Flat Urothelial Lesions Research Articles

HER2 Expression in Bladder Cancer: A Focused View on Its Diagnostic, Prognostic, and Predictive Role.

Bladder cancer (BC) is a heterogeneous disease from a molecular, morphological, and clinical standpoint. HER2 is a known oncogene involved in bladder carcinogenesis. Assessing HER2 overexpression as a result of its molecular changes in a routine pathology practice using immunohistochemistry might be a useful adjunct in several scenarios, namely (1) to correctly identify flat urothelial lesions and inverted urothelial lesions in the diagnostic setting; (2) to provide prognostic hints in both non-muscle invasive (NMI) and muscle invasive (MI) tumors, thus supplementing risk stratification tools, especially when evaluating higher-risk tumors such as those with variant morphology; (3) to improve antibody panels as a surrogate marker of BC molecular subtyping. Furthermore, the potential of HER2 as a therapeutic target has been only partly explored so far, in light of the ongoing development of novel target therapies.

Targeted Next-Generation Sequencing of Flat Urothelial Lesions Reveals Putative Pathobiological Pathways, Potential Biomarkers, and Rational Therapeutic Targets

Flat urothelial lesions are controversial diagnostic and prognostic urologic entities whose importance relies mainly on their ability to progress to muscle-invasive tumors via urothelial carcinoma in situ (CIS). However, the carcinogenetic progression of preneoplastic flat urothelial lesions is not well established. Moreover, predictive biomarkers and therapeutic targets of the highly recurrent and aggressive urothelial CIS lesion are lacking. Using a targeted next-generation sequencing (NGS) panel of 17 genes directly involved in bladder cancer pathogenesis, we investigated alterations of genes and pathways with clinical and carcinogenic implications on 119 samples of flat urothelium, including normal urothelium (n = 7), reactive atypia (n = 10), atypia of unknown significance ( n = 34), dysplasia ( n = 23), and CIS (n = 45). The majority of the flat lesions were tumor-associated but grossly/microscopically or temporally separated from the main tumor. Mutations were compared across flat lesions and concerning the concomitant urothelial tumor. Associations between genomic mutations and recurrence after intravesical bacillus Calmette–Guerin treatment were estimated with Cox regression analysis. TERT promoter mutations were highly prevalent in intraurothelial lesions but not in the normal or reactive urothelium, suggesting that it is a critical driver mutation in urothelial tumorigenesis. We found that synchronous atypia of unknown significance-dysplasia-CIS lesions without concomitant papillary urothelial carcinomas had a similar genomic profile that differed from atypia of unknown significance-dysplasia lesions associated with papillary urothelial carcinomas, which harbored significantly more FGFR3, ARID1A, and PIK3CA mutations. KRAS G12C and ERBB2 S310F/Y mutations were exclusively detected in CIS and were associated with recurrence after bacillus Calmette–Guerin treatment (P = .0006 and P = .01, respectively). This targeted NGS study revealed critical mutations involved in the carcinogenetic progression of flat lesions with putative pathobiological pathways. Importantly, KRAS G12C and ERBB2 S310F/Y mutations were identified as potential prognostic and therapeutic biomarkers for urothelial carcinoma.

Use of Artificial Intelligence for the Interpretable Prediction of the Pathologic Diagnosis and Molecular Abnormalities of Flat Urothelial Lesions

Flat urothelial lesions are important because of their potential for carcinogenesis and development into invasive urothelial carcinomas; however, it is difficult for pathologists to detect early flat urothelial changes and accurately diagnose flat urothelial lesions. To predict the pathologic diagnosis and molecular abnormalities of flat urothelial lesions from pathologic images, we used artificial intelligence with an interpretable method. We studied 110 hematoxylin-and-eosin-stained slides of normal urothelium and flat urothelial lesions, including atypical urothelium, dysplasia, and carcinoma in situ, and performed next-generation sequencing to detect 17 kinds of molecular abnormalities. To generate an interpretable prediction, we developed a new method for segmenting urothelium and a new pathologic criteria-based artificial intelligence (PCB-AI) model. κ Statistics and accuracy measurements were used to evaluate the ability of the model to predict the pathologic diagnosis. The likelihood ratio test was performed to evaluate the logistic regression models for predicting molecular abnormalities. The diagnostic prediction of the PCB-AI model was almost in perfect agreement with the pathologists' diagnoses (weighted κ=0.98). PCB-AI significantly predicted some molecular abnormalities in an interpretable manner, including abnormalities of TP53 (P=0.02), RB1 (P=0.04), and ERCC2 (P=0.04). We generated a new method of obtaining accurate urothelial segmentation, interpretable prediction of pathologic diagnosis, and interpretable prediction of molecular abnormalities.

Pathologic Image Classification of Flat Urothelial Lesions Using Pathologic Criteria-Based Deep Learning

Pathologic diagnosis of flat urothelial lesions is subject to high interobserver variability. We expected that deep learning could improve the accuracy and consistency of such pathologic diagnosis, although the learning process is a black box. We therefore propose a new approach for pathologic image classification incorporating the diagnostic process of the pathologist into a deep learning method. A total of 267 H&E-stained slides of normal urothelium and urothelial lesions from 127 cases were examined. Six independent convolutional neural networks were trained to classify pathologic images according to six pathologic criteria. We then used these networks in the main training for the final diagnosis. Compared with conventional manual analysis, our method significantly improved the classification accuracy of images of flat urothelial lesions. The automated classification showed almost perfect agreement (weighted κ = 0.98) with the consensus reading. In addition, our approach provides the advantages of reliable diagnosis corresponding to histologic interpretation. We used deep learning to establish an automated subtype classifier for flat urothelial lesions that successfully combines traditional morphologic approaches and complex deep learning to achieve a learning mechanism that seems plausible to the pathologist.

Early diagnosis of bladder cancer by photoacoustic imaging of tumor-targeted gold nanorods

Detection and removal of bladder cancer lesions at an early stage is crucial for preventing tumor relapse and progression. This study aimed to develop a new technological platform for the visualization of small and flat urothelial lesions of high-grade bladder carcinoma in situ (CIS).We found that the integrin α5β1, overexpressed in bladder cancer cell lines, murine orthotopic bladder cancer and human bladder CIS, can be exploited as a receptor for targeted delivery of GNRs functionalized with the cyclic CphgisoDGRG peptide (Iso4). The GNRs@Chit-Iso4 was stable in urine and selectively recognized α5β1 positive neoplastic urothelium, while low frequency ultrasound-assisted shaking of intravesically instilled GNRs@Chit-Iso4 allowed the distribution of nanoparticles across the entire volume of the bladder. Photoacoustic imaging of GNRs@Chit-Iso4 bound to tumor cells allowed for the detection of neoplastic lesions smaller than 0.5 mm that were undetectable by ultrasound imaging and bioluminescence.

Could double stain for p53/CK20 be a useful diagnostic tool for the appropriate classification of flat urothelial lesions?

BackgroundThe differential diagnosis between flat urothelial lesions [reactive urothelial atypia (RUA), atypia of unknown significance (AUS), urothelial dysplasia (UD) and carcinoma in situ (CIS)] has relevant prognostic and therapeutic implications. This crucial distinction could be very challenging but it is currently performed on hematoxylin and eosin (H&E) slides, with a great amount of partially discordant and/or not conclusive findings of the potential adjunctive role of immunohistochemistry. Herein, we tested double staining (DS) for p53/CK20 to verify if p53(+) cells, CK20(+) cells and double-positive cells (DPCs) are differentially expressed among these lesions and if p53/CK20 could be a useful tool in this diagnostic setting. MethodsWe tested 50, 9, 36 and 29 consecutive and retrospectively enrolled cases of RUA, AUS, UD and CIS, respectively. p53(+) cells, CK20(+) cells and DPCs were evaluated and compared by adopting the appropriate statistic tests (Mann-Whitney U and Kruskal-Wallis tests). ResultsWe found that p53(+) cells (p = 0.000), CK20(+) cells (p = 0.000) and DPCs (p = 0.000) showed statistically significant differences among the different flat urothelial lesions. Besides, when dichotomized, both CIS and RUA are easily differentiable from their histological mimickers adopting all these markers; by contrast, AUS and UD did not reach statistically significant differences able to differentiate them from each other [p53(+) cells, p = 0.123; CK20(+) cells, p = 0.567; DPCs, p = 0.409], except if compared to CIS [AUS VS CIS: p53(+) cells, p = 0.013; CK20(+) cells, p = 0.000; DPCs, p = 0.000; UD vs CIS: p53(+) cells, p = 0.000; CK20(+) cells, p = 0.000; DPCs, p = 0.000]. Conclusionsp53(+) cells, CK20(+) cells and DPCs are differently expressed by flat urothelial lesions and p53/CK20 could be a time- and money-saving tool for the appropriate management of these lesions if applied to a routine scenario.

THE ROLE OF EZH2 AND ARID1A IN THE DIAGNOSIS OF FLAT UROTHELIAL LESIONS WITH ATYP

Background. Diagnosis of urothelial carcinoma in situ is of great importance because it has prognostic and therapeutic value.We aim to determine the utility of EZH2 and ARID1A as a new tool in the diagnosis of carcinoma in situ.Material and Methods. This retrospective cross-sectional study included Twenty-four specimens of flat urothelial lesions, twenty specimens of CIS, and 10 of normal adjacent urothelium that was taken by cystoscopic resection biopsy procedure. immunohistochemical expression of EZH2 and ARID1A. were evaluated in all studied cases.Results. All normal urothelium specimens showed high nuclear staining for ARID1A and negative nuclear staining for EZH2. High EZH2 expression was observed in 80 % of CIS specimens compared to 20 % of flat urothelial lesions with atypia (p=0.001 ), while high ARID1A expression was observed in 70.8 % of flat urothelial lesions with atypia compared to 25 % of CIS specimens (p=0.001). EZH2 was more accurate and specific in the diagnosis of carcinoma in situ.Conclusion. EZH2 and ARID1A are promising diagnostic markers for urothelial CIS. EZH2 is more accurate and specific than ARID1A in the diagnosis of carcinoma in situ versus other flat urothelial lesions.

The Genitourinary Pathology Society Update on Classification and Grading of Flat and Papillary Urothelial Neoplasia With New Reporting Recommendations and Approach to Lesions With Mixed and Early Patterns of Neoplasia.

The Genitourinary Pathology Society (GUPS) undertook a critical review of the recent advances in bladder neoplasia with a focus on issues relevant to the practicing surgical pathologist for the understanding and effective reporting of bladder cancer, emphasizing particularly on the newly accumulated evidence post-2016 World Health Organization (WHO) classification. The work is presented in 2 manuscripts. Here, in the first, we revisit the nomenclature and classification system used for grading flat and papillary urothelial lesions centering on clinical relevance, and on dilemmas related to application in routine reporting. As patients of noninvasive bladder cancer frequently undergo cystoscopy and biopsy in their typically prolonged clinical course and for surveillance of disease, we discuss morphologies presented in these scenarios which may not have readily applicable diagnostic terms in the WHO classification. The topic of inverted patterns in urothelial neoplasia, particularly when prominent or exclusive, and beyond inverted papilloma has not been addressed formally in the WHO classification. Herein we provide a through review and suggest guidelines for when and how to report such lesions. In promulgating these GUPS recommendations, we aim to provide clarity on the clinical application of these not so uncommon diagnostically challenging situations encountered in routine practice, while also importantly advocating consistent terminology which would inform future work.

Immunohistochemical staining patterns of Ki-67 and p53 in florid reactive urothelial atypia and urothelial carcinoma in situ demonstrate significant overlap

Flat urothelial lesions with atypia may pose significant diagnostic challenges. Given frequent increased proliferation rates in florid reactive urothelial atypia and limited studies on the interpretation of p53 stains in the urothelium (following current standard guidelines for correlation with P53 mutation status), we sought to further study the discriminatory value of Ki-67 and p53 for florid reactive urothelial atypia versus urothelial carcinoma in situ (CIS). Bladder specimens diagnosed as reactive urothelial atypia (n=40) and CIS (n=40) were assessed by immunohistochemical staining with antibodies for Ki-67, p53, CD44, and CK20. Immunoreactivity was scored based on percent cells positive for Ki-67 and pattern of reactivity with p53 (aberrant: diffuse strong positive or negative; normal: patchy/wild type). CD44 and CK20 reactivity patterns served as adjunctive internal validation controls for reactive urothelial atypia and CIS, as previously described. In reactive urothelial atypia, Ki-67 ranged from 0% to 90% (mean, 34%±26) with 30 cases (75%) having >10%. In CIS, Ki-67 ranged from 5% to 95% (mean, 50%±25) with 17 cases (43%) having >50%. In all 40 cases (100%) of reactive urothelial atypia, p53 expression had a wild-type pattern. In CIS, aberrant p53 expression was identified in 15 cases (37%): 3 cases (7%) were p53 negative (i.e. null phenotype) and 12 cases (30%) showed strong and diffuse nuclear reactivity (in >85% of cells). The remaining 25 cases (63%) of CIS had a p53 wild-type pattern of expression. Cytoplasmic CK20 immunoreactivity in umbrella cells was seen in 34 cases (85%) of reactive urothelial atypia, and 6 cases (15%) were negative. In addition, 35 cases (88%) of reactive urothelial atypia demonstrated full-thickness CD44 expression, while 5 cases (12%) had expression confined to the basal/parabasal layers of the urothelium. Strong and diffuse CK20 positivity was present in 39 cases (98%) of CIS, and patchy positivity was detected in 1 case (2%). None of the CIS cases overexpressed CD44: 16 cases (40%) showed focal expression in the nonneoplastic basal cell layer; 24 cases (60%) demonstrated no staining. In summary, Ki-67 has poor discriminatory value for reactive urothelial atypia versus CIS and adds little to the classic CK20/CD44 immunophenotype. While p53 sensitivity for CIS is relatively low (30%) and interpretation as either wild type or negative may be challenging in a small subset of cases, strong and diffuse nuclear reactivity was 100% specific in the distinction from florid reactive urothelial atypia in this cohort.

Immunohistochemistry in the workup of bladder biopsies: Frequency, variation and utility of use at an academic center

IntroductionFlat urothelial lesions fall into one of four diagnostic categories including urothelial carcinoma in-situ (CIS). There is morphologic overlap between the categories leading to immunohistochemistry (IHC) utilization in difficult cases. The purpose of this study was to examine the frequency, variation and utility of IHC use in bladder biopsy specimens over a 17 year period. MethodsA search of “CD44”, “p53”, and “CK20” keywords was conducted from the pathology files (1/1/2003 to 12/31/2017) on bladder biopsy specimens at our institution. Atypical (AUS), dysplastic (UD) and CIS rates were calculated. ResultsA total of 4597 cases were identified. IHC was performed on 345 specimens (7.5%, 345/4597). For cases without IHC (H&E only), the AUS rate was 4.8% (206/4252), UD rate was 9.4% (399/4252), and the CIS rate was 8.4% (359/4252). For IHC cases, the AUS rate was 5.2% (18/345), the UD rate was 8.1% (28/345), and the CIS rate was 11.3% (39/345). There was no statistical difference between the H&E only or IHC rates (p > 0.05). The absolute number IHC orders per year increased until 2011 (60 cases) but drastically declined over the last five years (5 total cases in 2017). The CIS rates have remained relatively constant. ConclusionWe found the AUS, UD and CIS rates were similar regardless of IHC use. Our institution was an early adopter of IHC and it quickly fell out of favor. We agree with the ISUP in that IHC has limited clinical utility for flat urothelial lesions and morphology remains the gold standard.

Detection of genetic instability and proliferative activity in urinary bladder intraepithelial flat lesions using UroVysion fluorescence in-situ hybridization and flow cytometry

Background Chromosomal abnormalities play a key role in neoplastic potential for flat urothelial lesions, irrespective of whether cytologic atypia is present or not. Having information about lesions with malignant potential in the urinary bladder can help clinicians in taking logical actions concerning management after urinary bladder-conserving resections.Aim The aim of the study was to differentiate various flat urothelial lesions by detecting its genetic abnormalities and proliferative activity using UroVysion fluorescence in-situ hybridization and flow cytometry.Patients and methods Ninety-eight paraffin-embedded tissue sections from 35 patients with infiltrative bladder cancer and flat urothelial lesions [reactive atypia, hyperplasia, dysplasia, carcinoma in situ (CIS), squamous metaplasia, glandular metaplasia, histologically normal urothelium in patients with tumor and control] were assessed by flow cytometry and fluorescence in-situ hybridization using UroVysion targeted against the band 9p21 locus and pericentromeric areas of chromosomes 3, 7, and 17.Results Gain of at least one of the chromosomes was observed in 60% of samples being more than 90% in CIS, dysplasia, and infiltrating bladder tumors. The mean percentage of cells with polysomy in reactive atypia, flat urothelial hyperplasia, glandular metaplasia, keratinizing squamous metaplasia, and histologically normal urothelium in patients with tumors ranged from 12 to 20%, but in CIS and dysplasia it was 67 and 44%, respectively. Deletion of the P16 locus was detected in 49% of lesions. Flow cytometry displayed aneuploidy pattern in 60% of lesions with higher percentage in CIS, dysplasia, and infiltrating bladder tumors. High S-phase fraction (SPF) was observed in 44% of samples. High S phase showed significant association with polysomy and DNA ploidy (P=0.02 and 0.01, respectively). In addition, there was association between polysomy expression with del 9p21 (P=0.03) and between del 9p21 with DNA ploidy and SPF (P=0.003 and 0.02).Conclusions Our analysis has shown that combined evaluation of flat urothelial lesions using UroVysion, DNA ploidy, and SPF help in the recognition of these lesions, especially those with higher genetic abnormalities and having higher S phase because follow up of these lesions showed a higher tendency for malignant transformation. Furthermore, estimation of continuing chromosomal instability such as polysomy, DNA aneuploidy, and high SPF might deliver further interpretation of urinary bladder cancer risk assessment away from the position and histopathology of precancerous lesions.

Immunohistochemical Study of the Role of CK20, p53 and Ki-67 in Differentiation of Some Urothelial Lesions and Urothelial Carcinoma of the Urinary Bladder

AbstractBackground: Bladder cancer is the most common malig-nancy affecting the urinary tract. Distinguishing between urothelial dysplasia and carcinoma in situ based on histopatho-logical features alone is often difficult.Aim of Work: The aim of the current work is to distinguish between urothelial hyperplasia, urothelial dysplasia from urothelial carcinoma by using CK20, p53 and Ki-67 immu-nomarkers, determine the pattern and extent of their immuno-reactivity and correlate immuno-histochemical results with the clinicopathological parameters.Material and Methods: Fifty cases of urothelial carcinoma (38 cases) and some flat urothelial lesions (12 cases) were collected retrospectively. Tissue specimens were in the form of radical cystectomy (nine specimens) and transurethral resection of the tumor (TURT) (forty one specimens). They were stained by H&E, CK20, p53 and Ki-67 for immunohis-tochemical study. The relationship between their expression and the available clinicopathological features were evaluated.Results: CK20, p53 and Ki-67 expressions can signifi-cantly differentiate urothelial hyperplasia from urothelial dysplasia as the whole panel is negative in urothelial hyper-plasia and positive with scattered expression in urothelial dysplasia. Also pattern of expression of CK20, p53 and Ki-67 expression are suggesting for the diagnosis of either urothelial dysplasia or urothelial carcinoma in situ as their expressions show diffuse positivity throughout the urothelium in urothelial carcinoma in situ. These markers were statistically significant in grading of urothelial carcinoma as higher tumor grade associated with decreased CK20 expression and in-creased p53 and Ki-67 expression. CK20 expression was statistically significant in tumor stage as higher tumor stage was associated with decreased CK20 expression.Conclusions: Abnormal CK20 expression in urothelial cells plus overexpression of p53 and Ki-67 are indicators of dysplastic change in urothelial mucosa. A panel of CK20, p53 and Ki-67 can be a useful tool to confirm the diagnosis of CIS and can be helpful to distinguish it from dysplastic changes. Combined use of these markers may be helpful in assigning grade of urothelial carcinoma especially when histologic features are borderline.

AB065. Confocal laser endomicroscopy for diagnosing malignant bladder tumour: a pilot study

BackgroundTo perform an exploratory investigation on confocal laser endomicroscopy (CLE) in the diagnosis of malignant bladder tumour.MethodsFrom June 10 to July 11, 2017, patients initially diagnosed with bladder cancer by cystoscopy were hospitalised and had undergone white light cystoscopy (WLC) + CLE examination followed by transurethral resection of bladder tumour (TURBT) on suspected lesions. WLC and CLE imaging results were recorded and validated by pathologic specimens.ResultsLesions confirmed by histopathology were 3 low-grade non-invasive papillary urothelial carcinomas, 1 high-grade non-invasive papillary urothelial carcinoma, 1 low-grade invasive urothelial carcinoma, 1 high-grade invasive urothelial carcinoma, 1 carcinoma in situ (CIS), 1 high-grade dysplasia, 1 cystitis glandularis, 1 chronic inflammation, and 1 scar tissue. For CLE images in the normal urothelium, three layers of cells with different presentation were observed, namely, the superficial umbrella cells, the intermediate cells smaller in size and uniformly shaped, and the capillary network in the lamina propria. For non-invasive urothelial carcinoma, tumour cells appeared as papillary lesions growing from fibrovascular cores, with low-grade cells appearing monomorphic and more cohesively arranged, and high-grade cells relatively pleomorphic, more disorganised and with tortuous blood vessels in the fibrovascular core. For invasive urothelial carcinoma, tumour cells invaded the lamina propria, with uniform appearances, poor cohesion and indistinct cellular borders, and high-grade ones were more pleomorphic. CIS and inflammation both appeared as erythematous patch-like flat lesions under WLC and sometimes difficult to differentiate. Under CLE, the former appeared as dysplastic and disorganised cells with indistinct cellular borders, with intact lamina propria, and inflammatory cells were discovered as infiltrative clusters in the lamina propria that were uniformly shaped and loosely connected. Dysplasia appeared somewhat similar compared with CIS under WLC, but with lower cellular irregularity as confirmed with pathology. Cellular appearance and structure in scar tissue were similar to that in the normal urothelium, but superficial umbrella cells were more likely absent, with thinner cell layers, and inflammatory infiltration was sometimes discovered in the lamina propria.ConclusionsCLE provides real-time cellular imaging of the urothelium, and shows promising potential for clinical diagnosis, especially in differentiating flat urothelial lesions. Large prospective studies are required for further validation.

Confocal laser endomicroscopy for diagnosing malignant bladder tumour: a pilot study

Objective To perform an exploratory investigation on confocal laser endomicroscopy (CLE) in the diagnosis of malignant bladder tumour. Methods From June 10 to July 11, 2017, 6 male bladder cancer patients underwent white light cystoscopy (WLC)+ CLE examination, aging 64-86 years (median 72 years). All patients received TURBT on suspected lesions. WLC and CLE imaging results were recorded and validated by pathologic specimens. Results Lesions confirmed by histopathology were 3 low grade non-invasive papillary urothelial carcinomas, 1 high grade non-invasive papillary urothelial carcinoma, 1 low grade invasive urothelial carcinoma, 1 high grade invasive urothelial carcinoma, 1 carcinoma in situ (CIS), 1 high grade dysplasia, 1 cystitis glandularis, 1 chronic inflammation, and 1 scar tissue. For CLE images in the normal urothelium, three layers of cells with different presentation were observed, namely, the superficial umbrella cells, the intermediate cells smaller in size and uniformly shaped, and the capillary network in the lamina propria. For non-invasive urothelial carcinoma, tumour cells appeared as papillary lesions growing from fibrovascular cores, with low grade cells appearing monomorphic and more cohesively arranged, and high grade cells relatively pleomorphic, more disorganised and with tortuous blood vessels in the fibrovascular core. For invasive urothelial carcinoma, tumour cells invaded the lamina propria, with uniform appearances, poor cohesion and indistinct cellular borders, and high grade ones were more pleomorphic. CIS and inflammation both appeared as erythematous patch-like flat lesions under WLC and sometimes difficult to differentiate. Under CLE, the former appeared as dysplastic and disorganised cells with indistinct cellular borders, with intact lamina propria, and inflammatory cells were discovered as infiltrative clusters in the lamina propria that were uniformly shaped and loosely connected. Dysplasia appeared somewhat similar compared with CIS under WLC, but with lower cellular irregularity as confirmed with pathology. Cellular appearance and structure in scar tissue was similar to that in the normal urothelium, but superficial umbrella cells were more likely absent, with thinner cell layers, and inflammatory infiltration was sometimes discovered in the lamina propria. Conclusions CLE provides real-time cellular imaging of the urothelium, and shows promising potential for clinical diagnosis, especially in differentiating flat urothelial lesions. Large prospective studies are required for further validation. Key words: Bladder cancer; Cystoscopy; Confocal laser endomicroscopy; Pathological diagnosis

Flat Urothelial Lesions With Atypia: Interobserver Concordance and Added Value of Immunohistochemical Profiling.

Distinguishing urothelial carcinoma in situ (CIS) from other flat lesions of the urinary bladder with cytologic atypia is critically important for the management of patients with bladder neoplasia. However, there is high interpathologist variability in making these distinctions. The aim of this study is to assess interobserver agreement between general and specialized genitourinary pathologists, and to compare these diagnoses with those rendered after an immunohistochemical panel is performed. We hypothesized that addition of a set of immunohistochemical stains would reduce the number of cases classified within intermediate categories of atypia of uncertain significance and low-grade dysplasia. Two genitourinary pathologists independently assessed haematoxylin and eosin (H&E)-stained sections of 127 bladder biopsies from each of the 4 International Society of Urological Pathology/World Health Organization categories of flat lesions diagnosed by general pathologists. A subset of biopsies from 49 patients was reassessed after staining with a 3-antibody panel (CD44, CK20, and p53) and the results were correlated with patient follow-up. Based on these immunohistochemistry (IHC) stains, 26 cases (53.1%) were recategorized. Of most clinical importance, 5 of 27 cases (18.5%) originally diagnosed as either atypia of uncertain significance or low-grade dysplasia were recategorized as CIS, and recurrent disease was identified on subsequent biopsies. None of the 10 cases diagnosed as CIS based on H&E stains were recategorized. This triad of IHC stains can improve the precision of pathologic diagnosis of histologically atypical urothelial lesions of flat bladder mucosa. We recommend that pathologists apply this set of IHC stains to such lesions they find problematic based on H&E stains.

Significance of immunohistochemistry and DNA ploidy in differential diagnosis of flat urothelial lesions and conditions using CK20, p53, CD44, ErbB-2, and image cytometry

Significance of immunohistochemistry and DNA ploidy in differential diagnosis of flat urothelial lesions and conditions using CK20, p53, CD44, ErbB-2, and image cytometry

Nuclear size measurement for distinguishing urothelial carcinomas from reactive urothelium on tissue sections.

BackgroundPathological diagnosis of urothelial carcinoma (UC) is primarily based on cytological atypia. It has previously been shown that high-grade (HG) UC, particularly UC in situ cells (CIS), can be over five times the size of a lymphocyte. However, this has not been demonstrated in comparison to reactive urothelium. The objective of this study was to empirically compare the difference in nuclear size of UC cells with reactive urothelial cells.MethodsUsing CellSens imaging software, we measured urothelial nuclear length (l) and width (w) on digital images of H&E sections. The area (a) of a nucleus was calculated based on the oval shape of most urothelial cells. Lymphocytes were measured to calculate normalized urothelial linear and area ratios.ResultsA total of 1085 urothelial cell nuclei from 60 cases were measured from reactive urothelium, low grade (LG) UC, HG UC and CIS. CIS nuclei were found to have an a 2.75 times larger than reactive nuclei (p < 0.001). A nuclear size cut-off of 11 um for l and 7 um for w was found to be sensitive [98.09 % (95 % CI: 95.60–99.38 %) and 89.31 % (95 % CI: 83.6–91.82 %) for l and w, respectively] and specific [92.60 % (95 % CI: 87.13–95.82 %) and 85.71 % (95 % CI: 79.49–90.63 %) for l and w, respectively] for distinguishing CIS from reactive atypia.ConclusionsNuclear morphometry can be used to differentiate CIS from reactive atypia. A l over 11 um and a w over 7 um and is highly sensitive and specific for CIS compared to reactive urothelium. This difference in nuclear size may be used as a tool for differentiating the flat urothelial lesions from reactive urothelium in daily practice.

Reducing understaging of bladder cancer with the aid of photodynamic cystoscopy

BackgroundThe authors evaluated the role of photodynamic cystoscopy in the detection of additional urothelial lesions, mainly carcinoma in situ, that would not be detected solely with white light cystoscopy, leading to disease understaging. MethodsFrom 2009 to 2011, 70 patients underwent white light cystoscopy, followed by photodynamic cystoscopy (blue light system, Karl Storz, Tuttlingen, Germany). Preoperatively they were instilled intravesically with 50ml of Hexvix (Hexaminolevulinate hexylester). We recorded all lesions found with white light cystoscopy and the additional lesions revealed by blue light cystoscopy. Afterward all lesions were removed and sent for pathologic evaluation. ResultsSeventeen patients (24.3%) had primary tumors while 53 patients (75.7%) had recurrent disease. In 53 out of 70 patients (75.7%) white light cystoscopy revealed urothelial lesions. In the rest 17 patients who had no findings with white light cystoscopy, blue light cystoscopy revealed 7 cases of Cis (41.2%) and 8 cases of dysplasia (47%). In the group of patients with visible lesions in white light cystoscopy photodynamic cystoscopy revealed additional carcinoma in situ in 22 patients. Altogether additional carcinoma in situ cases found with the aid of photodynamic cystoscopy were 29 out of 70 cases (41.4%). ConclusionsPhotodynamic cystoscopy is the most efficient diagnostic procedure for flat urothelial lesions and especially for carcinoma in situ. It is significantly more sensitive than conventional white light cystoscopy in Cis diagnosis, thus reducing understaging of the disease and consequently improving recurrence and progression rates.

Are we closer to seeing carcinoma in situ in the upper urinary tract?

IntroductionThere is observed increase in detection rate of upper urinary tract urothelial cancer worldwide. This is a result of improved imaging as well as implementation of novel technologies of direct visualization of upper urinary tract. Standard techniques still remain insufficient to diagnose flat urothelial lesions. Carcinoma in situ is characterized by flat disordered proliferation of urothelial cells with marked cytologic abnormality, which occur within one cell layer as well as full thickness urothelium and therefore requires a better technology to pick up early and subtle mucosal changes.Material and methodsThe review presents available diagnostic tools in detection of upper urinary tract urothelial cancer and their ability to depict carcinoma in situ. ResultsUreterorenoscopy is an investigation of choice as various promising techniques are under pilot investigations to enhance visualization of upper urinary tract carcinoma in situ. So far only photodynamic diagnosis has been reported to be as effective in detection of carcinoma in situ in the upper as within the lower urinary tract.ConclusionsAlthough we are close to see upper urinary tract carcinoma in situ all new promising diagnostic techniques still require further validation in multicenter clinical trials to indicate any change to current recommendations.

Initial experience of combined use of photodynamic diagnosis and narrow band imaging for detection of flat urothelial lesion.

This preliminary study is the first report to compare photodynamic diagnosis (PDD) with narrow band imaging (NBI) in the same patients with flat urothelial lesions suspicious of carcinoma in situ (CIS) of the bladder. Between November 26, 2012 and April 1, 2013, 10 patients underwent transurethral resection of bladder tumor using PDD and NBI simultaneously because of suspicion of CIS. The bladder was mapped first under white light (WL), then under NBI, and subsequently under blue light in odd-numbered patients. The bladder was mapped first under WL, then under blue light, and subsequently under NBI in even-numbered patients. Biopsies were carried out from all suspicious areas, noting whether NBI, PDD or both detected lesions. Random cold cup biopsies from healthy mucosa of bladder were performed from lesions negative on PDD and NBI. The sensitivity and specificity of PDD for detection of CIS and dysplasia were 0.916 and 0.827, respectively. The sensitivity and specificity of NBI for detection of CIS and dysplasia were 0.625 and 0.879. The area under the curve (AUC) for detection of CIS and dysplasia was 0.872 with PDD and 0.752 with NBI. The AUC with combined use of PDD and NBI was 0.918. There was no cancer or dysplasia identified in 43 lesions that were both PDD- and NBI-negative. When both PDD and NBI are negative, the possibility of CIS or dysplasia might be very low. The usefulness of the combination of PDD with NBI was suggested in this study.