Abstract Details

BTK is a member of the TEC family of non-receptor tyrosine kinases expressed in cells of hematopoietic origin, including B lymphocytes and myeloid cells, but not T or NK cells. Selective BTK inhibitors (BTKi) have shown efficacy in Phase 2 trials in multiple sclerosis (MS).

We investigated the role of Bruton’s tyrosine kinase (BTK) in human and mouse myeloid cells by in vitro and in vivo studies.

The impact of a BTK inhibitor (BTKi) on markers for myeloid cell functions (activation, costimulation, adhesion and phagocytosis) was evaluated in PBMC cultures from healthy and MS subjects. The BTKi evobrutinib or vehicle was administered to MOG_35-55 peptide-immunized C57BL6 mice starting three days after disease onset. Mice were assessed for clinical signs, and frequency of TNF-producing macrophages by flow cytometry. BTK was inactivated by crossing mice carrying a floxable BTK allele with mice carrying the Tamoxifen (Tam)-inducible Cre-recombinase under Cx3CR1 promoter. Double-transgenic mice received repeated Tam injections, a 38-day washout, to allow reconstitution of short living Cx3CR1-positive cells expressing BTK, and then EAE immunization.

BTKi supported monocyte expression of VLA4/CD49d, an integrin directing immune cell migration towards the central nervous system, and CD163, a well-known M2 marker potentially involved in removal of myelin debris. This effect was maintained under all tested inflammatory settings and replicated with PBMC from MS subjects. Animals treated with evobrutinib showed significant clinical amelioration. Analysis of CNS-infiltrating myeloid cells indicated lower frequency of TNF-producing macrophages in evobrutinib-treated animals. BTK deletion significantly reduced disease severity, demyelination, and axonal damage, demonstrating a role for BTK in tissue-resident myeloid cells in experimental MS.

Overall, these experiments demonstrate that human and mouse myeloid cells may represent a cellular target of BTK inhibitors and that BTK in long-lived myeloid cells supports the expression of neuroinflammation.