STUDIES ON THE ROLE AND ONCOGENIC POTENTIAL OF HUMAN PAPILLOMAVIRUS GENOTYPES 16 AND 18 IN THE CAUSATION OF CARCINOMA OF THE CERVIX UTERI

Abstract

Human papillomaviruses (HPV) have been incriminated as an important co-factor in the causation of cervical cancer. Southern blot analysis of most benign and malignant lesions of the cervix uteri reveals the presence of HPV. HPV 16 is the predominant genotype worldwide and is associated with more than 50% of the malignant lesions, while other HPVs such as 18, 31 and 33 vary geographically. An aetiological role has been ascribed to HPV, supported by the epidemiologic implication of an infectious agent in cervical carcinogenesis and the properties of the transforming papillomavirus proteins. Papanicolaou smears from putatively 'healthy' women and 'high-risk' prostitutes permitted the screening for HPV DNA by filter in situ hybridization (FISH) and the polymerase chain reaction (PCR). The presence of HPV in genital lesions and lymph node material was determined by Southern blot and PCR. To identify possible HPV transmission pathways, genital biopsies of virginal versus sexually-active individuals, and samples from extra-genital sites were analysed, (1) FISH, Southern blot, and PCR were applied for the sensitivity-standardized detection of HPV DNA, and the limits were found to be 1 picogram, 1 copy per cell, and 10 molecules, respectively. Therefore, PCR is by far the most sensitive method. The kinetics and conditions for successful PCR amplification of HPV were delineated. (2) In the analysis by FISH of cervical smears from 740 and 225 women attending polyclinics, and 130 prostitutes, 4.1%, 6.2% and 6.9% were HPV-positive, with HPV 16 and 31 predominating. Among the 740 and 225 women, no significant differences between HPV positivity rates in Chinese and in Malays were found, even though Chinese have a higher cervical cancer incidence. This finding is noteworthy, and indicates that other factors in the Chinese population are contributory. The cervical smears of 52 and 50 randomly-sampled women analysed by PCR, revealed that 61% and 14% respectively were HPV 16- and 18-positive. Since the majority of the population is latently infected by genital HPVs, other factors are ultimately rate-limiting in cervical oncogenes is. Furthermore, subclinical HPV infection should be treated only in combination with HPV-associated disease. (3) Of 57 cervical neoplastic lesions, HPV was detected in 74% by Southern blot, the predominant types being HPV 16 and 18. After PCR with HPV 11, 16 and 18 primers, all samples were HPV-positive. Numerous negative controls were included to exclude artifacts, and the c-Ha-ras-1 proto-oncogene was co-amplified as an authenticity control. A sufficient diagnostic sensitivity has detected HPV DNA in most, if not all cervical neoplasia, thereby confirming the strong association between genital HPVs and cervical neoplasia. (4) Of 75 subclinical vulval HPV lesions, 76% were positive for HPV 11 and/or 16 by PCR, whereas only 1 was HPV-positive by Southern blot. HPV 16 was detected by PCR in 37.3%. 4 of 6 virginal women were HPV-positive. The degree of epithelial changes on colposcopy correlated well with HPV positivity. Such lesions constitute a reservoir for repeated cervical HPV infections, and a contamination for cervical HPV DNA detection by techniques. HPV infection in virgins suggests virus may also be spread by non-sexual modes. source of sensitive that the (5) Of 25 consorts of women with genital HPV infection or cervical intraepithelial neoplasia, colposcopy revealed subclinical HPV infection of the male lower genital tract in 22 cases. Although Southern blot detected HPV in only 1 case, PCR revealed HPV 11 and/or 16 in 80%. The prevalence of HPV 16 was high ( 40%). The high genital HPV carriage rate reiterates the sexual route as a major HPV infectious pathway, and emphasizes the detection sensitivity of PGR and the limitations of Southern blot. ( 6) Southern blot and PCR revealed HPV 11, and interestingly the oncogenic HPV 18 in a juvenile laryngeal papillomatosis biopsy. None of the skin and oral samples of 10 normal healthy individuals was positive for HPV 11 and HPV 16, emphasizing the greater predilection of these genital HPV types for genital sites. (7) Although PCR is by far the most sensitive technique, it is so sensitive that true association of oncogenic HPVs with cervical neoplasia cannot always be distinguished from contamination by HPV in surrounding tissues. In addition, miniscule quantities of HPV in a tissue may not be predictive of eventual disease.