical dysplasia]
V Janušicová1, I Kapustová, P Zúbor
1Gynekologicko-porodnicka klinika, Martin. veronika.janusicova@jfmed.uniba.sk
Objective:
To determine the presence of HPV infection and expression level of p16INK4A mRNA transcripts in cervical smears as adjunct biomarker in detection of cervical intraepithelial neoplasia or cancer.
Design:
Prospective pilot clinical study assessing clinical utility and validity of ddCt method for qPCR mRNA expression of p16ink4a in comparison to immunohistochemistry.
Setting:
Department of Molecular Biology, Department of Obstetrics and Gynecology, Jessenius Medical Faculty, Commenius University, Martin, Slovak Republic.
Methods:
Cervical smears (OC) from patients with different cervical lesions (L-SIL, H-SIL, SCA; n=45) and from healthy controls (n=45) were tested for the presence of HPV infection and p16INK4A mRNA transcripts using relative quantification (RQ). Results were compared to H&E and IHC histological findings from biopsies (conization, hysterectomy).
Results:
HPV 16 was the most frequent finding (53.3%) in the group of subjects with cervical dysplasia. The p16INK4A mRNA expression analysis revealed the slightly reduced expression in L-SIL group, 4-fold increased expression in H-SIL and 10-fold increase in women with SCA when compared to controls. The p16INK4A mRNA expression in OC was present in 30% of L-SIL, 75% of H-SIL and 85.7% of SCA samples, respectively. The test overall sensitivity was 81.48% (95% CI: 61.92-93.7) and specificity 60% (95% CI: 26.24- 87.84) with PPV of 84.62% and NPV of 54.55%. The likelihood ratio (LR) in case of test positivity was 2.04 and for negativity 0.31. The diagnostic accuracy of p16INK4A expression by RQ method in OC smears for prediction of p16 positivity in cervical dysplasia was 66.7% for the L-SIL lesions, 59.5% for H-SIL lesions, and 100% for SCA (r=0.9897, p<0.0913) when compared to IHC p16 positive findings in surgically treated samples.
Conclusion:
The relative quantification is able to determine the level of p16INK4A mRNA transcripts in cervical smear cells with active carcinogenesis nearly at the same level as IHC staining. The advance of biopsy sparing over IHC is qualifying this diagnostic approach for useful candidate in selective management of women with cervical dysplasia looking for cervix preservation or avoiding the unnecessary overtreatment.
