Structure-activity relationship studies of the lipophilic tail region of sphingosine kinase 2 inhibitors

Abstract

Sphingosine-1-phosphate (S1P) is a ubiquitous, endogenous small molecule that is synthesized by two isoforms of sphingosine kinase (SphK1 and 2). Intervention of the S1P signaling pathway has attracted significant attention because alteration of S1P levels is linked to several disease states including cancer, fibrosis, and sickle cell disease. While intense investigations have focused on developing SphK1 inhibitors, only a limited number of SphK2-selective agents have been reported. Herein, we report our investigations on the structure-activity relationship studies of the lipophilic tail region of SLR080811, a SphK2-selective inhibitor. Our studies demonstrate that the internal phenyl ring is a key structural feature that is essential in the SLR080811 scaffold. Further, we show the dependence of SphK2 activity and selectivity on alkyl tail length, suggesting a larger lipid binding pocket in SphK2 compared to SphK1.

Keywords: S1P; SphK1; SphK2; Sphingosine kinase; Sphingosine-1-phosphate.

Figure 1

Structure of sphingosine kinase 2 inhibitors.

Figure 2

Pharmacophore of guanidine-based inhibitors.

Scheme 1

a.) Alkyne (2 equiv.), TEA (5 equiv.), DMF, PdCl₂(PPh₃)₂ (0.05 equiv.), CuI (0.03 equiv.), 80 °C, 18 h, (72–93%); b.) i. Alkene, 0.5 M 9-BBN, in THF, rt, 12 h; ii. Pd(dppf)Cl₂, Cs₂CO₃, DMF, 70 °C, 18 h, (75–93%); c.) NH₂OH·HCl (3 equiv.), TEA (3 equiv.), EtOH, 80 °C, 6 h, (43–95%); d.) Boc-L-Proline (1.4 equiv.), DIEA (1.4 equiv.), HCTU (1.8 equiv.), DMF, 110 °C, 18 h, (25–65%); e.) DME (20 vol/wt), 4-toluenesulfonyl hydrazide (10 equiv.), TEA (5 equiv.), reflux, (67–71%); f.) HCl/MeOH, (35–100%); g.) DIEA (3 equiv.), N,N'-Di-Boc-1H-pyrazole-1-carboxamidine (1.05 equiv.), CH₃CN, rt, 3 days, (27–76%).

Scheme 2

a.) Boc-L-Azetidine (1.4 equiv.), DIEA (1.4 equiv.), HCTU (1.8 equiv.), DMF, 110 °C, 18 h, (63%); b.) Alkyne (2 equiv.), TEA (5 equiv.), DMF, PdCl₂(PPh₃)₂ (0.05 equiv.), CuI (0.03 equiv.), 80 °C, 18 h, (33–57%); c.) Phenylboronic acid (1.3 equiv.), Cs₂CO₃ (equiv.), DMF, PdCl₂(dppf) (0.04 equiv.), 80 °C, 18 h, (91%); d.) Amine, Pd(dba)₃, Cs₂CO₃, PtBu₃, toluene, 120 °C, 6 d, (81–83%); e.) DME (20 vol/wt), 4-toluenesulfonyl hydrazide (10 equiv.), TEA (5 equiv.), reflux, (60–71%); f.) HCl/MeOH, (78–96%); g.) DIEA (3 equiv.), N,N'-Di-Boc-1H-pyrazole-1-carboxamidine (1.05 equiv.), CH₃CN, rt, 3 days, (43–66%).

Scheme 3

a.) Piperazine (3 equiv.), Pd₂(dba)₃ (0.2 equiv.), PtBu₃ (0.8 equiv.), Cs₂CO₃ (1.2 equiv.), toluene, 120 °C, 3 days, (52%); b.) Acid chloride (2.5 equiv.) or benzyl bromide (1 equiv.), TEA (2 equiv.), CH₂Cl₂, 0 °C—rt, 2 h, (66–88%); c) HCl/MeOH, (76–95%); d) DIEA (3 equiv.), N,N'-Di-Boc-1H-pyrazole-1-carboxamidine (1.05 equiv.), CH₃CN, rt, 3 days, (23–74%).

Scheme 4

a.) NaH, EtOH, 0 °C—rt, (46%); b.) 4-cyanophenylboronic acid, Pd(PPh₃)₄ (10 mol%), Na₂CO₃, THF:H₂O, (94%); c.) NH₂OH·HCl (3 equiv.), TEA (3 equiv.), EtOH, 80 °C, 6 h, (71–93%); d.) Boc-L-Proline (1.4 equiv.), DIEA (1.4 equiv.), HCTU (1.8 equiv.), DMF, 110 °C, 18 h, (46–82%); e.) HCl/MeOH, (33–91%); f.) DIEA (3 equiv.), N,N'-Di-Boc-1H-pyrazole-1-carboxamidine (1.05 equiv.), CH₃CN, rt, 3 days, (53–83%); g.) sodium benzenesulfonate (1.5 equiv.), DMF, 60 °C, 2 h, (89%).

Scheme 5

a.) KCN (2 equiv.), 9:1 EtOH:H₂O, 80 °C, 18 h, (20–93%); b.) NH₂OH·HCl (3 equiv.), TEA (3 equiv.), EtOH, 80 °C, 12 h, (53–69%); c.) Boc-L-Proline (1.4 equiv.), DIEA (1.4 equiv.), HCTU (1.8 equiv.), DMF, 110 °C, 18 h, (50%); d.) Boc-L-Proline (1.4 equiv.), DIEA (1.4 equiv.), HCTU (1.8 equiv.), CH₂Cl₂, rt, 4 h, (57–80%); e.) TBAF (1.0 M, 1 equiv.), THF, rt, 1 h, (93–95%); f.) HCl/MeOH, (66–100%); g) DIEA (3 equiv.), N,N'-Di-Boc-1H-pyrazole-1-carboxamidine (1.05 equiv.), CH₃CN, rt, 3 days, (51–71%).