Monitoring Of Bladder Cancer Research Articles

From Detection to Cure - Emerging Roles for Urinary Tumor DNA (utDNA) in Bladder Cancer.

This review sought to define the emerging roles of urinary tumor DNA (utDNA) for diagnosis, monitoring, and treatment of bladder cancer. Building from early landmark studies the focus is on recent studies, highlighting how utDNA could aid personalized care. Recent research underscores the potential for utDNA to be the premiere biomarker in bladder cancer due to the constant interface between urine and tumor. Many studies find utDNA to be more informative than other biomarkers in bladder cancer, especially in early stages of disease. Points of emphasis include superior sensitivity over traditional urine cytology, broad genomic and epigenetic insights, and the potential for non-invasive, real-time analysis of tumor biology. utDNA shows promise for improving all phases of bladder cancer care, paving the way for personalized treatment strategies. Building from current research, future comprehensive clinical trials will validate utDNA's clinical utility, potentially revolutionizing bladder cancer management.

Comprehensive Urinary Proteome Profiling Analysis Identifies Diagnosis and Relapse Surveillance Biomarkers for Bladder Cancer.

Bladder cancer (BCa) is the predominant malignancy of the urinary system. Herein, a comprehensive urine proteomic feature was initially established for the noninvasive diagnosis and recurrence monitoring of bladder cancer. 279 cases (63 primary BCa, 87 nontumor controls (NT), 73 relapsed BCa (BCR), and 56 nonrelapsed BCa (BCNR)) were collected to screen urinary protein biomarkers. 4761 and 3668 proteins were qualified and quantified by DDA and sequential window acquisition of all theoretical mass spectra (SWATH-MS) analysis in two discovery sets, respectively. Upregulated proteins were validated by multiple reaction monitoring (MRM) in two independent combined sets. Using the multi-support vector machine-recursive feature elimination (mSVM-RFE) algorithm, a model comprising 13 proteins exhibited good performance between BCa and NT with an AUC of 0.821 (95% CI: 0.675-0.967), 90.9% sensitivity (95% CI: 72.7-100%), and 73.3% specificity (95% CI: 53.3-93.3%) in the diagnosis test set. Meanwhile, an 11-marker classifier significantly distinguished BCR from BCNR with 75.0% sensitivity (95% CI: 50.0-100%), 81.8% specificity (95% CI: 54.5-100%), and an AUC of 0.784 (95% CI: 0.609-0.959) in the test cohort for relapse surveillance. Notably, six proteins (SPR, AK1, CD2AP, ADGRF1, GMPS, and C8A) of 24 markers were newly reported. This paper reveals novel urinary protein biomarkers for BCa and offers new theoretical insights into the pathogenesis of bladder cancer (data identifier PXD044896).

The long noncoding RNA SUMO1P3 as urinary biomarker for monitoring bladder cancer progression

IntroductionUrothelial Bladder Cancer (BC) is the ninth most common cancer worldwide. It is classified into Non Muscle Invasive (NMIBC) and Muscle Invasive Bladder Cancer (MIBC), which are characterized by frequent recurrences and progression rate, respectively. The diagnosis and monitoring are obtained through invasive methods as cystoscopy and post-surgery biopsies. Thus, a panel of biomarkers able to discriminate BC based on grading or staging represents a significant step forward in the patients’ workup. In this perspective, long non-coding RNAs (lncRNAs) are emerged as reliable candidates as potential biomarker given their specific and regulated expression. In the present work we propose two lncRNAs, the Small Ubiquitin Modifier 1 pseudogene 3 (SUMO1P3), a poorly characterized pseudogene, and the Urothelial Carcinoma Associated 1 (UCA1) as candidates to monitor the BC progression.MethodsThis study was a retrospective trial enrolling NMIBC and MIBC patients undergoing surgical intervention: the expression of the lncRNA SUMO1P3 and UCA1 was evaluated in urine from 113 subjects (cases and controls). The receiver operating characteristic curve analysis was used to evaluate the performance of single or combined biomarkers in discriminating cases from controls.ResultsSUMO1P3 and UCA1 expression in urine was able to significantly discriminate low grade NMIBC, healthy control and benign prostatic hyperplasia subjects versus high grade NMIBC and MIBC patients. We also demonstrated that miR-320a, which binds SUMO1P3, was reduced in high grade NMIBC and MIBC patients and the SUMO1P3/miR-320a ratio was used to differentiate cases versus controls, showing a statistically significant power. Finally, we provided an automated method of RNA extraction coupled to ddPCR analysis in a perspective of clinical application.DiscussionWe have shown that the lncRNA SUMO1P3 is increased in urine from patients with high grade NMIBC and MIBC and that it is likely to be good candidate to predict bladder cancer progression if used alone or in combination with UCA1 or with miRNA320a.

Abstract A005: Exploring liquid biopsy for the detection of bladder cancer: Unraveling RNA fusions

Abstract Introduction: This study delves into the promising realm of liquid biopsy for the non-invasive detection of bladder cancer. Leveraging urine as a valuable source, our investigation focuses on identifying RNA fusions associated with bladder cancer, with specific attention to ETV6-NTRK3 and PRCC-TFE3. The utilization of liquid biopsy offers a less invasive and more accessible approach to diagnosing bladder cancer, potentially revolutionizing the field of cancer diagnostics. These specific RNA fusions in urine samples may serve as reliable biomarkers for early detection and monitoring of bladder cancer. This research contributes to the ongoing efforts to enhance diagnostic methodologies to improve patient outcomes and streamline cancer management. Materials and Methods: Urine was collected from healthy male donors using UAS, a urine preservative from Novosanis, for improved urine preservation and stability. The urine samples with UAS were centrifuged at 16,000 xg for 10 min to obtain cf-urine. Three 20 mL aliquots of the precleared supernatant urine samples were spiked with an RNA fusion reference standard from Anchor Molecular containing ETV6-NTRK3 and PRCC-TFE3RNA at a concentration of 20ng/mL. All samples were extracted using the nRichDX Revolution Max20 cfTNA Isolation Kit. The eluants from the extracted samples were DNAse treated. The extracted cfTNAs profile was assessed on 4150 TapeStation System, Agilent, using RNA ScreenTape. Extracted nucleic acids from urine were evaluated using a RT-qPCR assay to quantify the number of copies of the fusion RNAs. Results: The Renal RNA fusions ETV6-NTRK3 and PRCC-TFE3RNA were successfully extracted using the nRichDX Revolution cfTNA Isolation Kit. TapeStation analysis demonstrated prominent 18S and 28S peaks, confirming the successful extraction of RNA from 20 mL urine samples. The mean percent recovery of the recovered RNA Fusion mutation was about 70% via RT-qPCR. Conclusion: Our findings indicate a recovery rate of approximately 70% for spiked RNA fusions in the urine samples. This research holds the promise of revolutionizing cancer diagnostics by introducing a method that is not only reliable but also convenient for early detection and monitoring. Identifying specific RNA fusions in urine samples as biomarkers is a significant step in enhancing diagnostic methodologies. As we contribute to these ongoing efforts, we aim to improve patient outcomes and streamline the management of bladder cancer through innovative and effective diagnostic approaches. Citation Format: Mayer Saidian, Jason Saenz, Carlos Hernandez, Lauren Lee, Andrew Dunnigan, Yabin Lu, Nafiseh Jafari. Exploring liquid biopsy for the detection of bladder cancer: Unraveling RNA fusions [abstract]. In: Proceedings of the AACR Special Conference on Bladder Cancer: Transforming the Field; 2024 May 17-20; Charlotte, NC. Philadelphia (PA): AACR; Clin Cancer Res 2024;30(10_Suppl):Abstract nr A005.

Abstract LB327: DNA methylation biomarkers for early bladder cancer detection and treatment response monitoring

Abstract Objectives: Bladder cancer (BC) stands as the 5th most prevalent cancer in the USA, with over 83,000 new cases diagnosed in 2023. The absence of non-invasive diagnostic tools for sensitive early BC detection and treatment response monitoring poses a significant challenge. This study evaluates the clinical potential of the Bladder CARE™ Assay for early BC detection and monitoring treatment response in BC patients. Methods: Under an institutional review board-approved protocol, voided urine samples were prospectively collected from USC patients with prior BC history under surveillance/anticancer treatment, prior genitourinary manipulation. Enrollment spanned February 2019 to September 2021. Samples underwent analysis using the Bladder CARE™ Assay, a DNA methylation test for quantitative BC and upper tract urothelial carcinoma (UTUC) detection from urine. Results were reported as Bladder CARE Index (BCI) and samples were categorized as “positive” (BCI > 5), “low-positive” (2.5 < BCI < 5), or “negative” (BCI < 2.5). Correlations between BCI value and categories, and clinicopathological findings were assessed. Results: A total of 110 previously diagnosed BC patients (median age: 74; 86% male) were enrolled in this study. Within 36 months post-TURBT, 24 patients (21.8%) showed evidence of recurrence. Bladder CARE™ Assay detected all recurrences, averagely 7.35 months earlier than cystoscopy. Of 55 patients (50%) undergoing anticancer therapies (45 BCG, 4 MMC, 2 GEM, 1 9UT, 1 GEM/DOCE, 1 MMC/BCG, and 1 BCG, MMC, and GEM), 7 (12.7%) recur (non-responders) and 11 (20%) did not show evidence of recurrence by 18 months post-TURBT (responders); 18-month post-operative data was unavailable for 37 patients (67.3%), which could not be classified in responders or non-responders. Bladder CARE™ detected 85.7% of non-responders, with BCI increasing prior to positive histology (avg. BCI: 86.1). In responders, BCI remained stable in negative/low-positive range post-TURBT (avg. BCI: 2.5). In addition, data from pre- and post-TURBT assessments were available for 20 of the 110 enrolled patients. The Bladder CARE™ Assay demonstrated a decrease in BCI post-TURBT in 19 of these 20 patients (95%). Conclusions: This prospective pilot study underscores the Bladder CARE™ Assay's capacity to pre-emptively detect BC months ahead of the gold standard. Furthermore, its quantitative nature offers prognostic insight, enabling non-invasive monitoring of patient response to anticancer treatments. A larger-scale study is the next step to validating these promising findings. Citation Format: Paolo Piatti, Sia Daneshmand, Sanam Ladi Seyedian, Saum Ghodossipour, Hamed Ahmadi, Suzanne Roberts, Alireza Ghoreifi, Michael Basin, Simin Hajian, Yap Ching Chew, Jeffrey Bhasin, Benjamin Jara, Lucy Sanossian, Hooman Djaladat, Anne Shuckman, Sumeet Bhavandia, Gangning Liang. DNA methylation biomarkers for early bladder cancer detection and treatment response monitoring [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr LB327.

Plasma-Derived Cell-Free DNA as a Biomarker for Early Detection, Prognostication, and Personalized Treatment of Urothelial Carcinoma.

Bladder cancer (BC) is one of the most common malignancies in the United States, with over 80,000 new cases and 16,000 deaths each year. Urothelial carcinoma (UC) is the most common histology and accounts for 90% of cases. BC management is complicated by recurrence rates of over 50% in both muscle-invasive and non-muscle-invasive bladder cancer. As such, the American Urological Association (AUA) recommends that patients undergo close surveillance during and after treatment. This surveillance is in the form of cystoscopy or imaging tests, which can be invasive and costly tests. Considering this, there have been recent pushes to find complements to bladder cancer surveillance. Cell-free DNA (CfDNA), or DNA released from dying cells, and circulating tumor DNA (ctDNA), or mutated DNA released from tumor cells, can be analyzed to detect and characterize the molecular characteristics of tumors. Research has shown promising results for ctDNA use in the BC care realm. A PubMed literature review was performed finding studies discussing cfDNA and ctDNA in BC detection, prognostication, and monitoring for recurrence. Keywords used included bladder cancer, cell-free DNA, circulating tumor DNA, urothelial carcinoma, and liquid biopsy. Studies show that ctDNA can serve as prognostic indicators of both early- and late-stage BC, aid in risk stratification prior to major surgery, assist in detection of disease progression and metastatic relapse, and can assess patients who may respond to immunotherapy. The benefit of ctDNA is not confined to BC, as studies have also suggested its promise as a biomarker for neoadjuvant chemotherapy in upper-tract UC. However, there are some limitations to ctDNA that require improvements in ctDNA-specific detection methods and BC-specific mutations before widespread utilization can be achieved. Further prospective, randomized trials are needed to elucidate the true potential ctDNA has in advancements in BC care.

Results Obtained from a Pivotal Validation Trial of a Microsatellite Analysis (MSA) Assay for Bladder Cancer Detection through a Statistical Approach Using a Four-Stage Pipeline of Modern Machine Learning Techniques.

Several studies have shown that microsatellite changes can be profiled in urine for the detection of bladder cancer. The use of microsatellite analysis (MSA) for bladder cancer detection requires a comprehensive analysis of as many as 15 to 20 markers, based on the amplification and interpretations of many individual MSA markers, and it can be technically challenging. Here, to develop fast, more efficient, standardized, and less costly MSA for the detection of bladder cancer, we developed three multiplex-polymerase-chain-reaction-(PCR)-based MSA assays, all of which were analyzed via a genetic analyzer. First, we selected 16 MSA markers based on 9 selected publications. Based on samples from Johns Hopkins University (the JHU sample, the first set sample), we developed an MSA based on triplet, three-tube-based multiplex PCR (a Triplet MSA assay). The discovery, validation, and translation of biomarkers for the early detection of cancer are the primary focuses of the Early Detection Research Network (EDRN), an initiative of the National Cancer Institute (NCI). A prospective study sponsored by the EDRN was undertaken to determine the efficacy of a novel set of MSA markers for the early detection of bladder cancer. This work and data analysis were performed through a collaboration between academics and industry partners. In the current study, we undertook a re-analysis of the primary data from the Compass study to enhance the predictive power of the dataset in bladder cancer diagnosis. Using a four-stage pipeline of modern machine learning techniques, including outlier removal with a nonlinear model, correcting for majority/minority class imbalance, feature engineering, and the use of a model-derived variable importance measure to select predictors, we were able to increase the utility of the original dataset to predict the occurrence of bladder cancer. The results of this analysis showed an increase in accuracy (85%), sensitivity (82%), and specificity (83%) compared to the original analysis. The re-analysis of the EDRN study results using machine learning statistical analysis proved to achieve an appropriate level of accuracy, sensitivity, and specificity to support the use of the MSA for bladder cancer detection and monitoring. This assay can be a significant addition to the tools urologists use to both detect primary bladder cancers and monitor recurrent bladder cancer.

The Role of Checkpoint Inhibitor Expression Directly on Exfoliated Cells from Bladder Cancer: A Narrative Review.

Bladder cancer (BCa) is a common type of cancer that affects the urinary bladder. The early detection and management of BCa is critical for successful treatment and patient outcomes. In recent years, researchers have been exploring the use of biomarkers as a non-invasive and effective tool for the detection and monitoring of BCa. One such biomarker is programmed death-ligand 1 (PD-L1), which is expressed on the surface of cancer cells and plays a crucial role in the evasion of the immune system. Studies have shown that the PD-L1 expression is higher in BCa tumors than in healthy bladder tissue. Additionally, PD-L1 expression might even be detected in urine samples in BCa patients, in addition to the examination of a histological sample. The technique is being standardized and optimized. We reported how BCa patients had higher urinary PD-L1 levels than controls by considering BCa tumors expressing PD-L1 in the tissue specimen. The expression of PD-L1 in urinary BCa cells might represent both a diagnostic and a prognostic tool, with the perspective that the PD-L1 expression of exfoliate urinary cells might reveal and anticipate eventual BCa recurrence or progression. Further prospective and longitudinal studies are needed to assess the expression of PD-L1 as a biomarker for the monitoring of BCa patients. The use of PD-L1 as a biomarker for the detection and monitoring of BCa has the potential to significantly improve patient outcomes by allowing for earlier detection and more effective management of the disease.

Xpert bladder cancer monitor to predict the need for a second TURB (MoniTURB trial)

To determine whether Xpert bladder cancer monitor, a noninvasive PCR-based biomarker test, can predict the need for 2nd transurethral resection of the bladder (TURB) better than clinical assessment. Patients scheduled for TURB were prospectively screened. After initial TURB, patients were assigned to 2nd TURB or follow-up cystoscopy at 3 months (FU) by clinicians’ discretion. Central urine cytology and Xpert monitor tests were performed prior to the 1st TURB and 2nd TURB or FU, respectively. Statistical analysis to compare clinical assessment and Xpert monitor comprised sensitivity (SENS), specificity (SPEC), NPV and PPV. Of 756 screened patients, 171 were included (114 with 2nd TURB, 57 with FU). Residual tumors were detected in 34 patients who underwent 2nd TURB, and recurrent tumors were detected in 2 patients with FU. SENS and SPEC of Xpert monitor were 83.3% and 53.0%, respectively, PPV was 32.6% and NPV was 92.1%. Clinical risk assessment outperformed Xpert monitor. In patients with pTa disease at initial TURB, Xpert monitor revealed a NPV of 96%. Xpert monitor was not superior than clinical assessment in predicting the need for 2nd TURB. It might be an option to omit 2nd TURB for selected patients with pTa disease.

Field Cancerization Is Associated with Tumor Development, T-cell Exhaustion, and Clinical Outcomes in Bladder Cancer

BackgroundField cancerization is characterized by areas of normal tissue affected by mutated clones. Bladder field cancerization may explain the development and recurrence of bladder cancer and may be associated with treatment outcomes. ObjectiveTo investigate the predictive and prognostic roles of field cancerization in patients with high-risk non–muscle-invasive bladder cancer (NMIBC) treated with bacillus Calmette-Guérin (BCG). Design, setting, and participantsWe conducted comprehensive genomic and proteomic analyses for 751 bladder biopsies and 234 urine samples from 136 patients with NMIBC. The samples were collected at multiple time points during the disease course. Field cancerization in normal-appearing bladder biopsies was measured using deep-targeted sequencing and error correction models. Outcome measurements and statistical analysisEndpoints included the rates of recurrence and progression. Cox regression and Wilcoxon rank-sum and Fisher’s exact tests were used. Results and limitationsA high level of field cancerization was associated with high tumor mutational burden (p = 0.007), high tumor neoantigen load (p = 0.029), and high tumor-associated CD8 T-cell exhaustion (p = 0.017). In addition, high field cancerization was associated with worse short-term outcomes (p = 0.029). Nonsynonymous mutations in bladder cancer–associated genes such as KDM6A, ARID1A, and TP53 were identified as early disease drivers already found in normal-appearing bladder biopsies. Urinary tumor DNA (utDNA) levels reflected the bladder tumor burden and originated from tumors and field cancerization. High levels of utDNA after BCG were associated with worse clinical outcomes (p = 0.027) and with disease progression (p = 0.003). High field cancerization resulted in high urinary levels of proteins associated with angiogenesis and proliferation. Limitations include variation in the number of biopsies and time points analyzed. ConclusionsField cancerization levels are associated with tumor development, immune responses, and clinical outcomes. utDNA measurements can be used to monitor disease status and treatment response. Patient summaryMolecular changes in the tissue lining the bladder result in tumor recurrence. Urinary measurements may be used to monitor bladder cancer status and treatment responses.

Homeobox Gene Expression Dysregulation as Potential Diagnostic and Prognostic Biomarkers in Bladder Cancer.

Homeobox genes serve as master regulatory transcription factors that regulate gene expression during embryogenesis. A homeobox gene may have either tumor-promoting or tumor-suppressive properties depending on the specific organ or cell lineage where it is expressed. The dysregulation of homeobox genes has been reported in various human cancers, including bladder cancer. The dysregulated expression of homeobox genes has been associated with bladder cancer clinical outcomes. Although bladder cancer has high risk of tumor recurrence and progression, it is highly challenging for clinicians to accurately predict the risk of tumor recurrence and progression at the initial point of diagnosis. Cystoscopy is the routine surveillance method used to detect tumor recurrence. However, the procedure causes significant discomfort and pain that results in poor surveillance follow-up amongst patients. Therefore, the development of reliable non-invasive biomarkers for the early detection and monitoring of bladder cancer is crucial. This review provides a comprehensive overview of the diagnostic and prognostic potential of homeobox gene expression dysregulation in bladder cancer.

Ultrasensitive Electrochemiluminescence Immunosensor for Bladder Marker Human Complement Factor H-Related Protein Detection.

The development of noninvasive and sensitive detection methods for the early diagnosis and monitoring of bladder cancer is critical but challenging. Herein, an ultrasensitive electrochemiluminescence (ECL) immunosensor that uses Ru(bpy)32+-metal-organic framework (Ru-MOF) nanospheres and a DNA tetrahedral (TDN) probe was established for bladder cancer marker complement factor H-related protein (CFHR1) detection. The synthesized Ru(bpy)32+-metal-organic frameworks (Ru-MOFs) served as a linked substrate for immobilization of AuNPs and antibody (Ab2) to prepare the ECL signal probe (Ru-MOF@AuNPs-Ab2), exhibiting a stable and strengthened ECL emission. At the same time, the inherent advantages of TDN probes on the electrode as the capture probe (TDN-Ab1) improve the accessibility of targets to probes. In the presence of CFHR1, the signal probe Ru-MOF@AuNPs-Ab2 was modified on the electrode through immune binding, thereby obtaining an outstanding ECL signal. As expected, the developed ECL immunosensor exhibited splendid performance for CFHR1 detection in the range of 0.1 fg/mL to 10 pg/mL with a quite low detection limit of 0.069 fg/mL. By using the proposed strategy to detect CFHR1 from urine, it showed acceptable accuracy, which can effectively distinguish between bladder cancer patients and healthy samples. This work contributes to a novel, noninvasive, and accurate method for early clinical diagnosis of bladder cancer.

Xpert Bladder Cancer Monitor for the Early Detection of Non-Muscle Invasive Bladder Cancer Recurrences: Could Cystoscopy Be Substituted?

XBM was prospectively assessed in spontaneous urine collected just before flexible cystoscopy and washing cytology carried out within the first 2 years follow-up of 337 patients with NMIBC. Recurrences were pathologically confirmed in 49 patients (14.5%), 22 of them being high-risk (6.5%). The XBM sensitivity for detecting any type of recurrence was 69.4% and 63.6% in the cases of high-risk NMIBC. Negative predictive value (NPV) for XBM was 93% for all recurrences and 96.2% for high-risk recurrences. XBM could have avoided 213 invasive controls but missed the detection of 15 recurrences (30.6%)-8 of them of high-risk (36.4%). XBM false positive elevations were detected in 90 patients (26.7%), whereas 10 patients with the invasive method had a false positive result (3%), p <0.001. However, early detection of recurrences during the first year's follow-up after an XBM false positive result was observed in 18 patients (20%). On the other hand, 19 recurrences were detected during this period among the rest of the patients (7.7%)-p = 0.003, and odds ratio (OR) 3.0 (95% CI 1.5-6.0). Regarding one-year follow-up recurrences, 10% were high-risk recurrences in the XBM false positive group and 3.2% in the rest of the patients-p = 0.021, and OR 3.3 (95% CI 1.2-8.9). Additionally, 11.3% of the patients without false positive results developed a recurrence, p = 0.897, for any recurrence, being 10% and 5.2%, respectively, and high-risk and low-risk recurrences, p = 0.506. After searching for the best XBM cutoff for detecting the 38 high-risk initial recurrences and the early high-risk recurrences after a one-year follow-up, a linear discriminant analysis (LDA) of 0.13 could have avoided 11.3% of cystoscopies and bladder wash cytologies, as this cutoff missed only 1 high-risk recurrence (2.6%). More extensive and well-designed studies will confirm if XBM can improve the surveillance of NMIBC.

Off the fog to find the optimal choice: Research advances in biomarkers for early diagnosis and recurrence monitoring of bladder cancer.

Bladder cancer (BC) has high morbidity and mortality rates owing to challenges in clinical diagnosis and treatment. Advanced BC is prone to recurrence after surgery, necessitating early diagnosis and recurrence monitoring to improve the prognosis of patients. Traditional detection methods for BC include cystoscopy, cytology, and imaging; however, these methods have drawbacks such as invasiveness, lack of sensitivity, and high costs. Existing reviews on BC focus on treatment and management and lack a comprehensive assessment of biomarkers. Our article reviews various biomarkers for the early diagnosis and recurrence monitoring of BC and outlines the existing challenges associated with their application and possible solutions. Furthermore, this study highlights the potential application of urine biomarkers as a non-invasive, inexpensive adjunctive test for screening high-risk populations or evaluating patients with suspected BC symptoms, thereby alleviating the discomfort and financial burden associated with cystoscopy and improving patient survival.

Comet-FISH analysis of urothelial cells. A screening opportunity for bladder cancer?

ABSTRACT Introduction Bladder cancer (BCa) is the most frequent cancer of the urinary tract, with more than 500,000 reported cases and nearly 200,000 related deaths yearly. Cystoscopy is the standard examination used for the initial diagnosis and follow-up of BCa in the noninvasive stage. However, the American Cancer Society does not include BCa screening in its list of recommended cancer screenings. Areas covered Recently, several urine-based bladder tumor markers (UBBTMs) that identify genomic, transcriptomic, epigenetic, or protein alterations have been introduced, some of which have been approved by the Food and Drug Administration (FDA) to improve its diagnosis and surveillance. Several biomarkers have been found in the tissues and blood of individuals with BCa or predisposed to develop the disease, further enriching our information. Expert opinion From a prevention perspective, alkaline Comet-FISH could be a valuable tool with broad potential for clinical application. Furthermore, a comet assay could be more beneficial for diagnosing and monitoring bladder cancer and determining individual susceptibility. Thus, we recommend further studies to understand the potential of this combined assay in the general population as a potential screening test and in patients initiated into the diagnostic process.

Abstract 3326: The clinical validity of urinary pellet DNA monitoring of recurrent bladder cancer

Abstract Introduction: Approximately half of high-risk non-muscle-invasive bladder cancers (NMIBCs) recur after transurethral resection of the bladder tumor (TURBT), whereas conventional urological testing for bladder cancer (BC) patients has been considered invasive and with low sensitivity. Our previous reports have shown that circulating tumor DNA (ctDNA) monitored by digital PCR (dPCR) can predict relapse and evaluate treatment efficacy in digestive tract cancers. In BC patients, tumor-derived genetic mutations detected in urinary DNA are expected to be a diagnostic biomarker. This study evaluates the clinical validity of monitoring tumor-specific gene mutations in ctDNA and urine pellet DNA (upDNA) using dPCR as a biomarker for early relapse prediction and determination of treatment efficacy. Materials and Methods: Thirty-two previously treated and untreated BC patients were enrolled. Tumor DNA was extracted from archived TURBT tissues, and gene mutation analysis by Next Generation Sequencing (NGS) of tumors was performed to select patient-unique somatic mutations for both ctDNA and upDNA monitoring by dPCR. Longitudinal variant allele frequencies (VAFs) of ctDNA and upDNA were monitored by dPCR for up to two years. While dPCR is rapid and has a high sensitivity of less than 0.1% VAF, a mutation-specific primer/probe set is required. Therefore, we established over 1,000 primer/probe sets for frequent mutations in human cancer. This large-scale dPCR primer/probe library allow us to select patient specific mutations for immediate ctDNA monitoring. Clinical recurrence (Crec) based on imaging and urinary cell findings are compared with VAF dynamics of ctDNA and upDNA. Results: The median observation period was 516 days (30-733), and a total of 230 urine samples were collected. The selection of monitoring mutations based on NGS analysis was performed in 93.8% (30/32) cases, with 2.3 (range: 1-6) mutations monitored per case. The primer/probe sets covered 87.5% (28/32) of cases and 87.5% (42/48) of mutations. The VAFs of the upDNA of cases started showing constantly increasing trends 3-12 months earlier than the diagnosis of Crec in 71.4% (5/7) of the cases. The other two Crec cases, which did not show an elevated upVAF, were both pyuria cases. After local treatment, the upDNA VAFs remained high in the Crec cases. Conventional Crec did not seem to be fully reflected by ctDNA alone. Conclusion: Frequent VAF monitoring using upDNA by dPCR enables the prediction of local relapse and the evaluation of treatment efficacy in the management of BC patients. Citation Format: Masakazu Abe, Hayato Hiraki, Takashi Tsuyukubo, Sadahide Ono, Shigekatsu Maekawa, Daichi Tamura, Daiki Ikarashi, Renpei Kato, Tomohiko Matsuura, Mitsugu Kanehira, Ryo Takata, Hiromitsu Fujisawa, Takeshi Iwaya, Masashi Idogawa, Wataru Obara, Satoshi S. Nishizuka. The clinical validity of urinary pellet DNA monitoring of recurrent bladder cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3326.

Abstract 245: Development of a novel urine cell-free DNA preservation method for molecular profiling in genitourinary tumors

Abstract Introduction: Due to the benefits of its non-invasive testing, liquid biopsy technology has been applied therapeutically as an adjuvant to disease detection and monitoring in areas such as prenatal cell-free DNA (cfDNA) screening and cancer. Although the majority of current liquid biopsy techniques focus on DNA from blood, DNA from urine also conveys important genetic information from the human body. Studies have revealed that urine cfDNA can potentially be utilized for bladder cancer detection and monitoring. Urine samples can be quickly and painlessly collected; however, because urine DNA is readily damaged, maintaining high-quality cfDNA in urine is challenging. In this study, we developed a robust and user-friendly urine preservation method to maintain integrity of cfDNA in urine and to facilitate the transportation of urinary samples to clinical laboratories. Methods: Urine samples from 10 donors were collected with Predicine’s proprietary urine preservation method. Two urine samples were spiked with BL21(DE3) E.coli competent cells to simulate the urinary tract infection. All samples were incubated at 4°C, room temperature, and 37°C to mimic extreme storage or transporting environments. cfDNA was extracted after incubation of 0, 4, and 7 days and measured by Qubit and Fragment analyzer. Extracted DNA was also quantified by ddPCR with two markers: EGFR as an endogenous DNA marker and ARv7 (synthetic oligos containing intron-deleted sequence, post-incubation spike-in) as an inner control for the extraction and detection assays. Whole-genome sequencing was conducted to measure the quality of the cfDNA and bacterial contamination. Other commercial urine preserve buffers are used as control. Results: cfDNA derived from urine stored in Predicine’s proprietary urine preservation buffer remained stable till day 7 at different temperatures. There was no significant change in normalized copies of EGFR in the treated urine samples for up to 7 days. cfDNA yield was considerably higher in the novel preservation buffer than that using commercial buffers. Furthermore, the mapping rates in bacteria-contaminated samples were significantly improved. Conclusion: Predicine’s proprietary urine preservation method demonstrated excellent urine cfDNA preserving capacity at various temperatures for at least 7 days, outperforming commercially available preserve buffers in terms of DNA integrity and yield. The innovative technique offers an easy-to-use solution for obtaining high-quality urine cfDNA for clinical use. Citation Format: jie zhang, fang liu, min wang, cuishan deng, wei mo, seng tao, amy wang, shidong jia, zhixin zhao, pan du, binggang xiang, shujun luo. Development of a novel urine cell-free DNA preservation method for molecular profiling in genitourinary tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 245.

MP08-04 USE OF ANTIBIOTICS IN PATIENTS UNDERGOING FOLLOW-UP FOR NON-MUSCLE INVASIVE BLADDER CANCER WITH EITHER FLEXIBLE CYSTOSCOPY OR THE URINARY BIOMARKER TEST XPERT® BLADDER CANCER MONITOR: A SECONDARY OUTCOME FROM A RANDOMISED CLINICAL TRIAL

You have accessJournal of UrologyCME1 Apr 2023MP08-04 USE OF ANTIBIOTICS IN PATIENTS UNDERGOING FOLLOW-UP FOR NON-MUSCLE INVASIVE BLADDER CANCER WITH EITHER FLEXIBLE CYSTOSCOPY OR THE URINARY BIOMARKER TEST XPERT® BLADDER CANCER MONITOR: A SECONDARY OUTCOME FROM A RANDOMISED CLINICAL TRIAL Thomas Dreyer, Simone Brandt, Knud Fabrin, Nessn Azawi, Juan Luis Vásquez, Andreas Ernst, Lars Dyrskjøt, and Jørgen Bjerggaard Jensen Thomas DreyerThomas Dreyer More articles by this author , Simone BrandtSimone Brandt More articles by this author , Knud FabrinKnud Fabrin More articles by this author , Nessn AzawiNessn Azawi More articles by this author , Juan Luis VásquezJuan Luis Vásquez More articles by this author , Andreas ErnstAndreas Ernst More articles by this author , Lars DyrskjøtLars Dyrskjøt More articles by this author , and Jørgen Bjerggaard JensenJørgen Bjerggaard Jensen More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000003223.04AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Patients with non-muscle-invasive bladder cancer (NMIBC) have a high recurrence rate, which leads to frequent flexible cystoscopies (FC). Aside from the burden of undergoing FC, there is a risk of urinary tract infection (UTI) following FC. Follow-up of NMIBC with non-invasive urinary biomarker tests could reduce the number of FCs, and thus potentially the need for antibiotics due to UTI. Furthermore, because UTIs are among the most common infections, any reduction in incidence could reduce development of multiresistant uropathogenic bacteria.In this study, a secondary outcome from the DaBlaCa-15 trial is reported. The objective was to compare the number of patients with collected prescriptions for antibiotics for UTI in patients followed with FC versus patients followed primarily with the urinary biomarker test Xpert® Bladder Cancer Monitor (XBCM) (CE-IVD. In Vitro Diagnostic Medical Device. May not be available in all countries. Not available in the United States.). METHODS: A two-arm randomised non-inferiority trial enrolling 392 patients with previous high-grade NMIBC was conducted. In the intervention arm, patients were followed with XBCM every four months with only one yearly pre-scheduled FC. Patients in the control arm underwent FC every four months for two years. The number of patients who collected prescriptions for UTI during the study period was compared both peri- and post-FC and for visits where a FC was not pre-scheduled. RESULTS: During follow-up, 23.7% of patients in the intervention arm collected a prescription compared to 31.0% in the control arm (RR 0.76; 95% CI 0.55-1.06; p=0.11). When analysing visits without a pre-scheduled FC, 6.3% of patients in the intervention arm collected a prescription in the 14 days after a visit versus 13.4% in the control arm (RR 0.47; 95% CI 0.25-0.90; p=0.02). Similarly, the peri-FC use of antibiotics was 7.9% in the intervention arm, compared to 15.0% in the control arm (RR 0.53; 95% CI 0.29-0.94; p=0.03). A reduction in the number of FCs performed in the intervention arm was observed (39.1% versus 98.3%; p<0.0001). CONCLUSIONS: The findings of the present study support the hypothesis that replacing cystoscopy with XBCM can reduce the use of antibiotics for UTI during follow-up of NMIBC. Thus, the morbidity of follow-up of NMIBC with FC can be reduced, resulting perhaps in less development of multiresistant uropathogenic bacteria and increased quality of life. Source of Funding: Supported by Cepheid Inc. © 2023 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 209Issue Supplement 4April 2023Page: e94 Advertisement Copyright & Permissions© 2023 by American Urological Association Education and Research, Inc.MetricsAuthor Information Thomas Dreyer More articles by this author Simone Brandt More articles by this author Knud Fabrin More articles by this author Nessn Azawi More articles by this author Juan Luis Vásquez More articles by this author Andreas Ernst More articles by this author Lars Dyrskjøt More articles by this author Jørgen Bjerggaard Jensen More articles by this author Expand All Advertisement PDF downloadLoading ...

Recent development of urinary biomarkers for bladder cancer diagnosis and monitoring

AbstractUrine‐based liquid biopsy has emerged as a non‐invasive and effective tool for early screening and diagnosis of bladder cancer. This review provides a comprehensive overview of the current urine‐based biomarkers and methods for the detection and monitoring of bladder cancer. We focus on biomarkers including tumour DNAs, proteins, microbiome, tumour RNAs, long non‐coding RNAs, transfer RNA‐derived fragments, messenger RNAs, microRNAs, circular RNAs, exosomes and extrachromosomal circular DNA.

Noninvasive Detection of Bladder Cancer Markers Based on Gold Nanomushrooms and Sandwich Immunoassays

Bladder cancer is one of the most common malignancies in the urinary system. Cystoscopy is the traditional standard diagnostic method for bladder cancer with subsequent biopsy or surgery. However, this method is uncomfortable for most patients because it requires anesthesia and possibly causes infections. Because of the high recurrence rate of bladder cancer, a rapid, low-cost, high-sensitivity, and noninvasive sensing method is needed. This study employed gold nanomushroom (AuNM) chips for bladder cancer biomarker detection, combining the benefits of sandwich immunoassay and localized surface plasmon resonance (LSPR) sensing. With a metal nanotransfer printing technique, which is cheap and straightforward, the AuNMs were patterned on flexible polycarbonate (PC) sheets. The gold caps stood above PC stems and provided ample spatial areas for capturing the biomarkers to be sensed. Three biomarkers served as the antigens and analytes, including human complement factor H (CFH), hyaluronic acid (HA), and nuclear matrix protein 22 (NMP22). Different antibodies, against the same biomarker, were covalently conjugated to AuNMs or gold nanoparticles, respectively. When the antibody–antigen–antibody sandwich structure formed, the plasmonic coupling between the AuNM surface and the gold nanoparticles significantly enhanced LSPR signals. The LSPR red shifts correlated quantitatively with the concentrations of the biomarkers. The limits of detection were 6.5, 8.3, and 7.0 pg/mL for CFH, HA, and NMP22, respectively. The chip’s specificity was tested and confirmed, excluding the nonspecific binding and false-positive possibility. The sensing performance of this sandwich immunoassay-based AuNM chip was better than that of the commercialized enzyme-linked immunosorbent assay. It provided a rapid, label-free, and easy operating platform for diagnosing and monitoring bladder cancer.