Rosa Santopietro

Human Papillomavirus Testing with the Hybrid Capture 2 Assay and PCR as Screening Tools

ABSTRACT The recognition of high-risk human papillomaviruses (HPVs) as etiological agents of cervical cancer has increased the demands to use testing for HPV for the detection of abnormal cervical smears and for cervical cancer screening. The present study compared the performance of the Hybrid Capture 2 (HC2) assay with that of PCR for the detection of significant cervical lesions in 1,511 women with different risks for HPV infections in three New Independent States of the former Soviet Union. The results showed that the level of agreement between the HC2 assay and PCR was substantial, with a kappa (Cohen) value of 0.669 (95% confidence interval [CI], 0.629 to 0.709). Of the 228 samples with discrepant results, 92 were positive by the HC2 assay but negative by PCR, whereas 136 samples were PCR positive but HC2 assay negative. The positive predictive values (PPVs) of the HC2 assay and PCR in detecting high-grade intraepithelial lesions (HSILs) were 4.5% (95% CI, 3.5 to 5.5%) and 3.6% (95% CI, 2.7 to 4.5%), respectively, and the negative predictive values (NPVs) were 99.6% (95% CI, 99.3 to 99.9%) and 99.3% (95% CI, 98.9 to 99.7%), respectively. The sensitivities of the HC2 assay and PCR for the detection of HSILs were 85.2 and 74.0%, respectively, and the specificities were 67.2 and 64.1%, respectively. In receiver operating characteristic (ROC) analysis, the performance of the HC2 assay for the detection of HSILs was excellent (P = 0.0001); the area under the ROC analysis curve was 0.858 (95% CI, 0.811 to 0.905), and the optimal balance between sensitivity (86.5%) and specificity (80%) was obtained with an HC2 assay cutoff level of 15.6 relative light units/positive control. Use of this cutoff would increase the specificity of the HC2 assay to 80.0% without compromising sensitivity. In conclusion, the results of PCR and the HC2 assay were concordant for 85% of samples, resulting in substantial reproducibility. Both tests had low PPVs, equal specificities, and equal (almost 100%) NPVs for the detection of HSILs; but the sensitivity of the HC2 assay was slightly better.

Macrophage migration inhibitory factor in the human endometrium : Expression and localization during the menstrual cycle and early pregnancy

Abstract Macrophage migration inhibitory factor (MIF) was discovered as an activated T-lymphocyte-derived protein that inhibits the random migration of macrophages in vitro. Subsequently, knowledge of the physiological actions of MIF was extended to include its role as a proinflammatory cytokine that affects several functions of macrophages and lymphocytes. Previous reports have suggested an involvement of MIF in reproduction. However, no data are currently available on the presence of this cytokine in the human endometrium. In this study, the expression and tissue localization of MIF was evaluated in specimens of cycling endometrium, first trimester placenta bed biopsy, and isolated endometrial glands by Western blot analysis, immunohistochemistry, ELISA, and reverse transcription-polymerase chain reaction. The results demonstrated that MIF is expressed in human endometrium across the menstrual cycle and in early pregnancy. Immunohistochemical localization identified the protein in glandular epithelium, in stromal and predecidualized stromal cells of cycling endometrium, as well as in the decidua of first-trimester placenta. The proinflammatory features and specific actions of MIF on lymphoid cells suggest its potential involvement in several aspects of endometrial physiology.

Small thymus in very low birth weight infants born to mothers with subclinical chorioamnionitis

Chorioamnionitis, a major cause of preterm birth with significant neonatal morbidity and mortality, frequently occurs in mothers who are free of symptoms. A combined clinical, radiologic, and pathologic study of 129 very low birth weight infants indicated a significant association between a markedly decreased thymic size at birth and subclinical chorioamnionitis.

Nuclear morphometry as an important prognostic factor in stage I renal cell carcinoma

Although 60% of Stage I renal carcinoma patients die from tumor within 5 years postoperatively, a considerable percentage survive that period. Nuclear grading can help to predict the outcome, but many of the patients are Grade 2, and the prognosis of this subclass is uncertain. Therefore, nuclear morphometry was carried out in 41 patients with Stage I renal cell carcinoma. Of these, 24 died within 5 years and 17 have survived that period. Using a mean nuclear area of 32 μm2 as the decision threshold, none of the 24 short‐term survivors are below that threshold and three of the long‐term survivors exceed that value (18% false‐positives) (99% confidence limit). Separate analysis with sets for learning and testing and Grade 2 patients gave similar results. The results show the essential prognostic value of morphometry in this set of patients with Stage I renal cell carcinoma.

An interlaboratory study to determine the presence of human papillomavirus DNA in esophageal carcinoma from China

Esophageal‐carcinoma samples originating from the high‐incidence area of China were tested in 2 different laboratories, each using a different degenerate PCR approach. Results confirmed the notion that none of the PCR approaches available for HPV‐DNA detection today, is optimal for detecting all known HPV types at equal sensitivity and specificity. In combining results obtained in both laboratories, HPV DNA was demonstrated in 20/117 (17.1%) esophageal‐carcinoma samples analyzed. HPV DNA was detected in 3/70 (4.3%) diagnostic biopsies, 7/23 (30.4%) surgical specimen and 10/24 (41.6%) cytological scrapings originating from the entire surface of the esophagus. Mucosotropic HPV types were present in 7/117 (6%) samples, only 3 being of the high‐risk types (HPV 16, 18, 33). Other mucosal types found were HPV 6, 11, 13, 53 and 54. Cutaneous HPV types were present in 14/117 (12.0%) samples. HPVs 20 and 38 were present in 3 (2.6%) of the total samples and, in each case, together with another HPV type within one lesion. Two putative new HPV types, DL347 and DL 369, were identified. Int. J. Cancer 81:225–228, 1999.

The effects of HIV-1 Tat protein on cell cycle during cervical carcinogenesis

The role of HPV in the carcinogenesis of intraepithelial and invasive anogenital lesions is currently well established. E6 and E7 oncoproteins of high risk HPV genotypes are known to inactivate p53 and pRb pathways. Several studies have described an increased prevalence and recurrence of both cervical HPV infection and invasive cervical cancer among HIV-1 positive women compared to HIV-1 negative cases. For these reasons, cervical cancer is considered an AIDS-defining neoplasm. Unlike other AIDS-associated neoplasms, the occurrence of cervical cancer is independent of immune suppression. HIV-1 infection in patients with high grade precancerous lesions and cervical cancers results in a therapy refractory and more aggressive disease phenotype, which is not yet well understood at the molecular level. An upregulation of HPV E6 and E7 gene expressions by HIV-1 proteins such as Tat has been documented by some authors. However, the role of HIV-1 in cervical carcinomas is still unclear. It is already known that HIV-1 Tat protein is able to influence cell cycle progression. Altogether, these facts led us to investigate the effects of Tat on the expression of cell cycle regulator genes. After transfection of HeLa cells with Tat, we analyzed the expression of cell cycle regulators from these cells by IHC and Real-time PCR. A significant reduction in the expression of cell cycle inhibitors of transcription and an increase in the levels of proliferation markers were observed. These results suggest that HIV-1 may enhance cervical carcinogenesis by promoting cell cycle progression. We also found that this HIV-1 Tat-induced cell proliferation was not dependent on the E2F family of transcription factors, and therefore postulate that Sp factors may be involved.

Human papillomavirus testing and conventional pap smear cytology as optional screening tools of women at different risks for cervical cancer in the countries of the former soviet union.

Objectives. Human papillomavirus (HPV) infection is a sexually transmitted disease (STD) and the single most important etiological agent of cervical cancer. In parallel with the increase of STDs and because of the lack of any organized cancer screening in the new independent states of the former Soviet Union, the incidence and mortality rates of cervical cancer are rapidly rising. This is the first report from an ongoing European Commission-funded (INCO-Copernicus Program) cross-sectional and cohort study (focused on the key issues of this major health problem in the new independent states) analyzing the performance of the HPV DNA (Hybrid Capture II) test as a potential screening tool for cervical cancer in these countries. Materials and Methods. A series of 3,175 women (screening, gynecological, or STD patients) from six clinics in Russia, Belarus, and Latvia received routine cytology and HPV testing with Hybrid Capture II (HCII). All women with HPV-positive results or abnormalities in cytology were subjected to colposcopy and biopsy. The sensitivity, specificity, receiver operating characteristics, as well as positive (PPV) and negative predicting values (NPV), were determined for HCII and quality-controlled cytology in detecting significant pathology (cervical intraepithelial neoplasia [CIN] 3 and cancer). Results. Significant pathology was strongly associated with high-grade cytology (odds ratio [OR] = 8.5; 95% confidence interval [CI] = 4.1–17.8; chi-square, p < .0001). Pap smear cytology detected high-grade lesions with a sensitivity of 64.0% (44.8–83.2), specificity of 89.1% (84.5–93.7), PPV of 44.4% (28.8–61.0), and NPV of 94.8% (91.2–98.4). Of the 3,086 samples analyzed by HCII, 33.0% were positive for oncogenic HPV types, with a wide variation (from 23% to 45%) between the three patient groups (p < .0001). The presence of high-grade cytology was significantly associated with HCII positivity at all cutoff levels (OR = 14.4; 95% CI = 8.4–24.5; chi-square, p < .0001; 1 pg/mL threshold). In the receiver operating characteristics curve, the HCII cutoff point most closely balancing sensitivity (83.1%) and specificity (75.6%) was 2 pg/mL. The presence of high-grade histology was associated with HCII positivity (cutoff 1 pg/mL; OR = 4.8; 95% CI = 0.7–34.2;p = .047). At the cutoff (1 pg/mL), sensitivity of the HCII test was 96.6% (90.0–100), specificity was 15.9% (10.6–21.2), PPV was 15.1% (9.9–20.3), and NPV was 96.8% (90.3–100). Changing the cutoff significantly affected sensitivity at 20 pg/mL and NPV at 500 pg/mL. Conclusions. HCII assay is a sensitive tool in detecting significant pathology, but less specific than the Pap test. A negative HCII test practically precludes high-grade CIN (NPV, >95%). Because the performance characteristics of the HCII test depend on the prevalence of HPV and CIN in the study population, the cost-benefit issues in different settings will be the limiting factor for the application of this test as a screening tool.

Acquisition of high-risk human papillomavirus infections and Pap smear abnormalities among women in the New Independent States of the former Soviet Union

ABSTRACT The rates of acquisition and the times of incident high-risk (HR) human papillomavirus (HPV) infections and Pap smear abnormalities and their predictive factors were analyzed in women participating in a multicenter screening study in three countries of the New Independent States of the former Soviet Union. The 423 patients were prospectively monitored for a mean of 21.6 months. At the baseline, 118 women were HR HPV DNA negative (Hybrid Capture II assay) and Pap smear negative (group 1), 184 were HPV DNA positive and Pap smear negative (group 2), and 121 were HPV DNA negative and Pap smear positive (group 3). The time to the acquisition of an incident abnormal Pap smear (19.4 months) was significantly longer in group 1 than in group 2 (9.2 months) (P = 0.0001). The times of acquisition of incident HR HPV infection were 16.6 and 11.0 months in group 1 and group 3, respectively (P = 0.006). The monthly rates of acquisition of incident HR HPV infections were very similar in group 1 (1.0%) and group 3 (0.8%), whereas the rate of acquisition of an abnormal Pap smear was significantly higher in group 2 (3.1%) than in group 1 (1.5%) (P = 0.0001). The acquisition of HR HPV infection (but not a positive Pap smear result) was significantly (P = 0.0001) age dependent. The only significant independent (P = 0.001) predictor of the incidence of an abnormal Pap smear result was a high HR HPV load of >20 relative light units/control value (CO) (rate ratio, 2.050; 95% confidence interval, 1.343 to 3.129). Independent predictors of incident HR HPV infection were patient category (a sexually transmitted disease) and ever having been pregnant. The time of acquisition of HR HPV infection was 3 months shorter than that of an abnormal Pap smear. At the baseline the high load of a particular HR HPV type is the single most important predictor of an incident Pap smear abnormality, whereas young age and having a sexually transmitted disease predict incident HR HPV infections.

Quantitative in situ evaluation of telomeres in fluorescence in situ hybridization‐processed sections of cutaneous melanocytic lesions and correlation with telomerase activity

Summary Background Telomere length is correlated with cellular ageing and immortalization processes. In some human cancers telomere length measurement has proved to be of diagnostic and prognostic value. Results comparable with the traditional terminal restriction fragment length determination by Southern blotting have been obtained in metaphase and interphase cells in some studies by fluorescence in situ hybridization (FISH) analysis; FISH additionally allows for the quantification of telomeres at the cellular level.

Sexual habits and human papillomavirus infection among females in three New Independent States of the former Soviet Union.

Background On a global scale, the New Independent States (NIS) of the former Soviet Union have an intermediate incidence of cervical cancer, the main etiologic factor of which is human papillomavirus (HPV), a major sexually transmitted disease (STD). Recently, the prevalence of all STDs has exploded in these countries. Goal The goal of this study was to examine the sexual habits and HPV prevalence among females in three NIS countries. Study Design In this multinational (European Community-funded) trial, a series of 3,175 consecutive female patients were examined for HPV status (by Hybrid Capture II) at six clinics in Russia, Belarus, and Latvia. A meticulous survey of their sexual habits and other potential risk factors of HPV infections was made by structured questionnaire. Results Three categories of patients were examined: those attending STD clinics (n = 722), gynecological patients (n = 761), and those who participated in cervical cancer screening (n = 1,692). These three categories were significantly differentiated by a large number of key variables, including the HPV detection rate (44.9% of STD patients, 39.8% of gynecological patients, and 24.5% of those who were screened). A wide variety sexual habits of these subjects were predictors of the HPV status in univariate analysis. Binary logistic regression analysis found that six different variables remained as independent predictors of HPV status. Patient category (STD) and (young) age were two highly significant predictors of increased risk (P < 0.0001), whereas having a nonsmoking partner and having zero or one partner during the past 2 years were significant protective factors (P = 0.004 and P = 0.007, respectively). Conclusion The results of this study indicate that women and girls in these NIS countries are conservative in many key characteristics of “high-risk” sexual behavior, such as age at onset of sexual activity, number of partners, and casual sex partners. HPV-positive and HPV-negative groups are clearly distinguished by the same variables identified as the key risk factors of HPV infection and cervical intraepithelial neoplasia in Western countries.