Remibrutinib, a Novel Bruton’s Tyrosine Kinase Inhibitor, Exhibits Improved Target Selectivity and Potency In Vitro
Bernd Kieseier1, Bruno Cenni2, Robert Pulz2, Daniela Angst2, Denis Eichlisberger2, Marina Ziehn1
1Novartis Pharma AG, 2Novartis Institutes for Biomedical Research
To assess the potency and selectivity of Bruton’s tyrosine kinase inhibitors (BTKis) under comparable experimental conditions.
BTK is a key node in B-cell receptor and Fc receptor signaling. BTKis are an emerging oral option for patients with multiple sclerosis (MS). Several covalent and reversible BTKis are in clinical development for the treatment of MS. For covalent enzyme inhibitors, in vitro assays are influenced by experimental conditions and are time dependent.
In vitro binding of covalent BTKis was assessed over time and concentration in human blood. Inhibition of B cells and basophils, including the impact of drug washout on B-cell inhibition, was assessed for covalent and reversible BTKis. Kinase selectivity was studied (first screened kinome wide, then using the dissociation constant for select kinases) via a binding assay to allow for the direct comparison of covalent and reversible BTKis.
Covalent inhibitors demonstrated time- and concentration-dependent BTK binding in human blood; the IC50 at 1 hour was 21 nM for remibrutinib, 508 nM for evobrutinib, 165 nM for tolebrutinib, and 427 nM for orelabrutinib. These values correlated with B-cell inhibition; the IC50 was 18 nM for remibrutinib, 320 nM for evobrutinib, 74 nM for tolebrutinib, 185 nM for orelabrutinib, and 15 nM for fenebrutinib (reversible). Basophil inhibition potency was comparable. B-cell inhibition by remibrutinib (covalent) was not sensitive to washout, but B-cell inhibition by fenebrutinib was. Kinome selectivity screening at 1 µM showed the following ranking (from least to most off-target kinase binding): remibrutinib, fenebrutinib, evobrutinib, orelabrutinib, and tolebrutinib. A quantitative assessment of binding constants to a subset of kinases confirmed this ranking. 
BTKis undergoing development for MS exhibit varying selectivity across the human kinome, with the highest level of selectivity observed for remibrutinib. This distinction may translate into differences in clinical efficacy and safety.