Introduction
Inflammatory cell recruitment is a key step in the initiation of the acute immune response. Macrophage activation at the site of tissue injury results in the secretion of chemokines; these have a non-redundant role in leukocyte recruitment in pre-clinical models of inflammation. Activated macrophages are a major source of pro-inflammatory cytokines and chemokines in many chronic inflammatory diseases including rheumatoid arthritis, diet-induced diabetes or atherosclerosis (White et al., 2013). Chemokine-mediated recruitment, retention and activation of leukocytes, are attractive areas for the development of novel anti-inflammatory agents that could find application in a wide range of chronic inflammatory pathologies. We and others have demonstrated that genetic targeting of chemokine receptors shows promise in pre-clinical models of acute and chronic inflammation (McNeill et al., 2017). However, interventional studies in man using chemokine receptor antagonists have so far had limited therapeutic benefit (Schall and Proudfoot, 2011) (Noels et al., 2019). This could be due to high redundancy of chemokine receptor use in leukocytes to initiate monocyte/macrophage migration. Therefore, a therapeutic strategy that would affect both a myeloid cell’s ability to undergo chemotaxis and reduced chemokine secretion could limit leukocyte recruitment by two independent but complementary mechanisms.
Bruton’s tyrosine kinase (BTK) is a non-receptor bound intracellular signalling molecule best known for its signalling in B-cell development and malignancy. However, in recent years an alternative role for BTK is emerging in innate immune cells. BTK has been shown to be expressed at relatively high levels in myeloid cells (Mangla et al., 2004). BTK is activated in monocytes and macrophages in numerous acute inflammatory conditions, including polymicrobial sepsis and cerebral ischaemia (O’Riordan et al., 2019a)(Ito et al., 2015) but also in chronic inflammatory conditions such as obesity-induced diabetes, rheumatoid arthritis and lupus (Purvis et al., 2020)(Honigberg et al., 2010)(Hartkamp et al., 2015). During the COVID-19 pandemic BTK has been shown to be activated in monocytes and intervention with the BTK inhibitor Acalabrutinib has been reported to reduce systemic inflammation in patients with severe COVID-19 (Roschewski et al., 2020). We and others have reported inhibition of BTK reduces NF-kB and the NLRP3 inflammasome activity in both murine and human macrophages, and reduces pro-inflammatory cytokine and chemokine production (O’Riordan et al., 2020)(Mao et al., 2020).
Pharmacological inhibition of BTK reduced macrophage accumulation in inflamed tissues in a pre-clinical model of type II diabetes (Purvis et al. 2020) and a model of cerebral ischaemia (Ito et al 2015). To confirm and extend these studies we tested the hypothesis that BTK can directly regulate myeloid cell recruitment to sites of inflammation. To do this we used a combination of in vivo and in vitro cell recruitment assays using a range of European medicines agency (EMA) or US Food and Drug administration (FDA) approved BTK inhibitors and tool compounds in combination with BTK-deficient X-linked immunodeficient (XID) mice. This allowed us to fully explore the magnitude and kinetics of myeloid cell recruitment in acute inflammation. We demonstrate that inhibition of BTK activity reduces neutrophil and monocyte recruitment via two independent but complementary mechanisms a) reducing monocyte chemotaxis, and b) reducing chemokine production from tissue resident macrophages.