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In vitro profiling of rheumatic-disease-evaluated JAK inhibitors demonstrate differences in JAK isoform selectivity between different types of inhibitors

Arthritis Rheumatol. 2023 May 3. doi: 10.1002/art.42547 Aug. 18, 2023, midnight

Presented By

Dr Walter Maksymowych MB ChB, FRCP(C), FACP
Chief Medical Officer, CARE Arthritis; Professor of Medicine, University of Alberta

Guest Commentary By:

Dr. Joerg Ermann MD
Brigham and Women’s Hospital

Synopsis

Introduction
  • JAK inhibitor selectivity forms the basis for understanding differences in efficacy and safety of JAKinibs. Filgotinib and upadacitinib are considered to be more JAK1 selective.
  • Evaluation has been hampered by the lack of comprehensive JAKi head-to-head studies and assay-to-assay variation in individual studies.
Methods
  • 10 JAKi (5 globally approved for rheumatic diseases (pan-JAKi tofacitinib, baricitinib, peficitinib and JAK1i filgotinib and upadacitinib)) were analyzed for JAK-isoform selectivity by assaying:
    1. inhibition of catalytic activity
    2. binding to kinase and regulatory pseudokinase domains
    3. inhibition of cytokine signaling in peripheral blood of healthy volunteers
    4. inhibition of STAT phosphorylation at clinically relevant JAKi concentrations in PBMC of RA patients and healthy donors.
Results
  • In vitro inhibition of kinase activity, pan-JAKi (tofacitinib, baricitinib) targeted JAK1-2 with high potency and comparably with JAK1 selective agents. Filgotinib and upadacitinib also targeted JAK2 (2-fold selectivity for JAK1). TYK2 and JAK1 was targeted by deucravacitinib and beprocitinib (also JAK1).
  • In human leukocytes, JAKi predominantly inhibited JAK1-dependent cytokines IL-2, IL-6, and interferons. Inhibition of these cytokines was more pronounced in RA cells compared to healthy controls.
  • Pan-JAK and JAK1-targeted JAKi showed similar inhibition profiles at clinically applied dose in PBMCs of RA but the inhibition levels varied (Figure).

    Tap/click on the image for a larger view
    Figure.Cytokine inhibition profiles of JAKi in immune cells of RA patients. Inhibition percentages of cytokine-induced STAT phosphorylation for clinically relevant concentrations of JAKi. IL-2 and IFN-α inhibition was measured in CD4+ T cells, and IFN-γ and GM-CSF inhibition in CD33+ monocytes.

Commentary

This study was designed to provide a comprehensive in vitro analysis on direct JAK-related effects (JAK selectivity, cytokine inhibition profile in heathy and in patient cells) and comparison of JAKi evaluated for the treatment of rheumatic diseases. Inhibition of JAK kinase activity did not directly translate into cellular inhibition of JAK-STAT signaling. Despite differences in JAK-selectivity, the cytokine inhibition profiles of currently approved JAKi were highly similar with preference for targeting inhibition of JAK1-mediated cytokines. This basic work might explain why clinical efficacy data for tofacitinib and upadacitinib appears similar in phase III trials of bioDMARD-naïve radiographic axSpA. But does not shed light on the mechanism of action of JAKi in axSpA since we know from prior trials in AS that targeting IL6 is not beneficial.

Special Guest Commentary

JAK inhibitors are mysterious drugs. Despite the fact that they are designed to target a well-known enzymatic activity, we still don’t know why they work in axial spondyloarthritis or how to explain some of their side effects. The study by Virtanen et al. shows that the JAK inhibitors currently approved for rheumatic disease indications performed largely similar when tested in vitro for their ability to block cytokine receptor signaling in immune cells. Most JAK inhibitors work by binding non-covalently to the ATP binding site of the kinase domain. Ritlecitinib and Deucravacitinib have unique JAK targeting mechanisms giving them significantly higher selectivity for JAK3 and TYK2, respectively, and distinct cytokine receptor inhibition profiles in vitro. Whether a higher selectivity for individual JAKs results in better efficacy or reduced side effects in vivo remains to be shown.

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