>> P-425: Generation of Pan-HPV E4 antibodies as biomarkers of active HPV infections
19:00 PM - 19:00 PM
1MRC National Institute for Medical Research, Mill Hill, London, UK; 2GlaxoSmithKline Biologicals, Rixensart, Belgium.
Infection by high-risk HPV types causes cervical lesions that can progress to cancer, with HPV DNA being found in nearly all cases of cervical cancer. Detection of HPV DNA alone cannot however differentiate active infections from passive contamination or latency. Moreover, it is not possible to determine the causative HPV type in cases of multiple infections. Our recent work has focused on identifying biomarkers of active HPV infection. HPV E4 is an abundant viral protein with type-specific sequences. E4 is expressed in differentiated epithelial cells according to lesion grade, and is a good candidate as a disease-staging biomarker of active type-specific HPV infection, especially for CIN1 and CIN2 lesions. To examine the utility of E4 detection as a diagnostic tool, type specific anti-E4 antibodies (Abs) were generated against the E4 proteins of HPV-16, HPV-18 and HPV-58. These are the first type-specific reagents that have been described. Here we extend this work and describe the generation and testing of cross-reactive HPV E4 or pan-HPV E4 Ab. Rabbits were injected with several E4 proteins fused to maltose-binding protein, and polyclonal Abs were evaluated for HPV E4 cross-reactivity by immunohistochemistry. Qualification experiments have shown that these pan HPV E4 Abs can react with at least eight types of high risk HPV E4 proteins. The cross-reactivity and sensitivity of the Ab were confirmed in CIN lesions associated with HPV-16, 18, 31, 33, 35, 51, 52 and 58. These pan HPV E4 Abs, when combined with whole tissue section HPV DNA typing, could be used to identify regions of active infection by most high-risk HPV types in lesions with single infections. When combined with laser capture microdissection (LCM)/PCR or type-specific E4 reagents, the approach could identify active infection and confirm causative HPV types in lesional areas of multiple infections. In addition, these Abs could be used to detect HPV E4 expression in borderline or pre-CIN lesions, and therefore, could be used as an indicator for LCM sample collection.