Reproducibility of the Johns Hopkins Hospital template for urologic cytology samples.

INTRODUCTION: Cytologic screening for urothelial carcinoma is fraught with low sensitivity, a high indeterminate rate, and until recently, poor standardization of terminology. The Johns Hopkins Hospital John K. Frost Cytopathology Laboratory has recently developed and published a template for reporting urine cytopathology; herein, we evaluate its interobserver reproducibility. MATERIALS AND METHODS: Two sets of 100 cases each were deidentified; each set was reviewed by 5 of 10 observers in a randomized order at the direction of computerized data collection software that tracked observation time as well as observer classification of the atypia-no atypia, atypia (AUC-US), or atypia suggestive of high-grade urothelial carcinoma (AUC-H). Specific morphologic features were also recorded. Cases were grouped into low-, intermediate-, and high-agreement based on the number of observers who made the assessment. The findings were correlated against clinical outcomes. RESULTS: High agreement among observers about the presence or absence of high-grade features was possible in approximately two-thirds of indeterminate urine cases. Time and order did not factor significantly into observer propensity for identifying atypical features or favoring either AUC-US or AUC-H, and cases with high agreement about the presence of high-grade features were more likely to have a malignant follow-up. Furthermore, AUC-H diagnoses based on 2 or more high-grade features had a significantly higher malignancy risk than AUC-US diagnoses did. CONCLUSIONS: AUC-H is a valid diagnostic category with specific, reproducibly identified features that portend a higher risk of malignancy than the findings of AUC-US.

Utility of P16 expression and Ki-67 proliferation index in ASCUS and ASC-H pap tests.

Current cervical screening uses a combination of cytology and high-risk human papillomavirus (HR-HPV) analysis in cases of atypical squamous cells of undetermined significance (ASCUS) and atypical squamous cells cannot exclude high-grade intraepithelial lesion (ASC-H). These diagnoses are subject to interobserver variability and HR-HPV analysis can be limited by sampling inadequacy. This study correlates immunoexpression of P16 and Ki-67 in residual cervicovaginal material against cytology category and HR-HPV status. Eighteen pap tests were selected: 8 ASCUS, 4 ASC-H, and 6 controls (2 LSIL and 4 HSIL). Digene Hybrid Capture II test was used to detect HR-HPV. The cytospins were stained for P16/Ki-67. Pap tests, P16, Ki-67, HR-HPV result and available biopsies were correlated. P16 expression correlated with HR-HPV status in 15/17 cases. Discordant cases (1 ASCUS and 1 ASC-H) were +P16/-HR-HPV. Ki-67 correlated with HR-HPV in 8/15 cases. Discordant cases were +HR-HPV/- Ki-67 (HSIL, LSIL, and ASC-H one each), and -HR-HPV/+Ki-67 (3 ASCUS, 1 LSIL, 1 ASC-H). Two cases were + P16/+ Ki-67/- HR-HPV. None were - P16/- Ki-67/+ HR-HPV. Histologic follow-up in 13 cases varied from benign to CIN III. Two cases of +P16/ - Ki-67/- HR-HPV had benign cervical biopcies. Although a small sample size, our findings show a utility for adjunct P16/ Ki-67 in addition to HR-HPV testing in cases of squamous atypia when HR-HPVs are non-detected due to low DNA copies, or missed lesions in cervical biopsies.

Human papillomavirus prevalence and cytopathology correlation in young Ugandan women using a low-cost liquid-based Pap preparation.

Screening for HPV-driven cervical dysplasia and neoplasia is a significant public health concern in the developing world. The purpose of this study was to use a manual, low-cost liquid-based Pap preparation to determine HPV prevalence in HIV-positive and HIV-negative young women in Kampala, Uganda and to correlate cervical cytopathology with HPV-DNA genotype. About 196 post-partum women aged 18-30 years underwent rapid HIV testing and pelvic examination. Liquid-based cervical cytology samples were processed using a low-cost manual technique. A DNA collection device was used to collect specimens for HPV genotyping. HIV and HPV prevalence was 18 and 64%, respectively. Overall, 49% of women were infected with a high-risk HPV genotype. The most common high-risk HPV genotypes were 16 (8.2%), 33 (7.7%), 35 (6.6%), 45 (5.1%), and 58 (5.1%). The prevalence of HPV 18 was 3.6%. HIV-positive women had an HPV prevalence of 86% compared to 59% in HIV-negative women (P = 0.003). The prevalence of HPV 16/18 did not differ by HIV status. HIV-positive women were infected with a significantly greater number of HPV genotypes compared to HIV-negative women. By multivariate analysis, the main risk factor for HPV infection was coinfection with HIV. HIV-positive women were four times more likely to have abnormal cytology than HIV-negative women (43% vs. 11.6%, P < 0.001). These data highlight that HIV infection is a strong risk factor for HPV infection and resultant abnormal cervical cytology. Notably, the manual low-cost liquid-based Pap preparation is practical in this setting and offers an alternate method for local studies of HPV vaccine efficacy.

Detection of cervical high-grade squamous intraepithelial lesions from cytologic samples using a novel immunocytochemical assay (ProEx C).

BACKGROUND: Routine liquid-based cytology (LBC) provides excellent sensitivity for the detection of cervical high-grade squamous intraepithelial lesion (HSIL); however, its specificity is low. Consequently, many women who have atypical squamous cells of undetermined significance (ASC-US) or low-grade squamous intraepithelial lesion (LSIL) cytology undergo unnecessary colposcopy. The authors hypothesized that a novel immunocytochemical assay (ProEx C) that can be performed on LBC slides had a significantly higher positive predictive value (PPV) for biopsy-proven HSIL compared with routine LBC. METHODS: The ProEx C immunocytochemical assay utilizes a cocktail of monoclonal antibodies directed against proteins associated with aberrant S-phase cell cycle induction (topoisomerase IIA, minichromosome maintenance protein 2). The ProEx C reagents were validated in the authors' laboratory for staining and scoring reproducibility, open-vial stability, and accuracy before a retrospective analysis using these reagents was performed on 317 residual cytology samples. Sensitivity, specificity, PPV, and negative predictive value (NPV) for the detection of biopsy-proven HSIL were determined. RESULTS: The ProEx C assay was validated successfully in the authors' cytology laboratory. Using biopsy-proven HSIL as an endpoint, the ProEx C assay yielded a sensitivity of 85.3%, specificity of 71.7%, PPV of 44.6%, and NPV of 94.8%. Compared with the routine LBC results in the same cohort, the ProEx C sensitivity for biopsy-proven HSIL was 70.6% greater than HSIL+ cytology (50% vs. 85.3%). ProEx C also showed a 114% increase in PPV relative to ASC-US cytology (21.1% vs. 44.6%). CONCLUSIONS: The ProEx C immunocytochemical assay can be integrated into a clinical cytology laboratory and may increase the PPV of LBC for biopsy-proven HSIL.

Validation of a low-cost, liquid-based screening method for cervical intraepithelial neoplasia.

OBJECTIVE: The objective of the study was to validate a low-cost, liquid-based method for cervical cancer screening. STUDY DESIGN: We conducted a retrospective, split-sample comparison of 300 liquid-based cervical cytology samples from a group of 150 human immunodeficiency virus-seropositive women and 150 women from low-risk general gynecology clinics whose specimens were screened via standard liquid-based methodology as part of routine care. Residual samples from each specimen were used to prepare a slide using a novel, inexpensive manual membrane method of liquid-based cytology. These slides were screened by a cytotechnologist and abnormal cases were reviewed by a pathologist. Final diagnoses from the manual membrane method of liquid-based cytology slides were compared with the original diagnoses and available cervical biopsy data. RESULTS: There was good overall agreement between the manual membrane method of liquid-based cytology and original cytology diagnoses (76.3% agreement; kappa = 0.52, 95% confidence interval 0.44 to 0.59). Using available biopsy data to determine the accuracy of each method to identify high-grade squamous intraepithelial lesions, the manual membrane method of liquid-based cytology method was found to have a higher sensitivity (71.4% versus 57.1%) and lower specificity (82.1% versus 89.7%). The slightly higher referral rate to colposcopy using the manual membrane method of liquid-based cytology method was limited to women from the low-risk general gynecology clinics (16.7% versus 12.0%, P = .05). CONCLUSION: The low-cost manual membrane method of liquid-based cytology cervical cytology method is comparable with a standard commercial method. Consequently, it may be of value in alternative screening strategies in resource-limited settings.