Manu, G (2008) A Study of Viral and Immunological Markers of Human Papillomavirus (HPV) related Progression of Cervical Neoplasia. Doctoral thesis, The Tamilnadu Dr. M.G.R. Medical University, Chennai.
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Abstract
Cervical cancer is the second most common cancer among women worldwide with 493,000 new cases reported and 274,000 deaths in 2002. Eighty three percent of cervical cancer cases occur in the developing countries. In India, every year 132,082 new cases of cervical cancer cases are diagnosed and 74,118 women die from the disease. Thus, India has one-fourth of the global burden of cervical cancer. Human papillomavirus (HPV) infection is an identified cause for the development of cervical cancer. Epidemiological studies have shown that infections with certain HPV types (genotypes) are responsible for most cases of cervical cancer. This study attempted to evaluate the role of certain HPV markers with prognosis in women with cervical neoplasia. Three groups of women were studied. They were women with cervical neoplasia (Study group I), women without cervical neoplasia (Study group II) and healthy women from the community (Study group III). The major results obtained from this study are • HPV DNA was detected in 91.3% of women with cervical neoplasia using MY09/11 primers and in 93.3% of women using PGMY09/11 primers. • HPV genotypes could be ascertained in 91.2% of women with cervical neoplasia using PCR-RFLP. This technique was found to be a reliable and affordable tool for HPV genotyping. • The HPV types detected by PCR-RFLP in decreasing order of frequency were HPV 16 (66.4%), HPV 18 (13.9%), HPV 45 (4.3%), HPV 31 (2.9%), HPV 73 (1.5%) and HPV 59, 68 (1% each). • Line blot assay was able to type all samples (100%) that were amplified by PGMY09/11 primers. • Line blot assay was able to resolve all RFLP untypeable strains. It also picked up multiple infections (2 HPV genotypes) in 6.4% of cases studied. • The major high risk types detected in this study were HPV 16 (63.1%), 18 (13.4%), 45 (4.7%), 31 (3.3%), 52 (2.7%), 59 (2%), and 51, 58, 73, 84 (1.34% each). • Both MY09/11 and PGMY09/11 primers showed 100% specificity in amplifying HPVs. • Real time PCR was an efficient method of determining HPV 16 and 18 viral loads in cervical tissue. HPV viral loads (normalized against ERV3) were not found to correlate with clinical disease stage. • Both sets of primers in the HPV 16 and HPV 18 real time PCR showed 100% pecificity. • HPV DNA was detectable in 2 (2.1%) plasma samples of women who had HPV 16 DNA in their cervical tract and none of the 20 women who had HPV 18 in their cervical tract using conventional MY09/11 primers. • Real time PCR was able to detect HPV 16 DNA in plasma of 56.4% women who harboured HPV 16 in their cervix. HPV 16 plasma viremia detection rate increased with advancing disease stage suggesting poor disease prognosis. • There was no correlation between plasma HPV 16 viral load and corresponding HPV 16 virus load in tissue. • HPV 18 plasma viremia was detectable in 20% of HPV 18 positive women. However, HPV 18 plasma viral loads did not show any association with clinical disease stage. • HPV 16 E6/E7 mRNA transcripts were detectable in 89.3% of HPV 16 positive samples using nested PCR. The E7 transcript (E6*I, E6*II) was the major transcript detected in cervical tissue samples from women with neoplasia in this study. • There was no correlation between detection of HPV 16 E6/E7 mRNA transcripts using either conventional nested PCR or real time PCR and advancing cervical disease. • HPV 16 mRNA transcripts were detected in 86% of HPV 16 positive women using real time PCR. The median transcript level showed a trend to increase with advancing disease stage (not statistically significant). This indicates that oncogene mRNA levels may play an important role in progression of cervical disease. • There was no correlation between the HPV 16 mRNA transcript level (copies/20ng of cDNA) in cervical tissue and HPV 16 DNA level (copies/5000 cells) in the corresponding tissue sample. • mRNA transcripts were not detected in PBMC by both conventional nested PCR and real time PCR, suggesting that mRNA detection in PBMC is not a good marker of disease prognosis. • In terms of individual responses, reactivity to HPV 16 E7 peptides as studied by ELISPOT assay for interferon gamma (IFN γ) was low. This suggests that T cell responses are probably weak or absent in Indian patients or directed against other HPV proteins. The proportion of individuals showing T cell response as measured by IFN γ release to HPV 16 E7 peptides were not significantly different between the three study groups. • Antibody responses to E2, E6 and E7 proteins were detected in 18%, 19.1% and 34% of HPV 16 DNA positive women respectively. There was however, no correlation between antibody response to HPV 16 E2, E6 and E7 protein and advancing disease stage. • Antibodies to HPV 16 VLPs were found in 50% of HPV 16 DNA positive women of study group I, 45% of HPV DNA negative women in study group II and 30% of married women in study group III suggesting that anti-VLP antibodies might serve as a marker of viral exposure (sexual activity) in the Indian context. • Antibodies to HPV 18 VLPs were detected in 60%, 30% and 26.7% of HPV 18 DNA positive women in study group I, HPV DNA negative women of study group II and married women in study group III respectively. These findings suggest that anti-VLP antibody detection serves as a marker of viral exposure rather than a marker of disease prognosis.
| Item Type: | Thesis (Doctoral) |
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| Uncontrolled Keywords: | Human papillomavirus (HPV), Cervical neoplasia, viral and immunological markers. |
| Subjects: | Respiratory Medicine > Microbiology > Respiratory Medicine > Microbiology |
| Depositing User: | Devi S |
| Date Deposited: | 21 Jun 2017 09:26 |
| Last Modified: | 11 Sep 2022 05:56 |
| URI: | http://repository-tnmgrmu.ac.in/id/eprint/178 |
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