Abstract 321: Genome-wide CRISPR screens identify combination strategies for Capivasertib (AZD5363; AKT) and AZD8186 (PI3Kβ/δ) in PTEN-null breast cancer
Authors: Dunn Shanade , Yu Jason , Gris-Oliver Albert , Pilling James , Hopcroft Philip , Yelland Urs , Cureton Natalie , Staniszewsla Anna , Au Yan Zi , Ong Swee Hoe , Isherwood Beverley , Serra Violeta , Barry Simon , Davies Barry R. , Lynch James T. , Yusa Kosuke 
Published: August 2019

Abstract
The PI3K-AKT-mTOR pathway is frequently hyper-activated in breast cancer and several inhibitors targeting the PI3K pathway, including AKT (Capivasertib; AZD5363) and PI3Kβ/δ (AZD8186), are currently in clinical development (phase I/II) targeting tumours with mutational activation of the pathway including those with loss of PTEN. A better understanding of the mechanisms of resistance/sensitization to Capivasertib and AZD8186 in PTEN-null breast cancer is critical to fully exploit the anti-tumour activity of these compounds and to develop combination strategies for more effective therapy. Here we performed genome-wide CRISPR-Cas9 knockout screens to identify genes that when targeted promote resistance (gRNA enrichment) or sensitivity (gRNA depletion) to Capivasertib and AZD8186 in three PTEN-null breast cancer cell lines. Our screens identified five genes (NPRL2, DEPDC5, DDIT4, HNRNPD and ZC3H4) whose inactivation promoted resistance to Capivasertib; two resistance genes (PIK3R2 and INPPL1) for AZD8186 and three resistance genes (TSC2, TSC1 and FIBP) for both compounds across all three cell lines. While this result strongly suggests that mTOR pathway reactivation is the major resistance mechanism, we also identified novel resistance genes such as FIBP. Our screens also identified a number of genes that when inactivated sensitize cells to Capivasertib and AZD8186 treatment. Eighteen of these sensitizers have inhibitors in clinical development and we have performed a combination screen to identify compounds that give the best synergy with Capivasertib/AZD8186. The most striking combination effect was detected with the Mcl-1 inhibitor, AZD5991. Mechanistic work is ongoing and our data suggest that Capivasertib/AZD8186 prime cells for apoptosis by a previously undescribed mechanism and combined inhibition with AZD5991 drives a rapid apoptotic response. Importantly, the combination with AZD5991 remains effective in breast cancer cells with acquired resistance to Capivasertib/AZD8186. These combinations showed synergistic tumour suppressive effect in a PTEN-null TNBC breast cancer xenograft model (AZD5991 + AZD8186/Capivasertib) and activity in a patient-derived xenograft model (AZD5991 + Capivasertib). Overall, our CRISPR screening data provide new insights into the genes and pathways that drive resistance to inhibitors of the PI3K pathway in PTEN-null breast cancer and have identified novel synergistic combinations to maximize therapeutic response.
Citation Format: Shanade Dunn, Jason Yu, Albert Gris-Oliver, James Pilling, Philip Hopcroft, Urs Yelland, Natalie Cureton, Anna Staniszewsla, Yan Zi Au, Swee Hoe Ong, Beverley Isherwood, Violeta Serra, Simon Barry, Barry R. Davies, James T. Lynch, Kosuke Yusa. Genome-wide CRISPR screens identify combination strategies for Capivasertib (AZD5363; AKT) and AZD8186 (PI3Kβ/δ) in PTEN-null breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 321.