Clinical validation of full HR-HPV genotyping HPV Selfy assay according to the international guidelines for HPV test requirements for cervical cancer screening on clinician-collected and self-collected samples

Abstract

Background According to international guidelines, Human Papillomavirus (HPV) DNA tests represent a valid alternative to Pap Test for primary cervical cancer screening, provided that they guarantee balanced clinical sensitivity and specificity for cervical intraepithelial neoplasia grade 2 or more (CIN2+) lesions. The study aimed to assess whether HPV Selfy (Ulisse BioMed – Trieste, Italy), a full-genotyping HPV DNA test that detects and differentiates 14 high-risk HPV (HR-HPV) types, meets the criteria for primary cervical cancer screening described in the international guidelines, on clinician-collected as well as on self-collected samples. Methods For each participant woman, consecutively referring to Azienda Sanitaria Universitaria Giuliano Isontina (Trieste, Italy) and CRO—National Cancer Institute (Aviano, Italy) for the cervical cancer screening program, the following samples were tested: (a) a clinician-collected cervical specimen, analyzed with the reference test (Hybrid Capture®2 test, HC2) and HPV Selfy; and (b) a self-collected vaginal sample, analyzed with HPV Selfy. Enrolled women were also asked to fulfill a questionnaire about self-sampling acceptability. As required by guidelines, a non-inferiority test was conducted to compare the clinical performance of the test under evaluation with its reference test. Results HPV Selfy clinical sensitivity and specificity resulted non-inferior to those of HC2. By analysis of a total of 889 cervical liquid-based cytology samples from a screening population, of which 98 were from women with CIN2+, HPV Selfy showed relative sensitivity and specificity for CIN2+ of 0.98 and 1.00 respectively (non-inferiority score test: P = 0.01747 and P = 0.00414, respectively); the test reached adequate intra- and inter-laboratory reproducibility. Moreover, we demonstrated that the performance of HPV Selfy on self-collected vaginal samples was non-inferior to the performance obtained on clinician-collected cervical specimen (0.92 relative sensitivity and 0.97 relative specificity). Finally, through HPV Selfy genotyping, we were able to describe HPV types prevalence in the study population. Conclusions HPV Selfy fulfills all the requirements of the international Meijer’s guidelines and has been clinically validated for primary cervical cancer screening purposes. Moreover, HPV Selfy has also been validated for self-sampling according to VALHUDES guidelines. Therefore, at date, HPV Selfy is the only full-genotyping test validated both for screening purposes and for self-sampling. Trial registration ASUGI Trieste n. 16008/2018; CRO Aviano n.17149/2018

Can HPV Selfy be considered as a clinically validated HPV test for use in cervical cancer screening?

“…This may rise suspicion of a certain degree of histological over-classification. Nonetheless, we verified the data matrices in Table 2 in Avian et al [ 1 ] and confirm the correctness of the non-inferiority statistics (Table 1 ).

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“…In primary cervical cancer screening, it is crucial to use only hrHPV tests that are clinically validated according to international guidelines in order to reduce the risks of missing relevant disease and of over-treatment. In the recent J Transl Med [ 1 ] paper, Avian et al concluded that the HPV Selfy assay (Ulisse BioMed, Trieste, Italy) fulfils international validation criteria for hrHPV testing on clinician-collected cervical samples (Meijer guidelines) [ 2 ] as well as by extension on self-collected vaginal samples (VALHUDES) [ 3 ]. Our perception is that the study by Avian et al has certain limitations that are worthy of consideration and which may call into question certain conclusions.…”

“…Validation requires an appropriately composed study population comprising a sufficient number of diseased subjects, derived from a continuous screening population or from a clearly described selection of CIN2+ cases and < CIN2 controls [ 4 ]. Avian et al compiled cervical specimens for testing with HC2 (standard comparator test) and with the new HPV Selfy (index test) [ 1 ], but it remains unclear how the study population was composed. With 98 CIN2+ and 791 ≤ CIN1 subjects it was obviously not a continuous screening population, so more granularity on this would have been welcome.…”

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Fig. 1 Sensitivity of the HC2 assay (standard comparator used in validation of hrHPV assays) in studies included in the 2020 list of HPV assays validated for cervical screening [ 5 ] that applied the Meier [ 2 ] or VALGENT [ 4 ] validation protocols (on top) or included in the study of Avian et al [ 1 ] (at the bottom)

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“… In italics: non-inferiority statistic reported by Avian et al [ 1 ] which was due to erroneous switching the values 37 and 16. In fact this reported statistic reflects that Selfy on clin samples is not inferior to SS samples …”