Comparative Analysis of Single-Cell RNA Sequencing Methods with and without Sample Multiplexing.

Single-cell RNA sequencing (scRNA-seq) has emerged as a powerful technique for investigating biological heterogeneity at the single-cell level in human systems and model organisms. Recent advances in scRNA-seq have enabled the pooling of cells from multiple samples into single libraries, thereby increasing sample throughput while reducing technical batch effects, library preparation time, and the overall cost. However, a comparative analysis of scRNA-seq methods with and without sample multiplexing is lacking. In this study, we benchmarked methods from two representative platforms: Parse Biosciences (Parse; with sample multiplexing) and 10x Genomics (10x; without sample multiplexing). By using peripheral blood mononuclear cells (PBMCs) obtained from two healthy individuals, we demonstrate that demultiplexed scRNA-seq data obtained from Parse showed similar cell type frequencies compared to 10x data where samples were not multiplexed. Despite relatively lower cell capture affecting library preparation, Parse can detect rare cell types (e.g., plasmablasts and dendritic cells) which is likely due to its relatively higher sensitivity in gene detection. Moreover, a comparative analysis of transcript quantification between the two platforms revealed platform-specific distributions of gene length and GC content. These results offer guidance for researchers in designing high-throughput scRNA-seq studies.

A phase I study of an adenoviral vector delivering a MUC1/CD40-ligand fusion protein in patients with advanced adenocarcinoma.

Cancer vaccines as immunotherapy for solid tumours are currently in development with promising results. We report a phase 1 study of Ad-sig-hMUC1/ecdCD40L (NCT02140996), an adenoviral-vector vaccine encoding the tumour-associated antigen MUC1 linked to CD40 ligand, in patients with advanced adenocarcinoma. The primary objective of this study is safety and tolerability. We also study the immunome in vaccinated patients as a secondary outcome. This trial, while not designed to determine clinical efficacy, reports an exploratory endpoint of overall response rate. The study meets its pre-specified primary endpoint demonstrating safety and tolerability in a cohort of 21 patients with advanced adenocarcinomas (breast, lung and ovary). The maximal dose of the vaccine is 1 ×1011 viral particles, with no dose limiting toxicities. All drug related adverse events are of low grades, most commonly injection site reactions in 15 (71%) patients. Using exploratory high-dimensional analyses, we find both quantitative and relational changes in the cancer immunome after vaccination. Our data highlights the utility of high-dimensional analyses in understanding and predicting effective immunotherapy, underscoring the importance of immune competency in cancer prognosis.