Category: Assay Development and Screening

1372-C - Spinach Splice Sensor: A Fluorescent Drug Discovery Platform

Tuesday, February 6, 2018
2:00 PM - 3:00 PM

RNA splicing plays a central role in the generation of proteome diversity and in gene regulation. Splicing impacts vital cellular processes, such as cell-fate and differentiation, acquisition of tissue-identity, and organ development. Thus, defects in splicing has been linked to many diseases, including spinal muscular atrophy, Duchenne muscular dystrophy, Parkinson's disease, and several types of cancer. RNA splicing is even more relevant in cancer due to the higher incidence of mis-splicing events than in normal cells. Thus, it is not surprising that aberrant splicing has been linked to important hallmarks of cancer such as proliferation, proliferation, apoptosis evasion, metastasis, and angiogenesis, and in the development of cellular resistance against cancer therapeutics. Despite the significance of splicing in cancer and other diseases, drug discovery efforts targeting them are far and few. A critical bottleneck for such efforts is the lack of robust high-throughput assay tools to monitor endogenous spliced RNA in the cell. Even though there are excellent tools such as RT-qPCR and RNA-seq to study RNA splicing, they are very expensive and not readily adaptable for high-throughput screening (HTS) due to their complex and time-consuming methodology. While splice-mini-gene method offers the advantage of higher-throughput, it lacks in the ability to monitor the endogenous target RNA due its artificial design. Thus, there is an unmet need for simple and robust HTS-ready assay tools to monitor RNA splicing. Addressing this market need, Lucerna has developed of an easy-to-use, HTS-ready splice sensor platform that can specifically detect a spliced RNA isoform of interest. For example, we developed a pair of splice sensors that target pyruvate kinase isoforms M1 (PKM1) and M2 (PKM2). PKM isoform switching is one of the ways cancer cells reprogram their glycolytic pathways to meet cellular demands of energy and biosynthetic intermediates for growth and proliferation. Thus, targeting the alternative splicing of PKM is a new and untapped approach in cancer drug discovery. Validation experiments of the PKM HTS splice platform showed rapid response times (<30 min), high selectivity (300-fold specific fluorescence), sensitivity (can detect 10% changes in splicing), and extended readout window (16h). Further, the modular nature of the splice sensor platform enables easy customization to detect any RNA isoform of interest. In fact, we are also developing splice assays against mRNA isoforms involved in neurodegenerative diseases and circular RNAs that are known biomarkers for cancer. In summary, the Spinachâ„¢ splice sensor platform offers robust and homogenous assays for high-throughput drug screens and enable the discovery of a previously intractable drug targets.

Karen Wu

Lucerna, Inc.
Brooklyn, NY

Karen is a Co-Founder and the President at Lucerna. She has over 15 years of experience in RNA technology and cell biology and her work has been published in Science, Cell, and Nature.