| Objectives1.To study the expression of p16/Ki-67 and p16/mcm2 immunocytochemical dual staining in cervical lesions and its association with high-riskHPV infectionand discuss its clinical value in cervical cancer screening.2.To compare the clinical performance of these two biomarkers for triaging the HPV positive women in cervical cancer screening.Materials and MethodsCytology specimens were collected from two prospective studies implemented by Cancer Hospital,Chinese Academy of Medical Sciences(CICAMS)from 2015 to 2016.HPV DNA was tested by Liferiver HPV Detection Kit and Roche cobas4800 HPV Detection Kit.All specimens were tested for ThinPrep cytology test.Women who were positive for any of the results received colposcopy and biopsy.The residual specimens were tested for immunocytochemical dual staining by p16/Ki-67 Detection Kit and cocktail antibodies of p16/mcm2,respectively.We analyzed the association between p16/Ki-67,p16/mcm2 and high-risk HPV infection using the baseline data.Histology diagnosis was treated as the gold standard andthe clinical performance of these two biomarkers for detection of cervical precancer was evaluated in entire population and in women with positive HPV results.Results1.The association between p16/Ki-67 expression and high-risk HPV infection:The expression of p16/Ki-67 in HPV positive population was higher than HPV negatives(x2=95.98,P<0.001),with an positivity of 37.1%(122/329)and 8.9%(43/483),respectively.HPV positive women were divided into HPV 16/18 type and other 12HR-HPV types according to different HPV infection status and the corresponding positive rates were 34.5%(26/51)and 51.0%(96/278).Compared to HPV negative women,odds ratio for women with HPV 16/18type was nearly 2-folder higher than women with other 12 HR-HPV types(OR=10.7,95%CI:5.6-20.1 VS OR=5.4,95%CI:3.6-8.1).2.The association between pl6/mcm2 expression and high-risk HPV infection:HPV positive women had a higher risk of p16/mcm2 expression compared to HPV negative population,with an odds ratio and 95%CI of 11.5(8.2-16.1).The positive rates of p16/mcm2 in HPV negative g:roup,other 12 HR-HPV group and HPV 16/18 group were 9.8%(53/540),53.4%(188/352)and 63.4%(59/93),respectively.Compared to HPV negative women,odds ratio for women with HPV 16/18type was 1.5-folder higher than those with other 12 HR-HPV types(OR=16.0,95%CI:9.6-26.5 VS OR=10.5,95%CI:7.4-15.0).3.The clinical performance of pl6/Ki-67 for detection of CIN2+ and CIN3+:The positivity of p16/Ki-67 in histology confirmed inflammation,CIN1,CIN2,CEN3 and squamous cervical cancer was 28.2%(51/181),44.1%(15/34),81.8(9/11),93.3(14/15)and 100%(2/2),respectively.Linear trend test showed that p16/Ki-67 positivity increased with histologic severity(x2=38.57,ptrend<0.001).The sensitivity of p16/Ki-67 for detection of CIN2+ and CIN3+ were 89.3%and 94.1%,respectively,and the specificity were 69.3%and 66.8%,respectively.The sensitivity of Liquid-based cytology in CIN2+ and CIN3+ were 60.7%and 64.7%,respectively,and the specificity were 49.3%and 49.1%,respectively.4.The clinical performance of p16/mcm2 for detection of CIN2+ and CIN3+:The positive rate of p16/mcm2 increased with histologic severity(x2=15.4,Ptrend<0.001),from 53.3%(106/199)in inflammation,57.1%(24/42)in CIN1,77.8%(14/18)in CIN2 to 92.0%(23/25)in CIN3.The sensitivity of p16/mcm2 to detect CIN2+ and CIN3+ in entire population were 86.1%and 92.0%,respectively,and the specificity were 46.1%and 44.4%,respectively;in women with NILM cytology results,the sensitivity for CIN2+ and CIN3+ were 75.0%and 100%,respectively,and the specificity were 46.4%and 46.6%,respectively;in women with ASC and LSIL results,the sensitivity for CIN2+ and CIN3+ were 85.7%and 87.5%,respectively,and the specificity were 45.5%and 44.1%,respectively,with a referral rate of 53.8%.5.The clinical performance of p16/Ki-67 and p16/mcm2 for detection of CIN2+ and CIN3+ in women with positive HPV results:In histology confirmed normal,CIN1,CIN2,CIN3 and SCC,p16/Ki-67 positivity was 34.4%(41/119),46.2%(12/26),87.5(7/8),92,9(13/14)and 100%(2/2),respectively;pl6/mcm2 positivity was 56.3%(67/119),65.4%(17/26),75.0(6/8),92.9(13/14)and 100%(2/2),respectively.Positive rates of both biomarkers increased with histology severity(both Ptrend<0.001).In all HPV positive women,the sensitivity of p16/Ki-67 for CIN2+ and CIN3+ were 91.7%and 93.8%,respectively,and the specificity were 63.5%and 60.8%,respectively,with a referral rate of 36.2%;the sensitivity of p16/mcm2 for CIN2+and CIN3+ were 87.5%and 93.8%,respectively,and the specificity were 42.1%and 41.2%,respectively,with a referral rate of 58.4%.In women with HPV positive,cytology negative results,the sensitivity of p16/Ki-67 for CIN2+ and CIN3+ were 90.9%and 100%,respectively,and the specificity were 71.7%and 69.4%,respectively;the sensitivity of p16/mcm2 for CIN2+ and CIN3+ were 81.8%and 100%,respectively,and the specificity were 46.2%and 46.0%,respectively.Conclusions1.The expression of p16/Ki-67 and p16/mcm2 associated strongly with HR-HPV infection,especially with HPV 16/18 type.Due to the relationship of HR-HPV persistent infection,p16/Ki-67 and pl6/mcm2 testing could be considered as potential molecular biomarkers for cervical cancer screening.2.p16/Ki-67 had a higher sensitivity and similar specificity compared with cytology test,and could efficiently identify high-grade cervical lesions and guide the classification of CIN;the interpretation of p16/Ki-67 is independent of morphological criterion and could be performed by general cytotechnologists after a short period of training,with good reproductivity.It might be considered as a replacement of Liquid-based cytology in cervical cancer screening.p16/mcm2 had a high sensitivity and limited specificity.It could identify cervical precancerous lesions from women with normal cytology and be used as a triage method for ASC and LSIL population,with an efficient decrease in colposcopy referral rate.3.p16/Ki-67 provided a similar sensitivity and higher specificity compared to p16/mcm2 in women with positive HPV results,and could cut off the colposcopy referral rate in a greater degree.Thus,p16/Ki-67 is a promising triage biomarker for HPV positive women,very suitable for Chinese rural areas where medical resource is limited and the implementation of LBC program isn’t available... |
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