Analysis of the performance of 100% rapid review using an average time of 1 and 2 minutes according to the quality of cervical cytology specimens.

OBJECTIVES: To compare the performance of the 100% rapid review method carried out in a mean time of either 1 or 2 minutes according to cytological final result, and to assess whether the presence of obscuring factors in cervical smear samples affects the frequency of false-negative results. METHODS: A total of 5,235 smears classified as negative (93.0%) or unsatisfactory (2.1%) at routine screening were submitted to 100% rapid review using mean times of 1 and 2 minutes. RESULTS: Of the 5,235 smears submitted to 1-minute rapid review, 88 were considered suspect and of these, 45 were confirmed as abnormal in the cytological final result. When the time used was 2 minutes, 67 smears were considered suspect, 44 of which were confirmed as abnormal. Sensitivity and specificity were similar in the 1- and 2-minute reviews. In smears in which samples were satisfactory and had no obscuring factors, sensitivity and specificity were 64.2% and 98.9% and 61.5% and 99.5% for the 1- and 2-minute reviews, respectively. In comparison, in smears in which the sample was satisfactory for analysis but partially obscured (50-75%), sensitivity and specificity were 64.7% and 99.9% and 70.6% and 99.8% for the 1- and 2-minute reviews, respectively. CONCLUSIONS: The method of rapid review of 100% showed no difference in the detection of false-negative results using the time of 1 or 2 minutes. The quality of the sample did not influence the detection of false-negatives.

Excessive Pap smears due to opportunistic cervical cancer screening.

The study aimed to analyze the Pap smears carried out for cervical cancer screening according to Ministry of Health guidelines. All smear tests carried out within the public health system in Campinas in 2003 were analyzed. All tests that did not conform to the guidelines were considered excessive. The guidelines recommend screening once every three years for all women aged 25 to 59 after they have received two negative smears. This study showed that the majority of women initiated screening prior to 25 years of age and the periodicity was predominantly annual, followed by biannual tests. In conclusion, 63.4% of tests were excessive. The screening coverage was 14.76%, but if all the tests had been performed as recommended, the final coverage over three years could have reached 65.4%. Thus it is possible to increase the coverage with the available resources since the screening works like an organized program.

Stromal cells play a role in cervical cancer progression mediated by MMP-2 protein.

Metalloproteinases, especially metalloproteinase-2 (MMP-2), are known for their role in the degradation of the extracellular matrix. Nevertheless, a thorough understanding of MMP-2 expression in neoplastic lesions of the uterine cervix has yet to be accomplished. This study aimed to analyze the MMP-2 expression in cervical intraepithelial neoplasia III (CIN3) and in cervical squamous cell carcinoma, in tumor cells and adjacent stromal cells. MMP-2 expression was assessed by an immunohistochemical technique. MMP-2 expression was greater in the stromal cells of invasive carcinomas than in CIN3 (p < 0.0001). MMP-2 expression in stromal cells correlates with the clinical stage, gradually increasing as the tumor progresses (p = 0.04). This study corroborates that stromal cells play an important role in tumor invasion and progression, mediated by the progressive enhancement of MMP-2 expression from CIN3 to advanced invasive tumor. The intense MMP-2 expression most probably is associated with poor tumor prognosis.

The investigation of DNA repair polymorphisms with histopathological characteristics and hormone receptors in a group of Brazilian women with breast cancer.

The association of tumor differentiation and estrogen receptor expression with the prognosis of breast cancer has been well established. Nevertheless, little is yet reported about the association of morphological characteristics of the tumor, estrogen receptor status and polymorphisms in low penetrance genes. The aim of the present study was to investigate a possible association between DNA repair gene polymorphisms (XRCC1, XPD, XRCC3, and RAD51) with histological type, grade and hormone receptor expression in a series of breast cancers. A cross-sectional study was carried out to evaluate 94 women with breast carcinoma, who had already been selected and included in a study on the association of DNA repair gene polymorphisms. For immunohistochemistry, formalin-fixed, paraffin-embedded tissue samples from breast tumors were consecutively retrieved from the histopathology files of our institution. DNA obtained from blood samples of the same patients was investigated for the presence of the following polymorphisms: Arg-399Gln located in the XRCC1 gene; 135C/G located in the RAD51 gene; Lys751Gln located in the XPD gene and Thr241Met located in the XRCC3 gene. Polymorphisms were considered to be independent variables and hormone receptor expression and the morphological characteristics of the tumors comprised the dependent variables. No statistically significant association was found between gene polymorphisms and hormone receptor status. The association between XRCC1-Arg399Gln polymorphism and ductal carcinoma was statistically significant (P = 0.02). The association of the XPD-Lys751Gln polymorphism with histological grade was also tatistically significant (p = 0.05). In conclusion, the XRCC1 genotype was found to be associated with ductal carcinoma histotypes and XPD genotype with low histological grade, which is the most frequent pattern of sporadic breast carcinomas.

Rapid pre-screening of cervical smears as a method of internal quality control in a cervical screening programme.

OBJECTIVE: To evaluate the performance of rapid pre-screening (RPS) as a method of internal quality control in the cytopathological examination of cervical smears for cervical cancer screening. METHODS: The sample consisted of 6135 cervical smears submitted to RPS and routine screening (RS) methods. The smears classified as negative in RPS and RS were considered final diagnoses, and were not, therefore, submitted to any additional review. The smears identified as suspect or unsatisfactory according to RPS were analysed separately by two different cytologists irrespective of the diagnosis reached in RS. Smears considered abnormal or unsatisfactory at RS were also reviewed. When both cytologists issued concordant diagnoses, this was considered the final diagnosis. Discordant results were analysed by a third cytologist and a consensus meeting was held to define the final diagnosis. RESULTS: Taking abnormalities detected by RS as the denominator, RPS had a sensitivity of 63.0% for the detection of all abnormal smears and 96.7% for high grade squamous intraepithelial lesion (HSIL). When compared with the final diagnosis, sensitivity of RPS for all abnormal smears was 74.9% and for HSIL 95.0%. Of the 529 abnormal smears confirmed in the final diagnosis, 2.15% were detected only by the RPS. CONCLUSION: RPS is an effective alternative method of internal quality control with high sensitivity for the detection of more severe lesions. It also permits monitoring of the laboratory rate of false-negative results, and allows constant evaluation of the performance both of the pre-screening and RS cytologists.

Endocervical glandular cell abnormalities in conventional cervical smears: evaluation of the performance of cytomorphological criteria and HPV testing in predicting neoplasia.

OBJECTIVE: To analyse the correlation between cytomorphological criteria in smears with atypical glandular cells (AGC) or adenocarcinoma in situ (AIS) and human papillomavirus (HPV) reflex test results with different neoplastic histological diagnoses, particularly to distinguish between glandular and squamous neoplasia. METHODS: A series of 155 women with glandular abnormalities in their conventional cervical smears was included: 106 with AGC, 35 with AGC associated with high-grade squamous intraepithelial lesion (HSIL) and 14 with AIS. Two reviewers evaluated 35 cytomorphological criteria and hybrid capture II (HCII) was performed in all cases. Colposcopy was carried out in all cases and biopsy in 126/155. For statistical purposes, predictive values and odds ratio (OR) were calculated, followed by chi-square automatic interaction detection. RESULTS: Histology detected 56 cases of squamous and 17 of glandular intraepithelial or invasive neoplasia. Predictive values of the papillary groups and feathering criteria for glandular neoplasia were, respectively, 80.0% and 73.3%. Feathering was the criterion with the highest OR for distinguishing glandular from squamous neoplasia and also for distinguishing between glandular and non-neoplastic diagnosis. Rosettes and pseudostratified strips did not perform as well. Multivariant Classification and Regression Trees analysis identified feathering as the best criterion for distinguishing between glandular, squamous and non-neoplastic diagnoses regardless of HPV status. CONCLUSIONS: Feathering was the best criterion for predicting glandular neoplasia.

p16(INK4a) Expression does not predict the outcome of cervical intraepithelial neoplasia grade 2.

Spontaneous regression of cervical intraepithelial neoplasia grade 2 (CIN2) lesions has been recognized since 1955, but predictors of this are poorly understood. Among the predictive markers studied, p16(INK4a) has been suggested to be of some value in monitoring the diagnosis of CIN2. In this clinical trial, 90 Brazilian women, diagnosed to CIN2 and high-risk human papillomavirus infection, were randomized into two groups of equal size: 45 women whose lesions were excised and 45 women subjected to prospective follow-up at 3-month intervals at least for 1 year (mean 6.8 months). p16(INK4a) expression was analyzed in paraffin-embedded sections using immunohistochemical staining. Among the 45 women in the follow-up group, 42% experienced spontaneous regression, 11% showed persistence, 22% progressed to CIN3, and 20% had partial regression to CIN1 or ASCUS (atypical squamous cell undetermined signifiance). p16(INK4a) expression was detected in 68.9% of the patients. In univariate survival (Cox) analysis, no significant difference in regression was obtained between p16(INK4a)-negative and -positive CIN2 lesions (adjusted HR = 1.1; 95% CI 0.6-2.0). In conclusion, p16(INK4a) expression could be useful in the diagnosis of CIN2. However, it failed to predict the outcome of CIN2. Because of its high spontaneous regression rate, follow-up could be considered as a management option of CIN2 in young and compliant women.

Prediction of high-grade cervical disease with human papillomavirus detection in women with glandular and squamous cytologic abnormalities.

The objective of this study was to assess whether human papillomavirus (HPV) detection with hybrid capture II (HC II) can help predict the presence and the nature, glandular or squamous, of histologic cervical lesions in women referred due to atypical glandular cells (AGC) or high-grade squamous intraepithelial lesion (HSIL). A total of 247 women were included. Referral Pap smears comprised AGC (51 cases), AGC plus HSIL (28 cases), adenocarcinoma in situ (10 cases), and HSIL (158 cases). All patients were tested for high-risk HPV with HC II and had a histologic assessment of their cervix. Histologic analysis showed 38 women with (15.3%) cervicitis, 194 with (75.5%) squamous lesions, and 15 with (9.2%) glandular neoplasia. The overall rate of high-risk HPV detection was 77%. Almost 70% of AGC-HPV-negative patients did not have a pathologically proven cervical neoplasia, whereas 76% of women with AGC-HPV-positive result were diagnosed with a squamous or glandular neoplasia. Most (95%) of the lesions in patients with AGC-HSIL were of squamous nature, and HPV detection did not contribute to their differentiation from glandular lesions. We conclude that in women with AGC, HPV positivity strongly correlated with the presence of glandular or squamous cervical lesion but did not help distinguishing women with squamous from those with glandular neoplasia.

Evaluation of 100% rapid rescreening of negative cervical smears as a quality assurance measure.

OBJECTIVE: The objective of this study was to compare the performance of 100% rapid rescreening, 10% random rescreening and the review of smears selected on the basis of clinical criteria, as a method of internal quality control of cervical smears classified as negative during routine screening. METHODS: A total of 3149 smears were analysed, 173 of which were classified as positive and 2887 as negative, while 89 smears were considered unsatisfactory. The smears classified as negative were submitted to 100% rapid rescreening, 10% random rescreening, and rescreening based on clinical criteria. The rescreening stages were blinded and results were classified according to the Bethesda 2001 terminology. Six cytologists participated in this study, two of whom were responsible for routine screening while the other four alternated in carrying out rescreening so that no individual reviewed the same slide more than once. RESULTS: The 100% rapid rescreening method identified 92 suspect smears, of which 42 were considered positive at final diagnosis. Of the 289 smears submitted to the 10% rescreening method, four were considered abnormal but only one was confirmed positive in the final diagnosis. Of the 690 smears rescreened on the basis of clinical criteria, 10 were considered abnormal and eight received a positive final diagnosis. CONCLUSIONS: The 100% rapid rescreening method is more efficient at detecting false-negative results than 10% random rescreening or rescreening on the basis of clinical criteria, and is recommended as an internal quality control method.

Atypical glandular cells: criteria to discriminate benign from neoplastic lesions and squamous from glandular neoplasia.

OBJECTIVE: To evaluate the presence of some criteria in cervical smears with atypical glandular cells and their correlation with histological patterns to identify pre-neoplastic and neoplastic lesions. METHODS: Seventy-three women referred with an atypical glandular cell smear, who had undergone conization or hysterectomy, were included in this study. Referral Pap smears were reviewed using the set of 27 cyto-morphological criteria that was correlated with the histological diagnosis. RESULTS: Histological results showed intraepithelial or invasive neoplasia in 35 (48%) cases and benign lesions in 38 (52%) cases. After logistic regression and decision tree analysis an increased nuclear/cytoplasmic ratio and the presence of dyskeratotic cells were strongly associated with intraepithelial or invasive neoplasia and the differential cyto-morphological criteria for glandular lesions were decreased cytoplasm, irregular nuclear membranes and the presence of nucleoli. CONCLUSION: The analysis of individual cyto-morphological criteria can better predict intraepithelial or invasive neoplasia and differentiate glandular from squamous lesions.

Nuclear alterations of cells and atypical metaplastic cells in cervical smears are predictive criteria of high-grade cervical intraepithelial neoplasia.

Diagnoses based on the screening of cervical smears show low interobserver reproducibility and are frequently discordant with the final histological diagnosis. The aim of this study was to identify which of the cytomorphologic criteria used in the screening of cervical smears were most predictive of the histopathological grades of cervical intraepithelial neoplasia. The abnormal cervical smears of 206 women were reviewed blindly according to 22 pre-established cytomorphological criteria. Colposcopic evaluation was carried out in all cases. The marked presence of several nuclear criteria frequently found together in the same smear was associated with high grade intraepithelial neoplasia regardless of the presence of any other criteria. On the other hand, when the nuclear criteria were less evident, the cluster of criteria related to metaplastic cells was predictive of a diagnosis of high-grade intraepithelial neoplasia. Focusing on selected cytological criteria can aggregate predictive value to cervical smear diagnoses.

Aided visual inspection with acetic acid (VIA) and HPV detection as optional screening tools for cervical cancer and its precursor lesions.

PURPOSE OF INVESTIGATION: To assess the contribution of visual inspection with acetic acid (VIA) and Hybrid Capture II (HCII) as adjunct methods to the Pap test in detecting cervical neoplasia. SUBJECTS AND METHODS: This was a cross-sectional study with 809 women who consecutively attended gynecological consultations at Campinas University, Brazil, from January 2002 to July 2003. Pap test, HCII, VIA, and colposcopy were offered to all patients. Performance of tests (alone or in combination) in detecting histologically confirmed lesions was evaluated. RESULTS: Of the 40 patients with CIN, 69% had CIN1, 26% CIN2 or CIN3 and one patient had invasive carcinoma. VIA had the best performance in detecting CIN, yielding 72% sensitivity and 91% specificity. Considering only CIN2 or worse as significant lesions, HCII had the best sensitivity (73%), while the Pap test was the most specific (93%). Combining the three exams, 92% of the CIN1 or worse were detected. When CIN1 was excluded from the analysis, Pap smear plus HCII delivered 82% sensitivity and 79% specificity. However, this combination yielded a very low (5%) PPV. CONCLUSION: VIA and HCII contributed to the screening of cervical neoplasia in a group of Brazilian women, but the cost-effectiveness of conjoint screening modalities is still debatable.

DNA repair gene polymorphisms and susceptibility to familial breast cancer in a group of patients from Campinas, Brazil

Several studies have reported that the genes involved in DNA repair and in the maintenance of genome integrity play a crucial role in protecting against mutations that lead to cancer. Epidemiologic evidence has shown that the inheritance of genetic variants at one or more loci results in a reduced DNA repair capacity and in an increased risk of cancer. Polymorphisms have been identified in several DNA repair genes, such as XRCC1, XPD, XRCC3, and RAD51, but the influence of specific genetic variants on repair phenotype and cancer risk has not yet been clarified. This was a case-control study design with three case groups: 53 women with breast cancer and family history; 33 women with sporadic breast cancer; 175 women with no breast cancer but with family history. The control group included 120 women with no breast cancer and no family history. The PCR-RFLP method was used to analyze the XRCC1-Arg399Gln, XPD-Lys751Gln, XRCC3-Thr241Met, and RAD51-G135C polymorphisms. No statistically significant differences were found between the case groups and the control group for any of the polymorphisms analyzed, and also between the breast cancer and family history group and the sporadic breast cancer group. Sample sizes of women with breast cancer, whether familial or sporadic, were insufficient to show any small true differences between the groups, but we have to consider that currently there is no clear consensus with respect to the association of these polymorphisms with breast cancer risk. Considering the data available, it can be conjectured that if there is any risk association between these single-nucleotide polymorphisms and breast cancer, this risk will probably be minimal. The greater the risk associated with cancer, the smaller the sample size required to demonstrate this association, and the data of different studies are usually, therefore, more concordant.

Comparison of immunoexpression of 2 antibodies for estrogen receptors (1D5 and 6F11) in breast carcinomas using different antigen retrieval and detection methods.

The importance of in situ immunodetection of hormone receptors for therapy planning and prognostic evaluation in patients with breast carcinoma is well established. Sensitive detection methods are of utmost importance, especially in poorly fixed tissues, which are not uncommon in routine pathologic practice. The purpose of the present study is to compare immunoexpression of estrogen receptors in 20 cases of invasive ductal carcinoma using two antibodies, 1D5 and 6F11, and to verify the effect of different antigen retrieval solutions and detection systems. Immunoperoxidase was performed on paraffin sections using 1D5 and 6F11 as primary antibodies. Heat-induced antigen retrieval was performed using citrate buffer (pH 6.0) or Tris-EDTA buffer (pH 8.9). Detection was achieved using the following systems: EnVision, EnVision Plus, and labeled streptavidin-biotin peroxidase complex. Reaction was semiquantified from 0 to 4. There were no differences between the two markers, 1D5 and 6F11, except when 6F11 was used with EnVision and citrate buffer, in which case weaker reactivity was observed. Only in this combination (6F11/EnVision) was EDTA buffer significantly better than citrate. Labeled streptavidin-biotin peroxidase complex presented the best results, followed by EnVision Plus.

Prognostic value of P53 codon 72 polymorphism in invasive cervical cancer in Brazil.

OBJECTIVE: The prognostic value of p53 codon72 polymorphism was analyzed in Brazilian women with cervical cancer. METHODS: The present study consists of 148 women diagnosed and treated for invasive cervical carcinoma (FIGO stages Ib-IIIb) between 1992 and 2002. Demonstration of p53 polymorphism was performed in DNA extracted from paraffin-embedded sections using the polymerase chain reaction (PCR). RESULTS: Among the 148 women, arg/arg was found in 99 (67%) and, arg/pro in 49 (33%). The overall survival (OS) curves (univariate) were different between arg/arg and arg/pro patients (P = 0.01). There was slightly increased risk of death for arg/arg patients (crude HR 2.2 CI 95% 1.2-4.0), which was not confounded by FIGO stages (adjusted HR 2.4 CI 95% 1.3-4.3). For disease-free survival (DFS), two situations were considered: (1) 124 women who received any treatment, and (2) 118 who received FIGO-recommended treatment. In the first group, 59% of arg/arg patients presented recurrence as compared to 32% in the arg/pro group (P = 0.02), whereas in the second group, 61% of the arg/arg and 34% arg/pro showed recurrence (P = 0.04). The risk of recurrence adjusted by FIGO stage for the 124 patients was 2.4 (CI 95% 1.0-3.7) and for the 118 it was 1.9 (CI 95% 1.0-3.4). These adjusted models showed no confounding and no interaction. CONCLUSIONS: Despite the prognostic significance of p53 polymorphism in univariate survival analysis, there was no or only marginal evidence on the independent prognostic value of p53arg/arg in multivariate analysis. The more ominous prognosis of the homozygous (arg/arg) patients was explained by the primary treatment, independent on the FIGO stage.

P53 expression as a predictor of recurrence in cervical squamous cell carcinoma.

P53 protein function is frequently down-regulated in cervical cancer by complexing with human papillomavirus (HPV) E6 protein, leading to degradation of p53, genomic instability, and mutations. Results are controversial, however, on the prognostic value of p53 protein expression in cervical cancer. In this study, a cohort of 220 Brazilian women with FIGO stage IB-III cervical squamous cell carcinoma (SCC), followed for 5 years, was analyzed for p53 protein expression using immunohistochemistry. The disease-free survival (DFS) and relapse rate were analyzed using univariate (Kaplan-Meier) and multivariable (Cox's proportional hazards model) survival analyses. P53 protein expression was detected in 35% of the patients, including 21% in stage I, 28% in stage II and 51% in stage III of disease. Of 220 women, only 116 completed one of the treatment options standardized by FIGO within 120 days. There was a higher risk of relapse in stage II and III disease, that was not modified by p53 positivity; HR 3.0 (1.3-6.5) to stage II and HR 4.0 (1.9-8.5) to stage III. The multivariate analysis evidenced that p53 expression is not an independent factor exceeding the power of FIGO stage as the single most important determinant of the hazards for disease relapse.

c-Myc protein expression is not an independent prognostic predictor in cervical squamous cell carcinoma.

The c-myc protein is known to regulate the cell cycle, and its down-regulation can lead to cell death by apoptosis. The role of c-myc protein as an independent prognostic determinant in cervical cancer is controversial. In the present study, a cohort of 220 Brazilian women (mean age 53.4 years) with FIGO stage I, II and III (21, 28 and 51%, respectively) cervical squamous cell carcinomas was analyzed for c-myc protein expression using immunohistochemistry. The disease-free survival and relapse-rate were analyzed using univariate (Kaplan-Meier) survival analysis for 116 women who completed the standard FIGO treatment and were followed up for 5 years. Positive c-myc staining was detected in 40% of carcinomas, 29% being grade 1, 9% grade 2, and 2% grade 3. The distribution of positive c-myc according to FIGO stage was 19% (17 women) in stage I, 33% (29) in stage II, and 48% (43) in stage III of disease. During the 60-month follow-up, disease-free survival in univariate (Kaplan-Meier) survival analysis (116 women) was lower for women with c-myc-positive tumors, i.e., 60.5, 47.5 and 36.6% at 12, 36, and 60 months, respectively (not significant). The present data suggest that immunohistochemical demonstration of c-myc does not possess any prognostic value independent of FIGO stage, and as such is unlikely to be a useful prognostic marker in cervical squamous cell carcinoma.

C-Myc protein expression is not an independent prognostic predictor in cervical squamous cell carcinoma

The c-myc protein is known to regulate the cell cycle, and its down-regulation can lead to cell death by apoptosis. The role of c-myc protein as an independent prognostic determinant in cervical cancer is controversial. In the present study, a cohort of 220 Brazilian women (mean age 53.4 years) with FIGO stage I, II and III (21, 28 and 51 percent, respectively) cervical squamous cell carcinomas was analyzed for c-myc protein expression using immunohistochemistry. The disease-free survival and relapse-rate were analyzed using univariate (Kaplan-Meier) survival analysis for 116 women who completed the standard FIGO treatment and were followed up for 5 years. Positive c-myc staining was detected in 40 percent of carcinomas, 29 percent being grade 1, 9 percent grade 2, and 2 percent grade 3. The distribution of positive c-myc according to FIGO stage was 19 percent (17 women) in stage I, 33 percent (29) in stage II, and 48 percent (43) in stage III of disease. During the 60-month follow-up, disease-free survival in univariate (Kaplan-Meier) survival analysis (116 women) was lower for women with c-myc-positive tumors, i.e., 60.5, 47.5 and 36.6 percent at 12, 36, and 60 months, respectively (not significant). The present data suggest that immunohistochemical demonstration of c-myc does not possess any prognostic value independent of FIGO stage, and as such is unlikely to be a useful prognostic marker in cervical squamous cell carcinoma