Head of Laboratory, AZ-CRUK Functional Genomics Centre AstraZeneca
Disclosure: Disclosure information not submitted.
The identification of novel therapeutic targets that translate into clinical successes is needed now more than ever to deliver life changing medicines to patients across disease areas; from drug resistance in oncology through to cardiovascular and respiratory disease. In this effort the unbiased identification of targets through perturbation at the gene level is not new. However, the CRISPR/Cas9 revolution has enabled us to achieve this with higher efficiency, reduced off-target effects and has enabled new modes of perturbation such as gene activation (CRISPRa) and SNP mutation (base editing). We have used this technology to build a Target Discovery platform encompassing both pooled and arrayed screening techniques that, taken together, allow us to probe a wide range of biology, from the slow onset of drug resistance seen in oncology (pooled), through to understanding what role genes play in signalling and cell response (arrayed). Here we present the development and application of our Arrayed CRISPR Screening component of this platform. Using CRISPR libraries comprised of synthetic gRNAs arrayed into a “one-gene-per-well” format we demonstrate high efficiency of both gene knock out (CRISPRn) and gene activation (CRISPRa) at whole genome scale in cell-based assays. Coupled with the generation of high quality Cas9 expressing cells through the ObLiGaRe insertion of an inducible Cas9 expression cassettes (ODin), we show that CRISPR/Cas9 is a powerful and robust technology for arrayed screening. By combining this platform with high-content imaging and multivariate analysis technology we have been able to interrogate the phenotype resulting from gene perturbation to a much greater depth. This has significantly improved how we are able to rank, triage and progress hits into target validation and into the clinic. Here we demonstrate the application of this platform for the unbiased discovery of new therapeutic targets with two case studies that exemplify the capability and how it has impacted our new target pipeline in both oncology and advanced drug delivery.