Abstract 4795: Development of novel, selective, reversible inhibitors equipotent against wild-type and C481S Bruton's tyrosine kinase (BTK)
Authors: Guisot Nicolas E S , Walker Victoria , Best Stuart A. , Abet Valentina , Chappell Rose , Emmerich Juliette , Ho Kelvin , Kelly James R. , Lyons Kristina , Müller Melanie , Refuerzo Julienne , Sargent Louise , Talab Fatima , Waldron Madelene , Bingham Matilda , Campbell Mary-Ann , Phillips Caroline , Armer Richard 
Published: July 2016

Abstract
Redx Oncology has developed novel, differentiated, reversible small molecule inhibitors of BTK, combining current best-in-class potency with improved selectivity profiles, which are suitable for oral, once daily dosing, designed to be equipotent against wild-type and C481S BTK.
BTK is a member of the src-related Tec family of cytoplasmic tyrosine kinases. BTK plays a key role in the BCR signaling pathway, which is required for the development, activation and survival of B-cells. BTK inhibitors have therefore been developed with the aim of treating B-cell malignancies that are dependent on BCR signaling, such as CLL and NHL. Ibrutinib is an irreversible BTK inhibitor that has been approved for the treatment of CLL, MCL and WM.
Irreversible and covalent reversible BTK inhibitors specifically target a cysteine residue C481 within BTK. Following treatment with ibrutinib, cases of secondary resistance have emerged in both CLL and MCL patients. Acquired mutations within BTK such as C481S, C481Y, C481R, C481F have been reported in the literature and clearly interfere with covalent drug binding. It has been predicted that the incidence of observed resistance will increase as clinical use outside clinical trials expands over time. Here, we present our reversible BTK inhibitor series demonstrating subnanomolar binding affinity for WT and C481S forms of BTK, that inhibits formation of pBTK in both WT and C481S BTK expressing cells. In addition, the compound demonstrated significant in vitro potency against a number of lymphoma cell lines including inhibition of BCR signaling and proliferation in OCI-Ly10 cells at nanomolar concentrations. To further investigate the binding nature of these compounds, PBMC wash out studies, measuring CD69 as a marker of B-cell activation, were used to highlight the reversible activity of these compounds.
Some BTK inhibitors also inhibit ITK, which plays a critical role in FcR-stimulated NK cell function that is required for ADCC. ADCC is the mechanism that anti-CD20 antibodies, such as rituximab, are believed to activate, and ibrutinib has been shown to antagonize this mechanism in vitro. As rituximab-combination chemotherapy is today's standard of care in B-cell malignancies, it would be desirable to have a BTK inhibitor with high selectivity for BTK over ITK.
At 1 μM, our lead compound is highly selective when tested against 469 kinase and did not show significant inhibition against other kinases involved in BCR signaling (e.g. Syk, Lyn). Furthermore, the compound has high selectivity versus structurally related cysteine-containing kinases (including EGFR and ITK) both in enzyme and cellular assays.
Our lead compound has a favorable in vitro safety profile and drug-like properties, displaying an improved CYP profile to competitor compounds. In vivo PK demonstrated good oral bioavailability and in vivo efficacy has been demonstrated.
Citation Format: Nicolas E S Guisot, Victoria Walker, Stuart A. Best, Valentina Abet, Rose Chappell, Juliette Emmerich, Kelvin Ho, James R. Kelly, Kristina Lyons, Melanie Müller, Julienne Refuerzo, Louise Sargent, Fatima Talab, Madelene Waldron, Matilda Bingham, Mary-Ann Campbell, Caroline Phillips, Richard Armer. Development of novel, selective, reversible inhibitors equipotent against wild-type and C481S Bruton's tyrosine kinase (BTK). [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 4795.