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Development of cervical cancer is a long process of abnormal cancerous cell growth in the cervix and is primarily the result of infection with specific high-risk types of human papillomavirus (HPV). The cytokines tumor necrosis factor α (TNFα) and lymphotoxin α (LTA) have an important role in all stages of cervical cancer and have the ability to induce the regression or promote the development of human tumors. Biologically important single-nucleotide polymorphisms (SNPs) occur within the TNFα and LTA genes. Therefore, the purpose of this study was to investigate the SNPs in the TNFα promoter region (-163, -238, -244, -308, -376, -857, -863, and -1031) and in the first intron of LTA (+252) in women with precursor lesions of cervical cancer. Overall, we studied 396 women from Mexico City. A total of 191 patients with HPV infection and precursor cervical lesions were subdivided in two groups: those with low-grade squamous intraepithelial lesions (n = 132) and those with high-grade squamous intraepithelial lesions (n = 59). Women (n = 205) negative for HPV and without cervical lesions were also included in the study. DNA was extracted from peripheral white blood cells and from cervical samples, and detection of biallelic polymorphisms of TNFα and LTA was performed using the polymerase chain reaction-sequence-specific oligonucleotide probe and restriction fragment length polymorphism techniques, respectively. We demonstrated that risk is associated with the genotype G/A (odds ratio = 2.48) and that protection is associated with the genotype G/G of SNP TNFα -376 (odds ratio = 0.37).

Background

Human papillomavirus (HPV) infection plays an etiological role in the development of cervical dysplasia and cancer. Amplification of human telomerase gene (hTERC) and over expression of telomerase were found to be associated with cervical tumorigenesis. This study was performed to analyze genomic amplification of hTERC gene, telomerase activity in association with HPV infection in different stages of cervical intraepithelial neoplasia (CIN) and cervical cancer. We were studying the role of hTERC in the progression of uterine cervical dysplasia to invasive cancer, and proposed an adjunct method for cervical cancer screening.

Methods

Exfoliated cervical cells were collected from 114 patients with non neoplastic lesion (NNL, n=27), cervical intraepithelial neoplasia (CIN1, n=26, CIN2, n=16, CIN3, n=24) and cervical carcinoma (CA, n=21), and analyzed for amplification of hTERC with two-color fluorescence in situ hybridization (FISH) probe and HPV-DNA with Hybrid Capture 2.

From these patients, 53 were taken biopsy to analyze telomerase activity by telomeric repeat amplification protocol (TRAP) and expression of human telomerase reverse transcriptase (hTERT), with immunohistochemistry (IHC). All biopsies were clinically confirmed by phathologists.

Results

Amplification of hTERC was significantly associated with the histologic diagnoses (p<0.05). The positive correlation was found between the level of hTERC amplification and histologic grading of dysplasia (CIN2/3 from CIN1 or normal, P=0.03). A profounding increase in the accumulation of HPV and hTERC positive cases was observed in the CIN3 subgroup compared with the CIN2 group, 25% versus 62.96%, respectively (p=0.007).

Conclusions

hTERC ampliffication can be detected with FISH technique on exfoliated cervical cells. Amplification of hTERC and HPV infection are associated with more progressive CIN3 and CA. The testing of hTERC amplification might be a supplementary to cytology screening and HPV test, especially high-risk patients.

Virtual slides

The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/1857134686755648.

Cervical cancer (CxCa) is caused by persistent human papillomavirus (HPV) infection; genetic predisposition is also suspected to play a role. The present study is a targeted candidate gene follow-up based on: i) strong clinical evidence demonstrating that mutations in the TMC6 and TMC8 (EVER1 and EVER2) genes associate with the HPV-associated disease Epidermodysplasia Verruciformis (EV), and ii) recent epidemiological data suggesting a genetic susceptibility conferred by polymorphisms in such genes for skin and cervical cancer. Clarifying the association of the TMC6/8 genes with risk of CxCa will help in understanding why some HPV-infected women develop persistent infection, cervical lesions and eventually cancer while others do not. Twenty-two single nucleotide polymorphisms (SNP) harbouring the TMC6/8 genes were genotyped in 2,989 cases with cervical intraepithelial neoplasia grade III (CINIII) or invasive cervical cancer (ICC) and 2,281 controls from the Swedish population. Association was evaluated in logistic regression models. Two SNPs displayed association with cervical disease: rs2290907 (ORGGvsAA = 0.6, 95% CI: 0.3 - 0.9, p = 0.02) and rs16970849 (ORAGvsGG = 0.8, 95% CI: 0.66 - 0.98, p = 0.03). The present data supports the involvement of the TMC6/8 region in CxCa susceptibility but further analyses are needed to replicate our findings, fully characterize the region and understand the function of the genetic variants involved.

Background

Women with Human Papilloma Virus (HPV) persistence are characterized by high levels of IL-10 at cervix. We have determined whether polymorphisms of IL-10 gene promoter might be associated with increased risk of squamous intraepithelial cervical lesions (SICL) and whether exist significative differences of IL-10 mRNA expression at cervix and systemic and serum IL-10 protein between SICL cases and non-Cervical Lesions (NCL).

Methods

Peripheral blood samples from SICL (n = 204) and NCL (n = 166) were used to detect IL-10 promoter polymorphisms at loci -592A/C (rs1800872), -819C/T (rs1800871), -1082A/G (rs1800896), -1352A/G (rs1800893), by allelic discrimination and to evaluate serum IL-10 protein. Cervical epithelial scrapings from NCL and biopsies from SICLs were used for HPV-typing and to evaluate IL-10 mRNA expression level. The systemic and local IL-10 mRNA expression levels were measured by real time-PCR. Genotypic and allelic frequencies of the selected polymorphisms were analyzed by logistic regression, adjusting by age and HPV-genotype, to determine the association with SICL.

Results

No significant differences were found between genotype frequencies at loci −819, -1082, and −1352. Individuals carrying at least one copy of risk allele A of polymorphism −592 had a two-fold increased risk of developing SICL [adjusted odds ratio (OR), 2.02 (95% CI, 1.26-3.25), p = 0.003], compared to NCL. The IL-10 mRNA expression and serum IL-10 protein, were significantly higher in SICL cases (p < 0.01), being higher in patients carrying the risk allele A.

Conclusions

The −592 polymorphism is associated with increased risk of SICL and can serve as a marker of genetic susceptibility to SICL among Mexican women. According to IL-10 levels found in SICL, IL-10 can be relevant factor for viral persistence and progression disease.

Infection with certain types of human papillomavirus (HPV) presents a high risk for the subsequent development of cervical intraepithelial neoplasia (CIN) and cervical carcinoma. Immunological mechanisms are likely to play a role in control of cervical HPV lesions. The HPV E2 protein has roles in virus replication and transcription, and loss of E2 functions may be associated with progression of cervical neoplasia. Accordingly, it is of interest to monitor immune responses to the E2 protein, and previous studies have reported associations between serological reactivity to E2 peptide antigens and cervical neoplasia. In order to investigate serological responses to native, full-length E2 protein, we expressed HPV-16 E2 proteins with and without an N-terminal polyhistidine tag using the baculovirus system. Purified HPV-16 E2 protein was used to develop enzyme-linked immunosorbent assays to detect serological IgG and IgA responses in cervical neoplasia patients and controls. We found that serum IgA levels against the E2 protein were elevated in CIN patients relative to normal control subjects but were not elevated in cervical cancer patients. Moreover, there appeared to be a gradient of response within cervical neoplasia such that the highest antibody levels were seen in lower grades of neoplasia up to CIN 2, whereas lower levels were observed in CIN 3 and still lower levels in cervical carcinoma. These findings suggest that the IgA antibody response to E2 may associate with stage and progression in cervical neoplasia.

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Cervical cancer is almost invariably associated with infection by human papillomavirus. It is believed that the host genetic factors such as inflammation-induced cytokines may play a role in cervical carcinogenesis. The IL1B gene, encoding IL-1β cytokine, contains several single nucleotide polymorphisms. One of them which is in the positions -511 (C-T) related with promoter region has been associated with increased IL-1β production and with increased risk of developing a number of inflammatory diseases and gastric carcinoma. We assessed the association between the IL1B -511 polymorphism and cervical cancer risk in a hospital-based case-control study among 546 Korean women (182 cases; 364 age-matched controls). The allele frequencies of the case subjects (C, 0.42; T, 0.58) were not significantly different from those of control subjects (C, 0.43; T, 0.57). Control subjects were in Hardy-Weinberg equilibrium. The carriers with -511 C/T or T/T genotypes were at higher risk of cervical cancer with odds ratio of 2.42 (95% CI 1.31-4.46, p<0.005). However, there was no difference of cervical cancer risk between C/T heterologous genotypes and T/T homologous genotypes. In conclusion, in Korean population, IL1B -511 C/C genotypes were significantly associated with a decreased risk of cervical cancer.

Human papillomavirus (HPV) E6 and E7 oncogenes are expressed in the great majority of human cervical carcinomas, whereas the viral E2 regulatory gene is usually disrupted in these cancers. To investigate the roles of the papillomavirus E2 genes in the development and maintenance of cervical carcinoma, the bovine papillomavirus (BPV) E2 gene was acutely introduced into cervical carcinoma cell lines by infection with high-titer stocks of simian virus 40-based recombinant viruses. Expression of the BPV E2 protein in HeLa, C-4I, and MS751 cells results in specific inhibition of the expression of the resident HPV type 18 (HPV18) E6 and E7 genes and in inhibition of cell growth. HeLa cells, in which HPV gene expression is nearly completely abolished, undergo a dramatic and rapid inhibition of proliferation, which appears to be largely a consequence of a block in progression from the G1 to the S phase of the cell cycle. Loss of HPV18 gene expression in HeLa cells is also accompanied by a marked increase in the level of the cellular p53 tumor suppressor protein, apparently as a consequence of abrogation of HPV18 E6-mediated destabilization of p53. The proliferation of HT-3 cells, a human cervical carcinoma cell line devoid of detectable HPV DNA, is also inhibited by E2 expression, whereas two other epithelial cell lines that do not contain HPV DNA are not inhibited. Thus, a number of cervical carcinoma cell lines are remarkably sensitive to growth inhibition by the E2 protein. Although BPV E2-mediated inhibition of HPV18 E6 and E7 expression may contribute to growth inhibition in some of the cervical carcinoma cell lines, the BPV E2 protein also appears to exert a growth-inhibitory effect that is independent of its effects on HPV gene expression.

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Background

The causal association between persistent human papillomavirus (HPV) infection and cervical cancer has been established, but the mechanisms that favor HPV persistence in cervical cells are still unknown. The diminished capability of the immune system to control and resolve HPV infection is one of several hypotheses. The tolerogenic protein HLA-G has shown aberrant expression in a variety of cancers, which has been suggested as a mechanism for tumor escape from immunosurveillance. In the present study we evaluate the role of epigenetic modification (promoter de-methylation) of the HLA-G gene on susceptibility to HPV infection and development of high-grade cervical lesions.

Methods

A case–control study was carried out in Curitiba, Brazil, between February and June 2010. A total of 789 women aged 15–47 years were recruited: 510 controls with normal cervical cytology, and 279 cases with histologically confirmed cervical intraepithelial neoplasia grade 2 (CIN2, N = 150) or grade 3 (CIN3, N = 129). All women were administered a questionnaire by interview, which collected information on demographic and lifestyle factors, and a cervical sample was collected. HPV DNA detection was performed by GP5+/GP6+ primer-mediated PCR. HPV-positive samples were genotyped by multiplex PCR. A pilot analysis of HLA-G promoter methylation was carried out in a subset of the study population (96 cases and 76 controls) by pyrosequencing. HLA-G methylation and HPV infection status of cases and controls were compared, and confounding factors were computed by t Student and non-parametric Wilcoxon tests. Comparison of HLA-G methylation between cases and controls was assessed by the Bonferroni correction. The association of HLA-G methylation with CIN2/3 was evaluated by logistic regression.

Results

HPV prevalence was 19.6% in controls and 94.3% in CIN2/3 cases. HPV16, 31, 33, 35 and 18 were the most prevalent types. Methylation analysis of seven CpGs in the HLA-G promoter did not reveal any spontaneous de-methylation events in CIN2/3 cases (mean proportion of methylation: 75.8%) with respect to controls (mean 73.7%; odds ratio 1.01, 95% confidence interval 0.96, 1.07).

Conclusions

This study did not support the hypothesis that spontaneous de-methylation events in the HLA-G promoter play a primary role in promoting escape from immunosurveillance in the development of precancerous cervical lesions.

Persistent infections by high-risk types of human papillomavirus (HPV) have been established as the etiological agent of cervical cancer. The integration of the HPV genome into the host genome is a crucial step in cervical carcinogenesis, although, correct activation of DNA damage repair pathways will avoid the development of cancer. Recent data indicate that several polymorphisms of key regulators from the DNA damage repair pathway (i.e. 53BP1 and ATM) are associated with cancer development susceptibility. We have developed a hospital-based retrospective study considering 429 cervical specimens from women with different cervical lesions including invasive carcinoma. This study aimed to evaluate the role of the ATM D1853N (5557G>A) and 53bp1 D353E (1236C>G) polymorphisms in the development of cervical cancer, using TaqMan® SNP Genotyping Assays. Statistical analysis revealed that ATM 5557GG homozygous individuals (OR=1.94; p=0.044) are at increased risk of developing LSIL, while for the 53BP1 1236C>G polymorphism no association was found. Nevertheless, we observed a tendency for an increased risk of LSIL in 53BP1 1236C allele carriers (OR=1.63; p=0.069). Logistic regression adjusted for age revealed no significant differences from the non-adjusted analysis. This is the first study to evaluate the role of ATM G5557A and P53BP1 C1236G polymorphisms in cervical cancer susceptibility. This study reveals a possible trend of both polymorphisms for a genetic susceptibility pattern of cervical cancer development. Hence, our results may be of interest for future understanding of the progression of cervical cancer.

Different rate of development of productive infections (as low grade cervical intraepithelial neoplasias), or high grade lesions and cervical malignant tumors associated with infections of the Transformation zone (TZ) by High-Risk Human Papillomavirus (HR-HPV), could suggest that different epithelial host target cells could exist. If there is more than one target cell, their differential infection by HR-HPV may play a central role in the development of cervical cancer. Recently, the concept that cancer might arise from a rare population of cells with stem cell-like properties has received support in several solid tumors, including cervical cancer (CC). According to the cancer stem cell (CSC) hypothesis, CC can now be considered a disease in which stem cells of the TZ are converted to cervical cancer stem cells by the interplay between HR-HPV viral oncogenes and cellular alterations that are thought to be finally responsible for tumor initiation and maintenance. Current studies of CSC could provide novel insights regarding tumor initiation and progression, their relation with viral proteins and interplay with the tumor micro-environment. This review will focus on the biology of cervical cancer stem cells, which might contribute to our understanding of the mechanisms responsible for cervical tumor development.

Whilst there is strong evidence that human papillomavirus (HPV) is the principal aetiological agent in cervical neoplasia, some other sexually transmitted agents may either contribute or protect against cervical carcinogenesis, such as the herpes virus family (HSV), cytomegalovirus (CMV), Epstein-Barr virus (EBV), human immunodeficiency virus (HIV) or Chlamydia trachomatis (CT). Epidemiological studies suggest that HSV may have a role in cervical neoplasia, but there is no clear supportive experimental evidence. Serological studies have also failed to reveal a difference in the prevalence of antibodies to CMV and EBV between patients with cervical cancer and controls. However, longitudinal seroepidemiological studies have provided evidence that CT is an independent risk factor for the development of cervical squamous carcinoma and this association is serotype specific. The increased risk of cervical neoplasia in patients infected with HIV has been recognised for over a decade and HIV may interact with HPV either by alternating HPV gene transcription or by immunosuppression. Finally extensive experimental and limited epidemiological evidence suggests that adeno-associated viruses (AAV) may have antioncogenic activity in man and may protect against the development of cervical cancer. At present the mechanism of this action is unclear but may relate to AAV-induced regulation of HPV gene expression and the HPV life cycle. In this review we summarize the current literature relating to the associations and mechanisms of cervical carcinogenesis by each of these infectious microorganisms.

Abnormal fragile histidine triad transcripts were found in 20–30% of CIN2/3 lesions and 11% of normal cervical biopsies by RT–PCR. Bi-allelic loss of the fragile histidine triad gene and the loss of fragile histidine triad protein expression detectable by immunochemical staining with a polyclonal fragile histidine triad specific antibody was rare. The genomic changes showed no association with the presence of human papillomavirus types which carry high risk for cervical cancer (high risk human papillomavirus) as assessed by a type-specific multiplex PCR. The presence of abnormal fragile histidine triad transcripts in a subset of CIN2/3 lesions with no high risk human papillomavirus suggests that this could be an independent risk factor associated with an alternative carcinogenic pathway.

British Journal of Cancer (2002) 86, 376–381. DOI: 10.1038/sj/bjc/6600077 www.bjcancer.com

© 2002 The Cancer Research Campaign

Host genetic variability modifies the risk of cervical cancer in women infected with oncogenic human papillomavirus (HPV). Studies have reported an association of the TP53 codon 72 arginine and cervical cancer, but the results are inconsistent. We examined the association of this single nucleotide polymorphism (SNP) in women with cervical cancer and cervical intraepithelial neoplasia grade 3 (CIN3), using family-based association test. We further explored SNPs in two genes that regulate p53 stability: MDM2 (SNP309) and NQO1 (SNP609, SNP465). We also examined the relationship between host genotype and tumor HPV type. We genotyped 577 patients and their biological parents and/or siblings, using PCR-RFLP or TaqMan assays. HPVs were typed by sequence-based methods. The transmission/disequilibrium test was used to detect disease-susceptibility alleles. The arginine peptide of TP53 codon 72 was overtransmitted in Caucasian families (P=0.043), and the significance of this finding was enhanced in a subgroup of women infected with HPV16- and/or 18-related HPVs (P=0.026). Allele C of NQO1 SNP609 was also overtransmitted in all cases (P=0.026). We found no association between MDM2 SNP309 or NQO1 SNP465 and cervical cancer. Our results indicate that functional polymorphisms in TP53 codon 72 and NQO1 SNP609 associate with the risk of cervical cancer especially in women infected with type 16- and/or 18-related HPVs.

Cervical cancer is a common cancer inflicting women worldwide. Even though, persistent infection with oncogenic Human Papillomavirus (HPV) types is considered the most important risk factor for cervical cancer development, less than 5% of women with HPV will eventually develop cervical cancer supporting that other molecular events, like methylation-dependent inactivation of tumor suppressor genes, may cocontribute in cervical carcinogenesis. We analyzed promoter methylation of three candidate genes (p16, MGMT, and hMLH1) in 403 liquid-based cytology samples. Methylation was commonly identified in both benign and pathologic samples and correlated with higher lesion grade determined by cytological, colposcopical, or histological findings, with HPV DNA and mRNA positivity of specific HPV types and p16INK4A protein expression. Overall accuracy of methylation is much lower than traditional diagnostic tests ranking it as an ancillary technique with more data needed to identify the exact value of methylation status in cervical carcinogenesis.

Background: Infection with human papillomavirus (HPV) is the main cause of cervical cancer and its precursor lesion, cervical intraepithelial neoplasia (CIN). Variability in host immunogenetic background is important in determining the overall cellular immune response to HPV infections.

Objective: To determine whether the HLA-DQ or HLA-DR genes, or others in their vicinity, are associated with cervical cancer.

Methods: Markers covering the entire HLA region were genotyped in a large sample of CIN and cervical cancer patients and in controls (311 CIN, 695 cervical cancer, 115 family controls, and 586 unrelated controls).

Results: Two markers were associated with susceptibility to cervical neoplasia, G511525 and MICA. G511525, close to the region containing the HLA-DQ and HLA-DR genes, was most strongly associated, showing a decrease in frequency of allele 221 from 6.7% to 3.3% in patients with squamous cell cancer (SCC). An association was found for MICA (allele 184) with SCC (odds ratio (OR) = 1.31 (95% confidence interval, 1.13 to 1.53); homozygotes, OR = 1.48 (1.06 to 2.06)). No associations were observed with adenocarcinoma or CIN.

Conclusions: There is an association of the region containing the HLA-DQ and HLA-DR genes with the risk of developing squamous cell carcinoma. An increased risk was observed for carriers of allele 184 at the MICA locus, in particular for homozygotes, suggesting a recessive effect.

Cervical carcinomas are closely associated with high-risk human papillomavirus (HPV) types and are preceded by cervical intraepithelial neoplasia (CIN). Most CIN lesions regress spontaneously and will not evolve to invasive carcinoma. The cellular immune system mediated by cytotoxic T lymphocytes (CTLs) and natural killer (NK) cells are thought to play an important role in the ultimate decline of CIN lesions. Although TIA-1 is constitutively expressed in the majority of circulating T cells and defines a subpopulation of CD8+ T cells with cytotoxic potential, granzyme B is only expressed in CTLs upon activation. In the present study we have evaluated the expression of these proteins by lymphocytes present in 24 randomly chosen CIN lesions with increasing degree of atypia and in 14 cervical squamous cell carcinomas. As major histocompatibility complex (MHC) class I expression is frequently down-regulated in HPV-induced lesions, thus possibly frustrating tumour cell recognition by infiltrating CTLs, these lesions were also analysed for MHC class I expression. The results indicated that in most CIN lesions only a minority of CTLs are activated, whereas in some carcinomas a massive infiltration of activated, i.e. granzyme B-positive, CTLs were observed. The percentage of activated CTLs was not related to expression of MHC class I on neoplastic cells. These results suggest that in some carcinomas proper activation of CTLs occurs but that most likely local factors or immunoselection of resistant neoplastic cells inhibit a proper response of CTLs to these neoplastic cells.

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Previously, we have reported that the human papillomavirus (HPV) type 16 E6 D25E is the most prevalent variant in Korean women at high risk for cervical cancers. Several studies have identified an association between the increased frequency of this variant and the elevated risk of cervical intraepithelial neoplasia and invasive cervical carcinoma. To investigate whether the HPV-16 E6 D25E variant might influence cervical cancer progression, we used an oligonucleotide microarray approach to identify transcriptionally altered gene expression patterns in recombinant wild-type E6 or E6 D25E variant-expressing HPV-negative cancer cells. We found that 211 genes were significantly up- or down-regulated (at least 1.5-fold, p < 0.05). We identified 14 genes, nine down-regulated and five up-regulated upon E6 D25E expression, compared with wild-type E6 expression. These results further emphasize the unique biological activity of the HPV-16 E6 D25E variant.

Smoking is a known aetiological risk factor for cervical cancer. Smoking-related DNA damage (DNA adducts), in cervical epithelial cells, has recently been demonstrated to suggest a causal role in the development of cervical cancer. Human papillomavirus 16 (HPV 16) is a known oncogenic virus and is also implicated as a cause of cervical cancer. It has been suggested that both smoking and HPV may act synergistically in the development of cervical cancer. We have investigated the cervical DNA adduct level and the prevalence of HPV 16 (using polymerase chain reaction) in women who had normal cervical cytology. Both the DNA adduct assay and the HPV assay were carried out on exfoliated cervical cells recovered from cervical scrapes. In 87% of the cases there was enough DNA from the exfoliative cervical cells to analyse for DNA adducts. Smokers had higher DNA adduct levels than non-smokers (P = 0.002), confirming the previous data from cervical biopsy samples. Forty-two per cent of the specimens were found to be HPV 16 positive. There was no significant difference in smoking-related DNA damage (DNA adduct levels) between HPV-positive and HPV-negative smokers. This suggests that smoking DNA damage does not augment HPV infectivity. These results do not, therefore, support the molecular synergism theory.

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Human papillomavirus (HPV), the primary cause of cervical cancer, is also associated with the development of anal cancer. Relatively little is known about the epidemiology of anal HPV infection among healthy females and its relationship to cervical infection. We sought to characterize anal HPV infection in a cohort of adult women in Hawaii. Overall, 27% (372 of 1,378) of women were positive for anal HPV DNA at baseline compared with 29% (692 of 2,372) with cervical HPV DNA. Among women with paired anal and cervical samples, anal infection without accompanying cervical infection was observed in 14% (190 of 1,363). Concurrent anal and cervical HPV infections were observed in 13% (178 of 1,363) of women. Women with cervical HPV infection had >3-fold increased risk of concurrent anal infection. Concurrent anal and cervical HPV infection was most prevalent among the youngest women and steadily decreased through age 50 years. By contrast, the prevalence of anal infection alone remained relatively steady in all age groups. Compared with cervical infections, the overall distribution of HPV genotypes in the anus was more heterogeneous and included a greater proportion of nononcogenic types. A high degree of genotype-specific concordance was observed among concurrent anal and cervical infections, indicating a common source of infection. Nevertheless, the association of anal intercourse with anal HPV infection was limited to those women without accompanying cervical infection. The relationship of anal to cervical infection as described in this study has implications for the development of anal malignancies in women.

Of the many types of human papillomavirus (HPV), more than 30 infect the genital tract. The association between certain oncogenic (high-risk) strains of HPV and cervical cancer is well established. Although HPV is essential to the transformation of cervical epithelial cells, it is not sufficient, and a variety of cofactors and molecular events influence whether cervical cancer will develop. Early detection and treatment of precancerous lesions can prevent progression to cervical cancer. Identification of precancerous lesions has been primarily by cytologic screening of cervical cells. Cellular abnormalities, however, may be missed or may not be sufficiently distinct, and a portion of patients with borderline or mildly dyskaryotic cytomorphology will have higher-grade disease identified by subsequent colposcopy and biopsy. Sensitive and specific molecular techniques that detect HPV DNA and distinguish high-risk HPV types from low-risk HPV types have been introduced as an adjunct to cytology. Earlier detection of high-risk HPV types may improve triage, treatment, and follow-up in infected patients. Currently, the clearest role for HPV DNA testing is to improve diagnostic accuracy and limit unnecessary colposcopy in patients with borderline or mildly abnormal cytologic test results.

Background

Serial Analysis of Gene Expression (SAGE) is a new technique that allows a detailed and profound quantitative and qualitative knowledge of gene expression profile, without previous knowledge of sequence of analyzed genes. We carried out a modification of SAGE methodology (microSAGE), useful for the analysis of limited quantities of tissue samples, on normal human cervical tissue obtained from a donor without histopathological lesions. Cervical epithelium is constituted mainly by cervical keratinocytes which are the targets of human papilloma virus (HPV), where persistent HPV infection of cervical epithelium is associated with an increase risk for developing cervical carcinomas (CC).

Results

We report here a transcriptome analysis of cervical tissue by SAGE, derived from 30,418 sequenced tags that provide a wealth of information about the gene products involved in normal cervical epithelium physiology, as well as genes not previously found in uterine cervix tissue involved in the process of epidermal differentiation.

Conclusion

This first comprehensive and profound analysis of uterine cervix transcriptome, should be useful for the identification of genes involved in normal cervix uterine function, and candidate genes associated with cervical carcinoma.

Background

Cofactors might affect the risk of the rare progression from infection with carcinogenic human papillomavirus (HPV) to cervical premalignancy to invasive cancer. Some studies have observed that Chlamydia trachomatis infection is associated with increased risk for cervical cancer. In a large prospective cohort, we assessed the role of C trachomatis in cervical premalignancy and addressed confounding by HPV.

Methods

We identified 182 women with prevalent and 132 women with incident histological cervical intraepithelial neoplasia grade 2 (CIN2), grade 3 (CIN3), or cervical cancer (CIN2+) in the Costa Rica HPV Natural History Study. Control subjects were 995 (approximately 10% of the 10 049) subjects who were randomly selected from the same study. Cervical HPV status at enrollment was determined by MY09/MY11 polymerase chain reaction amplification and dot-blot hybridization. The presence of C trachomatis DNA in cervical exfoliated cells at enrollment was determined by a novel serovar-specific polymerase chain reaction–based C trachomatis detection and genotyping assay. Plasma drawn at enrollment from each subject was used to determine C trachomatis immunoglobulin G (IgG) status. Logistic regression was used to examine the association between C trachomatis and CIN2+, taking into account possible confounding by HPV.

Results

C trachomatis positivity at enrollment was associated with CIN2+ and concurrent and subsequent carcinogenic HPV infection. To account for confounding by HPV status, we restricted the analysis to women positive for carcinogenic HPV DNA at enrollment and found no association between C trachomatis status (as assessed by DNA or IgG) at enrollment and combined prevalent and/or incident CIN2+ (for C trachomatis DNA positivity, odds ratio = 0.77, 95% confidence interval = 0.42 to 1.41; for C trachomatis seropositivity, odds ratio = 1.09, 95% confidence interval = 0.85 to 1.41).

Conclusions

We found no association between C trachomatis status, as assessed by DNA or IgG, and risk of cervical premalignancy, after controlling for carcinogenic HPV-positive status. Previous positive associations between C trachomatis and cervical premalignancy could have been caused, in part, by an increased susceptibility to HPV infection.

Background

Cervical cancer is the second largest cause of cancer deaths in women worldwide. It is now evident that persistent infection with high-risk human papillomavirus (HPV) is necessary for the development and maintenance of cervical cancer. Thus, effective vaccination against HPV represents an opportunity to restrain cervical cancer and other important cancers. The FDA recently approved the HPV vaccine Gardasil for the preventive control of HPV, using HPV virus-like particles (VLP) to generate neutralizing antibodies against major capsid protein, L1. However, prophylactic HPV vaccines do not have therapeutic effects against pre-existing HPV infections and HPV-associated lesions. Furthermore, due to the considerable burden of HPV infections worldwide, it would take decades for preventive vaccines to affect the prevalence of cervical cancer. Thus, in order to speed up the control of cervical cancer and treat current infections, the continued development of therapeutic vaccines against HPV is critical. Therapeutic HPV vaccines can potentially eliminate pre-existing lesions and malignant tumors by generating cellular immunity against HPV-infected cells that express early viral proteins such as E6 and E7.

Objective

This review discusses the future directions of therapeutic HPV vaccine approaches for the treatment of established HPV-associated malignancies, with emphasis on current progress of HPV vaccine clinical trials.

Methods

Relevant literature is discussed.

Results/conclusion

Though their development has been challenging, many therapeutic HPV vaccines have been shown to induce HPV-specific antitumor immune responses in preclinical animal models and several promising strategies have been applied in clinical trials. With continued progress in the field of vaccine development, HPV therapeutic vaccines may provide a potentially promising approach for the control of lethal HPV-associated malignancies.

Certain high-risk (HR) human papillomavirus (HPV) types are a necessary cause for the development of cervical disorders. Women with persistent HR HPV infections have an increased risk of developing high-grade cervical lesions, compared with those who have no or low-risk HPV infections. Therefore, implementation of HPV detection into cervical screening programs might identify women at risk of cervical cancer. Several HPV detection methods with different sensitivities and specificities are available. Recently, a new PCR-based technique, the Roche AMPLICOR HPV Test, was developed. This test recognizes a group of 13 HR HPV types simultaneously. This study was undertaken to validate and compare HPV detection in 573 cervical scrape specimens by the AMPLICOR HPV Test and the INNO-LiPA HPV detection/genotyping assay (SPF10-LiPA system version 1). Human β-globin was not detected in nine specimens, which were therefore excluded from the comparison. Eleven scrape specimens containing HPV type 53 or 66 were also excluded from the comparison because these (probably) HR HPV types cannot be detected by the AMPLICOR HPV Test. The results of HPV detection by the Roche AMPLICOR HPV Test were confirmed by INNO-LiPA HPV detection/genotyping assay in 539/553 cases, showing an absolute agreement of 97.5% with a Cohen's kappa of 0.9327, indicating almost complete similarity of the two tests. Like the INNO-LiPA HPV detection/genotyping assay, the AMPLICOR HPV Test was sensitive, specific, feasible, and easy to handle. The value of the Roche AMPLICOR HPV Test with a broad-spectrum HR HPV detection has to be determined in prospective clinical studies.

Background

High-risk human papillomavirus (hrHPV) infections are causally related to cervical cancer development. The additional (epi)genetic alterations driving malignant transformation of hrHPV-infected cells however, are not yet fully elucidated. In this study we experimentally assessed the role of the PI3-kinase pathway and its regulator PIK3CA, which is frequently altered in cervical cancer, in HPV-induced transformation.

Methods

Cervical carcinomas and ectocervical controls were assessed for PIK3CA mRNA and protein expression by quantitative RT-PCR and immunohistochemical staining, respectively. A longitudinal in vitro model system of hrHPV-transfected keratinocytes, representing the immortal and anchorage independent phenotype, was assayed for PI3-kinase activation and function using chemical pathway inhibition i.e. LY294002 treatment, and PIK3CA RNA interference. Phenotypes examined included cellular viability, migration, anchorage independent growth and differentiation. mRNA expression of hTERT and HPV16 E6E7 were studied using quantitative RT-PCR and Northern blotting.

Results

Cervical carcinomas showed significant overexpression of PIK3CA compared to controls. During HPV-induced transformation in vitro, expression of the catalytic subunit PIK3CA as well as activation of downstream effector PKB/AKT progressively increased in parallel. Inhibition of PI3-kinase signalling in HPV16-transfected keratinocytes by chemical interference or siRNA-mediated silencing of PIK3CA resulted in a decreased phosphorylation of PKB/AKT. Moreover, blockage of PI3-kinase resulted in reduced cellular viability, migration, and anchorage independent growth. These properties were accompanied with a downregulation of HPV16E7 and hTERT mRNA expression. In organotypic raft cultures of HPV16- and HPV18-immortalized cells, phosphorylated PKB/AKT was primarily seen in differentiated cells staining positive for cytokeratin 10 (CK10). Upon PI3-kinase signalling inhibition, there was a severe impairment in epithelial tissue development as well as a dramatic reduction in p-PKB/AKT and CK10.

Conclusion

The present data indicate that activation of the PI3-kinase/PKB/AKT pathway through PIK3CA regulates various transformed phenotypes as well as growth and differentiation of HPV-immortalized cells and may therefore play a pivotal role in HPV-induced carcinogenesis.