August 19, 2022, Zhang Tiantai & Sun Lan's research group of the Institute of Materia Medica, Chinese Academy of Medical Sciences and Zhang Dayong's research group of China Pharmaceutical University jointly published their research results in Science Advances, titled A highly selective JAK3 inhibitor is developed for treating rheumatoid arthritis by suppressing γc cytokine–related JAK-STAT signal.
Autoimmune disease occurs when the regulatory mechanism of the immune system in an individual fails and that causes the immune system to attack self-antigens, including a series of diseases such as rheumatoid arthritis (RA), psoriasis, and alopecia areata. However, its pathogenesis is still unclear and there is a lack of effective therapeutic drugs. In 2012, with the launch of the drug tofacitinib for the treatment of RA, a new target for the treatment of autoimmune diseases, Junas kinase (JAK), was identified. JAK has become a confirmed new target for the treatment of autoimmune diseases and an ideal target for the development of autoimmune disease drugs. Currently, seven JAK small molecule inhibitors have been marketed for the treatment of different autoimmune diseases. However, with the clinical application of JAK inhibitors, side effects such as opportunistic infections and anemia have gradually emerged, mainly due to the non-selective inhibition of different isoforms of JAK kinase. JAK kinase includes four isoforms, JAK1, JAK2, JAK3 and Tyk2. The JAK-STAT signaling pathway composed of downstream effector molecule STAT (signal transducers and activators of transcription) regulates the inflammatory and immune signals of more than 50 cytokines in the body. Non-selective inhibition of JAK is bound to affect the physiological effects of various cytokines, resulting in corresponding side effects. Therefore, the development of highly selective JAK inhibitors is the current trend and direction of the development of such drugs.
The selectivity of kinases is a common difficulty in the development of kinase inhibitors. Among the JAK kinase subtypes, the JAK3 subtype only regulates the γc receptor subunit-related IL-2, IL-4, IL-7, IL-9, IL-15, IL-21 and other 6 cytokines, compared with other JAK subtypes that regulate multiple cytokines, selective inhibitors targeting JAK3 subtypes can minimize the effects of JAKs-related cytokines. Infections and other side effects caused by extensive inhibition of the drug, but there may also be a risk of less inhibitory cytokines and weak anti-inflammatory effects or drug efficacy. JAK2 subtypes mainly regulate hematopoietic system functions such as hemoglobin and platelet production, so inhibition of JAK2 subtypes will cause side effects such as anemia, which is why JAK2 inhibitors treat multiple myeloma. The authors' strategy to target JAK3 subtypes to develop small molecule inhibitors is expected to avoid the occurrence of opportunistic infection side effects by reducing broad inhibition of cytokines on the one hand, and reducing the risk of adverse effects due to JAK2 by increasing selectivity for JAK3 on the other hand. Inhibition of hematopoietic system side effects.
Based on the structural characteristics of JAK3 isoforms, the researchers designed, synthesized, screened and found a lead compound Z583 with strong inhibitory activity against JAK3 isoforms. Its IC50 for inhibiting the phosphorylation of JAK3 reaches 0.1nM (Km ATP concentration), and its selectivity for JAK3 is more than 4500 times that of other isoforms, showing very high selectivity. Studies on signaling pathways at the cellular level have shown that Z583 only has a clear inhibitory effect on STAT phosphorylation induced by six γc cytokines regulated by JAK3 isoforms, but has no inhibitory effect on STAT phosphorylation induced by other isoform-related cytokines , further confirmed the highly selective inhibition of JAK3 by Z583. In order to clarify the binding characteristics of Z583 and JAK3, the researchers confirmed through molecular docking, Washout experiments and protein profiling analysis that Z583 targets the ATP activity binding pocket of the JAK3 isoform and forms a covalent covalent relationship with the cysteine residue at position 909 (Cys909). combine. The authors further investigated the possible mechanism of Z583 targeting JAK3 isoforms to inhibit the maturation of DCs, the proliferation and differentiation of Th1 and Th17, and play an anti-inflammatory role.
Further, the researchers investigated the therapeutic effect of Z583 by establishing a classical collagen-induced RA mouse disease model. The results showed that Z583 significantly inhibited the progression and onset of the disease at a dose of 3 mg/kg for 3 weeks, and at a dose of 10 mg. Oral administration of /kg dose for 4 weeks can significantly reverse the existing joint swelling, showing a very strong anti-RA effect. In addition, the authors also examined the safety of Z583. The hERG assay and mini-Ames assay showed no cardiac or mutagenic effects. Importantly, the subacute toxicity assay in rats administered orally at high doses for 4 consecutive weeks showed that Z583 had no blood Abnormalities of routine, liver and kidney function, and organ pathology. The above in vivo studies confirmed the safety and efficacy of the selective JAK3 small molecule inhibitor Z583.
To sum up, this study found a novel and highly selective small molecule inhibitor of JAK3 against JAK, an important target for the treatment of autoimmune diseases. Most of the first and second generation JAK inhibitors that have been marketed have poor selectivity , there is no selective JAK3 inhibitor on the market. The selective JAK3 small molecule inhibitor Z583 discovered in this study shows great potential in the treatment of autoimmune diseases such as rheumatoid arthritis.
The first author of the paper is Dr. Chengjuan Chen, assistant researcher from the Chinese Academy of Medical Sciences. Researcher Tiantai Zhang and Lan Sun from the Institute of Materia Medica, Chinese Academy of Medical Sciences and Professor Dayong Zhang from China Pharmaceutical University are the co-corresponding authors.
Link: https://www.science.org/doi/10.1126/sciadv.abo4363
