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For cervical cancer (CC), circulating cell‐free HPV DNA (ccfHPV) may establish disease severity. Furthermore, HPV integration has been correlated to viral load and survival. In this study, pre‐treatment plasma from 139 CC cases (50 primary surgery patients, 22 primary surgery + adjuvant oncological therapy patients, and 67 primary oncological therapy patients) was collected (2018–2020). Furthermore, plasma from 25 cervical intraepithelial neoplasia grade 3 patients and 15 healthy women (negative controls) were collected. Two next‐generation sequencing (NGS) panels were used to establish ccfHPV presence and human papillomavirus type 16 (HPV16) integration status. ccfHPV was detected in four primary surgery (8.0%), eight primary surgery + adjuvant oncology (36.4%), and 54 primary oncology (80.6%) patients. For primary oncology patients with HPV16‐related cancer (n = 37), more ccfHPVneg than ccfHPVpos patients had HPV16 integration (P = 0.04), and in patients with HPV16 integration (n = 13), ccfHPVpos patients had higher disease stages than ccfHPVneg patients (P = 0.05). In summary, ccfHPV presence is related to disease severity and may add to the debated Sedlis criteria used for identifying patients for adjuvant oncological therapy. However, ccfHPV detection is influenced by HPV integration status and disease stage, and these factors need to be considered in ccfHPVneg patients.

For cervical cancer (CC), circulating cell‐free HPV DNA (ccfHPV) may establish disease severity. Furthermore, HPV integration has been correlated to viral load and survival. In this study, pre‐treatment plasma from 139 CC cases (50 primary surgery patients, 22 primary surgery + adjuvant oncological therapy patients, and 67 primary oncological therapy patients) was collected (2018–2020). Furthermore, plasma from 25 cervical intraepithelial neoplasia grade 3 patients and 15 healthy women (negative controls) were collected. Two next‐generation sequencing (NGS) panels were used to establish ccfHPV presence and human papillomavirus type 16 (HPV16) integration status. ccfHPV was detected in four primary surgery (8.0%), eight primary surgery + adjuvant oncology (36.4%), and 54 primary oncology (80.6%) patients. For primary oncology patients with HPV16‐related cancer (n = 37), more ccfHPVneg than ccfHPVpos patients had HPV16 integration (P = 0.04), and in patients with HPV16 integration (n = 13), ccfHPVpos patients had higher disease stages than ccfHPVneg patients (P = 0.05). In summary, ccfHPV presence is related to disease severity and may add to the debated Sedlis criteria used for identifying patients for adjuvant oncological therapy. However, ccfHPV detection is influenced by HPV integration status and disease stage, and these factors need to be considered in ccfHPVneg patients. [ABSTRACT FROM AUTHOR]

ABSTRACT Human papillomavirus (HPV) circulating tumor DNA (HPV ctDNA) was proposed as a biomarker for the detection and disease monitoring of HPV-related cancers. One hundred eighty plasma samples obtained from women diagnosed with HPV16-positive cervical cancer (CC) (n = 100), HPV16-positive premalignant lesions (cervical intraepithelial neoplasia grade 3 [CIN3]) (n = 20), and HPV DNA-negative controls (n = 60) were randomly selected from the archives for evaluating the performance of a bead-based HPV genotyping assay (E7 type-specific multiplex genotyping assay [E7-MPG]) in detecting HPV16 ctDNA. The performance of the E7-MPG was compared with those of DNA detection by droplet digital PCR (ddPCR) and detection of HPV16 E6 antibodies evaluated in an independent study. Internal controls to assess DNA quality were included in the molecular assays, i.e., beta-globin and ESR1, respectively. The sensitivity and specificity of E7-MPG and/or E6 antibodies to detect HPV16-positive CCs were evaluated. HPV16 ctDNA was detected using the E7-MPG in 42.3% of all plasma samples and in 74.7% of plasma samples from HPV16-positive CC cases. The validation of E7-MPG data by ddPCR showed that the sensitivity of the E7-MPG test for HPV16-positive CC detection was higher than that of ddPCR (74.7% versus 63.1%; P

A causal link between Human Papillomavirus (HPV) and breast cancer (BC) remains controversial. In spite of this, the observation that HPV DNA is over-represented in the Triple Negative (TN) BC has been reported. Here we remark the high prevalence of HPV DNA (44.4%) in aggressive BC subtypes (TN and HER2+) in a population of 273 Italian women and we convey the presence of HPV DNA in the epithelial and stromal compartments by in situ hybridization. As previously reported, we also found that serum derived-extracellular vesicles (EVs) from BC affected patients contain HPV DNA. Interestingly, in one TNBC patient, the same HPV DNA type was detected in the serum-derived EVs, cervical and BC tissue samples. Then, we report that HPV DNA can be transferred by EVs to recipient BC stromal cells that show an activated phenotype (e.g., CD44, IL6 expression) and an enhanced capability to sustain mammospheres (MS) formation. These data suggest that HPV DNA vehiculated by EVs is a potential trigger for BC niche aggressiveness.

A causal link between Human Papillomavirus (HPV) and breast cancer (BC) remains controversial. In spite of this, the observation that HPV DNA is over-represented in the Triple Negative (TN) BC has been reported. Here we remark the high prevalence of HPV DNA (44.4%) in aggressive BC subtypes (TN and HER2+) in a population of 273 Italian women and we convey the presence of HPV DNA in the epithelial and stromal compartments by in situ hybridization. As previously reported, we also found that serum derived-extracellular vesicles (EVs) from BC affected patients contain HPV DNA. Interestingly, in one TNBC patient, the same HPV DNA type was detected in the serum-derived EVs, cervical and BC tissue samples. Then, we report that HPV DNA can be transferred by EVs to recipient BC stromal cells that show an activated phenotype (e.g., CD44, IL6 expression) and an enhanced capability to sustain mammospheres (MS) formation. These data suggest that HPV DNA vehiculated by EVs is a potential trigger for BC niche aggressiveness. [ABSTRACT FROM AUTHOR]

Human papillomavirus (HPV) circulating tumor DNA (HPV ctDNA) was proposed as a biomarker for the detection and disease monitoring of HPV-related cancers. One hundred eighty plasma samples obtained from women diagnosed with HPV16-positive cervical cancer (CC) ( n  = 100), HPV16-positive premalignant lesions (cervical intraepithelial neoplasia grade 3 [CIN3]) ( n  = 20), and HPV DNA-negative controls ( n  = 60) were randomly selected from the archives for evaluating the performance of a bead-based HPV genotyping assay (E7 type-specific multiplex genotyping assay [E7-MPG]) in detecting HPV16 ctDNA. The performance of the E7-MPG was compared with those of DNA detection by droplet digital PCR (ddPCR) and detection of HPV16 E6 antibodies evaluated in an independent study. Internal controls to assess DNA quality were included in the molecular assays, i.e., beta-globin and ESR1, respectively. The sensitivity and specificity of E7-MPG and/or E6 antibodies to detect HPV16-positive CCs were evaluated. HPV16 ctDNA was detected using the E7-MPG in 42.3% of all plasma samples and in 74.7% of plasma samples from HPV16-positive CC cases. The validation of E7-MPG data by ddPCR showed that the sensitivity of the E7-MPG test for HPV16-positive CC detection was higher than that of ddPCR (74.7% versus 63.1%; P  < 0.001). When both HPV16 ctDNA and E6 antibodies were considered, the sensitivity for HPV16-positive CC detection increased from 74.7% to 86.1%, while the specificity was unchanged at 97.8%. The performance of E7-MPG for the detection of HPV16 ctDNA appears to be at least as sensitive as that of ddPCR, offering an additional tool for ctDNA detection of HPV16-positive CC. The use of an additional blood marker of HPV infection, such as E6 antibodies, further improved the detection of CC. IMPORTANCE The validity of HPV ctDNA as a marker of HPV-driven cancers has been previously reported. Herein we validated an alternative to ddPCR for HPV16 ctDNA detection, using a bead-based HPV genotyping assay that offers the potential advantage of reducing the cost of clinical management due to the multiplex capability of the test, thus facilitating its use in clinical settings. In addition, we analyzed HPV ctDNA in the context of E6 antibodies as an additional HPV marker. The HPV16 ctDNA biomarker appeared to be highly specific and, to a lesser extent, sensitive for the detection of CC, mainly indicated for those at an advanced tumor stage. In this proof-of-principle study, E6 antibodies were mainly detected in early tumor stages of CC, while HPV ctDNA was mainly positive at advanced tumor stages.

Human papillomaviruses (HPVs) are associated with invasive malignancies, including almost 100% of cervical cancers (CECs), and 35–70% of oropharyngeal cancers (OPCs). HPV infection leads to clinical implications in related tumors by determining better prognosis and predicting treatment response, especially in OPC. Currently, specific and minimally invasive tests allow for detecting HPV-related cancer at an early phase, informing more appropriately therapeutical decisions, and allowing for timely disease monitoring. A blood-based biomarker detectable in liquid biopsy represents an ideal candidate, and the use of circulating HPV DNA (ct-DNA) itself could offer the highest specificity for such a scope. Circulating HPV DNA is detectable in the greatest part of patients affected by HPV-related cancers, and studies have demonstrated its potential usefulness for CEC and OPC clinical management. Unfortunately, when using conventional polymerase chain reaction (PCR), the detection rate of serum HPV DNA is low. Innovative techniques such as droplet-based digital PCR and next generation sequencing are becoming increasingly available for the purpose of boosting HPV ct-DNA detection rate. We herein review and critically discuss the most recent and representative literature, concerning the role of HPV ctDNA in OPC and CEC in the light of new technologies that could improve the potential of this biomarker in fulfilling many of the unmet needs in the clinical management of OPC and CEC patients. [ABSTRACT FROM AUTHOR]

Objective: In our review article we focused on the circulating HPV DNA and its potential role in the pathogenesis of cervical cancer and in the evaluation of patients´ prognosis with cervical cancer Design: The article is a systematic review study analyzing available scientific articles focused on the circulating HPV DNA., Setting: Clinic of Obstetrics and Gynecology, Jesenius faculty of Medicine in Martin, Comenius University in Bratislava, Slovakia., Methods: In our study we searched the medical database PubMed with the key words: circulating HPV DNA, cervical cancer, cervical precanceroses. The core of our work is focused on the scientific articles published in English language since year 1995., Results: We identified 13 studies in PubMed database analyzing the circulating HPV DNA in the process of cervical carcinogenesis. It is clear from the results that circulating HPV DNA is a significant prognostic marker of cervical malignant diseases including the early stages., Conclusion: The results focused on circulating HPV DNA show the significance of molecular biology in assessing the prognosis of cervical cancer. This idea has to be supported by further relevant studies. The uniformity of studies and use of the most sophisticated methods could help to answer the question about the real role of circulating HPV DNA in the process of cervical carcinogenesis and disease progression.

Specific human papillomavirus genotypes are associated with most ano-genital carcinomas and a large subset of oro-pharyngeal carcinomas. Human papillomavirus DNA is thus a tumour marker that can be detected in the blood of patients for clinical monitoring. However, data concerning circulating human papillomavirus DNA in cervical cancer patients has provided little clinical value, due to insufficient sensitivity of the assays used for the detection of small sized tumours. Here we took advantage of the sensitive droplet digital PCR method to identify circulating human papillomavirus DNA in patients with human papillomavirus-associated carcinomas. A series of 70 serum specimens, taken at the time of diagnosis, between 2002 and 2013, were retrospectively analyzed in patients with human papillomavirus-16 or human papillomavirus-18-associated carcinomas, composed of 47 cases from the uterine cervix, 15 from the anal canal and 8 from the oro-pharynx. As negative controls, 18 serum samples from women with human papillomavirus-16-associated high-grade cervical intraepithelial neoplasia were also analyzed. Serum samples were stored at -80°C (27 cases) or at -20°C (43 cases). DNA was isolated from 200 µl of serum or plasma and droplet digital PCR was performed using human papillomavirus-16 E7 and human papillomavirus-18 E7 specific primers. Circulating human papillomavirus DNA was detected in 61/70 (87%) serum samples from patients with carcinoma and in no serum from patients with cervical intraepithelial neoplasia. The positivity rate increased to 93% when using only serum stored at -80°C. Importantly, the two patients with microinvasive carcinomas in this series were positive. Quantitative evaluation showed that circulating viral DNA levels in cervical cancer patients were related to the clinical stage and tumour size, ranging from 55 ± 85 copies/ml (stage I) to 1774 ± 3676 copies/ml (stage IV). Circulating human papillomavirus DNA is present in patients with human papillomavirus-associated invasive cancers even at sub-clinical stages and its level is related to tumour dynamics. Droplet digital PCR is a promising method for circulating human papillomavirus DNA detection and quantification. No positivity was found in patients with human papillomavirus-associated high grade cervical intraepithelial neoplasia.