Human papillomavirus (HPV) has been recognized as responsible of numerous neoplastic and preneoplastic disease in different anatomical district. Many researches have been conducted on these topic in these last few years, primarily in the field of gynecology but also in proctology, otorhinolaringoiatry, odontology, head and neck, dermatology and urology. Indeed many papers have been published also on the different diagnostic tools for HPV infection. Also the recent implementation of vaccines can change the history and the epidemiology of HPV related diseases.
Standing these premises the aim of these study is to approach the HPV infection in a multidisciplinary setting, with the objective to verify if a patient with a HPV lesion could present more HPV infections in different anatomical district and to give a comprehensive vision of the HPV disease to clinicians and patients. In particular, HPV genotyping represents an added value to assess whether positivity to HPV DNA in a second district is related to the primary HPV disease or it should be considered a different occurrence. The results of this study can be precious tools for the management of HPV patients in terms of epidemiology, treatment and prevention.
Considering the rising incidence of HPV-related cancer in different districts, these findings will be relevant to clinicians in their clinical knowledge in this area and for the clinical management of a HPV-positive patient.
Moreover, expected results will catalyze the development of new strong collaborations among the units, aiming at developing synergistic outcomes in patient clinical management and opening future prospective for scientific projects.
Indeed, in this study, we aim to implement sensitive and specific molecular methods for HPV-DNA detection in skin lesions, and in the oral and oropharyngeal mucosa taking advantage from the well-consolidated expertise in HPV detection in the genital tract. In previous studies on penile, laryngeal and dermatologic lesions (e.g. actinic keratoses and skin tags), biopsy specimens were mostly tested using L1-PCR, as well.
In another study in the oral cavity (Pierangeli et al , 2016), it has been shown that a brush biopsy performed with the same Cytobrush used by gynecologist, can obtain full transepithelial cells. However, many oral brushes resulted negative for the HLA gene amplification probably because of fragmented DNA, therefore resulting also not amplifiable with the consensus primer MY09/11 widely used in anogenital HPV detection in our Laboratory (Pierangeli et al., 2006, 2008). To overcome this problem, we then used quantitative real-time PCR fluorogenic assays (qPCR) that provide a greater analytical sensitivity without requiring intact genomic DNA. To avoid false negative results, all samples were tested for the quality of extracted DNA, amplifying the housekeeping gene glyceraldehyde-3-phosphate dehydrogenase (GAPDH) (Pierangeli et al , 2016).
Accordingly, HPV type-specific primers and TaqMan probes for the more common low-risk (HPV 6, 11) and high-risk (16, 18, 31, 33, 53, 58) HPV genotypes, were used and copy number measured in samples by mean of HPV plasmids external curves, as described (Pierangeli, 2008).
As detailed in state of the art, in published papers and on the market, there is a high heterogeneity in diagnostic criteria, collection methodology, molecular techniques, specific HPV genotypes detection. Therefore, the primary goal of the Virology Unit will be to validate HPV positivity results of different commercially available assays, in relation to in-house PCR/Real Time PCR assays. As a resulting achievement, testing of HPV DNA and eventually the specific genotype and viral load will be evaluated, together with the different clinicians, as a marker of risk of progression toward HPV-driven high-grade lesions and cancers of the different districts. Data on HPV genotype-specific infection will be evaluated in patients follow-up to clarify whether HPV-positivity in the absence of a clinical evident lesion could be associated to transient infections without oncogenic risk.
As to the clinical point of view we suppose that people who have HPV infection can have multiple anatomical sites of infection. In this optic, and if this idea is supported by data, HPV screening and prevention can change in the next future.Given that the overall purpose of the present Research Project is to design a comprehensive evaluation to give the patient a diagnosis for HPV, the specific aim of the virology Unit will be to set up a same, unique, molecular method to test HPV DNA with high sensitivity and specificity in the different mucosal and cutaneous districts.
In particular, this study will implement non-invasive sample collection methods with a sensitive molecular method, that will probably detect high rates of HPV-DNA. The fact that HPV infection in districts different from the cervical may represent a not uncommon condition represent a challenge to understand which are the factors that would predispose a still normal epithelia to eventually develop a clinically relent HPV-related lesion. As an example, it remains to be clarified whether low viral loads in the absence of a clinically evident lesion could be indicative of transient infections. Conversely, HPV infections at elevated viral loads, such those usually found in HPV16 and 18, could be associated with an increased risk of persistence and progression, to be tested in prospective studies.
In any case, detection of high-risk HPV genotype infections would pose the potential for prevention of developing precancerous lesions. Considering the rising incidence of HPV-related cancers other than cervico-carcinoma, these findings could be relevant to clinicians to enhance their knowledge in this area and for clinical patient management.