Makioka Daiki, Inada Mikio, Awano Masayuki, Saito Ema, Shinoda Takuya, Abe Satoko, Yoshimura Teruki, Müller Martin, Sasagawa Toshiyuki, Ito Etsuro
Department of Biology, Waseda University, Shinjuku, Tokyo 162-8480, Japan.
School of Pharmaceutical Sciences, Health Sciences University of Hokkaido, Tobetsu 061-0293, Hokkaido, Japan.
Microorganisms. 2024 Jun 14;12(6):1205. doi: 10.3390/microorganisms12061205.
We present the validity of using an ultrasensitive enzyme-linked immunosorbent assay (ELISA) for quantifying high-risk human papillomavirus (HPV) 16 E7 oncoproteins in urine specimens as a noninvasive method of analyzing the oncogenic activity of HPV. Some reports claim that the oncogenic activity of HPV is a more relevant clinical indicator than the presence of HPV DNA for estimating malignant potential. In the present study, urine containing HPV16 and related types were selected by uniplex E6/E7 polymerase chain reaction and classified according to the pathologic diagnosis of cervical intraepithelial neoplasia (CIN) in cervical biopsy specimens. Our ultrasensitive ELISA was able to detect attomole levels of HPV16 E7 oncoproteins, and it detected HPV16-positive SiHa cells at >500 cells/mL without detecting HPV18-positive cells. Our ELISA results showed E7 oncoproteins in 80% (4/5) of urine specimens from women with HPV16-positive CIN1, 71% (5/7) of urine specimens from CIN2 patients, and 38% (3/8) of urine specimens from CIN3 patients. Some urine specimens with undetectable E7 oncoproteins were thought to be negative for live HPV 16-positive cells or in an inactivated state of infection. These results provide the basis for assessing oncogenic activity by quantifying E7 oncoproteins in patient urine.
我们展示了使用超灵敏酶联免疫吸附测定（ELISA）对尿液标本中的高危型人乳头瘤病毒（HPV）16 E7癌蛋白进行定量分析作为一种分析HPV致癌活性的非侵入性方法的有效性。一些报告称，对于估计恶性潜能而言，HPV的致癌活性是比HPV DNA的存在更相关的临床指标。在本研究中，通过单重E6/E7聚合酶链反应选择含有HPV16及相关类型的尿液，并根据宫颈活检标本中宫颈上皮内瘤变（CIN）的病理诊断进行分类。我们的超灵敏ELISA能够检测到阿托摩尔水平的HPV16 E7癌蛋白，并且在>500个细胞/mL时检测到HPV16阳性的SiHa细胞，而未检测到HPV18阳性细胞。我们的ELISA结果显示，HPV16阳性CIN1女性的尿液标本中有80%（4/5）检测到E7癌蛋白，CIN2患者的尿液标本中有71%（5/7）检测到，CIN3患者的尿液标本中有38%（3/8）检测到。一些未检测到E7癌蛋白的尿液标本被认为HPV 16阳性活细胞为阴性或处于感染的失活状态。这些结果为通过定量患者尿液中的E7癌蛋白来评估致癌活性提供了依据。
