Comprehensive genomic characterization of early-stage bladder cancer.

Understanding the molecular landscape of nonmuscle-invasive bladder cancer (NMIBC) is essential to improve risk assessment and treatment regimens. We performed a comprehensive genomic analysis of patients with NMIBC using whole-exome sequencing (n = 438), shallow whole-genome sequencing (n = 362) and total RNA sequencing (n = 414). A large genomic variation within NMIBC was observed and correlated with different molecular subtypes.

Whole-genome Mutational Analysis for Tumor-informed Detection of Circulating Tumor DNA in Patients with Urothelial Carcinoma.

Background And Objective: Circulating tumor DNA (ctDNA) can be used for sensitive detection of minimal residual disease (MRD). However, the probability of detecting ctDNA in settings of low tumor burden is limited by the number of mutations analyzed and the plasma volume available. We used a whole-genome sequencing (WGS) approach for ctDNA detection in patients with urothelial carcinoma.

Beyond basics: Key mutation selection features for successful tumor-informed ctDNA detection.

Tumor-informed mutation-based approaches are frequently used for detection of circulating tumor DNA (ctDNA). Not all mutations make equally effective ctDNA markers. The objective was to explore if prioritizing mutations using mutational features-such as cancer cell fraction (CCF), multiplicity, and error rate-would improve the success rate of tumor-informed ctDNA analysis.

Multiomics profiling of urothelial carcinoma reveals CIS-specific gene signature and immune characteristics.

Urothelial carcinoma (CIS) is an aggressive phenotype of non-muscle-invasive bladder cancer. Molecular features unique to CIS compared to high-grade papillary tumors are underexplored. RNA sequencing of CIS, papillary tumors, and normal urothelium showed lower immune marker expression in CIS compared to papillary tumors.

Circulating Tumor DNA Analysis in Advanced Urothelial Carcinoma: Insights from Biological Analysis and Extended Clinical Follow-up.

Purpose: To investigate whether circulating tumor DNA (ctDNA) assessment in patients with muscle-invasive bladder cancer predicts treatment response and provides early detection of metastatic disease.

Experimental Design: We present full follow-up results (median follow-up: 68 months) from a previously described cohort of 68 neoadjuvant chemotherapy (NAC)-treated patients who underwent longitudinal ctDNA testing (712 plasma samples). In addition, we performed ctDNA evaluation of 153 plasma samples collected before and after radical cystectomy (RC) in a separate cohort of 102 NAC-naïve patients (median follow-up: 72 months).

Field Cancerization Is Associated with Tumor Development, T-cell Exhaustion, and Clinical Outcomes in Bladder Cancer.

Background: Field cancerization is characterized by areas of normal tissue affected by mutated clones. Bladder field cancerization may explain the development and recurrence of bladder cancer and may be associated with treatment outcomes.

Objective: To investigate the predictive and prognostic roles of field cancerization in patients with high-risk non-muscle-invasive bladder cancer (NMIBC) treated with bacillus Calmette-Guérin (BCG).

NGS-Based Tumor-Informed Analysis of Circulating Tumor DNA.

Accurate circulating tumor DNA (ctDNA) detection has an immense biomarker potential in all phases of the cancer disease course. Presence of ctDNA in the blood has been shown to have prognostic value in various cancer types as it may reflect the actual tumor burden. There are two main methods to consider, a tumor-informed and a tumor-agnostic analysis of ctDNA.

Single-nucleus and Spatially Resolved Intratumor Subtype Heterogeneity in Bladder Cancer.

Background: Current bulk transcriptomic classification systems for bladder cancer do not consider the level of intratumor subtype heterogeneity.

Objective: To investigate the extent and possible clinical impact of intratumor subtype heterogeneity across early and more advanced stages of bladder cancer.

Design Setting And Participants: We performed single-nucleus RNA sequencing (RNA-seq) of 48 bladder tumors and additional spatial transcriptomics for four of these tumors.

Improved protocol for single-nucleus RNA-sequencing of frozen human bladder tumor biopsies.

This paper provides a laboratory workflow for single-nucleus RNA-sequencing (snRNA-seq) including a protocol for gentle nuclei isolation from fresh frozen tumor biopsies, making it possible to analyze biobanked material. To develop this protocol, we used non-frozen and frozen human bladder tumors and cell lines. We tested different lysis buffers (IgePal and Nuclei EZ) and incubation times in combination with different approaches for tissue and cell dissection: sectioning, semi-automated dissociation, manual dissociation with pestles, and semi-automated dissociation combined with manual dissociation with pestles.

Cell-Free Urine and Plasma DNA Mutational Analysis Predicts Neoadjuvant Chemotherapy Response and Outcome in Patients with Muscle-Invasive Bladder Cancer.

Purpose: To investigate the use of plasma and urine DNA mutation analysis for predicting neoadjuvant chemotherapy (NAC) response and oncological outcome in patients with muscle-invasive bladder cancer.

Experimental Design: Whole-exome sequencing of tumor and germline DNA was performed for 92 patients treated with NAC followed by radical cystectomy (RC). A custom NGS-panel capturing approximately 50 mutations per patient was designed and used to track mutated tumor DNA in plasma and urine.

Elevated T-cell Exhaustion and Urinary Tumor DNA Levels Are Associated with Bacillus Calmette-Guérin Failure in Patients with Non-muscle-invasive Bladder Cancer.

Background: The functional status of immune cells in the tumor microenvironment and tumor characteristics may explain bacillus Calmette-Guérin (BCG) failure in high-risk non-muscle-invasive bladder cancer (NMIBC).

Objective: To characterize molecular correlates of post-BCG high-grade (HG) recurrence using multiomics analysis.

Design, Setting, And Participants: Patients with BCG-treated NMIBC (n = 156) were included in the study.

Circulating tumor DNA for prognosis assessment and postoperative management after curative-intent resection of colorectal liver metastases.

The recurrence rate of colorectal liver metastases (CRLM) patients treated with curative intent is above 50%. Standard of care surveillance includes intensive computed tomographic (CT) imaging as well as carcinoembryonic antigen (CEA) measurements. Nonetheless, relapse detection often happens too late to resume curative treatment.

, , Mutation Status and APOBEC Mutation Signatures are Predictive of Mitomycin C Response in Non-muscle-invasive Bladder Cancer.

Background: Currently, no biomarkers of response to mitomycin C have been identified in non-muscle-invasive bladder cancer patients. Predictive biomarkers could improve the treatment outcome and eliminate adverse events from unnecessary treatment.

Objective: To identify and validate predictive biomarkers of chemoresection with mitomycin C.

An integrated multi-omics analysis identifies prognostic molecular subtypes of non-muscle-invasive bladder cancer.

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.

Molecular correlates of cisplatin-based chemotherapy response in muscle invasive bladder cancer by integrated multi-omics analysis.

Overtreatment with cisplatin-based chemotherapy is a major issue in the management of muscle-invasive bladder cancer (MIBC), and currently none of the reported biomarkers for predicting response have been implemented in the clinic. Here we perform a comprehensive multi-omics analysis (genomics, transcriptomics, epigenomics and proteomics) of 300 MIBC patients treated with chemotherapy (neoadjuvant or first-line) to identify molecular changes associated with treatment response. DNA-based associations with response converge on genomic instability driven by a high number of chromosomal alterations, indels, signature 5 mutations and/or BRCA2 mutations.

Epigenetic and transcriptomic consequences of excess X-chromosome material in 47,XXX syndrome-A comparison with Turner syndrome and 46,XX females.

47,XXX (triple X) and Turner syndrome (45,X) are sex chromosomal abnormalities with detrimental effects on health with increased mortality and morbidity. In karyotypical normal females, X-chromosome inactivation balances gene expression between sexes and upregulation of the X chromosome in both sexes maintain stoichiometry with the autosomes. In 47,XXX and Turner syndrome a gene dosage imbalance may ensue from increased or decreased expression from the genes that escape X inactivation, as well as from incomplete X chromosome inactivation in 47,XXX.

Representative Sequencing: Unbiased Sampling of Solid Tumor Tissue.

Although thousands of solid tumors have been sequenced to date, a fundamental under-sampling bias is inherent in current methodologies. This is caused by a tissue sample input of fixed dimensions (e.g.

Analysis of Plasma Cell-Free DNA by Ultradeep Sequencing in Patients With Stages I to III Colorectal Cancer.

Importance: Novel sensitive methods for detection and monitoring of residual disease can improve postoperative risk stratification with implications for patient selection for adjuvant chemotherapy (ACT), ACT duration, intensity of radiologic surveillance, and, ultimately, outcome for patients with colorectal cancer (CRC).

Objective: To investigate the association of circulating tumor DNA (ctDNA) with recurrence using longitudinal data from ultradeep sequencing of plasma cell-free DNA in patients with CRC before and after surgery, during and after ACT, and during surveillance.

Design, Setting, And Participants: In this prospective, multicenter cohort study, ctDNA was quantified in the preoperative and postoperative settings of stages I to III CRC by personalized multiplex, polymerase chain reaction-based, next-generation sequencing.

Early Detection of Metastatic Relapse and Monitoring of Therapeutic Efficacy by Ultra-Deep Sequencing of Plasma Cell-Free DNA in Patients With Urothelial Bladder Carcinoma.

Purpose: Novel sensitive methods for early detection of relapse and for monitoring therapeutic efficacy may have a huge impact on risk stratification, treatment, and ultimately outcome for patients with bladder cancer. We addressed the prognostic and predictive impact of ultra-deep sequencing of cell-free DNA in patients before and after cystectomy and during chemotherapy.

Patients And Methods: We included 68 patients with localized advanced bladder cancer.

Cellular Requirements for Sensing and Elimination of Incoming HSV-1 DNA and Capsids.

Incoming viruses challenge the cell with diverse foreign molecules, which need to be sensed quickly to initiate immune responses and to remove the viral components. In this study, we investigate the cellular requirements for sensing and degradation of incoming viral DNA and capsids during herpes simplex virus type 1 (HSV-1) infections. Using click chemistry labeling of the viral genome, we found that HSV-1 DNA was released from a subset of capsids into the cytosol early in infection.

Discordant molecular subtype classification in the basal-squamous subtype of bladder tumors and matched lymph-node metastases.

Molecular subtypes of muscle-invasive bladder tumors have emerged as a promising research tool with potential to stratify patients for neoadjuvant treatment. Prior to radical cystectomy, the utility of molecular classification and biomarkers depend on concordance between tissue from transurethrally resected specimens and disseminated disease. We assess the concordance of molecular subtypes and a large number of potential biomarkers in 67 pairs of muscle-invasive bladder tumors and synchronous lymph-node metastases.

Optimized targeted sequencing of cell-free plasma DNA from bladder cancer patients.

Analysis of plasma cell-free DNA (cfDNA) may provide important information in cancer research, though the often small fraction of DNA originating from tumor cells makes the analysis technically challenging. Digital droplet PCR (ddPCR) has been utilized extensively as sufficient technical performance is easily achieved, but analysis is restricted to few mutations. Next generation sequencing (NGS) approaches have been optimized to provide comparable technical performance, especially with the introduction of unique identifiers (UIDs).