e17145 Background: The standard clinical practice to diagnose the presence and aggressiveness of prostate cancer, (PCa) consists of measuring serum prostate specific antigen (PSA), and performing image guided biopsies for histopathology. Due to the invasive nature potential risks and costs associated with prostate biopsy, urine based assays have been developed that require digital rectal examination (DRE) and to ascertain certain biomarkers present in the urine. These assays are not commonly integrated into clinical practice due to low sensitivity and high cost. Our goal was to develop a simple, noninvasive, reliable and affordable assay by targeting VPAC, a genomic receptor, expressed in high density on PCa malignant cells (MC), shed in non-DRE voided urine. Methods: Non DRE-Voided urine samples were collected from consenting patients (N=62, 56 ± 72 yrs) with histologically confirmed PCa and scheduled for radical prostatectomy. Urine was cytocentrifuged, cells on a glass slide fixed, and incubated with 0.5µg TP4303, a fluorophore containing small biomolecule developed in our laboratory with a high (3.1x10 -8 M) Kd value for VPAC. The excess TP4303 was washed, cells air dried, treated with DAPI, coverslip placed, and cells were analyzed using Zeiss microscope which calculated the % MC and the fluorophore intensity around MC. Data were corroborated with prostate grade group (GG) as determined by histology. Voided urine was also collected from consenting subjects (N= 97) clinically diagnosed with benign prostatic hyperplasia (BPH), with PSA <1.5 ng/ml (0.7±0.4, 60 8+ 6.3 yrs.). Results: Given in Table 1, indicate that the VPAC voided urine test was positive for all (N=62) PCa patients with increasing % MC shed in urine and their fluorescence intensity ,with increasing severity (GG) of PCa. Although these data points are small for statistical evaluations, the trend is remarkable that as the aggressiveness of disease increased, also increased were the % MC shed and the fluorescence intensity around them indicating increased VPAC density per MC. For BPH, the specificity of the assay was 89.6% to exclude prostate cancer with PPV 0.00% and NPV 100%. Conclusions: The data indicate i) using non-DRE voided urine, VPAC can be targeted to detect PCa, ii) the assay may also predict aggressiveness of PCa, iii) can distinguish PCa from BPH and iv) the assay is worthy of further investigation. Support: NIH 5RO1,CA249921 (MLT, ET). Patient clinical characteristics and results. PCa Grade Group No. of patients Age (yrs) PSA (ng/mL) Total cells Malignant cells % Malignant cells Fluorescence intensity (Mean ± Standard Deviation) 1 10 63.2±5.9 6.5 ±4.1 8588.±11679 2929±3356 41±28 31.8±3.1 2 31 63.90±7.3 7.2±3.8 5562±5207 2140±2286 49±29 33.7±4.0 3 13 65±9.2 13.1±14.5 4892±4143 2363±2979 37+27 33.0±6.3 4 2 65.50±6.3 6.5±2.1 4685±2616 1558±1351 48±56 32.2±5.6 5 6 68.20±4.1 50.2±104.9 8961±9877 7009±8237 66±27 35.7±5.7

OBJECTIVE:Contrast enhanced ultrasound (CEUS) is a cost-effective, safe, and accurate modality for monitoring renal cell carcinoma (RCC) recurrence following percutaneous ablation. However, ultrasound delineation of treated tumor borders can be challenging post-ablation. Here, we demonstrate the feasibility of using post-treatment CEUS fused to preablation MR/CT to detect RCC recurrence during long term follow up. METHODS:This study was performed as part of a larger ongoing prospective clinical trial for patients with biopsy-proven RCC treated with percutaneous ablation and receiving contrast enhanced (CE) -CT or -MRI for treatment response monitoring. CEUS with preablation CT/MR fusion was performed within 4 weeks of recurrence screening, and CE-CT/MRI was used as the reference standard. After intravenous injection of an ultrasound contrast agent, CEUS imaging of a single target lesion was performed with ultrasound fused to the patient's pretreatment CT or MRI. RCC recurrence was diagnosed at the bedside based on the presence of iso- to hyper-enhancement within the margins of the ablation cavity compared to normal renal parenchyma. RESULTS:To date, 50 participants have been recruited (76% male and 24% female). All participants tolerated the contrast injections without adverse events. CEUS was successfully fused to the participant's pretreatment cross-sectional imaging in all cases and was found to aid in the delineation of the original treatment zone. Additionally, CEUS correlated perfectly (100% agreement) with CE-CT/MRI findings for all the participants. CONCLUSION:Preliminary results demonstrate that post-treatment CEUS fused with a pretreatment CT/MRI is feasible and may aid in the correct localization of the treated tumor margins during long-term post-ablation monitoring.

PURPOSE:Prostate cancer (PCA) germline testing (GT) informs precision therapy, cancer screening, and hereditary cancer risk for patients and families. To support patient-centered PCA GT, studying patient-reported outcomes (PROs) is essential. METHODS:PROGRESS was a national patient-driven registry (January 2021-April 2022) for English-speaking males older than 18 years with previous/current PCA GT and Internet access. Surveys collected demographics, PCA history, family cancer history, mode of genetics care delivery, satisfaction with genetic counseling, decisional conflict, cancer genetics knowledge, and attitudes toward GT. Multiple linear regression modeling was used to estimate and draw inferences (α = .05) on strength of relationships between participant characteristics and PROs. RESULTS:Analyses focused on 414 participants: White (88%), Black (3%), Asian (6%), and mixed/other (3%). Most participants were non-Hispanic (95.2%) and 46.9% had PCA. Genetic results were positive (pathogenic/likely pathogenic variants; mutations) in 27.9%. The three most common modes of genetics care were meeting with genetics professional (in-person or remotely; 30.9%), discussing with doctor (21.1%), and using website (20.8%). In covariate-adjusted models, satisfaction scores were highest with pretest counseling by phone (β = 1.31; 95% CI, 0.26 to 2.36) or discussion with doctor (β = 1.25; 95% CI, 0.38 to 2.12). Lower decisional conflict scores were reported for pretest counseling by phone (β = -3.76; 95% CI, -7.28 to -0.24). Males with mutations reported higher GT benefit scores (β = .30; 95% CI, 0.02 to 0.59) and importance of GT (β = .34; 95% CI, 0.08 to 0.61). Asian Americans reported lower GT satisfaction (β = -2.91; 95% CI, -4.34 to -1.48) and higher decisional conflict (β = 8.93; 95% CI, 4.36 to 13.51). CONCLUSION:PROGRESS Registry informs the first comprehensive report of PROs among males undergoing PCA GT, providing insights into opportunities to improve patient experience and leverage the benefit of GT.

PURPOSE:To evaluate evidence on germline and somatic genomic testing for patients with metastatic prostate cancer and provide recommendations. METHODS:A systematic review by a multidisciplinary panel with patient representation was conducted. The PubMed database was searched from January 2018 to May 2024. Articles were selected for inclusion if they reported on patients with metastatic prostate cancer who received a germline or somatic genomic test and/or made comparisons between those tests, reported detection rates, prognostic information, or treatment implications. RESULTS:A total of 1,713 papers were identified in the literature search. After applying the eligibility criteria, 14 remained: eight systematic reviews and six clinical trials. RECOMMENDATIONS:Patients with metastatic prostate cancer should undergo both germline and somatic DNA sequencing using panel-based assays. These tests can guide the use of poly(ADP-ribose) polymerase inhibitors, which have a survival benefit in metastatic castration-resistant prostate cancer. In addition, germline testing may have screening implications for additional cancers for patients and cascade testing implications for family members. The data supporting when to perform repeat testing and optimal tissue type to use (eg, primary tumor v metastatic biopsy versus circulating tumor DNA [ctDNA] testing) are more limited, but this panel recommends considering retesting in patients whose results were previously negative or uninformative, and to consider either a metastatic biopsy or ctDNA when a significant change in clinical status occurs. Next-generation genomic sequencing findings that are associated with prognostic only (and not predictive) value should not be used to guide treatment outside of a clinical trial.Additional information is available at www.asco.org/genitourinary-cancer-guidelines.

PURPOSE:In QUILT-3.032, the efficacy of interleukin-15 receptor agonist, nogapendekin alfa inbakicept (NAI), in combination with bacillus Calmette-Guérin (BCG) for BCG-unresponsive high-grade papillary-only nonmuscle-invasive bladder cancer was assessed. In this study, we report the 36-month follow-up among participants with BCG-unresponsive papillary disease (cohort B). MATERIALS AND METHODS:NCT03022825 is an open-label, multicenter study of patients with BCG-unresponsive high-grade Ta/T1 papillary nonmuscle-invasive bladder cancer who received 400 μg NAI plus 50 mg BCG intravesically weekly for 6 consecutive weeks. The primary end point is disease-free survival (DFS) at 12 months. Progression-free survival (PFS), disease-specific survival (DSS), and cystectomy avoidance were assessed. Treatment-related adverse events were assessed. RESULTS:At July 15, 2024, data cutoff, the DFS rates at 12, 24, and 36 months were 58.2% (95% CI: 46.6, 68.2), 52.1% (95% CI: 40.3, 62.7), and 38.2% (95% CI: 25.6, 50.6), respectively. The PFS rates at 12 and 36 months were 94.9% (95% CI: 86.9, 98.0) and 83.1% (95% CI: 69.5, 91.0). The DSS rates at 12 and 36 months were 98.7% (95% CI: 91.4, 99.8) and 96.0% (95% CI: 88.2, 98.7). The median DSS has not been reached. Cystectomy avoidance rates at 12 and 36 months were 92.2% (95% CI: 83.4, 96.4) and 81.8% (95% CI: 68.1, 90.1), with median time to cystectomy not reached. Most treatment-related adverse events were grade 1 to 2 (61%) with 3% grade 3 and no grade 4 to 5. CONCLUSIONS:The 12-month and 36-month DFS, PFS, DSS, and cystectomy avoidance rates demonstrate the effectiveness and safety of NAI plus BCG in the management of BCG-unresponsive papillary disease. TRIAL REGISTRATION:ClinicalTrials.gov Identifier: NCT03022825.

Background: The standard diagnostic approach for prostate cancer (PCa) diagnosis consists of serum prostate-specific antigen (PSA) testing, digital rectal examination (DRE) and image-guided targeted biopsies. Given the invasive nature, potential adverse events and costs associated with these techniques, alternative approaches have been investigated, specifically with serum and urine assays. The work presented here is intended to further validate a novel noninvasive optical technique for PCa detection, targeting the VPAC genomic receptors that are overexpressed on prostate cancer’s malignant cells (MC), in non-DRE voided urine. Methods: Patients (N = 62) who had image-guided biopsy and histologically confirmed localized PCa, and who were scheduled for radical prostatectomy, provided a non-DRE voided urine sample prior to surgery. Urine was cytocentrifuged and cells fixed on a glass slide, incubated with 0.5 μg TP4303 (a receptor-specific fluorophore developed in our laboratory with high affinity for VPAC), excess washed and treated with 4,6-diamidodino-2-phenylindole (DAPI) for nuclear staining. The field of cells on each slide was analyzed using a Zeiss AX10 Observer microscope (20×). The total number of cells and MC were then counted, and the florescent intensity around each MC was measured using Zeiss software. Additionally, non-DRE voided urine samples collected from clinically determined BPH patients (N = 97), were also analyzed similarly. Results: Urine samples from 62 patients were processed and analyzed. Mean PSA levels by Gleason grade (GG) group were 6.5 ± 4.1 ng/mL for GG1 (N = 10), 7.2 ± 3.8 for GG2 (N = 31), 13.2 ± 14.6 for GG3 (N = 13), 6.2 ± 2.2 for GG4 (N = 2) and 50.2 ± 104.9 for GG5 (N = 6). Like the PSA, % MC shed (66.7 ± 27.7) in voided urine and the fluorescent intensity (35.8 ± 5.7) were highest in patients with GG5 prostate cancer. All PCa patients in GG1 to GG5 shed MC in voided urine with increasing % of MC and increasing fluorescence intensity which correlated with the increasing GG for PCa. For BPH, the specificity for the assay was 89.6% (95% CI:81.9–94.9%), PPV was 0.0% and NPV was 100% (95.9% CI, 95.9–100%). Conclusions: These data indicate the following: (i) PCa MC shed in non-DRE voided urine can be detected by targeting VPAC receptors, (ii) MC are shed in non-DRE voided urine with increasing quantity, corresponding to the severity of the disease, and (iii) this non-DRE voided urine optical assay provides a simple, noninvasive, and reliable method for the preliminary detection of PCa with potentially a lower cost than the currently available pre-biopsy detection technologies.

Multiparametric MRI (mpMRI) is currently the standard imaging modality for the diagnosis of prostate cancer; however, studies have reported that targeted biopsy based on mpMRI may miss approximately 30% of clinically significant cases. Recent advances in ultrasound imaging have improved its accuracy for detection of prostate cancer. Newer techniques such as MicroUS, elastography, contrast-enhanced ultrasound (CEUS), and contrast ultrasound dispersion imaging (CUDI) have enabled a comprehensive, real-time, and relatively inexpensive approach to evaluate the prostate gland. Multiparametric ultrasound (mpUS) integrates multiple parameters from these techniques to generate multiparametric maps akin to those produced by mpMRI, to localize prostate cancer. This review aims to explore the performance of modern ultrasound techniques and mpUS for diagnosis of prostate cancer, comparing them with mpMRI.

AbstractObjectivesThe objective of this study is to validate a hypothesis that a non‐invasive optical imaging assay targeting genomic VPAC receptors on malignant cells shed in voided urine will represent either benign prostatic hyperplasia (BPH) or prostatic cancer (PCa). Risk for BPH in men 50–70 years old is 50–70% and PCa is 17%. BPH and PCa can coexist in 20% of men with BPH. Most commonly practiced methods to diagnose BPH do not distinguish BPH from PCa.Patients (or Materials) and MethodsMales with BPH (N = 97, 60.8 ± 6.3 years, prostate‐specific antigen 0.7 ± 0.4 ng/mL) and without oncologic disease (N = 35, 63.4 ± 5.8 years, prostate‐specific antigen < 1.5 ng/mL) signed informed consent form and provided voided urine. Urine was cytocentrifuged, cells collected on glass slide, fixed, treated with VPAC specific fluorophore TP4303 (Kd 3.1 × 10−8M), washed, incubated with DAPI and observed using a fluorescence microscope. Cells with no VPAC did not fluoresce (BPH) and those with VPAC had red‐orange fluorescence (PCa). Real‐time polymerase chain reaction analyses for VPAC and NKX3.1 assay for cell origin were performed.ResultsEighty‐seven subjects were negative for VPAC expression. Positive VPAC expression was noted in 10 subjects. Patient chart review for clinical data on these 10 VPAC positive subjects showed five had nephrolithiasis, three had renal cysts, one had prostatitis and one was being treated with finasteride. Real‐time polymerase chain reaction analysis‐VPAC expressions for 7 normal and 12 BPH subjects were 1.31 ± 1.26 and 0.94 ± 0.89, respectively (P = 0.46). NKX3.1 showed cells of prostate origin for finasteride‐treated patient. Specificity for VPAC urine assay for excluding prostate cancer in this BPH cohort was 88.5%, positive predictive value 0.00% and negative predictive value 100%.ConclusionVPAC assay may contribute extensively for BPH diagnosis and warrant continued investigation.

478 Background: Radiation therapy (RT) is an integral component of the multimodal therapy of pelvic malignancies, either as primary treatment or in combination with surgical resection. In addition to local treatment effects on nearby pelvic organs, RT has been established to be a risk factor for delayed secondary malignancies. In this study, we examine the rate of any secondary malignancies following RT for primary pelvic malignancies, with a specific emphasis on secondary pelvic malignancies Methods: Using the SEER (Surveillance, Epidemiology, and Ends Results) database, we retrospectively examined 2,102,192 patients with primary pelvic malignancies (prostate, bladder, uterine, rectal, cervical). For each disease site, we compared the rate of all secondary malignancies in radiated patients to non-radiated patients. Secondary malignancies were then stratified as pelvic and non-pelvic, in order to determine the local effect of RT on malignancy risk. Results: A total of 2,102,192 patients were examined (1,189,108 prostate, 315,026 bladder, 88,809 cervical, 249,535 uterine, 259,714 rectal). A total of 113,322 patients developed secondary malignancies after RT (Table), with 26,299 developing secondary pelvic malignancies after RT (18,411 prostate, 1,026 bladder, 1,410 cervical, 2,179 uterine, 3,273 rectal) (Table). The overall relative risk (RR) of RT on developing a secondary malignancy was 1.79 (1.77-1.80 CI, P<0.0001), particularly in patients with prostate (RR 2.57), uterine (RR 1.24) and cervical cancer (1.09). The overall RR of RT on developing a secondary pelvic malignancy was 2.09 (2.06-2.13 CI, P<0.0001), particularly in patients with bladder (RR 6.90), prostate (RR 2.74), and uterine cancer (RR 1.21). Conclusions: Radiation treatment for pelvic malignancies increases the risk of developing secondary malignancies over the patient’s lifetime. Further work needs to done to identify at risk populations.[Table: see text]

Objective. To examine one academic institution’s experiences with SpaceOAR placement, its associated complications, and periprocedural characteristics that affect outcomes for the purpose of quality improvement. Materials and Methods. We conducted a retrospective review of 233 patients who received SpaceOAR from four surgeons and one radiation oncologist between 2018 and 2021. Variables such as demographics, oncologic parameters, radiation plan, and radiographic assessment of hydrogel placement were recorded. The Charlson Comorbidity Index (CCI) was used to assess comorbidity risk. Mann–Whitney and Fisher’s exact tests were performed to compare patients with and without complications. Results. Of the 233 patients who received SpaceOAR, 24 (10.3%) experienced toxicity. All complications were Clavien I or II, such as pelvic pain postplacement, pelvic fullness, bleeding, and lower urinary tract symptoms. 16 patients (6.9%) had some portion of the hydrogel injected into the rectal wall, but it was never clinically significant. The average CCI was 3.2 ± 0.95 for patients who experienced complications; the average CCI was 3.6 ± 1.6 (p = 0.48) in the group without complications. Of the physicians with higher procedure volumes, Physician #1 had the highest rate of patient‐reported complications at 11 out of 68 (16.2%) and Physician #2 had the lowest rate of complications at 4 out of 96 placements (4.2%). Multivariate analysis found that patients who had received hormone therapy previously had less odds of reporting complications after SpaceOAR placement. Conclusions. The listed attending on the procedure had a significant correlation to complications with SpaceOAR placement on univariate analysis, and hormone therapy had some benefits to the tolerance for the procedure on multivariate analysis. Overall, the hydrogel placement was well tolerated with low incidence of mild and transient procedure‐related toxicity.

Purpose: Targeted prostate biopsies are increasingly being performed by urologists in the United States includingthose in the Pennsylvania Urologic Regional Collaborative, a physician-led data-sharing and quality improvement collaborative. To evaluate the performance of MRI guided fusion needle prostate biopsies in the collaborative, we analyzed the variability by practice in rates of detection of clinically significant prostate cancer and patient characteristics associated with detection of clinically significant prostate cancer. Methods: We analyzed 857 first-time MRI fusion biopsy procedures performed at five practices (minimum 20 procedures) between 2015 and 2019. We used chi-square analysis for baseline patient characteristics and Grade Group (GG) >= 3 tumor detection rates by practice. Multivariable logistic regression was used to estimate the odds of clinically significant cancer detection when adjusting for baseline patient characteristics. Results: Approximately 15% of men undergoing targeted MRI guided biopsy were <= 59 years old. Median prostate specific antigen (PSA) was 6.8 ng/ml. Detection rates for GG >= 3 tumors ranged from 14.3% to 28.3% (P = 0.02) across practices. However, the odds of GG >= 3 tumor detection did not differ significantly between practices after adjusting for clinical and radiographic factors. Overall, increased likelihood of detecting a GG >= 3 tumor was associated with increased age, DRE abnormalities, higher PSA, smaller gland volume and PI-RADS >= 4 MRI lesions. There was an 81% concordance rate between PI-RADS >= 4 and Gleason grade >= 3 prostate cancer. Conclusion: We demonstrate the value of obtaining pre-biopsy MRI given high concordance between presence of suspicious lesions and MRI-targeted biopsy detection of clinically significant prostate cancer. Variability of baseline patient characteristics among practices may account for the observed differences in clinically significant cancer detection rates. These findings can aid standardization and quality improvement efforts within the collaborative.

You have accessJournal of UrologyProstate Cancer: Detection & Screening VI (MP74), Moderated Poster 741 May 2024MP74-20 VPAC TARGETED DIAGNOSTIC OPTICAL IMAGING ASSAY (VPAC ASSAY) FOR PROSTATE CANCER Mathew (Madhukar) Thakur, Vivek S. Tomar, Emma Dale, Leonard G. Gomella, Michael Chaga, Oleksandr Kolesnikov, Hector T. Navarro, Joon Yau Leong, Olivia Dahlgren, and Edouard J. Trabulsi Mathew (Madhukar) ThakurMathew (Madhukar) Thakur , Vivek S. TomarVivek S. Tomar , Emma DaleEmma Dale , Leonard G. GomellaLeonard G. Gomella , Michael ChagaMichael Chaga , Oleksandr KolesnikovOleksandr Kolesnikov , Hector T. NavarroHector T. Navarro , Joon Yau LeongJoon Yau Leong , Olivia DahlgrenOlivia Dahlgren , and Edouard J. TrabulsiEdouard J. Trabulsi View All Author Informationhttps://doi.org/10.1097/01.JU.0001008632.59099.b9.20AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Managing prostate cancer (PCa) effectively with a radical treatment is challenging. Risk stratification depends strongly, among other factors, on Gleason score or Grade Group (GG) of PCa. Neither PSA, nor other available ex-vivo tests can determine PCa, other than histopathology for which invasive biopsy must be performed. We have developed a VPAC Assay which targets genomic VPAC receptors expressed in high density on PCa malignant cells (MC) shed in voided urine and detects PCa with >95% sensitivity. The assay uses a receptor specific fluorophore (Kd 3.1x10-8M), and facilitates optical imaging of MC. The goal of this retrospective investigation was to examine if quantification of percent MC cells (with respect to total number of cells shed in voided urine) and the fluorescence intensity around MC can image PCa and determine severity of PCa. METHODS: Voided urine was collected from 101 consenting PCa patients scheduled for radical prostatectomy. Urine was cyto-centrifuged, cells on glass slide fixed, incubated with receptor specific fluorophore, washed, treated with DAPI (4,6-diamidodino-2-phenylindole), cover slip placed and analyzed with Zeiss Axio Observer microscope, coupled with ZENN Microscopy software, which determined percent PCa MC for each patient. The patient data, and the corresponding imaging results were analyzed as per GG, determined by post-surgical gold standard histopathology and evaluated statistically (Fig. 1). RESULTS: Although patient number is limited for a certain GG, it clearly depicts a trend, that with increasing GG, there was increased number of MC shed, with enhanced fluorophore intensity around them. Enhanced intensity around MC is thought to have resulted from increased VPAC receptor density on MC shed from more aggressive PCa. MC shed in urine increased from 13.1±14.4% for GG1 to 25.3±21.07% for GG3 (p=0.05) to 50.5±26.7% for GG5 (p=0.002). Similarly, fluorescence intensity increased from 43.8±2.2 for GG1 to 48.8±7.0 for GG3 (p=0.01) to 51.5±14.4 for GG5 (p=0.15). CONCLUSIONS: Data suggest that this simple, inexpensive, truly noninvasive diagnostic urinary VPAC assay may contribute to the management of PCa. Download PPT Source of Funding: NIH NC5R01CA249921 © 2024 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 211Issue 5SMay 2024Page: e1201 Advertisement Copyright & Permissions© 2024 by American Urological Association Education and Research, Inc.Metrics Author Information Mathew (Madhukar) Thakur More articles by this author Vivek S. Tomar More articles by this author Emma Dale More articles by this author Leonard G. Gomella More articles by this author Michael Chaga More articles by this author Oleksandr Kolesnikov More articles by this author Hector T. Navarro More articles by this author Joon Yau Leong More articles by this author Olivia Dahlgren More articles by this author Edouard J. Trabulsi More articles by this author Expand All Advertisement PDF downloadLoading ...

PURPOSEThere is significant interest in identifying complete responders to neoadjuvant chemotherapy (NAC) before radical cystectomy (RC) to potentially avoid removal of a pathologically benign bladder. However, clinical restaging after NAC is highly inaccurate. The objective of this study was to develop a next-generation sequencing-based molecular assay using urine to enhance clinical staging of patients with bladder cancer.METHODSUrine samples from 20 and 44 patients with bladder cancer undergoing RC were prospectively collected for retrospective analysis for molecular correlate analysis from two clinical trials, respectively. The first cohort was used to benchmark the assay, and the second was used to determine the performance characteristics of the test as it correlates to responder status as measured by pathologic examination.RESULTSFirst, to benchmark the assay, known mutations identified in the tissue (MT) of patients from the Accelerated Methotrexate, Vinblastine, Doxorubicin, Cisplatin trial (ClinicalTrials.gov identifier: NCT01611662, n = 16) and a cohort from University of California-San Francisco (n = 4) were cross referenced against mutation profiles from urine (MU). We then determined the correlation between MU persistence and residual disease in pre-RC urine samples from a second prospective clinical trial (The pT0 trial; ClinicalTrials.gov identifier: NCT02968732). Residual MU status correlated strongly with residual disease status (pT0 trial; n = 44; P = .0092) when MU from urine supernatant and urine pellet were assessed separately and analyzed in tandem. The sensitivity, specificity, PPV, and NPV were 91%, 50%, 86%, and 63% respectively, with an overall accuracy of 82% for this second cohort.CONCLUSIONMU are representative of MT and thus can be used to enhance clinical staging of urothelial carcinoma. Urine biopsy may be used as a reliable tool that can be further developed to identify complete response to NAC in anticipation of safe RC avoidance.

Abstract Introduction: Prostatic cancer (PCa) is the second most prevalent disease in men globally. In 2020 it lead to >14 million new cases and >375,000 deaths worldwide. Despite numerous detection methods, detecting PCa reliably and non-invasively continues to be challenging. PCa malignant cells (MC) express VPAC (combined for Vasoactive Intestinal Peptide and Pituitary Adenylate Cyclase Activating Peptide) in high density. A VPAC specific PACAP analogues (Kd=3.1 × 10−8M) developed in our laboratory was conjugated with a fluorophore and named TP4303. The goal was to assess if TP4303 will image PCa-MC shed in voided urine and present severity of PCa as grade group (GG). Method: Consenting PCa patients (N=110) scheduled for radical prostatectomy provided voided urine. Urine was cyto-centrifuged. Cells on glass slide were fixed, incubated with TP4303, washed, treated with DAPI (4,6-diamidodino-2-phenylindole), cover slip placed and analyzed with Zeiss Axio Observer microscope, coupled with ZENN Microscopy software. The software determined percentage of PCa MC shed by each patient. The results were corroborated as per GG, determined by post-surgical histopathology and analyzed statistically. Results:TP4303 imaged PCa in all subjects. The percent MC shed in urine increased from 13.1±14.4% for GG1 to 25.3±21.07% for GG3 (p=0.05) to 50.5±26.7% for GG5 (p=0.002). Similarly, the fluorescence intensity increased from 43.8±2.2 for GG1 to 48.8±7.0 for GG3 (p=0.01) to 51.5±14.4 for GG5 (p=0.15). Conclusions: The data strongly suggests that TP4303 may not only detect PCa reliably and noninvasively, but may also predict the severity of PCa. Supported by NIH: NC5R01CA249921 Citation Format: Mathew L. Thakur, Vivek S. Tomar, Emma Dale, Leonard G. Gomella, Hector T. Navarro, Oleksandr Kolesnikov, Joon Yau Leong, Olivia Dahlgren, Edouard J. Trabulsi. Targeting genomic biomarker in voided urine for prostate cancer detection [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1031.

<p>Supplementary Figures 1-6</p>

Supplemental Table 1: Structure-Activity-Relationship analysis.

124 Background: Nearly 50% of patients with persistently elevated PSA (PEP) have prostate cancer (PCa) despite previous negative biopsy. However not all patients agree to undergo repeat, invasive trans rectal or trans perineal prostate biopsy (PB). To identify men at high risk of PCa, the need is unmet of other reliable diagnostic methods. The goal of this investigation was to determine if the VPAC assay, which is being developed in our laboratory and detects PCa efficiently, by imaging PCa cells shed in voided urine, collected without digital rectal examination (DRE), can address the common clinical dilemma of when or if to repeat biopsy on PEP subjects. Methods: PEP patients (N=21, 55-75 yr. PSA 2.8 – 42 ng/ml), scheduled for PB and signed informed consent form were enrolled. They provided pre and clinically required, post DRE urine. VPAC assay was performed on both urine samples from each patient. VPAC, a genomic receptor, encodes G protein and expresses in high density on all malignant PCa cells, was targeted with TP4303, a biomolecule with fluorophore, designed in our laboratory with high affinity (Kd, 3.1x10 -9 M) for VPAC. Urine was centrifuged, cells on a glass slide fixed, incubated with TP4303 and excess was washed. Cells were then examined under a microscope. TP4303 fluorescence around cell nucleus allowed to identify malignant PCa cells. For urine collected post DRE, MDx Health assay, approved by FDA and determines mRNA levels of DXL1 and HOXC6 biomarkers and indicates risk of PCa, was also performed. Results were compared with gold standard histopathology of tissues acquired through PB, and evaluated statistically. Results: Of 21 PEP patients, who underwent PB, three (14%) had histopathologically proven, clinically significant PCa with Grade Group 2. All three (100%) patients were correctly identified as positive for PCa by the VPAC assay, while MDx reported low risk for two (9.5%) and high risk for one (53%). Of the remaining 18, histologically negative subjects, VPAC assay was disconcordant in five (28%), with positive for PCa. In these five patients, MDx also reported moderate to high risk (40%-59%). Of the remaining 13 histologically negative PCa patient, all (100%) had concordant results with VPAC assay while MDx reported low risk in 2 patients and moderate to high risk (range 30% to 90%) in other eleven. Results with sensitivity and specificity of the assays are given in the table. Conclusions: The VPAC assay data, in this small exploratory cohort, appear promising as an additional diagnostic in the clinical management of PEP patients, minimizing the number of unnecessary invasive PBs and eliminating possible overtreatment. Work is in progress. Support: NIH 5R01CA249921. [Table: see text]

Supplementary Table 5: IST5-002 regulation of known Stat5 target genes in K562 cells.