Expression and immunoreactivity of high-risk human papillomavirus E4 prtein

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Certain human papillomavirus types named high-risk types are thought to cause cervical cancer and cervical intraepithelial neoplasia (CIN). It has been recently known that E4 proteins of these types are abundantly expressed in CIN epithelium. However, such expression has not been investigated extensively in clinical samples. This study is to examine the expression pattern using E4 antibodies for specific HPV types.We obtained seven different polyclonal antibodies against E4 proteins of HPV types 11, 16, 18, 31, 33, 56, which were produced from immunized rabbits and purified. lb start this experiment, we tried to do HPV genotyping in cytological samples using three different HPV genotyping methods in women who were abnormal in cytology. These methods are now commercially available in Japan. We found that different assays have shown discordant results in some of the samples examined. Discordant results were more frequently observed in the cases infected with different HPV types. This type of multiple HPV type infection were more common in women having abnormal cytology as we have previously expected. Now we are going to analyze more detail in these eases showing discordant results. This is a crucial step for further experiment for E4 study.E4 expression has been confirmed on epithelium from middle to superficial layers in most of CIN1 and CIN2, 3 lesions in which HPV types were determined. Abundant expression was seen in some cases. However, some immune cells were also stained by these antibodies. We have to figure out the positive reaction to immune cell is merely due to cross-reaction or due to presence of E4 in these immune cells. Furthermore, we have to test type-specificity of these antibodies in immune-staining. E4 protein immunostaining method may be useful for diagnostic purpose of high-risk HPV infection, and furthermore E4 might be a good target for therapeutic HPV vaccine.