Adjunctive use of p16 immunohistochemistry for optimizing management of CIN lesions in a high-risk human papillomavirus-positive population.

INTRODUCTION: Immunostaining with p16INK4a (p16), a tumor-suppressor surrogate protein biomarker for high-risk human papillomavirus (hrHPV) oncogenic activity, may complement standard hematoxylin and eosin (H&E) histology review, and provide more objective criteria to support the cervical intraepithelial neoplasia (CIN) diagnosis. With this study we assessed the impact of p16 immunohistochemistry on CIN grading in an hrHPV-based screening setting. MATERIAL AND METHODS: In this post-hoc analysis, 326 histology follow-up samples from a group of hrHPV-positive women were stained with p16 immunohistochemistry. All H&E samples were centrally revised. The pathologists reported their level of confidence in classifying the CIN lesion. RESULTS: Combining H&E and p16 staining resulted in a change of diagnosis in 27.3% (n = 89) of cases compared with the revised H&E samples, with a decrease of 34.5% (n = 18) in CIN1 and 22.7% (n = 15) in CIN2 classifications, and an increase of 18.3% (n = 19) in no CIN and 20.7% (n = 19) in CIN3 diagnoses. The level of confidence in CIN grading by the pathologist increased with adjunctive use of p16 immunohistochemistry to standard H&E. CONCLUSIONS: This study shows that adjunctive use of p16 immunohistochemistry to H&E morphology reduces the number of CIN1 and CIN2 classifications with a proportional increase in no CIN and CIN3 diagnoses, compared with standard H&E-based CIN diagnosis alone. The pathologists felt more confident in classifying the material with H&E and p16 immunohistochemistry than by using H&E alone, particularly during assessment of small biopsies. Adjunctive use of p16 immunohistochemistry to standard H&E assessment of CIN would be valuable for the diagnostic accuracy, thereby optimizing CIN management and possibly decreasing overtreatment.

Cervical intraepithelial neoplasia and the risk of spontaneous preterm birth: A Dutch population-based cohort study with 45,259 pregnancy outcomes.

BACKGROUND: Excisional procedures of cervical intraepithelial neoplasia (CIN) may increase the risk of preterm birth. It is unknown whether this increased risk is due to the excision procedure itself, to the underlying CIN, or to secondary risk factors that are associated with both preterm birth and CIN. The aim of this study is to assess the risk of spontaneous preterm birth in women with treated and untreated CIN and examine possible associations by making a distinction between the excised volume of cervical tissue and having cervical disease. METHODS AND FINDINGS: This Dutch population-based observational cohort study identified women aged 29 to 41 years with CIN between 2005 and 2015 from the Dutch pathology registry (PALGA) and frequency matched them with a control group without any cervical abnormality based on age at and year of pathology outcome (i.e., CIN or normal cytology) and urbanization (<100,000 inhabitants or ≥100,000 inhabitants). All their 45,259 subsequent singleton pregnancies with a gestational age ≥16 weeks between 2010 and 2017 were identified from the Dutch perinatal database (Perined). Nineteen potential confounders for preterm birth were identified. Adjusted odds ratios (ORs) were calculated for preterm birth comparing the 3 different groups of women: (1) women without CIN diagnosis; (2) women with untreated CIN; and (3) women with treated CIN prior to each childbirth. In total, 29,907, 5,940, and 9,412 pregnancies were included in the control, untreated CIN, and treated CIN group, respectively. The control group showed a 4.8% (1,002/20,969) proportion of spontaneous preterm birth, which increased to 6.9% (271/3,940) in the untreated CIN group, 9.5% (600/6,315) in the treated CIN group, and 15.6% (50/321) in the group with multiple treatments. Women with untreated CIN had a 1.38 times greater odds of preterm birth compared to women without CIN (95% confidence interval (CI) 1.19 to 1.60; P < 0.001). For women with treated CIN, these odds 2.07 times increased compared to the control group (95% CI 1.85 to 2.33; P < 0.001). Treated women had a 1.51 times increased odds of preterm birth compared to women with untreated CIN (95% CI 1.29 to 1.76; P < 0.001). Independent from cervical disease, a volume excised from the cervix of 0.5 to 0.9 cc increased the odds of preterm birth 2.20 times (37/379 versus 1,002/20,969; 95% CI 1.52 to 3.20; P < 0.001). These odds further increased 3.13 times and 5.93 times for women with an excised volume of 4 to 8.9 cc (90/724 versus 1,002/20,969; 95% CI 2.44 to 4.01; P < 0.001) and ≥9 cc (30/139 versus 1,002/20,969; 95% CI 3.86 to 9.13; P < 0.001), respectively. Limitations of the study include the retrospective nature, lack of sufficient information to calculate odds of preterm birth <24 weeks, and that the excised volume could only be calculated for a select group of women. CONCLUSIONS: In this study, we observed a strong correlation between preterm birth and a volume of ≥0.5 cc excised cervical tissue, regardless of the severity of CIN. Caution should be taken when performing excisional treatment in women of reproductive age as well as prudence in case of multiple biopsies. Fertile women with a history of performing multiple biopsies or excisional treatment for CIN may benefit from close surveillance during pregnancy.

Hyperthermic Intraperitoneal Chemotherapy in Ovarian Cancer.

BACKGROUND: Treatment of newly diagnosed advanced-stage ovarian cancer typically involves cytoreductive surgery and systemic chemotherapy. We conducted a trial to investigate whether the addition of hyperthermic intraperitoneal chemotherapy (HIPEC) to interval cytoreductive surgery would improve outcomes among patients who were receiving neoadjuvant chemotherapy for stage III epithelial ovarian cancer. METHODS: In a multicenter, open-label, phase 3 trial, we randomly assigned 245 patients who had at least stable disease after three cycles of carboplatin (area under the curve of 5 to 6 mg per milliliter per minute) and paclitaxel (175 mg per square meter of body-surface area) to undergo interval cytoreductive surgery either with or without administration of HIPEC with cisplatin (100 mg per square meter). Randomization was performed at the time of surgery in cases in which surgery that would result in no visible disease (complete cytoreduction) or surgery after which one or more residual tumors measuring 10 mm or less in diameter remain (optimal cytoreduction) was deemed to be feasible. Three additional cycles of carboplatin and paclitaxel were administered postoperatively. The primary end point was recurrence-free survival. Overall survival and the side-effect profile were key secondary end points. RESULTS: In the intention-to-treat analysis, events of disease recurrence or death occurred in 110 of the 123 patients (89%) who underwent cytoreductive surgery without HIPEC (surgery group) and in 99 of the 122 patients (81%) who underwent cytoreductive surgery with HIPEC (surgery-plus-HIPEC group) (hazard ratio for disease recurrence or death, 0.66; 95% confidence interval [CI], 0.50 to 0.87; P=0.003). The median recurrence-free survival was 10.7 months in the surgery group and 14.2 months in the surgery-plus-HIPEC group. At a median follow-up of 4.7 years, 76 patients (62%) in the surgery group and 61 patients (50%) in the surgery-plus-HIPEC group had died (hazard ratio, 0.67; 95% CI, 0.48 to 0.94; P=0.02). The median overall survival was 33.9 months in the surgery group and 45.7 months in the surgery-plus-HIPEC group. The percentage of patients who had adverse events of grade 3 or 4 was similar in the two groups (25% in the surgery group and 27% in the surgery-plus-HIPEC group, P=0.76). CONCLUSIONS: Among patients with stage III epithelial ovarian cancer, the addition of HIPEC to interval cytoreductive surgery resulted in longer recurrence-free survival and overall survival than surgery alone and did not result in higher rates of side effects. (Funded by the Dutch Cancer Society; ClinicalTrials.gov number, NCT00426257 ; EudraCT number, 2006-003466-34 .).

Early prediction of post-molar gestational trophoblastic neoplasia and resistance to methotrexate, based on a single serum human chorionic gonadotropin measurement.

BACKGROUND: Clinicians are unable to provide individualized counseling regarding risk of progression for patients with a complete hydatidiform mole (CHM). We developed nomograms enabling early prediction of post-molar gestational trophoblastic neoplasia (GTN) and resistance to methotrexate (MTX) based on a single serum human chorion gonadotropin (hCG) measurement. METHODS: We generated two nomograms with logistic regression: to predict post-molar GTN, and MTX resistance. For patients with high probability to progress to post-molar GTN or MTX resistance, we determined hCG cut-offs at 97.5% specificity to select patients for additional- or adjustments in current treatment. RESULTS: The nomograms had a good to excellent ability to distinguish either between patients with uneventful hCG regression versus progression to post molar GTN, or between patients cured by MTX versus patients in whom resistance would occur. At 97.5% specificity, we identified 66% (95%CI 56-75) of the 149 patients who would progress to post-molar GTN, four weeks after initial curettage. For patients treated with MTX, we identified 55% (95%CI 23-83) of the 43 patients who would become resistant, preceding their third course at 97.5% specificity. CONCLUSION: The nomograms and cut-off levels can be used to assist in counseling for patients diagnosed with CHM.

Immune cell composition in the endometrium of patients with a complete molar pregnancy: Effects on outcome.

OBJECTIVE: In 15% of patients with complete hydatidiform mole (CHM), disease progresses to post-molar gestational trophoblastic neoplasia (GTN) after curettage. Tumor infiltrating lymphocytes (TILs) are essential in overcoming disease in many tumors. Infiltrating lymphocyte composition and density may influence trophoblast regression and development of post-molar GTN. We analyzed immune cell composition and density in curettaged endometrium of patients with CHM which spontaneously regressed, and of patients with CHM which progressed to post-molar GTN. METHODS: Sixteen patients with CHM and spontaneous regression, and 16 patients with CHM which progressed to post-molar GTN were selected. Immune cell composition and density of natural killer (NK) cells, natural killer T (NKT)-like cells, Cytotoxic T cells, T-Regulatory and T-Helper cells, were determined by multiplex immunohistochemistry (mIHC). RESULTS: Curettaged endometrium of patients with CHM and spontaneous regression contained a slightly higher number of immune cells compared to patients with CHM which progressed to post-molar GTN. NKT-like cell density was significantly higher in patients with spontaneous regression compared to patients with CHM which progressed to post-molar GTN (483 ± 296 vs.295 ± 143 (mean ± SD), p = 0.03) respectively. NKT-like cell density in the spontaneous regression group was split in 'high' and 'low' (i.e. above and below the median number of NKT-like cells). In patients with high NKT-like cell density, hCG normalized earlier than in patients with low NKT-like cell density (9.5 weeks, (range 3.7-14) vs. 12.9 weeks, (range 8.6-17.9), p = 0.05). CONCLUSION: A high number of NKT-like cells in the endometrium of CHMs may contribute to spontaneous regression of molar trophoblast cells.

The relative risk of noncervical high-risk human papillomavirus-related (pre)malignancies after recurrent cervical intraepithelial neoplasia grade 3: A population-based study.

Women with cervical intraepithelial neoplasia grade 3 (CIN3) have a long-lasting increased risk for noncervical high-risk human papillomavirus (hrHPV)-related (pre)malignancies. The aim of our study was to estimate this risk in women with recurrent CIN3 compared to women without a history of CIN3 and women with a single episode of CIN3. Women with a CIN3 diagnosis between 1990 and 2010 were obtained from the Dutch Pathology Registry (PALGA) and matched with a control group of women without CIN3. Analysis has been conducted in a subset of women with recurrent CIN3, defined as reoccurrence minimally 2 years post-treatment. Cases of noncervical hrHPV-related (pre)malignancies of the anus, vulva, vagina and oropharynx were identified until 2015 and incidence rate ratios (IRRs) were estimated. Then, 1,797 women with recurrent CIN3 were included with a median age of 34 years (range 18-76) and 31,594 person-years of follow-up. Women with recurrent CIN3 had an increased risk of developing noncervical hrHPV-related (pre)malignancies compared to women without CIN3 with an IRR of 25.96 (95%CI 6.32-106.58). The IRR was 2.48 (95% CI 1.87-3.30) compared to women with a single episode of CIN3. Studies on posttreatment follow-up and prophylactic hrHPV vaccination are warranted.

Mutational analysis of cervical cytology improves diagnosis of endometrial cancer: A prospective multicentre cohort study.

Endometrial carcinoma (EC) is traditionally diagnosed by a histopathological assessment of an endometrial biopsy, leaving up to 30% of patients undiagnosed due to technical failure or an inadequate amount of tissue. The aim of the current study is to assess whether mutational analysis of cervical cytology or pipelle endometrial biopsies improves the diagnostic accuracy of traditional histopathological diagnosis of EC. This prospective multicentre cohort study included patients surgically treated for EC or a benign gynaecological condition (control group). A Pap brush sample, cervicovaginal self-sample, pipelle endometrial biopsy and surgical specimen of either the EC or normal endometrium were obtained. A targeted next-generation sequencing panel was used to analyse these samples for mutations in eight genes. Sensitivity, specificity and predictive values were calculated. Fifty-nine EC patients and 31 control patients were included. In these patients, traditional histopathological diagnosis by pipelle had a sensitivity of 79% and a specificity of 100%. For EC patients, 97% of surgical specimens contained at least one mutation. Mutational analysis of Pap brush samples, self-samples and pipelle endometrial biopsies yielded a sensitivity of 78, 67 and 96% with a specificity of 97, 97 and 94%, respectively. Combining one of these three methods with histopathological pipelle endometrial biopsy evaluations yielded a sensitivity of 96, 93 and 96%, respectively. Our study has shown that mutational analysis of either cervical cytology or pipelle endometrial biopsies improves diagnosis of EC. Prospective validation will support implementation in clinical practice.

Preoperative risk stratification in endometrial cancer (ENDORISK) by a Bayesian network model: A development and validation study.

BACKGROUND: Bayesian networks (BNs) are machine-learning-based computational models that visualize causal relationships and provide insight into the processes underlying disease progression, closely resembling clinical decision-making. Preoperative identification of patients at risk for lymph node metastasis (LNM) is challenging in endometrial cancer, and although several biomarkers are related to LNM, none of them are incorporated in clinical practice. The aim of this study was to develop and externally validate a preoperative BN to predict LNM and outcome in endometrial cancer patients. METHODS AND FINDINGS: Within the European Network for Individualized Treatment of Endometrial Cancer (ENITEC), we performed a retrospective multicenter cohort study including 763 patients, median age 65 years (interquartile range [IQR] 58-71), surgically treated for endometrial cancer between February 1995 and August 2013 at one of the 10 participating European hospitals. A BN was developed using score-based machine learning in addition to expert knowledge. Our main outcome measures were LNM and 5-year disease-specific survival (DSS). Preoperative clinical, histopathological, and molecular biomarkers were included in the network. External validation was performed using 2 prospective study cohorts: the Molecular Markers in Treatment in Endometrial Cancer (MoMaTEC) study cohort, including 446 Norwegian patients, median age 64 years (IQR 59-74), treated between May 2001 and 2010; and the PIpelle Prospective ENDOmetrial carcinoma (PIPENDO) study cohort, including 384 Dutch patients, median age 66 years (IQR 60-73), treated between September 2011 and December 2013. A BN called ENDORISK (preoperative risk stratification in endometrial cancer) was developed including the following predictors: preoperative tumor grade; immunohistochemical expression of estrogen receptor (ER), progesterone receptor (PR), p53, and L1 cell adhesion molecule (L1CAM); cancer antigen 125 serum level; thrombocyte count; imaging results on lymphadenopathy; and cervical cytology. In the MoMaTEC cohort, the area under the curve (AUC) was 0.82 (95% confidence interval [CI] 0.76-0.88) for LNM and 0.82 (95% CI 0.77-0.87) for 5-year DSS. In the PIPENDO cohort, the AUC for 5-year DSS was 0.84 (95% CI 0.78-0.90). The network was well-calibrated. In the MoMaTEC cohort, 249 patients (55.8%) were classified with <5% risk of LNM, with a false-negative rate of 1.6%. A limitation of the study is the use of imputation to correct for missing predictor variables in the development cohort and the retrospective study design. CONCLUSIONS: In this study, we illustrated how BNs can be used for individualizing clinical decision-making in oncology by incorporating easily accessible and multimodal biomarkers. The network shows the complex interactions underlying the carcinogenetic process of endometrial cancer by its graphical representation. A prospective feasibility study will be needed prior to implementation in the clinic.

Oestrogen receptor pathway activity is associated with outcome in endometrial cancer.

BACKGROUND: Oestrogen receptor (ER) expression is a prognostic biomarker in endometrial cancer (EC). However, expression does not provide information about the functional activity of the ER pathway. We evaluated a model to quantify ER pathway activity in EC, and determined the prognostic relevance of ER pathway activity. METHODS: ER pathway activity was measured in two publicly available datasets with endometrial and EC tissue, and one clinical cohort with 107 samples from proliferative and hyperplastic endometrium and endometrioid-type EC (EEC) and uterine serous cancer (USC). ER pathway activity scores were inferred from ER target gene mRNA levels from Affymetrix microarray data (public datasets), or measured by qPCR on formalin-fixed paraffin-embedded samples (clinical cohort) and related to ER expression and outcome. RESULTS: ER pathway activity scores differed significantly throughout the menstrual cycle supporting the validity of the pathway test. The highest ER pathway scores were found in proliferative and hyperplastic endometrium and stage I EEC, whereas stage II-IV EEC and USCs had significantly lower levels. Low ER pathway activity was associated with recurrent disease, and added prognostic value in patients with low ER expression. CONCLUSION: The ER pathway test reflects activity of the ER pathway, and may improve prediction of outcome in EC patients.

RNA-based high-risk HPV genotyping and identification of high-risk HPV transcriptional activity in cervical tissues.

Nearly all cervical cancers are initiated by a persistent infection with one of the high-risk human papillomaviruses (high-risk HPV). High-risk HPV DNA testing is highly sensitive but cannot distinguish between active, productive infections and dormant infections or merely deposited virus. A solution for this shortcoming may be the detection of transcriptional activity of viral oncogenes instead of mere presence of high-risk HPVs. In this study, fresh-frozen cervical tissues (n = 22) were subjected to high-risk HPV DNA detection using the line probe assay and to targeted RNA next-generation sequencing using single-molecule molecular inversion probes. Targeted RNA sequencing was applied for (1) RNA-based genotyping of high-risk HPV, giving information on specific HPV-subtype (2) discrimination of E2, E6, and E7 transcripts and (3) discovery of possible non-HPV cancer biomarkers. Data were analyzed using computational biology. Targeted RNA sequencing enabled reliable genotyping of high-risk HPV subtypes and allowed quantitative detection of E2, E6, and E7 viral gene expression, thereby discriminating cervical lesions from normal cervical tissues. Moreover, targeted RNA sequencing identified possible cervical cancer biomarkers other than high-risk HPV. Interestingly, targeted RNA sequencing also provided high-quality transcription profiles from cervical scrape samples, even after 1 week of dry storage or storage in Preservcyt fixative. This proof of concept study shows that targeted RNA sequencing can be used for high-risk HPV genotyping and simultaneous detection of high-risk HPV gene activity. Future studies are warranted to investigate the potential of targeted RNA sequencing for risk assessment for the development of cervical lesions, based on molecular analysis of cervical scrapes.

Two types of primary mucinous ovarian tumors can be distinguished based on their origin.

The origin of primary mucinous ovarian tumors is unknown. We explore the hypothesis that they originate from either Brenner tumors or teratomas and examine differences between the tumors that arise in these settings. A total of 104 Brenner tumor-associated mucinous tumors and 58 teratoma-associated mucinous tumors were analyzed. Immunohistochemistry for 21 antigens and fluorescence in situ hybridization for ERBB2 and MYC were performed. Genome-wide copy number analysis and mutation analysis for 56 cancer-related genes was carried out on a subset of mucinous ovarian tumors and their complementary Brenner tumor or teratoma. Patients with teratoma-associated mucinous tumors were significantly younger than patients with Brenner tumor-associated mucinous tumors (43 vs. 61 years). During progression from cystadenoma to atypical proliferative mucinous (borderline) tumor to carcinoma expression of typical gastrointestinal markers was increased in both Brenner tumor-associated and teratoma-associated mucinous tumors. Brenner tumor-associated mucinous tumors showed more frequently calcifications and Walthard cell nests, rarely expressed SATB2 and showed more often co-deletion of CDKN2A and MTAP. Teratoma-associated mucinous tumors were characterized by mucinous stromal dissection, SATB2 expression and RNF43 mutations. Other frequent mutations in both Brenner tumor-associated and teratoma-associated mucinous tumors were TP53 and KRAS mutations. Based on identical mutations or copy number profiles clonal relationships were indicated in two mucinous tumors and their associated Brenner tumor. Teratomas and Brenner tumors give rise to different subtypes of mucinous ovarian tumors. Subsequent progression pathways are comparable since both Brenner tumor-associated and teratoma-associated mucinous tumors develop a gastrointestinal immunophenotype during progression and show early mutations in KRAS and TP53. Teratoma-associated mucinous tumors may more closely resemble true gastrointestinal tumors, indicated by their expression of SATB2 and the presence of RNF43 mutations.

The risk of cervical cancer after cervical intraepithelial neoplasia grade 3: A population-based cohort study with 80,442 women.

OBJECTIVE: To estimate the risk of cervical cancer in women with a history of cervical intraepithelial neoplasia (CIN) grade 3 and to review the compliance with post-treatment follow-up. METHODS: A population-based retrospective cohort study including 80,442 women with a median follow-up of 15.8 years, and 1,278,297 person years. Women with CIN3 between 1990 and 2010 were identified from the Dutch Pathology Registry (PALGA) and linked to the general female population from the Netherlands Cancer Registry. Cases of recurrent CIN3 and cervical cancer, defined as occurrence minimally two years post-treatment, were identified until 2016. Standardized incidence ratios (SIRs) were calculated for the risk of cervical cancer. RESULTS: 1554 women (1.9%) developed recurrent CIN3 and 397 women (0.5%) cervical cancer. Women with CIN3 were associated with a twofold increased risk of cervical cancer (SIR 2.29; 95%CI 2.07-2.52) compared with the general female population. Women aged ≥50 years during CIN3 diagnosis had a sevenfold and women with recurrent CIN3 a ninefold increased risk of developing cervical cancer. The increased risk up to 20 years of follow-up seems to be mostly attributable to ageing. 37.0% of women who developed cervical cancer after CIN3 did not complete the advised post-treatment follow-up. CONCLUSIONS: Women with CIN3 have a long-lasting twofold increased risk of developing cervical cancer, even when they complete the post-treatment follow-up and adhere to the regular screening program. This risk increases with CIN3 diagnosis at older age, further ageing during follow-up and in women with recurrent CIN3. Studies on optimizing follow-up strategies are warranted.

Fallopian tube abnormalities in uterine serous carcinoma.

OBJECTIVE: Uterine serous carcinoma (USC) is presumed to arise from endometrial intra-epithelial carcinoma (EIC), whereas tubo-ovarian high-grade serous carcinomas have similar precursor lesions in the Fallopian tube, i.e. serous tubal intra-epithelial carcinoma (STIC). The presence of Fallopian tube abnormalities and their clonal relationship to the concurrent USC was investigated. METHODS: In this multicenter study, all patients treated for USC between 1992 and 2017 were retrospectively identified. Histopathological diagnosis of USC, EIC and STIC was revised by an expert pathologist. Additionally, all Fallopian tube sections were immunohistochemically stained (p53 and Ki-67). Fallopian tube abnormalities were classified as either p53 signature, serous tubal intra-epithelial lesion (STIL) or STIC. The USCs and Fallopian tube abnormalities were analyzed by targeted next-generation sequencing. RESULTS: In 168 included patients, Fallopian tube abnormalities were found in 27.4% (46/168): p53-signatures in 17.9% (30/168), STILs in 3.0% (5/168) and STICs in 6.5% (11/168). In subgroup analysis, STICs were found in 9.5% (11/115) of patients with at least one section of the fimbriated end embedded. Next-generation sequencing showed identical TP53-mutations in the STIC and corresponding USC. CONCLUSIONS: In conclusion, the presence of Fallopian tube abnormalities was shown in a high percentage of patients with USC, representing either true precursor lesions or metastasized disease.

Clinical practice variation and overtreatment risk in women with abnormal cervical cytology in the Netherlands: two-step versus see-and-treat approach.

BACKGROUND: Only a few small studies have compared the 2-step method (biopsy followed by treatment) with a see-and-treat (immediate treatment) approach in women both low-grade or high-grade referral cytology. The clinical practice variation in the Netherlands has not been reviewed before. OBJECTIVES: To determine overtreatment rates in the 2-step versus see-and-treat approach in women referred for colposcopy because of abnormal cytology results, and to evaluate clinical practice variation in the Netherlands. MATERIALS AND METHODS: This was a population-based retrospective cohort study including 36,581 women with a histologic result of the cervix identified from the Dutch Pathology Registry (PALGA) between 2016 and 2017. Odds ratios for overtreatment, defined primarily as cervical intraepithelial neoplasia grade 1 or less, were determined for the 2-step and see-and-treat approach in relation to age, high-risk human papillomavirus status, and referral cytology. RESULTS: Of the included women 10,713 women (29.3%) received the 2-step method; 6,851 women (18.7%) underwent see-and-treat; and 19,017 women (52.0%) received conservative management after colposcopy with histologic assessment with cytologic follow-up or another type of treatment. Despite the existence of a national guideline advising see-and-treat only in case of suspected high-grade disease in women who have completed their childbearing, there is a wide practice variation between the 2 strategies in the Netherlands, with 7.0-88.3% of the women receiving see-and-treat per laboratory. The median time between cytology and treatment was 1-2 months (range, 0-12 months) in women receiving see-and-treat and the 2-step method, respectively. A total of 4119 women (23.5%) were overtreated, with older women, high-risk human papillomavirus-negative women, and women with low-grade cytology results being more likely to be overtreated. Women with low-grade cytology results and see-and-treat were associated with a higher overtreatment rate than women receiving the 2-step method (65.0% [1414 of 2174] versus 32.1% [1161 of 3613], respectively; odds ratio, 3.34; 95% confidence interval, 2.92-3.82). However, in women with high-grade cytology results, see-and-treat was inversely associated with overtreatment (11.3% [529 of 4677] versus 14.3% [1015 of 7100], respectively; odds ratio, 0.68; 95% confidence interval, 0.58-0.81). CONCLUSION: A see-and-treat approach is justified only in women with high-grade cytology results who have completed their childbearing. There is a wide practice variation between the 2 strategies in the Netherlands, and gynecologists should adhere to the guideline to prevent overtreatment.

Risk prediction of cervical abnormalities: The value of sociodemographic and lifestyle factors in addition to HPV status.

High-risk human papillomavirus (hrHPV) assessment as a primary screening test improves sensitivity but decreases specificity. Determining risk for cervical abnormalities and adapting policy accordingly may improve the balance between screening benefits and harms. Our aim is to assess the value of factors other than HPV in prediction of cervical abnormalities. Data from a Dutch prospective cohort were used. Women aged 18-29 years, not yet eligible for screening, were included in 2007. Data collection consisted of a questionnaire and a cervicovaginal self-sample. Linkage with PALGA (pathology database) was performed in 2017. The analyses included 1483 women. The full model, including sociodemographic and lifestyle factors, was compared to the null model, including baseline HPV only. The outcome of interest was cervical intraepithelial neoplasia 2 or worse (CIN2+). There were 86 women with CIN2+. Baseline hrHPV status was an important predictor (OR = 5.20, 95%CI = 3.27-8.27). The area under the ROC curve (AUC) of the null model was 0.67 (95%CI = 0.61-0.72). The full model had a slightly higher AUC of 0.73 (95%CI = 0.67-0.79). Bootstrap validation indicated that overfitting was present. This exploratory study has confirmed that a single hrHPV measurement is a strong predictor of cervical abnormalities, and additional risk factors in young women appeared to have limited added value. However, prediction based on hrHPV only does leave room for improvement. Future studies should therefore focus on women in the screening age range and search for other predictors to further enhance risk prediction. Adapting policy based on risk may eventually help optimise screening performance.

Primary Ovarian Tumors With Lymphogenic and Hematogenic Metastasis Express High MMP-14, Which Colocalizes With Highly Sulfated Chondroitin Sulfate in the Stroma.

Lymphogenic and hematogenic metastases are uncommon in ovarian cancer, especially at presentation. We hypothesized that MMP-14 and MMP-2, CD44, and highly sulfated chondroitin sulfate (CS-E) may be overexpressed in tumors with these metastatic patterns. These molecules are all present in the ovarian tumor microenvironment, wherein they may interact. In an ovarian cancer cohort of 44 patients with metastases in lymph nodes, spleen, and/or liver, the presence of MMP-14, MMP-2, CD44, and CS-E in both the primary tumor and the metastases was determined with immunohistochemistry and related to clinical characteristics. Immunohistochemical expression was found for MMP-14 in all primary tumors as well as in all metastases and for MMP-2 expression in most of the samples. Most primary tumors with synchronous metastases were positive for CS-E, as well as most primary tumors with metachronous lymphogenic metastases. The expression of the MMPs and CS-E in the stroma seemed to colocalize. For CD44 immunohistochemical expression, this relationship was not found. Epithelial MMP-14 on the one hand and stromal CS-E on the other hand seem to be essential players in ovarian cancer with lymphogenic and hematogenic metastases. CD44 expression is not correlated with the other markers. More research on the interaction of these molecules and their role in the process of dissimination of disease is warranted.

Short-term surgical complications after radical hysterectomy-A nationwide cohort study.

INTRODUCTION: Centralization has, among other aspects, been argued to have an impact on quality of care in terms of surgical morbidity. Next, monitoring quality of care is essential in identifying areas of improvement. This nationwide cohort study was conducted to determine the rate of short-term surgical complications and to evaluate its possible predictors in women with early-stage cervical cancer. MATERIAL AND METHODS: Women diagnosed with early-stage cervical cancer, 2009 FIGO stages IB1 and IIA1, between 2015 and 2017 who underwent radical hysterectomy with pelvic lymphadenectomy in 1 of the 9 specialized medical centers in the Netherlands, were identified from the Netherlands Cancer Registry. Women were excluded if primary treatment consisted of hysterectomy without parametrial dissection or radical trachelectomy. Women in whom radical hysterectomy was aborted during the procedure, were also excluded. Occurrence of intraoperative and postoperative complications and type of complications, developing within 30 days after surgery, were prospectively registered. Multivariable logistic regression analysis was used to identify predictors of surgical complications. RESULTS: A total of 472 women were selected, of whom 166 (35%) developed surgical complications within 30 days after radical hysterectomy. The most frequent complications were urinary retention with catheterization in 73 women (15%) and excessive perioperative blood loss >1000 mL in 50 women (11%). Open surgery (odds ratio [OR] 3.42; 95% CI 1.73-6.76), chronic pulmonary disease (OR 3.14; 95% CI 1.45-6.79), vascular disease (OR 1.90; 95% CI 1.07-3.38), and medical center (OR 2.83; 95% CI 1.18-6.77) emerged as independent predictors of the occurrence of complications. Body mass index (OR 0.94; 95% CI 0.89-1.00) was found as a negative predictor of urinary retention. Open surgery (OR 36.65; 95% CI 7.10-189.12) and body mass index (OR 1.15; 95% CI 1.08-1.22) were found to be independent predictors of excessive perioperative blood loss. CONCLUSIONS: Short-term surgical complications developed in 35% of the women after radical hysterectomy for early-stage cervical cancer in the Netherlands, a nation with centralized surgical care. Comorbidities predict surgical complications, and open surgery is associated with excessive perioperative blood loss.

Performance of human papillomavirus testing on self-collected versus clinician-collected samples for the detection of cervical intraepithelial neoplasia of grade 2 or worse: a randomised, paired screen-positive, non-inferiority trial.

BACKGROUND: Human papillomavirus (HPV) testing on self-collected samples is a potential alternative to HPV testing on clinician-collected samples, but non-inferiority of its clinical accuracy remains to be assessed in the regular screening population. The IMPROVE study was done to evaluate the clinical accuracy of primary HPV testing on self-collected samples within an organised screening setting. METHODS: In this randomised, non-inferiority trial, women aged 29-61 years were invited to participate in the study as part of their regular screening invitation in the Netherlands. Women who provided informed consent were randomly allocated (1:1, with a block size of ten stratified by age) to one of two groups: a self-sampling group, in which women were requested to collect their own cervicovaginal sample using an Evalyn Brush (Rovers Medical Devices BV, Oss, Netherlands); or a clinician-based sampling group, in which samples were collected by a general practitioner with a Cervex-Brush (Rovers Medical Devices BV). All samples were tested for HPV using the clinically validated GP5+/6+ PCR enzyme immunoassay (Labo Biomedical Products BV, Rijswijk, Netherlands). HPV-positive women in both groups were retested with the other collection method and triaged by cytology and repeat cytology in accordance with current Dutch screening guidelines. Primary endpoints were detection of cervical intraepithelial neoplasia (CIN) of grade 2 or worse (CIN2+) and grade 3 or worse (CIN3+). Non-inferiority of HPV testing on self-collected versus clinician-collected samples was evaluated against a margin of 90% for the relative sensitivity and 98% for the relative specificity. This study is registered at the Dutch Trial register (NTR5078) and has been completed. FINDINGS: Of the 187 473 women invited to participate, 8212 were randomly allocated to the self-sampling group and 8198 to the clinician-based sampling group. After exclusion of women who met the exclusion criteria or who did not return their sample, 7643 women were included in the self-sampling group and 6282 in the clinician-based sampling group. 569 (7·4%) self-collected samples and 451 (7·2%) clinician-collected samples tested positive for HPV (relative risk 1·04 [95% CI 0·92-1·17]). Median follow-up duration for HPV-positive women was 20 months (IQR 17-22). The CIN2+ sensitivity and specificity of HPV testing did not differ between self-sampling and clinician-based sampling (relative sensitivity 0·96 [0·90-1·03]; relative specificity 1·00 [0·99-1·01]). For the CIN3+ endpoint, relative sensitivity was 0·99 (0·91-1·08) and relative specificity was 1·00 (0·99-1·01). INTERPRETATION: HPV testing done with a clinically validated PCR-based assay had similar accuracy on self-collected and clinician-collected samples in terms of the detection of CIN2+ or CIN3+ lesions. These findings suggest that HPV self-sampling could be used as a primary screening method in routine screening. FUNDING: Ministry of Health, Welfare, and Sport (Netherlands), and the European Commission.

Genome-wide microRNA analysis of HPV-positive self-samples yields novel triage markers for early detection of cervical cancer.

Offering self-sampling for HPV testing improves the effectiveness of current cervical screening programs by increasing population coverage. Molecular markers directly applicable on self-samples are needed to stratify HPV-positive women at risk of cervical cancer (so-called triage) and to avoid over-referral and overtreatment. Deregulated microRNAs (miRNAs) have been implicated in the development of cervical cancer, and represent potential triage markers. However, it is unknown whether deregulated miRNA expression is reflected in self-samples. Our study is the first to establish genome-wide miRNA profiles in HPV-positive self-samples to identify miRNAs that can predict the presence of CIN3 and cervical cancer in self-samples. Small RNA sequencing (sRNA-Seq) was conducted to determine genome-wide miRNA expression profiles in 74 HPV-positive self-samples of women with and without cervical precancer (CIN3). The optimal miRNA marker panel for CIN3 detection was determined by GRridge, a penalized method on logistic regression. Six miRNAs were validated by qPCR in 191 independent HPV-positive self-samples. Classification of sRNA-Seq data yielded a 9-miRNA marker panel with a combined area under the curve (AUC) of 0.89 for CIN3 detection. Validation by qPCR resulted in a combined AUC of 0.78 for CIN3+ detection. Our study shows that deregulated miRNA expression associated with CIN3 and cervical cancer development can be detected by sRNA-Seq in HPV-positive self-samples. Validation by qPCR indicates that miRNA expression analysis offers a promising novel molecular triage strategy for CIN3 and cervical cancer detection applicable to self-samples.

Poor outcome in hypoxic endometrial carcinoma is related to vascular density.

BACKGROUND: Identification of endometrial carcinoma (EC) patients at high risk of recurrence is lacking. In this study, the prognostic role of hypoxia and angiogenesis was investigated in EC patients. METHODS: Tumour slides from EC patients were stained by immunofluorescence for carbonic anhydrase IX (CAIX) as hypoxic marker and CD34 for assessment of microvessel density (MVD). CAIX expression was determined in epithelial tumour cells, with a cut-off of 1%. MVD was assessed according to the Weidner method. Correlations with disease-specific survival (DSS), disease-free survival (DFS) and distant disease-free survival (DDFS) were calculated using Kaplan-Meier curves and Cox regression analysis. RESULTS: Sixty-three (16.4%) of 385 ECs showed positive CAIX expression with high vascular density. These ECs had a reduced DSS compared to tumours with either hypoxia or high vascular density (log-rank p = 0.002). Multivariable analysis showed that hypoxic tumours with high vascular density had a reduced DSS (hazard ratio [HR] 3.71, p = 0.002), DDFS (HR 2.68, p = 0.009) and a trend for reduced DFS (HR 1.87, p = 0.054). CONCLUSIONS: This study has shown that adverse outcome in hypoxic ECs is seen in the presence of high vascular density, suggesting an important role of angiogenesis in the metastatic process of hypoxic EC. Differential adjuvant treatment might be indicated for these patients.