Vaginal Microbiome and HPV Pre-malignant and Cervical Dysplasia

Infection with high-risk Human Papillomavirus (HPV) genotypes constitutes a well-recognized risk factor for cervical-carcinoma (CC). High-risk HPV features 10-15% persistence rate, consequently driving precancerous cervical-intraepithelial-neoplasia (CIN) and subsequent progression to CC. Multiple factors are believed to play permissive roles in the progression of CIN/CC , yet a molecular mechanism driving carcinogenesis across the CIN-CC continuum following persistent HPV infection remains elusive. The vaginal microbiome may play a role in the development of CIN-CC carcinogenesis, by modulation of host-immune-response and alteration of cervical microenvironment to become tumor-permissive. While suggested vaginal microbiome contributions include induction of altered epithelial cell adhesion and downregulation of DNA damage responses, no clear mechanism has been proven to date. The loss of Lactobacillus genus dominancy, and the switch to dysbiotic, high-diversity, high-pH, Bacterial Vaginosis (BV) is thought to play a key-role in HPV infection, persistence and carcinogenesis.

The current study consists of recruitment of a cohort of healthy, CIN and CC patients including vaginal samples and comprehensive metadata collection.

We will recruit three types of women as participants: 1) Individuals that underwent an HPV PCR cervical screening test and were found to be negative for HPV (healthy controls) 2) Women that were found to be positive for high risk-HPV but have no cervical dysplasia 3) Women who are positive for high risk-HPV and exhibit cervical pathology, either high-CIN (CIN 2-3) or CC.

After assessment of inclusion and exclusion criteria and patient's consent to participate, the patient will undergo gynecological examination. A swab for vaginal microbiome analysis and HPV testing will be conducted during the gynecological examination, as well as cytology test (as part as the standard examination). In case of a negative HPV testing, the patient will be considered a healthy control. In case of identification of high-risk HPV, the patient will be referred for colposcopy, and according to its results, will be assigned to group 2 (no cervical dysplasia) or group 3 (CIN 2-3 or CC).

Patients with either low-risk HPV or CIN1 will be excluded from the study. To allow recruitment of women with high grade CIN or CC, patients who attend the colposcopy clinic due to postcoital bleeding, abnormal HPV or abnormal cytology tests (which were conducted elsewhere) will be also requested to participate.

Sample and data collection : Patients will be asked questions evaluating demographics, medical and socioeconomic status, including age, marital status, gynecological and obstetrics history (hormonal status, parity, gravidity, age of sexual debut, method of contraception, previous STIs, genital herpes), smoking status (current/past), immune-deficiency status, medications, previous cervical pathologies and previous HPV-vaccination.

During gynecological exam and/or colposcopy, a sample for vaginal microbiome analysis will be obtained from those who were willing to participate in the study and who signed informed consent. Samples will be collected and stored in freezers at -80°.

Cytology/colposcopy/biopsy results will be collected following evaluation.

Samples for HPV as well as for vaginal microbiome analysis will be taken from HPV-positive patients (with or without CIN/CC) every 3-6 months, for 2 years, as well as documentation of clinical findings (colposcopy findings, cytology and histology) to evaluate for longitudinal alterations that may be parallel to persistence of HPV or its clearance. Follow-up appointments will be scheduled according to the accepted clinical guidelines.