The Diagnostic Accuracy of Electrical Impedance Spectroscopy-Assisted Colposcopy, HPV mRNA Test, and P16/Ki67 Immunostaining as CIN2+ Predictors in Greek Population.

OBJECTIVE: To evaluate the diagnostic accuracy of Electrical Impedance Spectroscopy (EIS)-assisted colposcopy in detecting CIN2+ Greek women towards standalone colposcopy, HPV mRNA testing, and p16/Ki67 immunostaining. METHODS: We conducted a cross-sectional observational study at the Cervical Pathology Clinic of the 2nd Obstetrics-Gynecology University Department of Hippokration Hospital Thessaloniki involving 316 patients from January 2022 to August 2023. All participants provided liquid-based cervical samples for cytology, HPV mRNA testing, and p16/Ki67 immunostaining. MAIN OUTCOME MEASURES: Subsequently, participants underwent both standalone colposcopy and EIS/ZedScan-assisted colposcopy, followed by cervical punch biopsies. RESULTS: The incorporation of EIS significantly enhanced the sensitivity of colposcopy, increasing it from 54.17% to 100%, equivalent to that of HPV mRNA testing and p16/Ki67 immunostaining, while achieving a high specificity (95.45%). The specificities observed with EIS/ZedScan-assisted and standalone colposcopy were notably superior to those of HPV-related biomarkers (HPV mRNA test and p16/Ki67 immunostaining). When compared to standalone colposcopy, HPV mRNA testing, and p16/Ki67 immunostaining, EIS/ZedScan-assisted colposcopy demonstrated the most favorable combination of Positive and Negative Predictive Values, at 90.57% and 100%, respectively. The inclusion of EIS/ZedScan in colposcopy led to the detection of 44 additional cases of true CIN2+ (100% of the total CIN2+ confirmed histologically) that were missed by standalone colposcopy. This discovery suggests a 45.83% increase in the detection of CIN2+ cases. CONCLUSIONS: The integration of EIS with colposcopy has demonstrated effectiveness in detecting cervical lesions, resulting in a significant detection increase of CIN2+ cases while offering optimal levels of sensitivity, specificity, and predictive values for CIN2+ detection.

Findings and Challenges in Replacing Traditional Uterine Cervical Cancer Diagnosis with Molecular Tools in Private Gynecological Practice in Mexico.

We have been encouraging practicing gynecologists to adopt molecular diagnostics tests, PCR, and cancer biomarkers, as alternatives enabled by these platforms, to traditional Papanicolaou and colposcopy tests, respectively. An aliquot of liquid-based cytology was used for the molecular test [high-risk HPV types, (HR HPV)], another for the PAP test, and one more for p16/Ki67 dual-stain cytology. A total of 4499 laboratory samples were evaluated, and we found that 25.1% of low-grade samples and 47.9% of high-grade samples after PAP testing had a negative HR HPV-PCR result. In those cases, reported as Pap-negative, 22.1% had a positive HR HPV-PCR result. Dual staining with p16/Ki67 biomarkers in samples was positive for HR HPV, and 31.7% were also positive for these markers. Out of the PCR results that were positive for any of these HR HPV subtypes, n 68.3%, we did not find evidence for the presence of cancerous cells, highlighting the importance of performing dual staining with p16/Ki67 after PCR to avoid unnecessary colposcopies. The encountered challenges are a deep-rooted social reluctance in Mexico to abandon traditional Pap smears and the opinion of many specialists. Therefore, we still believe that colposcopy continues to be a preferred procedure over the dual-staining protocol.

P16/Ki67 Dual Staining in Glandular Cell Abnormalities of the Uterine Cervix.

Very limited information exists about the role of p16/Ki67 dual staining on glandular cells in detecting glandular precancerous lesions and cervical adenocarcinoma. In this study, we investigated the diagnostic accuracy of p16/Ki67 dual staining for the detection of glandular and squamous lesions on the uterine cervix and for cancer of the upper reproductive tract. We performed a retrospective analysis of prospectively collected data on 96 patients with glandular cell abnormalities. We analyzed the diagnostic accuracy of p16/Ki67 dual staining for atypical glandular cells, not otherwise specified (AGC-NOS); atypical glandular cells, favor neoplastic (AGC-FN); adenocarcinoma in situ (AIS); and A-CA (cervical adenocarcinoma). A separate analysis for the detection of squamous precancerous lesions and squamous-cell carcinoma (CIN3+) and for cancer of the upper reproductive tract (EC/OC) was performed. Among patients who had normal histology or a low-grade lesion on final analysis, only 8.5% had positive dual staining. On the other hand, 85.7% of patients with AIS+ on final histology had positive dual staining. The respective specificities of p16/Ki67 dual staining on AGC-NOS for the detection of AIS+ (adenocarcinoma in situ or cervical adenocarcinoma), CIN3+ and EC/OC were 91.5%, 88.7% and 86.4%. High specificity values of p16/Ki67 dual staining on cervical smears labelled as AGC-NOS for the detection of CIN3+ and AIS+ suggest that this method might be a useful addition in cervical cancer screening.

Extended Genotyping to Stratify the Risk of CIN2+ in Women with Persistent HPV Infection, Negative Cytology and Type 3 Transformation Zone.

Persistent human papillomavirus (HPV) infection is recognized as a major risk factor for cervical cancer. Women with persistent HPV and negative cytology are at greater risk of CIN2+ than women with negative infection. The diagnosis becomes more complicated when the woman has a type 3 transformation zone at colposcopy. The aim of this study was to determine the prevalence of CIN2+ in women with persistent HPV, negative cytology and TZ3; how to stratify the risk of CIN2+; and what the best diagnostic strategy is, given TZ3. METHODS: In a multicenter retrospective cohort study, we enrolled women with negative cytology and TZ3 among the 213 women referred for colposcopy for persistent HPV. The average age of the women was 53 years; in particular, 83% were postmenopausal women. In the presence of a TZ3, the entire transformation zone cannot be explored, making colposcopy and targeted biopsy useless and inadequate, with great risks of underdiagnosis or missed diagnosis. Women with TZ3 underwent diagnostic LEEP to ensure correct diagnoses. RESULTS: The study highlighted 19% (16/84) of CIN2+ lesions, a higher frequency of non-HPV 16/18 genotypes (76.2%), and 50% of CIN2+ lesions being due to non-HPV 16/18 genotypes. Furthermore, more than half of the women (80.9%) had normal histopathological results in the LEEP sample. CONCLUSION: Women with viral persistence, negative cytology, and TZ3 have a 19% risk of CIN2+; genotyping helps stratify risk, but extensive genotyping is necessary instead of partial genotyping (16/18), referring to a population of women over 50 years old in which the prevalence of genotypes 16,18 decreases and the prevalence of other genotypes increases; diagnostic LEEP is excessive (only 16 cases of CIN2+ out of 48 cases treated), even though 83% of women had viral clearance after LEEP; p16/Ki67 double staining could be a potential risk marker, which would only highlight women at risk of CIN2+ to undergo LEEP. To individualize the diagnostic workup and treatment and minimize the risk of under diagnosis and overtreatment, future studies should explore the use of extended genotyping and new biomarkers for individual risk stratification.

Comparison of HPV-positive triage strategies combining extended genotyping with cytology or p16/ki67 dual staining in the Italian NTCC2 study.

BACKGROUND: Each high-risk HPV genotype has different oncogenic potential, and the risk of CIN3+ varies according to genotype. We evaluated the performance of different strategies of HPV-positivity triage combining cytology, p16/ki67 dual staining (DS), and extended genotyping. METHODS: Samples from 3180 consecutive women from the NTCC2 study (NCT01837693) positive for HPV DNA at primary screening, were retrospectively analyzed by the BD Onclarity HPV Assay, which allows extended genotyping. Genotypes were divided into three groups based on the risk of CIN3+. HPV DNA-positive women were followed up for 24 months or to clearance. FINDINGS: Combining the three groups of genotypes with cytology or DS results we identify a group of women who need immediate colposcopy (PPV for CIN3+ from 7.8 to 20.1%), a group that can be referred to 1-year HPV retesting (PPV in those HPV-positive at retesting from 2.2 to 3.8), and a group with a very low 24-month CIN3+ risk, i.e. 0.4%, composed by women cytology or DS negative and positive for HPV 56/59/66 or 35/39/68 or negative with the Onclarity test, who can be referred to 3-year retesting. INTERPRETATION: Among the baseline HPV DNA positive/cytology or DS negative women, the extended genotyping allows to stratify for risk of CIN3+, and to identify a group of women with a risk of CIN3+ so low in the next 24 months that they could be referred to a new screening round after 3 years. FUNDING: Italian Ministry of Health (grant number RF-2009-1536040). Hologic-Genprobe, Roche Diagnostics, and Becton & Dickinson provided financial and non-financial support.

A Prospective Study on the Progression, Recurrence, and Regression of Cervical Lesions: Assessing Various Screening Approaches.

(1) Background: The prediction of cervical lesion evolution is a challenge for clinicians. This prospective study aimed to determine and compare the predictive accuracy of cytology, HPV genotyping, and p16/Ki67 dual staining alone or in combination with personal risk factors in the prediction of progression, regression, or persistence of cervical lesions in human papillomavirus (HPV)-infected patients; (2) Methods: This prospective study included HPV-positive patients with or without cervical lesions who underwent follow-up in a private clinic. We calculated the predictive performance of individual tests (cervical cytology, HPV genotyping, CINtecPlus results, and clinical risk factors) or their combination in the prediction of cervical lesion progression, regression, and persistence; (3) Results: The highest predictive performance for the progression of cervical lesions was achieved by a model comprising a Pap smear suggestive of high-grade squamous intraepithelial lesion (HSIL), the presence of 16/18 HPV strains, a positive p16/Ki67 dual staining result along with the presence of at least three clinical risk factors, which had a sensitivity (Se) of 74.42%, a specificity of 97.92%, an area under the receiver operating curve (AUC) of 0.961, and an accuracy of 90.65%. The prediction of cervical lesion regression or persistence was modest when using individual or combined tests; (4) Conclusions: Multiple testing or new biomarkers should be used to improve HPV-positive patient surveillance, especially for cervical lesion regression or persistence prediction.

Correlation between P16/Ki67 in cervical cytology and diagnosis of cervical intraepithelial neoplasia 2-3 in Thai women infected with high-risk types of human papillomavirus.

OBJECTIVES: The addition of p16/Ki-67 dual immunostaining to human papilloma virus (HPV) screening tests has been shown to increase the detection rate of high-grade cervical intraepithelial neoplasia. Thus, the aim of this study was to evaluate the accuracy of p16/Ki67 dual staining in the detection of cervical intraepithelial neoplasia 2 (CIN2+) in women with high-risk HPV infection. MATERIALS AND METHODS: A cross-sectional study was conducted between August 2017 and August 2019 at the Chulabhorn Hospital in Bangkok, Thailand. Women aged 20-70 years who underwent co-testing and tested positive for high-risk (HR) HPV (N = 215) were invited to participate in the study. P16/Ki67 testing was performed on residual cytological materials. Colposcopic biopsies were performed on all patients, and the results were correlated with positive or negative p16/Ki-67 test results. RESULTS: The sensitivity and specificity of p16/Ki-67 dual staining in the detection of CIN2+ in the women with HR HPV infection were 74.4 % and 63.4 %, respectively. Compared with liquid-based cytology (LBC), p16/Ki67 cytology had similar sensitivity (p = 1.000) and specificity (p = 0.561) to LBC for detecting CIN2+. CONCLUSION: In this study, p16/Ki67 dual staining in HPV triage demonstrated a test performance similar to that of LBC.

Should we use risk selection tests for HPV 16 and/or 18 positive cases: Comparison of p16/Ki67 and cytology.

Major screening abnormalities in precolposcopic stage are tests results that imply direct referral to colposcopy (and/or expedited treatment) without performing additional high-grade squamous intraepithelial lesions or worse (HSIL+) risk selection testing. Currently, both clinically validated HSIL+ risk selection tests, reflex cytology and reflex p16/Ki67 dual staining (DS), are being compared for use in primary human papillomavirus (HPV)-based screening to avoid possible overtreatment, but there is still no sufficient data available for their performance. Among 30 066 liquid-based cervical cancer screening tests results, a group of 332 women was selected with available high-risk types of HPV tests results with 16/18 limited genotyping, liquid-based cytology, DS, and histology results from standardized colposcopy with biopsy. In HPV 16/18+ cases, three triage approaches were retrospectively analyzed. Predictive values for detection of HSIL+ were calculated and number of colposcopies required in each strategy. Both triage models with DS used (reflex cytology followed by DS, and reflex DS alone in all cases) had significantly higher positive predictive value for HSIL+ than strategy with reflex cytology alone (44.2%/45.7% vs. 28.3%; p < 0.0001). In models with DS, less colposcopies were required (95/92 vs. 152) and less colposcopies were needed per HSIL+ detection (2.26/2.19 vs. 3.54). Only one HSIL+ case was missed in both triage models with DS incorporation. p16/Ki67 dual-stain may be an effective, alone or combined with cytology, triage test to detect HSIL+ in patients with major screening abnormalities in primary HPV-based cervical cancer screening. Performing cytology as the first triage test improves the strategy by enabling referrals to expedited treatment in selected cases.

Enhancing Cervical Cancer Screening: Review of p16/Ki-67 Dual Staining as a Promising Triage Strategy.

Cervical cancer, primarily caused by high-risk human papillomavirus (HR-HPV) types 16 and 18, is a major global health concern. Persistent HR-HPV infection can progress from reversible precancerous lesions to invasive cervical cancer, which is driven by the oncogenic activity of human papillomavirus (HPV) genes, particularly E6 and E7. Traditional screening methods, including cytology and HPV testing, have limited sensitivity and specificity. This review explores the application of p16/Ki-67 dual-staining cytology for cervical cancer screening. This advanced immunocytochemical method allows for simultaneously detecting p16 and Ki-67 proteins within cervical epithelial cells, offering a more specific approach for triaging HPV-positive women. Dual staining and traditional methods are compared, demonstrating their high sensitivity and negative predictive value but low specificity. The increased sensitivity of dual staining results in higher detection rates of CIN2+ lesions, which is crucial for preventing cervical cancer progression. However, its low specificity may lead to increased false-positive results and unnecessary biopsies. The implications of integrating dual staining into contemporary screening strategies, particularly considering the evolving landscape of HPV vaccination and changes in HPV genotype prevalence, are also discussed. New guidelines and further research are necessary to elucidate the long-term effects of integrating dual staining into screening protocols.

Expression of p16 Tumor Suppressor Protein in Malignant Ovarian Germ Cell Tumors; Immunohistochemical Study.

BACKGROUND: Malignant ovarian germ cell tumors represent small percentage of malignant ovarian neoplasms but they affect significantly young age group. AIM OF THE STUDY: To investigate the immunohistochemical expression of p16 tumor suppressor protein in malignant ovarian germ cell tumors. MATERIALS AND METHODS: Twenty-two malignant ovarian germ cell tumors (five dysgerminoma, eight immature teratoma, and nine yolk sac tumors), twenty mature cystic teratoma tumors and twenty normal ovarian tissue were immunohistochemically stained with p16 monoclonal antibody. Ki67 immunohistochemical staining was done for malignant ovarian germ cell tumors to assess proliferation. RESULTS: We found that p16 tumor suppressor protein is overexpressed in all malignant ovarian germ cell tumors in both nuclear and cytoplasmic locations compared to control and to mature cystic teratoma (p-value <0.001). Cytoplasmic p16 expression was significantly correlated to Ki67 proliferation index in malignant ovarian germ cell tumors (p-value = 0.033, r = 0.445). CONCLUSION: Overexpression of p16 in malignant ovarian germ cell tumors denotes that dysfunction of the cyclin dependent kinase pathway is involved in tumorigenesis of malignant ovarian germ cell tumors.

Cumulative detection of anal high-grade squamous intraepithelial lesions over two-year follow-up in men who have sex with men living with HIV in France.

We assessed cumulative detection and determinants of anal high-grade squamous intraepithelial lesions (HSIL) in men who have sex with men living with HIV who underwent three visits over two years, with cytology and high-resolution anoscopy (HRA), within the ANRS-EP57-APACHES study. Cumulative HSIL detection was 33% (134/410), of which 48% were detected at baseline. HSIL detection varied considerably by center (13-51%). Strongest HSIL determinants were baseline HPV16 (adjusted odds ratio [aOR] 8.2; 95% confidence interval [95%CI] 3.6-18.9), and p16/Ki67 (aOR 4.6; 95%CI 2.3-9.1). Repeat annual cytology and HRA improved HSIL detection but did not fully compensate between-center heterogeneity.

p16/Ki67 dual stain triage versus cytology in primary human papillomavirus-based cervical cancer screening with limited genotyping.

The introduction of primary human papillomavirus (HPV) cervical cancer screening requires the implementation of an appropriate triage strategy that will be effective in detecting high-grade cervical disease without losing diagnostic specificity. From the 30.066 screening tests results, a total of 1086 with available high-risk human papillomavirus (HRHPV) with limited genotyping, cytology, and p16/Ki67 dual-stain were selected. Two triage strategies for primary HPV screening were analyzed retrospectively based on the study group. Performance characteristics for p16/Ki67 and cytology triage in the detection of cervical intraepithelial neoplasia grade 2 or worse (CIN2+) and grade 3 or worse (CIN3+) were calculated, detected in colposcopic biopsy. In HPV16/18-positive cases, primary HPV with p16/Ki67 triage was significantly more specific than cytology (53.1%/16.8% for CIN2+; p < 0.0001; 45.9%/17.0% for CIN3+; p < 0.0001), with yielded sensitivity (95.7%/84.8% for CIN2+; p = 0.0955; 100.0%/87.5% for CIN3+; p = 0.0832). In other HRHPV-positive cases (N16/N18), p16/Ki67 triage was also significantly higher specific (51.3%/15.3% for CIN2+; p < 0.0001; 44.5%/16.5% for CIN3+; p < 0.0001), with sensitivity (92.3%/74.4% for CIN2+; p = 0.0522; 90.9%/81.8% for CIN3+; p = 0.5637). Diagnostic predictive values were significantly higher for p16/Ki67 triage with the highest PPV in HPV16/18-positive cases for CIN2+ (45.4%; 95% confidence interval [CI]: 35.2-55.8; p < 0.0001) and very high NPV in all HPV-positive cases regardless of detected genotype (96.3%-100.0%). The risk (1-NPV) for CIN3+ in HRHPV16/18-positive/p16/Ki67-negative women was 0.0%. Superior diagnostic performance compared to cytology for detecting cervical cancer precursors indicates that p16/Ki67 dual-immunostain may be a highly effective tool of triage in primary HPV screening with limited HPV 16/18 genotyping in secondary cervical cancer prevention.

Triage Strategies for Non-16/Non-18 HPV-Positive Women in Primary HPV-Based Cervical Cancer Screening: p16/Ki67 Dual Stain vs. Cytology.

BACKGROUND: In the context of primary HPV cervical cancer screening, the identification of minor screening abnormalities necessitates triage tests to optimize management and mitigate overtreatment. Currently, reflex cytology and reflex p16/Ki67 dual-stain (DS) are under scrutiny for their applicability in primary HPV-based screening. However, there remains a dearth of comprehensive data for comparing their performance. METHODS: Among 30,066 results from liquid-based cervical cancer screening tests, a cohort of 332 cases was meticulously selected based on available high-risk human papillomavirus (HPV) test results, limited genotyping for HPV 16 and 18, liquid-based cytology, DS, and histology outcomes from standardized colposcopy with biopsy. For cases positive for 12 other high-risk HPV genotypes, three retrospective triage approaches were analyzed. We computed the positive predictive value (PPV) for the detection of high-grade squamous intraepithelial lesions or worse (HSIL+). RESULTS: Both triage models employing DS (reflex cytology followed by DS and reflex DS alone in all cases) exhibited significantly higher PPV for HSIL+ compared to the strategy with reflex cytology alone (35.9%/33.3% vs. 18.8%; p < 0.0001). Additionally, these DS-based models showed higher negative predictive values (NPV) (100%/96.2% vs. 69.2%; p = 0.0024/0.0079). In the DS-inclusive models, fewer colposcopies were necessitated (103/102 vs. 154), and fewer cases of HSIL+ were overlooked (0/3 vs. 8). CONCLUSIONS: Our findings suggest that p16/Ki67 dual-stain, either as a standalone or combined triage test, holds promise for the effective detection of HSIL+ in patients with minor screening abnormalities in primary HPV-based cervical cancer screening.

Evaluation of the clinical performance of p16/Ki-67 dual-staining cytology for cervical lesion detection in premenopausal and postmenopausal Chinese women.

BACKGROUND: Studies on the clinical performance of p16/Ki-67 dual-staining in detecting cervical lesions by menopausal status were limited. METHODS: 4364 eligible women were enrolled with valid p16/Ki-67, HR-HPV, and LBC test results, including 542 cancer and 217 CIN2/3 cases. The positivity rates of p16 and Ki-67 single staining and p16/ Ki-67 dual-staining were analyzed by different pathological grades and age groups. The sensitivity (SEN), specificity (SPE), positive predictive value (PPV), and negative predictive value (NPV) of each test in different subgroups were calculated and compared. RESULTS: P16/Ki-67 dual-staining positivity increased with histopathological severity in premenopausal and postmenopausal women (P < 0.05), while no increasing trends of individual expression of p16 single staining and Ki-67 single staining were observed in postmenopausal women. P16/Ki-67 showed higher SPE (88.09% vs. 81.91%, P < 0.001) and PPV (33.8% vs. 13.18%, P < 0.001) in detecting CIN2/3, and higher SEN (89.97% vs. 82.61%, P = 0.012) and SPE (83.22% vs. 79.89%, P = 0.011) in detecting cancer in premenopausal women than postmenopausal women. For triaging the HR-HPV+ population to identify CIN2/3, p16/Ki-67 performed comparably to LBC in the premenopausal women, and showed higher PPV (51.14% vs. 23.08%, P < 0.001) in premenopausal than postmenopausal women. For triaging ASC-US/LSIL population, p16/Ki-67 demonstrated higher SPE and lower colposcopy referral rate than HR-HPV in both premenopausal and postmenopausal women. CONCLUSIONS: Expressions of p16/Ki-67 dual-staining between premenopausal and postmenopausal women are varied. P16/Ki-67 performs better in detecting cervical lesions in premenopausal women. For triaging, p16/Ki-67 is suitable for HR-HPV+ women, especially premenopausal women, to identify CIN2/3 and women with ASC-US/LSIL.

Successful Preventive Treatment of Oncogenic Transforming HPV Infections in Low-Grade Cytology (ASC-US/LSIL) Patients with an Adsorptive and Antioxidant Vaginal Gel.

OBJECTIVE: This study aimed to investigate the preventive effect of a vaginal gel on p16/Ki-67-positive abnormal cytological cervical findings (ASC-US, LSIL) and hr-HPV in women. METHODS: The study included 134 women with p16/Ki-67-positive ASC-US or LSIL. Participants were selected from a randomized controlled trial that focused on women with histological diagnoses of p16-positive CIN1 lesions or CIN2. In the treatment group (TG), 57 patients applied the vaginal gel daily for three months, while 77 patients in the "watchful wait" control group (CG) received no treatment. The study's endpoints were cytological development, p16/Ki-67 and hr-HPV clearances. RESULTS: At three months, cytopathological results improved in 74% (42/57) of patients in the TG, compared with 18% (14/77) in the CG. Progression occurred in 7% (4/57) of TG patients compared with 18% (14/77) of CG patients. The p16/Ki-67 status changed statistically significantly in favor of the TG (p < 0.001), with 83% (47/57) becoming negative, compared with 18% (14/77) in the CG. The prevalence of hr-HPV decreased significantly in the TG by 51%, and by 9% in the CG (p < 0.001). CONCLUSIONS: Topical application of the gel resulted in statistically significant clearance of hr-HPV and p16/Ki-67 concomitant with amelioration of cytological findings, thus providing effective prevention and protection against oncogenic development. TRIAL REGISTRATION: ISRCTN11009040, on 10 December 2019.

Performance of p16/Ki67 Immunostaining for Triage of Elderly Women with Atypical Squamous Cells of Undetermined Significance.

BACKGROUND: The p16/Ki67 technique has been poorly studied in postmenopausal women with ASC-US cytology. The objective of this study was to compare the accuracy of p16/Ki67 staining, HPV testing and HPV 16 genotyping for the identification of CIN2 + lesions in postmenopausal women with ASC-US cytology. METHOD: A total of 324 postmenopausal women with positive ASC-US were included. The women underwent HPV test, colposcopy, and biopsy. The slides were discolored and then stained with the CINtec Plus Kit for p16/Ki67. The HPV test results were classified as HPV16 +, hrHPV+ (other hrHPV genotypes), or HPV negative. RESULTS: The p16/Ki67 sensitivity for CIN2+ was 94.5%, the specificity 86.6%, PPV of 59% and NPV of 95.9%. The HPV test showed a sensitivity of 96.4% for CIN2+, a specificity of 62.8%, a PPV of 35% and a NPV of 98.8%. In postmenopausal women, the prevalence of genotype 16 decreases in favor of the other high-risk genotypes. CONCLUSION: Given the low sensitivity of cytology and the low percentage of HPV16-positive cancers among elderly women, triage via cytology and genotyping is not the best strategy; double staining cytology shows high profiles of sensibility and specificity for CIN2+ in ASCUS postmenopausal women.

Clinical utility of p16/Ki67 dual-stain cytology for detection of cervical intraepithelial neoplasia grade two or worse in women with a transformation zone type 3: A cross-sectional study.

OBJECTIVE: To evaluate the clinical utility of p16/Ki67 dual-stain (DS) compared with cytology for detecting cervical intraepithelial lesion grade two or worse (CIN2+) in women with a transformation zone type 3 (TZ3). DESIGN: Cross-sectional study. SETTING: Colposcopy clinics in Central Denmark Region. POPULATION: Women aged 45 years or older referred for colposcopy because of an abnormal screening test. METHODS: All women had a cervical sample collected for cytology and DS testing and underwent large-loop excision of the transformation zone (LLETZ). MAIN OUTCOME MEASURE: Sensitivity, specificity and negative (NPV) and positive (PPV) predictive values of DS for CIN2+ detection were compared to those of cytology. RESULTS: Of 166 women eligible, 93 (56.0%) were included in the final analysis. Median age was 68 years (interquartile range [IQR] 63.4-70.5 years). Most women were postmenopausal (95.7%) and referred based on a positive human papillomavirus screening test (86.0%). Fifty-two women (55.9%) were DS-positive, 29 (55.8%) of whom had CIN2+ detected. Twenty-seven (29.0%) women had atypical squamous cells of undetermined significance or worse (ASC-US+), and CIN2+ was detected in 21 women (77.8%). DS had a higher sensitivity (96.7% versus 70.0% p = 0.021) and NPV (97.6% versus 86.4%, p = 0.018) compared with cytology for CIN2+ detection. In contrast, the specificity (63.5% versus 90.5% p < 0.001) and PPV (55.8% versus 77.8%, p = 0.001) were lower for DS compared with cytology. CONCLUSIONS: Dual stain may be a valuable risk marker to guide clinical management of women with a TZ3. The superior NPV of DS suggests that a diagnostic excision may safely be avoided in DS-negative women.

p16/Ki-67 dual stain, PAP cytology and HR-HPV test results prior to and 6 months after a LLETZ procedure: a prospective observational cohort study.

OBJECTIVES: To investigate the effect of a LLETZ procedure on p16/Ki-67 dual stain, PAP cytology and HR-HPV test results on cervical cytology samples obtained prior to and 6 months after the procedure. Secondary aims are to assess dependency between test results at the time of follow-up and explore dual stain positivity rates according to known risk factors for persistence/recurrence of cervical intra-epithelial neoplasia (CIN). STUDY DESIGN: Prospective observational cohort study conducted in the Department of Gynaecology at the University Hospitals of Leuven, Belgium. All patients referred for a LLETZ procedure were invited to participate. A cervical cytology sample was obtained just prior to and 6 months after the procedure. Every sample was used for PAP staining (cytology), p16/Ki-67 dual staining (dual stain test, DST) and HR-HPV genotyping. Test results were compared between both timepoints using the McNemar test. Dependency was assessed cross-sectionally at the time of follow-up using a chi-squared test. RESULTS: From the 110 participants originally included, 83 attended follow-up (75.5%). Mean duration of follow-up was 187.91 days (SD 21.47) and mean age was 41.4 years (SD 11.08). DST positivity rates were 70.9 and 30.1% prior to and 6 months after the procedure (p < 0.001). HR-HPV testing (positive or negative) and abnormal PAP cytology (evaluated at an ASCUS or worse threshold) showed a similar significant reduction in positivity rates (84.5 vs 42.2% and 72.7 vs 28.9%, respectively, p < 0.001). Results of all three assays showed high dependency at the time of follow-up (DST and PAP, PAP and HR-HPV test, DST and HR-HPV test-p values < 0.001). The highest proportion of positive DST results was seen in patients carrying HPV16 (84.6%), followed by any HR-HPV type (60%), those treated for CIN2 + (27.3%) and those with positive margins on the cone specimen (26.7%). CONCLUSION: A LLETZ procedure results in a significant decrease in abnormal DST, PAP cytology and HR-HPV test results in this diverse cohort of patients. The highest proportion of abnormal DST results was seen in patients carrying HR-HPV at the time of follow-up, especially HPV 16.

P16/Ki-67 Dual Staining in Positive Human Papillomavirus DNA Testing for Predictive Diagnosis of Abnormal Cervical Lesions in Northeastern Thai Women.

OBJECTIVE: Cervical cancer screening can effectively reduce new cervical cancer cases, including in Thailand. The abnormal results are subsequently referred for colposcopy. To avoid unnecessary colposcopy, an efficient triage is still needed for validation. This study aimed to investigate the overall positivity of cytology-based screening, HPV detection, and p16/Ki-67 dual staining and evaluate different triage strategies for predictive diagnosis of abnormal cervical lesions in northeastern Thailand. METHODS: Cervical cells were collected from 191 women who came for cervical screening in the gynecological outpatient department during March 2019-February 2020. Pap smear samples were classified into 6 groups including 17 atypical glandular cells (AGC), 21 atypical squamous cells of undetermined significance (ASC-US), 7 atypical squamous cells - cannot exclude HSIL (ASC-H), 26 low-grade squamous intraepithelial lesions (LSILs), 19 high-grade SILs (HSILs) and 101 no squamous intraepithelial lesion (noSIL). Polymerase chain reaction (PCR) was performed for HPV DNA detection. HPV genotyping was determined by reverse line blot hybridization. P16/Ki-67 dual staining was performed by using CINtec PLUS Cytology kit. Biopsies from abnormal screening were collected for surgical pathology classification. RESULTS: High-risk HPV (HR-HPV) infection was 2.97%, 29.41%, 38.10%, 57.14%, 46.15% and 84.21% in noSIL, AGC, ASC-US, ASC-H, LSIL and HSIL cytology respectively. P16/ Ki-67 in noSIL, AGC, ASC-US, ASC-H, LSIL and HSIL was 0.99%, 5.88%, 9.52%, 42.86%, 26.92% and 63.16%, respectively (P-value < 0.001). Among p16/Ki-67 positive cases, 96.15% (25/26) were infected with HPV and 84.62% (22/26) were HR-HPV. The overall positivity of each and co-testing between cytology or HPV DNA testing or p16/Ki-67 dual staining was evaluated. In each cervical lesion, primary HPV DNA testing showed the highest sensitivity, but low specificity. The combined all HPV/HR-HPV with p16/Ki-67 detection increased the specificity of abnormal cervical lesions. CONCLUSION: P16/Ki-67 dual stain cytology in HPV-positive women performs well for diagnosis of abnormal cervical lesions and should be considered for management of HPV-positive women to avoid unnecessary colposcopy referrals.

New Insights in the Diagnosis of Rare Adenocarcinoma Variants of the Cervix-Case Report and Review of Literature.

We report the case of a 29-year-old patient with low-grade squamous intraepithelial lesion (L-SIL), negative human papilloma virus (HPV), positive p16/Ki-67 dual-staining and colposcopy suggestive for severe dysplastic lesion. The patient underwent a loop electrosurgical excision procedure (LEEP), the pathology report revealing mesonephric hyperplasia and adenocarcinoma. The patient also opted for non-standard fertility-sparing treatment. The trachelectomy pathology report described a zone of hyperplasia at the limit of resection towards the uterine isthmus. Two supplementary interpretations of the slides and immunohistochemistry (IHC) were performed. The results supported the diagnosis of mesonephric adenocarcinoma, although with difficulty in differentiating it from mesonephric hyperplasia. Given the discordant pathology results that were inconclusive in establishing a precise diagnosis of the lesion and the state of the limits of resection, the patient was referred to a specialist abroad. Furthermore, the additional interpretation of the slides and IHC were performed, the results suggesting a clear cell carcinoma. The positive p16/Ki-67 dual-staining prior to LEEP, the non-specific IHC and the difficulties in establishing a diagnosis made the case interesting. Given the limitations of cytology and the fact that these variants are independent of HPV infection, dual staining p16/Ki-67 could potentially become useful in the diagnosis of rare adenocarcinoma variants of the cervix, however further documentation is required.