Hysteroscopic Evaluation of Endometrial Changes in Breast Cancer Women with or without Hormone Therapies: Results from a Large Multicenter Cohort Study.

STUDY OBJECTIVE: The primary aim of our study was to investigate the incidence of endometrial pathologies, especially endometrial cancer, in women with breast cancer treated with tamoxifen (TAM), aromatase inhibitors (AIs), or receiving no treatment (NT). The secondary aim was to identify, in this cohort, ultrasonographic findings that represent robust indications for hysteroscopy and endometrial biopsy, to avoid unnecessary second-level diagnostic procedures. DESIGN: Multicenter retrospective cohort study (Clinical Trial ID: NCT03898947). SETTING: Data were collected from different Italian centers: Regina Elena National Cancer Institute of Rome, Arbor Vitae Centre of Rome, Gaetano Martino University Hospital of Messina, and Villa Sofia-Cervello Hospital of Palermo. PATIENTS: We selected and consecutively included patients with a history of breast cancer who had undergone hysteroscopy for ultrasonographic or clinical indications between January 2007 and December 2016. INTERVENTIONS: Diagnostic hysteroscopy with endometrial biopsy or operative hysteroscopy, when clinically indicated. MEASUREMENTS AND MAIN RESULTS: A higher percentage of patients in the TAM and AI groups had a normal endometrium compared with those in the NT group, whereas the incidence of endometrial polyps was higher in the NT group than in the others; no significant differences were observed among the 3 groups for other benign conditions or for premalignant and malignant uterine diseases, such as endometrial atypical hyperplasia and adenocarcinoma. CONCLUSION: TAM treatment does not seem to be associated with a higher rate of endometrial cancer in women with breast cancer compared with women treated with AIs or NT.

Claspin as a biomarker of human papillomavirus-related high grade lesions of uterine cervix.

BACKGROUND: Claspin is a nuclear protein involved in DNA replication and damage response and is a key mediator for the S-phase checkpoint. Claspin expression is significantly high in several human solid tumors. Furthermore, high levels of claspin have been found in cervical cancer cell lines. Nevertheless, no data are available regarding claspin expression in cervical tissues. METHODS: In order to investigate whether claspin immunoreactivity is related to the lesion severity and High-Risk (HR) HPV infection, we analyzed claspin expression by immunohistochemistry in a series of cervical biopsies which represent the steps occurring during cervical carcinogenesis (normal tissues, Cervical Intraepithelial Neoplasias 1, 2 and 3, Squamous Cell Carcinomas). All patients also had a cervico-vaginal sample for HPV testing, collected immediately before the colposcopy-guided biopsy. The HR-HPV DNA detection was performed by the HR-HPV Hybrid Capture 2 test. HPV genotyping was performed using the Linear Array HPV Genotyping Test. RESULTS: Our results evidenced a constant and significant increase of the rate of claspin positivity from the normal tissues to carcinomas (pχ2(trend) < 0.0001). In fact, the normal tissues displayed either no or faint claspin immunoreactivity, whereas a moderate/high positivity was observed in 16% of the CIN1, 76% of the CIN2, 87.5% of the CIN3 and 93.3% of the cancers. Moreover, we found a statistically significant correlation between claspin expression and HR-HPV infection (pχ2 < 0.0001), irrespective of the genotype. Finally, we demonstrated the feasibility of claspin immunostaining in cervical cytology. CONCLUSIONS: Our findings indicate that in vivo claspin expression is significantly related to HR-HPV infection and lesion grade both in histological and cytological samples. Therefore, the analysis of claspin expression could be clinically relevant in the diagnosis of HPV-related cervical lesions, in particular when applied to cervico-vaginal cytology. Moreover, giving information on the proliferation rate of each lesion, claspin immunostaining may contribute to the evaluation of progression risk, thus being helpful in patient management. Nevertheless, only large prospective studies may clarify the true clinical usefulness of claspin expression in distinguishing lesions with different progression potential.

p16/Ki-67 dual staining in cervico-vaginal cytology: correlation with histology, Human Papillomavirus detection and genotyping in women undergoing colposcopy.

OBJECTIVES: 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. METHODS: 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. RESULTS: 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). CONCLUSIONS: 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.

Diagnostic and prognostic validity of the human papillomavirus E6/E7 mRNA test in cervical cytological samples of HC2-positive patients.

The study aimed to assess the clinical utility in identifying CIN2 or worse (CIN2+), of the Pretect HPV-Proofer test for E6/E7 mRNA detection in Hybrid Capture 2 (HC2)-positive patients, who underwent colposcopy. In particular, the study analyzed the mRNA test performance as the third test in a subgroup of HC2+ patients with less severe than high-grade squamous intraepithelial lesions (HSIL-). We analyzed 464 cervico-vaginal samples by liquid-based cytology (LBC) and PreTect HPV-Proofer. Moreover 231 patients also had a biopsy at baseline and 75, with HSIL-, were followed up within 2 years by LBC, colposcopy, and histology when indicated. The highest sensitivity for CIN2+ belonged to the mRNA compared to LBC, at the HSIL+ threshold (72% vs. 58%), whereas the LBC showed the highest specificity and positive predictive value (PPV) (99 and 93% vs. 73 and 39%, respectively). Focusing on the 408 HSIL- patients, the mRNA positivity was significantly more associated with CIN2+ than CIN2- lesions (p < 0.0001). Moreover, among the 75 HSIL- followed up patients, the mRNA displayed high longitudinal Specificity (89%), even if the sensitivity and the PPV were low (50 and 20%, respectively). The present data suggest that the mRNA test may have a diagnostic and a potentially prognostic role in HC2+/HSIL- patients.

Comparative evaluation of nm23 and p16 expression as biomarkers of high-risk human papillomavirus infection and cervical intraepithelial neoplasia 2(+) lesions of the uterine cervix.

AIMS: To investigate the clinical role of nm23 expression in identifying both high-risk human papillomavirus (HR-HPV) and high-grade cervical lesions or carcinomas [cervical intraepithelial neoplasia 2(+) (CIN2(+) )], and to compare it with p16 overexpression, as this latter biomarker has already been reported widely in HR-HPV infected cervical lesions. METHODS AND RESULTS: Immunohistochemical evaluation of nm23 and p16 in 143 cervical biopsy specimens including negative, low- and high-grade lesions and squamous carcinomas (SC). HR-HPV testing by Digene hybrid capture 2 (HC2) and polymerase chain reaction (PCR) on the cervico-vaginal samples of the same patients. In detecting CIN2(+) , p16 was significantly more sensitive and specific than nm23 (96.3% versus 81.8% and 66% versus 36.4%, respectively, both P < 0.0001). Concerning HR-HPV detection by HC2, p16 showed a significantly higher specificity than nm23 (82% versus 47%, P <0.0001), although the sensitivities were comparable (71% versus 76%). We found a significantly direct correlation between nm23 and HC2 findings. However, nm23 expression did not correlate with HPV16/18 infection. In contrast, we observed a significant association between p16 overexpression and HPV16/18 genotypes. CONCLUSIONS: We confirm the diagnostic value of p16 overexpression. Moreover, despite in vitro data regarding the interaction with the HPV-E7 protein, nm23 does not appear to be a more useful biomarker than p16 in identifying CIN2(+) or HR-HPV infection.

Development of new medical treatment for epithelial ovarian cancer recurrence.

Epithelial ovarian cancer (EOC) is the scariest gynaecological cancer. Many advances have been done with evolving knowledge, leading to the introduction of new drugs, most in maintenance setting. The antiangiogenic Bevacizumab and the three approved PARP-inhibitors-olaparib, niraparib and rucaparib-are gradually improving PFS of patients with EOC, with initial effects on OS too. But recurrence is still a heavy sentence and lethality continues to be high. Ovarian cancer is a complex disease, with different clinical presentation, histological aspect, and molecular expression, leading to disappointing results, when using a single drug. Implementation of biobanking and analysis of patients' tumour samples, before starting a treatment, could be a promising way to better understand molecular aspects of this disease, to identify markers predictive of response and to allow a better use of experimental drugs, as immunomodulators, targeted therapies, and combinations of these, to fight tumour growth and clinical progression. We reviewed the literature on the updated treatments for recurrent ovarian cancer, summarizing all the available drugs and combinations to treat patients with this diagnosis, and focusing the attention on the new approved molecules and the contemporary Clinical Trials, investigating new target therapies and new associations.

Practical recommendations for gynecologic surgery during the COVID-19 pandemic.

Surgery in suspected/confirmed COVID-19 patients is a high-risk venture. In infected patients, COVID-19 is present in the body cavity. During surgery it could be nebulized in the spray generated by surgical instruments and could theoretically infect members of the surgical team. Nevertheless, some surgical gynecologic pathologies cannot be postponed. We present a list of the most frequent gynecologic diseases and recommendations on their surgical management during the COVID-19 pandemic, based on expert opinion, current available information, and international scientific society recommendations to support the work of gynecologists worldwide. In brief, any kind of surgical treatment should be scrutinized and postponed if possible. Nonoperative conservative treatment including pharmacological therapies for hormone-sensitive pathologies should be implemented. Health risk assessment by patient history and COVID-19 test before elective surgery are pivotal to protect both patients and healthcare providers. In confirmed COVID-19 patients or highly suspected cases, elective surgery should be postponed until full recovery.

Clinical role of p16INK4a expression in liquid-based cervical cytology: correlation with HPV testing and histologic diagnosis.

p16INK4a is overexpressed in high-risk human papillomavirus (HR-HPV)-infected preneoplastic and neoplastic lesions of the uterine cervix. Our aim was to verify whether p16 is a diagnostic marker also in cervical liquid-based cytology. We performed p16 immunocytochemical analysis and the Hybrid Capture 2 (HC2) test (Digene, Gaithersburg, MD) for HR-HPV infection in 471 ThinPrep-processed (Cytyc, Boxborough, MA) cervicovaginal samples and correlated the results with histologic findings. A total of 32.3% of the specimens showed p16 immunoreactivity, whereas the HC2 test was positive in 41.2% of the cases (65.2% concordance rate). Correlating the cytologic, p16, and HPV results with histologic findings revealed HC2 as the most sensitive test for a diagnosis of cervical intraepithelial neoplasia 2 or worse, whereas cytologic examination was the most specific. The positive predictive value was significantly higher for cytologic examination than for p16 and HR-HPV testing. These data suggest that p16 evaluation in ThinPrep samples does not have better clinical effectiveness for identifying high-grade lesions than conventional morphologic examination and HPV testing.

Prognostic value of HER2 and progesterone receptor expression in endometrial carcinoma with positive peritoneal washing.

BACKGROUND: Although the majority of endometrial cancer (EC) patients can be cured by surgery, unexpected recurrent disease may also occur in early stage patients. In the present study, whether or not the analysis of multiple biopathological parameters might lead to more accurate predictions of the clinical outcome of EC patients with long-term follow-up (FU) was investigated. PATIENTS AND METHODS: Estrogen and progesterone receptor (ER and PgR) positivity and HER2 overexpression by immunohistochemistry were evaluated. The peritoneal washings (PWs) were analyzed by cytology and immunocytochemistry employing AR-3 and B72.3 monoclonal antibodies. RESULTS: The patients with positive PW and HER2 positive tumors showed shorter overall survival compared to those bearing HER2 negative tumors (p =0.004). HER2 overexpression also influenced the patient outcome in the group with tumors lacking PgR (p = 0.004). At multivariate analysis PgR and HER2 overexpression emerged as independent prognostic factors. CONCLUSION: The combined analysis of these biopathological markers could provide useful information for the selection of patients to be enrolled in innovative therapeutic strategies.

Prevalence of HPV infection among clinically healthy Italian males and genotype concordance between stable sexual partners.

BACKGROUND: HPV is one of the most common sexually transmitted infections. However little is known about its prevalence in the male population and concordance with female partners. OBJECTIVES: This cross-sectional study aimed to: (a) investigate HPV prevalence and genotype distribution among a series of stable male sexual partners of CIN/HPV positive women and (b) assess HPV infection and type-specific concordance between partners. STUDY DESIGN: 378 stable and monogamous male partners of CIN/HPV positive women were selected. Of these, 238 cases were enrolled at the same time as their female partner. All the subjects were tested by the Linear Array HPV genotyping assay. RESULTS: Overall, 153/378 men (40.5%) and 122/238 women (51.3%) were positive for at least one of the 37 HPV types detectable by the assay used. Among the HPV-positive participants, 69 of the 378 men (18.2%) and 54 of the 238 women (22.7%) harboured multiple genotypes. 75 couples (31.5%) were concordantly HPV positive, while 102 couples (42.9%) were concordantly negative (Kappa value: 0.491, p<0.0001). Among the couples in which both partners were HPV positive, 68% harboured at least one genotype in common. Results from a GEE model evidenced that when the male partner tested HPV positive for at least one genotype, this had a significant effect on the positivity of their relative female partner (p<0.0001). CONCLUSIONS: We evidenced a high prevalence of HPV male infections and a moderate concordance between partners. However, we observed a significant HPV type-specific correlation between partners, which is unlikely to be coincidental.

HPV prevalence among healthy Italian male sexual partners of women with cervical HPV infection.

Genital human papillomavirus (HPV) is the causative agent of cervical cancer and is the most common sexually transmitted infection. Only limited and controversial data are available regarding HPV transmission in male sexual partners of women with cervical intraepithelial neoplasia (CIN). The aim of this study was to investigate the prevalence and the genotype distribution of HPV in penile scrapings of a series of Italian men, who had no visible penile lesions and were partners of women who were affected, or had been affected previously by cervical intraepithelial neoplasia or who were infected with HPV. The concordance of the viral group in the infected partners was determined. A total of 77 penile scrapings were screened for HPV infection by the polymerase chain reaction, while 59 cervicovaginal brushings of their female partners were tested. 35% of evaluable male samples and 64% of female sexual partners were found to be HPV positive. In the 55 simultaneously evaluable couples, a concordance of 45% was found, 11 couples (20%) with both partners being HPV negative and 14 couples (25%) with both partners HPV positive (P=0.001). Six out of the 14 couples (43%), where both partners were HPV positive, harbored the same HPV genotype group. These data, although preliminary, could support further the hypothesis that male HPV infection is more frequent in sexual partners of HPV positive or women with cervical intraepithelial neoplasia indicating that men could represent an important source of HPV transmission between sex partners.

Immunohistochemical expression of p16(INK4a) is predictive of HR-HPV infection in cervical low-grade lesions.

The p16(INK4a) is a cyclin-dependent kinase inhibitor that decelerates the cell cycle by inactivating the cyclin-dependent kinases involved in the phosphorylation of the retinoblastoma protein (RB). Expression of E6 and E7 oncogenes of high-risk (HR) human papillomavirus (HPV), affecting the RB-p16 pathway, leads to p16 upregulation. Although it is widely reported that p16 is overexpressed in a high percentage of preneoplastic lesions and in almost all carcinomas of the uterine cervix, protein upregulation and its correlation with HPV infection in low-grade lesions is still being debated. In this study, we investigated in parallel, p16 expression and HPV infection in 100 cervical biopsies (17 normal tissues, 54 CIN1, 10 CIN2, 11 CIN3, eight invasive squamous cancers). Results obtained demonstrated that none of the 17 normal cervical tissues, evaluated by immunohistochemistry, presented p16 positivity whereas, starting from CIN1 (31%) to CIN2 (90%), CIN3 (100%) and carcinomas (100%), a constant and significant increase of protein overexpression (P<0.0001) was observed. In addition, p16 overexpression consistently showed elevated sensitivity (84%) and specificity (98%) in detecting HR-HPV infection with a high positive predictive value (97%) and negative predictive value (86%). Of interest, 93% of the p16-positive CIN1 were also HR-HPV infected. Our findings confirmed that p16 overexpression is associated to high-grade precancerous lesions and cervical carcinomas, and further demonstrated that immunohistochemical evaluation of p16 may be a useful biomarker in identifying HR-HPV-infected low-grade lesions.