Abstract LB-263: Preclinical efficacy of the ATR inhibitor AZD6738 in combination with the BTK inhibitor acalabrutinib in ABC-DLBCL models
Authors: Young Lucy A. , Delpuech Oona , Willis Brandon , Bussey Alexandra , Wilson Zena , Dupont Michelle , Grajales Carlos , Bloecher Andrew , Covey Todd , Wills Kate , Lau Alan , Hollingsworth Simon J. 
Published: August 2018

Abstract
Bruton tyrosine kinase (BTK) is an essential kinase in the B-cell receptor (BCR) signalling pathway. Acalabrutinib is a potent and highly selective irreversible BTK inhibitor that received accelerated approval by FDA for the treatment of Relapsed/Refractory Mantle Cell Lymphoma. It is proposed to be combined with the Ataxia telangiectasia and Rad3-related (ATR) kinase inhibitor AZD6738, in a Phase I/II proof of concept clinical trial to assess safety and efficacy in patients with B-cell malignancies. ATR is a key regulator of DNA replication, repair and cell cycle checkpoints. The Activated B-Cell (ABC) subtype of Diffuse Large B-Cell Lymphoma (DLBCL) is associated with chronic active BCR signalling, and given our previous identification of sensitivity to AZD6738 in ABC-DLBCL cell lines, we hypothesised that combination with acalabrutinib may have additional therapeutic benefit. Here we report preliminary data to support combination efficacy in models of ABC-type DLBCL. The growth inhibitory and cell kill effect of AZD6738 and acalabrutinib in combination was assessed in 3 ABC and 4 GCB DLBCL cell lines. Combination activity and cell kill was detected specifically in the ABC-type TMD8 cell line. Using flow cytometry, we show that the cell kill effects of the drug combination were dependent on dose and schedule. Single agent treatment with 5 nM acalabrutinib or 0.5 µM AZD6738 for 72 h, caused cell death in 12% and 17% of cells respectively. The equivalent doses given in combination for 72 h induced cell death in 39% of cells, indicative of a greater than additive effect. Cell killing was increased with longer co-exposure to the combination, shown by an increase in cell death from 18% at 24 h, to 30% at 48 h. Acalabrutinib is dosed on a BID schedule, thus we tested whether its activity in combination with AZD6738 was affected by pre- or post-addition of acalabrutinib. A 24 h pre-treatment of cells with acalabrutinib, followed by the combination of AZD6738 and acalabrutinib for 48 h resulted in 20% cell death, whereas 48 h combination treatment followed by 24 h acalabrutinib monotherapy caused a similar 27% increase in death, suggesting that combination efficacy is not dependent on sequence of drug administration. Furthermore, the mRNA expression of BTK target genes in ABC-DLBCL cell lines was modulated by acalabrutinib, however there was no additional modulation to this pathway by the combination with AZD6738, suggesting the two agents kill cells through different mechanisms of action. In the TMD8 xenograft mouse model, daily dosing of AZD6738 (25 mg/kg) with twice daily dosing of acalabrutinib (20 mg/kg) was well tolerated and resulted in complete and durable tumor regressions (8/8 tumors), whereas single agent treatments resulted in tumor growth delay. Collectively, our preliminary data support a rationale for combining BTK and ATR inhibitors for an effective and alternative treatment option in ABC-DLBCL.
Citation Format: Lucy A. Young, Oona Delpuech, Brandon Willis, Alexandra Bussey, Zena Wilson, Michelle Dupont, Carlos Grajales, Andrew Bloecher, Todd Covey, Kate Wills, Alan Lau, Simon J. Hollingsworth. Preclinical efficacy of the ATR inhibitor AZD6738 in combination with the BTK inhibitor acalabrutinib in ABC-DLBCL models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr LB-263.