Cystectomy is a standard treatment for muscle-invasive bladder cancer (MIBC), but it is life-altering. We initiated a phase 2 study in which patients with MIBC received four cycles of gemcitabine, cisplatin, plus nivolumab followed by clinical restaging. Patients achieving a clinical complete response (cCR) could proceed without cystectomy. The co-primary objectives were to assess the cCR rate and the positive predictive value of cCR for a composite outcome: 2-year metastasis-free survival in patients forgoing immediate cystectomy or <ypT1N0 in patients electing immediate cystectomy. Seventy-six patients were enrolled; of these, 33 achieved a cCR (43%, 95% confidence interval (CI): 32%, 55%), and 32 of 33 who achieved a cCR opted to forgo immediate cystectomy. The positive predictive value of cCR was 0.97 (95% CI: 0.91, 1), meeting the co-primary objective. The most common adverse events were fatigue, anemia, neutropenia and nausea. Somatic alterations in pre-specified genes (ATM,RB1,FANCCandERCC2) or increased tumor mutational burden did not improve the positive predictive value of cCR. Exploratory analyses of peripheral blood mass cytometry and soluble protein analytes demonstrated an association between the baseline and on-treatment immune contexture with clinical outcomes. Stringently defined cCR after gemcitabine, cisplatin, plus nivolumab facilitated bladder sparing and warrants further study. ClinicalTrials.gov identifier:NCT03558087.

Extracellular vesicles (EVs) are heterogeneous entities secreted by cells into their microenvironment and systemic circulation. Circulating EVs carry functional small RNAs and other molecular footprints from their cell of origin, and thus have evident applications in liquid biopsy, therapeutics, and intercellular communication. Yet, the complete transcriptomic landscape of EVs is poorly characterized due to critical limitations including variable protocols used for EV‐RNA extraction, quality control, cDNA library preparation, sequencing technologies, and bioinformatic analyses. Consequently, there is a gap in knowledge and the need for a standardized approach in delineating EV‐RNAs. Here, we address these gaps by describing the following points by (1) focusing on the large canopy of the EVs and particles (EVPs), which includes, but not limited to – exosomes and other large and small EVs, lipoproteins, exomeres/supermeres, mitochondrial‐derived vesicles, RNA binding proteins, and cell‐free DNA/RNA/proteins; (2) examining the potential functional roles and biogenesis of EVPs; (3) discussing various transcriptomic methods and technologies used in uncovering the cargoes of EVPs; (4) presenting a comprehensive list of RNA subtypes reported in EVPs; (5) describing different EV‐RNA databases and resources specific to EV‐RNA species; (6) reviewing established bioinformatics pipelines and novel strategies for reproducible EV transcriptomics analyses; (7) emphasizing the significant need for a gold standard approach in identifying EV‐RNAs across studies; (8) and finally, we highlight current challenges, discuss possible solutions, and present recommendations for robust and reproducible analyses of EVP‐associated small RNAs. Overall, we seek to provide clarity on the transcriptomics landscape, sequencing technologies, and bioinformatic analyses of EVP‐RNAs. Detailed portrayal of the current state of EVP transcriptomics will lead to a better understanding of how the RNA cargo of EVPs can be used in modern and targeted diagnostics and therapeutics. For the inclusion of different particles discussed in this article, we use the terms large/small EVs, non‐vesicular extracellular particles (NVEPs), EPs and EVPs as defined in MISEV guidelines by the International Society of Extracellular Vesicles (ISEV).

Post-acute sequelae of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection are debilitating, clinically heterogeneous and of unknown molecular etiology. A transcriptome-wide investigation was performed in 165 acutely infected hospitalized individuals who were followed clinically into the post-acute period. Distinct gene expression signatures of post-acute sequelae were already present in whole blood during acute infection, with innate and adaptive immune cells implicated in different symptoms. Two clusters of sequelae exhibited divergent plasma-cell-associated gene expression patterns. In one cluster, sequelae associated with higher expression of immunoglobulin-related genes in an anti-spike antibody titer-dependent manner. In the other, sequelae associated independently of these titers with lower expression of immunoglobulin-related genes, indicating lower non-specific antibody production in individuals with these sequelae. This relationship between lower total immunoglobulins and sequelae was validated in an external cohort. Altogether, multiple etiologies of post-acute sequelae were already detectable during SARS-CoV-2 infection, directly linking these sequelae with the acute host response to the virus and providing early insights into their development.