Some head and neck squamous cell carcinomas (HNSCC) are causally associated with high-risk human papillomavirus (HR-HPV) genotypes 16 and 18. Current HPV detection methods include surrogate p16 immunohistochemical (IHC p16) staining, in situ hybridization, and nucleic acid amplification assays. Arbor Vita Corp has developed an OncoE6™ Oral Test to detect the presence of HPV 16 and 18 E6 oncoproteins in samples.The study objective was to determine if the OncoE6™ Oral Test could detect HPV 16/18 E6 oncoproteins in cervical lymph node fine needle aspirates (FNAs), base of tongue (BOT) swabs, and saliva samples from patients with HNSCC who had HR-HPV E6/E7 mRNA in FNAs and/or IHC p16 positive tumors.
Clinic patients (n=68; 61 males and mean age 59.8 years), with confirmed or suspected HNSCC consented to collection of FNA, BOT swab, and saliva samples. FNAs were collected and preserved in 4 ml of PreservCyt ThinPrep fluid (TP; Hologic). A BOT swab was collected into a 15 ml tube. Saliva (1 ml) was collected into an OMNIgene Discover saliva collection kit (OM-505; DNA Genotek) for mRNA testing and another 1 ml was collected into a 15 ml tube for OncoE6™ testing. In the laboratory, TP was added to the FNA (1 ml) and BOT swab (5 ml), additional OMNIgene preservative (4 ml) was added to the Genotek saliva sample. The Aptima HPV (AHPV) assay was performed on 1 ml of sample added into 2.9 ml Aptima specimen transport media. For the OncoE6™ test, 1 ml of each sample type was used; total test time was 2.5 h.
Primary tumours were p16 positive in 58 of 68 patients. Overall agreement of the OncoE6™ Oral Test on FNA with p16 staining of tumors was 73.5% (50/68, k=0.40). IHC p16 staining of tumors and AHPV testing of FNA had an overall agreement of 83.8% (57/68, k=0.56). The AHPV assay detected E6/E7 HPV mRNA in 47 FNA samples, of which 39 were positive by OncoE6™. OncoE6™ and AHPV testing of FNAs showed 86.8% (59/68, k=0.72) overall agreement. The sensitivity of the OncoE6™ test for detecting HPV infected patients who were both p16 and AHPV positive was 83.0% (39/47); the specificity was 95.2% (20/21). AHPV mRNA was positive in 24 saliva samples, 3 of which were positive by OncoE6™. Thirteen BOT swabs were positive by AHPV, while none were positive by OncoE6™.
The OncoE6™ Oral Test was easy to perform and detected 83.0% of patients who had both p16 IHC positive tumors and AHPV E6/E7 mRNA positive FNA. The OncoE6™ Oral Test is the first commercial test for the detection of E6 oncoproteins in FNA and its strong agreement of 86.8% (k=0.72) with our reference standard should make it useful as a diagnostic test for HPV induced HNSCC.