Abstract LB-105: High-throughput target identification using CRISPR/Cas9

One of the major challenges of the drug discovery process is the identification of novel, validated targets, whose pharmacological modulation may yield the desired therapeutic outcomes. The use of CRISPR (clustered regularly interspaced short palindromic repeat)-associated Cas9 nuclease and guide RNA can potentially impact the entire drug discovery process from generation of models based on clinical findings through to the target identification. The ability of CRISPR/Cas9, to efficiently and precisely edit a cell9s DNA and introduce a complete genetic knockout, while minimizing off-target effects, offers an improved approach to target identification. Moreover, the ability to scale this approach through the generation of genome wide CRISPR libraries, coupled with the use of lentiviral delivery methods enables high-throughput (HTP) loss-of-function screens to be performed rapidly and identify genes whose activity is important for the specific endpoint being measured. For example, in this study, we have used CellSensor® cell lines targeting various signaling pathways (AP-1, c-Fos and NF-kB) to identify key targets critical to these pathways. Transcription factors like AP-1, c-Fos and NF-kB have shown to play an important role in cancer initiation and progression. Using the key signaling molecules along these pathways as controls we have data that supports HTP target identification. Furthermore, we will present our results from screening of a subset of a CRISPR library targeting 160 different kinases. Eventually, we plan to screen the entire CRISPR library targeting 750 kinases with 4 gRNA per gene against each of these signaling pathways. Similar screenings can be performed using other functional assay formats, like the cell health assays or high content imaging. These screenings can provide a wealth of data on the normal functioning of a target and in turn, should yield better validated targets for progression into full drug discovery. Ultimately, the hits from the HTP screenings can be followed up by using CRISPR technology to generate animal knockout models that would support translating the screen to the pre-clinical trials. This in turn could provide better correlation to the clinical setting and thereby reduce candidate drug attrition. Citation Format: Chetana M. Revankar, Julia Braun, LaiYee Wong, Justin Wetter, Jian-Ping Yang, Namritha Ravinder, Jon Chesnut, David Piper. High-throughput target identification using CRISPR/Cas9. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr LB-105.