Master of Science / Division of Biology / Nicholas A. Wallace / Cancer is not a single story, but rather numerous often interwoven tales, each with its own characters and progression. In the case of human papillomavirus (HPV) induced cervical cancer (CaCx), the narrative is about the relationship between virus and host, with the consequences of evolution’s shortsightedness driving the plot. Along with the increased proliferative state characteristic of cancer, cells experience frequent, inaccurate replication and replication stresses (ex. DNA damage and nucleotide starvation). To prevent replication fork stall and collapse generated by these stresses, the cell employs translesion synthesis (TLS). Notably, most of the genes in this pathway are upregulated in CaCx; however, the key protein polymerase eta is not. We have observed that upregulation in this pathway is complicated. It occurs at numerous levels, increasing both mRNA and protein abundance. This research further dissects how TLS upregulation occurs. Data shows that in CaCx-derived cell lines, the stability of some TLS proteins is increased, while the stability of other TLS proteins is unchanged. The increased proliferation, typical of these cell lines, cannot account for the enhanced stability. Despite increased TLS protein stability, these cells fail to adequately activate TLS increasing the risk of DNA damage. Genomic instability is a driving factor in HPV genome integration that prevents viral propagation and leads to cell transformation. It also raises mutagenesis rates, likely creating a selective pressure for tolerating failed TLS. The elevated mutation rate known to be associated with failed TLS could also provide a mechanism for acquired resistance to the drugs commonly used to treat CaCx. Changes in protein abundance are routinely used as biomarkers that can lead to the improved outcomes associated with early cancer detection. Elevated TLS protein could be leveraged to ensure cervical cancers are detected during Stage 1, when the 5-year survival rate is 80-90%, rather than at Stage IV, when the rate dips to around 15%.

Esta tese se fundamenta na necessidade de novos tratamentos para o câncer do colo de útero visando o aumento da adesão dos pacientes aos tratamentos, assim como à qualidade de vida dos mesmos. Nesse sentido, formulações nanotecnológicas foram desenvolvidas com o objetivo de carrear o fármaco imiquimode para um local específico – a mucosa vaginal – esperando gerar melhores desempenhos nesse tratamento quando comparados com a formulação comercial. Três nanoestruturas com morfologias distintas foram propostas visando potencializar o efeito do fármaco em células de câncer cervical (SiHa). As formulações desenvolvidas compreenderam: nanoemulsões (NEimiq), nanocápsulas poliméricas (NCimiq) e nanocápsulas poliméricas revestidas com quitosana (NCimiq-chit). Observou-se que nanocápsulas poliméricas produzidas com poli(ε-caprolactona) apresentaram efeito mais pronunciado frente às células SiHa. Para tanto, essas formulações (NCimiq e NCimiq-chit) foram incorporadas em hidrogéis de quitosana e de hidroxietilcelulose a fim de possibilitar uma melhor futura aplicação para o paciente. Estudos envolvendo mucosa vaginal suína demonstraram que ambas as formulações são mucoadesivas e permeiam a mucosa vaginal. Porém, a formulação produzida com hidrogel de quitosana (NCimiq) apresentou maior desempenho. Esta foi a formulação escolhida para dar continuidade aos estudos deste trabalho, sendo objeto de estudo posterior em cultura de células SiHa a fim de elucidar o mecanismo de ação da mesma. Esses estudos demonstraram que há uma ocorrência de processos combinados de diminuição da viabilidade celular de maneira tempo-dependente e que mecanismos como apoptose, autofagia e parada de ciclo celular estão presentes. Essa formulação (NCimiq) apresentou porcentagens de morte celular significativas, mesmo utilizando baixas concentrações do fármaco. Portanto, os achados desta tese constataram que nanoestruturas modulam efetivamente a interação do fármaco com as células. / This thesis deals with the need of new treatments for cervical cancer in order to increase the adherence of patients to the treatment as well as to improve their quality of life. In this sense, nanotechnological formulations were developed to carry imiquimod to a specific site – the vaginal mucosa – expecting to obtain better performance than the commercial drug in the cervical cancer treatment. Three nanostructures with different morphologies were proposed to potentilize the drug effect on cervical cancer cells (SiHa). The developed formulations are: nanoemulsions (NEimiq), polymeric nanocapsules (NCimiq) and polymeric nanocapsules coated with chitosan (NCimiq-chit). It was observed that polymeric nanocapsules produced with poly(ε-caprolactone) presented a stronger effect against SiHa cells. Therefore, formulations NCimiq and NCimiq-chit were incorporated into hydrogels of chitosan and hydroxyethylcellulose to enable a better future application on patients. The studies of this thesis involving porcine vaginal mucosa demonstrated that both formulations are mucoadhesive and that they provided a good drug permeation. However, the formulation produced with chitosan hydrogel (NCimiq) showed a better performance. This formulation was therefore chosen to follow the next steps of this work, conducted in SiHa cell culture to elucidate its action mechanism. This study demonstrated that there is an occurrence of combined processes of decreasing cell viability in a time-dependent type. The study also showed that mechanisms such as apoptosis, autophagy and cell cycle arrest are simultaneously present. The formulation NCimiq presented a significantly percentage of cellular death, even when low concentrations of the drug were used. Consequently, the findings of this thesis indicate that nanostructures effectively modulate the interaction of the drug with the cancer cells.

Orientador: Rogério Dias / Banca: Paulo Traiman / Banca: Ricardo Bassil Lasmar / Banca: Ildoberto Carneiro de Oliveira / Banca: José Carlos Meneguesi / Resumo: Avaliar a segurança oncológica da Histerectomia Radical laparoscópica através da análise da peça cirúrgica, comparando o material retirado pela via laparotômica com o extraído pela via laparoscópica, utilizando a mesma técnica operatória em ambas as vias. Pacientes e método: Estudoprospectivo e randomizado efetuado entre março/200ó e maio/2007, comparando 21 pacientes (grupo 1) submetidas a Histerectomia Radical Abdominal (HRA) com outras 21 (grupo 2) submetidas a Histerectomia Radical Laparoscópica (HRL).Todas eram portadoras de câncer do colo uterino, estadiamento clínico entre IA1 e liA. Foi feita a comparação da peça operatória retirada nos dois grupos e ainda variáveis c1inicas, epidemiológicas, de per e pós-operatório e de anatomia patológica. A técnica cirúrgica utilizada, o cirurgião e o patologista foram os mesmos nos dois grupos. Resultados: Não houve significância (NS) nas variáveis clínicas e epidemiológicas: cor, idade, IMe, hipertensão, diabetes, estadiamento clínico. Nas variáveis de per e pósoperatório houve diferença estatísticas (S) nas relacionadas com a cirurgia, mas foram NS com as complicações: tempo de cirurgia (S), perda sanguínea (S), dias de internação (S), dia que iniciou a deambulação (S), permanência do cateter vesical (S), infecção (NS), fístulas (NS), deiscência de ferida operatória (NS), trombose venosa profunda (NS) e evisceração (NS). As variáveis anatomopatológicas só mostraram significância no tamanho do tumor que foi maior no grupo 2 (Iaparoscópico): peso do útero (NS), tamanho do tumor(S), tipo anatomopatológico (NS), grau de diferenciação (NS), invasão angiolinfática (NS), invasão de paramétrios (NS), invasão da vagina (NS), número de gânglios pélvicos (NS), invasão de Iinfonodos à direita (NS) e à esquerda... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: To evaluate the oncological security of the Laparoscopic Radical Hysterectomy through analyzes of the surgical specimen, comparing the material removed from the laparotomic approach with the one extracted from the laparoscopy, using the same surgery technique in both. Patients and method: In a prospective and randomized study effected between march/2006 and may/2007, 21 submitted patients were compared Abdominal Radical Hysterectomy (ARH) with others 21 submitted Laparoscopic Radical Hysterectomy (LRH).Ali of them had cervical cancer with FIGO stage included IA1 and liA. Withdrawal in the two groups was made the comparison of the surgery specimens and still changeable clinics, epidemiologists, of per and postoperative and pathological datas. The used surgical technique, the surgeon and the pathologist were the same ones in both groups. Results: It did not have significance (NS) in the clinical and epidemiologists datas: color, age, BMI, hypertension, diabetes and FIGO stage. In the statistical datas of per and postoperative it had significance (s) in the related ones with the surgery, but they had· been NS with the complications: time of surgery (s), sanguineous loss (s), days of hospital stay (s), day that initiated the walking process(s), permanence of the vesical catheter (s), infection (NS), fistulas (NS), dehiscence of surgery wound (NS), deep venous coagulate(NS) and evisceration (NS). The pathological datas had only shown significance in the size of the tumor that was bigger in group 2 (Iaparoscopic): weight of the uterus (NS), size of the tumor (S), histological type (NS), degree of différentiation (NS), angiolinfática invasion (NS), invasion of parametrial (NS), invasion of the vagina (NS), number of pelvic ganglia (NS), invasion of Iinfonodos to the right (NS) and the left (NS) and type of Iinfonodo invaded (NS). The measure... (Complete abstract click electronic access below) / Doutor

Orientador: Agnaldo Lopes da Silva Filho / Coorientador: Andrezza Vilaça Belo / Banca: Paulo Traiman / Banca: Andrea Teixeira de Carvalho / Resumo: A resposta inflamatória é um processo ativo nas neoplasias de colo do útero, podendo atuar tanto na progressão quanto na regressão da lesão. No local da inflamação estão presentes macrófagos e neutrófilos, além de citocinas como o fator de necrose tumoral-alfa (TNF-α) e o interferon-gama (IFN-γ). Este estudo visa avaliar a resposta inflamatória sérica em mulheres com lesões pré-invasoras e invasoras do colo uterino. Foram analisadas amostras de soro obtidas de mulheres sem evidência de doença (n=30 - grupo controle), com lesões pré invasoras do colo uterino (NIC) (n=30) e com carcinoma invasor de colo uterino (CCE) (n=30). O sangue das pacientes foi coletado, centrifugado e armazenado até análise laboratorial. A atividade das enzimas inflamatórias N-acetilglucosaminidase (NAG) e mieloperoxidase (MPO) foram obtidas por ensaio enzimático. Utilizou-se o método imunoenzimático de ELISA para obtenção dos níveis séricos do TNF-α e IFN-γ. As diferenças entre os grupos foram avaliadas pelo teste do qui-quadrado, Kruskal-Wallis ou Mann Whitney. A correlação entre os grupos foi realizada pelo teste Kendall's Tau. As diferenças com valor de p<0,05 foram consideradas significativas. No grupo de pacientes com lesões pré- invasoras do colo uterino, duas mulheres apresentavam NIC I (6,7%), oito pacientes NIC II (26,7%) e 20 casos apresentavam NIC III (66,7%). O estadiamento (FIGO) das pacientes com CCE invasor foi IA2 em três casos (10,0%), IB1 em 19 (63,3%), IB2 em dois (6,7%) e IIA em seis (20,0%) mulheres pertencentes a esse grupo. As atividades das enzimas NAG, MPO e níveis de TNF-α foram maiores no grupo de mulheres com NIC, quando comparado com o grupo com CCE. Os níveis de IFN-γ foram menores no grupo de mulheres com NIC, quando comparado com o grupo com CCE. Não houve uma associação significativa entre o grau do NIC e o estadiamento do CCE invasor... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Introduction: The inflammatory response is an active process in cervical cancer and may act in the progression and/or regression of the lesion. At the site of inflammation cells are present, among the most important macrophages and neutrophils and some cytokines such as TNF- and IFN- . This study aims to evaluate the inflammatory response levels in women with pre-invasive lesions and invasive cervical cancer. Methods: Were analyzed serum samples obtained from women without evidence of disease (n = 30 - control group), with pre-invasive lesions of the cervix (n = 30) and with carcinoma of the cervix (n = 30). The activity of inflammatory enzymes Nacetylglucosaminidase (NAG) and myeloperoxidase (MPO) were obtained by enzymatic assay. We used the ELISA assay method for obtaining serum levels of TNF- and IFN- . The blood of the patients was collected, centrifuged and stored until laboratory analysis. Differences between groups were evaluated by chi-square, Kruskal-Wallis and Mann Whitney. The correlation between the groups was performed using Kendall's Tau. Differences with p <0.05 were considered significant. Results: The women age ranged from 30 to 79 years. In the group of patients with preinvasive lesions of the cervix, two had CIN I (6.7%), eight patients with CIN II (26.7%) and 20 cases had CIN III (66.7%). Staging (FIGO) of patients with SCC was IA2 in three cases (10.0%), IB1 in 19 (63.3%), IB2 in two (6.7%) and IIA in six women (20.0%). The activities of enzymes NAG, and MPO and the levels of TNF- were higher in women with CIN compared with the group with SCC. The levels of IFN- were lower in the group of women with CIN compared with the group with SCC. There was not a significant association between the degree of the CIN and the staging of the SCC of the cervix with inflammation assessed by the levels of inflammatory markers used... (Complete abstract click electronic access below) / Mestre

Made available in DSpace on 2014-06-11T19:29:51Z (GMT). No. of bitstreams: 0 Previous issue date: 2010-02-26Bitstream added on 2014-06-13T19:18:29Z : No. of bitstreams: 1 lages_el_me_botfm.pdf: 285358 bytes, checksum: 43b1317997b0df835998ac263f0477a7 (MD5) / A resposta inflamatória é um processo ativo nas neoplasias de colo do útero, podendo atuar tanto na progressão quanto na regressão da lesão. No local da inflamação estão presentes macrófagos e neutrófilos, além de citocinas como o fator de necrose tumoral-alfa (TNF-α) e o interferon-gama (IFN-γ). Este estudo visa avaliar a resposta inflamatória sérica em mulheres com lesões pré-invasoras e invasoras do colo uterino. Foram analisadas amostras de soro obtidas de mulheres sem evidência de doença (n=30 – grupo controle), com lesões pré invasoras do colo uterino (NIC) (n=30) e com carcinoma invasor de colo uterino (CCE) (n=30). O sangue das pacientes foi coletado, centrifugado e armazenado até análise laboratorial. A atividade das enzimas inflamatórias N-acetilglucosaminidase (NAG) e mieloperoxidase (MPO) foram obtidas por ensaio enzimático. Utilizou-se o método imunoenzimático de ELISA para obtenção dos níveis séricos do TNF-α e IFN-γ. As diferenças entre os grupos foram avaliadas pelo teste do qui-quadrado, Kruskal-Wallis ou Mann Whitney. A correlação entre os grupos foi realizada pelo teste Kendall's Tau. As diferenças com valor de p<0,05 foram consideradas significativas. No grupo de pacientes com lesões pré- invasoras do colo uterino, duas mulheres apresentavam NIC I (6,7%), oito pacientes NIC II (26,7%) e 20 casos apresentavam NIC III (66,7%). O estadiamento (FIGO) das pacientes com CCE invasor foi IA2 em três casos (10,0%), IB1 em 19 (63,3%), IB2 em dois (6,7%) e IIA em seis (20,0%) mulheres pertencentes a esse grupo. As atividades das enzimas NAG, MPO e níveis de TNF-α foram maiores no grupo de mulheres com NIC, quando comparado com o grupo com CCE. Os níveis de IFN-γ foram menores no grupo de mulheres com NIC, quando comparado com o grupo com CCE. Não houve uma associação significativa entre o grau do NIC e o estadiamento do CCE invasor... / Introduction: The inflammatory response is an active process in cervical cancer and may act in the progression and/or regression of the lesion. At the site of inflammation cells are present, among the most important macrophages and neutrophils and some cytokines such as TNF- and IFN- . This study aims to evaluate the inflammatory response levels in women with pre-invasive lesions and invasive cervical cancer. Methods: Were analyzed serum samples obtained from women without evidence of disease (n = 30 - control group), with pre-invasive lesions of the cervix (n = 30) and with carcinoma of the cervix (n = 30). The activity of inflammatory enzymes Nacetylglucosaminidase (NAG) and myeloperoxidase (MPO) were obtained by enzymatic assay. We used the ELISA assay method for obtaining serum levels of TNF- and IFN- . The blood of the patients was collected, centrifuged and stored until laboratory analysis. Differences between groups were evaluated by chi-square, Kruskal-Wallis and Mann Whitney. The correlation between the groups was performed using Kendall's Tau. Differences with p <0.05 were considered significant. Results: The women age ranged from 30 to 79 years. In the group of patients with preinvasive lesions of the cervix, two had CIN I (6.7%), eight patients with CIN II (26.7%) and 20 cases had CIN III (66.7%). Staging (FIGO) of patients with SCC was IA2 in three cases (10.0%), IB1 in 19 (63.3%), IB2 in two (6.7%) and IIA in six women (20.0%). The activities of enzymes NAG, and MPO and the levels of TNF- were higher in women with CIN compared with the group with SCC. The levels of IFN- were lower in the group of women with CIN compared with the group with SCC. There was not a significant association between the degree of the CIN and the staging of the SCC of the cervix with inflammation assessed by the levels of inflammatory markers used... (Complete abstract click electronic access below)

Esta tese se fundamenta na necessidade de novos tratamentos para o câncer do colo de útero visando o aumento da adesão dos pacientes aos tratamentos, assim como à qualidade de vida dos mesmos. Nesse sentido, formulações nanotecnológicas foram desenvolvidas com o objetivo de carrear o fármaco imiquimode para um local específico – a mucosa vaginal – esperando gerar melhores desempenhos nesse tratamento quando comparados com a formulação comercial. Três nanoestruturas com morfologias distintas foram propostas visando potencializar o efeito do fármaco em células de câncer cervical (SiHa). As formulações desenvolvidas compreenderam: nanoemulsões (NEimiq), nanocápsulas poliméricas (NCimiq) e nanocápsulas poliméricas revestidas com quitosana (NCimiq-chit). Observou-se que nanocápsulas poliméricas produzidas com poli(ε-caprolactona) apresentaram efeito mais pronunciado frente às células SiHa. Para tanto, essas formulações (NCimiq e NCimiq-chit) foram incorporadas em hidrogéis de quitosana e de hidroxietilcelulose a fim de possibilitar uma melhor futura aplicação para o paciente. Estudos envolvendo mucosa vaginal suína demonstraram que ambas as formulações são mucoadesivas e permeiam a mucosa vaginal. Porém, a formulação produzida com hidrogel de quitosana (NCimiq) apresentou maior desempenho. Esta foi a formulação escolhida para dar continuidade aos estudos deste trabalho, sendo objeto de estudo posterior em cultura de células SiHa a fim de elucidar o mecanismo de ação da mesma. Esses estudos demonstraram que há uma ocorrência de processos combinados de diminuição da viabilidade celular de maneira tempo-dependente e que mecanismos como apoptose, autofagia e parada de ciclo celular estão presentes. Essa formulação (NCimiq) apresentou porcentagens de morte celular significativas, mesmo utilizando baixas concentrações do fármaco. Portanto, os achados desta tese constataram que nanoestruturas modulam efetivamente a interação do fármaco com as células. / This thesis deals with the need of new treatments for cervical cancer in order to increase the adherence of patients to the treatment as well as to improve their quality of life. In this sense, nanotechnological formulations were developed to carry imiquimod to a specific site – the vaginal mucosa – expecting to obtain better performance than the commercial drug in the cervical cancer treatment. Three nanostructures with different morphologies were proposed to potentilize the drug effect on cervical cancer cells (SiHa). The developed formulations are: nanoemulsions (NEimiq), polymeric nanocapsules (NCimiq) and polymeric nanocapsules coated with chitosan (NCimiq-chit). It was observed that polymeric nanocapsules produced with poly(ε-caprolactone) presented a stronger effect against SiHa cells. Therefore, formulations NCimiq and NCimiq-chit were incorporated into hydrogels of chitosan and hydroxyethylcellulose to enable a better future application on patients. The studies of this thesis involving porcine vaginal mucosa demonstrated that both formulations are mucoadhesive and that they provided a good drug permeation. However, the formulation produced with chitosan hydrogel (NCimiq) showed a better performance. This formulation was therefore chosen to follow the next steps of this work, conducted in SiHa cell culture to elucidate its action mechanism. This study demonstrated that there is an occurrence of combined processes of decreasing cell viability in a time-dependent type. The study also showed that mechanisms such as apoptosis, autophagy and cell cycle arrest are simultaneously present. The formulation NCimiq presented a significantly percentage of cellular death, even when low concentrations of the drug were used. Consequently, the findings of this thesis indicate that nanostructures effectively modulate the interaction of the drug with the cancer cells.

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Background: Although subclinical persistent infections with the human polyomaviruses are ubiquitous worldwide, they are known to vary in relation to geographical location, diseases present and may associate with different human tumours, especially in immunocompromised patients. The current study hypothesised that there may be co-operativity between HPV and polyomaviruses, particularly in HIV positive women, that could influence the rate of progression to invasive cervical carcinoma. Patients and Methods: Novel PCR methods were developed for the detection of SV40, MCV, JCV and BKV polyomavirus DNA. These were used to test DNAs extracted from 220 cervical smears and 77 invasive cervical carcinomas (ICCs) from HIV positive and negative Kenyan women of known HPV status. An expression plasmid was constructed containing JCV Large T (LT) antigen and this, in addition to empty vector control, used to stably transfect HPV16 E6/E7 immortalised human keratinocytes. Expression of LT was analysed in transfected cell lines by PCR, immunocytology and Western blotting. These cells were then used to test for changes in Cell contact growth inhibition; Growth rate and Epithelial to Mesenchymal Transition (EMT). Screening of full transcriptome microarrays was carried out on vector and LT transfected cells and their sensitivity to the drug mefloquine tested by comparison of growth rates and live/dead cell assays. Results: PCR accurately detected ~18 copies of SV40, MCV, JCV and BKV DNA in addition to simultaneous detection of JCV and BKV. None of the clinical samples tested were positive for SV40, MCV, or BKV DNA. However, JCV DNA was detected in 24/297 (8%) of cervical specimens. Comparison of the incidence of JCV in cervical smears and ICCs showed a ~3-fold increase in samples from HIV positive women with ICC (P=0.025) whereas no significant difference was found between smears and ICCs from HIV negative women (P=0.553). Analysis of the consequences of ectopic expression of JCV LT in E6/E7 immortalised human keratinocytes showed no difference in either growth rates or contact inhibition and changes in the EMT marker vimentin were found to be related to cellular clonality. Microarray analysis showed LT related alterations in gene expression which could have bearing on its carcinogenic potential in addition to changes related to clonality. JCV LT expressing monoclonal cell were the most sensitive to mefloquine treatment. Conclusion: The simultaneous JCV/BKV detection method, described herein, is unique and has been evaluated by the WHO for this purpose. The results indicate the prevalence JCV and BKV with respect to the African geographical location and suggest that JCV may combine with high-risk HPV in a sub-set of HIV positive women to influence the rate of progression to invasive cervical carcinoma. In vitro JCV LT was found not to be an overt oncogene in the cell system used although cell cloning procedures clearly affected the assays. LT induced changes in total gene expression were consistent with neoplastic progression although a high proportion of genes with unknown function were dsyregulated with respect to clonality. The anti JCV drug mefloquine showed some selectivity for LT expressing cells and further investigation of this indication is warranted.

Great disparities exist in women’s health outcomes in rural indigenous communities in Guatemala; indigenous women are less likely to utilize family planning and cervical cancer screening services. This pilot study assessed baseline women’s health knowledge and the effects of a comprehensive women’s health course on women’s health knowledge and behaviors. From February to November 2013, 61 participants in a seven-week language-appropriate women’s health curriculum were evaluated before and after the course using a 10-item knowledge assessment. Cervical cancer screening and contraceptive utilization rates were assessed by self-report before and after the course. Several women’s health knowledge deficits were noted: belief that contraception causes cancer, inability to name symptoms of STIs, lack of understanding of pap smears, and lack of familiarity with condoms. The average pre-test score was 54.6%; increasing to 83.7% on the post-test (p<0.0001). 79% had received a pap smear prior to the course; this increased to 92% at the post-test (p=0.013). 53% had utilized contraception prior to the course; at the time of the post-test the percentage was 54% (p=1). This study reveals important patterns in women’s health beliefs in rural Guatemala, and shows that a seven-week curriculum was effective in improving scores on a knowledge assessment.

PROJECT ONE Introduction HPV is detected in about 99.7% of cervical cancers. However, the HPV type distribution in South African women is unknown. Objectives To determine HPV-type distribution among women with cervical dysplasia in relation to oral contraceptive usage. Methods Prospective cross-sectional study of four groups of patients according to oral contraceptive usage: non-users, users of less than five years duration, users of between five years and ten years and users of more than ten years duration. Swabs of the cervix were analysed for HPV DNA using polymerase chain reaction method. Results A total of 124 women were recruited for the study. There were 75 HIV-infected patients (seroprevalence 61%). Of the 102(82%) HPV-positive patients, 79 patients had high-risk HPV DNA (78%). In terms of the four oral contraceptive groups, high-risk HPV DNA was detected in 70% (n=21), 79% (n=22), 90% (n=21) and 71% (n=15) of patients, respectively. The odds of having HPV DNA was six times higher for the combination of contraceptive users of less than 5 years duration/non-users (OR 5.9, 95% CI: 1.87 - 18.77). There was no change when adjustment was made for age (OR 6.1, 95% CI: 1.9 - 19.4). HPV DNA types 16 and or 18 was present in a total of 21 patients (49%) (non-contraceptive users and users < 5years duration) versus 15 patients (42%) who used oral contraceptives of more than 5 years duration (p=0.524). HPV type 16 was the commonest HPV type detected (20.2%) and HPV type 58 was the next commonest high-risk HPV type (16.1%). HPV types 58 and 33 was detected in a much greater percentage of our population and HPV 16 in a much smaller percentage of our population compared with a non-South African population. Conclusion The findings of this study demonstrate an interesting distribution of HPV types in a South African population. PROJECT TWO Introduction Various risk factors have been implicated in the causation of cervical cancer including human papillomavirus (HPV), the early genes (E6 and E7) of which encode the main transforming proteins. Studies have suggested that steroid hormones may enhance the expression of these genes leading to loss of p53 gene-mediated cell apoptosis. Methods A total of 120 cervical tissue samples were obtained from patients with proven cervical cancer. Patients who used depo-medroxyprogesterone acetate steroid contraception were recruited as part of the study arm. Only HPV DNA type 16 samples were used for the study. Controls included three cell lines (CaSki, SiHa,&C33A) and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was used as an internal housekeeping gene. Of 120 patients, there were 111 patients with HPV type 16 identified. Of this number, RNA was present in 63 samples. There were 30 women (30/63) who used steroid contraception. In relation to patients who used contraception, HPV 16 E6 gene expression was present in 79% (n = 23) and 88% (n = 30) of steroid users compared to nonusers, respectively. In total there were 25 patients (40%) with expression of the HPV 16 E6*I gene and 30 patients with expression of the E6*II gene. There were 57% of steroid users (n = 17) who had expression of the E6*I/E6*II gene, compared to 52% (n = 17) of nonusers (P = 0.800). Conclusion From a molecular level, this study reflects almost similar distribution of the HPV 16 E6/E6*1 and E6*11 and does not confirm the role of injectable progesterones in cervical carcinogenesis. Further studies with larger patient numbers are needed. / Thesis (PhD)--University of Pretoria, 2011. / Obstetrics and Gynaecology / unrestricted