The immune system has been shown to control HPV infection in some women and not in others leading to pre-cancerous lesions and subsequently cancer. The relatively high regression rate of cervical intraepithelial lesions (CIN) has similarly been attributed to engagement of the immune response directed against neoplastic cells. Recent advances in immune-oncology have shown the dramatic effects of PD-1/PD-L1 inhibitors in epithelial tumors including squamous cell carcinoma and adenocarcinoma, the major cancer subtypes in the female genital tract. Here, we present a novel assay that combines RNA in situ hybridization for HPV E6, E7 mRNA and PD-L1 cell surface staining on squamous cells in liquid-base cervical cytology specimens. This assay could provide actionable data supporting therapeutic options.
ThinPrep liquid based cervical cytology samples were obtained from both normal patients and patients with HSIL cervical cytology. Cells were isolated and hybridized in suspension for HPV E6, E7 mRNA using the HPV OncoTect 3Dx (CE-IVD) kit. Following post-hybridization washes, cells were stained with PD-L1 using the OncoTect iO (CE-IVD) kit then counterstained with cell cycle reagent. Cells were analyzed on a CytoFlex flow cytometer (Beckman Coulter).
PD-L1 expression was low in the normal HPV negative samples and variable in the HSIL samples using a standard 1% cut-off for positivity in squamous cells. Of interest, PD-L1 expression was higher in cells expressing HPV E6, E7 mRNA compared to cells lacking transcriptional activity.
PD-L1 expression is variable in samples with abnormal cervical cytology. The prognostic implications of PD-L1 expression in cervical pre-cancer remains to be determined, however, we present in this study a means for fine quantification of PD-L1 expression in both HPV transcriptionally active and normal cells. This combination disease markers should facilitate further studies of the role in progression or regression of cervical pre-cancer.