Integrated analysis of multiple transcriptomic approaches and machine learning integration algorithms reveals high endothelial venules as a prognostic immune-related biomarker in bladder cancer.

BACKGROUND: Despite the availability of established surgical and chemotherapy options, the treatment of bladder cancer (BCa) patients remains challenging. While immunotherapy has emerged as a promising approach, its benefits are limited to a subset of patients. The exploration of additional targets to enhance the efficacy of immunotherapy is a valuable research direction. METHOD: High endothelial venules (HEV) ssGSEA analysis was conducted using BEST. Through the utilization of R packages Limma, Seurat, SingleR, and Harmony, analyses were performed on spatial transcriptomics, bulk RNA-sequencing (bulk RNA-seq), and single-cell RNA sequencing (scRNA-seq) data, yielding HEV-related genes (HEV.RGs). Molecular subtyping analysis based on HEV.RGs was conducted using R package MOVICS, and various machine learning-integrated algorithm was employed to construct prognostic model. LDLRAD3 was validated through subcutaneous tumor formation in mice, HEV induction, Western blot, and qPCR. RESULTS: A correlation between higher HEV levels and improved immune response and prognosis was revealed by HEV ssGSEA analysis in BCa patients receiving immunotherapy. HEV.RGs were identified in subsequent transcriptomic analyses. Based on these genes, BCa patients were stratified into two molecular clusters with distinct survival and immune infiltration patterns using various clustering-integrated algorithm. Prognostic model was developed using multiple machine learning-integrated algorithm. Low LDLRAD3 expression may promote HEV generation, leading to enhanced immunotherapy efficacy, as suggested by bulk RNA-seq, scRNA-seq analyses, and experimental validation of LDLRAD3. CONCLUSIONS: HEV served as a predictive factor for immune response and prognosis in BCa patients receiving immunotherapy. LDLRAD3 represented a potential target for HEV induction and enhancing the efficacy of immunotherapy.

Effects of combined radiotherapy with immune checkpoint blockade on immunological memory in luminal-like subtype murine bladder cancer model.

MIBC is a highly lethal disease, and the patient survival rate has not improved significantly over the last decades. UPPL is a cell line that can be used to recapitulate the luminal-like molecular subtype of bladder cancer and to discover effective treatments to be translated in patients. Here, we investigate the effects of combinational treatments of radiotherapy and immunotherapy in this recently characterized UPPL tumor-bearing mice. We first characterized the baseline tumor microenvironment and the effect of radiation, anti-PD-L1, and combinatorial treatments. Then, the mice were re-challenged with a second tumor (rechallenged tumor) in the contralateral flank of the first tumor to assess the immunological memory. Radiation slowed down the tumor growth. All treatments also decreased the neutrophil population and increased the T cell population. Anti-PD-L1 therapy was not able to synergize with radiation to further delay tumor growth. Furthermore, none of the treatments were able to generate immune memory. The treatments were not sufficient to induce a significant and lasting pool of memory cells. We show here that anti-PD-L1 treatment added to radiotherapy was not enough to achieve T cell-mediated memory in UPPL tumors. Stronger T cell activation signals may be required to enhance radiation efficacy in luminal-like bladder cancer.

Current Advances in the Management of Nonurothelial Subtypes of Bladder Cancer.

Urothelial cancer (UC) is the most common histology seen in bladder tumors. The 2022 WHO classification of urinary tract tumors includes a list of less common subtypes (formerly known as variants) for invasive UC which are considered high-grade tumors. This review summarizes the most recent advances in the management of selected nonurothelial subtypes of bladder cancer: squamous cell carcinoma, small cell carcinoma, sarcomatoid urothelial carcinoma, micropapillary carcinoma, plasmacytoid carcinoma, adenocarcinoma, and urachal carcinoma. The role of neoadjuvant and adjuvant chemotherapy has not been well characterized for most of these histologies, and prospective data are extremely limited. Participation in clinical trials is recommended in advanced disease.

Practice patterns and survival outcomes for muscle-invasive bladder cancer: real-life experience in a general population setting.

Bladder cancer (BC) is a common malignancy in Europe and North America. Among BCs, muscle-invasive BCs (MIBCs) are distinguished, as they require aggressive treatment due to their spreading potential and poor prognosis. Despite its clinical relevance, little information on MIBC in a general population setting is available. This study aims to report practice patterns and survival outcomes for MIBC patients in a general population setting. MIBCs among BC incidence in 2011 and 2012 recorded in a French population-based cancer registry (810 000 inhabitants) were included in the study. Data were extracted from the medical files. Individual, tumour-related characteristics and initial management including diagnostic tools, multidisciplinary team meeting (MDT) assessment, and treatment delivered were described. Cystectomy, chemoradiation, radiotherapy, and chemotherapy were considered as specific treatments. Matching between MDT decision and the treatment provided was detailed. Management practices were discussed according to the guideline's recommendations. Overall survival (using the Kaplan-Meier method) and net survival (using the Pohar-Perme estimator) were calculated. Among 538 incident BC cases, 147 (27.3%) were MIBCs. Diagnostic practices displayed a relevant locoregional assessment of BC. Almost all cases (n = 136, 92.5%) were assessed during an uro-oncological MDT with a median time from diagnosis of 18 days (first quartile:12-third quartile:32). Discrepancies appeared between MDT decisions and treatments delivered: 71 out of 86 subjects received the recommended cystectomy or chemoradiation (with or without neoadjuvant chemotherapy); 6 out of 11 had the recommended radio- or chemotherapy; and 9 patients did not undergo any specific treatment despite the MDT decision. Cystectomy was the most common treatment performed; the time to surgery appeared consistent with the guideline's recommendations. Forty people only received supportive care. Still, the 5-year overall and net survival was poor, with 19% (13-26) and 22% (14-31), respectively. The 5-year net survival was 35% (23-48) for people who underwent curative-intent treatments. MIBC management remains challenging even for cases assessed during an MDT. Many people did not undergo any specific treatment. Prognosis was poor even when curative-intent therapies were delivered. Efforts to reduce exposure to risk factors such as tobacco smoking and occupational exposures must be maintained.

Grade Heterogeneity in High-Grade Urothelial Carcinomas: Does It Have an Impact on the Survival of Patients With Intermediate/High-Risk Nonmuscle-Invasive Bladder Cancer Who Received Adequate Adjuvant Bacillus Calmette-Guérin Therapy?

PURPOSE: We aimed to compare recurrence-free survival (RFS) and progression-free survival (PFS) of the patients with pure high-grade (HG) vs mixed-grade (MG) nonmuscle-invasive bladder cancer who received adequate bacillus Calmette-Guérin therapy. MATERIALS AND METHODS: We conducted a retrospective cohort analysis using data from an institutional database. The study included patients diagnosed with HG nonmuscle-invasive bladder cancer at the initial transurethral resection specimen between 2010 and 2020. The initial transurethral resection specimens of all patients were reevaluated by a dedicated uropathologist. The percentage of low-grade tumor areas accompanying HG areas was determined for each case. Time-to-event analysis was performed using the Kaplan-Meier method. RFS and PFS rates were compared between groups. RESULTS: Of the 203 patients enrolled in the study, 69 (34%) had MG tumors. Recurrence was observed in 41 out of 134 patients (30.6%) in the HG group and in 19 out of 69 patients (27.5%) in the MG group. The 36-month RFS rates were 69% (CI: 62-77) and 72% (CI: 62-83) for the HG-urothelial carcinoma (UC) and MG-UC groups, respectively. The RFS rates were similar between groups (log-rank, P = .58). Progression was observed in 22 out of 134 patients (16.4%) in the HG group and in 4 out of 69 patients (5.8%) in the MG group. The 36-month PFS rates were 84% (CI: 77-90) and 94% (CI: 89-100) for the HG-UC and MG-UC groups, respectively. The pure HG-UC group had a worse PFS than the MG-UC group (log-rank, P = .042). Multivariate analysis demonstrated that age and tumor grade were significant risk factors for the development of progression. CONCLUSIONS: The indication of MG-UC category separately from pure HG carcinomas in the pathology report seems to be an important issue that can guide patient management. In this way, both more accurate risk classification and more accurate patient counseling can be performed. More importantly, the treatment plan can be made more accurately. For more precise conclusions, our results should be supported by prospective studies with larger sample size.

Defining oligometastatic state in uro-oncological cancers.

PURPOSE OF REVIEW: Oligometastatic tumors illustrate a distinct state between localized and systematic disease and might harbor unique biologic features. Moreover, these tumors represent a different clinical entity, with a potential of long-term disease control or even cure, therefore they receive growing attention in the field of urologic oncology. RECENT FINDINGS: Currently, there is no consensus on the definition of oligometastatic prostate cancer, most experts limit it to a maximum of three to five lesions and involvement of no more than two organs, excluding visceral metastases. Quality data on oligometastatic bladder cancer is scarce, however, a consensus of experts defined it as a maximum of three metastatic lesions, either resectable or suitable for stereotactic therapy, without restrictions to the number of organs involved. As for kidney cancer, a maximum number of five metastases, without limitations to the location are defined as oligometastatic, with an important implication of timing of developing metastases since diagnosis of the primary tumor. SUMMARY: Defining oligometastatic state among urological tumors reflecting their distinct biological and clinical behavior is crucial to establish a sound framework for future clinical trials, and to facilitate guideline and policy formulation for improved patient care. Advancements in molecular imaging are expected to transform the field of oligometastatic urologic tumors in the future.

Bladder Cancer, Version 3.2024.

Bladder cancer, the sixth most common cancer in the United States, is most commonly of the urothelial carcinoma histologic subtype. The clinical spectrum of bladder cancer is divided into 3 categories that differ in prognosis, management, and therapeutic aims: (1) non-muscle-invasive bladder cancer (NMIBC); (2) muscle invasive, nonmetastatic disease; and (3) metastatic bladder cancer. These NCCN Guidelines Insights detail recent updates to the NCCN Guidelines for Bladder Cancer, including changes in the fifth edition of the WHO Classification of Tumours: Urinary and Male Genital Tumours and how the NCCN Guidelines aligned with these updates; new and emerging treatment options for bacillus Calmette-Guérin (BCG)-unresponsive NMIBC; and updates to systemic therapy recommendations for advanced or metastatic disease.

Immune features are associated with response to neoadjuvant chemo-immunotherapy for muscle-invasive bladder cancer.

Neoadjuvant cisplatin-based chemotherapy is standard of care for muscle-invasive bladder cancer (MIBC). Immune checkpoint inhibition (ICI) alone, and ICI in combination with chemotherapy, have demonstrated promising pathologic response (

Advancements in Urothelial Cancer Care: Optimizing Treatment for Your Patient.

The standard treatment paradigm for muscle invasive bladder cancer has been neoadjuvant cisplatin-based chemotherapy followed by radical cystectomy. However, efforts are ongoing to personalize treatment by incorporating biomarkers to better guide treatment selection. In addition, bladder preservation strategies are aimed at avoiding cystectomy in well-selected patients. Similarly, in the metastatic urothelial cancer space, the standard frontline treatment option of platinum-based chemotherapy has changed with the availability of data from EV-302 trial, making the combination of enfortumab vedotin (EV) and pembrolizumab the preferred first-line treatment option. Here, we examine the optimization of treatment intensity and sequencing, focusing on the challenges and opportunities associated with EV/pembrolizumab therapy, including managing toxicities and exploring alternative dosing approaches. Together, these articles provide a comprehensive overview of contemporary strategies in bladder cancer management, highlighting the importance of individualized treatment approaches, ongoing research, and multidisciplinary collaboration to improve patient outcomes in this complex disease landscape.

Common immunological and prognostic features of lung and bladder cancer via smoking-related genes: PRR11 gene as potential immunotherapeutic target.

Smoking is a well-known risk factor for non-small-cell lung cancer (NSCLC) and bladder urothelial carcinoma (BLCA). Despite this, there has been no investigation into a prognostic marker based on smoking-related genes that could universally predict prognosis in these cancers and correlate with immune checkpoint therapy. This study aimed to identify smoking-related differential genes in NSCLC and BLCA, analyse their roles in patient prognosis and immune checkpoint therapy through subgroup analyses, and shed light on PRR11 as a crucial prognostic gene in both cancers. By examining PRR11 co-expressed genes, a prognostic model was constructed and its impact on immunotherapy for NSCLC and BLCA was evaluated. Molecular docking and tissue microarray analyses were conducted to explore the correlation between PRR11 and its reciprocal gene SPDL1. Additionally, miRNAs associated with PRR11 were analysed. The study confirmed a strong link between smoking-related genes, prognosis, and immune checkpoint therapy in NSCLC and BLCA. PRR11 was identified as a key smoking-associated gene that influences the efficacy of immune checkpoint therapy by modulating the stemness of these cancers. A prognostic model based on PRR11 co-expressed genes in BLCA was established and its prognostic value was validated in NSCLC. Furthermore, it was found that PRR11 regulates PDL1 via SPDL1, impacting immunotherapeutic efficacy in both cancers. The involvement of hsa-miR-200b-3p in the regulation of SPDL1 expression by PRR11 was also highlighted. Overall, the study elucidates that PRR11 modulates patient immunotherapy by influencing PDL1 expression through its interaction with SPDL1, with potential upstream regulation by hsa-miR-200b-3p.

Tumor microenvironment-modulating oncolytic adenovirus combined with GSK-3ß inhibitor enhances antitumor immune response against bladder cancer.

Bladder cancer is a common type of cancer around the world, and the majority of patients are diagnosed with non-muscle-invasive bladder cancer (NMIBC). Although low-risk NMIBC has a good prognosis, the disease recurrence rate and development of treatment-refractory disease remain high in intermediate- to high-risk NMIBC patients. To address these challenges for the treatment of NMIBC, a novel combination therapy composed of an oncolytic adenovirus (oAd) co-expressing interleukin (IL)-12, granulocyte-macrophage colony-stimulating factor (GM-CSF), and relaxin (RLX; HY-oAd) and a clinical-stage glycogen synthase kinase (GSK)-3ß inhibitor (9-ING-41; elraglusib) was investigated in the present report. Our findings demonstrate that HY-oAd and 9-ING-41 combination therapy (HY-oAd+9-ING-41) exerted superior inhibition of tumor growth compared with respective monotherapy in a syngeneic NMIBC tumor model. HY-oAd+9-ING-41 induced high-level tumor extracellular matrix (ECM) degradation and a more potent antitumor immune response than the respective monotherapy. In detail, HY-oAd+9-ING-41 induced superior accumulation of intratumoral T cells, prevention of immune cell exhaustion, and induction of tumor-specific adaptive immune response compared to either monotherapy. Collectively, these results demonstrate that the combination of HY-oAd and 9-ING-41 may be a promising approach to elicit a potent antitumor immune response against bladder cancer.

Predicting survival after brain metastases in patients with bladder cancer.

AIMS AND OBJECTIVES: Because of its rarity, limited data concerning brain metastasis (BM) from bladder cancer (BCa) are available, so this phenomenon remains unclear. We aimed to contribute to understanding this unique patient population's clinical behavior and outcomes. METHODS/MATERIALS: This retrospective cohort study included 27 BCa patients with BM treated at our Cancer Institute between April 2009 and December 2022. The time from initial diagnosis to BM and overall survival from BM diagnosis were calculated (Kaplan-Meier method). Cox regression was used to test key clinicopathologic associations. RESULTS: A total of 27 patients were included in the study (male/female = 23/4). The median patient age at BM diagnosis was 62.0 (47-79) years. The median interval from initial diagnosis to BM was 11.0 ± 2.59 (95 % CI, 5.91-16.08) months. Twenty (74.0 %) patients were diagnosed with BM by postsymptomatic imaging. The most common symptoms were headache-dizziness (n = 9, 33.3 %), seizure (n = 3, 11.1 %), hemiparesis (n = 2, 7.4 %), and vision defects (n = 2, 7.4 %). The most common sites of extracranial metastasis were the lung (n = 10, 52.6 %), bone (n = 7, 36.8 %), and lymph nodes (n = 6, 31.5 %). More than half of the patients (55.5 %) had multiple BMs. Eight (29.6 %) patients underwent surgery for BM. All of the patients received radiotherapy (RT) for BM (whole-brain radiotherapy (WBRT)/stereotactic radiotherapy (SRT) = 24/3), and eight patients received RT for the second time. Six patients were treated with systemic chemotherapy (CT) after BM. The median survival from BM was 3.0 ± 1.2 (95 % Cl, 0.4-5.5) months in the entire cohort. A low number of BMs (HR 0.270, 95 % CI 0.083-0.885; p = 0.031), surgery for BM (HR 0.174, 95 % CI 0.043-0.712; p = 0.015), CT after BM (HR 0.207, 95 % CI 0.057-0.755; p = 0.017), and better ECOG performance score (HR 0.248, 95 % CI 0.074-0.836; p = 0.025) were associated with better OS. CONCLUSIONS: Factors associated with improved survival in BCa patients with BM include a few brain lesions, intracranial resection, CT after BM, and better ECOG performance scores. Larger-scale prospective studies are needed to define the optimal management strategy further.

The Missing Trial Variable: A Case Report on Dietary Modification in a Patient with Durable Complete Remission After an Immunotherapy Clinical Trial for Metastatic Bladder Cancer.

A patient with metastatic bladder cancer has had an ongoing 9-year complete remission on a clinical trial of immunotherapy with nivolumab and ipimumab, but he also was following an intensive dietary and nutritional supplement regimen. Whether this combination or the immunotherapy alone brought about his good outcome is unknown but could be clarified in future trials by improved data collection about dietary and supplement choices.

Comparative analysis of recurrence rates between intravesical gemcitabine and bacillus Calmette-Guérin induction therapy following transurethral resection of bladder tumors in patients with intermediate- and high-risk bladder cancer: A retrospective multicenter study.

PURPOSE: This study investigated the efficacy of intravesical gemcitabine as an alternative to bacillus Calmette-Guérin (BCG) therapy. MATERIALS AND METHODS: Data were retrospectively collected across seven institutions from February 1999 to May 2023. Inclusion criteria included patients with intermediate- or high-risk non-muscle invasive bladder cancer (NMIBC) who underwent transurethral resection of bladder tumors (TURBT) and received at least four sessions of intravesical gemcitabine or BCG induction therapy. Patient characteristics, complete remission (CR), occurrence, and progression rates were compared. RESULTS: In total, 149 patients were included in this study (gemcitabine, 63; BCG, 86). No differences were apparent between the two groups in baseline characteristics, except for the follow-up period (gemcitabine, 9.2±5.9 months vs. BCG, 43.9±41.4 months, p<0.001). There were no consistent significant differences observed between the two groups in the 3-month (gemcitabine, 98.4% vs. BCG, 95.3%; p=0.848), 6-month (94.9% vs. 90.0%, respectively; p=0.793) and 1-year CR rates (84.2% vs. 83.3%, respectively; p=0.950). Also, there was no significant statistical difference in progression-free survival between the two groups (p=0.953). The occurrence rates of adverse events were similar between the groups (22.2% vs. 22.1%; p=0.989); however, the rate of Clavien-Dindo grade 2 or higher was significantly higher in the BCG group (1.6% vs. 16.3%, respectively; p<0.001). CONCLUSIONS: Intravesical gemcitabine demonstrated efficacy comparable to BCG therapy for the first year in patients with intermediate- and high-risk NMIBC. However, long-term follow-up studies are warranted.

Patient-centered development of a bladder cancer survivorship care plan.

PURPOSE: This study aimed to develop a patient-centered survivorship care plan (SCP) for US military Veteran bladder cancer (BC) survivors in accordance with the National Academy of Medicine recommendation that survivors receive an SCP at treatment completion. BC, which differentially impacts older men, is a costly and highly recurrent cancer associated with invasive procedures and long-term surveillance. Veteran BC survivors may face challenges navigating the patient-to-survivor transition due to their age and comorbidities. METHODS: We conducted 20 one-on-one qualitative interviews and 2 focus groups with Veteran BC survivors to understand their preferences for information and support to inform SCP development. Data were analyzed using rapid analysis. RESULTS: Participants voiced concerns about BC's impact on their psychosocial functioning and quality of life. They suggested information on BC recurrences, recommended surveillance schedules, long-term side effects and healthy living, and how and when to seek help if a medical problem arises should be included on the SCP to help manage expectations and access key resources during survivorship. Although participants had varying needs, many recommended including information on supportive resources (e.g., support groups, peer support programs) to manage cancer-related anxiety. Participants also suggested including Veteran-centered information (e.g., Veterans' BC risk factors). CONCLUSION: We developed an SCP to help Veteran BC survivors navigate the transition from patient to survivor. Adapting an SCP to address specific needs of the Veteran population was an important step in supporting Veteran BC survivors. Future research should evaluate the potential effectiveness of this SCP at improving Veterans' health outcomes and healthcare experiences.

The identification of a N6-methyladenosin-modifed immune pattern to predict immunotherapy response and survival in urothelial carcinoma.

BACKGROUND: Dysregulation of the immune system and N6-methyladenosine (m6A) contribute to immune therapy resistance and cancer progression in urothelial carcinoma (UC). This study aims to identify immune-related molecules, that are m6A-modified, and that are associated with tumor progression, poor prognosis, and immunotherapy response. METHODS: We identified prognostic immune genes (PIGs) using Cox analysis and random survival forest variable hunting algorithm (RSF-VH) on immune genes retrieved from the Immunology Database and Analysis Portal database (ImmPort). The RM2Target database and MeRIP-seq analysis, combined with a hypergeometric test, assessed m6A methylation in these PIGs. We analyzed the correlation between the immune pattern and prognosis, as well as their association with clinical factors in multiple datasets. Moreover, we explored the interplay between immune patterns, tumor immune cell infiltration, and m6A regulators. RESULTS: 28 PIGs were identified, of which the 10 most significant were termed methylated prognostic immune genes (MPIGs). These MPIGs were used to create an immune pattern score. Kaplan-Meier and Cox analyses indicated this pattern as an independent risk factor for UC. We observed significant associations between the immune pattern, tumor progression, and immune cell infiltration. Differential expression analysis showed correlations with m6A regulators expression. This immune pattern proved effective in predicting immunotherapy response in UC in real-world settings. CONCLUSION: The study identified a m6A-modified immune pattern in UC, offering prognostic and therapeutic response predictions. This emphasizes that immune genes may influence tumor immune status and progression through m6A modifications.

NIR-II light powered hydrogel nanomotor for intravesical instillation with enhanced bladder cancer therapy.

Intravesical instillation is the common therapeutic strategy for bladder cancer. Besides chemo drugs, nanoparticles are used as intravesical instillation reagents, offering appealing therapeutic approaches for bladder cancer treatment. Metal oxide nanoparticle based chemodynamic therapy (CDT) converts tumor intracellular hydrogen peroxide to ROS with cancer cell-specific toxicity, which makes it a promising approach for the intravesical instillation of bladder cancer. However, the limited penetration of nanoparticle based therapeutic agents into the mucosa layer of the bladder wall poses a great challenge for the clinical application of CDT in intravesical instillation. Herein, we developed a 1064 nm NIR-II light driven hydrogel nanomotor for the CDT for bladder cancer via intravesical instillation. The hydrogel nanomotor was synthesized via microfluidics, wrapped with a lipid bilayer, and encapsulates CuO2 nanoparticles as a CDT reagent and core-shell structured Fe3O4@Cu9S8 nanoparticles as a fuel reagent with asymmetric distribution in the nanomotor (LipGel-NM). An NIR-II light irradiation of 1064 nm drives the active motion of LipGel-NMs, thus facilitating their distribution in the bladder and deep penetration into the mucosa layer of the bladder wall. After FA-mediated endocytosis in bladder cancer cells, CuO2 is released from LipGel-NMs due to the acidic intracellular environment for CDT. The NIR-II light powered active motion of LipGel-NMs effectively enhances CDT, providing a promising strategy for bladder cancer therapy.

Efficacy of Intravesical Nadofaragene Firadenovec for Patients With Bacillus Calmette-Guérin-Unresponsive Nonmuscle-Invasive Bladder Cancer: 5-Year Follow-Up From a Phase 3 Trial.

PURPOSE: Nadofaragene firadenovec-vncg is a nonreplicating adenoviral vector-based gene therapy for bacillus Calmette-Guérin (BCG)-unresponsive carcinoma in situ (CIS) with/without high-grade Ta/T1. We report outcomes following 5 years of planned follow-up. MATERIALS AND METHODS: This open-label phase 3 trial (NCT02773849) enrolled patients with BCG-unresponsive nonmuscle-invasive bladder cancer in 2 cohorts: CIS ± Ta/T1 (CIS; n = 107) and Ta/T1 without CIS (Ta/T1 cohort; n = 50). Patients received 75 mL (3 × 1011 vp/mL) nadofaragene firadenovec intravesically once every 3 months with cystoscopy and cytology assessments, with continued treatment offered to those remaining high grade recurrence-free (HGRF). RESULTS: One hundred fifty-seven patients were enrolled from 33 US sites (n = 151 included in efficacy analyses). Median follow-up was 50.8 months (interquartile range 39.1-60.0), with 27% receiving ≥ 5 instillations and 7.6% receiving treatment for ≥ 57 months. Of patients with CIS 5.8% (95% CI 2.2-12.2) were HGRF at month 57, and 15% (95% CI 6.1-27.8) of patients with high-grade Ta/T1 were HGRF at month 57. Kaplan-Meier-estimated HGRF survival at 57 months was 13% (95% CI 6.9-21.5) and 33% (95% CI 19.5-46.6) in the CIS and Ta/T1 cohorts, respectively. Cystectomy-free survival at month 60 was 49% (95% CI 40.0-57.1): 43% (95% CI 32.2-53.7) in the CIS cohort and 59% (95% CI 43.1-71.4) in the Ta/T1 cohort. Overall survival at 60 months was 80% (71.0, 86.0): 76% (64.6-84.5) and 86% (70.9-93.5) in the CIS and Ta/T1 cohorts, respectively. Only 5 patients (4 with CIS and 1 with Ta/T1) experienced clinical progression to muscle-invasive disease. CONCLUSIONS: At 60 months, nadofaragene firadenovec-vncg allowed bladder preservation in nearly half of the patients and proved to be a safe option for BCG-unresponsive nonmuscle-invasive bladder cancer.