Discovery of an orally active nitrothiophene-based antitrypanosomal agent

Abstract

Human African Trypanosomiasis (HAT), caused by Trypanosoma brucei gambiense and rhodesiense, is a parasitic disease endemic to sub-Saharan Africa. Untreated cases of HAT can be severely debilitating and fatal. Although the number of reported cases has decreased progressively over the last decade, the number of effective and easily administered medications is very limited. In this work, we report the antitrypanosomal activity of a series of potent compounds. A subset of molecules in the series are highly selective for trypanosomes and are metabolically stable. One of the compounds, (E)-N-(4-(methylamino)-4-oxobut-2-en-1-yl)-5-nitrothiophene-2-carboxamide (10), selectively inhibited the growth of T. b. brucei, T. b. gambiense and T. b. rhodesiense, have excellent oral bioavailability and was effective in treating acute infection of HAT in mouse models. Based on its excellent bioavailability, compound 10 and its analogs are candidates for lead optimization and pre-clinical investigations.

Keywords: Cysteine protease; HAT; Inhibitors; Trypanosomes; Vinyl sulfone.

Figure 1.

Structure of previously reported covalent inhibitor of tCatL (1) [8]. Multiparameter structure-activity investigation identified 10 as an orally active antitrypanosomal agent. Please see the R groups in Table 1 or the experimental section.

Figure 2.

PK plot and parameters of 10. The IV dose was administered at 1 mpk, and the PO dose was at 5 mpk in 5% DMSO/10% Solutol/85% PBS (containing 20% Captisol).

Figure 3.

Mice were infected with 1 x 10⁵ T. b. brucei trypomastigotes and treated for 5 days via IP or oral gavage. Vehicle: 0.5% Hydroxymethyl cellulose + 0.5% Tween-80. RLU: relative luminescence reading for T. brucei-Luc parasites. *** P value is < 0.0001 relative to the vehicle. Parasite density color key: Red: very high; Yellow: high; Green: medium; Blue: low

Figure 4.. The survival plot of T. brucei rhodesiense-infected mouse models treated with 10.

The percentage survival is the ratio of surviving animals over the total number of mice in each group (5 animals per group). The points represent the percent survival at each day. Treatment started on day 3 after infection and continued until day 7, as described in the methods section. B.i.d = twice a day; q.d = once a day.

Scheme 1.

Synthesis of 2 and 3. Reagents and conditions: (a) CH₂Cl₂, CDI, 0°C, 1 hr. Then DIBAL-H, −78°C; 80% (b) NaH, THF, diethyl((phenylsulfonyl)methyl) phosphonate, 0°C, 25 min, 60%; (c) S2 (0.24 mmol), 33% TFA in DCM (2 mL), 0°C, 1.5 h. Then ACN (2 mL), R-COOH (0.18 mmol), Et₃N (0.36 mmol), HBTU (0.36 mmol), Room Temperature, 16 h, 30-40%.

Scheme 2.

Synthesis of 4-11. Reagents and conditions: (a) CH₂Cl₂:CH₃OH, Et₃N, Boc₂O, 23°C, 2 h; (b) H₂O, NaIO₄, RT, 1 h, >90%; (c) NaH, THF, diethyl((methylsulfonyl)methyl) phosphonate or diethyl((phenylsulfonyl)methyl) phosphonate, 0°C, 25 min, 40-70%; (d) tert-butyl (E)-(3-(phenylsulfonyl)allyl)carbamate (0.17 mmol), tert-butyl (E)-(3-(methylsulfonyl)allyl)carbamate (0.21 mmol), or S6 (0. 23 mmol), 33% TFA in DCM, 0°C, 1.5 h. Then ACN (2 mL), R-COOH (1.5 mol eq), Et₃N (2 mol eq), HBTU (2 mol eq), RT, 16 h, 35-45%; (e) methyl (triphenylphosphoranylidene)acetate, THF, RT, 2 hr; (f) LiOH, THF/H₂O, RT, 16 h; (g) N-methylmorpholine (NMM), isobutyl chloroformate, 0°C, 1 hr. Then, methylamine, room temperature, 15 h.

Scheme 3.

Synthesis of 12 and 13. Reagents and conditions: (a) CH₂Cl₂, CDI, 0°C, 1 hr. Then DIBAL-H, −78°C; 80%; (b) methyl (triphenylphosphoranylidene)acetate, THF, RT, 2 hr; (c) Li-OH, THF/H₂O, RT, 16 hr; (d) N-methylmorpholine (NMM), isobutyl chloroformate, 0°C, 1 hr. Then, methylamine, RT, 15 hr; (e) S7 (0.16 mmol), 33% TFA in DCM, 0°C, 1.5 hr. Then ACN (2 mL), R-COOH (1.5 mol eq), Et₃N (2 mol eq), HBTU (2 mol eq), RT, 16 hr.

Scheme 4.

Synthesis of 14-18. Reagents and conditions: (a) Amine (0.21 mmol), ACN (2 mL), 5-nitrofuran-2-carboxylic acid (14), 5-nitrothiophene-2-carboxylic acid (15 and 17) or thiophene-2-carboxylic acid (16 and 18), Et₃N (2 mol eq), HBTU (2 mol eq), RT, 16 hr.