Human papillomaviruses are a family of DNA viruses that infect the epithelium leading to the formation of lesions with the ability to progress into carcinoma in many forms. Research into causality of head and neck squamous cell carcinoma (HNSCC) have found a link to HPV infections affiliated with better survival than tobacco associated HNSCC. In addition, advances in immuno-oncology have brought immunotherapy to the forefront of cancer treatment across cancer types. Anti-PD-1 and anti-PD-L1 therapies are being explored for their efficacy in treatment of head and neck cancers. Here we present data that demonstrates a combined diagnostic approach to quantify both HPV E6,E7 and PD-L1, two markers important in the management of head and neck cancer.
Samples were collected from Institut Gustave Roussy patients with lesions of the oral pharynx. Swabs were collected and placed into a collection vial with a proprietary fixation/permeabilization solution (IncellCollect, IncellDx, Inc.) and shipped overnight on cold packs for processing. Upon receipt, samples were passed through a 35 uM filter to remove aggregates. Cells underwent RNA in situ hybridization with E6, E7 mRNA probes (HPV OncoTect) , were labeled with PD-L1 Ab (clone 28-8), and then stained with a cell cycle dye identify single nucleated cells, and to analyze cell cycle prior to analysis on the flow cytometer (CytoFLEX, Beckman Coulter, Inc).
We analyzed samples from 20 patients with oral cancer with the combined E6, E7 mRNA/PD-L1 protein assay by flow cytometry. The percentage of cells expressing E6,E7 mRNA and the percentage of cells expressing PD-L1 was calculated. E6,E7 result was compared to a second HPV test method as well as biopsy result. The study is ongoing and data will be presented at the meeting.
We report a novel flow cytometric assay to quantify both HPV E6, E7 mRNA and PD-L1 simultaneously in single cells from head and neck squamous cell carcinoma. The ability to characterize both markers in one test can provide clinicians with insight into the prognosis of the patient based on their HPV mRNA status and PD-L1 expression.