Kinetics of the Human Papillomavirus Type 16 E6 Antibody Response Prior to Oropharyngeal Cancer

Abstract

Background: In a European cohort, it was previously reported that 35% of oropharyngeal cancer (OPC) patients were human papillomavirus type-16 (HPV16) seropositive up to 10 years before diagnosis vs 0.6% of cancer-free controls. Here, we describe the kinetics of HPV16-E6 antibodies prior to OPC diagnosis.

Methods: We used annual serial prediagnostic blood samples from the PLCO Cancer Screening Trial. Antibodies to HPV were initially assessed in prediagnostic blood drawn at study enrollment from 198 incident head and neck cancer patients (median years to cancer diagnosis = 6.6) and 924 matched control subjects using multiplex serology, and subsequently in serial samples (median = 5/individual). Available tumor samples were identified and tested for HPV16 RNA to define HPV-driven OPC.

Results: HPV16-E6 antibodies were present at baseline in 42.3% of 52 OPC patients and 0.5% of 924 control subjects. HPV16-E6 antibody levels were highly elevated and stable across serial blood samples for 21 OPC patients who were seropositive at baseline, as well as for one OPC patient who seroconverted closer to diagnosis. All five subjects with HPV16-driven OPC tumors were HPV16-E6-seropositive, and the four subjects with HPV16-negative OPC tumors were seronegative. The estimated 10-year cumulative risk of OPC was 6.2% (95% confidence interval [CI] = 1.8% to 21.5%) for HPV16-E6-seropositive men, 1.3% (95% CI = 0.1% to 15.3%) for HPV16-E6-seropositive women, and 0.04% (95% CI = 0.03% to 0.06%) among HPV16-E6-seronegative individuals.

Conclusions: Forty-two percent of subjects diagnosed with OPC between 1994 and 2009 in a US cohort were HPV16-E6 seropositive, with stable antibody levels during annual follow-up for up to 13 years prior to diagnosis. Tumor analysis indicated that the sensitivity and specificity of HPV16-E6 antibodies were exceptionally high in predicting HPV-driven OPC.

Figure 1.

CONSORT flow diagram. HPV = human papillomavirus; HNSCC = head and neck squamous cell carcinoma. *The inclusion of cancer-free at random assignment excludes rare cancers amongst controls.

Figure 2.

Kinetics of HPV16-E6 in prediagnostic samples from oropharyngeal cancer cases and selected controls. A) This panel shows HPV16-E6 (MFI) values by years from blood draw to diagnosis among (OPC) cases. HPV16-E6 levels remained clearly positive and stable for all OPC cases who were HPV16-E6 seropositive at baseline. One OPC case who was HPV16-E6 seronegative at baseline displayed increasing levels during the first three visits, after which strong and stable levels were established, similar to the other OPC cases that were seropositive at baseline (highlighted in blue). One OPC case had HPV16-E6 levels that fluctuated between positive and negative (highlighted in orange). B) This panel shows HPV16-E6 MFI values by year from time since study enrollment among controls. Of the five HPV16-E6-seropositive controls, three were consistently positive across serial samples; one seroconverted over time (highlighted in purple), and one was HPV16-E6 positive in two of six serial samples (highlighted in green). HPV = human papillomavirus; MFI = median fluorescence intensity; OPC = oropharyngeal cancer.

Figure 3.

HPV16 antibody levels of multiple HPV proteins in serial samples leading up to diagnosis of oropharyngeal cancer (OPC), by HPV16-E6 serostatus in the baseline sample. HPV = human papillomavirus; MFI = median fluorescence intensity; OPC = oropharyngeal cancer.

Figure 4.

Cumulative 10-year risk of oropharyngeal cancer by sex and HPV16-E6 serostatus. Using the weighted Cox model, we estimated the 10-year cumulative risk for oropharyngeal cancer (OPC) overall at 3.4% (95% CI = 0.6% to 20.8%); by sex, the 10-year cumulative risk was 6.2% (95% CI = 1.8% to 21.5%) for HPV16-E6-seropositive males and 1.3% (95% CI = 0.1% to 15.3%) for HPV16-E6-seropositive females. The 10-year cumulative risk of OPC among HPV16-E6-seronegative subjects was extremely low (0.04%, 95% CI = 0.03% to 0.06%). A separate graph of the HPV16-E6-seronegative individuals is nested in the figure in order to better visualize the curve. Please note that the y-axis scale differs in the two graphs. HPV = human papillomavirus.