A Restricted Role for TYK2 Catalytic Activity in Human Cytokine Responses Revealed by Novel TYK2-Selective Inhibitors

Abstract

TYK2 is a JAK family protein tyrosine kinase activated in response to multiple cytokines, including type I IFNs, IL-6, IL-10, IL-12, and IL-23. Extensive studies of mice that lack TYK2 expression indicate that the IFN-α, IL-12, and IL-23 pathways, but not the IL-6 or IL-10 pathways, are compromised. In contrast, there have been few studies of the role of TYK2 in primary human cells. A genetic mutation at the tyk2 locus that results in a lack of TYK2 protein in a single human patient has been linked to defects in the IFN-α, IL-6, IL-10, IL-12, and IL-23 pathways, suggesting a broad role for TYK2 protein in human cytokine responses. In this article, we have used a panel of novel potent TYK2 small-molecule inhibitors with varying degrees of selectivity against other JAK kinases to address the requirement for TYK2 catalytic activity in cytokine pathways in primary human cells. Our results indicate that the biological processes that require TYK2 catalytic function in humans are restricted to the IL-12 and IL-23 pathways, and suggest that inhibition of TYK2 catalytic activity may be an efficacious approach for the treatment of select autoimmune diseases without broad immunosuppression.

Intestinal Epithelial Cell Tyrosine Kinase 2 Transduces IL-22 Signals To Protect from Acute Colitis

“…The complex Tyk2 functions in mice are also reflected in humans, because variation occurs between human populations with respect to Crohn's disease susceptibility and its correlation to the Tyk2 and STAT3 loci (64). In conclusion, a future use of Tyk2 inhibitors (65,66) for the treatment of colitis should carefully consider whether the primary therapeutic benefit is expected through the regeneration of the epithelial barrier or the dampening of the hyperinflammation.…”

Structure-Based Design and Synthesis of 3-Amino-1,5-dihydro-4H-pyrazolopyridin-4-one Derivatives as Tyrosine Kinase 2 Inhibitors

“…Reagents and conditions: (a) 2,2,6-trimethyl-4H-1,3-dioxin-4-one, NaOAc, THF, reflux, 74% for 22c; (b) diketene, MeOH, 0 °C, 96% for 22d; (c) K 2 CO 3 , DMF/DMA, DMF, rt, 69-84% for 23a-b, d; (d) DMF/DMA, DMF, rt, 95% for 23c; (e) POCl 3 , DMF, 0 °C→100-125 °C, 59-64% for 24a-c; (f) (chloromethylene)dimethylammonium chloride, DMF 100 °C, 57% for 24d, (g) hydroxylamine hydrochloride, NaOAc, MeOH-H 2 O, 75 °C, 77-92% for 25a, 25c; (h) hydroxylamine hydrochloride, conc HCl, 2-propanol, 100 °C, 68-95% for 25b, 25d; (i) POCl 3 , CH 3 CN, 90 °C, 53-100% for 26a-c; (j) SOCl 2 , CH 3 CN, rt, 68% for 26d; (k) NBS, DMF, 50-60 °C, 81-98%; (l) PhB(OH) 2 , Pd(PPh 3 ) 4 , 2 M Na 2 CO 3 aq., DME, 90 °C, 64%; (m) hydrazine monohydrate, EtOH, 70 °C, 27%; (n) hydrazine monohydrate, EtOH, 70-90 °C, 82%-94%; (o) chiral HPLC, CHIRALPAK AD, hexane/EtOH, 44%; (p) KOAc, PdCl 2 (dppf) (or Pd(PPh 3 ) 4 ), bis(pinacolato)diboron, DMF (or DMA), 110-120 °C, then R 2 -Br (or R 2 -I, or R 2 -OTf), Pd(PPh 3 ) 4 , 2 M Na 2 CO 3 aq., 120 °C, 8-41% for 5, 6, 13, 15, 17, 19, 20, 30; (q) 1-methyl-3-(4,4,5,5-tetramethy-1,3,2-dioxaborolan-2-yl)-1H-pyrazole, Pd(PPh 3 ) 4 , 2 M Na 2 CO 3 aq., DMF,120 °C,[11][12][13][14][15][16][17][18][19][20][21][22][23][24][25][26][27][28][29]9,16;(r) In order to acquire SAR of 5-position efficiently, an alternative synthetic route was developed (Scheme 2). Pyridone 31 was reacted with alkyl halides, followed by the separation of N-alkylated pyridones from the mixture of N-and O-alkylated products by chromatography to give 32a-c.…”

“…19 The formyl group of 24a-d was converted to cyano group via aldoxime, followed by bromination using N-bromosuccinimide (NBS) to give key intermediates, 1-substituted 5-bromo-pyridin-2(1H)-ones 27a-d. The targeted analogs (5-9, 13, [16][17][18][19][20] were synthesized from 27a-d via two step reactions: condensation with hydrazine to form the aminopyrazole part, and Suzuki-Miyaura coupling to introduce 7-substitutent. Optically pure 3-methylbutan-2-yl analogs were prepared by chiral HPLC resolution of the racemic analog 13, or the key chiral intermediate 29d…”

“…TYK2 inhibitory activity and JAK2 selectivity of a compound bearing propan-2-ol (20) slightly decreased compared to those of the morpholine derivative (17). The above results demonstrated that further improvement in TYK2 inhibitory activity was achieved by the optimization of the 5-and 7-positions of 3-amino-1,5-dihydro-4H-pyrazolopyridin-4-one, through (1) occupying the JAK pocket efficiently with a chiral alkyl group and (2) by additional interaction with the amino acid residues in the solvent exposed (14)(15)(16)(17)(18)(19) [7.6] 17 (13-22) [8.1] 3.3 (3.0-3.6) [1.6] 190 (75-480) a The enzyme inhibition assay using JAK1-3, or TYK2 enzymes. b Inhibitory activity of IL-23-induced luciferase gene expression in Jurkat cells.…”

“…TYK2 inhibitory activity of the (S)-form (14) with subnanomolar IC 50 values was 10-fold more potent than that of the (R)-form (15). Next the effect of introducing substitution at the 5'-position of the 1-methyl-3-pyrazolyl moiety (e.g., [16][17][18][19][20], which were expected to interact with Leu903 or Asp988, was investigated (Figure 3). Introduction of morpholine onto the 5'-position of 1-methyl-3-pyrazole (17) afforded a 10-fold increase in TYK2 inhibitory activity compared to the corresponding unsubstituted analog (16) with 25-fold selectivity over JAK2.…”