OBJECTIVES:To investigate the cumulative incidence proportion of disseminated or local Bacillus Calmette-Guérin (BCG) infections after adjuvant BCG instillations in patients with non-muscle-invasive bladder cancer (NMIBC). PATIENTS AND METHODS:We analysed the timing and occurrence of BCG infections and absolute and relative risk in relation to patient characteristics available in the Swedish nationwide database 'BladderBaSe 2.0'. The cumulative incidence proportion of a BCG infection was indicated by a reported diagnosis of tuberculosis (TB) in the patient registry or filing a prescription for tuberculostatic drugs. RESULTS:The cumulative incidence proportion was 1.1% at the 5-year follow-up in 5033 patients exposed to adjuvant BCG instillations. The incidence rate was highest during the first 2 years after start of BCG instillations. Women had a lower risk than men (hazard ratio 0.23, 95% confidence interval 0.07-0.74). Age and calendar time at diagnosis, comorbidity, tumour risk group, previous medication with corticosteroids, immunosuppressive drugs, or time between transurethral resection of the bladder tumour and commencing the adjuvant BCG instillation were not associated with risk. CONCLUSIONS:These data further supports that the overall risk of a BCG infection after BCG-instillation treatment for NMIBC is low. The great majority of infections occur in the first 2 years, calling for an awareness of the diverse symptoms of BCG infection during this period. We provide evidence for male sex as a risk factor; however, the statistical precision is low and with a risk of selection bias, making it difficult to rule out the other suggested risk factors without further studies with different approaches.

The most significant factors for a higher risk of residual disease at re-TUR in T1G3 patients are multifocal tumors and tumors more than 3 cm. Patients with concomitant CIS and those without muscle in the specimen also have a higher risk of residual disease.

Abstract Bladder cancer continues to be a healthcare challenge and for many years there have been very few new therapies providing benefit for patients either with non-muscle or muscle invasive bladder cancer. The molecular profiling of bladder cancer has evolved over the last years and different classifications systems have arisen; however, there is still a lack of knowledge on the prognostic or predictive therapeutic value based on these stratification methods. The clinical heterogeneity of the disease impacts the use of the molecular tools; in addition to that, the heterogeneity in the tumor microenvironment may hamper therapeutic strategies based on monotherapy approaches. As check-point blocking antibodies have entered the clinic for the treatment of muscle-invasive bladder cancer, and as novel therapeutic approaches are gaining interest in the non-muscle invasive treatment schedule, we may have novel therapeutic options for these patients ahead of us. But to be successful we also need to understand how to combine these therapies to improve overall survival and to impact relapse and progression of the disease. To enable an improved understanding of the landscape of the bladder cancer therapies we have developed a novel model based on a transgenic mouse with overexpression of Hgf and a dysregulated cell cycle. When exposing this transgenic strain to OH-BBN in drinking water these mice develop a non-muscle invasive bladder cancer that differ from a wt C57BL/6 mouse exposed to OH-BBN. The tumors are of squamous differentiated Tis/T1 type after a course of 10 weeks OH-BBN exposure. After another 5-10 weeks there is a noted muscle invasive squamous-like bladder cancer. The tumors show a strong inflammatory reaction. The muscle-invasive stage also leads to a marked change in the urine proteomic profile (as measured by the Proximity Extension Assay [PEA 92]). We have profiled both the micro and macro-hematuria and the time-course proteomic changes along with the pathology in this model. In addition, we recently performed single-cell sequencing on tumors from the non-muscle and muscle invasive model to dissect the molecular profile, immune landscape along with the heterogeneity. Thus, we can compare this novel model to the frequently used syngeneic MB49 tumor model. We are also currently assessing anti-PD1 therapy in the novel semi-spontaneous tumor model to evaluate the responsiveness to check-point blocking therapy. Ahead we will process the single cell sequencing data and dissect out what type of targeted therapies along with immunotherapies that should be combined, to improve the understanding of how to resolve the progression from a non-muscle invasive to a muscle-invasive deadly disease, with a hope to translate this into a clinically relevant therapeutic strategy. Citation Format: Iliana Kerzeli, Ramy Elgendy, Ivan Stepanek, Milena Doroszko, Aikaterini Chourlia, Sven Nelander, Per-Uno Malmstrom, Ulrika Segersten, Anca Dragomir, Martin Lord, Sara Mangsbo. Development and characterization of a novel semi-spontaneous bladder cancer model by pathological evaluation, single cell sequencing and proteomic profiling of urine and serum [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1610.

Cancer of the urinary bladder (UBC) primarily derives from the urothelium, encompassing the inner surface of the bladder. Tumors infiltrating the detrusor muscle are categorized as muscle-invasive bladder cancer (MIBC) and constitutes 20-25% of all newly diagnosed UBC. These tumors are more likely to spread to peripheral organs and lymph nodes, compared to the more prevalent (75-80%) non-muscle invasive bladder cancer subtype (NMIBC). To this end, cystoscopy and urine cytology are the current standards for diagnosing and surveying UBC, which provides limited information regarding their immunological profile. We performed a targeted human protein biomarker analysis from collected plasma and urine from a well-categorized UBC patient cohort. In short, 92 immuno-oncology related proteins were investigated by a novel Proximity Extension Assay multiplex technology (Olink). The assayed proteins depicted a heterogenic immune profile. Neither PCA or unsupervised hierarchical clustering identified distinct groups associated with clinical features. However, a direct comparison between non-invasive and invasive cases resulted in 4 differentially expressed proteins (p<0.05). MIBC patients had lower urine levels of the TNF-receptor superfamily members CD27 and CD40, and a systemic increase of matrix metallopeptidase 7 (MMP7) and C-C motif chemokine ligand 23 (CCL23). We further developed and trained a random forest machine learning classifier on our plasma samples. It consistently ranked CCL23 and MMP7 as the most important markers for invasiveness. When extended to survival analysis, we moreover found MMP levels to best predict a dismal survival status with MMP12 being the highest ranked variable. A finding supported by MMP12 being the most significant protein in individual cox-regression survival analyses. This study highlights the heterogenic nature of the immunogenic landscape in UBC and its potential role in immunotherapy treatment responses. Our findings indicate that systemic MMP levels should be further explored as a potential liquid biopsy marker for patient stratification purposes. The role of MMPs in UBC may be linked to an invasive tumor characteristic, as shown by previous UBC studies where high levels of MMPs correlated to reduced overall survival.

Purpose To evaluate the oncological impact of postponing radical cystectomy (RC) to allow further conservative therapies prior to progression in a large multicentre retrospective cohort of T1-HG/G3 patients initially treated with BCG. Methods According to the time of RC, the population was divided into 3 groups: patients who did not progress to muscle-invasive disease, patients who progressed before radical cystectomy and patients who experienced progression at the time of radical cystectomy. Clinical and pathological outcomes were compared across the three groups. Results Of 2451 patients, 509 (20.8%) underwent RC. Patients with tumors > 3 cm or with CIS had earlier cystectomies (HR = 1.79, p = 0.001 and HR = 1.53, p = 0.02, respectively). Patients with tumors > 3 cm, multiple tumors or CIS had earlier T3/T4 or N + cystectomies. In patients who progressed, the timing of cystectomy did not affect the risk of T3/T4 or N + disease at RC. Patients with T3/T4 or N + disease at RC had a shorter disease-specific survival (HR = 4.38, p < 0.001), as did patients with CIS at cystectomy (HR = 2.39, p < 0.001). Patients who progressed prior to cystectomy had a shorter disease-specific survival than patients for whom progression was only detected at cystectomy (HR = 0.58, p = 0.024) Conclusions Patients treated with RC before experiencing progression to muscle-invasive disease harbor better oncological and survival outcomes compared to those who progressed before RC and to those upstaged at surgery. Tumor size and concomitant CIS at diagnosis are the main predictors of surgical treatment while tumor size, CIS and tumor multiplicity are associated with extravesical disease at surgery.

PurposeThe goals of transurethral resection of a bladder tumor (TUR) are to completely resect the lesions and to make a correct diagnosis to adequately stage and treat the patient. Persistent disease after TUR is not uncommon and is why re-TUR is recommended in T1G3 patients. When there is T1 tumor in the re-TUR specimen, very high risks of progression (82%) have been reported. We analyze the risks of recurrence, progression to muscle-invasive disease and cancer-specific mortality (CSM) according to tumor stage at re-TUR in T1G3 patients treated with BCG.MethodsIn our retrospective cohort of 2451 T1G3 patients, 934 patients (38.1%) underwent re-TUR. 667 patients had residual disease (71.4%): Ta in 378 (40.5%), T1 in 289 (30.9%) patients. Times to recurrence, progression and CSM in the three groups were estimated using cumulative incidence functions and compared using the Cox regression model.ResultsDuring a median follow-up of 5.2years, 512 patients recurred. The recurrence rate was significantly higher in patients with a T1 at re-TUR (P<0.001). Progression rates differed according to the pathology at re-TUR, 25.3% in T1, 14.6% in Ta and 14.2% in case of no residual tumor (P<0.001). Similar trends were seen in both patients with and without muscle in the original TUR specimen.ConclusionsPatients with T1G3 tumors and no residual disease or Ta at re-TUR have better recurrence, progression and CSM rates than previously reported, with a CSM rate of 13.1 and a 25.3% progression rate in re-TUR T1 disease.

BACKGROUND:The oncoprotein-18/stathmin 1 (STMN1), involved in cell progression and migration, is associated with clinical outcome in breast cancer. Here we aim to investigate its clinical significance in urinary bladder cancer and its possibilities as a therapeutic target.METHODS:Immunohistochemical analyses of STMN1 protein expression were performed in three patient cohorts: cohort I (n=115 Ta, n=115 T1, n=112 T2-4 stages), cohort II, based on randomised controlled trials (n=239 T1-T4), and cohort III of primary tumour/matched metastasis (n=90 T1-T4). The effects of STMN1 on cell proliferation and migration were evaluated in the urinary bladder cancer cell line, T24, by inhibiting STMN1-cellular expression using siRNA.RESULTS:In cohort I, high STMN1 expression correlated to shorter disease-specific survival hazard ratio (HR)=2.04 (95% confidence interval (CI) 1.13-3.68; P=0.02), elevated p53- (P<0.001) and Ki67-protein levels (P<0.001). The survival result was validated in cohort II: HR=1.76 (95% CI 1.04-2.99; P=0.03). In the metastatic bladder cancer material, 70% of the patients were STMN1-positive in both the primary tumour and matched metastases. In vitro, the growth and migration of the T24 cells were significantly reduced (P<0.01, P<0.0001, respectively), when transfecting the cells with STMN1-siRNA.CONCLUSIONS:STMN1 protein expression has prognostic significance but is primarily a potential treatment target in urinary bladder cancer.