Regional transcriptomic profiling reveals immune system enrichment in nonfailing atria and all chambers of the failing human heart.

The different chambers of the human heart demonstrate regional physiological traits and may be differentially affected during pathological remodeling, resulting in heart failure. Few previous studies, however, have characterized the different chambers at a transcriptomic level. We, therefore, conducted whole tissue RNA sequencing and gene set enrichment analysis of biopsies collected from the four chambers of adult failing (n = 8) and nonfailing (n = 11) human hearts. Atria and ventricles demonstrated distinct transcriptional patterns. When compared with nonfailing ventricles, the transcriptional pattern of nonfailing atria was enriched for many gene sets associated with cardiogenesis, the immune system and bone morphogenetic protein (BMP), transforming growth factor-ß (TGF-ß), MAPK/JNK, and Wnt signaling. Differences between failing and nonfailing hearts were also determined. The transcriptional pattern of failing atria was distinct compared with that of nonfailing atria and enriched for gene sets associated with the innate and adaptive immune system, TGF-ß/SMAD signaling, and changes in endothelial, smooth muscle cell, and cardiomyocyte physiology. Failing ventricles were also enriched for gene sets associated with the immune system. Based on the transcriptomic patterns, upstream regulators associated with heart failure were identified. These included many immune response factors predicted to be similarly activated for all chambers of failing hearts. In summary, the heart chambers demonstrate distinct transcriptional patterns that differ between failing and nonfailing hearts. Immune system signaling may be a hallmark of all four heart chambers in failing hearts and could constitute a novel therapeutic target.NEW & NOTEWORTHY The transcriptomic patterns of the four heart chambers were characterized in failing and nonfailing human hearts. Both nonfailing atria had distinct transcriptomic patterns characterized by cardiogenesis, the immune system and BMP/TGF-ß, MAPK/JNK, and Wnt signaling. Failing atria and ventricles were enriched for gene sets associated with the innate and adaptive immune system. Key upstream regulators associated with heart failure were identified, including activated immune response elements, which may constitute novel therapeutic targets.

Sequence-based genotyping of extra-intestinal pathogenic Escherichia coli isolates from patients with suspected community-onset sepsis, Sweden.

Extra-intestinal pathogenic Escherichia coli (ExPEC) strains are responsible for a large number of human infections globally. The management of infections caused by ExPEC has been complicated by the emergence of antimicrobial resistance, most importantly the increasing recognition of isolates producing extended-spectrum ß-lactamases (ESBL). Herein, we used whole-genome sequencing (WGS) on ExPEC isolates for a comprehensive genotypic characterization. Twenty-one ExPEC isolates, nine with and 12 without ESBL-production, from 16 patients with suspected sepsis were sequenced on an Illumina MiSeq platform. Analysis of WGS data was performed with widely used bioinformatics software and tools for genotypic characterization of the isolates. A higher number of plasmids, virulence and resistance genes were observed in the ESBL-producing isolates than the non-ESBL-producing, although not statistically significant due to the low sample size. All nine ESBL-producing ExPEC isolates presented with at least one bla gene, as did three of the 12 without ESBL-production. Multi-locus sequence typing analysis revealed a diversity of sequence types whereas phylogroup A prevailed among isolates both with and without ESBL-production. In conclusion, this limited study shows that analysis of WGS data can be used for genotypic characterization of ExPEC isolates to obtain in-depth information of clinical relevance.

Detection of mobile genetic elements associated with antibiotic resistance in Salmonella enterica using a newly developed web tool: MobileElementFinder.

OBJECTIVES: Antimicrobial resistance (AMR) in clinically relevant bacteria is a growing threat to public health globally. In these bacteria, antimicrobial resistance genes are often associated with mobile genetic elements (MGEs), which promote their mobility, enabling them to rapidly spread throughout a bacterial community. METHODS: The tool MobileElementFinder was developed to enable rapid detection of MGEs and their genetic context in assembled sequence data. MGEs are detected based on sequence similarity to a database of 4452 known elements augmented with annotation of resistance genes, virulence factors and detection of plasmids. RESULTS: MobileElementFinder was applied to analyse the mobilome of 1725 sequenced Salmonella enterica isolates of animal origin from Denmark, Germany and the USA. We found that the MGEs were seemingly conserved according to multilocus ST and not restricted to either the host or the country of origin. Moreover, we identified putative translocatable units for specific aminoglycoside, sulphonamide and tetracycline genes. Several putative composite transposons were predicted that could mobilize, among others, AMR, metal resistance and phosphodiesterase genes associated with macrophage survivability. This is, to our knowledge, the first time the phosphodiesterase-like pdeL has been found to be potentially mobilized into S. enterica. CONCLUSIONS: MobileElementFinder is a powerful tool to study the epidemiology of MGEs in a large number of genome sequences and to determine the potential for genomic plasticity of bacteria. This web service provides a convenient method of detecting MGEs in assembled sequence data. MobileElementFinder can be accessed at https://cge.cbs.dtu.dk/services/MobileElementFinder/.

IL-16 processing in sentinel node regulatory T cells is a factor in bladder cancer immunity.

In the effort of developing new immunotherapies, the sentinel node (SN) has proven a promising source from which to harness an effective antitumour T cell response. However, tumour immune escape, a process in which regulatory T cells (Tregs) play a central role, remains a major limiting factor. Therefore, there is a clear need to increase the knowledge of Treg function and signalling in sentinel nodes. Here, we set out to explore whether the proteome in SN-resident T cells is altered by the tumour and to identify key proteins in SN T cell signalling, focusing on Tregs. Five patients with muscle-invasive urothelial bladder cancer were prospectively included. Mass spectrometry was performed on two patients, with validation and functional studies being performed on three additional patients and four healthy donors. At cystectomy, SN, non-SN lymph nodes and peripheral blood samples were collected from the patients and T cell subsets isolated through flow cytometry before downstream experiments. Proteomic analysis indicated that growth and immune signalling pathways are upregulated in SN-resident Tregs. Furthermore, centrality analysis identified the cytokine IL-16 to be central in the SN-Treg signalling network. We show that tumour-released factors, through activating caspase-3, increase Treg IL-16 processing into bioactive forms, reinforcing Treg suppressive capacity. In conclusion, we provide evidence that Tregs exposed to secreted factors from bladder tumours show increased immune and growth signalling and altered IL-16 processing which translates to enhanced Treg suppressive function, indicating altered IL-16 signalling as a novel tumour immune escape mechanism.

Fewer tumour draining sentinel nodes in patients with progressing muscle invasive bladder cancer, after neoadjuvant chemotherapy and radical cystectomy.

PURPOSE: To examine the relationship between the number of tumour draining sentinel nodes (SNs) and pathoanatomical outcomes, in muscle-invasive bladder cancer (MIBC), in patients undergoing neoadjuvant chemotherapy (NAC) and radical cystectomy (RC). MATERIALS AND METHODS: In an ongoing prospective multicenter study, we included 230 patients with suspected urothelial MIBC from ten Swedish urological centers. All underwent TURb and clinical staging. From the cohort, 116 patients with urothelial MIBC; cT2-cT4aN0M0, underwent radical cystectomy (RC) and lymphadenectomy with SN-detection (SNd). 83 patients received cisplatin-based NAC and 33 were NAC-naïve. The number and locations of detected SNs and non-SNs were recorded for each patient. The NAC treated patients were categorized by pathoanatomical outcomes post-RC into three groups: complete responders (CR), stable disease (SD) and progressive disease (PD). Selected covariates with possible impact on SN-yield were tested in uni -and multivariate analyses for NAC-treated patients only. RESULTS: In NAC treated patients, the mean number of SNs was significantly higher in CR patients (3.3) and SD patients (3.6) compared with PD patients (1.4) (p = 0.034). In a linear multivariate regression model, the number of harvested nodes was the only independent variable that affected the number of SNs (p = 0.0004). CONCLUSIONS: The number of tumor-draining SNs in NAC-treated patients was significantly lower in patients with progressive disease.

Doxorubicin enhances the capacity of B cells to activate T cells in urothelial urinary bladder cancer.

Cancer is currently treated by a combination of therapies, including chemotherapy which is believed to suppress the immune system. Combination of immunotherapy and chemotherapy correlates with improved survival but needs careful planning in order to achieve a synergistic effect. In this study, we have demonstrated that doxorubicin treatment of B cells resulted in increased expression of CD86 and concordantly increased CD4+ T cell activation in the presence of superantigen, an effect that was inhibited by the addition of a CD86 blocking antibody. Furthermore, doxorubicin resulted in decreased expression of the anti-inflammatory cytokines IL-10 and TNF-α. Finally, B cells from urinary bladder cancer patients, treated with a neoadjuvant regiment containing doxorubicin, displayed increased CD86-expression. We conclude that doxorubicin induces CD86 expression on B cells and hence enhances their antigen-presenting ability in vitro, a finding verified in patients. Development of tailored time and dose schedules may increase the effectiveness of combining chemotherapy and immunotherapy.

Constructive Method for Detecting the Information Backflow of Non-Markovian Dynamics.

We investigate the relation between two approaches to the characterization of quantum Markovianity, divisibility and lack of information backflow. We show that a bijective dynamical map is completely positive divisible if and only if a monotonic nonincrease of distinguishability is observed for two equiprobable states of the evolving system and an ancilla. Moreover, our proof is constructive: given any such map that is not completely positive divisible, we give an explicit construction of two states that, when taken with the same a priori probability, exhibit information backflow. Finally, while an ancilla is necessary for the equivalence to hold in general, we show that it is always possible to witness the non-Markovianity of bijective maps without using any entanglement between the system and ancilla.

Sentinel node detection in muscle-invasive urothelial bladder cancer is feasible after neoadjuvant chemotherapy in all pT stages, a prospective multicenter report.

PURPOSE: To determine whether sentinel node detection (SNd) in muscle-invasive urothelial bladder cancer (MIBC) can be performed in patients undergoing neoadjuvant chemotherapy (NAC) and determine whether SNd is feasible in all pT stages, including pT0. BACKGROUND: Previous published series of SNd in MIBC have not included patients undergoing NAC, and systematic reports of pT0 patients w/wo NAC were absent. Translational immunological tumor research on MIBC focusing on SNd, in the era of NAC, requires technical feasibility. Additionally, SNd in MIBC requests further evaluations as a method for nodal staging. MATERIALS AND METHODS: Ninety-nine patients with suspected urothelial MIBC were prospectively selected from six urological centers. After TUR-B and primary staging, 65 MIBC patients qualified for radical cystectomy. Precystectomy staging was cT2a-T4aN0M0, including 47 NAC patients and 18 chemo-naïve patients. All 65 patients underwent intraoperative SNd by peritumoral injection of 80 Mbq Technetium and Geiger probe detection. Postcystectomy staging was pT0-T4aN0-N2M0. SNs were defined by two calculations, SNdef1 and SNdef2. RESULTS: Totally 1063 lymph nodes were removed (total SNs; 222-227). NAC patients with pT0 (n = 24) displayed a true positive detection in 91.7 % by either SNdef, with a median of 3.0 SNs. NACpT >0 patients had a true positive detection in 87 % (SNdef1) and 91.3 % (SNdef2). In a univariate analysis, patient group neither NAC nor tumor downstaging influenced detection rates, regardless of SN definition. In total eight patients, 4/22 metastatic nodes were SNs while 18/22 were non-SNs. CONCLUSIONS: Sentinel node detection in MIBC is feasible also in NAC patients, regardless of pT stage. SNd played no role in nodal staging.

Metagenomic analysis of sewage for surveillance of bacterial pathogens: A release experiment to determine sensitivity.

Accurate monitoring of gastro-enteric and other diseases in large populations poses a challenge for public health management. Sewage represents a larger population, is freely obtainable and non-subject to ethical approval. Metagenomic sequencing offers simultaneous, multiple-target analysis. However, no study has demonstrated the sensitivity of metagenomics for detecting bacteria in sewage. In this study, we spot-released 1013 colony-forming units (CFU) of Staphyloccus hyicus (non-pathogenetic strain 842J-88). The strain was flushed down a toilet into the sewer in the catchment area of a public wastewater treatment plant (WWTP), serving a population of 36,000 people. Raw sewage was continuously sampled at the WWTP's inlet over 30- and 60-minute intervals for a total period of seven hours. The experiment was conducted twice with one week in-between release days and under comparable weather conditions. For the metagenomics analyses, the pure single isolate of S. hyicus was sequenced, assembled and added to a large database of bacterial reference sequences. All sewage samples were analyzed by shotgun metagenome sequencing and mapped against the reference database. S. hyicus was identified in duplicate samples at both of two release days and these sequence fragment counts served as a proxy to estimate the minimum number of sick people or sensitivity required in order to observe at least one sick person at 95% probability. We found the sensitivity to be in the range 41-140 and 16-36 sick people at release days 1 and 2, respectively. The WWTP normally serves 36,000 people giving a normalized sensitivity in the range of one in 257 to 2,250 persons.

Importance of mobile genetic elements for dissemination of antimicrobial resistance in metagenomic sewage samples across the world.

We are facing an ever-growing threat from increasing antimicrobial resistance (AMR) in bacteria. To mitigate this, we need a better understanding of the global spread of antimicrobial resistance genes (ARGs). ARGs are often spread among bacteria by horizontal gene transfer facilitated by mobile genetic elements (MGE). Here we use a dataset consisting of 677 metagenomic sequenced sewage samples from 97 countries or regions to study how MGEs are geographically distributed and how they disseminate ARGs worldwide. The ARGs, MGEs, and bacterial abundance were calculated by reference-based read mapping. We found systematic differences in the abundance of MGEs and ARGs, where some elements were prevalent on all continents while others had higher abundance in separate geographic areas. Different MGEs tended to be localized to temperate or tropical climate zones, while different ARGs tended to separate according to continents. This suggests that the climate is an important factor influencing the local flora of MGEs. MGEs were also found to be more geographically confined than ARGs. We identified several integrated MGEs whose abundance correlated with the abundance of ARGs and bacterial genera, indicating the ability to mobilize and disseminate these genes. Some MGEs seemed to be more able to mobilize ARGs and spread to more bacterial species. The host ranges of MGEs seemed to differ between elements, where most were associated with bacteria of the same family. We believe that our method could be used to investigate the population dynamics of MGEs in complex bacterial populations.

The Risk of Thromboembolism in Patients with Muscle Invasive Bladder Cancer before and after Cystectomy Depending on Blood Group and Neoadjuvant Chemotherapy-A Multicentre Retrospective Cohort Study.

PURPOSE: Previous studies have indicated that patients with muscle-invasive bladder cancer with non-O blood types have an increased risk of experiencing thromboembolic events (TEEs). This is finding is in relation to neoadjuvant-chemotherapy (NAC)-naïve patients. AIM: to establish the risk of TEEs and any association with blood types among NAC patients as well as NAC-naïve patients. METHODS: Cystectomized patients at four centres treated from 2009 to 2018 (n = 244) were analysed. The quantities of patients corresponding to each blood group were as follows: A-108 (44%); O-99 (41%); B-30 (12%); and AB-7 (3%). NAC patients (n = 167) and NAC-naïve NAC-eligible patients (n = 77) were assessed. In total, 54 women (22%) and 190 men (78%), with a median age of 69 years, were included in the study. The occurrence of any type of TEE from six months pre-cystectomy to 12-24 months after was analysed using logistic regression adjusted for NAC and confounders. RESULTS: Sixty-six TEEs were detected in 21% of the patients (n = 52). Pulmonary embolus (n = 33) and deep venous thrombosis (n = 11) were the most common forms. No significant differences between blood types were found in the analysis, although B blood type had a nearly significant increased crude risk compared with O blood type, for which there was an OR of 2.48 (95% CI 0.98-6.36). Adjustment for NAC and covariates weakened the OR, which plummeted to 1.98 (95% CI 0.71-5.51). CONCLUSIONS: No significant associations were found between blood types and TEE occurrences in this cohort including both NAC and NAC-naïve NAC-eligible patients.

Data Mining Identifies CCN2 and THBS1 as Biomarker Candidates for Cardiac Hypertrophy.

Cardiac hypertrophy is a condition that may contribute to the development of heart failure. In this study, we compare the gene-expression patterns of our in vitro stem-cell-based cardiac hypertrophy model with the gene expression of biopsies collected from hypertrophic human hearts. Twenty-five differentially expressed genes (DEGs) from both groups were identified and the expression of selected corresponding secreted proteins were validated using ELISA and Western blot. Several biomarkers, including CCN2, THBS1, NPPA, and NPPB, were identified, which showed significant overexpressions in the hypertrophic samples in both the cardiac biopsies and in the endothelin-1-treated cells, both at gene and protein levels. The protein-interaction network analysis revealed CCN2 as a central node among the 25 overlapping DEGs, suggesting that this gene might play an important role in the development of cardiac hypertrophy. GO-enrichment analysis of the 25 DEGs revealed many biological processes associated with cardiac function and the development of cardiac hypertrophy. In conclusion, we identified important similarities between ET-1-stimulated human-stem-cell-derived cardiomyocytes and human hypertrophic cardiac tissue. Novel putative cardiac hypertrophy biomarkers were identified and validated on the protein level, lending support for further investigations to assess their potential for future clinical applications.

Multi-Omics Characterization of a Human Stem Cell-Based Model of Cardiac Hypertrophy.

Cardiac hypertrophy is an important and independent risk factor for the development of cardiac myopathy that may lead to heart failure. The mechanisms underlying the development of cardiac hypertrophy are yet not well understood. To increase the knowledge about mechanisms and regulatory pathways involved in the progression of cardiac hypertrophy, we have developed a human induced pluripotent stem cell (hiPSC)-based in vitro model of cardiac hypertrophy and performed extensive characterization using a multi-omics approach. In a series of experiments, hiPSC-derived cardiomyocytes were stimulated with Endothelin-1 for 8, 24, 48, and 72 h, and their transcriptome and secreted proteome were analyzed. The transcriptomic data show many enriched canonical pathways related to cardiac hypertrophy already at the earliest time point, e.g., cardiac hypertrophy signaling. An integrated transcriptome-secretome analysis enabled the identification of multimodal biomarkers that may prove highly relevant for monitoring early cardiac hypertrophy progression. Taken together, the results from this study demonstrate that our in vitro model displays a hypertrophic response on both transcriptomic- and secreted-proteomic levels. The results also shed novel insights into the underlying mechanisms of cardiac hypertrophy, and novel putative early cardiac hypertrophy biomarkers have been identified that warrant further investigation to assess their potential clinical relevance.

Central Venous Access and the Risk for Thromboembolic Events in Patients Undergoing Neoadjuvant Chemotherapy and Radical Cystectomy for Muscle-Invasive Bladder Cancer.

Thromboembolic events (TEE) are high-risk complications in patients undergoing neoadjuvant chemotherapy (NAC) and radical cystectomy (RC) for urothelial muscle-invasive bladder cancer (MIBC). The purpose of the study was to investigate any differences in TEE-incidence, comparing peripherally inserted central catheter (PICC) versus a totally implanted port (PORT) as CVA (central venous access) during NAC. We identified 947 cystectomized MIBC-patients from four Swedish medical centers in 2009-2021. Inclusion criteria were cT2-T4aN0M0 and 375 patients were finally eligible and evaluated, divided into: NAC-administered (n = 283) resp. NAC-naïve-NAC-eligible (n = 92), the latter as tentative control group. Data on TEEs and types of CVA were retrospectively collected and individually validated, from final transurethral resection of the bladder tumor (TUR-B) to 30 days post-RC. Adjusted logistic regression and log rank test were used for statistical analyses. Amongst NAC-administered, 83% (n = 235) received PICCs and 15% (n = 42) PORTs. Preoperative TEEs occurred in 38 PICC-patients (16.2%) and in one PORT-patient (2.4%), with 47 individual events registered. We found a significantly increased odds ratio of TEE in NAC-administered PICC-patients compared to in PORT-patients (OR: 8.140, p-value: 0.042, 95% CI 1.078-61.455). Our findings indicate a greater risk for pre-RC TEEs with PICCs than with PORTs, suggesting favoring the usage of PORTs for MIBC-NAC-patients.

Thromboembolic Events in Patients Undergoing Neoadjuvant Chemotherapy and Radical Cystectomy for Muscle-Invasive Bladder Cancer: A Study of Renal Impairment in Relation to Potential Thromboprophylaxis.

Recent studies on patients with muscle-invasive bladder cancer (MIBC) undergoing neoadjuvant chemotherapy (NAC) have shown an association between NAC and thromboembolic events (TEE) prior to radical cystectomy (RC). Recent studies suggest that central venous access catheters (CVAs) may induce TEEs, and low-molecular-weight heparin (LMWH) has been mentioned as possible prophylaxis. However, other studies have shown a high incidence of decreased renal function in these patients. The purpose of this study was to determine the portion of MIBC patients with NAC-induced TEEs who had decreased preoperative renal function for whom LMWH potentially would not be beneficial as prophylaxis. We identified 459 cystectomized MIBC patients from two Swedish medical centers from 2009 to 2021. The inclusion criterion of cT2-T4aN0M0 resulted in 220 eligible patients, who were further divided into NAC-administered (n = 187) and NAC-eligible (n = 33), the tentative control group. Values of renal function before, during, and after each NAC cycle were retrospectively collected from individual medical records. Amongst the NAC-administered patients with TEE (n = 29), 41% (95% CI 23.5-61.1%) of patients had decreased renal function. Thus, a substantial portion of NAC-administered patients who developed TEEs had reduced renal function and would have been less likely to have benefited from renal clearance-dependent LMWH prophylaxis.

A Retrospective Analysis of the De Ritis Ratio in Muscle Invasive Bladder Cancer, with Focus on Tumor Response and Long-Term Survival in Patients Receiving Neoadjuvant Chemotherapy and in Chemo Naïve Cystectomy Patients-A Study of a Clinical Multicentre Database.

Background: A high pre-treatment De Ritis ratio, the aspartate transaminase/alanine aminotransferase ratio, has been suggested to be of prognostic value for mortality in muscle-invasive bladder cancer (MIBC). Our purpose was to evaluate if a high ratio was associated with mortality and downstaging. Methods: A total of 347 Swedish patients with clinically staged T2-T4aN0M0, with administered neoadjuvant chemotherapy (NAC) or eligible for NAC and undergoing radical cystectomy (RC) 2009−2021, were retrospectively evaluated with a low ratio < 1.3 vs. high ratio > 1.3, by Log Rank test, Cox regression and Mann−Whitney U-test (MWU), SPSS 27. Results: Patients with a high ratio had a decrease of up to 3 years in disease-free survival (DFS), cancer-specific survival (CSS) and overall survival (OS) (p = 0.009, p = 0.004 and p = 0.009) and 5 years in CSS and OS (p = 0.019 and p = 0.046). A high ratio was associated with increased risk of mortality, highest in DFS (HR, 1.909; 95% CI, 1.265−2.880; p = 0.002). No significant relationship between downstaging and a high ratio existed (p = 0.564 MWU). Conclusion: A high pre-treatment De Ritis ratio is on a population level, associated with increased mortality post-RC in endpoints DFS, CSS and OS. Associations decrease over time and require further investigations to determine how strong the associations are as meaningful prognostic markers for long-term mortality in MIBC. The ratio is not suitable for downstaging-prediction.

A prospective multicenter study of visual response-evaluation by cystoscopy in patients undergoing neoadjuvant chemotherapy for muscle invasive urinary bladder cancer.

PURPOSE: To evaluate a method of transurethral visual response-staging in patients with urothelial muscle-invasive urinary bladder cancer (MIBC), undergoing neoadjuvant chemotherapy (NAC) and radical cystectomy (RC). METHODS: A prospective study at four Swedish cystectomy centers, cystoscopy was performed after final NAC-cycle for MIBC. Fifty-six participants underwent cystoscopy for visual staging of the tumor immediately pre-RC. Visual assessments were correlated to pathoanatomical outcomes post-RC. RESULTS: Seventeen tumors were classified as complete response (CR), i.e. pT0. Twenty-five patients had residual MIBC and 14 had non-muscle invasive residual tumors (NMIBC). Of the 39 patients with residual tumor, 25 were correctly identified visually (64%). Eleven patients were pN+. The diagnostic accuracy of cystoscopy to correctly identify complete response or remaining tumor was 70% (CI = 56-81%) with a sensitivity of 64% (CI = 47-79%), specificity 82% (CI = 57-96%), PPV 89% (CI = 74-96%) and NPV 50% (CI =38-61%). Twenty-eight cystoscopy evaluations showed signs of residual tumors and 3/28 (11%) were false positive. In 4/14 patients assessed having residual NMIBC the estimates were correct, 8/14 had histopathological MIBC and 2/14 had CR. In 11/14 patients (79%), the suggested visual assessment of MIBC was correct, 2/14 had NMIBC and 1/14 had CR. Twenty-eight cystoscopies had negative findings, 14 were false negatives (50%), when cystoscopy falsely predicted pT0. Among them there were eight patients with pTa, pT1 or pTis and six MIBC-tumors. In 17 patients with histopathological pT0, 14 were correctly identified with cystoscopy (82%). CONCLUSION: Cystoscopy after the final NAC-cycle cannot robustly differentiate between NAC-responders and non-responders. Visually, negative MIBC-status cannot be determined safely.

Adverse events during neoadjuvant chemotherapy for muscle invasive bladder cancer-a Swedish retrospective multicentre study of a clinical database.

Background: Adverse events (AEs) during neoadjuvant chemotherapy (NAC) for muscle invasive bladder cancer (MIBC) are known but insufficiently reported. Clinical implications include affected cardiac, pulmonary, urinary, vascular and haematological organ systems. The main purpose was to evaluate the incidence and severity of AEs for ascertaining possible clinical significance. Further investigating possible effects of AEs on downstaging outcomes-downstaging is considered a surrogate marker for overall survival (OS). Methods: A retrospective evaluation of AEs during ongoing NAC for MIBC patients analysing individual patient data in a clinical database. We identified 687 cystectomies between 2009-2020 at four Swedish urological centres. Inclusion criteria were cT2-4aN0M0 in 261 NAC patients undergoing radical cystectomy (RC). Medical files were reviewed and AEs were assessed and graded, including detailed measurements by the Common Terminology Criteria for Adverse Events (CTCAE) v.5. Data were retrospectively analysed in SPSS statistics 27.0 with Spearman rank-order correlation coefficient and Mann-Whitney U-test (MWU). Results: A total of 251/261 patients [95% confidence interval (CI), 93-98%] experienced AEs during NAC pre-RC (mean two AEs/patient). In total, 208 (80%) patients received methotrexate, vinblastine, adriamycin (doxorubicin) and cisplatin (MVAC). In the total cohort, 200 (76.6%) received all pre-planned NAC-cycles. Most common AEs were anaemia (88.9%), thrombocytopenia (44.8%) and acute kidney injury (40.6%). Patients with prematurely terminated cycles had higher AE-grades (P=0.042 MWU). A correlation between higher AE-grades and decrease in downstaging existed, in the entire cohort (-0.133; P=0.033) and in patients undergoing all pre-planned NAC-cycles (-0.148; P=0.038). Anaemia and acute kidney injury were individually associated with decreased downstaging (-0.360, P=0.025 and -0.183, P=0.010, respectively). Conclusions: NAC in MIBC poses a significant risk for AEs before RC with clinical implications. For instance, patients terminating chemotherapy prematurely, have higher AE-grades and decreased downstaging. Further, acute kidney injury and anaemia are individually associated with decreased downstaging. We propose that early detection and prevention of AEs may increase downstaging of the primary tumour.

Proteomic Profiling of Tissue Exosomes Indicates Continuous Release of Malignant Exosomes in Urinary Bladder Cancer Patients, Even with Pathologically Undetectable Tumour.

Invasive urothelial bladder cancer (UBC) has high recurrence rates even after radical cystectomy (RC). Exosomes are membrane-bound nanovesicles, which have been shown to contribute to carcinogenesis and metastasis. We previously showed that urinary exosomes display a malignant profile in UBC patients despite the absence of detectable tumour. Here, we investigated exosomes from sampling sites close to or distant from the former tumour, aiming to understand the effect of the tumour on the local milieu. Ten patients scheduled for cystectomy after transurethral bladder resection (TUR-B), without remaining detectable tumour, were included. Exosomes were isolated from tissue explants of both the previous tumour site and distant bladder tissue. Proteins were quantified by mass spectrometry in seven patients. Exosomes from the previous tumour site were enriched in inflammatory but not cancer-related pathways compared to distant tissue. However, the 69 most abundant proteins in tissue-derived exosomes regardless of site, 20 of which were also found in urinary exosomes from our previous study, were enriched for cancer-related metabolic pathways and associated with poor prognosis in an external mRNA dataset. The enrichment of cancer-related pathways in the most abundant proteins, regardless of sampling site, confirms our hypothesis that despite the absence of detectable tumour, the entire bladder releases exosomes that contribute to metastasis and highlights the need for early RC.

Genomic evolution of antimicrobial resistance in Escherichia coli.

The emergence of antimicrobial resistance (AMR) is one of the biggest health threats globally. In addition, the use of antimicrobial drugs in humans and livestock is considered an important driver of antimicrobial resistance. The commensal microbiota, and especially the intestinal microbiota, has been shown to have an important role in the emergence of AMR. Mobile genetic elements (MGEs) also play a central role in facilitating the acquisition and spread of AMR genes. We isolated Escherichia coli (n = 627) from fecal samples in respectively 25 poultry, 28 swine, and 15 veal calf herds from 6 European countries to investigate the phylogeny of E. coli at country, animal host and farm levels. Furthermore, we examine the evolution of AMR in E. coli genomes including an association with virulence genes, plasmids and MGEs. We compared the abundance metrics retrieved from metagenomic sequencing and whole genome sequenced of E. coli isolates from the same fecal samples and farms. The E. coli isolates in this study indicated no clonality or clustering based on country of origin and genetic markers; AMR, and MGEs. Nonetheless, mobile genetic elements play a role in the acquisition of AMR and virulence genes. Additionally, an abundance of AMR was agreeable between metagenomic and whole genome sequencing analysis for several AMR classes in poultry fecal samples suggesting that metagenomics could be used as an indicator for surveillance of AMR in E. coli isolates and vice versa.