Novel TYK2 Inhibitors with an N-(Methyl-d (3))pyridazine-3-carboxamide Skeleton for the Treatment of Autoimmune Diseases.

pubs.acs.org/acsmedchemlett Letter Novel TYK2 Inhibitors with an N‑(Methyl‑d3)pyridazine-3carboxamide Skeleton for the Treatment of Autoimmune Diseases Fei Liu,§ Bin Wang,§ Yanlong Liu, Wei Shi, Xujing Tang, Xiaojin Wang, Zhongyuan Hu, Ying Zhang, Yahui Guo, Xiayun Chang, Xiangyi He, Hongjiang Xu, and Ying He* Cite This: ACS Med. Chem. Lett. 2022, 13, 1730−1738 ACCESS Metrics & More Read Online Article Recommendations sı Supporting Information * ABSTRACT: Tyrosine kinase 2 (TYK2) mediates the interleukin-23 (IL-23), IL-12, and type I interferon (IFN)-driven signal responses that are critical in autoimmune diseases. Here, a series of novel derivatives with an N-(methyl-d3)pyridazine-3carboxamide skeleton that bind to the TYK2 pseudokinase domain were designed, synthesized, and evaluated. Among them, compound 30 demonstrated more excellent inhibitory potency against STAT3 phosphorylation than the positive control deucravacitinib. In addition to JAK isoform selectivity, compound 30 exhibited good in vivo and in vitro pharmacokinetic properties. Furthermore, compound 30 was orally highly effective in both IL-23-driven acanthosis and anti-CD40-induced colitis models. Together, these findings support compound 30 as a promising candidate for therapeutic applications in autoimmune diseases. KEYWORDS: TYK2, pseudokinase, inhibitor, autoimmune diseases A mechanism.26−28 Based on this mechanism, deucravacitinib achieves higher selectivity for TYK2 than JAK 1−3, avoiding undesirable side effects resulting from the high homology of the adenosine triphosphate (ATP) active site (JH1) within the JAK family.13,29 Besides, BMS-986202 with a six-membered pyrimidine group is also a TYK2 JH2 inhibitor and in a phase II clinical trial.30 While brepocitinib31,32 and PF-0682664733 are also under development, they both bind to the TYK2 JH1 domain. Herein, to discover structurally diverse TYK2 inhibitors that bind to the JH2 domain, 28 new compounds have been designed, synthesized, and tested with various biological assays. Most compounds have moderate to good potencies in the TYK2 JH2 binding affinity assay and the inhibition of STAT3 utoimmune diseases are characterized by dysregulated cytokine signaling. Targeting cytokines or their receptors has proved to be effective in multiple autoimmune diseases. Many pathogenic cytokines transmit signals through the JAKSTAT pathway.1−5 As a member of the JAK family, tyrosine kinase 2 (TYK2) regulates downstream signal pathways of the interleukin-23 (IL-23), IL-12, and type I and III interferon (IFN) receptors,6−9 which are critical in the pathobiology of psoriasis, systemic lupus erythematosus (SLE), and inflammatory bowel disease (IBD). 10−13 The human IL-12/23 monoclonal antibody ustekinumab has been approved by the U.S. Food and Drug Administration for the treatment of psoriasis,14 ulcerative colitis,15 and Crohn’s disease.16 As a result, TYK2 is a promising target for the development of orally active small molecules for autoimmune diseases.17−19 According to previous studies and available data, there is currently no commercially available inhibitor for clinical use.20−25 Some of the TYK2 inhibitors are summarized in Figure 1. Deucravacitinib is currently in the registration stage for the treatment of psoriasis. Unlike previous JAK inhibitors, deucravacitinib selectively binds to the TYK2 pseudokinase (JH2) domain and inhibits its signaling pathway by an allosteric © 2022 American Chemical Society Received: July 19, 2022 Accepted: October 3, 2022 Published: October 6, 2022 1730 https://doi.org/10.1021/acsmedchemlett.2c00334 ACS Med. Chem. Lett. 2022, 13, 1730−1738 ACS Medicinal Chemistry Letters pubs.acs.org/acsmedchemlett Letter Figure 1. Representative structures of TYK2 inhibitors. Figure 2. Design and modification strategies for the target compounds and X-ray crystal structure of TYK2 JH2 complexed with deucravacitinib (green) (PDB ID 6NZP). phosphorylation activity assay. Among them, compound 30 demonstrated excellent inhibitory potency against STAT3 phosphorylation. To better understand the selectivity of 30, binding affinity and cellular function assays for other members of JAK family were performed. In addition, compound 30 had reasonable pharmacokinetic (PK) exposure in mice. Finally, 30 was orally effective in both IL-23-driven acanthosis (psoriasislike) and anti-CD40-induced colitis models. Based on the crystal structure of TYK2 JH2 in complex with deucravacitinib (PDB ID 6NZP),13,34,35 we found that the N(methyl-d3)pyridazine-3-carboxamide skeleton of the ligand could form key hydrogen-bonding interactions with Val690 and Glu688 in the hinge region (Figure 2). In this way, the deuteromethyl group of the C3 amide could be anchored toward the adjacent Ala671, which is critical for maintaining the high selectivity. Thus, we preserved the N-(methyl-d3)pyridazine-3carboxamide skeleton and tried to introduce other substituents. Although the triazole nitrogen atom N2 engages in a direct hydrogen bond with Arg738, we found that there was still an unoccupied pocket below the P loop, which was particularly attractive since it might improve the potency upon the introduction of other groups. In order to facilitate the study of the structure−activity relationship (SAR) of this pocket, we replaced the terminal triazole group of deucravacitinib with an amide. Subsequently, the introduction of different substituents was carried out to investigate the effect of the electronic properties and steric hindrance on the activity. In this case, the target compounds 5−20, 23−30, and 32 were generated. Previous work by BMS showed that pyridylamine could be substituted for the C6 pendent cyclopropylamide with retention of activity using both in silico and traditional techniques.13 In addition, we found that the cyclopropylamide reached out into the solvent based on the crystal structural information. Hence, we designed the target compounds 21, 22, and 31 substituted with pyridylamine groups. Here we describe the synthesis and SAR of compounds with an N-(methyl-d3)pyridazine-3carboxamide skeleton. As shown in Scheme 1, compounds 5−27 were first prepared starting from commercially available methyl 4,6-dichloropyridazine-3-carboxylate (33), which was hydrolyzed and then amidated to afford intermediate 35 in high yield. The C4 chloride of 35 was selectively displaced by 3-amino-2methoxybenzoic acid to give key acid 36. Under palladiumcatalyzed conditions, 36 reacted with cyclopropanecarboxamide to give intermediate 37, which was condensed with various amines using EDCI to provide the desired amides 5−19 and 23−27. Alternatively, treatment of intermediate 37 with ammonium chloride easily gave amide 40, which was further converted to the expected compound 20. In addition, pyridines 41 were prepared by the coupling reaction of acid 36 with the corresponding amines. The condensation reactions of 41 with various amines gave compounds 21 and 22 in mod (...truncated)