Background/Purpose: The Non-receptor tyrosine kinases, JAK3 and ITK are key regulators of cytokine pathways and are important clinically validated targets, which offer the potential for developing targeted therapeutics in treating Rheumatoid Arthritis (RA) and other immunological, inflammatory and autoimmune diseases such as systemic lupus erythematosus, psoriasis, psoriatic arthritis, atherosclerosis, ulcerative colitis and crohns disease. Development of selective small-molecule inhibitors for both JAK3 and ITK is challenging due to the highly conserved ATP binding pocket within the Janus and TEC-kinase family members. Three variable amino acids within the ATP binding pocket of JAK3 and ITK were targeted and inhibitors were rationally designed by employing FIELDS platform. The FIELDS assisted in identifying fragments/scaffolds to a series of lead compounds, and SAR efforts lead to the discovery of the first-in-class irreversible dual JAK3 and ITK inhibitor ARN-4079 for the treatment of RA.

Methods: The JAK3 and ITK enzymatic inhibition and selectivity studies were performed at DiscoveRx. The cell-based profiling using SelectScreen was performed at Life Technologies. In vivocytokine and CIA efficacy studies were conducted using BALB/c and DBA/1OlaHsd mouse models.   

Results: Fragment-based discovery and lead optimization of a series of highly selective JAK3 and ITK inhibitors in tandem with control of physicochemical and ADME-Tox properties culminated in selecting clinically ready ARN-4079 from ~300 new chemical entities. ARN-4079 potently inhibited JAK3 and ITK activity with an IC₅₀ = 5 nM, and 33 nM in biochemical kinase assay. The ScanKIENTIC Kds were estimated to be 2.5 and 23 nM. ARN-4079 exhibited over 100-fold cellular selectivity within the JAK family and potently inhibited IL-4 induced STAT6 phosphorylation with an IC₅₀ = 70 nM. Its low mM inhibition in SelectScreen IL-6, IFN-γ, and EPO supported the JAK3 selectivity and inhibition of PLCγ1-mediated calcium release from CD4⁺T cells (EC₅₀ = 630 nM) via TCR engagement supports its ITK selectivity in cells. In a set of in vivo experiments, ARN-4079 potently inhibited IL-2, IL-4 and IFN-λ production in mice which are the key characteristics supporting its dual activity. Additionally, ARN-4079 was optimized to have many drug like characteristics in terms of solubility, cell permeation, and DMPK properties. ARN-4079 is an orally available entity (%F = 23), demonstrated efficacy 84% at 60 mg/kg which was equivalent to Tofacitinib in the mouse CIA model. PK studies indicated that, after oral dosing of ARN-4079 reached plasma levels (1080 ng/mL) that are higher or comparable to the therapeutic dose of Tofacitinib (627 ng/mL). Moreover, ARN-4079 showed no toxicity up to doses of 300 mg/kg in rats. On the basis of its strong in vivoefficacy and dose tolerability, ARN-4079 was selected for IND enabling studies.

Conclusion: ARN-4079 is a novel, potent, selective small molecule irreversible dual target inhibitor of JAK3 and ITK that has demonstrated dose linear pharmacokinetic, and in vivo efficacy in CIA models on par with Tofacitinib. Safety and IND studies are ongoing to develop ARN-4079 as a therapeutic agent for RA.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/discovery-of-arn-4079-a-potent-orally-available-dual-target-inhibitor-of-janus-kinase-3-jak3-and-interleukin-2-inducible-t-cell-kinase-itk-for-rheumatoid-arthritis/