BIOMARKER DISCOVERY FOR HPV-ASSOCIATED CERVICAL AND OROPHARYNGEAL CANCER

Despite the efficacy of vaccination and cervical cancer screening, human papillomavirus (HPV) continues to be a major public health epidemic and the causative agent of cervical cancer and a subset of oropharyngeal cancer (HPVOPCs). In 2012, over 500,000 new cases of HPV-associated cervical cancer were diagnosed worldwide. To develop biomarkers for early disease detection, we analyzed the systemic IgG antibody responses to HPV antigens using a protein microarray platform. Custom protein microarrays displaying 98 proteins of two low-risk HPV types (HPV 6,11) and ten onocgenic high-risk HPV types (HPV 16, 18, 31, 33, 35, 39, 45, 51, 52, and 58) were generated. Protein expression was verified by GST detection for 96 of the 98 antigens. IgG antibody responses were evaluated in serum from patients with invasive cervical cancer (n=80), CIN II/III (n=60), CIN 0/I (n=60), HPVOPC (n=64), and healthy controls (n=30). Each slide was scanned for signal intensity of individual spots. A qualititative review of the array images was performed to identify positive responses by ranking the signal intensity around each spot on the microarray. A total of 63,504 array spot images were visually inspected. IgG antibodies to antigens of one low-risk and nine high-risk HPV types were detected in the serum of patients with CIN II/III, with 45% sensitivity and 89% specificity. Specific antibody response to one or more early antigens was present in 31% of invasive cervical cancer sera and 43% of CIN II/III sera. In addition to the known immunogenicity of HPV16 E2, E6, and E7 in HPVOPC, 14.5% of HPVOPC cases had antibody responses specifically to non-HPV 16 antigens. Serum IgG antibody detection against HPV antigens may provide potential biomarkers for HPV-associated cancers. The use of protein microarrays facilitates rapid detection of these antibodies in sera against a broad spectrum of HPV types and antigens.