Discovery of GLPG3667, a Selective ATP Competitive Tyrosine Kinase 2 Inhibitor for the Treatment of Autoimmune Diseases

Abstract

Tyrosine kinase 2 (TYK2) mediates cytokine signaling through type 1 interferon, interleukin (IL)-12/IL-23, and the IL-10 family. There appears to be an association between TYK2 genetic variants and inflammatory conditions, and clinical evidence suggests that selective inhibition of TYK2 could produce a unique therapeutic profile. Here, we describe the discovery of compound 9 (GLPG3667), a reversible and selective TYK2 adenosine triphosphate competitive inhibitor in development for the treatment of inflammatory and autoimmune diseases. The preclinical pharmacokinetic profile was favorable, and TYK2 selectivity was confirmed in peripheral blood mononuclear cells and whole blood assays. Dermal ear inflammation was reduced in an IL-23-induced in vivo mouse model of psoriasis. GLPG3667 also completed a phase 1b study (NCT04594928) in patients with moderate-to-severe psoriasis where clinical effect was shown within the 4 weeks of treatment and it is now in phase 2 trials for the treatment of dermatomyositis (NCT05695950) and systemic lupus erythematosus (NCT05856448).

Figure 1

Examples of approved selective and nonselective JAK1 inhibitors. (Note: This is not an exhaustive list of all licensed JAK inhibitors targeting JAK1.)

Figure 2

Investigational tyrosine kinase 2 inhibitors.^,,

Figure 3

Selectivity comparison between the (A) 1H-imidazo[4,5-c]pyridine and the 3H-imidazo[4,5-b]pyridine scaffolds and (B) matched pairs 10 and 11. IC₅₀ values obtained from fluorescence-based biochemical assays using the catalytic domains of the four JAK members. Selectivity determined as the ratio of the IC₅₀ values for JAK1/TYK2.

Figure 4

(A) Compound 11 docked in TYK2 structure (Protein Data Bank code 3LXN). Compound 11 is displayed in green ball-and-sticks. Only key amino acids of TYK2 are shown (gray sticks). Hydrogen bonds are highlighted with yellow dotted lines, and the aromatic H-bond is materialized with a dotted blue line. (B) Quantum mechanics minimized structures of 10 (pink) and 11 (green), overlaid on the imidazole ring. A distance of 1.5 Å could be measured between the nitrogen atoms of the cyano group.

Figure 5

Compound 13 docked in TYK2 structure (Protein Data Bank code 3LXN). Compound 13 is displayed in green ball-and-sticks. Only key amino acids of TYK2 are shown (gray sticks). Hydrogen bonds are highlighted with yellow dotted lines.

Figure 6

Relationship between calculated distribution coefficient (cLog D) and unbound intrinsic clearance (CL_int,u) from incubations with mouse liver microsomes (mLM). ^aCalculated by Simulation Plus. Solid lines show regions with equal lipophilic metabolic efficiency (LipMetE) (LipMetE = cLog D – log10[CL_int,u], where CL_int,u is expressed in mL/min/kg).

Figure 7

Representative frame of a molecular dynamics simulation of compound 9 docked in TYK2 structure (Protein Data Bank code 3LXN). Compound 9 is displayed in green sticks. Only key amino acids of TYK2 are shown (gray sticks). Bridging water molecules that were stable during the simulation are displayed. Hydrogen bonds are highlighted with black dotted lines.

Figure 8

Pharmacokinetics after 4 days of treatment with 9, dosed at 3, 10, and 30 mg/kg (q.d.). IC₅₀ values of 9 for the two mouse pathways are deduced from mouse whole blood assays (see Supporting Information).

Figure 9

Effect of 9 dosed at 3, 10, and 30 mg/kg (q.d.) and TYK2 inhibitor used as positive control dosed at 30 mg/kg (q.d.) on IL-23-induced psoriasis-like inflammation in the ear skin of mice on (A) quantification of ear thickening and (B) pictures of inflammation and STAT3 phosphorylation in the ear tissue, (C) quantification of phosphorylated STAT3, (D) pictures of neutrophil (NIMP-R14-positive cells) infiltration in the ear tissue, and (E) quantification of neutrophil percentage area. Data shown are average ± standard error of the mean of n = 9–10 mice per group. Groups were compared using a one-way analysis of variance followed by Dunnett’s multiple comparisons test and, for part B, t test for two groups comparison: *p < 0.05, **p < 0.01, ***p < 0.001 versus vehicle group. ^#p < 0.05 for TYK2 inh. group versus Cpd 9 30 mg/kg group. ^£p < 0.05 for Cpd 9 10 mg/kg group versus Cpd 9 30 mg/kg group. BSA, bovine serum albumin; C, cartilage; D, dermis; E, epidermis.

Scheme 1. Synthesis of Core 37

Reagents and conditions: (a) HNO₃, H₂SO₄, 10 °C for 30 min then 80 °C for 30 min, 86%; (b) benzyl bromide, K₂CO₃, DMF, 80 °C, 1 h, 84%; (c) 33% MeNH₂ in EtOH, Et₃N, THF, rt, 16 h; (d) Na₂S₂O₄, NaHCO₃, THF/H₂0, rt, 1 h, crude; (e) triethyl orthoformate, MeCN, 80 °C, 16 h, 55%.

Scheme 2. Synthesis of 11, 12, and 13

Reagents and conditions: (a) XantPhos, XantPhos Pd G3, Cs₂CO₃, 1,4-dioxane, 110 °C, 16 h; (b) MeI, NaH, THF, 0 °C then rt, 1 h, 74%; (c) TfOH, DCM, rt, 16 h; (d) for 11: cyclopropanecarbonyl chloride, pyridine, DCM, rt, 2 h; (e) for 12: CDT, pyridine, DCM, 50 °C, 1 h, then 2 M MeNH₂ in THF, 1 h; (f) for 13: 6-chloropyrimidin-4-amine, BrettPhos, BrettPhos Pd G3, Cs₂CO₃, 1,4-dioxane, 110 °C, 16 h.

Scheme 3. Synthesis of Core 45

Reagents and conditions: (a) N-iodosuccinimide, TFA, THF, rt, 16 h, 88%; (b) 2 M MeNH₂ in THF, NMP, 180 °C, 48 h, 61%; (c) trimethyl orthoformate, formic acid, 60 °C, 1 h, 89%.

Scheme 4. Synthesis of 14, 15, 16, 17, 19, 20, and 21

Reagents and conditions: (a) RuPhos, RuPhos Pd G3, K₃PO₄, 1,4-dioxane, 110 °C, 18 h, 43%; (b) pyrimidine-4,6-diamine, MorDalPhos, MorDalPhos Pd G3, Cs₂CO₃, 1,4-dioxane, 110 °C, 18 h; (c) for 49a and 49d: RNH₂, XantPhos, XantPhos Pd G3, Cs₂CO₃, 1,4-dioxane, 110 °C, 16 h, 39–65%; for 49b: 2b, XantPhos, XantPhos Pd G3, Cs₂CO₃, 1,4-dioxane, 70 °C, 18 h, 78%; for 49c: RNH₂, XantPhos, XantPhos Pd G3, K₃PO₄, diglyme, 110 °C, 18 h; for 49e: 3b, XantPhos, XantPhos Pd G3, K₃PO₄, diglyme, 80 °C, 18 h; (d) NaH, MeI, 0 °C to rt; (e) for 19: pyrimidine-4,6-diamine, tBuXPhos, tBuXPhos Pd G3, K₃PO₄, 1,4-dioxane, 110 °C, 18 h, 73%.

Scheme 5. Synthesis of 18, 22, and 23

Reagents and conditions: (a) For 51a: ROH, CuI, 3,4,7,8-tetramethyl-1,10-phenanthroline, Cs₂CO₃, DMF, 80 °C, 3 h; for 51b (using 4b) and 51c (using 5e), CuI, 2,2,6,6-tetramethyl-3,5-heptanedione, Cs₂CO₃, DMF, 85 °C, 72 h; (b) pyrimidine-4,6-diamine, MorDalPhos, MorDalPhos Pd G3, Cs₂CO₃, 1,4-dioxane, 110 °C, 18 h.

Scheme 6. Synthesis of Compounds 9, 24, 25, 26, 29, 30, and 31

Reagents and conditions: (a) For 24–25: ArNH₂, MorDalPhos, MorDalPhos Pd G3, Cs₂CO₃, 1,4-dioxane, 110 °C, 18 h; for 9, 26, 30 (using 6c), and 31 (using 7c): ArNH₂, MorDalPhos, [(Allyl)PdCl]₂, Cs₂CO₃, 1,4-dioxane, 110 °C, 18 h; for 29: ArNH₂, MorDalPhos Pd G4, Cs₂CO₃, 1,4-dioxane, 110 °C, 18 h.

Scheme 7. Synthesis of Compounds 27 and 28

Reagents and conditions: (a) MorDalPhos, [(Allyl)PdCl]₂, Cs₂CO₃, 1,4-dioxane, 110 °C, 75%; (b) LiI, pyridine, 115 °C, 24 h, quantitative; (c) RNH₂, HATU, Et₃N, NMP, rt, 4–18 h.