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Purpose: Immunotherapy with Bacillus Calmette-Guerin (BCG) instillation is recommended for high-risk, non–muscle invasive bladder cancer. Bacillus Calmette-Guerin is not effective in advanced tumors, and better alternatives are warranted. Immunostimulating gene therapy with adenoviral vectors expressing CD40 ligand (AdCD40L) has shown efficacy in tumor models. CD40 ligand stimulates systemic immunity and may be effective in local and invasive human disease.Experimental Design: Patients with invasive bladder cancer scheduled for cystectomy or patients with Ta tumors were enrolled in a phase I/IIa trial. Patients were treated with three cycles of intrabladder Clorpactin WCS-90 prewash, followed by AdCD40L instillation 1 week apart. Safety, gene transfer, immune effects, and antitumor responses were monitored.Results: All eight recruited patients were treated as scheduled, and therapy was well tolerated. The main adverse effect was transient local pain during prewash. Postoperatively, urinary tract infections and one case of late septicemia with elevated potassium were reported. No adverse events were ascribed to vector therapy. Gene transfer was detected in biopsies, and bladders were heavily infiltrated with T cells. The effector marker IFN-γ increased in biopsies, whereas levels of circulating T regulatory cells were reduced. Histologic evaluation indicated that AdCD40L therapy reduced the load of malignant cells.Conclusions: To our knowledge, this is the first report on immunogene therapy in bladder cancer and the first using AdCD40L in vivo. Local AdCD40L gene therapy was safe, boosted immune activation, and should be further evaluated as a single or an adjuvant therapy for urothelial malignancies. Clin Cancer Res; 16(12); 3279–87. ©2010 AACR.

Supplementary figure S8. Progression-free survival curve in patients with EAU high risk tumors of a combination of FGFR3 and GATA2 status. Blue: good status (hypomethylated GATA2 and mutated FGFR3), green; moderate status (either hypermethylated GATA2 and mutated FGFR3 or hypomethylated GATA2 and wild type FGFR3) or red: poor status (hypermethylated GATA2 and wild type FGFR3) compared to a progression free survival curse in all EAU high risk patients. Progressive disease is defined as progression to stage T2 or higher stage disease.

Supplementary Table S2. Cut-offs for dichotomization of the methylation marker results were chosen based on the desired sensitivity-specificity trade-off for progression to MIBC as visualized by the receiver-operating characteristic curve (ROC curve); the point on the curve closest to the upper left corner.

Purpose: Clinically useful molecular markers predicting the clinical course of patients diagnosed with non–muscle-invasive bladder cancer are needed to improve treatment outcome. Here, we validated four previously reported gene expression signatures for molecular diagnosis of disease stage and carcinoma in situ (CIS) and for predicting disease recurrence and progression.Experimental Design: We analyzed tumors from 404 patients diagnosed with bladder cancer in hospitals in Denmark, Sweden, England, Spain, and France using custom microarrays. Molecular classifications were compared with pathologic diagnosis and clinical outcome.Results: Classification of disease stage using a 52-gene classifier was found to be highly significantly correlated with pathologic stage (P < 0.001). Furthermore, the classifier added information regarding disease progression of Ta or T1 tumors (P < 0.001). The molecular 88-gene progression classifier was highly significantly correlated with progression-free survival (P < 0.001) and cancer-specific survival (P = 0.001). Multivariate Cox regression analysis showed the progression classifier to be an independently significant variable associated with disease progression after adjustment for age, sex, stage, grade, and treatment (hazard ratio, 2.3; P = 0.007). The diagnosis of CIS using a 68-gene classifier showed a highly significant correlation with histopathologic CIS diagnosis (odds ratio, 5.8; P < 0.001) in multivariate logistic regression analysis.Conclusion: This multicenter validation study confirms in an independent series the clinical utility of molecular classifiers to predict the outcome of patients initially diagnosed with non–muscle-invasive bladder cancer. This information may be useful to better guide patient treatment.

Supplementary Table 2 from Gene Expression Signatures Predict Outcome in Non–Muscle-Invasive Bladder Carcinoma: A Multicenter Validation Study

Purpose: The European Association of Urology (EAU) guidelines for non–muscle-invasive bladder cancer (NMIBC) recommend risk stratification based on clinicopathologic parameters. Our aim was to investigate the added value of biomarkers to improve risk stratification of NMIBC.Experimental Design: We prospectively included 1,239 patients in follow-up for NMIBC in six European countries. Fresh-frozen tumor samples were analyzed for GATA2, TBX2, TBX3, and ZIC4 methylation and FGFR3, TERT, PIK3CA, and RAS mutation status. Cox regression analyses identified markers that were significantly associated with progression to muscle-invasive disease. The progression incidence rate (PIR = rate of progression per 100 patient-years) was calculated for subgroups.Results: In our cohort, 276 patients had a low, 273 an intermediate, and 555 a high risk of tumor progression based on the EAU NMIBC guideline. Fifty-seven patients (4.6%) progressed to muscle-invasive disease. The limited number of progressors in this large cohort compared with older studies is likely due to improved treatment in the past two decades. Overall, wild-type FGFR3 and methylation of GATA2 and TBX3 were significantly associated with progression (HR = 0.34, 2.53, and 2.64, respectively). The PIR for EAU high-risk patients was 4.25. On the basis of FGFR3 mutation status and methylation of GATA2, this cohort could be reclassified into a good class (PIR = 0.86, 26.2% of patients), a moderate class (PIR = 4.32, 49.7%), and a poor class (PIR = 7.66, 24.0%).Conclusions: We conclude that the addition of selected biomarkers to the EAU risk stratification increases its accuracy and identifies a subset of NMIBC patients with a very high risk of progression. Clin Cancer Res; 24(7); 1586–93. ©2018 AACR.

Supplementary Figures S1-S2 from Gene Expression Signatures Predict Outcome in Non–Muscle-Invasive Bladder Carcinoma: A Multicenter Validation Study

Supplementary Data from AdCD40L Immunogene Therapy for Bladder Carcinoma—The First Phase I/IIa Trial

Supplementary Table 1 from Gene Expression Signatures Predict Outcome in Non–Muscle-Invasive Bladder Carcinoma: A Multicenter Validation Study

Introduction: We aimed to compare short term outcomes after robot assisted radical cystectomy (RARC) and open radical cystectomy (ORC) for urinary bladder cancer in a large population. Materials and methods: We included all patients without distant metastases who underwent either RARC or ORC with ileal conduit between 2011 and 2019 registered in the Bladder cancer data Base Sweden (BladderBaSe) 2.0. Primary outcome was unplanned readmissions within 90 days, and secondary out-comes within 90 days of surgery were reoperations, Clavien 3-5 complications, total days alive and out of hospital, and mortality. The analysis was carried out using multivariate regression models. Results: Out of 2905 patients, 832 were operated with RARC and 2073 with ORC. Robotic procedures were to a larger extent performed during later years, at high volume centers (47% vs 17%), more often for organ-confined disease (82% vs. 72%) and more frequently in patients with high socioeconomic status (26% vs. 21%). Patients operated with RARC were more commonly readmitted (29% vs. 25%). In multi -variable analysis RARC was associated with decreased risk of Clavien 3-5 complications (OR 0.58, 95% CI 0.47-0.72), reoperations (OR 0.53, 95% CI 0.39-0.71) and had more days alive and out of hospital (mean difference 3.7 days, 95% CI 2.4-5.0). Conclusion: This study illustrates the "real-world" effects of a gradual and nation-wide introduction of RARC. Patients operated with RARC had fewer major complications and reoperations but were more frequently readmitted compared to ORC. The observed differences were largely due to more wound related complications among patients treated with ORC.

Background Urothelial bladder cancer is most frequently diagnosed at the non-muscle-invasive stage (NMIBC). However, recurrences and interventions for intermediate and high-risk NMIBC patients impact quality of life. Biomarkers for patient stratification could help to avoid unnecessary interventions whilst indicating aggressive measures when required. Methods In this study, immuno-oncology focused, multiplexed proximity extension assays were utilised to analyse plasma (n=90) and urine (n=40) samples from 90 newly-diagnosed and treatment-naïve bladder cancer patients. Public single-cell and bulk RNA-sequencing data from patient tumour tissues and murine OH-BBN-induced urothelial carcinomas were also explored. Results Plasma from muscle-invasive, urothelial bladder cancer patients displayed higher levels of MMP7 (p=0.028) and CCL23 (p=0.03) compared to NMIBC patients, whereas urine displayed higher levels of CD27 (p=0.044) and CD40 (p=0.04) in the NMIBC group by two-sided Wilcoxon rank-sum tests. Random forest survival and multivariable regression analyses identified increased MMP12 plasma levels as an independent marker (ppConclusions The measurable levels of tumour-localised, immune-cell-derived MMP12 in blood suggest MMP12 as an important, tumour-microenvironment-related biomarker that could complement histopathology-based risk stratification and represent a pharmacological target in urothelial bladder cancer.

27-hydroxycholesterol (27HC), synthesized from cholesterol by the enzyme CYP27A1, differentially impacts estrogen receptor positive (ER+) breast cancer (BC) cell growth depending on estrogen levels. This study examined the association between CYP27A1 expression and prognosis in a cohort of 193 premenopausal patients with lymph node-negative primary BC with limited exposure to adjuvant systemic cancer treatments. In multivariable analyses among patients with ER+ tumors, high CYP27A1 protein and mRNA expressions were associated with four- and eight-fold reductions in the incidence of distant recurrence-free survival events: HRadj = 0.26, 95% CI = 0.07–0.93 and HRadj = 0.13, 95% CI = 0.03–0.60, respectively. In vitro studies revealed that 27HC treatment potently inhibited ER+ BC cell proliferation under lipid-depleted conditions regardless of estradiol levels, transcriptionally mediated through the downregulation of ER signaling with a concomitant upregulation of cholesterol export. Importantly, if validated, these results may have implications for adjuvant treatment decisions in premenopausal patients, especially when de-escalation of therapy is being considered.

Clinical utility of cisplatin based neoadjuvant chemotherapy (NAC) prior to radical cystectomy is limited because of lack of tools that can guide for a better patient selection. We aim to explore if a combination of biomarkers is superior to a single marker. Pretreatment tumor specimens and clinical data from two randomized trials including 250 patients with T2-T4 urothelial bladder cancer, were used. The information on the expressions on tumor tissue of four biomarkers; CCTα, emmprin, survivin, and BCL-2, detected by immunohistochemistry in our previous studies, was used. Cox proportional hazard models, including treatment-by-biomarker interaction terms, were used to assess the predictive value of the biomarkers for efficacy of NAC on overall survival. CCTα provided predictive information about the efficacy of NAC (interaction P=0.009). None of the other biomarkers provided statistically significant information additional to CCTα. The adjusted hazard ratio for NAC treated versus no-NAC was 0.42 (95% CI: 0.27-0.64) for patients with negative CCTα expression, when adding information about emmprin it decreased to 0.33 (95% CI: 0.19-0.56) for patients with both negative CCTα and emmprin. This corresponds to a decrease in number needed to treat from 4 to 3 patients. The combination of CCTα with survivin or BCL-2 yielded similar results. In a group of patients with muscle invasive bladder cancer a combination of two biomarkers might improve the possibility to identify patients most likely to benefit from the use of NAC. Further studies designed to have sufficient power to detect an interaction effect are needed.

Purpose: To investigate whether outpatient blue-light flexible cystoscopy could solve the diagnostic challenge of positive or suspicious urine cytology findings despite normal white-light flexible cystoscopy results and normal findings on computerized tomography urography, in patients investigated for urothelial cancer. Material and methods: In a multicentre study, a total of 70 examinations were performed with the use of blue-light flexible cystoscopy (photodynamic diagnosis) after intravesical instillation of the fluorescence agent hexaminolevulinate. The examination started with a conventional white-light flexible cystoscopy and then the settings were switched to use blue light. Suspicious lesions were biopsied. Afterwards, the patients were interviewed regarding their experience of the examinations. Results: Bladder cancer was diagnosed in 29 out of 70 (41%) cases, among them 14/29 (48%) had malignant lesions seen only in blue light. The majority had carcinoma in situ (21/29). Normal findings were seen in 41 cases that underwent BLFC. During the further course, malignancy of the bladder was detected in six cases (9%) and malignancy of the upper urinary tract was detected in one case (1%). The majority of patients (93%) preferred the blue-light flexible cystoscopy performed at the outpatient clinic instead of the transurethral resection under general anaesthesia. Conclusion: Blue-light flexible cystoscopy at the outpatient clinic may be a useful tool to solve unclear cases of a malignant or suspicious urinary cytology suggestive of bladder cancer. The procedure was well tolerated by the patients.

Purpose Predictive biomarkers are needed to aid the individualization of radiotherapy (RT) in breast cancer. Cancer-associated fibroblasts have been implicated in tumor radioresistance and can be identified by platelet-derived growth factor receptor-beta (PDGFRb). This study aims to analyze how PDGFRb expression affects RT benefit in a large randomized RT trial. Methods PDGFRb was assessed by immunohistochemistry on tissue microarrays from 989 tumors of the SweBCG91RT trial, which enrolled lymph node-negative, stage I/IIA breast cancer patients randomized to RT after breast-conserving surgery. Outcomes were analyzed at 10 years for ipsilateral breast tumor recurrence (IBTR) and any recurrence and 15 years for breast cancer specific death (BCSD). Results PDGFRb expression correlated with estrogen receptor negativity and younger age. An increased risk for any recurrence was noted in univariable analysis for the medium (HR 1.58, CI 95% 1.11–2.23, p = 0.011) or PDGFRb high group (1.49, 1.06–2.10, p = 0.021) compared to the low group. No differences in IBTR or BCSD risk were detected. RT benefit regarding IBTR risk was significant in the PDGFRb low (0.29, 0.12–0.67, p = 0.004) and medium (0.31, 0.16–0.59, p p = 0.110) in multivariable analysis. Likewise, risk reduction for any recurrence was less pronounced in the PDGFRb high group. No significant interaction between RT and PDGFRb-score could be detected. Conclusion A higher PDGFRb-score conferred an increased risk of any recurrence, which partly can be explained by its association with estrogen receptor negativity and young age. Reduced RT benefit was noted among patients with high PDGFRb, however without significant interaction.

Background: The genomic landscape of cancer is complex and includes mutations and copy number alterations (CNAs) that affect several cancer related pathways and drive tumor evolution. Non-muscle-invasive bladder cancers (NMIBC) are largely orphan for integrative genomic studies. Large studies are needed to delineate the genomic complexity and heterogeneity of NMIBC. Methods: A total of 438 patients with NMIBC were analyzed, 296 of which were part of the UROMOL cohort (PMID: 27321955). The median follow-up was 5 years. The progression rate was 13% (n= 56). Whole exome sequencing (WES) was performed on DNA from tumor (~150x) and matched germline samples to call somatic mutations. Additionally, shallow whole genome sequencing (sWGS; ~2x) was performed on DNA from 362 of the tumors to quantify CNAs. RNA-sequencing was available for 414 of the samples, and tumors were classified according to the UROMOL2021 transcriptomic classes. We identified significantly mutated genes by mutsigCV and significantly amplified or deleted regions by GISTIC2. Results: The median tumor mutation burden (TMB) was 3.7/Mb. TMB was not associated with progression (p=0.28). A total of 61 genes were significantly mutated in the cohort, the most frequent being FGFR3 (61%), KDM6A (44%) and KMT2D (38%). Mutations in EP300 and RHOB were significantly associated with an increased risk of progression after adjusting for grade and stage (p=0.040 and 0.044, respectively). Several mutations showed a strong transcriptomic class dependent occurrence: mutations in RB1, TP53, ERCC2 and ERBB2 were enriched in the aggressive class 2a, FGFR3 and STAG2 in class 1 and class 3, and KMT2C and KMT2D in class 3. Genome doubling was identified in 15% of the tumors. These tumors were enriched in the aggressive classes 2a and 2b and were associated with increased risk of progression (p=0.0049). In addition, we observed several significantly altered genomic regions, the most significant being deletions in 9p21.3 (CDKN2A & CDKN2B, 64%), 2q37.1 (GIGYF2 & EIF4E2, 28%) and amplification in 11q13.3 (CCND1, 9%). Class 2a tumors were enriched for genomic alterations in most of the significant regions. 9p21.3 was the only region with frequent homozygous losses (22%). High-level gains were prognostic of progression, independently of ploidy, stage and grade, for several regions, including 4p16.3 (FGFR3, p=0.00013), 17q23.2(TBX2, p=0.0004) and 8p11.23(ZNF703, p=0.011). In addition, we observed an enrichment of uniparental disomy in 4p16.3 (FGFR3, 8%). Conclusion: Here we investigated the landscape of DNA alterations in NMIBC in a large patient cohort of NMIBC samples with paired transcriptomic data and detailed clinical follow-up. We identified several novel genomic alterations; specifically, we showed that 15% of the tumors had genome doublings, and we identified a complex underlying copy number landscape of the region containing FGFR3. Citation Format: Frederik Prip, Philippe Lamy, Iver Nordentoft, Sia Viborg Lindskrog, Trine Strandgaard, Karin Birkenkamp-Demtröder, Gregers G. Hermann, Astrid C. Petersen, Veronika Bahlinger, Marc-Oliver Grimm, Marcus Horstmann, Karin Mogensen, Roman Nawroth, Ulrika Segersten, Danijel Sikic, Kim E. M van Kessel, Tobias Maurer, Tatjana Simic, Arndt Hartmann, Ellen C. C. Zwarthoff, Per-Uno Malmström, Torben Steiniche, Jørgen Bjerggaard Jensen, Núria Malats, Francisco X. Real, Lars Dyrskjøt. Comprehensive genomic characterization of early-stage bladder cancer from 438 patients by whole genome- and exome sequencing. [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 6075.

We investigated the importance of HGF, pMET and pAkt for the risk of ipsilateral breast tumour recurrence after adjuvant radiotherapy in primary breast cancer in tumours from the SweBCG91‐RT randomised trial. Low levels of HGF and pMet in the tumour cell cytoplasm and high levels of pAkt in the tumour cell nucleus are associated with a larger benefit from RT, indicating that these proteins may be used as biomarkers or treatment targets., Experimental studies suggest that hepatocyte growth factor (HGF) and its transmembrane tyrosine kinase receptor, Met, in part also relying on Akt kinase activity, mediate radioresistance. We investigated the importance of these biomarkers for the risk of ipsilateral breast tumour recurrence (IBTR) after adjuvant radiotherapy (RT) in primary breast cancer. HGF, phosphorylated Met (pMet) and phosphorylated Akt (pAkt) were evaluated immunohistochemically on tissue microarrays from 1004 patients in the SweBCG91‐RT trial, which randomly assigned patients to breast‐conserving therapy, with or without adjuvant RT. HGF was evaluated in the stroma (HGFstr); pMet in the membrane (pMetmem); HGF, pMet and pAkt in the cytoplasm (HGFcyt, pMetcyt, pAktcyt); and pAkt in the nucleus (pAktnuc). The prognostic and treatment predictive effects were evaluated to primary endpoint IBTR as first event during the first 5 years. Patients with tumours expressing low levels of HGFcyt and pMetcyt and high levels of pAktnuc derived a larger benefit from RT [hazard ratio (HR): 0.11 (0.037–0.30), 0.066 (0.016–0.28) and 0.094 (0.028–0.31), respectively] compared to patients with high expression of HGFcyt and pMetcyt, and low pAktnuc [HR: 0.36 (0.19–0.67), 0.35 (0.20–0.64) and 0.47 (0.32–0.71), respectively; interaction analyses: P = 0.052, 0.035 and 0.013, respectively]. These differences remained in multivariable analysis when adjusting for patient age, tumour size, histological grade, St Gallen subtype and systemic treatment (interaction analysis, P‐values: 0.085, 0.027, and 0.023, respectively). This study suggests that patients with immunohistochemically low HGFcyt, low pMetcyt and high pAktnuc may derive an increased benefit from RT after breast‐conserving surgery concerning the risk of developing IBTR.

PURPOSE: To study how patients with urinary bladder cancer (UBC) with previous or concomitant other primary cancers (OPC) were treated, and to investigate their prognosis.METHODS: Using nationwide ...

Purpose Radiation therapy (RT) after breast-conserving surgery reduces locoregional recurrences and improves survival but may cause late side effects. The main purpose of this paper was to investigate long-term side effects after whole breast RT in a randomized clinical trial initiated in 1991 and to report dose-volume data based on individual 3-dimensional treatment plans for organs at risk. Methods and Materials The trial included 1187 patients with T1-2 N0 breast cancer randomized to postoperative tangential whole breast RT or no further treatment. The prescription dose to the clinical target volume was 48 to 54 Gy. We present 20-year follow-up on survival, cause of death, morbidity, and later malignancies. For a cohort of patients (n = 157) with accessible computed tomography–based 3-dimensional treatment plans in Dicom-RT format, dose-volume descriptors for organs at risk were derived. In addition, these were compared with dose-volume data for a cohort of patients treated with contemporary RT techniques. Results The cumulative incidence of cardiac mortality was 12.4% in the control group and 13.0% in the RT group (P = .8). There was an increase in stroke mortality: 3.4% in the control group versus 6.7% in the RT group (P = .018). Incidences of contralateral breast cancer and lung cancer were similar between groups. The median Dmean (range) heart dose for left-sided treatments was 3.0 Gy (1.1-8.1), and the corresponding value for patients treated in 2017 was 1.5 Gy (0.4-6.0). Conclusions In this trial, serious late side effects of whole breast RT were limited and less than previously reported in large meta-analyses. We observed no increase in cardiac mortality in irradiated patients. Doses to the heart were a median Dmean of 3.0 Gy for left-sided RT. The observed increase in stroke mortality may partly be secondary to cardiac side effects, complications to anticoagulant treatment, or to chance, rather than a direct side effect of tangential whole breast irradiation.

Background: Although preclinical studies have indicated that stroma cells can modulate the radiosensitivity of tumor cells, non-leukocytic stroma cells have not yet been explored as potential predictive markers for radiotherapy (RT) through systematic analyses of clinical samples. Platelet-derived growth factor receptor beta (PDGFRb) is a key regulator of fibroblasts, pericytes and smooth muscle cells. A high expression of PDGFRb in tumor-associated stroma has previously been associated with worse recurrence free and breast cancer specific survival as well as reduced tamoxifen sensitivity in invasive breast cancer. Purpose: To analyze the prognostic and predictive impact of stromal PDGFRb regarding ipsilateral breast tumor recurrence (IBTR) and any recurrence in a large randomized RT trial. Methods: Stromal PDGFRb expression was assessed by immunohistochemical staining of tissue microarrays (TMAs) with 958 tumors from the SweBCG91RT trial (Swedish Breast Cancer Group 91 Radiotherapy). The trial enrolled node-negative, stage I or IIA breast cancer patients, who were randomized to postoperative RT or not after breast conserving surgery. Stained TMAs were evaluated by two independent raters for PDGFRb staining intensity (0/negative; 1/low; 2/moderate; 3/high) and positive stroma fraction (0/0%; 1/1-10%; 2/11-50; 3/51-75%; 4/76-100%). The evaluations of both raters were averaged and the product between the two scoring metrics was formed. For final analysis the data was split in tertiles, as predefined, and referred to as PDGFRb low, medium or high score group. Outcomes were analyzed at 10 years of follow-up. Results: A high PDGFRb score was correlated with ER negativity (p=0.003,), young age (p With regard to prognosis, a significantly increased risk for any recurrence was detected in univariable analysis for patients with a medium (HR 1.57, CI95% 1.11-2.23, p=0.011) or high PDGFRb score (HR 1.49, CI95% 1.06-2.10, p=0.021) compared to the PDGFRb low score group. The significance remained for the PDGFRb medium (HR 1.47, CI95% 1.03-2.11, p=0.034) but not the PDGFRb high score group (HR 1.32, CI95% 0.93-1.88, p=0.125) in a multivariable analysis including grade, age and RT. The benefit of RT regarding the risk of IBTR was significant in the PDGFRb low (HR 0.28, CI95% 0.12-0.65, p=0.003) and medium (HR 0.32, CI95% 0.16-0.61, p=0.001) score groups but not in the PDGFRb high (HR 0.61, CI95% 0.35-1.07, p=0.086) score group in a multivariable analysis including grade and age. The risk reduction from RT regarding any recurrence was also less pronounced in the PDGFRb high (HR 0.73, CI95% 0.48-1.12, p=0.149) score group compared to the PDGFRb low (HR 0.57, CI95 0.32-1.03, p=0.065) and medium (HR 0.46, CI95% 0.28-0.75, p=0.002) score groups in multivariable analysis. No significant interaction between RT and PDGFRb expression could however be detected for IBTR (p=0.14) or any recurrence (p=0.33). Conclusion: A higher PDGFRb score conferred a negative prognosis with increased risk of any recurrence in univariable analyses, which in part can be explained by its association with ER negativity, young age and grade. A predictive effect of RT benefit from PDGFRb could not be confirmed although a tendency toward less benefit was observed in the patient group with the highest PDGFRb score. Citation Format: Carina Strell, Reidunn J Edelmann, Axel Stenmark Tullberg, Per Malmström, Mårten Fernö, Arne Östman, Erik Holmberg, Per Karlsson. Prognostic and predictive impact of stroma cells defined by platelet derived growth factor receptor beta in early breast cancer: Results from the randomized SweBGR91RT trial [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P3-08-08.

Background: Experimental evidence suggests that pathogenicity caused by cholesterol in breast cancer is partly elicited by 27-hydroxycholesterol (27HC), a primary cholesterol metabolite produced by the sterol 27-hydroxylase cytochrome p450 27A1 (CYP27A1). Targeting CYP27A1 abrogated the adverse effects of cholesterol and 27HC in preclinical studies. The prognostic relevance of CYP27A1 expression in patients with breast cancer is however not clearly defined. Previous studies suggest that the intratumoral expression of CYP27A1 might have diverse effects on prognosis depending on menopausal status. The aim of this study was to comprehensively evaluate the impact of both CYP27A1 protein and mRNA expression on tumor pathobiology and prognosis in pre-menopausal, node-negative breast cancer. Materials and Methods: CYP27A1 expression at the protein and transcript level was evaluated in a prospective cohort of 237 premenopausal women with node-negative, primary invasive breast cancer. Protein and mRNA expression were assessed by immunohistochemistry (IHC) and in situ hybridization technique-RNAscope, respectively. Associations of CYP27A1 expression with tumor pathological features and prognosis were assessed with chi-square test and Cox regression models, respectively. Multivariable models were adjusted for age, hormone receptor status, Ki67, and histological grade. Results: CYP27A1 protein was overexpressed in 28% and CYP27A1 mRNA in 23% of the tumors. The concordance between CYP27A1 protein and mRNA expression was 77.8% (rs=0.430). Among the discordant cases, 13.9% were only positive by IHC. Elevated expression of both protein and mRNA was significantly associated with aggressive tumor pathological features like high histological grade and high Ki67 expression (P Conclusion: These results indicate that high intratumoral expression of CYP27A1 is a strong indicator for an extensive recurrence-free interval and overall survival in premenopausal women with ER positive, lymph node-negative breast cancer. CYP27A1 transcript expression may serve as an even more sensitive prognostic biomarker. Citation Format: Maria Inasu, Pär Ola Bendahl, Mårten Fernö, Per Malmström, Signe Borgquist, Siker Kimbung. High CYP27A1 expression at protein and transcript level predicts favorable outcome in node negative, premenopausal breast cancer [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P3-08-21.

A 71-yr-old man was transferred to our institution with multiple and recurrent high-grade pTa bladder cancer 26 mo after an initial presentation of multiple and large pTa low-grade tumors and concomitant carcinoma in situ, treated with transurethral resection plus 6-mo postoperative mitomycin C. This case discusses several treatment options in the absence of bacillus Calmette-Guerin (BCG). Immediate radical cystectomy is an option with excellent survival, since there is a substantial risk of understaging and disease progression; however, this results in overtreatment in similar to 50% of these patients. Therefore, a conservative approach could be intravesical combination therapy such as gemcitabine/docetaxel or epirubicin/interferon. In addition, device-assisted intravesical therapy is becoming an option to consider. Finally, patients could be included in trials such as immunotherapy trials. Patient summary: This 71-yr-old patient was diagnosed with recurrent, moderately severe noninvasive bladder tumors, which were removed. The recommended additional therapy, intravesical bacillus Calmette-Guerin (BCG) instillations, was not available. Both the pros and the cons of radical surgery (bladder removal) and a more conservative approach (other intravesical treatments) are discussed. (c) 2019 The Authors. Published by Elsevier B.V. on behalf of European Association of Urology. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).

OBJECTIVE: The most common form of urinary bladder cancer is the low and intermediate risk categories of stage Ta. This patient group has a high recurrence rate, but progression is rare. The aim of this study was to investigate recurrence and survival in a large population-based setting, with respect to possible prognostic factors and during different time periods. PATIENTS AND METHODS: BladderBaSe is a database which links information from the Swedish National Register of Urinary Bladder Cancer with national healthcare and demographic registers. Between 1997 and 2014, 16,599 were diagnosed with low and intermediate risk of Ta cancer in Sweden. The times to recurrence and cancer-specific death were analysed concerning the differences in age, gender, grade, region and hospital type. For temporal analysis, we divided the material into 6-year periods. RESULTS: The mean age was 70 years and 75% were males. Low risk according to grade constituted 56%, whilst 44% had intermediate risk. With a median follow-up time of 63 months the recurrence rates were 47% and 59% for the respective categories and overall 52%. The rate was similar between the first two time periods, but became substantially lower in the most recent period. Five percent of patients died of the disease and risk category was the main prognostic variable. CONCLUSIONS: The risk of recurrence decreased in the last time period. Risk category based on grade was the most important prognostic indicator for outcome.

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

Alexander Schultz needs little introduction to readers of Scandinavian Journal of Urology. One of the leading Scandinavian urologists of his generation, Alexander was a constant standard bearer for...

PURPOSE Most patients with early-stage breast cancer are treated with adjuvant radiotherapy (RT) after breast-conserving surgery (BCS) to prevent locoregional recurrence (LRR). However, no genomic tools are used currently to select the optimal RT strategy. METHODS We profiled the transcriptome of primary tumors on a clinical grade assay from the SweBCG91-RT trial, in which patients with node-negative breast cancer were randomly assigned to either whole-breast RT after BCS or no RT. We derived a new classifier, Adjuvant Radiotherapy Intensification Classifier (ARTIC), comprising 27 genes and patient age, in three publicly available cohorts, then independently validated ARTIC for LRR in 748 patients in SweBCG91-RT. We also compared previously published genomic signatures for ability to predict benefit from RT in SweBCG91-RT. RESULTS ARTIC was highly prognostic for LRR in patients treated with RT (hazard ratio [HR], 3.4; 95% CI, 2.0 to 5.9; P < .001) and predictive of RT benefit ( Pinteraction = .005). Patients with low ARTIC scores had a large benefit from RT (HR, 0.33 [95% CI, 0.21 to 0.52], P < .001; 10-year cumulative incidence of LRR, 6% v 21%), whereas those with high ARTIC scores benefited less from RT (HR, 0.73 [95% CI, 0.44 to 1.2], P = .23; 10-year cumulative incidence of LRR, 25% v 32%). In contrast, none of the eight previously published signatures were predictive of benefit from RT in SweBCG91-RT. CONCLUSION ARTIC identified women with a substantial benefit from RT as well as women with a particularly elevated LRR risk in whom whole-breast RT was not sufficiently effective and, thus, in whom intensified treatment strategies such as tumor-bed boost, and possibly regional nodal RT, should be considered. To our knowledge, ARTIC is the first classifier validated as predictive of benefit from RT in a phase III clinical trial with patients randomly assigned to receive or not receive RT.

Purpose: To investigate the clinical management and outcome of patients with muscle-invasive bladder cancer with clinical lymph node involvement, using longitudinal nationwide population-based data ...

Objective: To evaluate trends in bladder cancer incidence, survival and mortality in Sweden from 1997-2016.Patients and methods: The Swedish National Registry of Urinary Bladder Cancer is a nation- ...

OBJECTIVE: To describe the management of patients with hematuria in the Nordic countries in relation to bladder cancer epidemiology, especially in the context of introducing fast track pathways with the aim of proposing a common guideline.MATERIALS AND METHODS: Epidemiological data on bladder cancer from each country, and the combined cancer registry, Nordcan, were analyzed. The evolution of the different national recommendations and the introduction of fast track pathways were assessed. Patients' demographics, type of hematuria and cancer detection rates were analysed if available.RESULTS: The crude incidence of bladder cancer has increased substantially since the 1960s, while the age standardized incidence has been stable during recent decades. The relative survival has increased in all countries, while the mortality has been stable. For those with microscopic hematuria there has been a clear trend towards less rigorous investigations. In the fast track pathways, introduced in three of five countries, about one in five patients with macroscopic hematuria had a cancer diagnosis. Data show that time to diagnosis has been reduced.CONCLUSIONS: The number of patients with bladder cancer is increasing in the Nordic region. The introduction of fast track pathways has been important in improving the management of patients with suspicion of the disease. Our recommendation is to focus on macroscopic hematuria in the fast track pathways. Microhematuria without any symptoms should not be an indication for cystoscopy. However, urinary tract symptoms accompanied by microhematuria can still be investigated according to respective guidelines but not necessarily within fast track pathways.

BackgroundPrognostic factors in breast cancer are often measured on a continuous scale, but categorized for clinical decision-making. The primary aim of this study was to evaluate if accounting for continuous non-linear effects of the three factors age at diagnosis, tumor size, and number of positive lymph nodes improves prognostication. These factors will most likely be included in the management of breast cancer patients also in the future, after an expected implementation of gene expression profiling for adjuvant treatment decision-making.MethodsFour thousand four hundred forty seven and 1132 women with primary breast cancer constituted the derivation and validation set, respectively. Potential non-linear effects on the log hazard of distant recurrences of the three factors were evaluated during 10years of follow-up. Cox-models of successively increasing complexity: dichotomized predictors, predictors categorized into three or four groups, and predictors transformed using fractional polynomials (FPs) or restricted cubic splines (RCS), were used. Predictive performance was evaluated by Harrells C-index.ResultsUsing FP-transformations, non-linear effects were detected for tumor size and number of positive lymph nodes in univariable analyses. For age, non-linear transformations did, however, not improve the model fit significantly compared to the linear identity transformation. As expected, the C-index increased with increasing model complexity for multivariable models including the three factors. By allowing more than one cut-point per factor, the C-index increased from 0.628 to 0.674. The additional gain, as measured by the C-index, when using FP- or RCS-transformations was modest (0.695 and 0.696, respectively). The corresponding C-indices for these four models in the validation set, based on the same transformations and parameter estimates from the derivation set, were 0.675, 0.700, 0.706, and 0.701.ConclusionsCategorization of each factor into three to four groups was found to improve prognostication compared to dichotomization. The additional gain by allowing continuous non-linear effects modeled by FPs or RCS was modest. However, the continuous nature of these transformations has the advantage of making it possible to form risk groups of any size. Funding Agencies|Swedish Cancer Society; Swedish Research Council; Gunnar Nilsson Cancer Foundation; Swedish Breast Cancer Association; Swedish Cancer and Allergy Foundation; Mrs. Berta Kamprad Foundation; Anna and Edwin Bergers Foundation; Skane County Councils Research and Development Foundation; Governmental Funding of Clinical Research within the National Health Service

Investigate symptoms and how they affect daily life in patients with Non-Muscle Invasive Bladder Cancer (NMIBC) treated with Bacillus Calmette-Guérin (BCG) instillations.Patients treated with BCG were included. After an initial transurethral resection (TURB) followed by a second-look resection, the patients were given an induction course with BCG for 6 weeks followed by maintenance therapy for 2 years. The patients answered a questionnaire before, during and after the treatment. The questionnaire contained questions about specific symptoms combined with bother questions on how each symptom affected patients' life.In total, 113 of 116 patients responded to the first questionnaire. Thirty per cent of all patients were bothered by disease-specific symptoms before the start of BCG. Few patients reported fever, haematuria, illness or urinary tract symptoms. No difference in symptoms was found between patients with or without concomitant CIS (carcinoma in situ). Patients younger than 65 years of age reported a greater worry about the symptom burden in the future than those who were older. Patients younger than 65 years reported a decreased level of mental well-being.Patients with bladder cancer T1G2-G3 had disease-specific symptoms present already before the start of the BCG. The burden of symptoms was reduced over time and showed that the bladder might recover. BCG instillations had side-effects that negatively affected the patient's well-being. It is important to record the patients' baseline bladder and voiding status before as well as during the BCG-instillation period in order to understand symptoms caused by the treatment.

The molecular landscape in non-muscle-invasive bladder cancer (NMIBC) is characterized by large biological heterogeneity with variable clinical outcomes. Here, we perform an integrative multi-omics analysis of patients diagnosed with NMIBC (n = 834). Transcriptomic analysis identifies four classes (1, 2a, 2b and 3) reflecting tumor biology and disease aggressiveness. Both transcriptome-based subtyping and the level of chromosomal instability provide independent prognostic value beyond established prognostic clinicopathological parameters. High chromosomal instability, p53-pathway disruption and APOBEC-related mutations are significantly associated with transcriptomic class 2a and poor outcome. RNA-derived immune cell infiltration is associated with chromosomally unstable tumors and enriched in class 2b. Spatial proteomics analysis confirms the higher infiltration of class 2b tumors and demonstrates an association between higher immune cell infiltration and lower recurrence rates. Finally, the independent prognostic value of the transcriptomic classes is documented in 1228 validation samples using a single sample classification tool. The classifier provides a framework for biomarker discovery and for optimizing treatment and surveillance in next-generation clinical trials., Multiple molecular profiling methods are required to study urothelial non-muscle-invasive bladder cancer (NMIBC) due to its heterogeneity. Here the authors integrate multi-omics data of 834 NMIBC patients, identifying a molecular subgroup associated with multiple alterations and worse outcomes.

Immune cells in the microenvironment shape tumor development and progression. The prognostic value of T-cell-based immune scores exceeds those of clinical parameters in colon cancer, but reflects only a part of the anti-tumor immune response. Here, we assessed 15 distinct immune cell classes and identified a simple prognostic signature based on pro- and anti-tumoral immune cells in the tumor microenvironment. The ratio of cytotoxic lymphocytes to tumor supportive macrophages predicted survival better than the state-of-art immune score in colon cancer and had the highest relative contribution to survival prediction when compared to established clinical parameters. This signature was prognostic also in other cancers with high mutation burden, such as those of the lung, bladder, esophagus, and melanomas, supporting broad clinical applicability.One Sentence SummaryThe CD8/M2 ratio in tumor tissue defines prognosis in immunogenic cancers

Background The development of a reactive tumour stroma is a hallmark of tumour progression and pronounced tumour stroma is generally considered to be associated with clinical aggressiveness. The variability between tumour types regarding stroma fraction, and its prognosis associations, have not been systematically analysed. Methods Using an objective machine-learning method we quantified the tumour stroma in 16 solid cancer types from 2732 patients, representing retrospective tissue collections of surgically resected primary tumours. Image analysis performed tissue segmentation into stromal and epithelial compartment based on pan-cytokeratin staining and autofluorescence patterns. Findings The stroma fraction was highly variable within and across the tumour types, with kidney cancer showing the lowest and pancreato-biliary type periampullary cancer showing the highest stroma proportion (median 19% and 73% respectively). Adjusted Cox regression models revealed both positive (pancreato-biliary type periampullary cancer and oestrogen negative breast cancer, HR(95%CI)=0.56(0.34-0.92) and HR(95%CI)=0.41(0.17-0.98) respectively) and negative (intestinal type periampullary cancer, HR(95%CI)=3.59(1.49-8.62)) associations of the tumour stroma fraction with survival. Interpretation Our study provides an objective quantification of the tumour stroma fraction across major types of solid cancer. Findings strongly argue against the commonly promoted view of a general associations between high stroma abundance and poor prognosis. The results also suggest that full exploitation of the prognostic potential of tumour stroma requires analyses that go beyond determination of stroma abundance. Funding The Swedish Cancer Society, The Lions Cancer Foundation Uppsala, The Swedish Government Grant for Clinical Research, The Mrs Berta Kamprad Foundation, Sweden, Sellanders foundation, P.O.Zetterling Foundation, and The Sjoberg Foundation, Sweden.

BACKGROUND: Routine VTE prophylaxis within 30 days of radical cystectomy (RC) for urinary bladder cancer (UBC) is used to protect from venous thromboembolism (VTE). However, randomized studies and nationwide population-based studies are lacking. OBJECTIVE: To study VTE and risk factors for VTE in muscle-invasive UBC in a nationwide population-based series, with a focus on the association with RC with and without chemotherapy. MATERIALS AND METHODS: We studied all patients with clinical stage T2-T4 UBC diagnosed 1997 to 2014 in the Bladder Cancer Data Base Sweden (BladderBaSe). Previous VTE events and risk factors for VTE were registered from 1987. Cox regression analyses and Kaplan-Meier curves were performed to study risk factors for VTE and cumulative incidence of VTE. RESULTS: In 9720 patients (71% males) with a median age of 74 years 546 (5.6%) had VTE after diagnosis. In Cox analyses controlling for patients and tumour characteristics, and risk factors for VTE, VTE after diagnosis and first treatment date were associated with chemotherapy with or without RC. Cumulative incidence of VTE increased during 24 months after diagnosis and first treatment date. VTE were less common in patients with previous cardiovascular disease. CONCLUSION: VTE was commonly observed after 30 days from diagnosis and from first treatment date in patients with T2-T4 UBC, particularly after chemotherapy. The findings suggest that long-term intervention studies of benefit and possible harms of VTE prophylaxis after UBC should be undertaken. Funding Agencies|Swedish Cancer SocietySwedish Cancer Society [CAN 2016/470, CAN 2017/278]