A new technology that analyzes gene expression in cells reveals new insights into immune cell activity. This new single-cell approach doesn’t just capture genetic information, but also cell function. The new technique is known as Cytokine Intracellular Protein High-throughput Expression with RNA (CIPHER)-sequencing and is a flexible workflow of network-based designs to analyze high throughput sequencing data. More specifically, it enables the quantification of proteins and transcriptomes from cells. This novel technique measures RNA and proteins concurrently to provide information on cytokine secretion. The results more accuracy inform scientists about cancer biology, the immune response, and treatment resistance.
The recent article published in Scientific Reports, by Dr. Justin Taylor and others, demonstrates the use of novel CIPHER-sequencing to analyze cytokine profiling in cells. This platform has the capability to improve immunotherapy design and patient response. Taylor is a physician scientist, and Associate Professor in the Division of Hematology at the University of Miami Miller School of Medicine. His clinical practice is at the Sylvester Comprehensive Cancer Center where he investigates genetic mutations in hematological malignancies. Taylor’s lab utilizes several approaches including single-cell genomic technology, animal models, and lab bench assays to drive clinical discoveries. The goal of the lab is to improve treatment outcomes in patients with blood cancer.
The team analyzed stimulated peripheral blood mononuclear cells (PBMCs) to understand and profile cytokine production and metabolic activity. Single cell sequencing allows scientists to analyze thousands of cells at once in real time. Ribonucleic acid (RNA) or genetic information provides information about what the future activity of that cell, while proteins indicate current cell function. By combining genetic information and protein expression, clinicians have a better opportunity to predict which therapies will have most benefit in patients.
Cytokines are secreted proteins that drive communication between cells. Specifically, these small proteins control inflammation, direct cell function, and remodel how tumors behave. However, gaps remain in the field of cytokine biology. RNA expression does not necessarily correlate with the production of protein in a cell. The inconsistency is not an error, but the nature of each cellular component. RNA is transient and disappears quickly, while proteins take more time to be produce. Unfortunately, studying RNA alone can restrict the information available and limit scientific discovery. CIPHER-sequencing fills the gap to capture RNA information.
CIPHER-sequencing measures cell profiles on multiple levels, including RNA across the genome, surface proteins, intracellular proteins, and cytokines within the cell. This new analysis provides a more realistic and in-depth analysis of what is going on in a cell. Another benefit to using CIPHER-sequencing is the limited stress it puts on the cells. Other methods damage cells during preparation, which inherently change the cell’s characteristics and readout – skewing results. CIPHER-sequencing allows scientists to clearly observe true immune behavior compared to stress signals interfering with results and causing technical artifacts.
Researchers found, using CIPHER-sequencing, that RNA signals increased first before protein indicating RNA is necessary before proteins can be generated. This confirms previously known knowledge in the field but helps to establish a working platform for future tests. As RNA and protein are measured simultaneously, researchers can track the step-by-step process of RNA to protein. Overall, this approach helps scientists move beyond educated guesses and helps to fully understand immune response to improve clinical care.
Article, Scientific Reports, Justin Taylor, University of Miami Miller School of Medicine, Sylvester Comprehensive Cancer Center
