Jasenka Matisic

Exploratory Analysis of Quantitative Histopathology of Cervical Intraepithelial Neoplasia: Objectivity, Reproducibility, Malignancy-Associated Changes, and Human Papillomavirus

Background: As part of a project to evaluate emerging optical technologies for cervical neoplasia, our group is performing quantitative histopathological analyses of biopsy specimens from 1,190 patients. Objectives in the interim analysis are (a) quantitatively assessing progression of the neoplastic process of cervical intraepithelial neoplasia (CIN)/squamous intraepithelial lesions (SIL), (b) detecting malignancy‐associated changes (MACs), and (c) phenotypically measuring human papillomavirus (HPV) detected by DNA testing.

Quantitative Histopathological Analysis of Cervical Intra-Epithelial Neoplasia Sections: Methodological Issues

Objectives: As part a Program Project to evaluate emerging optical technologies for cervical neoplasia, our group is performing quantitative histopathological analysis of biopsies from 1800 patients. Several methodological issues have arisen with respect to this analysis: (1) Finding the most efficient way to compensate for staining intensity variation with out losing diagnostic information; (2) Assessing the inter‐ and intra‐observer variability of the semi‐interactive data collection; and (3) the use of non‐overlapping cells from the intermediate layer only. Methods: Non‐overlapping quantitatively stained nuclei were selected from 280 samples with histopathological characteristics of normal (199), koilocytosis (37), CIN 1 (18), CIN 2 (10) and CIN 3 (16). Linear discriminant analysis was used to assess the diagnostic information in three different feature sets to evaluate and compare staining intensity normalization methods. Selected feature values and summary scores were used to evaluate intra‐ and inter‐observer variability. Results: The features normalized by the internal subset of the imaged cells had the same discriminatory power as those normalized by the control cells and by both normalization methods seem to have additional discriminatory power over the set of features which do not require normalization. The use of the internal subset decreased the image acquisition time by ∼50% at each center, respectively. The intra‐ and inter‐observer variability was of a similar size. Good performance was obtained by measuring the intermediate layer only. Conclusion: The use of intensity normalization from a subset of the imaged non‐overlapping intermediate layer cells works as well as or better than any of the other methods tested and provides a significant timesaving. Our intra‐ and inter‐observer variability do not seem to affect the diagnostic power of the data. Although this must be tested in a larger data set, the use of intermediate layer cells only may be acceptable when using quantitative histopathology.

Accuracy of optical spectroscopy for the detection of cervical intraepithelial neoplasia: Testing a device as an adjunct to colposcopy

Testing emerging technologies involves the evaluation of biologic plausibility, technical efficacy, clinical effectiveness, patient satisfaction, and cost‐effectiveness. The objective of this study was to select an effective classification algorithm for optical spectroscopy as an adjunct to colposcopy and obtain preliminary estimates of its accuracy for the detection of CIN 2 or worse. We recruited 1,000 patients from screening and prevention clinics and 850 patients from colposcopy clinics at two comprehensive cancer centers and a community hospital. Optical spectroscopy was performed, and 4,864 biopsies were obtained from the sites measured, including abnormal and normal colposcopic areas. The gold standard was the histologic report of biopsies, read 2 to 3 times by histopathologists blinded to the cytologic, histopathologic, and spectroscopic results. We calculated sensitivities, specificities, receiver operating characteristic (ROC) curves, and areas under the ROC curves. We identified a cutpoint for an algorithm based on optical spectroscopy that yielded an estimated sensitivity of 1.00 [95% confidence interval (CI) = 0.92–1.00] and an estimated specificity of 0.71 [95% CI = 0.62–0.79] in a combined screening and diagnostic population. The positive and negative predictive values were 0.58 and 1.00, respectively. The area under the ROC curve was 0.85 (95% CI = 0.81–0.89). The per‐patient and per‐site performance were similar in the diagnostic and poorer in the screening settings. Like colposcopy, the device performs best in a diagnostic population. Alternative statistical approaches demonstrate that the analysis is robust and that spectroscopy works as well as or slightly better than colposcopy for the detection of CIN 2 to cancer.

The quality of community Colposcopic practice

OBJECTIVE: To estimate the quality of community colposcopic practice in British Columbia through an assessment of the degree of correlation between colposcopy, cytology, and histology. METHOD: We reviewed all new-patient colposcopies in British Columbia during 2001 by 37 gynecologists in 24 hospital-based clinics. RESULTS: Colposcopic impression closely mirrored the referral cytology diagnosis in 89.8% of cases. As with cytology-biopsy comparisons, discordant cases were more likely to be overestimates of disease rather than underestimates, 18.8% versus 1.8%. Overestimates were usually biopsy sampling errors rather than false positive cytology. The overall correlation between cytology and biopsy was considered satisfactory in 79.4% of cases. Satisfactory agreement between the colposcopic diagnosis and accompanying biopsies occurred in 86.8% of patients. Five colposcopists had performance scores below this standard. Colposcopy with a sensitivity of 90.3% and a specificity of 57.3% as practiced in this provincial program would appear to be of a satisfactory level. The rate of intraepithelial or invasive disease increased from 40.6% in patients with low-grade squamous intraepithelial changes to 91.9% in patients with suspicious or malignant cytology. The value of the colposcopic impression to identify disease correlated best with the higher the grade of disease predicted (64.6% to 92.6%). CONCLUSION: A measure of the colposcopic proficiency in the community can be estimated by comparing the level of agreement between the presenting cytology, colposcopic impression, and corresponding directed biopsies. The results of this study would indicate that 5 individuals had practice standards that were below average. An integrated cytology-colposcopy program facilitates the assessment and identification of below-average practice standards in a community. LEVEL OF EVIDENCE: III

DNA-cytometry of progressive and regressive cervical intraepithelial neoplasia.

A retrospective analysis was performed on archival cervical smears from a group of 56 women with cervical intraepithelial neoplasia (CIN), who had received follow‐up by cytology only. Automated image cytometry of Feulgen‐stained DNA was used to determine the differences between progressive and regressive lesions. The first group of 30 smears was from women who had developed cancer after initial smears with dysplastic changes (progressive group). The second group of 26 smears with dysplastic changes had shown regression to normal (regressive group). The goal of the study was to determine if differences in cytometric features existed between the progressive and regressive groups. CIN categories I, II and III were represented in both groups, and measurements were pooled across diagnostic categories. Images of up to 700 intermediate cells were obtained from each slide, and cells were scanned exhaustively for the detection of diagnostic cells. Discriminant function analysis was performed for both intermediate and diagnostic cells. The most significant differences between the groups were found for diagnostic cells, with a cell classification accuracy of 82%. Intermediate cells could be classified with 60% accuracy. Cytometric features which afforded the best discrimination were characteristic of the chromatin organization in diagnostic cells (nuclear texture). Slide classification was performed by thresholding the number of cells which exhibited progression associated changes (PAC) in chromatin configuration, with an accuracy of 93 and 73% for diagnostic and intermediate cells, respectively. These results indicate that regardless of the extent of nuclear atypia as reflected in the CIN category, features of chromatin organization can potentially be used to predict the malignant or progressive potential of CIN lesions.

Up regulation in gene expression of chromatin remodelling factors in cervical intraepithelial neoplasia.

BackgroundThe highest rates of cervical cancer are found in developing countries. Frontline monitoring has reduced these rates in developed countries and present day screening programs primarily identify precancerous lesions termed cervical intraepithelial neoplasias (CIN). CIN lesions described as mild dysplasia (CIN I) are likely to spontaneously regress while CIN III lesions (severe dysplasia) are likely to progress if untreated. Thoughtful consideration of gene expression changes paralleling the progressive pre invasive neoplastic development will yield insight into the key casual events involved in cervical cancer development.ResultsIn this study, we have identified gene expression changes across 16 cervical cases (CIN I, CIN II, CIN III and normal cervical epithelium) using the unbiased long serial analysis of gene expression (L-SAGE) method. The 16 L-SAGE libraries were sequenced to the level of 2,481,387 tags, creating the largest SAGE data collection for cervical tissue worldwide. We have identified 222 genes differentially expressed between normal cervical tissue and CIN III. Many of these genes influence biological functions characteristic of cancer, such as cell death, cell growth/proliferation and cellular movement. Evaluation of these genes through network interactions identified multiple candidates that influence regulation of cellular transcription through chromatin remodelling (SMARCC1, NCOR1, MRFAP1 and MORF4L2). Further, these expression events are focused at the critical junction in disease development of moderate dysplasia (CIN II) indicating a role for chromatin remodelling as part of cervical cancer development.ConclusionWe have created a valuable publically available resource for the study of gene expression in precancerous cervical lesions. Our results indicate deregulation of the chromatin remodelling complex components and its influencing factors occur in the development of CIN lesions. The increase in SWI/SNF stabilizing molecule SMARCC1 and other novel genes has not been previously illustrated as events in the early stages of dysplasia development and thus not only provides novel candidate markers for screening but a biological function for targeting treatment.

Cytologic features of papillary serous adenocarcinoma of the uterine cervix

The cytologic features of papillary serous carcinoma of the cervix (PSCC) have not been described in detail previously. In this study the cytologic features of primary, pure PSCC are described and correlated with the histology. Comparison is made with papillary serous ovarian carcinoma metastatic to the cervix.

Apocrine carcinoma of the lip: A case report including immunohistochemical and ultrastructural study, discussion of differential diagnosis, and review of the literature

The light microscopic, immunohistochemical, and ultrastructural features of a case of apocrine carcinoma of the upper lip of a 54-year-old white man are described. The neoplasm had a cribriform intraductal component resembling apocrine carcinoma of the breast. The tumor had irregular borders and infiltrated skeletal muscle. The neoplastic cells had abundant eosinophilic granular cytoplasm and showed apical decapitation secretion characteristic of apocrine differentiation. The differential diagnosis is discussed with particular reference to distinction of the tumor from oncocytic carcinoma and ductal carcinoma of minor salivary gland.

Comprehensive serial analysis of gene expression of the cervical transcriptome

BackgroundMore than half of the approximately 500,000 women diagnosed with cervical cancer worldwide each year will die from this disease. Investigation of genes expressed in precancer lesions compared to those expressed in normal cervical epithelium will yield insight into the early stages of disease. As such, establishing a baseline from which to compare to, is critical in elucidating the abnormal biology of disease. In this study we examine the normal cervical tissue transcriptome and investigate the similarities and differences in relation to CIN III by Long-SAGE (L-SAGE).ResultsWe have sequenced 691,390 tags from four L-SAGE libraries increasing the existing gene expression data on cervical tissue by 20 fold. One-hundred and eighteen unique tags were highly expressed in normal cervical tissue and 107 of them mapped to unique genes, most belong to the ribosomal, calcium-binding and keratinizing gene families. We assessed these genes for aberrant expression in CIN III and five genes showed altered expression. In addition, we have identified twelve unique HPV 16 SAGE tags in the CIN III libraries absent in the normal libraries.ConclusionEstablishing a baseline of gene expression in normal cervical tissue is key for identifying changes in cancer. We demonstrate the utility of this baseline data by identifying genes with aberrant expression in CIN III when compared to normal tissue.

Aberrant Expression of Long Noncoding RNAs in Cervical Intraepithelial Neoplasia

Objective Long noncoding RNAs (lncRNAs) are a unique class of messenger RNA–like transcripts of at least 200 nucleotides in length with no significant protein-coding capacity. Aberrant lncRNA expression is emerging as a major component of the cancer transcriptome. Here, we sought to determine if differential lncRNA expression is a feature of the human cervical intraepithelial neoplasia (CIN) transcriptome. Methods Sequence data were derived from 16 long serial analyses of gene expression (L-SAGE) libraries constructed from cervical specimens representing mild (CIN1), moderate (CIN2), and severe (CIN3) histopathologic grades of CIN. A novel lncRNA discovery pipeline was developed to query the expression of lncRNAs within the SAGE data sets. Results A total of 2,230,370 sequence tags were delineated from the 16 SAGE libraries, representing the expression of 367,482 unique tags at varying abundance. Using a novel stepwise filtering strategy, we analyzed the cervical SAGE libraries and identified the expression profiles of 1056 lncRNAs in the human cervix. We present the first lncRNA expression profile derived from nonneoplastic cervical tissue and establish that changes in lncRNA expression do occur in cervical intraepithelial lesions. Our analysis also shows statistically significant aberrant expression of lncRNAs in the 3 CIN grades, suggesting that these unique noncoding RNA transcripts may contribute to the development and progression of precursor lesions. Conclusions Through the analysis of L-SAGE libraries constructed from cervical specimens, we provide the first lncRNA expression profile of the cervix and demonstrate aberrant expression in early-stage neoplasia.