Real-world performance of Uromonitor® in urothelial bladder cancer detection: a multicentric trial.

OBJECTIVES: To compare Uromonitor® (U-Monitor Lda, Porto, Portugal), a multitarget DNA assay that detects mutated proto-oncogenes (telomerase reverse transcriptase [TERT], fibroblast growth factor receptor 3 [FGFR-3], Kirsten rat sarcoma viral oncogene homologue [KRAS]), with urine cytology in the urine-based diagnosis of urothelial carcinoma of the bladder (UCB) within a multicentre real-world setting. PATIENTS AND METHODS: This multicentre, prospective, double-blind study was conducted across four German urological centres from 2019 to 2024. We evaluated the diagnostic performance of Uromonitor compared to urine cytology in a cohort of patients with UCB and in healthy controls within a real-world setting. Sensitivity, specificity, positive-predictive value (PPV), negative-predictive value (NPV), and accuracy of the tests were measured, in addition to multivariate analyses to assess the ability of individual proto-oncogene mutations in detecting UCB. The biometric sample size was designed to achieve a 10% difference in sensitivity. RESULTS: Patients with UCB comprised 63.7% (339/532) of the study group. Uromonitor showed a sensitivity, specificity, PPV, NPV, accuracy, and an area-under-the-curve of 49.3%, 93.3%, 92.8%, 51.1%, 65.2%, and 0.713%, respectively. These metrics did not demonstrate statistical superiority over urine cytology in terms of sensitivity (44.6%; P = 0.316). Moreover, the comparison of additional test parameters, as well as the comparison within various sensitivity analyses, yielded no significant disparity between the two urinary tests. Multivariate logistic regression underscored the significant predictive value of a positive Uromonitor for detecting UCB (odds ratio [OR] 9.03; P < 0.001). Furthermore, mutations in TERT and FGFR-3 were independently associated with high odds of UCB detection (OR 13.30 and 7.04, respectively), while KRAS mutations did not exhibit predictive capability. CONCLUSION: Despite its innovative approach, Uromonitor fell short of confirming the superior results anticipated from previous studies in this real-world setting. The search for an optimal urine-based biomarker for detecting and monitoring UCB remains ongoing. Results from this study highlight the complexity of developing non-invasive diagnostic tools and emphasise the importance of continued research efforts to refine these technologies.

Automated Ki-67 labeling index assessment in prostate cancer using artificial intelligence and multiplex fluorescence immunohistochemistry.

The Ki-67 labeling index (Ki-67 LI) is a strong prognostic marker in prostate cancer, although its analysis requires cumbersome manual quantification of Ki-67 immunostaining in 200-500 tumor cells. To enable automated Ki-67 LI assessment in routine clinical practice, a framework for automated Ki-67 LI quantification, which comprises three different artificial intelligence analysis steps and an algorithm for cell-distance analysis of multiplex fluorescence immunohistochemistry (mfIHC) staining, was developed and validated in a cohort of 12,475 prostate cancers. The prognostic impact of the Ki-67 LI was tested on a tissue microarray (TMA) containing one 0.6 mm sample per patient. A 'heterogeneity TMA' containing three to six samples from different tumor areas in each patient was used to model Ki-67 analysis of multiple different biopsies, and 30 prostate biopsies were analyzed to compare a 'classical' bright field-based Ki-67 analysis with the mfIHC-based framework. The Ki-67 LI provided strong and independent prognostic information in 11,845 analyzed prostate cancers (p < 0.001 each), and excellent agreement was found between the framework for automated Ki-67 LI assessment and the manual quantification in prostate biopsies from routine clinical practice (intraclass correlation coefficient: 0.94 [95% confidence interval: 0.87-0.97]). The analysis of the heterogeneity TMA revealed that the Ki-67 LI of the sample with the highest Gleason score (area under the curve [AUC]: 0.68) was as prognostic as the mean Ki-67 LI of all six foci (AUC: 0.71 [p = 0.24]). The combined analysis of the Ki-67 LI and Gleason score obtained on identical tissue spots showed that the Ki-67 LI added significant additional prognostic information in case of classical International Society of Urological Pathology grades (AUC: 0.82 [p = 0.002]) and quantitative Gleason score (AUC: 0.83 [p = 0.018]). The Ki-67 LI is a powerful prognostic parameter in prostate cancer that is now applicable in routine clinical practice. In the case of multiple cancer-positive biopsies, the sole automated analysis of the worst biopsy was sufficient. © 2023 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

PD-L1 expression in tumor and inflammatory cells is associated with favorable tumor features and favorable prognosis in muscle-invasive urothelial carcinoma of the bladder not treated by immune checkpoint inhibitors.

BACKGROUND: A high level of PD-L1 expression is the most relevant predictive parameter for response to immune checkpoint inhibitor (CPI) therapy in urinary bladder cancer. Existing data on the relationship between PD-L1 expression and the natural course of disease are controversial and sparse. METHODS: To expand our understanding of the relationship between PD-L1 expression and parameters of cancer aggressiveness, PD-L1 was analyzed on tissue microarrays containing 2710 urothelial bladder carcinomas including 512 patients with follow-up data who underwent radical cystectomy and follow-up therapies in the pre-immune checkpoint inhibitor therapy era. RESULTS: Tumor cell positivity in ≥10% of cells were seen in 513 (20%) and an immune cell positivity occurred in 872 (34%) of 2566 interpretable cancers. PD-L1 positivity in tumor cells increased from pTaG2 low grade (0.9% positive) to pTaG3 high grade (4.1%; p = 0.0255) and was even higher in muscle-invasive (pT2-4) carcinomas (29.3%; p < 0.0001). However, within pT2-4 carcinomas, PD-L1 positivity was linked to low pT stage (p = 0.0028), pN0 (p < 0.0001), L0 status (p = 0.0005), and a better prognosis within 512 patients with cystectomy who never received CPIs (p = 0.0073 for tumor cells and p = 0.0086 for inflammatory cells). PD-L1 staining in inflammatory cells was significantly linked to PD-L1 staining in tumor cells (p < 0.0001) and both were linked to a positive p53 immunostaining (p < 0.0001). CONCLUSION: It cannot be fully excluded that the strong statistical link between PD-L1 status and favorable histological tumor features as well as better prognosis could influence the outcome of studies evaluating CPIs in muscle-invasive urothelial carcinoma.

Cytokeratin 20 expression is linked to stage progression and to poor prognosis in advanced (pT4) urothelial carcinoma of the bladder.

Cytokeratin 20 (CK20) expression is limited to umbrella cells in the normal urothelium. Since CK20 is often upregulated in neoplastic urothelial cells including dysplasia and carcinoma in situ, immunohistochemical CK20 analysis is often used for the assessment of bladder biopsies. CK20 expression is a feature of luminal bladder cancer subtype, but its prognostic relevance is disputed. In this study, we investigated CK20 on >2700 urothelial bladder carcinomas in a tissue microarray format by immunohistochemistry. Cytoplasmic and membranous CK20 staining was seen in 1319 (51.8%) cancers. The fraction of CK20 positive and especially strongly positive cases increased from pTaG2 low grade (44.5% strongly positive) and pTaG2 high grade (57.7%) to pTaG3 high grade (62.3%; p = 0.0006) but was lower in muscle-invasive (pT2-4) carcinomas (51.1% in all pTa vs. 29.6% in pT2-4; p < 0.0001). Within pT2-4 carcinomas, CK20 positivity was linked to nodal metastasis and lymphatic vessel invasion (p < 0.0001 each) and to venous invasion (p = 0.0177). CK20 staining was unrelated to overall patient survival if all 605 pT2-4 carcinomas were jointly analyzed but subgroup analyses revealed a significant association of CK20 positivity with favorable prognosis in 129 pT4 carcinomas (p = 0.0005). CK20 positivity was strongly linked to the expression of GATA3 (p < 0.0001), another feature of luminal bladder cancer. The combined analysis of both parameters showed best prognosis for luminal A (CK20+/GATA3+, CK20+/GATA3-) and worst outcome for luminal B (CK20-/GATA3+) and basal/squamous (CK20-/GATA3-) in pT4 urothelial carcinomas (p = 0.0005). In summary, the results of our study demonstrate a complex role of CK20 expression in urothelial neoplasms including neoexpression in pTa tumors, a subsequent loss of CK20 expression in a subset of tumors progressing to muscle-invasion, and a stage dependent prognostic role in muscle-invasive cancers.

Altered p53/p16 expression is linked to urothelial carcinoma progression but largely unrelated to prognosis in muscle-invasive tumors.

BACKGROUND: Most inactivating p53 mutations result in a nuclear p53 accumulation - detectable by immunohistochemistry (IHC). p53 alterations leading to a complete lack of p53 protein and absence of immunostaining do also occur - not easily detectable by IHC. p16 is upregulated in p53 inactivated cells. We hypothesized that a positive p16 IHC may help to distinguish p53 inactivation in IHC negative cases. MATERIAL AND METHODS: We investigated p53 and p16 immunostaining on 2710 urothelial bladder carcinomas in a tissue microarray format to understand their impact in relation to clinicopathological parameters of disease progression and patient outcome. RESULTS: p16 immunostaining was absent in normal urothelium but occurred in 63.5% (30.4% strong) of cancers. p16 strongly positive cases increased from pTaG2 low-grade (9.6%) to pTaG3 high-grade tumors (46.5%, p < .0001) but decreased from pTaG3 to pT4 (33.3%; p = .0030). Among pT2-4 carcinomas, p16 positivity was linked to high-grade (p = .0005) but unrelated to overall survival. p53 staining was negative in 8.4%, very weak in 15.4%, weak in 55.3%, strong in 4.7%, and very strong in 16.2% cancers. p53 negative (potentially p53 null phenotype), strong, and very strong p53 positivity increased from pTaG2 low-grade to pTaG3 high-grade tumors (p < .0001) and from pTaG3 to pT2-4 cancers (p = .0007). p53 staining was largely unrelated to histopathological parameters or patient prognosis among pT2-4 carcinomas, except of p53 strong/very strong immunostaining. p16 expression predominated in tumors with very strong, strong, and negative p53 staining and the combination of p53 negative/p16 strongly positive cancers was linked to features of tumor aggressiveness. CONCLUSION: Aberrant p53 and p16 immunostaining increases during grade and stage progression although p53 negative and p16 positive immunostaining lack prognostic significance in pT2-4 carcinomas. Potential diagnostic features are that high level p16 expression is limited to neoplastic urothelium and p53 null phenotype to aggressive cancers (grade 3 and invasive).

Subjective Outcome after Discontinuation of Botulinum Toxin-A Detrusor Injection for Lower Urinary Tract Disorders: The Majority Suffers!

INTRODUCTION: According to the EAU guidelines, botulinum toxin type A (BoNT-A) detrusor injections are one of the last options in the management of overactive bladder before opting for invasive surgery. So far, there have been no studies dedicated exclusively to such patients who have undergone this treatment and in whom this treatment has presumably failed. From the patient's point of view, there are questions regarding what reasons led to discontinuation, how patients perceive their BoNT-A treatment in hindsight, what further treatment do these patients receive, and how satisfied such patients are with their current situation. METHODS: A database of clinical and inpatient records was searched, and 695 records from 406 patients were identified in a 6-year period, who had received BoNT-A detrusor injections. There were 255 cases that were treated with BoNT-A injections into the detrusor muscle where the therapy was not repeated for at least 12 months (= suspected treatment failures). Interviews with these patients were conducted by mail and phone, and 115 questionnaires could be included in the analysis. RESULTS: From the subjective and prospective points of view of the patients, the most common reason for stopping the therapy was a lack of efficacy of BoNT-A injections (39.1%). For 26.1% of all patients, side effects were a reason for dissatisfaction but never a reason for discontinuation. For 10.4%, the reason for stopping the therapy was spontaneous improvement. 35.6% of the respondents had no follow-up therapy. Those with a follow-up therapy mostly returned to anticholinergic treatment (33%). Operations were carried out on 13%, of which about half were highly invasive. For 71.3% of those patients, who were under any current therapy, this therapy led to no improvement or only some improvement of the symptoms. Surprisingly, 50.4% of the respondents would choose to undergo BoNT-A injection therapy again. DISCUSSION/CONCLUSION: The majority of patients who did not continue BoNT-A therapy are still suffering from lower urinary tract symptoms. The lack of efficacy was the reason for stopping the BoNT-A injection therapy for less than half of the patients. From the patient's point of view, reasons other than the effectiveness also seem to be relevant in the choice of the treatment. When changing therapy, most returned to drug treatment. However, for the majority of patients with any follow-up therapy, this therapy could not improve the symptoms.

A non-inferiority comparative analysis of micro-ultrasonography and MRI-targeted biopsy in men at risk of prostate cancer.

OBJECTIVE: To compare the efficacy of multiparametric magnetic resonance imaging (mpMRI)-directed and micro-ultrasonography (micro-US)-directed biopsy for detecting clinically significant (Grade Group >1) prostate cancer (csPCa). MATERIALS AND METHODS: A total of 203 patients were prospectively enrolled at three institutions across Germany and Austria in the period from January 2019 to December 2019. During each biopsy, the urologist was blinded to the mpMRI report until after the micro-US targets had been assessed. After unblinding, targets were then sampled using software-assisted fusion, followed by systematic samples. The primary outcome measure was non-inferiority of micro-US to detect csPCa, with a detection ratio of at least 80% that of mpMRI. RESULTS: A total of 79 csPCa cases were detected overall (39%). Micro-US-targeted biopsy detected 58/79 cases (73%), while mpMRI-targeted biopsy detected 60/79 (76%) and non-targeted (completion sampling) samples detected 45/79 cases (57%). mpMRI-targeted samples alone detected 7/79 (9%) csPCa cases which were missed by micro-US-targeted and non-targeted samples. Three of these seven were anterior lesions with 2/7 in the transition zone. Micro-US-targeted samples alone detected 5/79 (6%) and completion sampling alone detected 4/79 cases (5%). Micro-US was non-inferior to mpMRI and detected 97% of the csPCa cases detected by mpMRI-targeted biopsy (95% CI 80-116%; P = 0.023). CONCLUSIONS: This is the first multicentre prospective study comparing micro-US-targeted biopsy with mpMRI-targeted biopsy. The study provides further evidence that micro-US can reliably detect cancer lesions and suggests that micro-US biopsy might be as effective as mpMRI for detection of csPCA. This result has significant implications for increasing accessibility, reducing costs and expediting diagnosis.

Nomograms including the UBC® Rapid test to detect primary bladder cancer based on a multicentre dataset.

OBJECTIVES: To evaluate the clinical utility of the urinary bladder cancer antigen test UBC® Rapid for the diagnosis of bladder cancer (BC) and to develop and validate nomograms to identify patients at high risk of primary BC. PATIENTS AND METHODS: Data from 1787 patients from 13 participating centres, who were tested between 2012 and 2020, including 763 patients with BC, were analysed. Urine samples were analysed with the UBC® Rapid test. The nomograms were developed using data from 320 patients and externally validated using data from 274 patients. The diagnostic accuracy of the UBC® Rapid test was evaluated using receiver-operating characteristic curve analysis. Brier scores and calibration curves were chosen for the validation. Biopsy-proven BC was predicted using multivariate logistic regression. RESULTS: The sensitivity, specificity, and area under the curve for the UBC® Rapid test were 46.4%, 75.5% and 0.61 (95% confidence interval [CI] 0.58-0.64) for low-grade (LG) BC, and 70.5%, 75.5% and 0.73 (95% CI 0.70-0.76) for high-grade (HG) BC, respectively. Age, UBC® Rapid test results, smoking status and haematuria were identified as independent predictors of primary BC. After external validation, nomograms based on these predictors resulted in areas under the curve of 0.79 (95% CI 0.72-0.87) and 0.95 (95% CI: 0.92-0.98) for predicting LG-BC and HG-BC, respectively, showing excellent calibration associated with a higher net benefit than the UBC® Rapid test alone for low and medium risk levels in decision curve analysis. The R Shiny app allows the results to be explored interactively and can be accessed at www.blucab-index.net. CONCLUSION: The UBC® Rapid test alone has limited clinical utility for predicting the presence of BC. However, its combined use with BC risk factors including age, smoking status and haematuria provides a fast, highly accurate and non-invasive tool for screening patients for primary LG-BC and especially primary HG-BC.

Transurethral suturing of bladder following transurethral excision of female vesico-urachal diverticula: a novel technique with long-term follow-up results.

BACKGROUNDS: This study aims to describe and evaluate outcomes of transurethral surgery-natural orifice transluminal endoscopic surgery (TUS-Notes) technique in patients treated with transurethral excision of vesico-urachal diverticula (VD). METHODS: Patients who underwent TUS-Notes following transurethral VD excision due to recurrent urinary tract infection (rUTI) since 2013 were included in this prospective non-randomized cohort study. Under cystoscopic guidance VD and surrounding bladder wall was resected until the fatty tissue using monopolar resectoscope. The specimen was removed with a grasper through the cystoscope. TUS-Notes technique was performed with Minimal Suturing Device (MSD-Ney®). The needle of the suture was shaped according to suturing position and loaded into MSD-Ney. They were inserted into the bladder under cystoscopic guidance transurethrally. Once the defect was sutured properly, an extracorporeal knot was prepared and tied. The length of the operation (LOO), and perioperative complications according to the Clavien-Dindo grading system were noted. The integrity of the bladder was checked with cystography to assess objective cure. Subjective cure was evaluated with Patient Global Impression of Improvement (PGI-I) scale. RESULTS: The follow-up period of 65 participants varied from three months to eight years. The median LOO was 37 min. A Clavien grade-3 complication was observed in one patient. Peroperative failure was not noted. The median duration of hospital stays, and catheterization time was three days. Objective cure rate and subjective cure rates were 100%. UTI was not noted after surgery. CONCLUSIONS: Transurethral complete excision of VD is an acceptable technique to prevent rUTI. The TUS-Notes technique provides a successful minimal invasive treatment option for the treatment of bladder defects. CONCISE: Transurethral suturing of urinary bladder.

Prediction of Response to Cisplatin-Based Neoadjuvant Chemotherapy of Muscle-Invasive Bladder Cancer Patients by Molecular Subtyping including KRT and FGFR Target Gene Assessment.

Patients with muscle-invasive urothelial carcinoma achieving pathological complete response (pCR) upon neoadjuvant chemotherapy (NAC) have improved prognosis. Molecular subtypes of bladder cancer differ markedly regarding sensitivity to cisplatin-based chemotherapy and harbor FGFR treatment targets to various content. The objective of the present study was to evaluate whether preoperative assessment of molecular subtype as well as FGFR target gene expression is predictive for therapeutic outcome­rate of ypT0 status­to justify subsequent prospective validation within the "BladderBRIDGister". Formalin-fixed paraffin-embedded (FFPE) tissue specimens from transurethral bladder tumor resections (TUR) prior to neoadjuvant chemotherapy and corresponding radical cystectomy samples after chemotherapy of 36 patients were retrospectively collected. RNA from FFPE tissues were extracted by commercial kits, Relative gene expression of subtyping markers (e.g., KRT5, KRT20) and target genes (FGFR1, FGFR3) was analyzed by standardized RT-qPCR systems (STRATIFYER Molecular Pathology GmbH, Cologne). Spearman correlation, Kruskal−Wallis, Mann−Whitney and sensitivity/specificity tests were performed by JMP 9.0.0 (SAS software). The neoadjuvant cohort consisted of 36 patients (median age: 69, male 83% vs. female 17%) with 92% of patients being node-negative during radical cystectomy after 1 to 4 cycles of NAC. When comparing pretreatment with post-treatment samples, the median expression of KRT20 dropped most significantly from DCT 37.38 to 30.65, which compares with a 128-fold decrease. The reduction in gene expression was modest for other luminal marker genes (GATA3 6.8-fold, ERBB2 6.3-fold). In contrast, FGFR1 mRNA expression increased from 33.28 to 35.88 (~6.8-fold increase). Spearman correlation revealed positive association of pretreatment KRT20 mRNA levels with achieving pCR (r = 0.3072: p = 0.0684), whereas pretreatment FGFR1 mRNA was associated with resistance to chemotherapy (r = −0.6418: p < 0.0001). Hierarchical clustering identified luminal tumors of high KRT20 mRNA expression being associated with high pCR rate (10/16; 63%), while the double-negative subgroup with high FGFR1 expression did not respond with pCR (0/9; 0%). Molecular subtyping distinguishes patients with high probability of response from tumors as resistant to neoadjuvant chemotherapy. Targeting FGFR1 in less-differentiated bladder cancer subgroups may sensitize tumors for adopted treatments or subsequent chemotherapy.

Chromosome 5 harbors two independent deletion hotspots at 5q13 and 5q21 that characterize biologically different subsets of aggressive prostate cancer.

Deletion of chromosome 5q is common in prostate cancer and is linked to aggressive disease. Most previous studies focused on 5q21 where CHD1 is located, but deletion of mapping studies has identified a second deletion hotspot at 5q13. To clarify the prevalence and clinical relevance of 5q13 deletions and to determine the relative importance of 5q13 and 5q21 abnormalities, a tissue microarray containing samples from 12 427 prostate cancers was analyzed by fluorescence in situ hybridization. Deletion of 5q13 and 5q21 was found in 13.5% and 10%, respectively, of 7932 successfully analyzed cancers. Deletion was restricted to 5q13 in 49.4% and to 5q21 in 32.0% of cancers with a 5q deletion. Only 18.6% of 5q-deleted cancers had deletions of both loci. Both 5q13 and 5q21 deletions were significantly linked to advanced tumor stage, high Gleason grade, nodal metastasis and early biochemical recurrence (P < .005 each). Cancers with co-deletion of 5q13 and 5q21 had a worse prognosis than cancers with isolated 5q13 or 5q21 deletion (P = .0080). Comparison with TMPRSS2:ERG fusion status revealed that 5q21 deletions were tightly linked to ERG negativity (P < .0001) while 5q13 deletions were unrelated to the ERG status. In summary, 5q13 deletion and 5q21 deletion are common, but independent genomic alterations with different functional effects lead to aggressive prostate cancer.

Loss of the adhesion molecule CEACAM1 is associated with early biochemical recurrence in TMPRSS2:ERG fusion-positive prostate cancers.

Altered expression of the carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1) has been linked to adverse tumor features in various cancer types. To better understand the role of CEACAM1 in prostate cancer, we analyzed a tissue microarray containing tumor spots from 17,747 prostate cancer patients by means of immunohistochemistry. Normal prostate glands showed intense membranous CEACAM1 positivity. Immunostaining was interpretable in 13,625 cancers and was considered high in 28%, low in 43% and absent in 29% of tumors. Low and lost CEACAM1 expression was strongly linked to adverse tumor features including high classical and quantitative Gleason grade, lymph node metastasis, advanced tumor stage, positive surgical margin, a high number of genomic deletions and early biochemical recurrence (p < 0.0001 each). Subset analysis of molecularly defined cancer subsets revealed that these associations were strongest in V-ets avian erythroblastosis virus E26 oncogene homolog (ERG) fusion-positive cancers and that CEACAM1 loss was prognostic even in tumors harboring genomic deletions of the phosphatase and tensin homolog tumor suppressor (p < 0.0001). Multivariate analysis suggested that CEACAM1 analysis can provide independent prognostic information beyond established prognosis parameters at the stage of the initial biopsy when therapy decisions must be taken. In conclusion, loss of CEACAM1 expression predicts poor prognosis in prostate cancer and might provide clinically useful prognostic information particularly in cancers harboring the TMPRSS2:ERG fusion.

Reduced KLK2 expression is a strong and independent predictor of poor prognosis in ERG-negative prostate cancer.

BACKGROUND: Kallikrein-related peptidase 2 (KLK2)-like KLK3 (prostate-specific antigen [PSA])-belongs to the highly conserved serine proteases of the glandular kallikrein protein family (KLK family). Studies suggested that measurement of KLK2 serum levels advanced the predictive accuracy of PSA testing in prostate cancer. METHODS: To clarify the potential utility of KLK2 as a prognostic tissue biomarker, KLK2 expression was analyzed by immunohistochemistry in more than 12 000 prostate cancers. RESULTS: Normal epithelium cells usually showed weak to moderate KLK2 immunostaining, whereas KLK2 was negative in 23%, weak in 38%, moderate in 35%, and strong in 4% of 9576 analyzable cancers. Lost or reduced KLK2 immunostaining was associated with advanced tumor stage, high Gleason score, lymph node metastasis, increased cell proliferation, positive resection margin, and early PSA recurrence (P < .0001). Comparison with previously analyzed molecular alterations revealed a strong association of KLK2 loss and presence of TMPRSS2:ERG fusion (P < .0001), most of all analyzed common deletions (9 of 11; P ≤ .03), and decreased PSA immunostaining (P < .0001 each). Cancers with combined negative or weak immunostaining of KLK2 and PSA showed worse prognosis than cancers with at least moderate staining of one or both proteins (P < .0001). Multivariate analyses including established preoperative and postoperative prognostic parameters showed a strong independent prognostic impact of KLK2 loss alone or in combination of PSA, especially in erythroblast transformation-specific-negative cancers (P ≤ .006). CONCLUSIONS: Loss of KLK2 expression is a potentially useful prognostic marker in prostate cancer. Analysis of KLK2 alone or in combination with PSA may be useful for estimating cancer aggressiveness at the time of biopsy.

A pre-specified model based on four kallikrein markers in blood improves predictions of adverse pathology and biochemical recurrence after radical prostatectomy.

BACKGROUND: A pre-specified model based on four kallikrein markers in blood, commercially available as 4Kscore, predicts Gleason Grade (GG) 3 + 4 or higher prostate cancer on biopsy. However, sampling error and variation in pathology reporting may miss aggressive disease. METHODS: The 4Kscore was measured in cryopreserved blood from 2330 men obtained before prostatectomy at a single institution between 2002 and 2010. Adverse surgical pathology and biochemical recurrence (BCR) were pre-specified to be assessed in all men, biopsy GG 3 + 3, and 3 + 4. RESULTS: Adjusted for established clinical predictors, the 4Kscore was significantly associated with adverse pathology (OR 1.49; 95% CI 1.32, 1.67; p < 0.0001). Adding 4Kscore increased discrimination from (AUC) 0.672 to 0.718 and 0.644 to 0.659 within biopsy GG 3 + 3 and 3 + 4, respectively. Higher 4Kscore was associated with higher risk of BCR (HR 1.16, 95% CI 1.06, 1.26; p = 0.001). Adding 4Kscore improved the prediction of BCR (C-index 0.630-0.660) within GG 3 + 3, but not GG 3 + 4. CONCLUSIONS: The 4Kscore can help guide the clinical decision whether additional risk assessment-such as confirmatory biopsy-is needed to decide between active surveillance versus curative therapy. Evidence that the panel could influence management in biopsy GG 3 + 4 is less strong and requires further investigation.

EGFR as a stable marker of prostate cancer dissemination to bones.

BACKGROUND: Prostate cancer (PCa) is among the most commonly diagnosed malignancies in men. Although 5-year survival in patients with localised disease reaches nearly 100%, metastatic disease still remains incurable. Therefore, there is a need for markers indicating metastatic dissemination. METHODS: EGFR overexpression (EGFRover) was tracked in 1039 primary tumours, circulating tumour cells from 39 d'Amico high-risk patients and metastatic samples from 21 castration-resistant PCa cases. EGFR status was compared to clinical parameters and multiple molecular factors were assessed using immunohistochemistry and gene ontology analysis. The functional aspect of EGFR was evaluated by plating PC-3 cells on soft and rigid matrices. RESULTS: EGFRover was found in 14% of primary tumours, where it was associated with shorter metastasis-free survival and was an independent indicator of worse overall survival. EGFRover correlated with a pro-migratory and pro-metastatic phenotype of tumour cells as well as rich collagen fibre content. All circulating tumour cells (detected in 13% of cases) were positive for EGFR, independent of their EMT-related phenotype. EGFRover was more prevalent in castration-resistant bone metastases (29% of patients) and supported growth of human PCa cells on rigid matrices mimicking bone stiffness. CONCLUSIONS: EGFRover is a stable, EMT-independent marker of PCa disseminating to rigid organs, preferentially bones.

Solid organ transplantation programs facing lack of empiric evidence in the COVID-19 pandemic: A By-proxy Society Recommendation Consensus approach.

The ongoing severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic has a drastic impact on national health care systems. Given the overwhelming demand on facility capacity, the impact on all health care sectors has to be addressed. Solid organ transplantation represents a field with a high demand on staff, intensive care units, and follow-up facilities. The great therapeutic value of organ transplantation has to be weighed against mandatory constraints of health care capacities. In addition, the management of immunosuppressed recipients has to be reassessed during the ongoing coronavirus disease 2019 (COVID-19) pandemic. In addressing these crucial questions, transplant physicians are facing a total lack of scientific evidence. Therefore, the aim of this study was to offer an approach of consensus-based guidance, derived from individual information of 22 transplant societies. Key recommendations were extracted and the degree of consensus among different organizations was calculated. A high degree of consensus was found for temporarily suspending nonurgent transplant procedures and living donation programs. Systematic polymerase chain reaction-based testing of donors and recipients was broadly recommended. Additionally, more specific aspects (eg, screening of surgical explant teams and restricted use of marginal donor organs) were included in our analysis. This study offers a novel approach to informed guidance for health care management when a priori no scientific evidence is available.

Upregulation of the transcription factor TFAP2D is associated with aggressive tumor phenotype in prostate cancer lacking the TMPRSS2:ERG fusion.

BACKGROUND: TFAP2D is a transcription factor important for modulating gene expression in embryogenesis. Its expression and prognostic role in prostate cancer has not been evaluated. METHODS: Therefore, a tissue microarray containing 17,747 prostate cancer specimens with associated pathological, clinical, and molecular data was analyzed by immunohistochemistry to assess the role of TFAP2D. RESULTS: TFAP2D expression was typically increased in prostate cancer as compared to adjacent non-neoplastic glands. TFAP2D staining was considered negative in 24.3% and positive in 75.7% of 13,545 interpretable cancers. TFAP2D staining was significantly linked to advanced tumor stage, high classical and quantitative Gleason grade, lymph node metastasis, and a positive surgical margin (p ≤ 0.0045). TFAP2D positivity was more common in ERG fusion positive (88.7%) than in ERG negative cancers (66.8%; p < 0.0001). Subset analyses in 3776 cancers with and 4722 cancers without TMPRSS2:ERG fusion revealed that associations with tumor phenotype and patient outcome were largely driven by the subset of ERG negative tumors. Multivariate analysis did not identify TFAP2D protein expression levels as a robust independent prognostic parameter. Positive TFAP2D immunostaining was significantly associated with 10 of 11 previously analyzed chromosomal deletions in ERG negative cancers (p ≤ 0.0244 each) indicating that elevated TFAP2D expression parallels genomic instability in prostate cancer. CONCLUSION: These data demonstrate that TFAP2D protein overexpression is linked to prostate cancer progression and genomic instability in ERG negative prostate cancers.

Tumor-Associated Release of Prostatic Cells into the Blood after Transrectal Ultrasound-Guided Biopsy in Patients with Histologically Confirmed Prostate Cancer.

BACKGROUND: Transrectal ultrasound-guided prostate biopsy (TRUS) is a standard procedure for prostate cancer diagnosis. Because prostate cancer is a multifocal disease in many patients, multiple sampling (n ≥ 10) is required, which may bear the risk of systemic spread of cancer cells. DESIGN: Using the standardized CellSearch® system that allows for the detection of single epithelial cell adhesion molecule-positive circulating tumor cells (CTCs) in blood, we investigated whether prostate biopsy is associated with release of prostatic tumor cells into the circulation. Peripheral blood was obtained before and within 30 min after performing prostate biopsy from 115 men with increased serum prostate-specific antigen. RESULTS: The number of CTCs significantly increased after biopsy in men with histologically confirmed prostate cancer (odds ratio, 7.8; 95% CI, 4.8-12.8), whereas no biopsy-related changes could be detected in men without confirmed prostate cancer. Multivariable analysis showed that biopsy-related increase of CTCs was significantly correlated with a worse progression-free survival (hazard ratio, 12.4; 95% CI, 3.2-48.6) within the median follow-up of 41 months. CONCLUSIONS: Prostate biopsies may lead to a tumor-associated release of CTCs into the blood circulation. Larger confirmatory trials with longer follow-up periods are required before any change in clinical practice can be recommended.

Epithelial splicing regulatory protein 1 and 2 (ESRP1 and ESRP2) upregulation predicts poor prognosis in prostate cancer.

BACKGROUND: Epithelial splicing regulatory protein 1 (ESRP1) and 2 (ESRP2) regulate alternative splicing events of various pre-mRNAs. Some of these targets play a role in cancer-associated processes, including cytoskeleton reorganization and DNA-repair processes. This study was undertaken to estimate the impact of ESRP1 and ESRP2 alterations on prostate cancer patient prognosis. METHODS: A tissue microarray made from 17,747 individual cancer samples with comprehensive, pathological, clinical and molecular data was analyzed by immunohistochemistry for ESRP1 and ESRP2. RESULTS: Nuclear staining for ESRP1 was seen in 38.6% (36.0% low, 2.6% high) of 12,140 interpretable cancers and in 41.9% (36.4% low, 5.3% high) of 12,962 interpretable cancers for ESRP2. Nuclear protein expression was linked to advanced tumor stage, high Gleason score, presence of lymph node metastasis, early biochemical recurrence, and ERG-positive cancers (p < 0.0001 each). Expression of ESRPs was significantly linked to 11 (ESRP1)/9 (ESRP2) of 11 analyzed deletions in all cancers and to 8 (ESRP1)/9 (ESRP2) of 11 deletions in ERG-negative cancers portending a link to genomic instability. Combined ESRPs expression analysis suggested an additive effect and showed the worst prognosis for cancers with high ESRP1 and ESRP2 expression. Multivariate analyses revealed that the prognostic impact of ESRP1, ESRP2 and combined ESRP1/ESRP2 expression was independent of all established pre- and postoperative prognostic features. CONCLUSIONS: Our data show a striking link between nuclear ESRP expression and adverse features in prostate cancer and identifies expression of ESRP1 and/or ESRP2 as independent prognostic markers with a potential for routine application.

Claudin-1 upregulation is associated with favorable tumor features and a reduced risk for biochemical recurrence in ERG-positive prostate cancer.

BACKGROUND: Claudin-1 is a membrane-tight junction protein and important for the sealing of the paracellular cleft in epithelial and endothelial cells. Differential expression of Claudin-1 is linked to disease outcome in various cancers. MATERIAL AND METHODS: To evaluate the potential relevance of Claudin-1 expression in prostate cancer, a tissue microarray containing samples of 17,747 tumors with annotated clinico-pathological and molecular data was immunohistochemically analyzed for Claudin-1 expression. RESULTS: In normal prostate, glandular cells were always Claudin-1-negative while there was a strong staining of gland-surrounding basal cells. In contrast to normal prostatic glands, a positive Claudin-1 immunostaining, was found, however, in 38.7% of 12,441 interpretable cancers and was considered weak in 12.7%, moderate in 13.2%, and strong in 12.8% of cases. Positive Claudin-1 immunostaining was associated with favorable tumor features like low pT (p = 0.0032), low Gleason grade (p< 0.0001), and a reduced risk of PSA recurrence (p = 0.0005). A positive Claudin-1 staining was markedly more frequent in ERG-positive (63%) than in ERG-negative cancers (23%; p < 0.0001). Subset analyses revealed that all associations of Claudin-1 expression and favorable phenotype and prognosis were driven by ERG-positive cancers. Multivariate analyses revealed, however, that even in ERG-positive cancers, the prognostic impact of high Claudin-1 expression was not independent of established clinico-pathological parameters. Comparison with 12 previously analyzed chromosomal deletions identified conspicuous associations with PTEN and 12p13 deletions potentially indicating functional interactions. CONCLUSION: These data identify a peculiar role for Claudin-1 in prostate cancer. The protein is overexpressed in a fraction of prostate cancers and increased Claudin-1 expression levels predict a favorable prognosis in ERG-positive cancer.