Human Papillomavirus 16 Load and E2/E6 Ratio in HPV16-Positive Women: Biomarkers for Cervical Intraepithelial Neoplasia ≥2 in a Liquid-Based Cytology Setting?

Abstract

This retrospective case-control study assessed human papillomavirus 16 (HPV16) viral load and E2/E6 ratio as risk markers for cervical intraepithelial neoplasia (CIN) ≥2 lesions in HPV16-positive women in a routine liquid-based cytology setting. Triplex quantitative PCR for HPV16 E6, E2, and β-globin was done to determine the HPV16 load and the E2/E6 ratio, as a surrogate marker for integration, for women with a negative histologic endpoint (200 controls: 83 normal histology and 117 CIN1) and women with a ≥CIN2 endpoint (180 cases: 41 CIN2, 122 CIN3, and 17 invasive carcinoma). Our analysis showed a significantly higher HPV16 load in the case group, which was completely attributable to the high viral load of samples with invasive carcinoma as histologic endpoint. There was no significant difference in viral load between the other histologic groups. The E2/E6 ratio proved to be lower for the cases. However, the E2/E6 ratio indicated the presence of HPV integration in a considerable amount of control samples (44.3%), which suggests that HPV integration occurs early in the development of cancer and undermines the clinical value of viral integration. Overall, the intrinsic heterogeneous nature of the cervical cytology samples caused a substantial overlap of the HPV16 load and the E2/E6 ratio between controls and cases, which precludes the determination of cutoff values for risk prediction and hampers the clinical applicability in a cervical screening setting. (Cancer Epidemiol Biomarkers Prev 2009;18(11):2992-9)

Pre-analytical and post-analytical evaluation in the era of molecular diagnosis of sexually transmitted diseases: cellularity control and internal control

“…Al contrario, la maggior parte dei NAATs per la ricerca di HPV presenta il CC per la valutazione qualitativa della cell ularità ma non presenta l'IC per la valutazione di inibitori parziali (29), ad eccezione di alcuni kit in RT-PCR (Dx HR-HPV Auto Assay, Biorad; RealLine HPV High RiskGentotype Fla-Format, Bioron) e microarray (PAPILLOCHECK HPV Screening Test, Greiner) che presentano entrambi i controlli. Praticamente inesistenti sono comunque lavori che riportano dei cut-off di cellularità che definiscano l'idoneità del prelievo e spesso i valori di cellularità riportati non sono espressi come numero assoluto di cellule, ma come unità di misura relative alla metodica usata (10,12,14,16,39). Le stesse linee guida dei requisiti di un test HPV per lo screening primario del cervicocarcinoma (24) non menzionano niente riguardo l'idoneità del prelievo e il test Hybrid Capture II (Qiagen, Germania), con cui sono stati condotti tutti gli studi che hanno dimostrato l'importanza dell'inserimento della ricerca dell'HPV nello screening, non ha un CC (31).…”

“…Diversi sono invece i lavori in letteratura che valutano l'influenza dell'arricchimento del pellet mediante centrifugazione dei tamponi cervicali per la citologia su strato sottile (ThinPrep, SurePath) per aumentare la sensibilità del test molecolare rispetto all'estrazione diretta del DNA (5,12,35), mentre praticamente inesistenti sono lavori che valutino l'idoneità del prelievo eseguito con i tamponi E-Swab (Copan, Italia), i più utilizzati attualmente nelle microbiologie perché adatti all'esecuzione automatizzata degli esami colturali. Inesistenti sono anche lavori condotti con metodiche NAATs che quantificano la carica definendo dei cut-off per il trattamento dei patogeni commensali opportunisti (Ureaplasma spp.…”

Human papillomavirus viral load in cervical intraepithelial neoplasia as a prognostic factor in a Mexican population

“…HR-HPV types (including are associated with more than 99% of all cervical cancers as well as with high-grade CIN (CIN2-3) and abnormal Papanicolaou test (Pap) results. Low-risk HPV types are responsible for additional abnormal Pap test results and almost all cases of genital warts (Winer et al, 2003;Boulet et al, 2009;Hesselink et al, 2009;Seaman et al, 2010).…”

“…Recent studies have described viral load as an effective diagnostic and prognostic marker for CIN (Castle et al, 2005;Moberg et al, 2005;Lillo and Uberti-Foppa, 2006;Fiander et al, 2007;Boulet et al, 2009;Hesselink et al, 2009). On the spectrum of cervical disease, CIN2 and 3 are considered cervical cancer precursors.…”

“…However, the use of HPV testing combined with cervical cytology to detect cervical cancer or its precursors is still controversial and insufficient (Cain and Howett, 2000;Fiander et al, 2007;Boulet et al, 2009;Hesselink et al, 2009;Grce et al, 2010). Given the positive relationship between viral load and the likelihood of persistent HPV infection and the strong association between the latter and the risk of cervi-cal neoplasia, a single measurement of viral load in cervical specimens may be a suitable biomarker (Schlecht et al, 2003;Boulet et al, 2009).…”