Prognostic importance of DNA from human papillomavirus in patients with oral squamous cell carcinoma.

BACKGROUND: Survival of patients with oral squamous cell carcinoma (OSCC) is generally low, with the likelihood of locoregional recurrence or disease progression (LR/DP). Knowledge of prognostic factors for survival is key to achieving an understanding and increased survival. The present study aimed to identify prognostic factors for patients with OSCC, especially the presence of DNA from human papillomavirus (HPV). MATERIAL AND METHODS: Retrospective cohort study including 119 patients with OSCC treated at the National Cancer Institute in Mexico City (2009-2013). Clinical information was obtained from patient records including LR/DP. Formalin-fixed, paraffin-embedded tissues were obtained and used for detecting DNA from different types of HPV. Potential prognostic factors for Overall Survival (OS) were analyzed using the Cox proportional hazards model. RESULTS: After model adjustment, factors associated with longer OS were a pre-treatment platelet count above 400,000/mm3 (HR=0.09, p=0.026) and response to primary treatment (HR=0.26, p=0.001). HPV DNA was present in 23 (19.3%) of the patients and importantly, type 16 found in 19 of them. Although survival of HPV-positive patients was longer, difference was not significant. However, among patients with LR/DP, HPV positivity was significantly associated with increased survival (HR=0.23, p=0.034). Importantly, survival was significantly different for HPV-positive patients with LR/DP > 6 months (HR=0.20, p=0.002), had higher absolute lymphocyte count at start of treatment (HR=0.50, p=0.028) or had local rescue treatment (HR=0.24, p=0.019). CONCLUSIONS: Although HPV positivity was not associated with a longer OS of OSCC patients, a better prognosis was significantly associated with HPV positivity and recurring or progressing disease, particularly with HPV type 16.

Resveratrol decreases Rad51 expression and sensitizes cisplatin­resistant MCF­7 breast cancer cells.

Resveratrol (RES), a polyphenol compound with anti­proliferative properties, has been previously evaluated for its beneficial effects against a variety of tumour cells. The current study elucidated the means by which RES enhances the anti­proliferative effects of cisplatin (CIS) on MCF­7 cells, focusing on the inhibitory effects on DNA repair of double­strand breaks (DSBs). Chemoresistant MCF­7 cells (MCF­7R) were generated by continuous exposure to low concentrations of CIS (10 µM CIS­IC40) during 5 passages, with the IC50 value increasing ~3­fold. Using an MTT assay, we estimated the changes in IC50 for CIS in MCF­7, T47­D, MDA­MB­231 and MCF­7R cells in the presence of RES. The relative transcript level of Nbs­1, Mre­11 and Rad­50 genes was assessed using RT­qPCR analysis. Rad51 and H2AX [pSer139] protein expression was determined by western blot analysis. RES at 50 and 100 µM significantly enhanced the anti­proliferative effects of CIS in both MCF­7 and MCF­7R cells, decreasing the IC50 values for CIS to one­tenth and one­sixth, respectively. A total of 100 µM RES decreased the relative transcript levels of homologous recombination (HR) initiation complex components and the Rad51 protein level in MCF­7 and MCF­7R cells. After 48 h of CIS DNA damage, the levels of Rad51 protein increased, but this effect was inhibited by 100 µM RES. RES also maintained serine 139 phosphorylation of histone H2AX, suggesting that RES prevents the repair of DSBs. It was observed that RES exerts an antagonistic effect over CIS on the activation of Rad51 and sustained phosphorylation of H2AX. The results suggest that RES in combination with DNA damage­based therapy has potential as a strategy to overcome resistance and provide much safer and more effective treatment for breast cancer.

Human papillomavirus load in benign HPV-associated oral lesions from HIV/AIDS individuals.

BACKGROUND: Although HPV emerged as a crucial carcinogenic and prognostic biomarker in head and neck cancer, and considering the increase in HPV-associated oral lesions (HPV-OLs) in HIV individuals, molecular information about HPV-OLs is scarce; thus, our aim was to determine viral loads in HPV-OLs from HIV/AIDS individuals. METHODS: HIV/AIDS subjects with HPV-OL were included in this cross-sectional study. Following informed consent, biopsies were obtained. HPV detection and typing were carried out by PCR and sequencing (MY09/11, GP5+/6+). HPV-13 and HPV-32 loads were determined by a high-resolution melting assay. For statistical analysis, X2 , Fisher's exact, and Mann-Whitney U tests were applied, using SPSS software (v.23). RESULTS: Twenty-nine HIV subjects (median age 38 years, 93% males) were included. Most were AIDS individuals (72.4%) under HAART (89.7%). Twenty-two (75.9%) participants had more than one HPV-OL (four with florid presentations), mostly multifocal epithelial hyperplasia (62%), being HPV-13 (26%) and HPV-32 (31%) the most frequent types. HPV load was higher in individuals with multiple HPV-OLs than in solitary lesions (4.9 vs. 3.2 Log10 copies/ml, p = .090) and in HPV-32+ than in HPV-13+ (8.3 vs. 6.4 Log10 copies/ml, p = .014). CONCLUSIONS: Multiple HPV-OLs showed high HPV loads, possibly indicating transcriptional activity of the virus; however, in the HIV setting, the individual and local immunological response could be the key process.

Risk allelic load in Th2 and Th3 cytokines genes as biomarker of susceptibility to HPV-16 positive cervical cancer: a case control study.

BACKGROUND: Alterations in the host cellular immune response allow persistent infections with High-Risk Human Papillomavirus (HR-HPV) and development of premalignant cervical lesions and cervical cancer (CC). Variations of immunosuppressive cytokine levels in cervix are associated with the natural history of CC. To assess the potential role of genetic host immunity and cytokines serum levels in the risk of developing CC, we conducted a case-control study paired by age. METHODS: Peripheral blood samples from patients with CC (n = 200) and hospital controls (n = 200), were used to evaluate nine biallelic SNPs of six cytokine genes of the adaptive immune system by allelic discrimination and cytokines serum levels by ELISA. RESULTS: After analyzing the SNP association by multivariate logistic regression adjusted by age, CC history and smoking history, three Th2 cytokines (IL-4, IL-6 and IL-10) and one Th3 (TGFB1) cytokine were significantly associated with CC. Individuals with at least one copy of the following risk alleles: T of SNP (-590C > T IL-4), C of SNP (-573G > C IL-6), A of SNP (-592C > A IL-10), T of SNP (-819C > T IL-10) and T of SNP (-509C > T TGFB1), had an adjusted odds ratio (OR) of 2.08 (95 % CI 1.475-2.934, p = 0.0001), an OR of 1.70 (95 % CI 1.208-2.404, p = 0.002), an OR of 1.87 (95 % CI 1.332-2.630, p = 0.0001), an OR of 1.67 (95 % CI 1.192-2.353, p = 0.003) and an OR of 1.91 (95 % CI 1.354-2.701, p = 0.0001), respectively, for CC. The burden of carrying two or more of these risk alleles was found to have an additive effect on the risk of CC (p trend = 0.0001). Finally, the serum levels of Th2 and Th3 cytokines were higher in CC cases than the controls; whereas IFNG levels, a Th1 cytokine, were higher in controls than CC cases. CONCLUSION: The significant associations of five SNPs with CC indicate that these polymorphisms are potential candidates for predicting the risk of development of CC, representing a risk allelic load for CC and can be used as a biomarker of susceptibility to this disease.

The P2RY2 Receptor Induces Carcinoma Cell Migration and EMT Through Cross-Talk With Epidermal Growth Factor Receptor.

Extracellular nucleotides are signaling elements present in the tumor microenvironment; however, their role in tumor growth is not completely understood. In the present study, we asked whether nucleotides regulate cell migration in ovarian carcinoma-derived cells. We observed that 100 µM UTP induced migration in SKOV-3 cells (1.57 ± 0.08 fold over basal), and RT-PCR showed expression of transcripts for the P2RY2 and P2RY4 receptors. Knockdown of P2RY2 expression in SKOV-3 cells (P2RY2-KD) abolished the UTP-induced migration. The mechanism activated by UTP to induce migration involves transactivation of the epidermal growth factor receptor (EGFR) since we observed that the EGFR kinase inhibitor AG1478 and the PI3K inhibitor Wortmannin inhibit this response (to 0.76 ± 0.23 and 0.46 ± 0.14 relative to the control, respectively). In agreement with these observations, UTP was able to modify the phosphorylation state of the EGFR; likewise, the induction of ERK1/2 phosphorylation promoted by UTP was abolished by a 30-60 min treatment with AG1478. Our data also suggested that the enhanced cell migration involves the epithelium to mesenchymal transition (EMT) process, since a 12 h stimulation of SKOV-3 cells with 100 µM UTP showed an increase in vimentin and SNAIL protein levels (459.8 ± 132.4% over basal for SNAIL). Interestingly, treatment with apyrase (10 U/mL) reduces the migration of control cells and induces a considerable enrichment of E-cadherin in the cell-cell contacts, favoring an epithelial phenotype and strongly suggesting that the nucleotides released by tumor cells and acting through the P2RY2 receptor are potential regulators of invasiveness.

Cervical cancer-associated promoter polymorphism affects akna expression levels.

Cervical cancer (CC) is responsible for >260,000 deaths worldwide each year. Efforts are being focused on identifying genetic susceptibility factors, especially in genes related to the immune response. Akna has been proposed to be one of them, but data regarding its functional role in the disease is scarce. Supporting the notion of akna as a CC susceptibility gene, we found two polymorphisms associated with squamous intraepithelial lesion (SIL) and CC; moreover, we identified an association between high akna expression levels and CC and SIL, but its direction differs in each disease stage. To show the potential existence of a cis-acting polymorphism, we assessed akna allelic expression imbalance for the alleles of the -1372C>A polymorphism. We found that, regardless of the study group, the number of transcripts derived from the A allele was significantly higher than those from the C allele. Our results support the hypothesis that akna is a CC susceptibility genetic factor and suggest that akna transcriptional regulation has a role in the disease. We anticipate our study to be a starting point for in vitro evaluation of akna transcriptional regulation and for the identification of transcription factors and cis-elements regulating AKNA function that are involved in carcinogenesis.

Differential expression of functionality markers in mid-secretory endometrium of infertile women under treatment with ovulation-inducing agents.

OBJECTIVES: To analyze the expression of protein markers related to cell proliferation and death, as well as oestrogen and progesterone receptors in the endometrium of infertile women with hypothalamic-pituitary dysfunction treated with clomiphene citrate (CC) or recombinant follicle-stimulating hormone (rFSH), and compare them with ovulatory women. STUDY DESIGN: The study included 12 control ovulatory women and 29 anovulatory women, 19 of whom underwent ovulation induction with CC (n = 12) or rFSH (n = 5). Endometrial biopsies were obtained by Pipelle during the mid-secretory phase. Samples were stained with haematoxylin and eosin. Immunohistochemistry of proteins related to cell proliferation and cell death, as well as steroid receptors, was undertaken, and apoptosis was determined using TUNEL analysis. RESULTS: Immunohistochemical analysis of Ki67 expression showed significantly higher expression in the glandular epithelium of ovulatory women compared with the other groups. Glandular oestrogen receptor α expression was significantly lower in rFSH-treated women compared with ovulatory women. The number of apoptotic cells, Bax expression and progesterone receptor expression were similar in all groups. In contrast, Bcl-2 expression was significantly lower in the glandular epithelium of rFSH-treated women. CONCLUSIONS: In infertile women with hypothalamic-pituitary dysfunction, treatment with ovulation-inducing agents modifies the expression of proteins involved in cell proliferation and death, as well as the expression of steroid hormone receptors in the endometrium. These differences may help to explain, at the molecular level, the functionality of the endometrium during the implantation window, and may help to optimize pregnancy rates obtained with these treatments.

Association between age and high-risk human papilloma virus in Mexican oral cancer patients.

OBJECTIVE: Studies reporting low prevalence of HPV in OSCC with declining age at presentation are increasing. The aim of this study was to determine the prevalence of HPV in a group of OSCC cases and controls in a Mexican population. METHODS: The matched case-control study included 80 OSCC cases and 320 controls. HPV/DNA presence was evaluated through PCR amplification using three sets of consensus primers for the L1 gene. A conditional logistic regression analysis was carried out for the matched OSCC cases and controls. Interactions between risk factors and OCSS were tested in the construction process of the models. RESULTS: HPV prevalence was 5% in OSCC cases and 2.5% in controls. HPV-detected types were 16, 18 and 56. According to conditional logistics regression model, an association was detected between HR-HPV and OSCC. All HR-HPV-positive OSCC cases corresponded to young patients (<45 years), non-smokers and non-alcohol drinkers. CONCLUSIONS: The HR-HPV can be a contributing factor to oral carcinogenesis, especially in younger individuals without known risk factors such as tobacco and alcohol.

Differential expression of the P2X7 receptor in ovarian surface epithelium during the oestrous cycle in the mouse.

Purinergic signalling has been proposed as an intraovarian regulatory mechanism. Of the receptors responsible for purinergic transmission, the P2X7 receptor is an ATP-gated cationic channel that displays a broad spectrum of cellular functions ranging from apoptosis to cell proliferation and tumourigenesis. In the present study, we investigated the functional expression of P2X7 receptors in ovarian surface epithelium (OSE). P2X7 protein was detected in the OSE layer of the mouse, both in situ and in primary cultures. In cultures, 2'(3')-O-(4-Benzoylbenzoyl)adenosine-5'-triphosphate (BzATP) activation of P2X7 receptors increased [Ca(2+)]i and induced apoptosis. The functionality of the P2X7 receptor was investigated in situ by intrabursal injection of BzATP on each day of the oestrous cycle and evaluation of apoptosis 24h using the terminal deoxyribonucleotidyl transferase-mediated dUTP-fluorescein nick end-labelling (TUNEL) assay. Maximum effects of BzATP were observed during pro-oestrus, with the effects being blocked by A438079, a specific P2X7 receptor antagonist. Immunofluorescence staining for P2X7 protein revealed more robust expression during pro-oestrus and in OSE regions behind the antral follicles, strongly supporting the notion that the differences in apoptosis can be explained by increased receptor expression, which is regulated during the oestrous cycle. Finally, P2X7 receptor expression was detected in the OSE layer of human ovaries, with receptor expression maintained in human ovaries diagnosed with cancer, as well as in the human ovarian carcinoma SKOV3 cell line.

HPV 16 E2 protein induces apoptosis in human and murine HPV 16 transformed epithelial cells and has antitumoral effects in vivo.

OBJECTIVE: Our aims were to examine the ability of the human papillomaviruse (HPV) 16 E2 protein to induce apoptosis in a murine HPV-transformed cell line, and to evaluate its antitumor properties on HPV-associated tumors in vivo in immunocompetent mice. METHODS: HPV-transformed murine BMK-16/myc cells and human SiHa cells were transfected with the HPV 16 E2 gene to examine the effects of the E2 protein on cell growth and on the E6 and E7 oncogenes as well as DNA fragmentation and activation of the extrinsic pathway of apoptosis. Finally, to test the antitumor effect of the E2 protein on an experimental mouse tumor model, we generated a recombinant adenovirus expressing the E2 protein. RESULTS: The E2 protein inhibited the growth of SiHa and BMK-16/myc cell lines, and repressed the E6 and E7 oncogenes. Moreover, the E2 protein induced DNA fragmentation and apoptosis through activation of caspases 8 and 3 in BMK-16/myc cells. On the other hand, E2 also showed antitumor effects in vivo. CONCLUSIONS: Our findings indicate that E2 exerts pro-apoptotic activity in a murine HPV-transformed cell line as well as an antitumor effect in vivo.

Mucosal IgG and IgA responses to human papillomavirus type 16 capsid proteins in HPV16-infected women without visible pathology.

Human papillomavirus type 16 (HPV16) may infect the cervical epithelium without producing pathological changes for a long time. To investigate if mucosal antibodies are induced in HPV16-infected women without visible pathology, cervical mucus from HPV16-infected patients with and without evident pathology, along with mucus from uninfected women were analyzed for the presence of mucosal IgG and secretory IgA (sIgA) antibodies to HPV16 capsid proteins by ELISA. sIgA and IgG antibodies were found in a significantly higher proportion of infected patients compared with uninfected women (p < 0.0001). sIgA antibodies were present in 13.1% of infected patients without visible pathology, the proportion of positivity increased to 27.0% in patients with visible pathology (p = 0.001). Mucosal IgG response was observed in 6.5% of patients without and 27.5% of patients with visible pathology (p = 0.00005). The antibody mean signal strength was significantly higher in patients with than in patients without pathological evidence (p < 0.005). In conclusion, both sIgA and IgG are found in patients without pathological signs of infection, however, the response increases significantly in patients with pathological evidence, suggesting that the appearance of these changes might be associated with a more vigorous antibody-mediated mucosal reaction.

High-level expression of NRAMP1 in peripheral blood cells and tuberculous granulomas from Mycobacterium bovis-infected bovines.

By Western blotting, we demonstrate high-level expression of NRAMP1 proteins in peripheral blood cells and granulomas of Mycobacterium bovis-infected bovines. Immunohistochemistry of granulomatous lesions showed heavily labeled epithelioid macrophages and Langhans cells. These data suggest that M. bovis infection enhances NRAMP1 expression and that active tuberculosis can occur despite this response.

Asian-American variants of human papillomavirus 16 and risk for cervical cancer: a case-control study.

BACKGROUND: Human papillomavirus 16 (HPV16) has a number of variants, each with a different geographic distribution and some that are associated more often with invasive neoplasias. We investigated whether the high incidence of cervical cancer in Mexico (50 cases per 100 000 women) may be associated with a high prevalence of oncogenic HPV16 variants. METHODS: Cervical samples were collected from 181 case patients with cervical cancer and from 181 age-matched control subjects, all from Mexico City. HPV16 was detected with an E6/E7 gene-specific polymerase chain reaction, and variant HPV classes and subclasses were identified by sequencing regions of the E6 and L1/MY genes. Clinical data and data on tumor characteristics were also collected. All statistical tests were two-sided. RESULTS: HPV16 was detected in cervical scrapes from 50.8% (92 of 181) of case patients and from 11% (20 of 181) of control subjects. All HPV16-positive samples, except one, contained European (E) or Asian-American (AA) variants. AA and E variants were found statistically significantly more often in case patients (AA = 23.2% [42 of 181]; E = 27.1% [49 of 181]) than in control subjects (AA = 1.1% [two of 181]; E = 10% [18 of 181]) (P<.001 for case versus control subjects for either E or AA variants, chi2 test). However, the frequency of AA variants was 21 times higher in cancer patients than in control subjects, whereas that ratio for E variants was only 2.7 (P =.006, chi2 test). The odds ratio (OR) for cervical cancer associated with AA variants (OR = 27.0; 95% confidence interval [CI] = 6.4 to 113.7) was higher than that associated with E variants (OR = 3.4; 95% CI = 1.9 to 6.0). AA-positive case patients (46.2 +/- 12.5 years [mean +/- standard deviation]) were 7.7 years younger than E-positive case patients (53.9 +/- 12.2 years) (P =.004, Student's t test). AA variants were associated with squamous cell carcinomas and adenocarcinomas, but E variants were associated with only squamous cell carcinomas (P =.014, Fisher's exact test). CONCLUSIONS: The high frequency of HPV16 AA variants, which appear to be more oncogenic than E variants, might contribute to the high incidence of cervical cancer in Mexico.

Cytotoxic effect of three arsenic compounds in HeLa human tumor and bacterial cells.

Numerous epidemiological studies suggest that arsenic (As) compounds are carcinogens, however, recent data have renewed the interest in their anticarcinogenic properties. The cytotoxic effects of three arsenic compounds were assessed: sodium arsenite, sodium arsenate and sodium cacodylate, representing the trivalent and pentavalent species of arsenic, along with a dimethylated pentavalent arsenic species. HeLa cells and Salmonella typhimurium (strains TA98 and TA100) were exposed to As compounds and the cytotoxic effects were evaluated. Alterations on RNA and DNA synthesis in HeLa cells were also examined. All arsenic compounds produced a dose-dependent inhibition on colony formation and DNA synthesis in HeLa cells, yet any of them significantly influenced RNA synthesis in these cells. No evidence of arsenic-induced mutagenicity or antimutagenicity was observed using the Ames assay. In bacterial cells, only sodium arsenite caused a dose-dependent inhibition of colony formation.Collectively, these results indicate that in both, HeLa and S. typhimurium cell systems, only trivalent sodium arsenite can act as an effective inhibitor of cell growth. The possible mechanism(s) of the cytotoxic effect of arsenite in these two different cell systems might be due to its reactivity with intracellular sulfhydryl groups.

Cervical secretory immunoglobulin A to human papillomavirus type 16 (HPV16) from HPV16-infected women inhibit HPV16 virus-like particles-induced hemagglutination of mouse red blood cells.

Secretory immunoglobulin A (sIgA) antibodies are the first line of defence at the genital mucosa, and are thought to hinder viral infections by binding to conformational epitopes on the viral capsid. To investigate if cervical sIgA binds to conformational epitopes of the Human papillomavirus type 16 (HPV16), cervical mucus samples from 109 HPV16-infected patients were examined in a HPV16 virus-like particles-induced hemagglutination inhibition assay. 48 (44.1%) patients were able to inhibit hemagglutination. Inhibition of hemagglutination was associated with the presence of sIgA (P=0.001). In conclusion, naturally occurring cervical anti-HPV16 sIgA binds to and hinders conformational epitopes on the viral capsid, suggesting that these antibodies might have a neutralizing capacity.

High prevalence of oncogenic human papillomavirus in the genital tract of women with human immunodeficiency virus.

OBJECTIVE: The goal of this study was to determine the prevalence of human papillomavirus (HPV) and squamous intraepithelial lesions (SILs) in women infected with human immunodeficiency virus (HIV) in Mexico. METHODS: Cases included women who were positive for human immunodeficiency virus (HIV) and accepted to participate. There were two control groups in this study: group A, heterosexual partners of HIV+ men; group B, commercial sex workers. Gynecologic examination was performed in all participants. Also, a cervical smear with colposcopy and a sample for detection of HPV DNA by polymerase chain reaction (PCR) were obtained in all subjects, as were CD4+ counts. Relative risks (RR) and 95% confidence interval were calculated. RESULTS: Eighty-five HIV+ women agreed to participate in this study; the route of HIV infection was heterosexual in 78.8%; transfusion in 8.2%; paid donors in 3.5%; and 9.4% unknown. A total of 9 controls were included: 4 from group A and 5 from group B. HPV DNA was detected by PCR in 57 (69%) cases and in 26 (29%) controls from both groups (P < 0.0001). The RR of HPV infection was 5.5 (2.7-11.5). Also, a significant difference in the prevalence of high-risk HPV types was observed between cases and controls, RR = 12.8 (4.07-42.9). These associations were independent of CD4+ counts and antiretroviral therapy. No association was observed between HIV infection and the risk for high-grade SIL. CONCLUSIONS: We observed a high prevalence of oncogenic HPV types in HIV-positive women. These women should be screened regularly for early diagnosis of premalignant lesions and prevention of cervical cancer.

Human tumor growth is inhibited by a vaccinia virus carrying the E2 gene of bovine papillomavirus.

BACKGROUND: Papillomavirus is the etiologic agent associated with cervical carcinoma. The papilloma E2 protein is able to regulate negatively the expression of E6 and E7 papilloma oncoproteins. Therefore, a new, highly attenuated vaccinia virus known as modified vaccinia virus Ankara (MVA), which carries the papillomavirus E2 gene, was used for the treatment of tumors associated with human papillomavirus. METHODS: Analysis of expression of the E2 gene from the recombinant vaccinia virus was performed by reverse transcription-polymerase chain reaction of RNA isolated from infected cells. Detection of the E2 protein was done by immunoprecipitation from proteins labeled with [(35)S]-methionine, isolated from infected cells. The therapeutic effect of the MVA E2 recombinant virus over human tumors was tested in nude mice bearing tumors generated by inoculation of HeLa cells. Series of 10 nude mice with tumors of different sizes were injected with MVA, MVA E2, or phosphate-buffered saline. Tumor size was monitored every week to assess growth. RESULTS: The MVA E2 recombinant virus efficiently expressed the E2 protein in BS-C-1 cells. This protein was able to repress, in vivo, the papillomavirus P105 promoter, which controls the expression of the E6 and E7 oncoproteins. In nude mice the MVA E2 virus reduced tumor growth very efficiently. In contrast, tumors continued to grow in mice treated with MVA or PBS. The life expectancy of MVA E2-treated mice was also increased three- to fourfold compared with that of animals that received MVA or PBS. CONCLUSIONS: The growth of human tumors was efficiently inhibited by the MVA E2 recombinant vaccinia virus. The absence of side effects in treated animals suggested that the MVA E2 virus is a safe biologic agent that could in the future be used in humans for the treatment of cervical carcinoma.

[Molecular variants of human papillomavirus (HPV) types 16 and 18 in adenocarcinomas of the cervix]. / Variantes moleculares de virus papiloma humano (HPV) tipos 16 y 18 en adenocarcinomas de cérvix.

Human Papillomavirus (HPV), placebo clinical trial particularly types 16 and 18, are considered human carcinogens since an etiological association has been demonstrated between these viruses and the development of cervical cancer. While the viral role in squamous carcinoma has been largely studied, the information available on adenocarcinoma is scarce, partly because of its lower frequency. In this paper we investigated the presence of HPV types and intratype variants in adenocarcinomas of the cervix. A total of 23 archive samples, fixed and paraffin embedded biopsies, were included. The detection and viral typing was performed by generic PCR and subsequent single stranded conformational polymorphism analysis (SSCP). Genetic variability was investigated in a 450 bp-fragment corresponding to L1 gene by post-PCR direct sequencing. We detected 11 HPV 16 positive samples (9 prototypes and 2 variants: 1 European and 1 Asiatic-American), 10 HPV 18 (9 prototypes and 1 European variant), 1 HPV 31 and 1 negative. The high risk HPV association with this neoplasia was confirmed with a high prevalence (43%) of HPV 18, (but) without predominance over the other types as previously published. The demonstrated variability in L1 protein epitopes originated aminoacidic changes which could have implications on the immune response and therefore should be considered in a vaccine design.

[Ha-ras and p53 gene mutations scanned by PCR-SSCP in premalignant and malignant lesions of the uterine cervix associated with human papillomavirus]. / Mutaciones en genes Ha-ras y p53 detectadas mediante PCR-SSCP en lesiones premalignas y malignas de cuello uterino asociadas con virus papiloma humano.

The aim of this study w trial randomized as to investigate the frequencies of human papillomavirus (HPV) and mutation in Ha-ras oncogene and tumour suppressor p53 gene in cervical cancer and precursor lesions. A total of 30 invasive carcinomas (IC), 36 cervical intraepithelial neoplasia grade III (CIN III) and 12 normal tissues adjacent to the tumor (NT) were included. HPV typification and scanning of possible mutations in Ha-ras and p 53 genes were made by SSCP-PCR. The IC cases showed 93% HPV positivity, 41% having mobility shifts for Ha-ras mutations and 17% for p53 mutations while in CIN III, these percentages were 80%, 18% and 11%, respectively. In normal tissues HPV frequency was 17%. All Ha-ras mutated samples were HPV positive but 33% of p53 mutated cases were HPV negative. All mutations were heterozygous. HPV 16 was more prevalent (44%) than HPV 18 (15%) and the high rate of undetermined HPV types (18%) would indicate the circulation in our country of other types different from the assayed HPV controls (6, 11, 16, 18, 31 and 33), being variants or mixed infections. The low frequency of p53 mutations (17%) strengthens the view that wild type p53 inactivation by HPV probably plays a major role in the pathogenesis of cervical cancer. Because mutated Ha-ras was found in HPV associated premalignant lesions, we speculate that it represents an early marker for progression. Our findings provide additional evidence for an interactive effect between high risk types of HPV and oncogene activation in the development of uterine cervical cancer.

Variantes moleculares de virus papiloma humano (HPV) tipos 16 y 18 en adenocarcinomas de cérvix. / [Molecular variants of human papillomavirus (HPV) types 16 and 18 in adenocarcinomas of the cervix]

Human Papillomavirus (HPV), placebo clinical trial particularly types 16 and 18, are considered human carcinogens since an etiological association has been demonstrated between these viruses and the development of cervical cancer. While the viral role in squamous carcinoma has been largely studied, the information available on adenocarcinoma is scarce, partly because of its lower frequency. In this paper we investigated the presence of HPV types and intratype variants in adenocarcinomas of the cervix. A total of 23 archive samples, fixed and paraffin embedded biopsies, were included. The detection and viral typing was performed by generic PCR and subsequent single stranded conformational polymorphism analysis (SSCP). Genetic variability was investigated in a 450 bp-fragment corresponding to L1 gene by post-PCR direct sequencing. We detected 11 HPV 16 positive samples (9 prototypes and 2 variants: 1 European and 1 Asiatic-American), 10 HPV 18 (9 prototypes and 1 European variant), 1 HPV 31 and 1 negative. The high risk HPV association with this neoplasia was confirmed with a high prevalence (43

) of HPV 18, (but) without predominance over the other types as previously published. The demonstrated variability in L1 protein epitopes originated aminoacidic changes which could have implications on the immune response and therefore should be considered in a vaccine design.