Heteroaryl amide derivatives as sodium channel inhibitors

Abstract

The present invention relates generally to compounds of formula (I) that inhibit sodium channel (NaV) activity; in particular, the present invention relates to compounds which are heteroaryl amide derivatives, including zwitterionic forms and pharmaceutically acceptable salts thereof, methods of preparing said compounds, and therapeutic uses thereof. The compounds of the invention are useful, for example, in the treatment of numerous disorders associated with NaV, such as respiratory diseases. (I).

Description

(19) State Intellectual Property Office (12) Invention Patent Application (10) Application Publication Number (43) Application Publication Date (21) Application Number 202480069977.0 (22) Application Date 2024.10.30 (30) Priority Data 23207536.6 2023.11.02 EP (85) PCT International Application Entering National Phase Date 2026.04.30 (86) PCT International Application Application Data PCT / EP2024 / 080663 2024.10.30 (87) PCT International Application Publication Data WO2025 / 093589 EN 2025.05.08 (71) Applicant: Chis Pharmaceuticals Co., Ltd. Address: Italy (72) Inventors: C. Fiorelli, G. Fossati, L. Mengozchi, G. Mosca, R. Patachini, A. Ritz (74) Patent Agency: China Council for the Promotion of International Trade Patent & Trademark Office Co., Ltd. 11038 Patent Attorney: Xu Da (51) Int.Cl. C07D 409 / 06 (2006.01) A61K 31 / 4025 (2006.01) A61K 31 / 381 (2006.01) A61K 31 / 42 (2006.01) A61K 31 / 5377 (2006.01) A61P 11 / 14 (2006.01) A61P 11 / 00 (2006.01) C07D 333 / 38 (2006.01) C07D 409 / 14 (2006.01) C07D 413 / 12 (2006.01) C07D 413 / 14 (2006.01) (54) Invention Title: Heteroarylamide Derivatives as Sodium Channel Inhibitors (57) Abstract: This invention generally relates to compounds of formula (I) that inhibit sodium channel (NaV) activity; in particular, this invention relates to compounds that are heteroarylamide derivatives, including zwitterionic forms and pharmaceutically acceptable salts thereof, methods for preparing said compounds, and their therapeutic uses. The compounds of this invention can be used, for example, to treat many disorders related to NaV, such as respiratory diseases. (I). Claims 7 pages, Description 106 pages, CN 122180674 A 2026.06.09 CN 1 22 18 06 74 A 1. Compound (I) of formula (I) wherein at least one of Y and Z is S, and the other is CR4; R1 and R2 are independently -(C1-C6)alkyl or fused to form -(C3-C7)heteroalkyl, wherein the -(C3-C7)heteroalkyl is optionally substituted by one or more groups selected from halogens, -(C1-C6)alkyl and -(C1-C6)alkyl-OR8;R3 and R4 are independently H or -(C1-C6)alkyl, or, in the case that Y is CR4, R3 and R4 are fused to form an aryl group; R5 and R6 are independently H or selected from -(C1-C6)alkyl, -(C1-C6)aminoalkyl, -(C1-C6)alkyl-OR8, -(C1-C6)alkyl-NR7R8, -(C1-C4)alkyl-NR7-C(O)OR8, -(C1-C4)alkyl-(C3-C7)heterocyclic alkyl and -(C3-C7)heterocyclic alkyl, or fused to form -(C3-C7)heterocyclic alkyl, wherein the -(C1-C4)alkyl-(C3-C7)heterocyclic alkyl, -(C3-C7)heterocyclic alkyl, -(C1-C6)aminoalkyl or -(C1-C6)alkyl-NR7R8 is optionally substituted by one or more groups selected from - (C1-C6)alkyl, -C(O)OR8 and -SO2R7; R7 and R8 are independently H or selected from -(C1-C6)alkyl, -(C3-C6)cycloalkyl and -(C1-C6)haloalkyl, or fused to form -(C3-C7)heteroalkyl; L1 is a bond or selected from -(C1-C6)alkylene- and -(C1-C6)cycloalkylene-; and A is a ring selected from aryl and heteroaryl, wherein any of the aryl or heteroaryl groups are optionally substituted by one or more groups selected from -OR8, -C(O)OR8, -C(O)NR7R8 and -(C1-C6)alkyl; provided that, if L1 is a bond, any of the aryl groups are not substituted by -(C1-C6)alkyl; in zwitterionic or pharmaceutically acceptable salt form. 2. The compound of formula (I) according to claim 1, wherein Z, Y, R1, R2, R3, R4, R5, R6, R7 and R8 are as defined in claim 1; L1 is selected from -(C1-C6)alkylene- and -(C1-C6)cycloalkylene-; and A is a ring selected from aryl and heteroaryl, wherein any of the aryl or heteroaryl groups are optionally substituted by one or more groups selected from -OR8, -C(O)OR8, -C(O)NR7R8 and -(C1-C4)alkyl; and is in zwitterionic or pharmaceutically acceptable salt form. 3. The compound of formula (I) according to claim 1, wherein L1 is a bond, represented by formula (Ia) as defined in claim 1 / 7 page 2 CN 122180674 A (Ia) wherein Z, Y, R1, R2, R3, R4, R5, R6, R7 and R8 are as defined in claim 1; and A is a ring selected from aryl and heteroaryl, wherein any of the aryl groups are optionally substituted by one or more groups selected from -OR8, -C(O)OR8 and -C(O)NR7R8, and wherein any of the heteroaryl groups are optionally substituted by one or more groups selected from -OR8, -C(O)OR8, -C(O)NR7R8 and -(C1-C6)alkyl;It is in the form of a zwitterion or a pharmaceutically acceptable salt. 4. A compound according to any one of the preceding claims, wherein R1 and R2 are independently -(C1-C4)alkyl or fused to form -(C3-C7)heteroalkyl, wherein the -(C3-C7)heteroalkyl is optionally substituted by one or more groups selected from halogens, -(C1-C4)alkyl, and -(C1-C4)alkyl-OR8; R3 and R4 are independently H or -(C1-C6)alkyl, or, in the case where Y is CR4, R3 and R4 are fused to form an aryl group; R5 and R6 are independently H or selected from -(C1-C4)alkyl, -(C1-C4)aminoalkyl, -(C1-C4)alkyl-OR8, and -(C1-C4)alkyl-OR8. Alkyl-NR7R8, -(C1-C4)alkyl-NR7-C(O)OR8, -(C1-C4)alkyl-(C3-C7)heterocyclic alkyl, -(C3-C7)heterocyclic alkyl, or fused to form -(C3-C7)heterocyclic alkyl, wherein the -(C1-C4)alkyl-(C3-C7)heterocyclic alkyl, -(C3-C7)heterocyclic alkyl, -(C1-C4)aminoalkyl or -(C1-C4)alkyl-NR7R8 is optionally substituted by one or more groups selected from -(C1-C4)alkyl, -C(O)OR8 and -SO2R7; and R7 and R8 are independently H or selected from -(C1-C4)alkyl, -(C3-C6)cycloalkyl and -(C1-C4)haloalkyl, or fused to form -(C6-C7)heterocyclic alkyl; in zwitterionic or pharmaceutically acceptable salt form. 5. A compound according to any one of the preceding claims, wherein R1 and R2 are fused into a -(C6-C7) heterocyclic alkyl group, wherein the -(C6-C7) heterocyclic alkyl group is optionally substituted by one or more groups selected from -OR8, -(C1-C4) alkyl, and -(C3-C7) heterocyclic alkyl; and is in a zwitterionic or pharmaceutically acceptable salt form. 6. A compound according to any one of the preceding claims, wherein R3 and R4 are independently H or -(C1-C4) alkyl; and is in a zwitterionic or pharmaceutically acceptable salt form. 7. A compound according to any one of the preceding claims, wherein Z is S and Y is CH, and R3 is H or methyl; and is in a zwitterionic or pharmaceutically acceptable salt form. 8. A compound according to any one of the preceding claims, wherein claim AA is a heteroaryl group, optionally substituted by one or more groups selected from -OR8, -C(O)OR8, -C(O)NR7R8 and -(C1-C6)alkyl; and is in zwitterionic or pharmaceutically acceptable salt form. 9. A compound according to any one of claims 1, selected from:2-(benzylamino)-N,N-dimethyl-N-(2-((4-methyl-2-(pyrrolidine-1-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)-2-oxoethyl-1-ammonium (Example 1); 2-(benzylamino)-N-(2-((2-(dimethylcarbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-N,N-dimethyl-2-oxoethyl-1-ammonium (Example 2); 2-(benzylamino)-N-(2-((2-(tert-butylcarbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-N,N-dimethyl-2-oxoethyl-1-ammonium (Example 3); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(morpholin-4-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 4); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(methylcarbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 5); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(pyrrolidine-1-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 6); 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(pyrrolidine-1-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 7); 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(morpholin-4-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 8); 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(methylcarbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 9); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-(dimethylcarbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 10); 1-(2-((2-(dimethylcarbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-(isoxazo-3-ylamino)-2-oxoethyl)azacycloheptane-1-onium (Example 11); 1-(2-((4-methyl-2-(methylcarbamoyl)thiophen-3-yl)amino)-2-oxoethyl)-1-(2-((4-methylisoxazo-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 12);1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((2-((2-methoxyethyl)carbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)piperidine-1-onium (Example 13); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-(dimethylcarbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)piperidine-1-onium (Example 14); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-((2-((tert-butoxycarbonyl)(methyl)amino)ethyl)carbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 15); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-((2-methoxyethyl)carbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 16); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-((2-methoxyethyl)carbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)piperidine-1-onium (Example 17); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-((2-hydroxyethyl)carbamoyl)-4-methylthiophene-... Claims 3 / 7, page 4, CN 122180674 A 3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 18); 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((4-methyl-2-((2-(methylamino)ethyl)carbamoyl)thiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 19); 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((2-((2-methoxyethyl)carbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 20); 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(oxetane-3-ylcarbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 21); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-((2-(methylamino)ethyl)carbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 22); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(oxetane-3-ylcarbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 22); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(oxetane-3-ylcarbamoyl)thiophen-3-yl)thiophen-3-yl)thiophen-3-yl)thiophen-3-yl)thiophen-3-yl)thiophen-3-yl)thiophen-3-yl)azacycloheptane-1-onium (Example 22);Pheno-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 23); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-((2-hydroxyethyl)carbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)piperidine-1-onium (Example 24); 1-(2-((2-((2-hydroxyethyl)carbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-(isoxazo-3-ylamino)-2-oxoethyl)azacycloheptane-1-onium (Example 25); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-((2-(methylamino)ethyl)carbamoyl)thiophen-3-yl)amino)-2-oxoethyl)piperidine-1-onium (Example 26); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(4-methylpiperazin-1-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 27); 1-(2-((2-(dimethylcarbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((4-methylisoxazol-3-yl)amino)-2-oxoethyl)piperidine-1-onium (Example 28); 1-(2-((2-((2-methoxyethyl)carbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)-1-(2-((4-methylisoxazol-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 29); 1-(2-(isoxazol-3-ylamino)-2-oxoethyl)-1-(2-((4-methyl-2-((2-morpholinoethyl)carbamoyl)thiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 30); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-(4-(tert-butoxycarbonyl)piperazine-1-carbonyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 31); 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(4-(methanesulfonyl)piperazine-1-carbonyl)thiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 32); 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(piperazine-1-carbonyl)thiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 32); 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(piperazine-1-carbonyl)thiophene-3-yl) (Example 33) 1-(2-((2-(4-(tert-butoxycarbonyl)piperazine-1-carbonyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)-1-(2-(isoxazo-3-ylamino)-2-oxoethyl)azacycloheptane-1-onium (Example 34); 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(4-methylpiperazine-1-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 35); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(4-(methanesulfonyl)piperazine-1-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 36); 1-(2-(((2-carbamoyl-4-methylthiophene-3-yl)methyl)amino)-2-oxoethyl)-1-(2-(isoxazo-3-ylamino)-2-oxoethyl)azacycloheptane-1-onium (Example 37); Claims 4 / 7, page 5, CN 122180674 A 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(piperazin-1-carbonyl)thiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 38); 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((2-(morpholino-4-carbonyl)benzo[b]thiophene-3-yl) (R)-1-(2-(((2-carbamoyl-4-methylthiophen-3-yl)methyl)amino)-2-oxoethyl)-1-(2-oxo-2-(((1-phenylethyl)amino)ethyl)azacycloheptane-1-on-(Example 40); 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((4-methyl-2-((2-(trimethylammonium)ethyl)carbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-on-(Example 41); 1-(2-((2-(dimethylcarbamoyl)benzo[b]thiophen-3-yl)amino)-2-oxoethyl)-1-(2-(isoxazo-3-ylamino)-2-oxoethyl)azacycloheptane-1-onium (Example 42); 1,1-di(2-((2-(dimethylcarbamoyl)benzo[b]thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 43); 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((2-((2-methoxyethyl)(methyl)carbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 44); 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((4-methyl-2-((3-(methylamino)propyl)amino)1-(2-(benzylamino)-2-oxoethyl)azacycloheptane-1-onium (Example 45); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-((2-(trimethylammonium)ethyl)carbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 46); 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((4-methyl-2-((2-(4-methylpiperazin-1-yl)ethyl)carbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 47); 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((4-methyl-2-((2-(piperazin-1-yl)ethyl)carbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 48); 1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((4-methyl-2-(4-methylpiperazin-1-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 49); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-((2-methoxyethyl)(methyl)carbamoyl)-4- Methylthiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 50); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-((2-(piperazin-1-yl)ethyl)carbamoyl)thiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 51); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-((2-(4-methylpiperazin-1-yl)ethyl)carbamoyl)thiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 52); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-((3-(methylamino)propyl)carbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 53); 1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((4-methyl-2-(morpholin-4-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 54); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-((2-hydroxyethyl)(methyl)carbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 55);1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(methyl(2-(methylamino)ethyl)carbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 56); 1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((4-methyl-2-(piperazine-1-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 57); (S)-1-(2-((2-((6-amino-1-methoxy-1-oxohexane-2-yl)carbamoyl)-4-methylthiophen-3-yl) (Example 58) 1-(2-(benzylamino)-2-oxoethyl)azacycloheptane-1-onium; 1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((4-methyl-2-((2-(methylamino)ethyl)carbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 59); 4-fluoro-1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-4-methyl-1-(2- ((4-methyl-2-((2-(methylamino)ethyl)carbamoyl)thiophene-3-yl)amino)-2-oxoethyl)piperidine-1-onium (Example 60); 1-(2-((4-methyl-2-((2-(methylamino)ethyl)carbamoyl)thiophene-3-yl)amino)-2-oxoethyl)-1-(2-oxo-2-((2-(trifluoromethoxy)benzyl)amino)ethyl)azacycloheptane-1-onium (Example 61); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-((piperidine-4-ylmethyl)carbamoyl)thiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 62); 1-(2-((2-((2-(cyclopropylamino)ethyl)carbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 63); 1-(2-((2-(di(2-hydroxyethyl)carbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 64);1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((4-methyl-2-(piperidin-4-ylcarbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 65); 1-(2-((2-(dimethylcarbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 66); 1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-4,4-dimethyl-1-(2- ((4-Methyl-2-(methylcarbamoyl)thiophen-3-yl)amino)-2-oxoethyl)piperidine-1-onium (Example 67); 1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((4-methyl-2-(methylcarbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 68); 1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((4-methyl-2-(methylcarbamoyl)thiophen-3-yl)amino)-2-oxoethyl)piperidine-1-onium (Example 69); 1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((4-methyl-2-(oxetane-3-ylcarbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 70); 1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-4,4-dimethyl-1-(2-((4-methyl-2-((2-(methylamino)ethyl)carbamoyl)thiophen-3-yl)amino)-2-oxoethyl)piperidine-1-onium (Example 71); 1-(2-((2-(ethoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-4,4-dimethyl-1-(2-((4-methyl-2-((3-(methylamino)propyl)carbamoyl)thiophen-3-yl)amino)-2-oxoethyl)piperidine-1-onium (Example 72); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-(dimethylcarbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-4,4-di(hydroxymethyl)piperidine-1-onium (Example 73); (2-(4,4-dimethyl-1-(2-((4-methyl-2-(methylcarbamoyl)thiophen-3-yl)amino)-2-oxoethyl) (Claims 6 / 7)7 CN 122180674 A (Isoxazol-1-onthiol-1-yl)acetyl(isoxazo-3-yl)amide (Example 74); 1-(2-((2-(dimethylcarbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-4,4-di(hydroxymethyl)-1-(2-oxo-2-((1-phenylcyclopropyl)amino)ethyl)piperidine-1-onthiol (Example 75); 1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((4-methyl-2-yl)) ((piperidin-4-ylmethyl)carbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 76); and (R)-1-(2-((2-((6-amino-1-methoxy-1-oxohexane-2-yl)carbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-(benzylamino)-2-oxoethyl)azacycloheptane-1-onium (Example 77); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-methyl-4-(morpholin-4-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 78) 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-methyl-4-(pyrrolidine-1-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 79) 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-((2-methoxyethyl)(methyl)carbamoyl)-2-methylthiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 80) are in zwitterionic or pharmaceutically acceptable salt form. 10. Use of an intermediate compound selected from compounds (V), (VIII), (X), (XI), (XV), and (XVI) for the preparation of a compound of formula (I) according to any one of claims 1 to 9, wherein R1, R2, R3, R4, R5, L1, and A are as defined in claims 1 to 9, X2 is a halogen, preferably chlorine or bromine, and PG is a -(C1-C6)alkyl group. 11. A pharmaceutical composition comprising a compound according to any one of claims 1 to 9 and one or more pharmaceutically acceptable carriers or excipients mixed therewith. 12. A pharmaceutical composition according to claim 11, formulated for administration by inhalation. 13. A compound of formula (I) according to any one of claims 1 to 9 or a pharmaceutical composition according to claim 11 or 12, used as a medicament. 14. A compound of formula (I) or a pharmaceutical composition according to claim 13 for the prevention and / or treatment of diseases, disorders, or conditions related to sodium channel receptor mechanisms.15. A compound or pharmaceutical composition of formula (I) according to claim 13 or 14 for the prevention and / or treatment of respiratory diseases selected from the following: cough, subacute or chronic cough, treatment of resistant cough, intractable chronic cough, idiopathic chronic cough, post-viral cough, iatrogenic cough, asthma, idiopathic pulmonary fibrosis (IPF), chronic obstructive pulmonary disease (COPD), and cough associated with respiratory diseases such as COPD, asthma, and bronchospasm. 16. A compound or pharmaceutical composition of formula (I) according to claim 15 for the prevention and / or treatment of chronic cough. Claims 7 / 7 pages 8 CN 122180674 A Heteroarylamide derivative as a sodium channel inhibitor

[0001] Field of the Invention

[0002] The present invention generally relates to compounds (hereinafter referred to as NaV inhibitors) that inhibit the activity of voltage-gated sodium channels (NaV); the present invention relates to compounds that are heteroarylamide derivatives, including pharmaceutically acceptable salts thereof, methods for preparing said compounds, and their therapeutic uses.

[0003] The compounds of the present invention can be used, for example, to treat many disorders related to sodium channel receptor mechanisms, such as respiratory diseases.

[0004] Background of the Invention

[0005] Sensory afferent nerves carry impulses originating from various stimuli (heat, cold, thermal, mechanical, and chemical) and mediate the cough reflex.

[0006] Coughing is a defensive reflex designed to protect the airway from foreign objects and help remove debris from the airway. Chronic cough is defined as a cough lasting more than 8 weeks. Hypersensitive and parasitic cough states are often chronic, lasting more than three months and can manifest as many respiratory disease states, including asthma, chronic obstructive pulmonary disease (COPD), asthma-COPD overlap syndrome (ACOS), cough variant asthma, gastroesophageal reflux disease (GERD), idiopathic pulmonary fibrosis (IPF), and lung cancer. In addition, an inappropriate cough reflex can manifest acutely and persistently after viral infection. Ultimately, chronic cough can be idiopathic with an unknown etiology (Dicpinigaitis, P.V. et al., Pharmacol Rev., 2014, 66, 468-512; Patil, MJ et al., Pulmonary Pharmacology & Therapeutics, 2019, 56, 15-19).

[0007] Due to the failure of many cough suppression treatment strategies in clinical practice, increased attention in recent years has focused on potential inhibitors of voltage-gated sodium channels (NaV), which are absolutely necessary for conducting action potentials originating in nerve endings in response to various stimuli, including many inflammatory mediators. Voltage-gated sodium channels currently exist.The (NaV)α subunit family comprises at least nine known members. Phylogenetically, the NaV family has been divided into two subfamilies: NaV 1.x (all except SCN6A) and NaV 2.x (SCN6A). The NaV 1.x subfamily can be further functionally divided into two groups: those sensitive to tetrodotoxin blockade (TTX-sensitive or TTX-s) and those resistant to tetrodotoxin blockade (TTX-resistant or TTX-r). Coughing is a complex process triggered and regulated by two independent neural pathways that can interfere with each other: mechanosensitive vagal nodal Aδ fibers and chemosensitive vagal jugular C-fibers. Of the various known NaV 1.x subunits, only a few are expressed in the C-fibers of the vagus nerve and jugular vein in the respiratory tract, involving TTX-sensitive NaV 1.7 and TTX-resistant NaV 1.8 and NaV 1.9 (Kollarik, M., et al., J Physiol, 2018, 596, 1419-1432). NaV 1.7 (PN1, SCN9A) is sensitive to blockade by tetrodotoxin and is preferentially expressed in peripheral sympathetic and sensory neurons.

[0008] The cough reflex can be eliminated by effective sodium channel blockers (e.g., lidocaine). Lidocaine blocks all sensory input (e.g., in local anesthesia) and mechanoreceptors and motor neurons, resulting in numbness and paralysis. Lidocaine spray is routinely used to suppress the cough reflex prior to bronchoscopy. The inhalation of lidocaine for relieving intractable cough was first reported in 1977 (C Fletcher et al, Br Med J, 1977 1(6077), 1645-8), and then more recently confirmed in clinical trials as a throat spray for patients with intractable chronic cough (RCC) (Abdulqawi, R. et al., The Journal of Allergy and Clinical Immunology: In Practice, 2021, 9, 4, 1640-1647). However, lidocaine is short-acting, non-selective, weakly potent, and has off-target effects (burning sensation at injection), as indicated in the product information leaflet (page 1 / 106, CN 122180674 A), and its use (as a cough suppressant) is limited by its inferior risk / benefit profile (blocking the cardiac NaV 1.5 channel).

[0009] Sensory neurons express several channels with macropores (TRPV1, TRPA1, ASICS, P2X3, etc.), which, upon activation, allow charged molecules that normally cannot permeate the neuronal membrane to pass through. This differs from neutral molecules such as lidocaine, which can penetrate the lipid membrane surrounding the channel to reach the Nav binding site within the channel pore (Frazier et al., JPET,2015; 171, 1.; Brenneis et al, J Neuroscience, 2013, 33(1):315-26).

[0010] In 1999, Nortran Pharmaceutical filed a patent application (WO99 / 63985) relating to a pharmaceutical composition having antitussive activity and a method of treating warm-blooded animals affected by cough or bronchial stenosis by administering an effective amount of the inventive pharmaceutical composition. An example active antitussive agent reported therein is a charged quaternary ammonium compound, namely N,N-di(phenylcarbamoylmethyl)dimethylammonium chloride (i.e., carbochloride).

[0011] Carbochloride was first disclosed by Astra in 1962 for the treatment of arrhythmias (US 3255207). The antitussive effects of carcainium have been studied in various animal models: in guinea pig models, carcainium showed high antitussive efficacy and a mechanism of action somewhat different from lidocaine (J.J. Adcock, British Journal of Pharmacology, 2003, 138, 407-416). In 2013, Verona Pharma filed a patent application (WO2013 / 03490) concerning carcainium in the form of a salt (anion of a pharmaceutically acceptable acid) for the treatment and / or suppression of cough, cough attack, or cough flare-up in patients. This compound was found to be effective in preliminary studies in patients with chronic cough and idiopathic interstitial pneumonia (Lavorini et al, Pulmonary Pharmacology & Therapeutics, 2016, 40, 91-94) but was never commercialized because it failed to reduce the frequency and severity of cough in larger follow-up studies (Satia et al, Thorax, 2015;70 (Suppl 3): A1-A254).

[0012] Currently available cough medications often have limited efficacy. The quality of life of patients with cough can be severely affected, with physical (including fatigue, wheezing, sleep disturbances, and incontinence), psychological (including anger, frustration, depression, and anxiety), and social consequences. Therefore, continuous efforts are needed to find new and promising therapies and targets for the treatment of cough, and chronic cough remains an unmet medical need.

[0013] The prior art does not describe or suggest the general formula (I) amide derivative compounds of the present invention, which may represent a solution to the unmet medical needs mentioned above.

[0014] Summary of the Invention

[0015] In a first aspect, the present invention relates to compounds of formula (I)

[0016]

[0017] (I)

[0018] in the form of zwitterions or pharmaceutically acceptable salts,

[0019] Wherein at least one of Y and Z is S, and the other is CR4;

[0020] R1 and R2 are independently -(C1-C6)alkyl or fused to form -(C3-C7)heterocyclic alkyl, wherein the -(C3-C7)heterocyclic alkyl may optionally be substituted by one or more groups selected from halogens, -(C1-C6)alkyl and -(C1-C6)alkyl-OR8;

[0021] R3 and R4 are independently H or -(C1-C6)alkyl, or, in the case where Y is CR4, R3 and R4 are fused to form an aryl;

[0022] R5 and R6 are independently H or selected from -(C1-C6)alkyl, -(C1-C6)aminoalkyl, -(C1-C6)alkyl-OR8, - (C1-C6)alkyl-NR7R8, -(C1-C4)alkyl-NR7-C(O)OR8, -(C1-C4)alkyl-(C3-C7)heterocyclic alkyl and -(C3-C7)heterocyclic alkyl, or fused to form -(C3-C7)heterocyclic alkyl, wherein the -(C1-C4)alkyl-(C3-C7)heterocyclic alkyl, -(C3-C7)heterocyclic alkyl, -(C1-C6)aminoalkyl or -(C1-C6)alkyl-NR7R8 may optionally be substituted by one or more groups selected from -(C1-C6)alkyl, -C(O)OR8 and -SO2R7;

[0023] R7 and R8 are independently H or selected from -(C1-C6)alkyl, -(C3-C6)cycloalkyl and -(C1-C6)haloalkyl, or fused to form -(C3-C7)heterocyclic alkyl;

[0024] L1 is a bond or selected from -(C1-C6)alkylene- and -(C1-C6)cycloalkylene-;

[0025] A is a ring selected from aryl and heteroaryl, wherein any of the aryl or heteroaryl groups may optionally be substituted by one or more groups selected from -OR8, -C(O)OR8, -C(O)NR7R8 and -(C1-C6)alkyl; provided that, in the case that L1 is a bond, any of the aryl groups are not substituted by -(C1-C6)alkyl.

[0026] In a second aspect, the present invention relates to pharmaceutical compositions comprising a compound of formula (I) and one or more pharmaceutically acceptable carriers or excipients mixed therewith.

[0027] In a third aspect, the present invention relates to the use of a compound of formula (I) as a pharmaceutical remedy.

[0028] In yet another aspect, the present invention relates to the use of a compound of formula (I) for the treatment of diseases, disorders or conditions associated with voltage-gated sodium channel (NaV) dysregulation.

[0029] In another aspect, the present invention relates to compounds of formula (I) for the prevention and / or treatment of respiratory diseases selected from: cough, subacute or chronic cough, treatment of resistant cough, intractable chronic cough, idiopathic chronic cough, viral infection.Post-infection cough, iatrogenic cough, asthma, idiopathic pulmonary fibrosis (IPF), chronic obstructive pulmonary disease (COPD), and cough associated with respiratory diseases such as COPD, asthma, and bronchospasm.

[0030] Detailed Description of the Invention

[0031] Unless otherwise specified, the term "compound of formula (I)" means its stereoisomers, tautomers, pharmaceutically acceptable salts, zwitterionic forms, or solvates.

[0032] The term "stereoisomer" refers to isomers with identical constitutions but different atomic spatial arrangements. Enantiomers and diastereomers are examples of stereoisomers.

[0033] The term "enantiomer" refers to one of a pair of molecular species that are mirror images of each other and do not overlap.

[0034] The term "diastereomer" refers to a stereoisomer that is not a mirror image of the other.

[0035] The term "racemic mixture" or "racemic mixture" refers to a composition consisting of two enantiomers in equimolar amounts, wherein the composition is optically inactive.

[0036] The symbols "R" and "S" represent the configuration of the substituents surrounding one or more chiral carbon atoms and are intended to be used as defined in the literature (IUPAC Recommendations 1996, Pure and Applied Chemistry, 68:2193-2222 (1996)).

[0037] The term "tautomer" refers to each of two or more isomers of a compound, which are present together in equilibrium and readily interconvertible by the migration of intramolecular atoms or groups.

[0038] The compound of formula (I) is a quaternary ammonium compound. The term "pharmaceutically acceptable salt" as used herein refers to the compound of formula (I), wherein the compound is provided as an addition salt of any acid conventionally expected to be pharmaceutically acceptable. The salt described in the specification 3 / 106 pages 11 CN 122180674 A is an example that may include inorganic or organic acid addition salts of quaternary ammonium fractions, such as salts of hydrochloric acid, hydrobromic acid, iodic acid, formic acid, benzoic acid, sulfuric acid, nitric acid, phosphoric acid, methanesulfonic acid, camphorsulfonic acid, p-toluenesulfonic acid, acetic acid, trifluoroacetic acid, oxalic acid, maleic acid, fumaric acid, succinic acid, 2-naphthoic acid, tartaric acid, 1-hydroxy-2-naphthoic acid, naphthalene-2,7-disulfonic acid, and citric acid.

[0039] Alternatively, the compound of formula (I) according to the invention may be provided as an "amphoteric form," also known as an internal salt or dipole ion, which is a form in which both positive and negative charges are present.

[0040] The term "solvent" means the physical combination of the compound of the invention with one or more solvent molecules (whether organic or inorganic). This physical combination includes the formation of hydrogen bonds. In some cases, the solvate may be separated by crystallization, for example in aThis is true when one or more solvent molecules are incorporated into the lattice of a crystalline solid. The solvate may contain stoichiometric or non-stoichiometric amounts of solvent molecules.

[0041] The terms "halogen" or "halogen atom" or "halogenated" as used herein include fluorine, chlorine, bromine, and iodine atoms.

[0042] The term "-(Cx-Cy)alkyl" where x and y are integers refers to a straight-chain or branched alkyl group having x to y carbon atoms. Thus, for example, in the case where x is 1 and y is 6, the term includes methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, and n-hexyl.

[0043] The term "-(Cx-Cy)alkylene-" where x and y are integers refers to a (Cx-Cy)alkyl residue having a total of two unsatisfied valences. Examples of the "-(Cx-Cy)alkylene-" group may include, for example, divalent methylene residues and -CH(CH3)-.

[0044] The term "-(Cx-Cy)haloalkyl" where x and y are integers refers to the "(Cx-Cy)alkyl" group as defined above, wherein one or more hydrogen atoms are replaced by one or more halogen atoms that can be the same or different. Examples of the "-(Cx-Cy)haloalkyl" group may thus include halogenated, polyhalogenated and fully halogenated alkyl groups, wherein all hydrogen atoms are replaced by halogen atoms, such as trifluoromethyl.

[0045] The term "-(Cx-Cy)aminoalkyl" where x and y are integers refers to the "(Cx-Cy)alkyl" group as defined above, wherein one or more hydrogen atoms are replaced by one or more amino groups that can be the same or different. Examples of "-(Cx-Cy)aminoalkyl" include, for example, aminomethyl.

[0046] The term "-(Cx-Cy)hydroxyalkyl" where x and y are integers refers to the "-(Cx-Cy)alkyl" group as defined above, wherein one or more hydrogen atoms are replaced by one or more hydroxyl (OH) groups. Examples of "-(Cx-Cy)hydroxyalkyl" include, for example, hydroxymethyl.

[0047] The term "partially unsaturated" refers to a ring moiety comprising at least one double or triple bond. The term "partially unsaturated" is intended to cover rings having multiple unsaturated sites but is not intended to include aryl or heteroaryl moieties as defined herein.

[0048] The term "-(Cx-Cy)cycloalkyl" where x and y are integers refers to a monocyclic or polycyclic hydrocarbon group containing the stated number of ring carbon atoms and either saturated or partially unsaturated. Examples include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, and cycloheptyl.

[0049] The term "(Cx-Cy)cycloalkylene" where x and y are integers refers to a monocyclic or polycyclic cycloalkyl group containing the stated number of ring carbon atoms and either saturated or partially unsaturated, having a total of two unsatisfied valences on the same carbon atom.Examples of "cycloalkyl-" include "-(C1-C4)cycloalkylene-" and "-(C1-C6)cycloalkylene-", such as cyclopropylene.

[0050] The term "-(Cx-Cy)heterocyclic alkyl", where x and y are integers, refers to a saturated or partially unsaturated monocyclic or polycyclic or polycyclic spirocyclic group containing the stated number of ring carbon atoms, wherein at least one ring carbon atom is replaced by at least one heteroatom or heterogroup (e.g., N, NH, S, or O) or carries an oxo (=O) substituent. The heterocyclic alkyl (i.e., heterocyclic residue or group) may be optionally substituted by one or more specific groups at possible positions in the ring, i.e., on a carbon atom, or on a possible substituted heteroatom or heterogroup. In the presence of more than one substituent, they may involve the same or adjacent positions in the ring, for example, "(C3-C10)heterocyclic alkyl" may be 1,1- or 1,2- or 1,3-disubstituted. Carbon Atom Specification 4 / 106 pages 12 CN The substitution on 122180674 A includes spirodisubstituted groups, wherein an additional spirofused 3- to 6-membered ring is formed. The spirodisubstituted groups are included as polycyclic spiro groups in the meaning of "(C3-C10) heterocyclic alkyl". Non-limiting examples of "(Cx-Cy) heterocyclic alkyl" are represented by pyrrolidinyl, piperazinyl, piperidinyl, morpholinyl, thiomorpholinyl, dihydro- or tetrahydro-pyridinyl, oxocyclic butyl, isoindolyl, tetrahydropyranyl, pyranyl, dihydro- or tetrahydrofuranyl.

[0051] The term "-(Cx-Cy) heterocyclic alkyl-" where x and y are integers refers to a saturated or partially unsaturated mono- or poly(Cx-Cy) heterocyclic alkyl group containing the stated number of cyclic carbon atoms, having a total of two unsatisfied valences on the same carbon atom.

[0052] The group may be optionally substituted, wherein the term "optionally substituted" means substituted or unsubstituted. In the case where the term "one or more" refers to any atom or group acting as a substituent for a compound of formula (I), it is desirable that 1 to 3, preferably 1 to 2, more preferably 1, said substituent can replace the hydrogen on said variable.

[0053] The term "aryl" refers to a monocyclic or bicyclic unsaturated or partially unsaturated carbocyclic system, wherein at least one ring is aromatic. Examples of suitable aryl ring systems include, for example, phenyl, dihydro-1H-indenyl, or naphthyl.

[0054] The term "heteroaryl" refers to a monocyclic, bicyclic, or tricyclic system having 5 to 20, preferably 5 to 15 ring atoms, wherein at least one ring is aromatic and wherein at least one ring atom is a heteroatom (e.g., N, NH, S, or O). Examples of "heteroaryl" include thiophene, benzothiophene, pyrrole, pyrazolyl, imidazolyl, isoxazolyl, oxazolyl, isothiazolyl, thiazolyl, pyridinyl, pyrazinyl, and triazinyl.

[0055] Keys pointing to waveforms or wavy lines, such as those used in the structural formulas herein, describe such keys, which are...The attachment point of the substituent or substituent to the core or skeletal structure.

[0056] The term "bond" used to define a substituent refers to a situation where the two functional groups to which the substituent is attached are directly connected to each other without any additional atoms between them.

[0057] The dash ("-") not between two letters or symbols is intended to represent the attachment point of the substituent.

[0058] The terms "sodium channel receptor" or "voltage-gated sodium channel receptor" or "NaV receptor" used to refer to the inhibitory activity of the compounds of the present invention are intended to include members of the voltage-gated sodium channel (NaV) α subunit family expressed in the C-fibers of the jugular vein of the vagus nerve of the respiratory tract, namely NaV 1.7, NaV 1.8 and NaV 1.9.

[0059] The term "NaV inhibitor" means that the compounds of the present invention are inhibitors of NaV receptors, especially but not limited to NaV 1.7 receptors.

[0060] The term "IC50" refers to the half-maximal inhibitory concentration, which serves as a measure of the efficacy of a substance in inhibiting a specific biological or biochemical function.

[0061] The term "pIC50" refers to the negative logarithm of the IC50 value expressed as a molar concentration.

[0062] Whenever other basic amino or quaternary ammonium groups are present in the compound of formula (I), pharmaceutically acceptable anions selected from chloride, bromide, iodide, trifluoroacetate, formate, sulfate, phosphate, methanesulfonate, nitrate, maleate, acetate, citrate, fumarate, tartrate, oxalate, succinate, benzoate, p-toluenesulfonate, emipamoate, sine, and naphthalenedisulfonate may be present. Similarly, in the presence of acidic groups such as COOH groups, corresponding pharmaceutically acceptable cations may be present, including alkali metal or alkaline earth metal ions, such as sodium, potassium, calcium, etc.

[0063] As noted above, the present invention relates to a series of compounds represented by general formula (I) having inhibitory activity against voltage-gated sodium channel receptors, as described in detail below.

[0064] The inhibitory activity of compounds of formula (I) was tested in CHO cells stably expressing human NaV 1.7 channels in an automated patch-clamp assay, as described in the experimental section. Specification 5 / 106 pages 13 CN 122180674 A

[0065] pIC50 values ​​were calculated for some of the compounds of the present invention, demonstrating the intracellular inhibition of NaV 1.7 receptors by the test compounds (see Qian, Binbin et al, Current protocols in pharmacology, 2020, 89, 1).

[0066] Indeed, the compounds of formula (I) of the present invention are capable of acting as inhibitors of NaV 1.7 in a substantial and effective manner. In particular, as noted in the experimental section, the compounds of formula (I) of the present invention show NaVIntracellular inhibition of NaV 1.7, as reported in Table 2 for representative compounds.

[0067] Compounds of formula (I) are heteroaryl quaternary ammonium amide derivatives in salt or zwitterionic form, exhibiting particularly good inhibitory activity against NaV 1.7.

[0068] As noted in the comparative compound section of the experimental section, particularly in Table 4, the presence of an amide-substituted tiophene or benzothiophene ring in the compounds of the present invention unexpectedly and significantly determines the associated increase in intracellular inhibitory activity against the NaV 1.7 receptor, contrary to carbaryl, which is considered a comparative compound C1.

[0069] Furthermore, as noted in the same experimental section, the reported data show that, contrary to comparative compound C2, which is characterized by a different heteroaryl ring, the presence of an amide-substituted tiophene or benzothiophene ring in the compounds of the present invention unexpectedly and notably determines the associated increase in inhibitory activity against the NaV receptor.

[0070] Compounds of formula (I) in pharmaceutically acceptable salt or zwitterionic form, hereinafter alternatively referred to as compounds of the present invention, can be used as medicines. Accordingly, the present invention also provides the use of compounds of formula (I) in pharmaceutically acceptable salt or zwitterionic form as medicines.

[0071] Compounds of formula (I) of the present invention can act as inhibitors of NaV 1.7 receptors in a substantial and effective manner particularly recognized by those skilled in the art (in searching for compounds that may treat respiratory diseases such as cough, asthma, IPF and COPD, and especially chronic cough).

[0072] Accordingly, compounds of formula (I) can be used to treat respiratory diseases selected from cough, asthma, IPF and COPD, and especially chronic cough, provided that NaV receptors are involved.

[0073] As can be appreciated, the compounds of the present invention according to Table 2 show their efficacy in terms of intracellular inhibitory activity against NaV 1.7 receptors, expressed as pIC50 values ​​equal to or greater than 5, preferably 5 to 5.5, more preferably equal to or greater than 5.5.

[0074] More advantageously, compounds represented by general formula (I) are able to inhibit one or more voltage-gated ion channels when exposed to or administered to cells, but show little or no inhibition when administered extracellularly.

[0075] A similar protocol was used for extracellular inhibition, i.e., after exchanging the external solution for a solution containing the test compound, compounds of formula (I) were also tested in an automated patch-clamp assay: the experimental values ​​obtained were used to calculate pIC50 values ​​representing extracellular inhibition of the NaV 1.7 receptor.

[0076] As noted in the experimental section, compounds of formula (I) of the present invention exhibit lower extracellular inhibition of NaV 1.7 than intracellular inhibition, as shown in Table 2.

[0077] Accordingly, the compounds of the present invention according to Table 2 show intracellular efficacy versus extracellular efficacy regarding NaV 1.7 inhibition.The difference between the inhibitors is defined as DELTA, which is equal to or greater than 0.5, preferably between 0.5 and 1, and more preferably equal to or greater than 1.

[0078] This difference between intracellular and extracellular inhibition can result in higher safety of the compound for sensory neurons.

[0079] The compounds of the present invention have also been tested in safety pharmacology tests, particularly in in vitro tests of human ether-a-go-go related gene (hERG) channels. Safety pharmacology studies are non-clinical studies that test the safety of new drugs in humans from a pharmacological perspective. Guidelines for safety pharmacology studies aimed at examining the safety of test substances in humans and predicting their adverse effects have been established at the International Conference on Harmonisation of Technical Requirements for Registration of Pharmaceuticals for Human Use (ICH) between Europe, Japan, and the United States. Specification 6 / 106 pages 14 CN 122180674 A According to these guidelines, as part of safety pharmacology studies, it is necessary to examine the arrhythmic effects of the test substance, especially the presence or absence of QT interval prolongation on electrocardiogram. The QT interval is an electrocardiogram measurement used to evaluate certain electrical properties of the heart and approximates the time it takes for the ventricles to begin systole and then to end diastole. To protect patients from ventricular tachycardia, torsades de pointes, and lethal arrhythmias associated with drug-induced QT interval prolongation, it is important in drug development to detect QT interval prolongation effects that can induce these serious adverse effects.

[0080] To date, many drugs with QT interval prolongation effects have been known to inhibit delayed rectifier potassium channels in cardiomyocytes. hERG channels are believed to act as major component proteins in delayed rectifier potassium channels. Therefore, in the draft guidelines for non-clinical evaluation of the ability of human drugs to delay ventricular repolarization (QT interval prolongation), the use of hERG channel-transfer cell ion channel assays as non-clinical studies is recommended.

[0081] In addition to its significantly effective inhibitory activity against NaV receptors, the in vitro inhibition of hERG channels by the compound of formula (I) of the present invention was also tested and showed low toxicity to hERG, thereby reducing the likelihood of adverse events observed during clinical studies.

[0082] Thus, in one aspect, the present invention relates to compounds of general formula (I)

[0083]

[0084] (I)

[0085] in zwitterionic or pharmaceutically acceptable salt form,

[0086] wherein at least one of Y and Z is S, and the other is CR4;

[0087] R1 and R2 are independently -(C1-C6)alkyl or fused to -(C3-C7)heteroalkyl, wherein the -(C3-C7)heteroalkylThe alkyl group may optionally be substituted with one or more groups selected from halogens, -(C1-C6)alkyl, and -(C1-C6)alkyl-OR8;

[0088] R3 and R4 are independently H or -(C1-C6)alkyl, or, in the case that Y is CR4, R3 and R4 are fused to form an aryl group;

[0089] R5 and R6 are independently H or selected from -(C1-C6)alkyl, -(C1-C6)aminoalkyl, -(C1-C6)alkyl-OR8, -(C1-C6)alkyl-NR7R8, -(C1-C4)alkyl-NR7-C(O)OR8, -(C1-C4)alkyl-(C3-C7)heterocyclic alkyl, and -(C3-C7) Heterocyclic alkyl groups, or fused to form -(C3-C7) heterocyclic alkyl groups, wherein the -(C1-C4)alkyl-(C3-C7) heterocyclic alkyl groups, -(C3-C7) heterocyclic alkyl groups, -(C1-C6) aminoalkyl groups, or -(C1-C6) alkyl-NR7R8 may optionally be substituted by one or more groups selected from -(C1-C6) alkyl groups, -C(O)OR8, and -SO2R7;

[0090] R7 and R8 are independently H or selected from -(C1-C6) alkyl groups, -(C3-C6) cycloalkyl groups, and -(C1-C6) haloalkyl groups, or fused to form -(C3-C7) heterocyclic alkyl groups;

[0091] L1 is a bond or selected from -(C1-C6) alkylene- and -(C1-C6) cycloalkylene-;

[0092] A is a ring selected from aryl and heteroaryl groups, wherein any of the aryl or heteroaryl groups may optionally be substituted by one or more groups selected from -OR8, -C(O)OR8, -C(O)NR7R8 and -(C1-C6)alkyl; provided that, in the case that L1 is a bond, any of the aryl groups are not substituted by -(C1-C6)alkyl groups.

[0093] All the listed groups in the variable parts Y, Z, R1, R2, R3, R4, R5, R6, R7, R8, L1 and A of the compounds of the present invention must be considered as alternatives and can be combined with each other to form embodiments included within the scope of the present invention.

[0094] The 5-membered ring containing Y and Z is an aromatic ring, i.e. a thiophene ring, substituted by the -C(O)NR5R6 group.

[0095] Preferred halogens, individually and in the -(Cx-Cy) haloalkyl substituents, are fluorine and bromine, wherein fluorine is more preferred.

[0096] In a preferred embodiment, Z is S and Y is CR4. In a more preferred embodiment, Z is S and Y is CH.

[0097] In yet another embodiment, Y is S and Z is CR4, wherein R4 is preferably hydrogen.

[0098] In one embodiment, R1 and R2 are independently -(C1-C6) alkyl or fused to form -(C3-C7) heterocyclic alkyl, whereinThe -(C3-C7) heterocyclic alkyl group described herein may optionally be substituted with one or more groups selected from halogens, -(C1-C6)alkyl, and -(C1-C6)alkyl-OR8. In a preferred embodiment, R1 and R2 are independently -(C1-C6)alkyl. In a more preferred embodiment, R1 and R2 are independently -(C1-C4)alkyl. In an even more preferred embodiment, R1 and R2 are methyl. In yet another preferred embodiment, R1 and R2 are fused to form a -(C3-C7) heterocyclic alkyl group, optionally substituted with one or more groups selected from halogens, -(C1-C6)alkyl, and -(C1-C6)alkyl-OR8. In a more preferred embodiment, R1 and R2 are fused to form a -(C3-C7) heterocyclic alkyl group, optionally substituted with one or more groups selected from halogens, methyl, and -(C1-C4)alkyl-OR8. In the most preferred embodiment, R1 and R2 are fused to optionally substituted piperidinyl or aziridine heptyl.

[0099] In one embodiment, R3 and R4 are independently H or -(C1-C6)alkyl, or, if Y is CR4, R3 and R4 are fused to form an aryl group. In yet another embodiment, R3 and R4 are H. In a more preferred embodiment, R3 and R4 are independently H or -(C1-C4)alkyl; even more preferably, R3 and R4 are independently H or methyl, most preferably R3 is methyl and R4 is H. In the most preferred embodiment, Z is S and Y is CH, and R3 is H or methyl.

[0100] In yet another preferred embodiment, R3 and R4 are fused to form an optionally substituted phenyl ring, which is fused to a 5-membered ring to form an optionally substituted benzothiophene ring system.

[0101] In one embodiment, R5 and R6 are independently H or selected from -(C1-C6)alkyl, -(C1-C6)aminoalkyl, -(C1-C6)alkyl-OR8, -(C1-C6)alkyl-NR7R8, -(C1-C4)alkyl-NR7-C(O)OR8, -(C1-C4)alkyl-(C3-C7)heterocyclic alkyl and -(C3-C7)heterocyclic alkyl, or fused to form -(C3-C7)heterocyclic alkyl, wherein the -(C1-C4)alkyl-(C3-C7)heterocyclic alkyl, -(C3-C7)heterocyclic alkyl, -(C1-C6)aminoalkyl or -(C1-C6)alkyl-NR7R8 may optionally be substituted by one or more groups selected from -(C1-C6)alkyl, -C(O)OR8 and -SO2R7. In yet another embodiment, R5 and R6 are independently H or selected from -(C1-C6)alkyl, -(C1-C6)aminoalkyl, -(C1-C6)alkyl-OR8, -(C1-C6)alkyl-NR7R8, -(C1-C4)alkyl-NR7-C(O)OR8, -(C1-C4)alkyl-(C3-C7)heterocyclic alkyl, and -(C3-C7)heterocyclic alkyl, wherein the -(C1-C4)alkyl-(C3-C7)heterocyclic alkyl, -(C3-C7)heterocyclic alkyl, -(C1-C6)aminoalkyl, or -(C1-C6)alkyl-NR7R8 may optionally be substituted with one or more groups selected from -(C1-C6)alkyl, -C(O)OR8, and -SO2R7. In a preferred embodiment, R5 and R6 are independently H or -(C1-C6)alkyl. In a more preferred embodiment, R5 and R6 are independently H or -(C1-C4)alkyl. In an even more preferred embodiment, R5 and R6 are independently H or Me. In yet another preferred embodiment, R5 and R6 are independently selected from H, -(C1-C4)aminoalkyl, -(C1-C4)alkyl-OR8, -(C1-C4)alkyl-NR7R8, -(C1-C4)alkyl-NR7-C(O)OR8, -(C1-C4)alkyl-(C3-C7)heterocyclic alkyl, and -(C3-C7)heterocyclic alkyl, wherein the -(C1-C4)alkyl-(C3-C7)heterocyclic alkyl, -(C3-C7)heterocyclic alkyl, -(C1-C4)aminoalkyl, or -(C1-C4)alkyl-NR7R8 may optionally be substituted by one or more groups selected from -(C1-C4)alkyl, -C(O)OR8, and -SO2R7. In another embodiment, R5 and R6 are fused into a -(C3-C7) heterocyclic alkyl group, wherein the -(C3-C7) heterocyclic alkyl group may optionally be substituted with one or more groups selected from -(C1-C6) alkyl, -C(O)OR8, and -SO2R7. In a preferred embodiment, R5 and R6 are fused into a -(C3-C7) heterocyclic alkyl group, wherein the -(C3-C7) heterocyclic alkyl group may optionally be substituted with one or more groups selected from -(C1-C4) alkyl. In a more preferred embodiment, R5 and R6 are fused into a -(C4-C7) heterocyclic alkyl group selected from pyrrolidine, piperazine, and morpholine, wherein the -(C3-C7) heterocyclic alkyl group may optionally be substituted with one or more groups selected from -(C1-C4) alkyl.

[0102] In one embodiment, R7 and R8 are independently H or selected from -(C1-C6)alkyl, -(C3-C6)cycloalkyl, and -(C1-C6)haloalkyl, or fused to form -(C3-C7)heteroalkyl. In a preferred embodiment, R7 and R8 are independently H or -(C1-C4)alkyl. In yet another preferred embodiment, R7 and R8 are independently H or methyl. In still a preferred embodiment, R7 and R8 are H or R7 and R8 are methyl.

[0103] In one embodiment, L1 is a bond or selected from -(C1-C6)alkylene- and -(C1-C6)cycloalkylene-. In a preferred embodiment, L1 is a bond or selected from -(C1-C4)alkylene- and -(C1-C4)cycloalkylene-. In yet another preferred embodiment, L1 is a bond. In yet another more preferred embodiment, L1 is selected from methylene, cyclopropylene, and -CH(CH3)-.

[0104] In one embodiment, A is a ring selected from aryl and heteroaryl groups, wherein any of the aryl or heteroaryl groups may optionally be substituted by one or more groups selected from -OR8, -C(O)OR8, -C(O)NR7R8, and -(C1-C6)alkyl; provided that if L1 is a bond, any of the aryl groups are not substituted by -(C1-C6)alkyl groups. In a preferred embodiment, A is a heteroaryl group, optionally substituted with one or more groups selected from -OR8, -C(O)OR8, -C(O)NR7R8, and -(C1-C6)alkyl. In yet another preferred embodiment, A is a heteroaryl group selected from optionally substituted thiophene, benzothiophene, pyrrole, pyrazolyl, imidazolyl, isoxazolyl, oxazolyl, isothiazolyl, thiazolyl, pyridinyl, pyrazinyl, and triazinyl. In yet another preferred embodiment, A is an aryl group, optionally substituted with one or more groups selected from -OR8, -C(O)OR8, -C(O)NR7R8, and -(C1-C6)alkyl, provided that, if L1 is a bond, none of the aryl groups are substituted with -(C1-C6)alkyl. In a preferred embodiment, A is an aryl group, wherein any of the aryl group may optionally be substituted by one or more groups selected from -OR8, -C(O)OR8, and -C(O)NR7R8. In a more preferred embodiment, A is selected from phenyl, tiophenyl, and isoxazolyl, which may optionally be substituted by one or more groups selected from -OR8, -C(O)OR8, and -C(O)NR7R8. In other preferred embodiments, A is phenyl or selected from unsubstituted isoxazolyl or isoxazolyl substituted with one or more methyl groups.

[0105] All preferred groups of the variable parts Y, Z, R1, R2, R3, R4, R5, R6, R7, R8, L1, and A of the compounds of the present invention listed above may be combined with each other to form embodiments included within the scope of the present invention.

[0106] In yet another preferred embodiment, the present invention relates to compounds of formula (I) as defined above, wherein

[0107] Z, Y, R1, R2, R3, R4, R5, R6, R7, R8 are as defined above;

[0108] L1 is selected from -(C1-C4)alkylene- and -(C1-C4)cycloalkylene-; and

[0109] A is a ring selected from aryl and heteroaryl groups, wherein any aryl or heteroaryl group may optionally be substituted by one or more groups selected from -OR8, -C(O)OR8, -C(O)NR7R8, and -(C1-C4)alkyl.

[0110] In a preferred embodiment, L1 is selected from -(C1-C4)alkylene and -(C1-C4)cycloalkylene. In an even more preferred embodiment, L1 is selected from methylene, cyclopropylene, and -CH(CH3)-.

[0111] In yet another preferred embodiment, A is a ring selected from aryl and heteroaryl groups, wherein any aryl or heteroaryl group may optionally be substituted by one or more groups selected from -OR8, -C(O)OR8, and -C(O)NR7R8. In a more preferred embodiment, A is a heteroaryl group, wherein any heteroaryl group may optionally be substituted by one or more groups selected from -OR8, -C(O)OR8, and -C(O)NR7R8. In yet another preferred embodiment, A is an aryl group, wherein any of the aryl group may optionally be substituted by one or more groups selected from -OR8, -C(O)OR8 and -C(O)NR7R8.

[0112] In yet another embodiment, the present invention relates to a compound of formula (I), wherein L1 is a bond, represented by formula (Ia): Specification 9 / 106 pages 17 CN 122180674 A

[0113]

[0114] (Ia)

[0115] in zwitterionic or pharmaceutically acceptable salt form,

[0116] wherein

[0117] Z, Y, R1, R2, R3, R4, R5, R6, R7 and R8 are as defined above; and

[0118] A is a ring selected from aryl and heteroaryl, wherein any of the aryl groups may optionally be substituted by one or more groups selected from -OR8, -C(O)OR8 and -C(O)NR7R8, and wherein any of the heteroaryl groups may optionally be substituted by one or more groups selected from -OR8, -C(O)OR8, -C(O)NR7R8 and -(C1-C6)alkyl.

[0119] In a more preferred embodiment, A is a heteroaryl group, wherein any of the heteroaryl groups may optionally be substituted with one or more -OR8, -C(O)OR8, -C(O)NR7R8, and -(C1-C6)alkyl groups. In an even more preferred embodiment, A is a heteroaryl group, wherein any of the heteroaryl groups are selected from thiophene, pyrrolo, pyrazolyl, imidazolyl, isoxazolyl, oxazolyl, isothiazolyl, thiazolyl, pyridinyl, pyrimidinyl, pyrazinyl, and triazinyl groups, and any of the heteroaryl groups may optionally be substituted with one or more methyl, ethyl, isopropyl, or tert-butyl groups.

[0120] In yet another preferred embodiment, the present invention relates to compounds of formula (I) as defined above,

[0121] wherein

[0122] At least one of Y and Z is S, and the other is CR4;

[0123] R1 and R2 are independently -(C1-C4)alkyl or fused to form -(C3-C7)heteroalkyl, wherein the -(C3-C7)heteroalkyl may optionally be substituted by one or more groups selected from halogens, -(C1-C4)alkyl and -(C1-C4)alkyl-OR8;

[0124] R3 and R4 are independently H or -(C1-C6)alkyl, or wherein Y is CR4, R3 and R4 are fused to form an aryl, preferably optionally a substituted aryl;

[0125] R5 and R6 are independently H or selected from -(C1-C4)alkyl, -(C1-C4)aminoalkyl, -(C1-C4)alkyl-OR8, - (C1-C4)alkyl-NR7R8, -(C1-C4)alkyl-NR7-C(O)OR8, -(C1-C4)alkyl-(C3-C7)heterocyclic alkyl, -(C3-C7)heterocyclic alkyl, or fused to form -(C3-C7)heterocyclic alkyl, wherein the -(C1-C4)alkyl-(C3-C7)heterocyclic alkyl, -(C3-C7)heterocyclic alkyl, -(C1-C4)aminoalkyl or -(C1-C4)alkyl-NR7R8 may optionally be substituted by one or more groups selected from -(C1-C4)alkyl, -C(O)OR8 and -SO2R7;

[0126] R7 and R8 are independently H or selected from -(C1-C4)alkyl, -(C3-C6)cycloalkyl and -(C1-C4)haloalkyl, or fused to form -(C6-C7)heterocyclic alkyl;

[0127] L1 is a bond or selected from -(C1-C4)alkylene and -(C1-C4)cycloalkylene;

[0128] A is a ring selected from aryl and heteroaryl, wherein any of the aryl or heteroaryl groups may optionally be substituted by one or more groups selected from -OR8, -C(O)OR8, -C(O)NR7R8 and -(C1-C4)alkyl; provided that if L1 is a bond, any of the aryl groups are not substituted by -(C1-C4)alkyl.

[0129] In a preferred embodiment, R1 and R2 are fused to form -(C6-C7)heteroalkyl, wherein the -(C6-C7)heteroalkyl group may optionally be substituted by one or more groups selected from -OR8, -(C1-C6)alkyl and -(C3-C7)heteroalkyl.

[0130] In yet another preferred embodiment, the present invention relates to compounds of formula (I) as defined above, specification 10 / 106 pages 18 CN 122180674 A

[0131] wherein Z is S and Y is CH or Z is CH and Y is S; R3 is H or methyl, and R1, R2, R5, R6, R7, R8, L1, A are as defined above.

[0132] In a preferred embodiment, the present invention relates to at least one of the compounds listed in Table 1 below, which are zwitterionic.In its form or as a pharmaceutically acceptable salt.

[0133] Table 1 - List of Preferred Compounds

[0134] Specification 11 / 106 pages 19 CN 122180674 A

[0135] Specification 12 / 106 pages 20 CN 122180674 A

[0136] Specification 13 / 106 pages 21 CN 122180674 A

[0137] Specification 14 / 106 pages 22 CN 122180674 A

[0138] Specification 15 / 106 pages 23 CN 122180674 A

[0139] Specification 16 / 106 pages 24 CN 122180674 A

[0140] Specification 17 / 106 pages 25 CN 122180674 A

[0141] Specification 18 / 106 pages 26 CN 122180674 A

[0142] Specification 19 / 106 pages 27 CN 122180674 A

[0143] Specification 20 / 106 pages 28 CN 122180674 A

[0144] Specification 21 / 106 pages 29 CN 122180674 A

[0145] Specification 22 / 106 pages 30 CN 122180674 A

[0146] Specification 23 / 106 pages 31 CN 122180674 A

[0147] Specification 24 / 106 pages 32 CN 122180674 A

[0148]

[0149] It should be understood that all individual enantiomers, diastereomers and mixtures thereof in any proportion of the compounds of formula (I) of the present invention are covered within the scope of the present invention.

[0150] In a preferred embodiment, the present invention relates to compounds of formula (I) as NaV inhibitors. In this regard, compounds of formula (I) of the present invention have been found to have inhibitory pharmacological efficacy equal to or greater than 5, expressed as a pIC50 against the NaV 1.7 receptor.

[0151] Preferably, compounds of the present invention have a pIC50 of 5 to 5.5 against NaV 1.7. More preferably, compounds of the present invention have a pIC50 of 5.5 to 5.5 against NaV 1.7. Even more preferably, compounds of the present invention have a pIC50 of 5.5 to 6 against NaV 1.7.

[0152] The present invention also relates to pharmaceutical compositions comprising compounds of formula (I) in zwitterionic or pharmaceutically acceptable salt form, and at least one or more pharmaceutically acceptable carriers and / or excipients mixed therewith.

[0153] As used in this application, regarding compounds of formula (I) (in zwitterionic or pharmaceutically acceptable salt form) or otherThe "effective amount" of a pharmaceutically active agent means an amount of compound sufficient to treat a patient's condition but low enough to avoid serious side effects, and it can still be routinely determined by those skilled in the art.

[0154] Compounds of Formula (I) may be administered once or according to a dosage regimen in which multiple doses are administered at varying time intervals for a given duration. Typical daily doses may vary depending on the chosen route of administration.

[0155] In one embodiment, the present invention relates to pharmaceutical compositions of compounds of Formula (I) mixed with one or more pharmaceutically acceptable carriers or excipients (e.g., those described in Remington's Pharmaceutical Sciences Handbook, XVII Ed., Mack Pub., NY, USA).

[0156] In therapeutic applications, compounds of Formula (I) may be administered by any convenient, suitable, or effective route. Administration of the compounds of the present invention and pharmaceutical compositions thereof may be achieved as needed by the patient, for example, orally, nasally, parenterally (subcutaneously, intravenously, intramuscularly, intrasternally, and by infusion), and by inhalation.

[0157] Preferably, the compounds of the present invention are administered orally or by inhalation.

[0158] In a more preferred embodiment, the compound of the present invention or a pharmaceutical composition thereof is administered by inhalation.

[0159] In yet another preferred embodiment, the compound of the present invention or a pharmaceutical composition thereof is administered orally.

[0160] In a preferred embodiment, the pharmaceutical composition comprising a compound of formula (I) in the form of a zwitterion or a pharmaceutically acceptable salt is a solid oral dosage form such as tablets, gel caps, capsules, tablets, granules, sugar tablets, and packaged powders. Specification 25 / 106 pages 33 CN 122180674 A

[0161] The compounds of the present invention can be administered alone or in combination with various pharmaceutically acceptable carriers, diluents (e.g., sucrose, mannitol, lactose, starch) and known excipients, including suspending agents, solubilizers, buffers, binders, disintegrants, preservatives, colorants, flavoring agents, lubricants, etc.

[0162] In another embodiment, the pharmaceutical composition comprising the compound of formula (I) is a liquid oral dosage form such as a solution of water and non-aqueous substances, emulsions, suspensions, or syrups. Such liquid dosage forms may also contain suitable known inert diluents (e.g., water) and suitable known excipients (e.g., preservatives, wetting agents, sweeteners, flavoring agents, etc.), as well as agents for emulsifying and / or suspending the compounds of the present invention.

[0163] In another embodiment, the pharmaceutical composition comprising the compound of formula (I) is an inhalable formulation such as an inhalable powder, a metered-dose aerosol containing propellant, or an inhalable formulation without propellant. The inhalable formulation can be administered via a suitable inhalation device, which may be selected from dry powder inhalers, pressurized metered-dose inhalers, or nebulizers.

[0164] In another embodiment, the invention also relates to an apparatus comprising a pharmaceutical composition comprising a compound of formula (I) according to the invention (obtained according to the invention as described above) in the form of a zwitterion or a pharmaceutically acceptable salt, in the form of a single-dose or multi-dose dry powder inhaler or a metered-dose inhaler.

[0165] For administration as a dry powder, single-dose or multi-dose inhalers known in the art can be used. In this case, the powder can be filled into gelatin, plastic or other capsules, cartridges or blister packs or reservoirs.

[0166] A diluent or carrier chemically inert to the compound of the invention, such as lactose or any other additive suitable for improving the inhalable fraction, can be added to the powdered compound of the invention.

[0167] Inhalation aerosols containing propellant gases such as hydrofluorocarbons can contain the compound of the invention in solution or dispersion form. Propellant-driven formulations may also contain other components, such as cosolvents, stabilizers and optional other excipients.

[0168] Propellant-free inhalable formulations containing compounds of the present invention may be solutions or suspensions in water, alcohol, or water / alcohol media and may be delivered by jet nebulizers or ultrasonic nebulizers known from the prior art or by soft mist nebulizers.

[0169] Compounds of the present invention can be administered as the sole active agent or in combination with other pharmaceutical active ingredients.

[0170] The dosage of compounds of the present invention depends on various factors, including, in particular, the specific disease to be treated, the severity of symptoms, the route of administration, etc.

[0171] In another aspect, the present invention relates to the use of compounds of formula (I) in the preparation of pharmaceuticals. In another aspect, the present invention relates to compounds of formula (I) as pharmaceuticals.

[0172] Thus, the present invention relates to compounds of formula (I) in the preparation of pharmaceuticals, preferably for the treatment of disorders related to voltage-gated sodium channel (NaV) receptor mechanisms.

[0173] In yet another embodiment, the present invention relates to compounds of formula (I) for the prevention and / or treatment of diseases, disorders, or conditions related to voltage-gated sodium channel (NaV) dysregulation.

[0174] In one aspect, the invention also relates to a method for preventing and / or treating disorders related to the NaV receptor mechanism, the method comprising administering a therapeutically effective amount of a compound of formula (I) to a patient requiring treatment.

[0175] Preferably, the compounds of the invention are used for treating and / or preventing respiratory diseases.

[0176] In a preferred embodiment, the invention provides a method for preventing and / or treating respiratory diseases, the method comprising administering a compound of formula (I).

[0177] In yet another aspect, the invention relates to pharmaceutical compositions comprising a compound of formula (I) and one or more pharmaceutically acceptable carriers and / or excipients for the prevention and / or treatment of respiratory diseases. Specification 26 / 106 pages 34 CN 122180674 A

[0178] In another embodiment, the present invention provides a method for preventing and / or treating respiratory diseases, the method comprising administering a pharmaceutical composition comprising a compound of formula (I).

[0179] In yet another aspect, the respiratory diseases mentioned above are selected from cough, subacute or chronic cough, treatment-resistant cough, idiopathic chronic cough, refractory chronic cough, post-viral cough, iatrogenic cough, asthma, IPF, COPD, and cough associated with respiratory diseases such as COPD, asthma, and bronchospasm.

[0180] In one embodiment, the present invention relates to the use of a compound of formula (I) or a pharmaceutical composition thereof for the preparation of a medicament for treating and / or preventing cough, subacute or chronic cough, treatment-resistant cough, idiopathic chronic cough, refractory chronic cough, post-viral cough, iatrogenic cough, asthma, IPF, COPD, and cough associated with respiratory diseases such as COPD, asthma, and bronchospasm.

[0181] In another embodiment, the present invention relates to a compound or pharmaceutical composition of formula (I) for the prevention and / or treatment of respiratory diseases, wherein the respiratory diseases are selected from: cough, subacute or chronic cough, treatment-resistant cough, idiopathic chronic cough, refractory chronic cough, post-viral cough, iatrogenic cough, asthma, IPF, COPD, and cough associated with respiratory diseases such as COPD, asthma, and bronchospasm.

[0182] In another embodiment, the present invention relates to a method for treating and / or preventing respiratory diseases, wherein the respiratory diseases are selected from cough, subacute or chronic cough, treatment-resistant cough, idiopathic chronic cough, refractory chronic cough, post-viral cough, iatrogenic cough, asthma, IPF, COPD, and cough associated with respiratory diseases such as COPD, asthma, and bronchospasm, the method comprising administering a compound of formula (I) or a pharmaceutical composition comprising a compound of formula (I) and one or more pharmaceutically acceptable carriers and / or excipients.

[0183] In a preferred embodiment, the respiratory disease mentioned above is chronic cough.

[0184] In yet another preferred embodiment, the present invention relates to the use of a compound of formula (I) or a pharmaceutical composition thereof for the prevention and / or treatment of chronic cough.

[0185] In yet another preferred embodiment, the present invention relates to the use of a compound of formula (I) or a pharmaceutical composition thereof for the preparation of a medicament for the treatment and / or prevention of chronic cough.

[0186] In yet another preferred embodiment, the present invention relates to a method for the treatment and / or prevention of chronic cough, the method comprising administering a compound of formula (I) or a pharmaceutical composition comprising a compound of formula (I) and one or more pharmaceutically acceptable carriers and / or excipients.

[0187] An effective dose of the compound of formula (I) can be delivered to mammals, particularly humans, using any suitable route of administration.

[0188] The dose of the prophylactic or therapeutic dose of the compound of formula (I) will, of course, vary depending on the severity of the condition to be treated and the route of administration, and will generally be determined through clinical trials required in the pharmaceutical field. It will also vary depending on the individual patient's age, weight, and response.

[0189] All preferred groups or embodiments described above for the compound of formula (I) can also be combined with each other and applied with necessary modifications.

[0190] The compounds of the present invention can be prepared in a variety of ways known to those skilled in the art of organic synthesis. It should be understood that the functionalities present on the molecule should correspond to the transformations mentioned. Sometimes this will require modification of the order of synthetic steps to obtain the desired compound of the present invention. Although optimal reaction conditions may vary depending on the specific reactants or solvents used, these conditions can be readily determined by those skilled in the art through conventional optimization methods. Therefore, the processes described below should not be considered as limiting the scope of synthetic methods that can be used to prepare the compounds of the present invention. In some cases, according to general chemical principles, a generally known protecting group (PG) can be used to mask or protect sensitive or reactive parts when needed (Protective Specification 27 / 106 pages 35 CN 122180674 A group in organic syntheses, 3rd ed. TW Greene, PGM Wuts).

[0191] Compounds of formula (I), including all the compounds listed above, can generally be prepared according to the procedures described in Schemes 1 to 2 below, wherein at least one non-limiting synthetic route is provided for the preparation of the illustrative compounds (i.e., examples).

[0192] Scheme 1

[0193] Specification 28 / 106 pages 36 CN 122180674 A

[0194] In one embodiment of the invention, compound (I) can be prepared from compounds (VI) and (VIII) according to Scheme 1.

[0195] Compound (VI) can be prepared starting from compound (II). Compound (V) can be obtained from compound (II) via a two-step procedure: step a) involves an acylation reaction with a suitable haloacetyl halide (III), wherein X1 and X2 are halogens, preferably and independently chlorides or bromides. Step b) involves an alkylation reaction with a suitable amine NHR1R2 (IV). Compound (VI) can then be prepared from compound (V) via a two-step procedure: step c) involves an ester hydrolysis reaction mediated by a base such as NaOH or KOH or an acid such as HCl or H2SO4. The subsequent step d) involves an amide formation reaction with a suitable amine NHR5R6 (IX) in the presence of a suitable coupling agent such as chloro-N,N,N′,N′-tetramethylformamidinium hexafluorophosphate (TCFH). Compound (VIII)Compound (VII) can be prepared by acylation with a suitable haloacetyl halide (III), wherein X1 and X2 are halogens, preferably and independently chlorides or bromides. Compound (I) can then be obtained from compounds (VI) and (VIII) by a nitrogen quaternization reaction (Menshutkin reaction).

[0196] Scheme 2

[0197]

[0198] In another embodiment of the invention, compound (I) can be prepared according to Scheme 2.

[0199] Compound (X) can be prepared from compound (V) by a two-step procedure: step a) involves an ester hydrolysis reaction mediated by a base such as NaOH or KOH or an acid such as HCl or H2SO4, and step b) involves the introduction of a suitable carboxylic acid ester protecting group (=PG), such as tert-butyl. Compound (XI) can be prepared by reacting compound (X) and compound (VIII) via a nitrogen quaternization reaction (Menshutkin reaction). Compound (I) can then be obtained from compound (XI) via a two-step procedure: step c) involves a carboxylic acid protecting group removal reaction, and step d) involves an amide formation reaction with a suitable amine NHR5R6 (IX) in the presence of a suitable coupling agent such as TCFH.

[0200] Scheme 3 Specification 29 / 106 pages 37 CN 122180674 A

[0201]

[0202] In another embodiment of the invention, compound (I) can be prepared according to Scheme 3. Compound (XIV) can be prepared from compound (XII): an amide formation reaction with a suitable amine NHR5R6 (IX) in the presence of a suitable coupling agent such as TCFH. Specification 30 / 106 pages 38 CN 122180674 A. Compound (XVI) can be prepared from compound (XIV): an acylation reaction with a suitable haloacetyl halide (III), wherein X1 and X2 are halogens, preferably and independently chlorides or bromides. Compound (XV) can be prepared from compound (VIII): by alkylation with a suitable amine NHR1R2 (IV). Ultimately, compound (I) can be prepared from compounds (XV) and (XVI) by alkylation.

[0203] Accordingly, the present invention provides intermediate compounds (V), (VI), (VIII), (X), (XI), (XIV), (XV), and (XVI) as defined above, and their use in the preparation of compound (I).

[0204] In another aspect, the present invention provides the use of intermediate compounds selected from compounds (V), (VIII), (X), (XI), (XV), and (XVI) as defined above (particularly in schemes 1, 2, and 3) for the preparation of compound (I) as defined above.

[0205] In the case of separation of Formula (I) compounds by precipitation from the reaction mixture followed by grinding or crystallization, they are generally obtained as salts, wherein the anion is a chloride anion or a bromide anion, depending on the meaning of X2 in compounds (III), (VIII), and (XVI) used to form the quaternary ammonium salt (see schemes 1, 2, and 3). The salt form of Formula (I) compounds can be obtained as zwitterions by subsequent treatment under basic conditions (e.g., purification on an amino-functionalized silica gel chromatography). The zwitterion form of Formula (I) compounds can be treated with acids such as hydrochloric acid to obtain the corresponding salt form of Formula (I) compounds.

[0206] Illustrated preparation methods are provided in the experimental section below.

[0207] Preparation of intermediates and examples

[0208] The chemical names of the compounds were generated using the PerkinElmer ChemDraw® Pro app (v. 20.0.0.41.) structure-to-name tool or common chemical names. All reagents not described in the experimental section are commercially available or are known compounds or can be formed from known compounds by methods known to those skilled in the art.

[0209] In the following methods, some of the raw materials are identified by the number indicated in the "Intermediate" or "Example" step number. This is provided solely to assist chemists in the art.

[0210] In the context of references to the use of "similar" or "analogous" procedures, those skilled in the art will recognize that the above procedures may involve minor variations that will be recognized by those skilled in the art, such as reaction temperature, reagent / solvent volume, reaction time, post-treatment conditions, or chromatographic purification conditions. All final compounds are obtained as pharmaceutically acceptable salts unless otherwise stated.

[0211] All intermediates and examples reported were characterized by LC-MS and / or 1H-NMR analysis as described herein. Optimal proton frequencies and solvent conditions can vary and can be readily determined by those skilled in the art through conventional optimization procedures.

[0212] Abbreviations

[0213] ACN = Acetonitrile; AMU = Atomic mass unit; CDCl3 = Deuterated chloroform; CyHex = Cyclohexane; DCM = Dichloromethane; DMF = Dimethylformamide; DMSO = Dimethyl sulfoxide; DMSO-d6 = Deuterated dimethyl sulfoxide; EGTA = Ethyl glycol-di(β-aminoethyl ether)-N,N,N′,N′-tetraacetic acid; Et2O = Diethyl ether; EtOAc = Ethyl acetate; eq. = Equivalent; FCC = Rapid column chromatography; h = Hour; HATU = 1-[Di(dimethylamino)methylene]-1H-1,2,3-triazolo[4,5-b]pyridinium 3-oxide hexafluorophosphate; HCOOH = Formic acid; HEPES = N-2-hydroxyethylpiperazine-N′-2-ethanesulfonic acid; HPLC = High performance liquid chromatography; MeOH=Methanol; min =minutes; NMR =nuclear magnetic resonance; RT / rt =room temperature; tR =retention time; ss = saturated solution; TCFH =chloro-N,N,N′,N′-tetramethylformamidinium hexafluorophosphate; TFA = trifluoroacetic acid; THF = tetrahydrofuran; LC-MS = liquid chromatography / mass spectrometry; XPhos = 2-dicyclohexylphosphino-2′,4′,6′-triisopropylbiphenyl; XPhos Pd G3 = (2-dicyclohexylphosphino-2′,4′,6′-triisopropyl-1,1′-biphenyl)[2-(2′-amino-1,1′-biphenyl)]palladium(II)methanesulfonate. Instruction manual 31 / 106 pages 39 CN 122180674 A

[0214] Analytical methods

[0215] NMR characterization:

[0216] 1H-NMR spectra were performed as follows: on a 400 Agilent INOVA, 500 Agilent VNMRS, or 600 Agilent INOVA spectrometer, at 400, 500, and 600 MHz (proton frequency), or on a Bruker Avance III 400 spectrometer.

[0217] Chemical shifts are reported as δ values ​​in ppm relative to the internal standard tetramethylsilane (TMS). Coupling constants (J values) are provided in Hertz (Hz), and multiplicity is reported using the following abbreviations (s = singlet, d = doublet, t = triplet, quin = quintet, m = multiplet, br = broad).

[0218] Information on the zwitterionic or salt forms of compounds of formula (I) is obtained by 1H-NMR spectroscopy: zwitterionic forms generally show a missing amide NH signal. HBr or HCl salts generally show the full amide NH signal. Formate (i.e., formate salts) are examined and quantified by the 1H-NMR signal of formic acid.

[0219] LC-MS:

[0220] LCMS can be recorded under the following conditions: on a Waters LC / PDA / MS Acquity™ system coupled to a Micromass ZQ™ or Waters SQD single quadrupole mass spectrometer, in positive and / or negative electron spray ES ionization mode, diode array DAD chromatograms, mass spectrometry chromatograms, and mass spectra can be acquired.

[0221] LC / UV / MS characterization:

[0222] It is estimated that the LC / MS retention time (tR) is affected by experimental error by ±0.5 minutes.

[0223] LC-MS Method 1: Acquity CSH C18 2.1 x 50 mm 1.7 μm, column temperature 40℃; mobile phase solvent A was milliQ water + 0.1% HCOOH, mobile phase solvent B was ACN + 0.1% HCOOH. Flow rate was 1 mL / min.

[0224] Gradient table: t=0 min 97%A 3% B, t=1.5 min 0.1% A 99.9% B, t=1.9 min 0.1% A 99.9% B and t=2 min 97% A 3% B. The UV detection range is 210-350 nm and the ES+ / ES- range is 100 to 1000 AMU.

[0225] LC-MS method 2: Column: Kinetex EVO C18 2.1 x 50 mm 1.7 μm, column temperature is 40℃; mobile phase solvent A is 10 mM NH4HCO3 aqueous solution (adjusted to pH=10 with ammonia), mobile phase solvent B is ACN. The flow rate is 1 mL / min. The gradient table is 97% A 3% B at t=0 min, 0.1% A 99.9% B at t=1.5 min, 0.1% A 99.9% B at t=1.9 min, and 97% A 3% B at t=2 min. The UV detection range is 210–350 nm, and the ES+ / ES- range is 100–1200 AMU.

[0226] Purification Methods

[0227] Purification of some compounds was achieved by semi-preparative HPLC: using an MDAP Waters device with mass spectrometry detection (MS: ZQ2000) and DAD detection 220, equipped with a CSH C18 (30 x 100 mm, 5 µm) column and a 1 mL injection loop. The gradient was ACN / H2O + 0.1% HCOOH from 50% to 70% over 10 min (flow rate = 40.00 mL / min.), used as the mobile phase.

[0228] Other compounds were purified by FCC using a Biotage® column and the conditions specified in each example.

[0229] General Synthetic Procedure

[0230] Intermediate 1: Methyl 3-(2-bromoacetamido)-4-methylthiophene-2-carboxylic acid 32 / 106 pages 40 CN 122180674 A

[0231]

[0232] 2-bromoacetyl bromide (6.11 mL, 71.0 mmol) was added dropwise to a suspension of methyl 3-amino-4-methylthiophene-2-carboxylic acid (10.0 g, 58.4 mmol) in water (235 mL) at 0 °C, and a creamy white solid was observed to form. The reaction was warmed to RT and stirred overnight. The solid was filtered, washed with water and dried under high vacuum to give the title compound (Intermediate 1: 16.0 g, 55.0 mmol, 94% yield) as a creamy white powder.

[0233] LC-MS Method 1: tR = 0.81 min, MS (ESI) m / z = 292.0 / 294.0 [M+H]+

[0234] Intermediate 2: 2-bromo-N-(isoxazo-3-yl)acetamide

[0235]

[0236] To potassium carbonate (6.58 g, 47.6 g)A suspension of 2-bromoacetyl bromide (1.82 mL, 20.9 mmol) in DCM (73 mL) and isoxazol-3-amine (1.44 mL, 19.0 mmol) was added dropwise. After 3 hours, the reaction mixture was washed with water (2x) and then with a saturated aqueous solution of NaHCO3. The organic phase was separated, dried and concentrated under reduced pressure on a phase separator to provide a crude product, which was purified by FCC (KP Sfär silica, 10–80% EtOAc / cyclohexane) to provide the title compound (intermediate 2: 1.60 g, 7.80 mmol, 41% yield) as an orange solid.

[0237] LC-MS method 1: tR = 0.53 min, MS (ESI) m / z = 205.0 / 207.0 [M+H]+

[0238] Intermediate 3: (R)-2-bromo-N-(1-phenylethyl)acetamide

[0239]

[0240] Similar to intermediate 1, intermediate 3 was prepared as follows: starting from a suitable amine:

[0241] (R)-1-phenylethyl-1-amine: 1.00 g, 8.25 mmol (1.0 equivalent)

[0242] Amount / yield: 1.69 g, 7.00 mmol, 84% yield.

[0243] LC-MS Method 2: tR = 0.83 min, MS (ESI) m / z = 242.0 / 244.0 [M+H]+

[0244] Example 1: 2-(benzylamino)-N,N-dimethyl-N-(2-((4-methyl-2-(pyrrolidine-1-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)-2-oxoethyl-1-ammonium (zwitterion)

[0245] Step 1 - 3-(2-chloroacetamido)-4-methylthiophen-2-carboxylic acid methyl ester (intermediate 4) Specification 33 / 106 pages 41 CN 122180674 A

[0246]

[0247] 3-amino-4-methylthiophen-2-carboxylic acid methyl ester (5.00 g, 29.0 mmol) was added to water (117) at 0°C. 2-Chloroacetyl chloride (9.30 mL, 116 mmol) was added dropwise to the solution. The reaction was allowed to warm to RT overnight, and a white precipitate was observed to form. The precipitate was filtered and washed with water (3x) and pentane (2x) to provide the title compound (intermediate 4: 7.40 g, quantitative yield) as a creamy white powder.

[0248] LC-MS Method 1: tR = 0.82 min, MS (ESI) m / z = 248.0 [M+H]+

[0249] Step 2 - 3-(2-(dimethylamino)acetamido)-4-methylthiophene-2-carboxylic acid methyl ester (intermediate 5)

[0250]

[0251] Dimethylamine (2.0 M, in THF) (2.42 mL, 4.84 mmol) was added to a solution of methyl 3-(2-chloroacetamido)-4-methylthiophene-2-carboxylic acid intermediate 4 (1.00 g, 4.0 mmol) and sodium bicarbonate (0.68 g, 8.1 mmol) in a container (10 mL) under stirring at RT. The mixture was stirred overnight at RT. Water was added, and the reaction was extracted with EtOAc (3x). The collected organic layer was dried on Na2SO4, filtered, and concentrated to provide the title compound (intermediate 5: 0.91 g, 3.6 mmol, 88% yield), which is a yellow oil.

[0252] LC-MS Method 1: tR = 0.38 min, MS (ESI) m / z = 257.1 [M+H]+

[0253] Step 3 - 2-(dimethylamino)-N-(4-methyl-2-(pyrrolidine-1-carbonyl)thiophene-3-yl)acetamide (intermediate 6)

[0254]

[0255] Four balls were loaded into a 10 mL ball mill reactor with pyrrolidine (64 mL, 0.78 mmol), potassium tert-butoxide (39 mg, 0.35 mmol) and methyl 3-(2-(dimethylamino)acetamido)-4-methylthiophene-2-carboxylic acid (intermediate 5, 100 mg, 0.39 mmol). The reactor was sealed and shaken at 13 Hz for 4 hours. Then the reactor was opened and water and EtOAc were added, the reactor was sealed and shaken. The organic layer was separated and the aqueous layer was extracted with EtOAc (2x). The collected organic layer was dried on Na2SO4, filtered and concentrated to provide the title compound (intermediate 6: 104 mg, 0.35 mmol, 90% yield).

[0256] LC-MS Method 1: tR = 0.43 min, MS (ESI) m / z = 296.1 [M+H]+

[0257] Step 4 - 2-(benzylamino)-N,N-dimethyl-N-(2-((4-methyl-2-(pyrrolidine-1-carbonyl)thiophene-3-yl)amino)-2-oxoethyl)-2-oxoethyl-1-ammonium (zwitterion) (Example 1)

[0258] N-benzyl-2-chloroacetamide (46 mg, 0.25 mmol) and 2-(dimethylamino)-N-(4-methyl-2-(pyrrolidine-1-carbonyl)thiophene-3-yl)acetamide (intermediate 6, 70 mg, 0.23 mmol) were stirred and mixed with ACN (0.8 Potassium iodide (3.8 mg, 0.020 mmol) was added to the solution. The reaction was stirred overnight at 80 °C. After this time, the reaction was cooled to RT and concentrated under high vacuum. The residue was subjected to FCC.(Sfär amino, 0-5% MeOH / DCM) purified to provide the title compound (Example 1: 35 mg, 0.079 mmol, 32% yield), as a pale yellow powder.

[0259] LC-MS Method 2: tR = 0.98 min, MS (ESI) m / z = 443.2 [M]+

[0260] 1H NMR (400 MHz, CDCl3) δppm 9.78 (br s, 1 H), 7.21-7.38 (m, 5 H), 6.92 (s, 1 H), 4.71 (s, 2 H), 4.45 (s, 2 H), 4.26 (s, 2 H), 3.46 (s, 6 H), 3.13-3.80 (m, 4 H), 2.13 (s, 3 H), 1.85 (br s, 4 H).

[0261] Example 2: 2-(benzylamino)-N-(2-((2-(dimethylcarbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-N,N-dimethyl-2-oxoethyl-1-ammonium carboxylate

[0262] Step 1 - Sodium chloride of 3-(2-(dimethylamino)acetamido)-4-methylthiophen-2-carboxylic acid (intermediate 7)

[0263]

[0264] Water (0.4 mL) and 2 M aqueous solution of sodium hydroxide (0.42 mL, 0.84 mmol) were added to a THF (2 mL) solution of intermediate 5 (180 mg, 0.700 mmol), and the reaction was heated at 50 °C for 16 hours. The reaction was then cooled to 0 °C and neutralized by adding 2 M aqueous solution of HCl until pH = 7. Volatile substances were removed under reduced pressure, yielding the title compound (intermediate 7: 230 mg, 92% wt, quantitative yield) as an orange solid.

[0265] LC-MS Method 1: tR = 0.32 min, MS (ESI) m / z = 243.1 [M+H]+

[0266] Step 2 - 3-(2-(dimethylamino)acetamido)-N,N,4-trimethylthiophene-2-carboxamide (intermediate 8)

[0267]

[0268] HATU (315 mg, 0.83 mmol) was added to a DMF (2 mL) solution of intermediate 7 (230 mg, 92% wt, 0.70 mmol). After stirring for 5 min, a 2 M THF solution of dimethylamine (0.69 mL, 1.38 mmol) was added. The reaction was stirred overnight at RT. An aqueous solution of NaOH at pH = 9 was added, and the reaction was extracted with EtOAc (3x). The collected organic layer was dried on Na2SO4, filtered, and concentrated to provide a crude product, which was purified by FCC (Sfär amino, 1% MeOH / DCM) to provide the title compound.Product (Intermediate 8: 176 mg, 0.65 mmol, 93% yield).

[0269] LC-MS Method 1: tR = 0.36 min, MS (ESI) m / z = 270.1 [M+H]+

[0270] Step 3 - 2-(benzylamino)-N-(2-((2-(dimethylcarbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)-N,N-dimethyl-2-oxoethyl-1-ammonium carbamate (Example 2)

[0271] Similar to Example 1, Example 2 was prepared as follows: starting from a suitable amine and chloride. Instructions for Use, Page 35 / 106, CN 122180674 A

[0272] Intermediate 8: 176 mg, 0.65 mmol (1.0 equivalent)

[0273] N-Benzyl-2-chloroacetamide: 120 mg, 0.65 mmol (1.0 equivalent)

[0274] Purified by FCC (Sfär-C18, 10-30% ACN + 0.1% HCOOH / H2O + 0.1% HCOOH).

[0275] Amount / Yield: 9.9 mg, 0.021 mmol, 3% yield.

[0276] LC-MS method 2: tR = 0.87 min, MS (ESI) m / z = 417.5 [M]+

[0277] 1H NMR (600 MHz, CDCl3) δppm 11.92 (br s, 1 H), 9.13 (s, 1 H), 8.66 (br s, 1 H), 7.30–7.35 (m, 4 H), 7.26–7.29 (m, 1 H), 6.97 (s, 1 H), 4.69 (s, 2 H), 4.61 (s, 2 H), 4.45 (d, J=5.6 Hz, 2 H), 3.53 (s, 6 H), 2.69–3.34 (m, 6 H), 2.22 (s, 3 H).

[0278] Example 3: 2-(benzylamino)-N-(2-((2-(tert-butylcarbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)-N,N-dimethyl-2-oxoethyl-1-ammonium carboxylate

[0279] Step 1 - N-(tert-butyl)-3-(2-(dimethylamino)acetamido)-4-methylthiophene-2-carboxamide (intermediate 9)

[0280]

[0281] Similar to intermediate 8, intermediate 9 was prepared as follows: starting from a suitable carboxylic acid and amine:

[0282] Intermediate 7: 150 mg, 0.62 mmol (1.0 equivalent)

[0283] 2-methyl-2-propylamine: 0.33 mL, 3.1 mmol (5 equivalent)

[0284] Yield / Quantity: 140 mg, 0.47 mmol, 76% yield.

[0285] LC-MS Method 1: tR = 0.45 min, MS (ESI) m / z = 298.1 [M+H]+

[0286] Step 2 - 2-(benzylamino)-N-(2-((2-(tert-butylcarbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)-N,N-dimethyl-2-oxoethyl-1-ammonium carbamate (Example 3)

[0287] N-benzyl-2-bromoacetamide (118 mg, 0.52 mmol) was added to a 1.7 mL solution of ACN containing intermediate 9 (140 mg, 0.47 mmol) under stirring. The reaction was stirred overnight at 80 °C. The residue was purified by a semi-preparative HPLC method to provide the title compound (Example 3: 10 mg, 0.021 mmol, 4% yield).

[0288] LC-MS method 2: tR = 0.66 min, MS (ESI) m / z = 445.2 [M]+

[0289] 1H NMR (400 MHz, CDCl3) δppm 10.37–12.99 (m, 1 H), 9.24 (br s, 1 H), 8.72 (s, 1 H), 7.23–7.37 (m, 5 H), 6.92 (s, 1 H), 5.89 (s, 1 H), 4.79 (s, 2 H), 4.61 (s, 2 H), 4.47 (br d, J=5.3 Hz, 2 H), 3.57 (s, 6 H), 2.17 (s, 3 H), 1.34 (s, 9 H).

[0290] Example 4: 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(morpholino-4-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion)

[0291] Step 1 - 3-(2-(azacycloheptane-1-yl)acetamido)-4-methylthiophene-2-carboxylic acid methyl ester (intermediate 11) Specification 36 / 106 pages 44 CN 122180674 A

[0292]

[0293] Perpiperidine (0.99 mL, 8.81 mmol) and potassium carbonate (1.21 g, 8.81 mmol) were added to a 27 mL solution of intermediate 1 (2.0 g, 6.78 mmol) in ACN and the mixture was stirred at RT for 16 hours. The reaction was filtered to remove salt and concentrate the organic phase. The residue was dissolved in EtOAc and water. The phases were mixed in an extractor, the organic layer was separated, and the aqueous layer was extracted with EtOAc (2x). The collected organic layer was dried on Na2SO4, filtered, and concentrated to provide the title compound (intermediate).11: 1.77 g, 5.69 mmol, 84% yield), is an orange oily substance.

[0294] LC-MS Method 1: tR = 0.48 min, MS (ESI) m / z = 311.2 [M+H]+

[0295] Step 2 - 2-(azacycloheptane-1-yl)-N-(4-methyl-2-(morpholine-4-carbonyl)thiophene-3-yl)acetamide (intermediate 12)

[0296]

[0297] Similar to intermediate 6, intermediate 12 is prepared as follows: starting from suitable amines and esters:

[0298] Intermediate 11: 200 mg, 0.64 mmol (1.0 equivalent)

[0299] Morpholine: 0.11 mL, 1.29 mmol (2.0 equivalent)

[0300] Potassium tert-butoxide: 65 mg, 0.58 mmol (0.9 equivalent)

[0301] Amount / Yield: 182 mg, 0.5 mmol, 77% yield.

[0302] LC-MS Method 1: tR = 0.36 min, MS (ESI) m / z = 366.2 [M+H]+

[0303] Step 3 - 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(morpholin-4-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion) (Example 4)

[0304] 2-(azacycloheptane-1-yl)-N-(4-methyl-2-(morpholin-4-carbonyl)thiophen-3-yl)acetamide (intermediate 12, 60 mg, 0.16 mmol) was added to a stirred solution of N-benzyl-2-bromoacetamide (41 mg, 0.18 mmol) in ACN (0.9 mL). The reaction was stirred overnight at 80 °C. The reaction was then cooled and concentrated under high vacuum. The residue was purified by FCC (Sfär amino, 1–5% MeOH / DCM) to provide the title compound (Example 4: 42 mg, 0.082 mmol, 45% yield).

[0305] LC-MS Method 1: tR = 0.63 min, MS (ESI) m / z = 513.3 [M]+

[0306] 1H NMR (500 MHz, DMSO-d6) δppm 9.13–10.77 (m, 1 H), 7.19–7.38 (m, 5 H), 7.09 (br s, 1 H), 4.48 (s, 2 H), 4.35 (d, J=5.6 Hz, 2 H), 4.08 (br s, 2 H), 3.75–3.83 (m, 2 H), 3.72 (br d, J=6.0 Hz, 2 H), 3.40–3.57 (m, 8 H), 1.77–2.07(m, 7 H), 1.62 (br s, 4 H).

[0307] Example 5: 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(methylcarbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion)

[0308] Step 1 - 3-(2-(azacycloheptane-1-yl)acetamido)-4-methylthiophen-2-carboxylic acid sodium chloride (intermediate specification 37 / 106 pages 45 CN 122180674 A body 13)

[0309]

[0310] Similar to intermediate 7, intermediate 13 was prepared as follows: starting from the corresponding ester:

[0311] Intermediate 11: 5.0 g, 16.1 mmol (1.0 equivalent)

[0312] Sodium hydroxide (2 M, in water): 9.7 mL, 19.3 mmol (1.2 equivalents)

[0313] Amount / yield: 6.3 g, 90% wt, quantitative yield.

[0314] LC-MS method 1: tR = 0.43 min, MS (ESI) m / z = 297.2 [M+H]+

[0315] Step 2 - 3-(2-(azacycloheptane-1-yl)acetamido)-N,4-dimethylthiophene-2-carboxamide (intermediate 14)

[0316]

[0317] Similar to intermediate 8, intermediate 14 is prepared as follows: starting from the corresponding acid and amine.

[0318] Intermediate 13: 284 mg, 0.80 mmol (1.0 equivalent)

[0319] Methylamine 2 M, in THF: 0.80 mL, 1.60 mmol (2.0 equivalent)

[0320] Purified by FCC (Sfäramino, 20% EtOAc / DCM)

[0321] Volume / Yield: 212 mg, 0.69 mmol, 86% yield

[0322] LC-MS Method 1: tR = 0.38 min, MS (ESI) m / z = 310.2 [M+H]+

[0323] Step 3 - 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(methylcarbamoyl)) Thiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion) (Example 5)

[0324] Similar to Example 4, Example 5 was prepared as follows: starting from the corresponding bromide and amine.

[0325] Intermediate 14: 70 mg, 0.23 mmol (1.0 equivalent)

[0326] N-Benzyl-2-bromoacetamide: 52 mg, 0.23 mmol (1.0 equivalent)

[0327] Purified by FCC (Sfäramino, 10-20% EtOAc / DCM)

[0328] Yield / Quantity: 34 mg, 0.074 mmol, 32% yield

[0329] LC-MS Method 1: tR = 0.59 min, MS (ESI) m / z = 457.3 [M]+

[0330] 1H NMR (400 MHz, DMSO-d6) δppm 9.19 (t, J=5.7 Hz, 1 H), 8.99 (q, J=4.4 Hz, 1 H), 7.18–7.37 (m, 5 H), 6.97 (s, 1 H), 4.61 (s, 2 H), 4.34 (d, J=5.7 Hz, 2 H), 4.25 (s, 2 H), 3.68–3.88 (m, 4 H), 2.73 (d, J=4.6 Hz, 3 H), 1.80– 2.03 (m, 7 H), 1.63 (br s, 4 H).

[0331] Example 6: 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(pyrrolidine-1-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion)

[0332] Step 1 - 2-(azacycloheptane-1-yl)-N-(4-methyl-2-(pyrrolidine-1-carbonyl)thiophen-3-yl)acetyl Specification 38 / 106 pages 46 CN 122180674 A Amine (intermediate 15)

[0333]

[0334] Similar to intermediate 6, intermediate 15 is prepared as follows: starting from the corresponding amine and ester.

[0335] Intermediate 11: 200 mg, 0.63 mmol (1.0 equivalent)

[0336] Pyrrolidine: 0.10 mL, 1.26 mmol (2.0 equivalent)

[0337] Potassium tert-butoxide: 64 mg, 0.57 mmol (0.9 equivalent)

[0338] Yield / Quantity: 134 mg, 0.38 mmol, 61% yield

[0339] LC-MS Method 1: tR = 0.51 min, MS (ESI) m / z = 350.2 [M+H]+

[0340] Step 2 - 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(pyrrolidine-1-carbonyl)thiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion) (Example 6)

[0341] Similar to Example 4, Example 6 was prepared as follows: starting from the corresponding bromide and amine.

[0342] Intermediate 15: 70 mg, 0.20 mmol (1.0 equivalent)

[0343] N-benzyl-2-bromoacetamide: 58 mg, 0.22 mmol (1.0 equivalent)

[0344] by FCC(Sfär amino, 0-5% MeOH / DCM) Purification

[0345] Volume / Yield: 60 mg, 0.10 mmol, 52% yield

[0346] LC-MS Method 1: tR = 0.70 min, MS (ESI) m / z = 497.2 [M]+

[0347] 1H NMR (400 MHz, DMSO-d6) δppm 10.38 (br s, 1 H), 6.96-7.50 (m, 6 H), 4.44 (s, 2 H), 4.35 (d, J=5.5 Hz, 2 H), 4.05-4.66 (m, 2 H), 3.64-3.91 (m, 4 H), 3.34-3.56 (m, 4 H), 2.00 (br s, 3 H) δppm 1.91 (br s, 4 H), 1.75 (br s, 4 H), 1.62 (br s, 4 H).

[0348] Example 7: 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(pyrrolidine-1-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion)

[0349] Similar to Example 4, Example 7 was prepared as follows: starting from the corresponding bromide and amine.

[0350] Intermediate 15: 70 mg, 0.20 mmol (1.0 equivalent)

[0351] Intermediate 2: 58 mg, 0.26 mmol (1.3 equivalent)

[0352] Purified by FCC (Sfär amino, 0-5% MeOH / DCM)

[0353] Amount / Yield: 33 mg, 0.069 mmol, 35% yield. LC-MS Method 1: tR = 0.58 min, MS (ESI) m / z = 474.2 [M]+

[0355] H NMR (400 MHz, CDCl) δ ppm 8.20 (d, J = 1.4 Hz, 1 H), 6.96 (s, 1 H), 6.89 (d, J = 1.4 Hz, 1 H), 4.39 (br s, 4 H), 3.78 - 3.98 (m, 2 H), 3.58 - 3.75 (m, 4 H), 3.47 (br s, 2 H), 2.28 (s, 3 H), 1.98 (br s, 4 H), 1.86 - 1.94 (m, 4 H), 1.77 (br s, 4 H).

[0356] Example 8: 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(morpholine-4-carbonyl))(3-yl)thiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion)

[0357] Similar to Example 4, Example 8 was prepared as follows: starting from the corresponding bromide and amine. Specification 39 / 106 pages 47 CN 122180674 A

[0358] Intermediate 12: 60 mg, 0.16 mmol (1.0 equivalent)

[0359] Intermediate 2: 44 mg, 0.21 mmol (1.3 equivalent)

[0360] Purified by FCC (Sfäramino, 0-5% MeOH / DCM)

[0361] Amount / Yield: 11 mg, 0.022 mmol, 14% yield. LC-MS method 1: tR = 0.51 min, MS (ESI) m / z = 490.2 [M]+

[0363] 1H NMR (400 MHz, CDCl3) δ ppm 8.64 (br s, 1 H), 8.31 (s, 1 H), 6.99 (s, 1 H), 6.86 (s, 1 H), 4.98 (br s, 2 H), 4.70 (br s, 2 H), 3.95 - 4.26 (m, 4 H), 3.64 - 3.80 (m, 8 H), 2.23 (s, 3 H), 2.05 (br s, 4 H), 1.80 (br s, 4 H).

[0364] Example 9: 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(methylcarbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onthionate

[0365] Similar to Example 4, Example 9 was prepared as follows: starting from the corresponding bromide and amine.

[0366] Intermediate 14: 70 mg, 0.23 mmol (1.0 equivalent)

[0367] Intermediate 2: 60 mg, 0.29 mmol (1.3 equivalent)

[0368] Purified by FCC (Sfär-C18, 10%-25% ACN + 0.1% HCOOH / H2O + 0.1% HCOOH)

[0369] Amount / Yield: 14 mg, 0.029 mmol, 13% yield.

[0370] LC-MS Method 1: tR = 0.46 min, MS (ESI) m / z = 434.2 [M]+

[0371] 1H NMR (500 MHz, CDCl3) δppm 8.65 (s, 1 H), 8.25 (d, J = 1.1 Hz, 1 H), 7.96 (br s, 1 H), 6.96 (s, 1 H), 6.83 (s, 1 H), 4.51–4.97(m, 4 H), 3.91 (br s, 4 H), 2.83 (d, J=4.4 Hz, 3 H), 1.89-2.10 (m, 7 H), 1.65-1.85 (m, 4 H).

[0372] Example 10: 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-(dimethylcarbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion)

[0373] Step 1-3-(2-(azacycloheptane-1-yl)acetamido)-N,N,4-trimethylthiophene-2-carboxamide (intermediate 16)

[0374]

[0375] Similar to intermediate 8, intermediate 16 is prepared as follows: starting from the corresponding acid and amine.

[0376] Intermediate 13: 250 mg, 0.70 mmol (1.0 equivalent)

[0377] N-methylmethylamine 2 M, in THF: 0.70 mL, 1.41 mmol (2.0 equivalent)

[0378] Purified by FCC (Sfär amino, 20-40% EtOAc / cyclohexane)

[0379] Amount / Yield: 157 mg, 0.49 mmol, 69% yield.

[0380] LC-MS Method 1: tR = 0.44 min, MS (ESI) m / z = 324.2 [M+H]+

[0381] Step 2 - 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-(dimethylcarbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion) (Example 10)

[0382] Similar to Example 4, Example 10 was prepared as follows: starting from the corresponding bromide and amine.

[0383] Intermediate 16: 77 mg, 0.24 mmol (1.0 equivalent)

[0384] N-Benzyl-2-bromoacetamide: 54 mg, 0.24 mmol (1.0 equivalent) Product Specification 40 / 106 pages 48 CN 122180674 A

[0385] Purified by FCC (Sfäramino, 1-5% MeOH / DCM)

[0386] Yield / Quantity: 65 mg, 0.14 mmol, 58% yield

[0387] LC-MS Method 1: tR = 0.64 min, MS (ESI) m / z = 471.2 [M]+

[0388] 1H NMR (400 MHz, CDCl3) δppm 10.51 (br s, 1 H), 7.23-7.45 (m, 5 H), 6.90 (s, 1 H), 4.58 (br s, 2 H), 4.45 (s, 2 H), 4.23(br s, 2 H), 3.94–4.09 (m, 2 H), 3.60–3.74 (m, 2 H), 3.02 (br s, 6 H), 2.12 (s, 3 H), 1.64–2.05 (m, 8 H).

[0389] Example 11: 1-(2-((2-(dimethylcarbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)-1-(2-(isoxazol-3-ylamino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion)

[0390] Similar to Example 4, Example 11 was prepared as follows: starting from the corresponding bromide and amine.

[0391] Intermediate 16: 77 mg, 0.24 mmol (1.0 equivalent)

[0392] Intermediate 2: 59 mg, 0.29 mmol (1.2 equivalent)

[0393] Purified by FCC (Sfär amino, 1-5% MeOH / DCM)

[0394] Amount / Yield: 22 mg, 0.049 mmol, 20% yield.

[0395] LC-MS method 1: tR = 0.50 min, MS (ESI) m / z = 448.2 [M]+

[0396] 1H NMR (500 MHz, CDCl3) δppm 12.01 (br s, 1 H), 8.29 (s, 1 H), 6.97 (s, 1 H), 6.84 (s, 1 H), 4.70–5.06 (m, 4 H), 3.90–4.14 (m, 4 H), 2.88–3.25 (m, 6 H), 2.19 (s, 3 H), 2.01 (br d, J=1.9 Hz, 4 H), 1.77 (br s, 4 H).

[0397] Example 13: 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((2-((2-methoxyethyl)carbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)piperidine-1-onthionylcarboxylate

[0398] Step 1 4-methyl-3-(2-(piperidine-1-yl)acetamido)thiophen-2-carboxylic acid methyl ester (intermediate 17)

[0399]

[0400] Similar to intermediate 11, intermediate 17 is prepared as follows: starting from the corresponding bromide and amine.

[0401] Intermediate 1: 2.0 g, 6.85 mmol (1.0 equivalent)

[0402] Piperidine: 0.88 mL, 8.9 mmol (1.3 equivalent)

[0403] Potassium carbonate: 1.23 g, 8.9 mmol (1.3 equivalent)

[0404] Amount / Yield: 1.3 g, 4.5 mmol, 65% yield

[0405] LC-MS Method 1: tR =0.45 minutes, MS (ESI) m / z = 297.1 [M+H]+

[0406] Step 2 - N-(2-methoxyethyl)-4-methyl-3-(2-(piperidin-1-yl)acetamido)thiophene-2-carboxamide (intermediate 18) Specification 41 / 106 pages 49 CN 122180674 A

[0407]

[0408] Similar to intermediate 6, intermediate 18 is prepared as follows: starting from a suitable amine and ester.

[0409] Intermediate 17: 300 mg, 0.93 mmol (1.0 equivalent)

[0410] 2-Methoxyethylamine: 0.28 ml, 2.1 mmol (2.20 equivalent)

[0411] Potassium tert-butoxide: 102 mg, 0.91 mmol (0.98 equivalent)

[0412] Amount / Yield: 110 mg, 0.32 mmol, 35% yield.

[0413] LC-MS Method 1: tR = 0.39 min, MS (ESI) m / z = 340.2 [M+H]+

[0414] Step 3 - 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((2-((2-methoxyethyl)carbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)piperidine-1-onthionate (Example 13)

[0415] Similar to Example 4, Example 13 was prepared as follows: starting from the corresponding bromide and amine.

[0416] Intermediate 18: 55 mg, 0.16 mmol (1.0 equivalent)

[0417] Intermediate 2: 110 mg, 0.16 mmol (1.0 equivalent)

[0418] Purified by FCC (Sfär-C18, 5-25% ACN + 0.1% HCOOH / H2O + 0.1% HCOOH)

[0419] Amount / Yield: 6.2 mg, 0.08 mmol, 7.5% yield

[0420] LC-MS Method 1: tR = 0.47 min, MS (ESI) m / z = 464.2 [M]+

[0421] 1H NMR (500 MHz, CDCl3) δppm 8.72 (s, 1H), 8.27 (d, J = 1.6 Hz, 1H), 7.35 (br s, 1 H), 6.94 (s, 1 H), 6.86 (d, J=1 .6 Hz, 1 H), 4.88 (br s, 4 H), 3.90 - 4.08 (m, 4 H), 3.38 - 3.54 (m, 4 H), 3.28 (s, 3 H), 2.02 (s, 3 H), 1.91 - 2.08 (m, 4 H), 1.62 - 1.82 (m, 2 H).

[0422] Example 14: 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-(dimethylcarbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)piperidine-1-onium (zwitterion)

[0423] Step 1: Sodium chloride of 4-methyl-3-(2-(piperidine-1-yl)acetamido)thiophen-2-carboxylic acid (intermediate 19)

[0424]

[0425] Similar to intermediate 7, intermediate 19 is prepared as follows: starting from the corresponding ester.

[0426] Intermediate 17: 0.60 g, 2.0 mmol (1.0 equivalent)

[0427] Sodium hydroxide (2 M, in water): 1.2 mL, 2.4 mmol

[0428] Amount / Yield: 0.62 g, 1.83 mmol, 90% yield

[0429] LC-MS Method 1: tR = 0.39 min, MS (ESI) m / z = 283.0 [M+H]+

[0430] Step 2 - N,N,4-trimethyl-3-(2-(piperidin-1-yl)acetamido)thiophene-2-carboxamide (Intermediate 20) Specification 42 / 106 pages 50 CN 122180674 A

[0431]

[0432] Similar to intermediate 8, intermediate 20 is prepared as follows: starting from the corresponding acid and amine.

[0433] Intermediate 19: 400 mg, 1.42 mmol (1.0 equivalent)

[0434] N-methylmethylamine 2 M, in THF: 1.4 mL, 2.84 mmol (2.0 equivalent)

[0435] Purified by FCC (Sfäramino, 20-40% EtOAc / cyclohexane)

[0436] Volume / Yield: 100 mg, 0.32 mmol, 23% yield

[0437] LC-MS Method 1: tR = 0.41 min, MS (ESI) m / z = 310.1 [M+H]+

[0438] Step 3 - 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-(dimethylcarbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)piperidine-1-onium (zwitterion) (Example 14)

[0439] Similar to Example 4, Example 14 was prepared as follows: starting from the corresponding bromide and amine.

[0440] Intermediate 20: 60 mg, 0.19 mmol (1.0 equivalent)

[0441] N-Benzyl-2-bromoacetamide: 51 mg, 0.19 mmol (1.0 equivalent)

[0442] Purified by FCC (Sfäramino, 0-5% MeOH / DCM)

[0443] Amount / Yield: 34 mg, 0.074 mmol, 39% yield

[0444] LC‑MS Method 1: tR = 0.61 min, MS (ESI) m / z = 457.2 [M]+

[0445] 1H NMR (400 MHz, CDCl3) δppm 9.41 (br s, 1 H), 7.25 – 7.36 (m, 5 H), 6.94 (s, 1 H), 4.72 (s, 2 H), 4.52 (br d, J=5.2 Hz, 2 H), 4.46 (s, 2 H), 4.01 - 4.16 (m, 2 H), 3.80 (br d, J=12.5 Hz, 2 H), 2.98 (br s, 6 H), 2.18 (s, 3 H), 1.38 - 2.10 (m, 6 H).

[0446] Example 15: 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-((2-((tert-butoxycarbonyl)(methyl)amino)ethyl)carbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onthionyl carbamate

[0447] Step 1-(2-(3-(2-(azacycloheptane-1-yl)acetamido)-4-methylthiophene-2-carbamate)ethyl)(methyl)carbamate tert-butyl ester (intermediate 10)

[0448]

[0449] Similar to intermediate 6, intermediate 10 is prepared as follows: starting from a suitable amine and ester.

[0450] Intermediate 11: 600 mg, 1.72 mmol (1.0 equivalent)

[0451] N-(2-aminoethyl)-N-methylcarbamate tert-butyl ester: 600 mg, 3.44 mmol (2.0 equivalent)

[0452] Potassium tert-butoxide: 193 mg, 1.72 mmol (1.0 equivalent)

[0453] Purification by FCC (Sfäramino, 10-30% EtOAc / cyclohexane) Instructions 43 / 106 pages 51 CN 122180674 A

[0454] Amount / Yield: 110 mg, 0.32 mmol, 15% yield.

[0455] LC-MS Method 1: tR = 0.63 min, MS (ESI) m / z = 453.2 [M+H]+

[0456] Step 2 - 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-((2-((tert-butoxycarbonyl)(methyl)amino)ethyl)carbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onthionyl carbamate (Example 15)

[0457] Similar to Example 4, Example 15 was prepared as follows: starting from the corresponding bromide and amine.

[0458] Intermediate 10: 95 mg, 0.21 mmol (1.0 equivalent)

[0459] N-Benzyl-2-bromoacetamide: 61 mg, 0.23 mmol (1.0 equivalent)

[0460] Purified by FCC (Sfär-C18, 5-40% ACN + 0.1% HCOOH / H2O + 0.1% HCOOH). Yield: 29 mg, 0.045 mmol, 21% yield.

[0461] LC-MS Method 1: tR = 0.77 min, MS (ESI) m / z = 600.2 [M]+

[0462] 1H NMR (500 MHz, CDCl3) δppm 10.66–13.72 (m, 1 H), 9.32–10.46 (m, 1 H), 8.77 (s, 1 H), 7.37–7.59 (m, 1 H), 7.25–7.35 (m, 5 H), 7.02 (s, 1 H), 4.50–4.98 (m, 4 H), 4.38–4.49 (m, 2 H), 3.76–4.30 (m, 4 H), 3.26–3.49 (m, 4 H), 2.87 (s, 3 H) H), 2.08 (s, 3 H), 1.89-2.15 (m, 4 H), 1.72-1.87 (m, 4 H), 1.36 (br s, 9 H).

[0463] Example 16: 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-((2-methoxyethyl)carbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion)

[0464] Step 1-3-(2-(azacycloheptane-1-yl)acetamido)-N-(2-methoxyethyl)-4-methylthiophene-2-carboxamide (intermediate 21)

[0465]

[0466] Similar to intermediate 8, intermediate 21 is prepared as follows: starting from the corresponding acid and amine.

[0467] Intermediate 13: 350 mg, 0.99 mmol (1.0 equivalent)

[0468] 2-Methoxyethylamine: 133 mg, 1.78 mmol (1.8 equivalent)

[0469] Purified by FCC (Sfäramino, 10-40% EtOAc / cyclohexane)

[0470] Yield / Quantity: 200 mg, 0.57 mmol, 57% yield

[0471] LC-MS Method 1: tR = 0.43 min, MS (ESI) m / z = 354.2 [M+H]+

[0472] Step 2 - 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-((2-methoxyethyl)carbamoyl)carbamoyl)(4-methylthiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion) (Example 16)

[0473] Similar to Example 4, Example 16 was prepared as follows: starting from the corresponding bromide and amine.

[0474] Intermediate 21: 65 mg, 0.18 mmol (1.0 equivalent)

[0475] N-Benzyl-2-bromoacetamide: 45 mg, 0.20 mmol (1.1 equivalent)

[0476] Purified by FCC (Sfäramino, 1-5% MeOH / DCM)

[0477] Yield / Quantity: 65 mg, 0.13 mmol, 72% yield

[0478] LC-MS Method 1: tR = 0.61 min, MS (ESI) m / z = 501.2 [M] + Specification 44 / 106 pages 52 CN 122180674 A

[0479] 1H NMR (400 MHz, DMSO-d6) δppm 9.32 (br s, 1 H), 9.16 (br t, J=5.5 Hz, 1 H), 7.19 - 7.39 (m, 5 H), 6.99 (s, 1 H), 4.65 (s, 2 H), 4.33 (d, J=5.7 Hz, 2 H), 4.22 (s, 2 H), 3.78 (br dd, J=6.0, 3.2 Hz, 4 H), 3.34 - 3.43 (m, 4 H), 3.27 (s, 3 H), 1.93 (s, 3 H), 1.91 (br s, 4 H), 1.63 (br s, 4 H).

[0480] Example 17: 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-((2-methoxyethyl)carbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)piperidine-1-onium (zwitterion)

[0481] Similar to Example 4, Example 17 was prepared as follows: starting from the corresponding bromide and amine.

[0482] Intermediate 18: 55 mg, 0.16 mmol (1.0 equivalent)

[0483] N-Benzyl-2-bromoacetamide: 43 mg, 0.16 mmol (1.0 equivalent)

[0484] Purified by FCC (Sfäramino, 0-5% MeOH / DCM)

[0485] Yield / Quantity: 14 mg, 0.028 mmol, 18% yield

[0486] LC-MS Method 1: tR = 0.61 min, MS (ESI) m / z = 487.2 [M]+

[0487] 1H NMR (400 MHz, CDCl3) δppm 9.86 (br d, J = 0.8 Hz, 1 H), 8.87(br s, 1 H), 7.21 - 7.34 (m, 5H), 6.96 (s, 1 H), 4.38 (s, 2 H), 4.34 (br s, 2 H), 4.00 (s, 2 H), 3.60 - 3.73 (m, 2 H), 3.43 - 3.59 (m, 6 H), 3.34 (s, 3 H), 1.99 (s, 3 H), 1.91 - 2.02 (m, 4 H), 1.75 - 1.87 (m, 1 H), 1.65 - 1.73 (m, 1 H).

[0488] Example 18: 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-((2-hydroxyethyl)carbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion)

[0489] Step 1 - 3-(2-(azacycloheptane-1-yl)acetamido)-N-(2-((tert-butyldimethylsilyl)oxy)ethyl)-4-methylthiophene-2-carboxamide (intermediate 22)

[0490]

[0491] Similar to intermediate 8, intermediate 22 is prepared as follows: starting from the corresponding acid and amine.

[0492] Intermediate 13: 513 mg, 1.45 mmol (1.0 equivalent)

[0493] 2-Aminoethoxy(tert-butyl)dimethylsilane: 507 mg, 2.89 mmol (2.0 equivalent)

[0494] Purified by FCC (Sfäramino, 10-30% EtOAc / cyclohexane)

[0495] Yield / Quantity: 409 mg, 0.90 mmol, 62% yield

[0496] LC-MS Method 1: tR = 0.80 min, MS (ESI) m / z = 454.3 [M+H]+

[0497] Step 2 - 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-((2-((tert-butyldimethylsilyl)oxy)ethyl)carbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion) (intermediate 23) Specification 45 / 106 pages 53 CN 122180674 A

[0498]

[0499] Similar to intermediate 4, intermediate 23 is prepared as follows: starting from the corresponding bromide and amine.

[0500] Intermediate 22: 136 mg, 0.30 mmol (1.0 equivalent)

[0501] N-Benzyl-2-bromoacetamide: 75 mg, 0.33 mmol (1.1 equivalent)

[0502] Purified by FCC (Sfäramino, 1-5% MeOH / DCM)

[0503] Yield / Quantity: 71 mg, 0.12mmol, 39% yield.

[0504] LC-MS Method 1: tR = 0.94 min, MS (ESI) m / z = 601.3 [M]+

[0505] Step 3 - 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-((2-hydroxyethyl)carbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion) (Example 18)

[0506] 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-((2-hydroxyethyl)carbamoyl)oxy)carbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium bromide intermediate 23 (70 mg, 0.10 mg) A solution of 2 mL of DCM (1 mmol) was added dropwise to a 2 M Et2O solution of hydrogen chloride (0.5 mL, 1.0 mmol). After 3 hours, the evaporation was evaporated, and the residue was purified by column chromatography (Sfär amino, 0–10% MeOH / DCM) to provide the title compound (Example 18: 10 mg, 0.020 mmol, 19% yield) as a white powder. LC-MS Method 1: tR = 0.56 min, MS (ESI) m / z = 487.2 [M]+

[0508] 1H NMR (400 MHz, DMSO-d6) δppm 9.39 (br t, J=5.5 Hz, 1 H), 9.16 (t, J=5.6 Hz, 1 H), 7.20 - 7.36 (m, 5 H), 6.98 (d, J=0.9 Hz, 1 H), 4.82 (t, J=5.0 Hz, 1 H), 4.66 (s, 2 H), 4.34 (d, J=5.7 Hz, 2 H), 4.17 - 4.28 (m, 2 H), 3.76 (br s, 4 H), 3.48 (q, J= 5.5 Hz, 2 H), 3.23-3.34 (m, 2 H), 1.78-1.98 (m, 7 H), 1.63 (br d, J=3.3 Hz, 4 H).

[0509] Example 19: 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((4-methyl-2-((2-(methylamino)ethyl)carbamoyl)thiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium chloride hydrochloride

[0510] Step 1-1-(2-((2-((2-((tert-butoxycarbonyl)(methyl)amino)ethyl)carbamoyl)-4-methylThiophene-3-yl)amino)-2-oxoethyl)-1-(2-(isoxazol-3-ylamino)-2-oxoethyl)azacycloheptane-1-onthionate (intermediate 24) Specification 46 / 106 pages 54 CN 122180674 A

[0511]

[0512] Similar to intermediate 4, intermediate 24 is prepared as follows: starting from a suitable bromide and amine.

[0513] Intermediate 10: 95 mg, 0.21 mmol (1.0 equivalent)

[0514] Intermediate 2: 67 mg, 0.23 mmol (1.1 equivalent)

[0515] Purified by FCC (Sfär amino, 1-5% MeOH / DCM) and then by FCC (Sfär-C18, 5-30% ACN + 0.1% HCOOH / H2O + 0.1% HCOOH)

[0516] Amount / yield: 63 mg, 0.10 mmol, 49% yield.

[0517] LC-MS Method 1: tR = 0.66 min, MS (ESI) m / z = 577.3 [M]+

[0518] Step 2 - 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((4-methyl-2-((2-(methylamino)ethyl)carbamoyl)thiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium chloride hydrochloride (Example 19)

[0519] Similar to Example 18, Example 19 was prepared as follows: starting from a suitable Boc-protected amine. After removing volatiles, a pure product was obtained without FCC.

[0520] Intermediate 24: 61 mg, 0.10 mmol (1.0 equivalent)

[0521] Amount / Yield: 47 mg, 0.090 mmol, 86% yield

[0522] LC-MS Method 2: tR = 0.67 min, MS (ESI) m / z = 239.1 [(M+H) / 2]+

[0523] 1H NMR (500 MHz, DMSO-d6) δppm 11.82 (br s, 1 H), 10.99 (br s, 1 H), 8.89 (d, J=1.6 Hz, 1 H), 8.76 (br s, 2 H), 8.31 (t, J=5.6 Hz, 1 H), 7.41 (d, J=0.8 Hz, 1 H), 6.90 (d, J=1.6 Hz, 1 H), 4.80 (s, 2 H), 4.67 - 4.74 (m, 2 H), 3.77 - 4.03 (m, 4 H), 3.51 (q, J=5.8 Hz, 2 H), 3.01 - 3.14 (m, 2 H), 2.57 (br s, 32.02 (s, 3 H), 1.89-2.15 (m, 4 H), 1.66 (br s, 4 H).

[0524] Example 20: 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((2-((2-methoxyethyl)carbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onthionyl carbamate

[0525] Similar to Example 4, Example 20 was prepared as follows: starting from the corresponding bromide and amine.

[0526] Intermediate 21: 65 mg, 0.18 mmol (1.0 equivalent)

[0527] Intermediate 2: 59.24 mg, 0.200 mmol (1.1 equivalent)

[0528] Purified by FCC (Sfär-C18, 5-30% ACN + 0.1% HCOOH / H2O + 0.1% HCOOH) Instructions for Use 47 / 106 pages 55 CN 122180674 A

[0529] Amount / Yield: 4.4 mg, 0.0090 mmol, 5% yield)

[0530] LC-MS Method 1: tR = 0.49 min, MS (ESI) m / z = 478.2 [M]+

[0531] 1H NMR (400 MHz, CDCl3) δppm 8.70 (s, 1H), 8.29 (d, J=1.3 Hz, 1 H), 7.39 (br s, 1 H), 6.99 (s, 1 H), 6.86 (s, 1 H), 4.80 - 4.99 (m, 4 H), 3.87 - 4.06 (m, 4 H), 3.43 - 3.58 (m, 4 H), 3.30 (s, 3 H), 2.07 (s, 3 H), 1.91 - 2.15 (m, 4 H), 1.68 - 1.86 (m, 4 H).

[0532] Example 21: 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(oxetane-3-ylcarbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion)

[0533] Step 1 - 3-[[2-(azacycloheptane-1-yl)acetyl]amino]-4-methyl-N-(oxetane-3-yl)thiophen-2-carboxamide (intermediate 25)

[0534]

[0535] Similar to intermediate 8, intermediate 25 is prepared as follows: starting from a suitable acid and amine.

[0536] Intermediate 13: 400 mg, 1.13 mmol (1.0 equivalent)

[0537] 3-oxacyclobutanamine: 165 mg, 2.25 mmol (2.0 equivalent)

[0538] Purification by FCC (Sfäramino, 5-20% EtOAc / DCM)

[0539] Amount / Yield: 256 mg, 0.73 mmol, 65% yield

[0540] LC-MS Method 1: tR = 0.80 min, MS (ESI) m / z = 352.1 [M+H]+

[0541] Step 2 - 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(oxecyclobutane-3-ylcarbamoyl)thiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion) (Example 21) Similar to Example 4, Example 21 was prepared as follows: starting from a suitable bromide and amine.

[0542] Intermediate 25: 80 mg, 0.23 mmol (1.0 equivalent)

[0543] Intermediate 2: 64 mg, 0.27 mmol (1.2 equivalent)

[0544] Purified by FCC (Sfär amino, 1-5% MeOH / DCM)

[0545] Amount / Yield: 10 mg, 0.021 mmol, 9% yield

[0546] LC-MS Method 1: tR = 0.48 min, MS (ESI) m / z = 476.2 [M]+

[0547] 1H NMR (500 MHz, DMSO-d6) δppm 11.91 (br s, 1 H), 9.59 (br d, J=5.9 Hz, 1 H), 8.76 (d, J=1.5 Hz, 1 H), 7.09 (s, 1 H) H), 6.90 (s, 1 H), 4.83 - 4.94 (m, 1 H), 4.74 (t, J=6.9 Hz, 2 H), 4.69 (s, 2 H), 4.48 (t, J=6.4 Hz, 2 H), 4.38 (s, 2 H), 3.76 - 3.90 (m, 4 H), 1.96 (s, 3 H), 1.87 - 2.03 (m, 4 H), 1.64 (br s, 4 H).

[0548] Example 22: 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-((2-(methylamino)ethyl)carbamoyl)thiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onthionyl carbamate

[0549] Step 1 - N-[2-[[3-[[2-[1-[2-(benzylamino)-2-oxo-ethyl]azacycloheptane-1-onthion-1-yl]acetyl]amino]-4-methyl-thiophene-2-carbonyl]amino]ethyl]-N-methyl-carbamate tert-butyl ester (zwitterion) Specification 48 / 106 pages 56 CN 122180674 A (Intermediate 26)

[0550]

[0551] Similar to intermediate 4, intermediate 26 was prepared as follows: starting from a suitable bromide and amine.

[0552] Intermediate 10: 247 mg, 0.55 mmol (1.0 equivalent)

[0553] N-Benzyl-2-bromoacetamide: 125 mg, 0.55 mmol (1.0 equivalent)

[0554] Purified by FCC (Sfär amino, 0-10% MeOH / DCM)

[0555] Amount / Yield: 100 mg, 0.17 mmol, 30% yield.

[0556] LC-MS method 1: tR = 0.75 min, MS (ESI) m / z = 600.5 [M]+.

[0557] Step 2 - 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-((2-(methylamino)ethyl)carbamoyl)thiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onthionyl carboxylate (Example 22)

[0558] Similar to Example 18, Example 22 was prepared as follows: starting from a suitable Boc-protected amine.

[0559] Intermediate 26: 100 mg, 0.17 mmol (1.0 equivalent)

[0560] Amount / Yield: 66 mg, 0.12 mmol, 71% yield.

[0561] Purification by FCC (Sfär-C18, 5-50% ACN + 0.1% HCOOH / H2O + 0.1% HCOOH)

[0562] LC-MS method 2: tR = 0.98 min, MS (ESI) m / z = 500.4 [M]+

[0563] 1H NMR (400 MHz, DMSO-d6) δppm 9.26 (br s, 1 H), 8.46-8.64 (m, 1 H), 8.42 (s, 1 H), 7.24-7.36 (m, 5 H), 7.22 (br s, 1 H), 4.48-4.63 (m, 4 H), 4.29-4.40 (m, 2 H), 3.72-3.90 (m, 4 H) 3.23–3.35 (m, 2H), 2.63 (t, J = 6.2 Hz, 2H), 2.30 (s, 3H), 1.98 (s, 3H), 1.85–1.97 (m, 4H), 1.53–1.73 (m, 4H).

[0564] Example 23: 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(oxecyclobutane-3-ylcarbamoyl)thiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion)

[0565] Similar to Example 4, Example 23 was prepared as follows: starting from a suitable bromide and amine.

[0566] Intermediate 25: 80 mg, 0.22 mmol (1.0 equivalent)

[0567] N-Benzyl-2-bromoacetamide: 55 mg, 0.24 mmol (1.1 equivalent)

[0568] Purified by FCC (Sfäramino, 1-5% MeOH / DCM)

[0569] Yield / Quantity: 29 mg, 0.058 mmol, 26% yield

[0570] LC-MS Method 1: tR = 0.62 min, MS (ESI) m / z = 499.2 [M]+

[0571] 1H NMR (500 MHz, DMSO-d6) δppm 9.85 (br d, J=6.7 Hz, 1 H), 9.09 (t, J=5.7 Hz, 1 H), 7.18-7.36 (m, 5 H), 7.03 (d, J=0.8 Hz, 1 H), 4.90 (sxt, J=6.8 Hz, 1 H), 4.73 (t, J=6.9 Hz, 2 H), 4.64 (s, 2 H), 4.47 (t, J=6.4 Hz, 2 H), 4.28 - 4.37 (m, 4 H), 3.72 - 3.92 (m, 4 H), 1.94 (s, 3 H), 1.80 - 2.03 (m, 4 H), 1.62 (br s, 4 H).

[0572] Example 24: 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-((2-hydroxyethyl)carbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)piperidine-1-onthiocarbamate

[0573] Step 1 - N-(2-((tert-butyldimethylsilyl)oxy)ethyl)-4-methyl-3-(2-(piperidine-1-yl)acetamido)thiophen-2-carboxamide (intermediate 27)

[0574]

[0575] Similar to intermediate 8, intermediate 27 is prepared as follows: starting from a suitable carboxylic acid and amine.

[0576] Intermediate 19: 365 mg, 1.30 mmol (1.0 equivalent)

[0577] (2-aminoethoxy)(tert-butyl)dimethylsilane: 0.53 mL, 2.6 mmol (2.0 equivalent)

[0578] Purified by FCC (Sfäramino, 0-40% EtOAc / cyclohexane)

[0579] Volume / Yield: 260 mg, 0.59 mmol, 46% yield

[0580] LC-MS Method 1: tR = 0.82 min, MS (ESI) m / z = 440.2[M+H]+

[0581] Step 2 -1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-((2-hydroxyethyl)carbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)piperidine-1-onthionate (Example 24)

[0582] Similar to Example 4, Example 24 was prepared as follows: starting from a suitable bromide and amine.

[0583] Intermediate 27: 90 mg, 0.21 mmol (1.0 equivalent)

[0584] N-Benzyl-2-bromoacetamide: 48 mg, 0.21 mmol (1.0 equivalent)

[0585] Purified by FCC (Sfär-C18, 5-28% ACN + 0.1% HCOOH / H2O + 0.1% HCOOH)

[0586] Amount / Yield: 12 mg, 0.022 mmol, 11% yield

[0587] LC-MS Method 1: tR = 0.55 min, MS (ESI) m / z = 473.2 [M]+

[0588] 1H NMR (400 MHz, CDCl3) δppm 8.97-9.13 (m, 1H), 8.68 ppm (s, 1H), 7.60 (br s, 1 H), 7.27 - 7.44 (m, 5 H), 7.05 (s, 1 H), 4.63 (s, 3 H), 4.37 - 4.49 (m, 4 H), 3.82 - 4.23 (m, 4 H), 3.55 - 3.65 (m, 2 H), 3.32 - 3.43 (m, 2 H), 2.05 (s, 3 H), 1.52-2.03 (m, 6 H).

[0589] Example 26: 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-((2-(methylamino)ethyl)carbamoyl)thiophene-3-yl)amino)-2-oxoethyl)piperidine-1-onium chloride hydrochloride

[0590] Step 1 1-methyl(2-(4-methyl-3-(2-(piperidine-1-yl)acetamido)thiophene-2-carbamate)ethyl) tert-butyl carbamate (intermediate 28)

[0591] Specification 50 / 106 pages 58 CN 122180674 A

[0592] Similar to intermediate 8, intermediate 28 is prepared as follows: starting from the corresponding acid and amine.

[0593] Intermediate 19: 384 mg, 1.3 mmol (1.0 equivalent)

[0594] N-(2-aminoethyl)-N-methylcarbamate tert-butyl ester: 0.46 mL, 2.6 mmol (2.0 equivalent)

[0595] Passed by FCC (Sfäramino, 0-40%)EtOAc / cyclohexane) purification

[0596] Amount / yield: 250 mg, 0.57 mmol, 44% yield.

[0597] LC-MS method 1: tR = 0.60 min, MS (ESI) m / z = 439.2 [M+H]+

[0598] Step 2 - 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-((2-((tert-butoxycarbonyl)(methyl)amino)ethyl)carbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)piperidine-1-onium (zwitterion) (intermediate 29)

[0599]

[0600] Similar to intermediate 4, intermediate 29 is prepared as follows: starting from a suitable bromide and amine.

[0601] Intermediate 28: 100 mg, 0.23 mmol (1.0 equivalent)

[0602] N-Benzyl-2-bromoacetamide: 61 mg, 0.23 mmol (1.0 equivalent)

[0603] Purified by FCC (Sfäramino, 0-5% MeOH / DCM)

[0604] Yield / Quantity: 73 mg, 0.11 mmol, 48% yield

[0605] LC-MS Method 1: tR = 0.78 min, MS (ESI) m / z = 586.2 [M]+

[0606] Step 3 - 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-((2-(methylamino)ethyl)carbamoyl)thiophene-3-yl)amino)-2-oxoethyl)piperidine-1-onium chloride hydrochloride (Example 26)

[0607] Similar to Example 18, Example 26 was prepared as follows: starting from a suitable Boc-protected amine.

[0608] Intermediate 29: 73 mg, 0.11 mmol (1.0 equivalent)

[0609] Amount / Yield: 43 mg, 0 mm.19 mmol, 71% yield.

[0610] LC-MS Method 1: tR = 0.41 min, MS (ESI) m / z = 486.3 [M]+

[0611] 1H NMR (500 MHz, DMSO-d6) δppm 11.03 (s, 1 H), 9.35 (t, J=5.8 Hz, 1 H), 8.82–9.04 (m, 2 H), 8.22 (t, J=5.7 Hz, 1 H), 7.43 (d, J=0.8 Hz, 1 H), 7.31–7.36 (m, 2 H), 7.24–7.31 (m, 3 H), 4.90 (s, 2 H), 4.58 (s, 2 H), 4.36 (d, J=5.6 Hz, 2 H), 3.64–3.91 (m, 4H), 3.51(q, J=5.6 Hz, 2 H), 2.99-3.17 (m, 2 H), 2.55 (t, J=5.4 Hz, 3 H), 2.02 (s, 3 H), 1.81-2.09 (m, 4 H), 1.52-1.70 (m, 2 H).

[0612] Example 25: 1-(2-((2-((2-hydroxyethyl)carbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)-1-(2-(isoxazol-3-ylamino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion) Specification 51 / 106 pages 59 CN 122180674 A

[0613] Similar to Example 4, Example 25 was prepared as follows: starting from a suitable bromide and amine.

[0614] Intermediate 22: 136 mg, 0.30 mmol (1.0 equivalent)

[0615] Intermediate 2: 85 mg, 0.36 mmol (1.2 equivalent)

[0616] Purified by FCC (Sfär amino, 1-10% MeOH / DCM) followed by grinding with DCM

[0617] Amount / Yield: 14 mg, 0.030 mmol, 10% yield

[0618] LC-MS Method 1: tR = 0.44 min, MS (ESI) m / z = 464.2 [M]+

[0619] 1H NMR (500 MHz, DMSO-d6) δppm 11.88 (br s, 1 H), 9.05 (br s, 1 H), 8.76 (d, J=1.4 Hz, 1 H), 7.05 (s, 1 H) H), 6.90 (d, J=0.8 Hz, 1 H), 4.70 (s, 1 H), 4.82 (br s, 1 H), 4.32 (s, 2 H), 3.72-3.86 (m, 4 H), 3.47 (t, J=5.8 Hz, 2 H), 3.26 (q, J=5.6 Hz, 2 H), 1.93 (s, 7 H), 1.63 (br s, 4 H).

[0620] Example 28: 1-(2-((2-(dimethylcarbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)-1-(2-((4-methylisoxazol-3-yl)amino)-2-oxoethyl)piperidine-1-onium (zwitterion)

[0621] Step 1 2-bromo-N-(4-methylisoxazol-3-yl)acetamide (intermediate 30)

[0622]

[0623] 2-bromoacetyl bromide (0) was added dropwise to a suspension of potassium carbonate (1.34 g, 9.69 mmol) and 4-methylisoxazol-3-amine (0.56 g, 5.70 mmol) in ACN (20 mL).0.78 mL, 8.55 mmol). Water was added after 3 h, and the reaction was extracted with EtOAc (3x). The collected organic layer was dried on Na2SO4, filtered, and concentrated to provide a crude product, which was purified by FCC (KP Sfär silica, 0-60% EtOAc / cyclohexane) to provide the title compound (intermediate 30: 1.23 g, 94% wt).

[0624] LC-MS Method 1: tR = 0.54 min, MS (ESI) m / z = 219.0 / 221.0 [M+H]+

[0625] Step 2 - 1-(2-((2-(dimethylcarbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)-1-(2-((4-methylisoxazol-3-yl)amino)-2-oxoethyl)piperidine-1-onium (zwitterion) (Example 28)

[0626] Similar to Example 4, Example 28 was prepared as follows: starting from a suitable bromide and amine.

[0627] Intermediate 20: 26 mg, 0.084 mmol (1.0 equivalent)

[0628] Intermediate 30: 18.4 mg, 0.084 mmol (1.0 equivalent)

[0629] Purified by FCC (Sfär amino, 0-5% MeOH / DCM).

[0630] Amount / Yield: 4 mg, 0.009 mmol, 11% yield

[0631] LC-MS Method 1: tR = 0.51 min, MS (ESI) m / z = 448.2 [M]+

[0632] 1H NMR (400 MHz, CDCl3) δppm 8.69 (br s, 1 H), 8.10 (s, 1 H), 6.97 (s, 1 H), 5.07 (br s, 2 H), 4.75 (s, 2 H), 3.86–4.28 (m, 4 H), 3.08 (br s, 6 H), 2.19 (s, 3 H), 2.05 (s, 3 H), 1.92–2.04 (m, 4 H), 1.61–1.90 (m, 2 H).

[0633] Example 29: 1-(2-((2-((2-methoxyethyl)carbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)-1-(2-((4-methylisoxazol-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onthionyl carboxylate

[0634] Similar to Example 4, Example 29 was prepared as follows: starting from suitable bromides and amines.

[0635] Intermediate 21: 65 mg, 0.18 mmol (1.0 equivalent)

[0636] Intermediate 30: 47 mg, 0.20 mmol (1.2 equivalent)

[0637] Purified by a semi-preparative HPLC method. (Instructions for use, page 52 / 106, CN 122180674 A

[0638] ) Amount / Yield: 31 mg, 0.054 mmol, 29% yield. LC-MS method 1: tR = 0.51 min, MS (ESI) m / z = 492.3 [M]+

[0640] H NMR (500 MHz, CDCl) δ ppm 8.72 (s, 1 H), 8.10 (s, 1 H), 7.08 (br s, 1 H), 7.01 (s, 1 H), 5.01 (br s, 2 H), 4.77 (br s, 2 H), 3.88 - 4.21 (m, 4 H), 3.45 - 3.58 (m, 4 H), 3.33 (s, 3 H), 2.09 (s, 3 H), 2.06 (s, 3 H), 1.96 - 2.07 (m, 4 H), 1.78 (br s, 4 H).

[0641] Example 12: 1-(2-((4-methyl-2-(methylcarbamoyl)thiophen-3-yl)amino)-2-oxoethyl)-1-(2-((4-methylisoxazol-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion)

[0642] Similar to Example 4, Example 12 was prepared as follows: starting from a suitable bromide and amine.

[0643] Intermediate 14: 70 mg, 0.23 mmol (1.0 equivalent)

[0644] Intermediate 30: 50 mg, 0.23 mmol (1.0 equivalent)

[0645] Purified by FCC (Sfär amino, 1-5% MeOH / DCM) and then by FCC (Sfär-C18, 10-25% ACN + 0.1% HCOOH / H2O + 0.1% HCOOH)

[0646] Amount / yield: 2.7 mg, 0.006 mmol, 3% yield. LC-MS method 1: tR = 0.48 min, MS (ESI) m / z = 448.1 [M] + H), 5.04 (br s, 2 H), 4.82 (br s, 2 H), 3.86 - 4.06 (m, 4 H), 2.89 (d, J =4.7 Hz, 3 H), 2.09 (s, 6 H), 2.05 (br s, 4 H), 1.78 (br s, 4 H).

[0649] Example 39: 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((2-(morpholin-4-carbonyl)benzo[b]thiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium bromide

[0650] Step 1 - 3-(2-bromoacetamido)benzo[b]thiophene-2-carboxylic acid methyl ester (intermediate 31)

[0651]

[0652] Similar to intermediate 1, intermediate 31 is prepared as follows: starting from a suitable amine.

[0653] Methyl 3-aminobenzo[b]thiophene-2-carboxylate: 2.50 g, 12.0 mmol (1.0 equivalent)

[0654] Amount / Yield: 1.80 g, 5.50 mmol, 45% yield

[0655] LC-MS Method 1: tR = 1.00 min, MS (ESI) m / z = 328.0 / 330.1 [M+H]+

[0656] Step 2 - Methyl 3-(2-(azacycloheptane-1-yl)acetamido)benzo[b]thiophene-2-carboxylate (Intermediate 32)

[0657] Specification 53 / 106 pages 61 CN 122180674 A

[0658] Similar to intermediate 11, intermediate 32 is prepared as follows: starting from a suitable bromide.

[0659] Intermediate 31: 1.30 g, 3.96 mmol (1.0 equivalent)

[0660] Homopiperidine: 0.47 mL, 3.96 mmol (1.0 equivalent)

[0661] Amount / Yield: 1.37 g, 3.95 mmol, 99% yield.

[0662] LC-MS Method 1: tR = 0.61 min, MS (ESI) m / z = 347.1 [M+H]+

[0663] Step 3 - 3-(2-(azacycloheptane-1-yl)acetamido)benzo[b]thiophene-2-carboxylic acid (intermediate 33)

[0664]

[0665] Lithium hydroxide (86 mg, 3.59 mmol) was added to a suspension of methyl 3-(2-(azacycloheptane-1-yl)acetamido)benzo[b]thiophene-2-carboxylic acid (intermediate 32, 829 mg, 2.39 mmol) in water (2.7 mL) / MeOH (2.7 mL) / THF (2.7 mL), and the reaction was stirred at 45 °C for 6 h. The organic phase was evaporated under reduced pressure, and 2M HCl aqueous solution was added to the remaining aqueous phase until pH = 7. The aqueous phase was then extracted with DCM and DCM / MeOH (9 / 1). The collected organic layer was evaporated under reduced pressure, providing the title compound (33: 790 mg, 2.38 mmol, 99% yield).

[0666] LC-MS Method 1: tR = 0.57 min, MS (ESI) m / z = 333.3 [M+H]+

[0667] Step 4 – 2-(azacycloheptane-1-yl)-N-(2-(morpholine-4-carbonyl)benzo[b]thiophene-3-yl)acetamide (intermediate 34)

[0668]

[0669] At 0°C, thionyl chloride (320 mL, 4.51 mmol) was added to a DCM (6 mL) suspension of 3-(2-(azacycloheptane-1-yl)acetamido)benzo[b]thiophene-2-carboxylic acid (intermediate 33, 750 mg, 2.26 mmol) and the reaction mixture was stirred at RT for 2 hours. Morpholine (0.78 mL, 9.00 mmol) was added, and the reaction mixture was stirred at 35°C for 4 hours. Volatile substances were removed under reduced pressure. The crude product was purified by FCC (KP Sfär silica, 0–10% MeOH / DCM) to provide the title compound (intermediate 34: 95 mg, 0.24 mmol, 10% yield) as an orange solid.

[0670] LC-MS Method 1: tR = 0.57 min, MS (ESI) m / z = 402.3 [M+H]+

[0671] Step 5 - 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((2-(morpholino-4-carbonyl)benzo[b]thiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium bromide (Example 39)

[0672] 2-bromo-N-(isoxazo-3-yl)acetamide intermediate 2 (16 mg, 0.070 mmol) was added to a solution of intermediate 34 (30 mg, 0.070 mmol) in ACN (0.5 mL) and the reaction mixture was stirred overnight at 70 °C. The reaction was cooled to RT and the solid precipitate was allowed to settle. The liquid phase was removed using a Pasteur pipette, and the solid was ground with cyclohexane / EtOAc (7 / 3 v / v) and then with cold ACN to obtain the title compound (Example 39: 13 mg, 0.021 mmol, 28% yield). Instruction manual, pages 54 / 106, CN 122180674 A

[0673] LC-MS method 1: tR = 0.63 min, MS (ESI) m / z = 526.38 [M]+

[0674] 1H NMR (400 MHz, DMSO-d6) δppm 11.73 (br s, 1 H), 10.99 (br s, 1 H), 8.90 (d, J=1.5 Hz, 1 H), 8.03 (d, J=8.1 Hz, 1 H), 7.82 (d, J=7.8 Hz, 1 H), 7.47–7.54 (m, 1 H), 7.41–7.46 (m, 1 H), 6.94 (d, J=1.0 Hz, 1 H), 4.77 (s, 2 H), 4.67 (s, 2H), 3.78 - 4.03 (m, 4 H), 3.42 - 3.73 (m, 8 H), 1.85 - 2.11 (m, 4 H), 1.55 - 1.80 (m, 4 H).

[0675] Example 43: 1,1-Di(2-((2-(dimethylcarbamoyl)benzo[b]thiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion)

[0676] Step 1 - 3-aminobenzo[b]thiophene-2-carboxylic acid (intermediate 35)

[0677]

[0678] Lithium hydroxide (115 mg, 4.83 mmol) was added to a suspension of methyl 3-aminobenzo[b]thiophene-2-carboxylic acid (500 mg, 2.41 mmol) in isopropanol (20 mL) / water (4 mL), and the reaction mixture was stirred under reflux for 1.5 hours. It was then cooled to RT, diluted with water, and the volatile organic compounds were evaporated under reduced pressure. The remaining aqueous phase was neutralized with 3 M HCl aqueous solution until pH = 6, and a white precipitate was observed to form. The solid was filtered, washed with cold water (2x), and dried to give the title compound (intermediate 35: 460 mg, 2.38 mmol, 99% yield) as a creamy white solid.

[0679] LC-MS Method 1: tR = 0.83 min, MS (ESI) m / z = 194.2 [M+H]+

[0680] Step 2 - 3-amino-N,N-dimethylbenzo[b]thiophene-2-carboxamide (intermediate 36)

[0681]

[0682] Dimethylamine hydrochloride (155 mg, 1.90 mmol) was added to a DMF (9 mL) solution of 3-aminobenzo[b]thiophene-2-carboxylic acid (intermediate 35, 175 mg, 0.91 mmol), HATU (482 mg, 1.27 mmol), and diisopropylethylamine (160 mL, 0.91 mmol). The reaction mixture was stirred overnight at RT. Water was added, and the aqueous phase was extracted with EtOAc (3x). The collected organic layer was washed with a saturated aqueous solution of NaHCO3, a 2 M aqueous solution of potassium carbonate, and then brine. The organic phase was dried and evaporated under reduced pressure on a phase separator to obtain the title compound (intermediate 36, 170 mg, 0.77 mmol, 85% yield).

[0683] LC-MS method 1: tR = 0.93 min, MS (ESI) m / z = 221.0 [M+H]+

[0684] Step 3 - 3-(2-bromoacetamido)-N,N-dimethylbenzo[b]thiophene-2-carboxamide (intermediate 37) Specification 55 / 106 pages 63 CN 122180674 A

[0685]

[0686] Similar to intermediate 1, intermediate 37 was prepared as follows: starting from a suitable amine.

[0687] Intermediate 36: 170 mg, 0.77 mmol (1.0 equivalent)

[0688] Amount / Yield: 260 mg, 0.76 mmol, 99% yield.

[0689] LC-MS Method 1: tR = 0.79 min, MS (ESI) m / z = 341.1 / 343.2 [M+H]+

[0690] Step 4 - 3-(2-(azacycloheptan-1-yl)acetamido)-N,N-dimethylbenzo[b]thiophene-2-carboxamide (intermediate 38)

[0691]

[0692] Add azacycloheptanane (0.13 mL, 1.11 mmol) to a solution of 3-(2-bromoacetamido)-N,N-dimethylbenzo[b]thiophene-2-carboxamide (intermediate 37, 100 mg, 0.53 mmol) in ACN (4.2 mL), and stir the reaction mixture at RT for 1.5 h. Volatile substances and crude product were removed under reduced pressure and purified by FCC (KP Sfär silica, 0-50% EtOAc / cyclohexane) to provide the title compound (intermediate 38: 150 mg, 0.42 mmol, 79% yield).

[0693] LC-MS Method 1: tR = 0.55 min, MS (ESI) m / z = 360.3 [M+H]+

[0694] Step 5 - 1,1-di(2-((2-(dimethylcarbamoyl)benzo[b]thiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion) (Example 43)

[0695] Similar to Example 4, Example 43 was prepared as follows: starting from a suitable amine and bromide.

[0696] Intermediate 38: 35 mg, 0.10 mmol (1.0 equivalent)

[0697] Intermediate 37: 3-(2-bromoacetamido)-N,N-dimethylbenzo[b]thiophene-2-carboxamide: 33 mg, 0.10 mmol (1.0 equivalent)

[0698] Amount / Yield: 3.8 mg, 0.006 mmol, 6% yield.

[0699] Purified by FCC (Sfäramino, 0-5% MeOH / DCM).

[0700] LC-MS Method 1: tR = 0.80 min, MS (ESI) m / z = 620.2 [M]+

[0701] 1H NMR (500 MHz, DMSO-d6) δppm 13.87 (br s, 1 H), 7.88 (d, J=8.1 Hz, 2 H), 7.67 (br d, J=8.1 Hz, 2 H), 7.35 (t, J=7.3 Hz, 2 H), 7.20 (t, J=7.5 Hz, 2 H), 4.40 (br s, 4 H)H), 3.78 (br s, 4 H), 2.97 (br s, 12 H), 1.98 (br s, 4 H), 1.67 (br s, 4 H).

[0702] Example 68: 1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((4-methyl-2-(methylcarbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion) Specification 56 / 106 pages 64 CN 122180674 A

[0703] Similar to Example 4, Example 68 was prepared as follows: starting from a suitable bromide and amine.

[0704] Intermediate 14: 81 mg, 0.26 mmol (1.0 equivalent)

[0705] Intermediate 1: 76 mg, 0.26 mmol (1.0 equivalent)

[0706] Purified by FCC (Sfär amino, 1-5% MeOH / DCM)

[0707] Amount / Yield: 69 mg, 0.13 mmol, 50% yield.

[0708] LC-MS Method 1: tR = 0.61 min, MS (ESI) m / z = 521.2 [M]+

[0709] 1H NMR (500 MHz, DMSO-d6) δppm 11.24 (br s, 1H), 8.73 (br s, 1H), 7.53 (s, 1H), 7.04 (s, 1H), 4.68 (br s, 2H), 4.39 (s, 2H), 3.85–3.77 (m, 2H), 3.73 (s, 3H), 3.77–3.70 (m, 2H), 2.62 (d, J = 4.7 Hz, 3H), 2.01 (s, 3H), 1.96 (s, 3H), 2.00–1.90 (m, 4H), 1.69 - 1.58 (m, 2H)

[0710] Example 69: 1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((4-methyl-2-(methylcarbamoyl)thiophen-3-yl)amino)-2-oxoethyl)piperidine-1-onium (zwitterion)

[0711] Step 1 - 3-amino-N,4-dimethylthiophen-2-carboxamide (intermediate 55)

[0712]

[0713] Similar to intermediate 36, intermediate 55 is prepared as follows: starting from a suitable acid and amine.

[0714] 3-Amino-4-methylthiophene-2-carboxylic acid: 640 mg, 4.07 mmol (1.0 equivalent)

[0715] Methylamine hydrochloride: 550 mg, 8.14 mmol (2.0 equivalent)

[0716] FCC certified(Sfäramino, 20% EtOAc / cyclohexane) Purification

[0717] Amount / Yield: 259 mg, 1.5 mmol, 37% yield

[0718] LC-MS Method 1: tR = 0.57 min, MS (ESI) m / z = 171.3 [M+H]+

[0719] Step 2 - 3-(2-chloroacetamido)-N,4-dimethylthiophene-2-carboxamide (intermediate 56)

[0720]

[0721] 2-chloroacetyl chloride (0.097 mL, 1.22 mmol) was slowly added to a suspension of intermediate 55 (190 mg, 1.1 mmol) and TEA (0.155 mL, 0.89 mmol) in ACN (9 mL) at 0 °C. The reaction was stirred at RT overnight. The reaction mixture was then washed with a saturated aqueous solution of NaHCO3 and water. The organic phase was dried and concentrated under reduced pressure on a phase separator to provide a crude product, which was purified by FCC (Sfär silica, 20-50% EtOAc / cyclohexane) to provide the title compound (intermediate 56:200 mg, 0.81 mmol, 74% yield, specification 57 / 106 pages 65 CN 122180674 A).

[0722] LC-MS Method 1: tR = 0.58 min, MS (ESI) m / z = 247.4 [M+H]+

[0723] Step 3 - N,4-dimethyl-3-(2-(piperidin-1-yl)acetamido)thiophene-2-carboxamide (intermediate 57)

[0724]

[0725] Piperidine (0.028 mL, 0.28 mmol) and potassium carbonate (58 mg, 0.42 mmol) were added to a stirred solution of intermediate 56 (69 mg, 0.28 mmol) in ACN (1 mL), and the mixture was stirred at RT for 3 h. The reaction was quenched with a saturated aqueous solution of NaHCO3, the organic layer was separated, and the aqueous layer was extracted with DCM (3x). The collected organic layer was filtered through a phase separator and concentrated to provide the title compound (intermediate 57: 62 mg, 0.21 mmol, 75% yield).

[0726] LC-MS Method 2: tR = 0.73 min, MS (ESI) m / z = 296.1 [M+H]+

[0727] Step 4 - 1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((4-methyl-2-(methylcarbamoyl)thiophen-3-yl)amino)-2-oxoethyl)piperidine-1-onium (zwitterion) (Example 69)

[0728] Similar to Example 4, Example 69 was prepared as follows: starting from suitable bromides and amines.

[0729] Intermediate 57: 62 mg, 0.21mmol (1.0 equivalent)

[0730] Intermediate 1: 61.3 mg, 0.21 mmol (1.0 equivalent)

[0731] Purified by FCC (Sfär amino, 0-5% MeOH / DCM)

[0732] Amount / yield: 41 mg, 0.081 mmol, 38% yield.

[0733] ​​LC-MS Method 1: tR = 0.59 min, MS (ESI) m / z = 507.2 [M]+

[0734] 1H NMR (500 MHz, DMSO-d6) δppm 11.24 (br s, 1H), 8.73 (br s, 1H), 7.53 (s, 1H), 7.04 (s, 1H), 4.68 (br s, 2H), 4.39 (s, 2H), 3.85–3.77 (m, 2H), 3.73 (s, 3H), 3.77–3.70 (m, 2H), 2.62 (d, J = 4.7 Hz, 3H), 2.01 (s, 3H), 1.96 (s, 3H), 2.00–1.90 (m, 4H), 1.69 - 1.58 (m, 2H)

[0735] Example 73: 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-(dimethylcarbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-4,4-di(hydroxymethyl)piperidin-1-onthionyl carboxylate

[0736] Step 1 - N-benzyl-2-(4,4-di(hydroxymethyl)piperidin-1-yl)acetamide (intermediate 58)

[0737]

[0738] Similar to intermediate 11, intermediate 58 is prepared as follows: starting from a suitable bromide and amine. Instructions for Use, Page 58 / 106, CN 122180674 A

[0739] N-Benzyl-2-bromoacetamide: 150 mg, 0.66 mmol (1.0 equivalent)

[0740] Piperidine-4,4-imidedimethylethanol hydrochloride: 120 mg, 0.66 mmol (1.0 equivalent)

[0741] Potassium carbonate: 227 mg, 1.6 mmol (2.5 equivalent)

[0742] Yield / Quantity: 190 mg, 0.64 mmol, 98% yield

[0743] LC-MS Method 2: tR = 0.52 min, MS (ESI) m / z = 293.2 [M+H]+

[0744] Step 2 - 3-Amino-N,N,4-trimethylthiophene-2-carboxamide (Intermediate 59)

[0745]

[0746] Similar to intermediate 36, intermediate 59 is prepared as follows: starting from a suitable acid and amine.

[0747] 3-Amino-4-methylthiophene-2-carboxylic acid: 900mg, 5.72 mmol (1.0 equivalent)

[0748] Dimethylamine hydrochloride: 934 mg, 11.45 mmol (2.0 equivalent)

[0749] Purified by FCC (Sfär silica, 70% EtOAc / cyclohexane)

[0750] Yield / volume: 410 mg, 1.5 mmol, 39% yield

[0751] LC-MS method 1: tR = 0.68 min, MS (ESI) m / z = 185.0 [M+H]+

[0752] Step 3 - 3-(2-bromoacetamido)-N,N,4-trimethylthiophene-2-carboxamide (intermediate 60)

[0753]

[0754] Similar to intermediate 30, intermediate 60 is prepared as follows: starting from a suitable amine, at 0°C:

[0755] Intermediate 59: 190 mg, 1.03 mmol (1.0 equivalent)

[0756] Purified by FCC (Sfär silica, 90% EtOAc / cyclohexane)

[0757] Amount / yield: 187 mg, 0.61 mmol, 59% yield.

[0758] LC-MS Method 1: tR = 0.64 min, MS (ESI) m / z = 305.0 / 307.0 [M+H]+

[0759] Step 4 - 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-(dimethylcarbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)-4,4-di(hydroxymethyl)piperidine-1-onthionyl carbamate (Example 73)

[0760] Similar to Example 4, Example 73 was prepared as follows: starting from a suitable bromide and amine.

[0761] Intermediate 58: 180 mg, 0.61 mmol (1.0 equivalent)

[0762] Intermediate 60: 187 mg, 0.61 mmol (1.0 equivalent)

[0763] Purified by FCC (Sfär-C18, 10-30% ACN + 0.1% HCOOH / H2O + 0.1% HCOOH)

[0764] Crystallized by vapor diffusion into iPrOH via Et2O. Specification 59 / 106 pages 67 CN 122180674 A

[0765] Amount / Yield: 131 mg, 0.23 mmol, 38% yield.

[0766] LC-MS Method 1: tR = 0.59 min, MS (ESI) m / z = 517.2 [M]+

[0767] 1H NMR (400 MHz, DMSO-d6) δppm 10.98 (br s, 1H), 9.65–9.22 (m, 1H), 8.51 (s, 1H), 7.41–7.21(m, 6H), 5.25 - 4.56 (m, 4H), 4.54 (s, 2H), 4.35 (d, J = 5.7 Hz, 2H), 3.89 - 3.64 (m, 4H), 3.46 - 3.33 (m, 4H), 2.89 (s, 6H), 2.06 (s, 3H), 1.80 - 1.63 (m, 4H).

[0768] Example 74: (2-(4,4-dimethyl-1-(2-((4-methyl-2-(methylcarbamoyl)thiophen-3-yl)amino)-2-oxoethyl)piperidin-1-onthiol-1-yl)acetyl)(isoxazol-3-yl)amide formate

[0769] Step 1 - 2-chloro-N-(isoxazol-3-yl)acetamide (intermediate 53)

[0770]

[0771] 2-chloroacetyl chloride (0.40 mL, 5.0 mmol) was added to a DCM (34 mL) solution of isoxazol-3-amine (400 mg, 4.76 mmol) and triethylamine (0.73 mL, 5.23 mmol) under stirring at 0 °C. After 10 minutes, the reaction mixture was warmed to RT and stirred for 2 h. Water was then added, the organic phase was separated, and the aqueous phase was extracted with DCM (2x). The collected organic phase was dried and concentrated under reduced pressure on a phase separator. The crude product was purified by FCC (Sfär amino, 5-50% cyclohexane / EtOAc) to give the title compound (intermediate 53: 610 mg, 3.80 mmol, 80% yield) as a white powder.

[0772] LC-MS Method 2: tR = 0.50 min, MS (ESI) m / z = 161.2 [M+H]+

[0773] Step 2 - 2-(4,4-dimethylpiperidin-1-yl)-N-(isoxazo-3-yl)acetamide (intermediate 61)

[0774]

[0775] Similar to intermediate 57, intermediate 61 was prepared as follows: starting from a suitable amine and chloride.

[0776] Intermediate 53: 2.3 g, 12.9 mmol (1.0 equivalent)

[0777] 4,4-Dimethylpiperidine hydrochloride: 1.92 g, 12.9 mmol (1.0 equivalent)

[0778] Potassium carbonate: 4.46 g, 32.2 mmol (2.5 equivalent)

[0779] Purified by FCC (Sfäramino, 10% EtOAc / cyclohexane)

[0780] Amount / Yield: 2.88 g, 12.1 mmol, 94% yield.

[0781] LC-MS Method 1: tR = 0.39 min, MS (ESI) m / z = 238.2 [M+H]+

[0782] Step 3 - 3-(2-bromoacetamido)-N,4-dimethylthiophene-2-carboxamide (Intermediate 62)

[0783] Specification 60 / 106 pages 68 CN 122180674 A

[0784] Similar to intermediate 30, intermediate 62 was prepared as follows: starting from a suitable amine, at 0°C:

[0785] Intermediate 55: 265 mg, 1.55 mmol (1.0 equivalent)

[0786] Purified by FCC (Sfär silica, 50% EtOAc / cyclohexane)

[0787] Amount / yield: 340 mg, 1.34 mmol, 86% yield.

[0788] LC-MS Method 1: tR = 0.65 min, MS (ESI) m / z = 290.9 / 292.9 [M+H]+

[0789] Step 4 - (2-(4,4-dimethyl-1-(2-((4-methyl-2-(methylcarbamoyl)thiophen-3-yl)amino)-2-oxoethyl)piperidin-1-on-1-yl)acetyl)(isoxazol-3-yl)amide formate (Example 74)

[0790] Similar to Example 4, Example 74 was prepared as follows: starting from a suitable bromide and amine.

[0791] Intermediate 62: 100 mg, 0.34 mmol (1.0 equivalent)

[0792] Intermediate 61: 81 mg, 0.34 mmol (1.0 equivalent)

[0793] Semi-preparative purification by HPLC

[0794] Amount / yield: 26 mg, 0.053 mmol, 15% yield.

[0795] LC-MS Method 1: tR = 0.60 min, MS (ESI) m / z = 448.2 [M]+

[0796] 1H NMR (500 MHz, DMSO-d6) δppm 8.84 (d, J = 1.5 Hz, 1H), 8.35 (s, 1H), 8.17 (br q, J = 4.0 Hz, 1H), 7.29 (s, 1H), 6.91 (s, 1H), 4.85–4.64 (m, 4H), 3.89–3.73 (m, 4H), 2.67 (d, J = 4.5 Hz, 3H), 1.98 (s, 3H), 1.87–1.62 (m, 4H), 1.07 (d, J = 6.9 Hz, 3H) Hz, 6H)

[0797] Example 70: 1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((4-methyl-2-(oxetane-3-ylcarbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion)

[0798] Similar to Example 4, Example 70 was prepared as follows: starting from suitable bromides and amines.

[0799] Intermediate 25: 32mg, 0.09 mmol (1.0 equivalent)

[0800] Intermediate 1: 27 mg, 0.09 mmol (1.0 equivalent)

[0801] Purified by FCC (Sfär amino, 0-2% MeOH / DCM)

[0802] Amount / Yield: 13 mg, 0.02 mmol, 25% yield.

[0803] LC-MS Method 1: tR = 0.66 min, MS (ESI) m / z = 563.3 [M]+

[0804] 1H NMR (400 MHz, DMSO-d6) δppm 11.26 (s, 1H), 9.58 (s, 1H), 7.54–7.07 (s, 2H), 4.80 (m, 9H), 4.70–4.41 (m, 4H), 4.64–4.41 (m, 4H), 3.85 (s, 4H), 3.72 (s, 3H), 2.00–1.97 (m, 10H), 1.67 (s, 4H)

[0805] Example 55: 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-((2-hydroxyethyl)(methyl)carbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium chloride

[0806] Step 1 - 3-(2-(azacycloheptane-1-yl)acetamido)-N-(2-hydroxyethyl)-N,4-dimethylthiophen-2-carboxamide (intermediate 63)

[0807] Specification 61 / 106 pages 69 CN 122180674 A

[0808] Similar to intermediate 36, intermediate 63 is prepared as follows: starting from a suitable acid and amine.

[0809] Intermediate 13: 700 mg, 1.97 mmol (1.0 equivalent)

[0810] 2-(methylamino)ethanol-1-ol: 222 mg, 2.96 mmol (1.5 equivalent)

[0811] Purified by FCC (Sfär silica, 0-5% MeOH / DCM) followed by FCC (Sfär-C18, 5-40% ACN + 0.1% (33% aq NH4OH) / H2O + 0.1% (33% aq NH4OH)

[0812] Yield / volume: 336 mg, 0.95 mmol, 48% yield

[0813] LC-MS Method 1: tR = 0.40 min, MS (ESI) m / z = 354.2 [M+H]+

[0814] Step 2 - 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-((2-hydroxyethyl)(methyl)amino)(Formyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium chloride (Example 55)

[0815] N-benzyl-2-bromoacetamide (114 mg, 0.48 mmol) was added to a solution of intermediate 63 (168 mg, 0.48 mmol) in ACN (4.7 mL) under stirring. The mixture was stirred in a sealed vial at 80 °C for 18 hours. The reaction mixture was concentrated under reduced pressure, and the residue was purified by FCC (Sfäramino, 0-5% MeOH / DCM) to provide the desired product, which is zwitterionic. It was dissolved in DCM (2.0 mL) and HCl 2 MEt2O solution (4 mL) was added under stirring at RT. After 5 minutes, the volatiles were removed under reduced pressure to provide the title compound (Example 55: 101 mg, 0.19 mmol, 40% yield).

[0816] LC-MS Method 1: tR = 0.60 min, MS (ESI) m / z = 501.3 [M]+

[0817] 1H NMR (500 MHz, DMSO-d6) δppm 10.44 (br s, 1H), 9.12 (br t, J = 5.7 Hz, 1H), 7.38–7.23 (m, 6H), 4.80 (br s, 1H), 4.65 (s, 2H), 4.45 (s, 2H), 4.35 (d, J = 5.6 Hz, 2H), 3.86–3.71 (m, 4H), 3.53 (br s, 2H), 3.38 (br s, 2H), 2.94 (br s, 3H), 2.07 (s, 3H), 1.95 (br d, J = 4.3 Hz, 4H), 1.64 (br s, 4H)

[0818] Example 67: 1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-4,4-dimethyl-1-(2-((4-methyl-2-(methylcarbamoyl)thiophen-3-yl)amino)-2-oxoethyl)piperidine-1-onthionate

[0819] Step 1-3-(2-(4,4-dimethylpiperidine-1-yl)acetamido)-N,4-dimethylthiophen-2-carboxamide (intermediate 64)

[0820]

[0821] Similar to intermediate 57, intermediate 64 is prepared as follows: starting from a suitable amine and chloride.

[0822] Intermediate 56: 69 mg, 0.28 mmol (1.0 equivalent)

[0823] 4,4-Dimethylpiperidine hydrochloride: 42 mg, 0.28 mmol (1.0 equivalent)

[0824] Yield / Quantity: 86 mg, 0.27 mmol, 95% yield.

[0825] LC-MS Method 2: tR = 0.88 min, MS (ESI) m / z = 324.1 [M+H]+

[0826] Step 2 - 1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-4,4-dimethyl-1-(2-((4-methyl-2-(methylcarbamoyl)thiophen-3-yl)amino)-2-oxoethyl)piperidine-1-onthionic acid Specification 62 / 106 pages 70 CN 122180674 A Salt (Example 67)

[0827] Similar to Example 4, Example 67 was prepared as follows: starting from a suitable bromide and amine.

[0828] Intermediate 64: 86 mg, 0.27 mmol (1.0 equivalent)

[0829] Intermediate 1: 78 mg, 0.27 mmol (1.0 equivalent)

[0830] Purified by FCC (Sfär-C18, 5-20% ACN + 0.1% HCOOH / H2O + 0.1% HCOOH).

[0831] Amount / Yield: 71 mg, 0.12 mmol, 45% yield. LC-MS method 1: tR = 0.63 min, MS (ESI) m / z = 535.6 [M]+ H NMR (500 MHz, DMSO-d6) δ ppm 11.05 (br s, 1 H), 8.41 (s, 1 H), 8.20 (br s, 1 H), 7.58 (s, 1 H), 7.26 (br s, 1 H), 4.80 (br s, 2 H), 4.69 (br s, 2 H), 3.77-3.90 (m, 4 H), 3.74 (s, 3 H), 2.62 (d, J=4.5 Hz, 3 H), 2.05 (s, 3 H), 2.00 (s, 3 H), 1.78 (br s, 4 H), 1.09 (s, 6 H)

[0834] Example 75: 1-(2-((2-(dimethylcarbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)-4,4-di(hydroxymethyl)-1-(2-oxo-2-((1-phenylcyclopropyl)amino)ethyl)piperidine-1-onthionyl carboxylate

[0835] Step 1 2-bromo-N-(1-phenylcyclopropyl)acetamide (intermediate 65)

[0836]

[0837] 2-bromoacetyl bromide (0.13 mL, 1.47 mmol) was added to a solution of 1-phenylcyclopropyl-1-amine hydrochloride (250 mg, 1.47 mmol) and TEA (0.41 mL, 2.94 mmol) in DCM (15 mL) at 0 °C.After stirring at 0 °C for 1 hour, a saturated aqueous solution of NH4Cl was added to separate the organic phase, and the aqueous phase was extracted with DCM (2x). The collected organic phase was dried on a phase separator and concentrated under reduced pressure. The crude product was purified by FCC (Sfär silica, 40% EtOAc / cyclohexane) to provide the title compound (intermediate 65: 279 mg, 1.09 mmol, 74% yield) as a white solid.

[0838] LC-MS Method 2: tR = 0.64 min, MS (ESI) m / z = 253.1 / 255.1 [M+H]+

[0839] Step 2 - 3-(2-(4,4-di(hydroxymethyl)piperidin-1-yl)acetamido)-N,N,4-trimethylthiophene-2-carboxamide (Intermediate 66)

[0840]

[0841] Piperidin-4,4-imidedimethylethanol hydrochloride (30 mg, 0.16 mmol) and potassium carbonate (56 mg, 0.41 mmol) were added to a stirred solution of intermediate 60 (50 mg, 0.16 mmol) in ACN (4 mL), and the mixture was stirred at RT for 16 h. The reaction was filtered to remove the salt, and the organic phase was concentrated under reduced pressure to provide the title compound (intermediate 66: 55 mg, 0.15 mmol, 90% yield) as a white solid.

[0842] LC-MS Method 2: tR = 0.50 min, MS (ESI) m / z = 370.2 [M+H]+ Specification 63 / 106 pages 71 CN 122180674 A

[0843] Step 3 - 1-(2-((2-(dimethylcarbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)-4,4-di(hydroxymethyl)-1-(2-oxo-2-((1-phenylcyclopropyl)amino)ethyl)piperidine-1-onthionate (Example 75)

[0844] Similar to Example 4, Example 75 was prepared as follows: starting from a suitable bromide and amine.

[0845] Intermediate 66: 55 mg, 0.15 mmol (1.0 equivalent)

[0846] Intermediate 65: 38 mg, 0.15 mmol (1.0 equivalent)

[0847] Purified by FCC (Sfär-C18, 5-35% ACN + 0.1% HCOOH / H2O + 0.1% HCOOH).

[0848] Amount / Yield: 39 mg, 0.066 mmol, 44% yield.

[0849] LC-MS Method 1: tR = 0.60 min, MS (ESI) m / z = 543.5 [M]+

[0850] 1H NMR (400 MHz, DMSO-d6) δppm 11.00 (brs, 1H), 9.68 (br s, 1H), 8.51 (s, 1H), 7.32 (s, 1H), 7.28 - 7.21 (m, 2H), 7.20 - 7.11 (m, 3H), 4.70 (br s, 2H), 4.52 (s, 2H), 4.78 (br s, 2H), 3.73 (br t, J = 4.9 Hz, 4H), 3.45 - 3.36 (m, 4H), 2.90 (s, 6H), 2.06 (s, 3H), 1.82 - 1.61 (m, 4H), 1.30 - 1.09 (m, 4H)

[0851] Example 66: 1-(2-((2-(dimethylcarbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion)

[0852] Similar to Example 4, Example 66 was prepared as follows: starting from suitable bromides and amines.

[0853] Intermediate 11: 90 mg, 0.26 mmol (1.0 equivalent)

[0854] Intermediate 60: 55 mg, 0.26 mmol (1.0 equivalent)

[0855] Purified by FCC (Sfär amino, 0-2% MeOH / DCM)

[0856] Amount / Yield: 55 mg, 0.10 mmol, 39% yield. LC-MS method 1: tR = 0.65 min, MS (ESI) m / z = 535.2 [M]+

[0858] 1H NMR (400 MHz, DMSO-d6) δppm 13.99 (br s, 1 H), 7.29 (s, 1 H), 7.21 (s, 1 H), 4.61 (br s, 2 H), 4.10 (s, 2 H), 3.73 - 3.89 (m, 2 H), 3.65 (s, 3 H), 3.57 - 3.69 (m, 2 H), 2.91 (br s, 6 H), 1.94 (br s, 4 H), 1.87 - 1.90 (m, 6 H), 1.64 (br s, 4 H)

[0859] Example 64: 1-(2-((2-(di(2-hydroxyethyl)carbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium carboxylate

[0860] Step 1-3-(2-(azacycloheptane-1-yl)acetamido)-N,N-di(2-hydroxyethyl)-4-methylthiophen-2-carboxamide (intermediate 67)

[0861] Specification 64 / 106 pages 72 CN 122180674 A

[0862] 2,2'-azaalkylidene di(ethylene-1-ol) (0.41 mL, 0.42 mmol) was added to a DMF (1.4 mL) suspension of sodium chloride (intermediate 13, 100 mg, 0.28 mmol), HATU (129 mg, 0.34 mmol), and DIPEA (147 mL, 0.85 mmol), and the reaction mixture was stirred overnight at 80 °C. The mixture was diluted with EtOAc. The two phases were separated, and the organic layer was washed with a saturated aqueous solution of NaHCO3 followed by brine. The aqueous phase was back-extracted (2x) with a DCM:iPrOH 2:1 solution. The collected organic phase was dried on Na2SO4, filtered, and concentrated under reduced pressure. The residue was purified by FCC (Sfär-C18, 0-80% ACN / H2O) to provide the title compound (intermediate 67: 35 mg, 0.09 mmol, 32% yield).

[0863] LC-MS Method 1: tR = 0.37 min, MS (ESI) m / z = 384.2 [M+H]+

[0864] Step 2 - 1-(2-((2-(di(2-hydroxyethyl)carbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onthionyl carbamate (Example 64)

[0865] Similar to Example 4, Example 64 was prepared as follows: starting from a suitable bromide and amine.

[0866] Intermediate 67: 35 mg, 0.09 mmol (1.0 equivalent)

[0867] Intermediate 1: 27 mg, 0.09 mmol (1.0 equivalent)

[0868] Purified by FCC (Sfär-C18, 5-30% ACN + 0.1% HCOOH / H2O + 0.1% HCOOH).

[0869] Amount / Yield: 14.1 mg, 0.022 mmol, 24% yield.

[0870] LC-MS Method 1: tR = 0.59 min, MS (ESI) m / z = 595.2 [M]+

[0871] 1H NMR (500 MHz, DMSO-d6) δppm 9.79–11.60 (m, 2H), 8.45 (s, 1H), 7.55 (br s, 1H), 7.28 (s, 1H), 4.44–4.81 (m, 4H), 4.08–5.93 (m, 2H), 3.77–3.89(m, 4 H), 3.74 (s, 3 H), 3.48-3.62 (m, 4 H), 3.39-3.48 (m, 4 H), 2.00-2.07 (m, 6 H), 1.92-2.09 (m, 4 H), 1.67 (br s, 4 H).

[0872] Example 42: 1-(2-((2-(dimethylcarbamoyl)benzo[b]thiophene-3-yl)amino)-2-oxoethyl)-1-(2-(isoxazol-3-ylamino)-2-oxoethyl)azacycloheptane-1-onium bromide

[0873] Similar to Example 39, Example 42 was prepared as follows: starting from a suitable amine and bromide.

[0874] Intermediate 38: 50 mg, 0.14 mmol (1.0 equivalent)

[0875] Intermediate 2: 29 mg, 0.14 mmol (1.0 equivalent)

[0876] Precipitated from EtOAc and then ground with DCM:MeOH (9 / 1 v / v).

[0877] Amount / Yield: 31 mg, 0.054 mmol, 39% yield.

[0878] LC-MS Method 1: tR = 0.62 min, MS (ESI) m / z = 484.2 [M]+

[0879] 1H NMR (500 MHz, DMSO-d6) δppm 11.77 (s, 1 H), 10.86 (s, 1 H), 8.91 (d, J = 1.6 Hz, 1 H), 8.02 (d, J = 8.1 Hz, 1 H), 7.78 (d, J = 8.0 Hz, 1 H), 7.46–7.54 (m, 1 H), 7.38–7.45 (m, 1 H), 6.95 (d, J = 1.5 Hz, 1 H), 4.77 (s, 2 H), 4.67 (s, 2 H), 3.79–3.97 (m, 4 H), 2.85-3.03 (m, 6 H), 1.99 (br d, J=3.4 Hz, 4 H), 1.67 (br s, 4 H).

[0880] Intermediate 40: 3-(2-(azacycloheptane-1-yl)acetamido)-4-methylthiophene-2-carboxylic acid tert-butyl ester

[0881] Specification 65 / 106 pages 73 CN 122180674 A

[0882] The stirred suspension of sodium chloride of 3-(2-(azacycloheptane-1-yl)acetamido)-4-methylthiophene-2-carboxylic acid (intermediate 13, 6.30 g, 16.0 mmol) in toluene (113 mL) was refluxed for 15 minutes. 1,1-di-tert-butoxy-N,N-dimethylmethylamine (7.66 mL, 32.0 mmol) was added dropwise to the resulting solution.The reaction was stirred under reflux for 24 hours. The solvent was evaporated to produce a crude mixture, which was purified by FCC (Sfär amino, 0–20% DCM / EtOAc (1 / 1 v / v), in cyclohexane) to provide the title compound (intermediate 40: 3.15 g, 8.93 mmol, 56% yield).

[0883] LC-MS Method 1: tR = 0.65 min, MS (ESI) m / z = 353.2 [M+H]+

[0884] Intermediate 39: 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-(tert-butoxycarbonyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion)

[0885]

[0886] Intermediate 40 (2.10 g, 5.84 mmol) was added to a stirred solution of N-benzyl-2-bromoacetamide (1.33 g, 5.84 mmol) in ACN (30 mL). The reaction was stirred at 80 °C for 36 h. The reaction was cooled and concentrated under high vacuum. The residue was purified by FCC (Sfär amino, 0–5% MeOH / DCM) to provide the title compound (intermediate 39: 2.54 g, 5.08 mmol, 87% yield).

[0887] LC-MS Method 1: tR = 0.82 min, MS (ESI) m / z = 500.3 [M]+

[0888] Intermediate 41: 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-carboxy-4-methylthiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium chloride

[0889]

[0890] Complete dissolution was observed when HCl in 2 M Et2O solution (30 mL, 60 mmol) was added dropwise to the suspension of intermediate 39 (2.50 g, 5.00 mmol) in THF (20 mL) and DCM (30 mL) at RT. After 48 hours, the evaporation was evaporated under reduced pressure, providing a solid, which was then ground with Et2O to provide the title compound (intermediate 41: 1.92 g, 4.00 mmol, 80% yield), a white solid as per specification 66 / 106 pages 74 CN 122180674 A.

[0891] LC-MS Method 1: tR = 0.64 min, MS (ESI) m / z = 445.2 [M]+

[0892] Intermediate 42: 1-(2-((2-(tert-butoxycarbonyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)-1-(2-(isoxazol-3-ylamino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion)

[0893]

[0894] Intermediate 2 (789 mg, 3.85 g) was stirred.Intermediate 40 (1.40 g, 3.85 mmol) was added to a 20 mL solution of ACN. The reaction was stirred overnight at 80 °C. The reaction was cooled and concentrated under high vacuum. The residue was purified by FCC (Sfär amino, 0–5% MeOH / DCM) to give the title compound (Intermediate 42: 1.42 g, 2.98 mmol, 77% yield).

[0895] LC-MS Method 1: tR = 0.74 min, MS (ESI) m / z = 477.2 [M]+

[0896] Intermediate 43 - 1-(2-((2-carboxy-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-(isoxazo-3-ylamino)-2-oxoethyl)azacycloheptane-1-onium chloride

[0897]

[0898] 3-[[2-[1-[2-(isoxazo-3-ylamino)-2-oxoethyl]azacycloheptane-1-onium-1-yl]acetyl]amino]-4-methyl-thiophen-2-carboxylic acid tert-butyl ester; bromide (intermediate 42, 1.42 g, 2.98 mmol) in DCM (37 mL) solution was added dropwise with 2 M HCl. Et2O solution (37 mL, 74 mmol). The reaction was stirred at RT for 16 h, and then the evaporation was evaporated under reduced pressure to provide the title compound (intermediate 43: 1.15 g, 2.52 mmol, 86% yield).

[0899] LC-MS Method 1: tR = 0.51 min, MS (ESI) m / z = 421.2 [M]+

[0900] Example 27: 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(4-methylpiperazine-1-carbonyl)thiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium chloride hydrochloride

[0901] 1-methylimidazole (205 mg, 2.50 mmol) was added to a suspension of intermediate 41 (400 mg, 0.83 mmol) in ACN (2.6 ml), followed by the addition of TCFH (257 mg, 0.92 mmol) at 0 °C. After 1 hour, 1-methylpiperazine (125 mg, 1.25 mmol) was added, and the mixture was stirred at 70 °C for 14 hours. The evaporated volatiles and residues were purified by FCC (Sfär amino, 0-5% MeOH / DCM) followed by FCC (Sfär-C18, 10-30% ACN / H2O + 0.05% HCl 37%), as per specification page 67 / 106, 75 CN 122180674 A, which provides the title compound (Example 27: 32 mg, 0.053 mmol, 6% yield).

[0902] LC-MS Method 2: tR =1.16 min, MS (ESI) m / z = 526.5 [M]+

[0903] 1H NMR (400 MHz, DMSO‑d6) δppm 11.03 (br s, 1 H), 10.82 (s, 1 H), 9.21 (t, J =5.8 Hz, 1 H), 7.41 (s, 1 H), 7.23‑ 7.37 (m, 5 H), 4.74 (s, 2 H), 4.46 (s, 2 H), 4.36 (d, J=5.7 Hz, 2 H), 4.13 (br s, 2 H), 3.68 - 3.93 (m, 4 H), 3.21 - 3.51 (m, 4 H), 2.93 - 3.10 (m, 2 H), 2.74 (br s, 3 2.13 (s, 3 H), 1.85-2.09 (m, 4 H), 1.64 (br s, 4 H)

[0904] Similar to the description of Example 27, the following examples were prepared by starting from suitable intermediates and corresponding commercially available amines.

[0905] Specification 68 / 106 pages 76 CN 122180674 A

[0906] Specification 69 / 106 pages 77 CN 122180674 A

[0907] Specification 70 / 106 pages 78 CN 122180674 A

[0908]

[0909] Intermediate 44: 1-(2-((2-((2-(4-(tert-butoxycarbonyl)piperazine-1-yl)ethyl)carbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)-1-(2-(isoxazol-3-ylamino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion)

[0910]

[0911] Similar to Example 27, intermediate 44 was prepared as follows: starting from intermediate 43.

[0912] Intermediate 43: 100 mg, 0.22 mmol (1.0 equivalent)

[0913] 4-(2-aminoethyl)piperazine-1-carboxylic acid tert-butyl ester: 50 mg, 0.22 mmol (1.0 equivalent)

[0914] Purified by FCC (Sfäramino, 0-5% MeOH / DCM).

[0915] Yield / Quantity: 75 mg, 0.12 mmol, 54% yield.

[0916] LC-MS Method 1: tR = 0.46 min, MS (ESI) m / z = 632.3 [M]+

[0917] Intermediate 45: 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-((2-(4-(tert-butyloxycarbonyl)piperazine)(1-(2-aminoethyl)piperazine-1-carbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion)

[0918] Specification 71 / 106 pages 79 CN 122180674 A

[0919] Similar to Example 27, intermediate 45 was prepared as follows: starting from a suitable carboxylic acid and amine.

[0920] Intermediate 41: 125 mg, 0.23 mmol (1.0 equivalent)

[0921] 4-(2-aminoethyl)piperazine-1-carboxylic acid tert-butyl ester: 50 mg, 0.22 mmol (1.0 equivalent)

[0922] Purified by FCC (Sfär amino, 0-5% MeOH / DCM).

[0923] Amount / Yield: 137 mg, 0.21 mmol, 95% yield

[0924] LC-MS Method 1: tR = 0.63 min, MS (ESI) m / z = 655.5 [M]+

[0925] Intermediate 68: methyl 3-(2-(4,4-dimethylpiperidin-1-yl)acetamido)-4-methylthiophene-2-carboxylic acid

[0926]

[0927] 4,4-dimethylpiperidin hydrochloride (1.00 g, 8.9 mmol) and potassium carbonate (2.46 g, 17.8 mmol) were added to a stirred solution of intermediate 1 (2 g, 6.85 mmol) in ACN (38 mL), and the mixture was stirred at RT for 2 h. The reaction was diluted with water and EtOAc, the organic layer was separated, and the aqueous layer was extracted with EtOAc (3x). The collected organic layer was dried on Na2SO4, filtered, concentrated, and yielded methyl 3-(2-(4,4-dimethylpiperidin-1-yl)acetamido)-4-methylthiophene-2-carboxylic acid (intermediate 68: 2.2 g, 6.78 mmol, 99% yield).

[0928] LC-MS Method 1: tR = 0.58 min, MS (ESI) m / z = 325.2 [M+H]+

[0929] Intermediate 69: 3-(2-(4,4-dimethylpiperidin-1-yl)acetamido)-4-methylthiophene-2-carboxylic acid sodium chloride

[0930]

[0931] Similar to intermediate 7, intermediate 69 was prepared as follows: starting from the corresponding ester:

[0932] Intermediate 68: 2.2 g, 6.3 mmol (1.0 equivalent)

[0933] Sodium hydroxide (2 M, in water): 3.8 mL, 7.6 mmol (1.2 equivalent)

[0934] Amount / yield: 2.4 g, quantitative yield.

[0935] LC-MS Method 1: tR = 0.52 minutes, MS (ESI) m / z = 311.2 [M+H]+

[0936] Intermediate 70: (2-(3-(2-(4,4-dimethylpiperidin-1-yl)acetamido)-4-methylthiophene-2-carbamate)ethyl)tert-butyl carbamate specification 72 / 106 pages 80 CN 122180674 A

[0937]

[0938] Similar to intermediate 36, intermediate 70 is prepared as follows: starting from a suitable acid and amine.

[0939] Intermediate 69: 0.60 g, 1.63 mmol (1.0 equivalent)

[0940] (2-aminoethyl) tert-butyl carbamate: 0.43 g, 2.44 mmol (1.5 equivalent)

[0941] Purified by FCC (Sfäramino, 50% EtOAc / cyclohexane)

[0942] Amount / Yield: 1.10 g, quantitative yield (impure compounds were used as is in subsequent steps)

[0943] LC-MS Method 1: tR = 0.67 min, MS (ESI) m / z = 467.3 [M+H]+

[0944] Intermediate 71: 1-(2-((2-((2-((tert-butoxycarbonyl)(methyl)amino)ethyl)carbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-4,4-dimethylpiperidine-1-onthionyl carbamate

[0945]

[0946] Similar to Example 4, intermediate 71 was prepared as follows: starting from the corresponding bromide and amine.

[0947] Intermediate 70: 130 mg, 0.28 mmol (1.0 equivalent)

[0948] Intermediate 1: 81 mg, 0.28 mmol (1.0 equivalent)

[0949] Purified by FCC (Sfär amino, 0-5% MeOH / DCM) followed by FCC (Sfär-C18, 5%-40% ACN + 0.1% HCOOH / H2O + 0.1% HCOOH)

[0950] Amount / Yield: 34 mg, 0.050 mmol, 18% yield

[0951] LC-MS Method 1: tR = 0.82 min, MS (ESI) m / z = 678.5 [M]+

[0952] Intermediate 72: 1-(2-((2-((2-((tert-butoxycarbonyl)(methyl)amino)ethyl)carbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onthionyl carbamate Specification 73 / 106 pages 81 CN 122180674 A

[0953]

[0954] Similar to Example 4, intermediate 72 was prepared as follows: starting from the corresponding bromide and amine.

[0955] Intermediate 10: 108 mg, 0.24 mmol (1.0 equivalent)

[0956] Intermediate 1: 88 mg, 0.29 mmol (1.2 equivalent)

[0957] Purified by FCC (Sfär amino, 0-2% MeOH / DCM) and then by FCC (Sfär-C18, 5%-50% ACN + 0.1% HCOOH / H2O + 0.1% HCOOH)

[0958] Amount / Yield: 80 mg, 0.113 mmol, 47% yield

[0959] LC-MS Method 1: tR = 0.78 min, MS (ESI) m / z = 664.4 [M]+

[0960] Intermediate 73: (2-(3-amino-4-methylthiophene-2-carbamoyl)ethyl)(cyclopropyl)carbamate tert-butyl ester

[0961]

[0962] Similar to intermediate 36, intermediate 73 is prepared as follows: starting from a suitable acid and amine.

[0963] (2-Aminoethyl)(cyclopropyl)carbamate tert-butyl ester: 0.51 mL, 2.67 mmol (1.0 equivalent)

[0964] 3-Amino-4-methylthiophene-2-carboxylic acid: 600 mg, 3.80 mmol (1.4 equivalent)

[0965] Purified by FCC (Sfär silica, 40% EtOAc / cyclohexane)

[0966] Yield / volume: 490 mg, 1.44 mmol, 54% yield

[0967] LC-MS method 1: tR = 1.02 min, MS (ESI) m / z = 340.5 [M+H]+

[0968] Intermediate 74: (2-(3-(2-(azacycloheptane-1-yl)acetamido)-4-methylthiophene-2-carbamate)ethyl)(cyclopropyl)tert-butyl carbamate

[0969] Specification 74 / 106 pages 82 CN 122180674 A

[0970] 2-bromoacetyl bromide (0.10 mL, 1.22 mmol) was added dropwise to a suspension of potassium carbonate (340 mg, 2.45 mmol) and (2-(3-amino-4-methylthiophene-2-carbamate)ethyl)(cyclopropyl)tert-butyl carbamate (intermediate 73, 490 mg, 1.22 mmol) in ACN (12 mL) at 0 °C. After stirring at RT for 1 hour, azacycloheptane (0.41 mL, 3.66 mmol) was added, and the reaction was stirred at RT for 4 hours. The solid and the filtrate were filtered off and concentrated under reduced pressure. The crude product was purified by FCC (KP Sfär silica, 5-40% EtOAc / cyclohexane) to provide the title compound (intermediate 74: 550).mg, 94% wt, quantitative yield).

[0971] LC-MS method 1: tR = 0.68 min, MS (ESI) m / z = 479.3 [M+H]+

[0972] Intermediate 75: 1-(2-((2-((2-((tert-butoxycarbonyl)(cyclopropyl)amino)ethyl)carbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion)

[0973]

[0974] Similar to Example 4, intermediate 75 was prepared as follows: starting from the corresponding bromide and amine.

[0975] Intermediate 74: 150 mg, 0.31 mmol (1.0 equivalent)

[0976] Intermediate 1: 110 mg, 0.29 mmol (1.0 equivalent)

[0977] Purified by FCC (Sfäramino, 0-10% MeOH / DCM)

[0978] Yield / Quantity: 70 mg, 0.091 mmol, 29% yield

[0979] LC-MS Method 1: tR = 0.82 min, MS (ESI) m / z = 694.4 [M]+

[0980] Intermediate 102: 4-((3-(2-(azacycloheptane-1-yl)acetamido)-4-methylthiophene-2-carboxamido)methyl)piperidine-1-carboxylic acid tert-butyl ester

[0981]

[0982] Similar to intermediate 36, intermediate 102 was prepared as follows: starting from a suitable acid and amine.

[0983] Intermediate 13: 500 mg, 1.41 mmol (1.0 equivalent) Specification 75 / 106 pages 83 CN 122180674 A

[0984] 4-(aminomethyl)piperidine-1-carboxylic acid tert-butyl ester: 0.450 mL, 2.11 mmol (1.5 equivalent)

[0985] Purified by FCC (Sfär amino, 20% EtOAc / cyclohexane)

[0986] Amount / Yield: 140 mg, 0.28 mmol, 20% yield

[0987] LC-MS Method 1: tR = 0.79 min, MS (ESI) m / z = 493.3 [M+H]+

[0988] Intermediate 76: 1-(2-((2-(((1-(tert-butoxycarbonyl)piperidin-4-yl)methyl)carbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion)

[0989]

[0990] Similar to Example 4, intermediate 76 was prepared as follows: starting from the corresponding bromide and amine.

[0991] Intermediate 102: 90 mg, 0.180 mmol (1.0 equivalent)

[0992] Intermediate 1: 53 mg, 0.180 mmol (1.0 equivalent)

[0993] Purified by FCC (Sfäramino, 0-2.5% MeOH / DCM)

[0994] Amount / Yield: 60 mg, 0.085 mmol, 47% yield

[0995] LC-MS Method 1: tR = 0.78 min, MS (ESI) m / z = 704.8 [M]+

[0996] Intermediate 77: 4-(3-(2-(azacycloheptane-1-yl)acetamido)-4-methylthiophene-2-carbonyl)piperazine-1-carboxylic acid tert-butyl ester

[0997]

[0998] Similar to intermediate 36, intermediate 77 was prepared as follows: starting from the corresponding acid and amine. The reaction was carried out at 60 °C.

[0999] Intermediate 13: 560 mg, 1.68 mmol (1.0 equivalent)

[1000] Piperazine-1-carboxylic acid tert-butyl ester: 470 mg, 2.52 mmol (1.5 equivalent)

[1001] Purified by FCC (Sfär silica, 30% EtOAc / cyclohexane) and then by FCC (Sfär-C18, 10-30% ACN / H2O)

[1002] Amount / yield: 84 mg, 0.18 mmol, 10% yield

[1003] LC-MS method 1: tR = 0.67 min, MS (ESI) m / z = 465.4 [M+H]+

[1004] Intermediate 78: 1-(2-((2-(4-(tert-butoxycarbonyl)piperazine-1-carbonyl)-4-methylthiophen-3-yl)amino)- 2-oxoethyl)-1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion)

[1005]

[1006] Similar to Example 4, intermediate 78 was prepared as follows: starting from the corresponding bromide and amine.

[1007] Intermediate 77: 760 mg, 1.64 mmol (1.0 equivalent)

[1008] Intermediate 1: 493 mg, 1.64 mmol (1.0 equivalent)

[1009] Purified by FCC (Sfär amino, 0–10% MeOH / DCM) followed by Sfär-C18 (5–100% ACN / H2O).

[1010] Amount / Yield: 107 mg, 0.14 mmol, 9% yield

[1011] LC-MS Method 1: tR = 0.82 min, MS (ESI) m / z = 676.3 [M]+

[1012] Intermediate 79: 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-((2-((tert-butoxycarbonyl)(methyl)amino)ethyl)(methyl)carbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion)

[1013]

[1014] Similar to Example 27, intermediate 79 was prepared as follows: starting from the corresponding acid and amine.

[1015] Intermediate 41: 200 mg, 0.42 mmol (1.0 equivalent)

[1016] Methyl (2-(methylamino)ethyl)carbamate tert-butyl ester: 118 mg, 0.63 mmol (1.5 equivalent)

[1017] Purified by FCC (Sfäramino, 0-5% MeOH / DCM)

[1018] Yield / Quantity: 77 mg, 0.12 mmol, 28% yield

[1019] LC-MS Method 1: tR = 0.81 min, MS (ESI) m / z = 614.4 [M]+

[1020] Intermediate 80: N2-(3-amino-4-methylthiophene-2-carbonyl)-N6-(tert-butoxycarbonyl)-D-lysine methyl ester Specification 77 / 106 pages 85 CN 122180674 A

[1021]

[1022] Similar to intermediate 36, intermediate 80 was prepared as follows: starting from a suitable acid and amine.

[1023] N6-(tert-butoxycarbonyl)-D-lysine methyl ester hydrochloride: 1.96 g, 6.62 mmol (1.3 equivalents)

[1024] 3-amino-4-methylthiophene-2-carboxylic acid: 800 mg, 5.09 mmol (1.0 equivalents)

[1025] Purified by FCC (Sfär silica, 20% (EtOAc / EtOH) / cyclohexane)

[1026] Amount / yield: 0.75 g, 1.88 mmol, 37% yield

[1027] LC-MS method 1: tR = 0.99 min, MS (ESI) m / z = 400.2 [M+H]+

[1028] Intermediate 81: N2-(3-(2-bromoacetamido)-4-methylthiophene-2-carbonyl)-N6-(tert-butoxycarbonyl)-D-lysine methyl ester

[1029]

[1030] 2-bromoacetyl bromide (0.164 mL, 1.88 mmol) was added dropwise to a suspension of intermediate 80 (0.75 g, 1.88 mmol) in ACN (17 mL) at 0 °C. A creamy white solid was observed to form. The reaction was stirred at RT for 1 h. The solid and the liquid phase were then filtered off by vacuum evaporation. The crude product was purified by FCC (Sfär silica, 10% EtOAc / cyclohexane) to provide the title compound (intermediate 81: 0.48 g).g, 0.92 mmol, 49% yield), is a colorless oil.

[1031] LC-MS method 1: tR = 1.00 min, MS (ESI) m / z = 520.2 / 522.2 [M+H]+

[1032] Intermediate 101: N2-(3-(2-(azacycloheptane-1-yl)acetamido)-4-methylthiophene-2-carbonyl)-N6-(tert-butoxycarbonyl)-D-lysine methyl ester Specification 78 / 106 pages 86 CN 122180674 A

[1033]

[1034] Similar to intermediate 66, intermediate 101 is prepared as follows: starting from a suitable bromide and amine.

[1035] Intermediate 81: 700 mg, 0.77 mmol (1.0 equivalent)

[1036] Azacycloheptanine: 0.17 mL, 1.53 mmol (2.0 equivalent)

[1037] Amount / yield: 170 mg, 0.32 mmol, 41% yield.

[1038] LC-MS Method 1: tR = 0.65 min, MS (ESI) m / z = 540.4 [M+H]+

[1039] Intermediate 82: (R)-1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-((6-((tert-butoxycarbonyl)amino)-1-methoxy-1-oxohexane-2-yl)carbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion)

[1040]

[1041] Similar to Example 4, intermediate 82 was prepared as follows: starting from the corresponding bromide and amine.

[1042] Intermediate 101: 170 mg, 0.32 mmol (1.0 equivalent)

[1043] N-Benzyl-2-bromoacetamide: 72 mg, 0.32 mmol (1.0 equivalent)

[1044] Purified by FCC (Sfäramino, 0-10% MeOH / DCM)

[1045] Amount / Yield: 85 mg, 0.11 mmol, 35% yield

[1046] LC-MS Method 1: tR = 0.78 min, MS (ESI) m / z = 686.8 [M]+

[1047] Intermediate 83: methyl 3-(2-(4-fluoro-4-methylpiperidin-1-yl)acetamido)-4-methylthiophene-2-carboxylate

[1048]

[1049] To the stirred intermediate 1 (570 mg, 1.95 mmol) in DCM (13 mL) of solution was added to 4-fluoro-4-methylpiperidine hydrochloride (300 mg, 1.95 mmol) and potassium carbonate (674 mg, 4.88 mmol), and the mixture was stirred at RT for 16 hours. (See instruction manual, pages 79 / 106, 87 CN)122180674 A The reaction was carried out to remove salts, and the organic phase was concentrated. The residue was dissolved in EtOAc and water. The phases were mixed in an extractor, the organic layer was separated, and the aqueous layer was extracted with EtOAc (2x). The collected organic layer was dried on Na2SO4, filtered, and concentrated to provide the title compound (intermediate 83: 600 mg, 1.83 mmol, 94% yield), as a yellow solid.

[1050] LC-MS Method 1: tR = 0.47 min, MS (ESI) m / z = 329.2 [M+H]+

[1051] Intermediate 84: 3-(2-(4-fluoro-4-methylpiperidin-1-yl)acetamido)-4-methylthiophene-2-carboxylic acid

[1052]

[1053] Similar to intermediate 7, intermediate 84 was prepared as follows: starting from the corresponding ester:

[1054] Intermediate 83: 600 mg, 1.83 mmol (1.0 equivalent)

[1055] Sodium hydroxide (2 M, in water): 1.1 mL, 2.19 mmol (1.2 equivalent)

[1056] Purified by FCC (Sfär-C18, 0-10% ACN + 0.1% HCOOH / H2O + 0.1% HCOOH)

[1057] Dosage / Yield: 150 mg, 0.48, 26% yield.

[1058] LC-MS Method 1: tR = 0.43 min, MS (ESI) m / z = 315.1 [M+H]+

[1059] Intermediate 85: (2-(3-(2-(4-fluoro-4-methylpiperidin-1-yl)acetamido)-4-methylthiophene-2-carbamate)ethyl)(methyl)carbamate tert-butyl ester

[1060]

[1061] Similar to Example 27, Intermediate 85 was prepared as follows: starting from a suitable carboxylic acid and amine:

[1062] Intermediate 84: 150 mg, 0.48 mmol (1.0 equivalent)

[1063] (2-aminoethyl)(methyl)carbamate tert-butyl ester: 83 mg, 0.48 mmol (1.0 equivalent)

[1064] Purified by FCC (Sfäramino, 0-60% EtOAc / cyclohexane)

[1065] Yield / Quantity: 100 mg, 0.21 mmol, 45% yield

[1066] LC-MS Method 1: tR = 0.62 min, MS (ESI) m / z = 471.4 [M+H]+

[1067] Intermediate 86: 1-(2-((2-((2-((tert-butoxycarbonyl)(methyl)amino)ethyl)carbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)-4-fluoro-1-(2-((2-((methoxycarbonyl)-4-methylthiophene-3-yl)amino)-2-Oxoethyl)-4-methylpiperidine-1-onium (zwitterion) (mixture of diastereomers) Specification 80 / 106 pages 88 CN 122180674 A

[1068]

[1069] Similar to Example 4, intermediate 86 was prepared as follows: starting from the corresponding bromide and amine.

[1070] Intermediate 85: 100 mg, 0.21 mmol (1.0 equivalent)

[1071] Intermediate 1: 62 mg, 0.21 mmol (1.0 equivalent)

[1072] Purified by FCC (Sfär amino, 0-5% MeOH / DCM)

[1073] Amount / Yield: 47 mg, 0.062 mmol, 29% yield

[1074] LC-MS Method 1: tR = 0.79 min, MS (ESI) m / z = 682.4 [M]+

[1075] Intermediate 87: N2-(3-amino-4-methylthiophene-2-carbonyl)-N6-(tert-butoxycarbonyl)-L-lysine methyl ester

[1076]

[1077] Similar to intermediate 36, intermediate 87 was prepared as follows: starting from a suitable acid and amine. The reaction was carried out in ACN.

[1078] N6-(tert-butoxycarbonyl)-L-lysine methyl ester hydrochloride: 1.88 g, 6.36 mmol (1.0 equivalent)

[1079] 3-amino-4-methylthiophene-2-carboxylic acid: 1.00 g, 6.36 mmol (1.0 equivalent)

[1080] Purified by FCC (Sfär silica, 10% ACN / DCM)

[1081] Yield / volume: 910 mg, 2.28 mmol, 36% yield

[1082] LC-MS method 1: tR = 1.00 min, MS (ESI) m / z = 300.2 [M+H]+

[1083] Intermediate 88: N2-(3-(2-(azacycloheptane-1-yl)acetamido)-4-methylthiophene-2-carbonyl)-N6-(tert-butoxycarbonyl)-L-lysine methyl ester Specification 81 / 106 pages 89 CN 122180674 A

[1084]

[1085] Similar to intermediate 74, intermediate 88 is prepared as follows: starting from a suitable amine.

[1086] Intermediate 87: 910 mg, 2.28 mmol (1.0 equivalent)

[1087] Purified by FCC (Sfär silica, 80% (EtOAc:EtOH 9:1) / cyclohexane)

[1088] Amount / Yield: 500 mg, 0.93 mmol, 40% yield

[1089] LC-MS Method 1: tR = 0.69 min, MS (ESI) m / z = 539.5 [M+H]+

[1090] Intermediate 89: (S)-1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-((6-((tert-butoxycarbonyl)amino)-1-methoxy-1-oxohexane-2-yl)carbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion)

[1091]

[1092] Similar to Example 4, intermediate 89 was prepared as follows: starting from the corresponding bromide and amine.

[1093] Intermediate 88: 250 mg, 0.27 mmol (1.0 equivalent)

[1094] N-Benzyl-2-bromoacetamide: 73 mg, 0.27 mmol (1.0 equivalent)

[1095] Purified by FCC (Sfäramino, 0-100% ACN / DCM)

[1096] Yield / Quantity: 40 mg, 0.058 mmol, 22% yield

[1097] LC-MS Method 1: tR = 0.80 min, MS (ESI) m / z = 686.3 [M]+

[1098] Intermediate 90: (3-(3-(2-(4,4-dimethylpiperidin-1-yl)acetamido)-4-methylthiophene-2-carbamate)propyl)(methyl)tert-butyl carbamate, 82 / 106 pages, 90 CN 122180674 A

[1099]

[1100] Similar to intermediate 36, intermediate 90 is prepared as follows: starting from a suitable acid and amine.

[1101] Intermediate 69: 1.00 g, 2.71 mmol (1.0 equivalent)

[1102] (3-aminopropyl)(methyl)carbamate tert-butyl ester: 766 mg, 4.07 mmol (1.5 equivalent)

[1103] Purified by FCC (Sfäramino, 50% EtOAc / cyclohexane)

[1104] Amount / yield: 1.60 g, quantitative yield (82 % wt)

[1105] LC-MS method 1: tR = 0.71 min, MS (ESI) m / z = 484.1 [M+H]+

[1106] Intermediate 91: methyl 3-((2-bromoacetamido)methyl)benzoate

[1107]

[1108] Similar to intermediate 65, intermediate 91 was prepared as follows: starting from a suitable amine.

[1109] Methyl 3-(aminomethyl)benzoate hydrochloride: 300 mg, 1.49 mmol (1.0 equivalent)

[1110] Purified by FCC (Sfär silica, 0-10% MeOH / DCM)

[1111] Yield / volume: 334 mg, 1.17 mmol, 79% yield

[1112] LC-MS method 1: tR = 0.76 min, MS (ESI) m / z =284.4 / 286.4[M+H]+

[1113] Intermediate 92: 1-(2-((2-((3-((tert-butoxycarbonyl)(methyl)amino)propyl)carbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)-1-(2-((3-(methoxycarbonyl)benzyl)amino)-2-oxoethyl)-4,4-dimethylpiperidine-1-onium (zwitterion)

[1114] Specification 83 / 106 pages 91 CN 122180674 A

[1115] Similar to Example 4, intermediate 92 was prepared as follows: starting from the corresponding bromide and amine.

[1116] Intermediate 90: 152 mg, 0.31 mmol (1.0 equivalent)

[1117] Intermediate 91: 90 mg, 0.31 mmol (1.0 equivalent)

[1118] Purified by FCC (Sfär amino, 0-2% MeOH / DCM)

[1119] Amount / Yield: 56 mg, 0.08 mmol, 26% yield

[1120] LC-MS Method 1: tR = 0.81 min, MS (ESI) m / z = 686.9 [M]+

[1121] Intermediate 93: ethyl 3-(2-bromoacetamido)-4-methylthiophene-2-carboxylate

[1122]

[1123] Similar to intermediate 1, intermediate 93 was prepared as follows: starting from a suitable amine:

[1124] ethyl 3-amino-4-methylthiophene-2-carboxylate: 820 mg, 4.41 mmol (1.0 equivalent)

[1125] Amount / yield: 1.36 g, quantitative yield.

[1126] LC-MS method 1: tR = 0.92 min, MS (ESI) m / z = 306.1 / 308.1 [M+H]+

[1127] Intermediate 94: 1-(2-(2-((3-((tert-butoxycarbonyl)(methyl)amino)propyl)carbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((2-((2-(ethoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-4,4-dimethylpiperidine-1-onium (zwitterion)

[1128]

[1129] Similar to Example 4, intermediate 94 was prepared as follows: starting from the corresponding bromide and amine.

[1130] Intermediate 90: 120 mg, 0.21 mmol (1.0 equivalent)

[1131] Intermediate 93: 77 mg, 0.21 mmol (1.0 equivalent)

[1132] Purified by FCC (Sfär amino, 0–2% MeOH / DCM)

[1133] Amount / Yield: 43 mg, 0.06 mmol, 29% yield

[1134] LC-MS Method 1: tR = 0.93 min, MS(ESI) m / z = 706.3 [M]+

[1135] Intermediate 95: 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-(((1-(tert-butoxycarbonyl)piperidin-4-yl)methyl)carbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion) Specification 84 / 106 pages 92 CN 122180674 A

[1136]

[1137] Similar to Example 4, intermediate 95 was prepared as follows: starting from the corresponding bromide and amine.

[1138] Intermediate 102: 130 mg, 0.26 mmol (1.0 equivalent)

[1139] N-Benzyl-2-bromoacetamide: 60 mg, 0.26 mmol (1.0 equivalent)

[1140] Purified by FCC (Sfäramino, 0-5% MeOH / DCM)

[1141] Amount / Yield: 76 mg, 0.11 mmol, 40% yield

[1142] LC-MS Method 1: tR = 0.85 min, MS (ESI) m / z = 640.4 [M]+

[1143] Intermediate 96: 4-(3-(2-(azacycloheptane-1-yl)acetamido)-4-methylthiophene-2-carboxamido)piperidine-1-carboxylic acid tert-butyl ester

[1144]

[1145] Similar to intermediate 36, intermediate 96 is prepared as follows: starting from a suitable acid and amine.

[1146] Intermediate 13: 500 mg, 1.41 mmol (1.0 equivalent)

[1147] 4-Aminopiperidine-1-carboxylic acid tert-butyl ester: 423 mg, 2.11 mmol (1.3 equivalent)

[1148] Purified by FCC (Sfär silica, 30% EtOAc / cyclohexane)

[1149] Yield / volume: 400 mg, 0.84 mmol, 59% yield

[1150] LC-MS method 1: tR = 0.67 min, MS (ESI) m / z = 479.3 [M+H]+

[1151] Intermediate 97: 1-(2-((2-((1-(tert-butyloxycarbonyl)piperidine-4-yl)carbamoyl)-4-methylthiophene-3- (2-(methoxycarbonyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion) Specification 85 / 106 pages 93 CN 122180674 A

[1152]

[1153] Similar to Example 4, intermediate 97 was prepared as follows: starting from the corresponding bromide and amine.

[1154] Intermediate 1: 128 mg, 0.41 mmol (1.0 equivalent)

[1155] Intermediate 96: 199 mg, 0.41 mmol (1.0 equivalent)

[1156] Purified by FCC (Sfär amino, 0-3% MeOH / DCM)

[1157] Amount / yield: 110 mg, 0.16 mmol, 38% yield

[1158] LC-MS method 1: tR = 1.02 min, MS (ESI) m / z = 690.7 [M]+

[1159] Intermediate 98: (2-(3-(2-(azacycloheptane-1-yl)acetamido)-4-methylthiophene-2-carbamate)ethyl)(methyl)carbamate tert-butyl ester

[1160]

[1161] Similar to intermediate 36, intermediate 98 was prepared as follows: starting from a suitable amine and carboxylic acid.

[1162] Intermediate 13: 2.0 g, 6.75 mmol (1.0 equivalent)

[1163] (2-aminoethyl)(methyl)carbamate tert-butyl ester: 1.76 g, 10.0 mmol (1.5 equivalent)

[1164] Purified by FCC (Sfäramino, 65% (EtOAc:EtOH 9:1) / DCM)

[1165] Amount / Yield: 700 mg, 1.54 mmol, 23% yield.

[1166] LC-MS Method 1: tR = 0.60 min, MS (ESI) m / z = 453.7 [M]+

[1167] Intermediate 99: 2-bromo-N-(2-(trifluoromethoxy)benzyl)acetamide Specification 86 / 106 pages 94 CN 122180674 A

[1168]

[1169] 2-bromoacetyl bromide (0.78 mL, 8.55 mmol) was added dropwise to a suspension of potassium carbonate (1.34 g, 9.69 mmol) and (2-(trifluoromethoxy)phenyl)methylamine (0.56 g, 5.70 mmol) in ACN (20 mL) at 0 °C. After stirring at RT for 2 h, the solid and the fraction filtered by vacuum evaporation were filtered off. The crude product was purified by FCC (KP Sfäramino, 3% MeOH / DCM) to give the title compound (Intermediate 99: 737 mg, 2.36 mmol, 45% yield).

[1170] LC-MS Method 1: tR = 0.92 min, MS (ESI) m / z = 312.0 / 314.0 [M+H]+

[1171] Intermediate 100: 1-(2-((2-((2-(tert-butoxycarbonyl)(methyl)amino)ethyl)carbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)-1-(2-oxo-2-((2-(trifluoromethoxy)benzyl)amino)ethyl)azacycloheptane-1-onium (zwitterion)

[1172]

[1173] Similar to Example 4, intermediate 100 was prepared as follows: starting from the corresponding bromide and amine.

[1174] Intermediate 98: 200 mg, 0.42 mmol (1.0 equivalent)

[1175] Intermediate 99: 170 mg, 0.42 mmol (1.0 equivalent)

[1176] Purified by FCC (Sfär amino, 10% MeOH / DCM)

[1177] Amount / Yield: 68 mg, 0.099 mmol, 24% yield

[1178] LC-MS Method 1: tR = 0.84 min, MS (ESI) m / z = 684.3 [M]+

[1179] Example 33: 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(piperazine-1- Carbonyl)thiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium chloride hydrochloride

[1180] was added to a solution of DCM (1 mL) containing HCl in 2 M Et2O (0.5 mL, 1 mmol) as the chloride salt in Example 34 (17 mg, 0.030 mmol) at RT. The evaporation was evaporated after 16 and the solid was washed with Et2O to provide the title compound (Example 33: 12 mg, 0.021 mmol, 79% yield).

[1181] LC-MS Method 2: tR = 0.66 min, MS (ESI) m / z = 489.2 [M]+

[1182] 1H NMR (500 MHz, DMSO-d6) δppm 11.82 (br s, 1 H), 10.84 (s, 1 H), 9.16 (br s, 2 H), 8.89 (d, J=1.8 Hz, 1 H), 7.40 (d, J=1.0 Hz, 1 H), 6.92 (d, J=1.6 Hz, 1 H), 4.71 (s, 2 H), 4.66 (s, 2 H), 3.77-4.03 (m, 4 H), 3.67-3.75 (m, 4 H) H), 3.13 (br s, 4 H), 2.11 (s, 3 H), 1.92-2.09 (m, 4 H), 1.66 (br s, 4 H).

[1183] Similar to the description of Example 33, the following examples were prepared by starting from suitable intermediates or examples.

[1184] Specification 88 / 106 pages 96 CN 122180674 A

[1185] Specification 89 / 106 pages 97 CN 122180674 A

[1186] Specification 90 / 106 pages 98 CN122180674 A

[1187]

[1188] Example 49: 1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((4-methyl-2-(4-methylpiperazin-1-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion)

[1189] Step 1 - 1-(2-((2-(tert-butoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium bromide (intermediate 46) Specification 91 / 106 pages 99 CN 122180674 A

[1190]

[1191] Similar to intermediate 42, intermediate 46 was prepared as follows: starting from a suitable amine and bromide:

[1192] Intermediate 40: 200 mg, 0.57 mmol (1.0 equivalent)

[1193] Intermediate 1: 167 mg, 0.57 mmol (1.0 equivalent)

[1194] Amount / yield: 226 mg, 0.35 mmol, 62% yield.

[1195] LC-MS Method 1: tR = 0.83 min, MS (ESI) m / z = 564.3 [M]+

[1196] Step 2 - 1-(2-((2-carboxy-4-methylthiophene-3-yl)amino)-2-oxoethyl)-1-(2-((2-(methoxycarbonyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium chloride (intermediate 47)

[1197]

[1198] Similar to intermediate 43, intermediate 47 was prepared as follows: starting from a suitable ester.

[1199] Intermediate 46: 223 mg, 0.35 mmol (1.0 equivalent)

[1200] Amount / yield: 226 mg, 83% wt, quantitative yield.

[1201] Method 1: tR = 0.65 min, MS (ESI) m / z = 508.2 [M]+

[1202] Step 3 - 1-(2-((2-(methoxycarbonyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)-1-(2-((4-methyl-2-(4-methylpiperazine-1-carbonyl)thiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion) (Example 49)

[1203] Similar to Example 27, Example 49 was prepared as follows: starting from suitable amines and carboxylic acids. Specification 92 / 106 pages 100 CN 122180674 A

[1204] Intermediate 47: 115 mg, 0.21 mmol (1.0 equivalent)

[1205] 1-Methylpiperazine: 42 mg, 0.42 mmol (2.0 equivalent)

[1206] Purified by FCC (Sfäramino, 0-5% MeOH / DCM).

[1207] Amount / yield: 12 mg, 0.020 mmol, 9% yield.

[1208] LC‑MS Method 1: tR = 1.04 min, MS (ESI) m / z = 590.4 [M]+

[1209] 1H NMR (500 MHz, DMSO‑d6) δppm 7.36 (br s, 1 H), 7.25 (s, 1 H), 4.61 (br s, 2 H), 4.05‑ 4.43 (m, 2 H), 3.67 - 3.87 (m, 4 H), 3.68 (s, 3 H), 3.40 - 3.50 (m, 4 H), 2.27 (br t, J=4.7 Hz, 4 H), 2.15 (s, 3 H), 1.93 - 2.04 (m, 4 H), 1.93 (br s, 6 H), 1.58- 1.73 (m, 4 H)

[1210] Example 54: 1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((4-methyl-2-(morpholin-4-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion)

[1211] Similar to Example 27, Example 54 was prepared as follows: starting from suitable amines and carboxylic acids.

[1212] Intermediate 47: 115 mg, 0.21 mmol (1.0 equivalent)

[1213] Morpholine: 0.027 mL, 0.32 mmol (1.5 equivalent)

[1214] Purified by FCC (Sfär amino, 0-5% MeOH / DCM)

[1215] Amount / Yield: 38 mg, 0.066 mmol, 29% yield.

[1216] LC‑MS Method 1: tR = 0.66 min, MS (ESI) m / z = 577.3 [M]+

[1217] 1H NMR (500 MHz, DMSO‑d6) δppm 14.14 (br s, 1 H), 7.33 (br s, 1 H), 7.24 (s, 1 H), 4.60 (br s, 2 H), 4.15 (br s, 2 H), 3.66 (s, 3 H), 3.61 - 3.86 (m, 4 H), 3.55 - 3.62 (m, 4 H), 3.45 - 3.52 (m, 4 H), 1.91 - 2.04 (m, 4 H), 1.84 - 1.92 (m, 6 H), 1.65 (brs, 4 H)

[1218] Example 37: 1-(2-(((2-carbamoyl-4-methylthiophen-3-yl)methyl)amino)-2-oxoethyl)-1-(2-(isoxazol-3-ylamino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion)

[1219] Step 1 3-iodo-4-methylthiophen-2-carboxylic acid methyl ester (intermediate 48)

[1220]

[1221] Diiodomethane (5.65 mL, 70.1 mmol) was added to a solution of 3-amino-4-methylthiophen-2-carboxylic acid methyl ester (3.00 g, 17.5 mmol) in ACN (44 mL) under a nitrogen atmosphere and the reaction mixture was stirred at 70 °C for 20 minutes. After this time, the reaction mixture was cooled to RT and tert-butyl nitrite (3.80 mL, 35.0 mmol) was added dropwise over 10 minutes. After the addition was complete, the reaction mixture was stirred at 60 °C for 4 hours. The mixture was then slowly poured into 0.5 M aqueous HCl (30 mL) and extracted with EtOAc (3x). The collected organic layer was dried over MgSO4 and concentrated under reduced pressure to give a crude product, which was purified by FCC (KP Sfär silica, 0–10% EtOAc / cyclohexane) to provide the title compound (intermediate 48: 2.90 g, 10.3 mmol, 59% yield).

[1222] LC-MS Method 1: tR = 1.15 min, MS (ESI) m / z = 283.0 [M+H]+ Instructions 93 / 106 pages 101 CN 122180674 A

[1223] Step 2 - methyl 3-cyano-4-methylthiophene-2-carboxylate (intermediate 51)

[1224]

[1225] A round-bottom flask was loaded with methyl 3-iodo-4-methylthiophene-2-carboxylate (intermediate 48, 2.90 g, 10.3 mmol), potassium acetate (1.41 g, 14.4 mmol), XPhos (0.49 g, 1.03 mmol) and XPhos Pd G3 (0.87 g, 1.03 mmol). The flask was purged and backfilled with nitrogen (3x), and then potassium hexacyanoferrate(II) 0.1 N standard solution (60.0 mL, 6.00 mmol) and 1,4-dioxane (52 mL) were added under a nitrogen atmosphere. Three vacuum-nitrogen cycles were then performed, and the reaction mixture was heated at 100 °C with stirring for 4 hours. XPhos Pd G3 (0.16 mg, 0.19 mmol) and potassium hexacyanoferrate(II) 0.1 N standard solution (60.0 mL, 6.00 mmol, degassed before addition) were then added at RT. The mixture was then heated at 100 °C with stirring.The reaction mixture was heated for 5 hours. It was then cooled to RT and extracted with EtOAc. The collected organic layer was concentrated under reduced pressure to provide a crude product, which was purified by FCC (KP Sfär silica, 0-10% EtOAc / cyclohexane) to give the title compound (intermediate 51: 0.78 g, 4.30 mmol, 42% yield).

[1226] LC-MS Method 1: tR = 0.89 min, MS (ESI) m / z = 181.9 [M+H]+

[1227] Step 3 - 3-(aminomethyl)-4-methylthiophene-2-carboxamide (intermediate 49)

[1228]

[1229] In a hydrogenation flask under nitrogen atmosphere, Raney® nickel (70.0 mg, 50% aqueous slurry) followed by a 7 M MeOH solution of ammonia (15.0 mL, 185 mmol) of methyl 3-cyano-4-methylthiophene-2-carboxylic acid (intermediate 51, 680 mg, 3.75 mmol) was added to a solution of methyl 3-cyano-4-methylthiophene-2-carboxylic acid (intermediate 51, 680 mg, 3.75 mmol) in 100 mL. The flask was emptied, backfilled with hydrogen (1 atm), and stirred for 24 h. The solid was then allowed to settle, and the organic phase was removed under nitrogen atmosphere. The remaining solid was washed with MeOH and DCM. The collected organic layer was concentrated under reduced pressure to provide a crude product, which was purified by FCC (Sfär amino, 0–100% EtOAc / MeOH (9 / 1 v / v), in cyclohexane) to provide the title compound (intermediate 49: 173 mg, 1.01 mmol, 27% yield).

[1230] LC-MS Method 2: tR = 0.46 min, MS (ESI) m / z = 171.1 [M+H]+

[1231] Step 4 - 3-((2-(azacycloheptane-1-yl)acetamido)methyl)-4-methylthiophene-2-carboxamide (intermediate 50)

[1232] Specification 94 / 106 pages 102 CN 122180674 A

[1233] 2-chloroacetyl chloride (70 mL, 0.89 mmol) was slowly added to a solution of 3-(aminomethyl)-4-methylthiophene-2-carboxamide (intermediate 49, 152 mg, 0.89 mmol) and triethylamine (0.12 mL, 0.89 mmol) in ACN (9 mL) at 0 °C. The reaction was stirred at RT for 4 hours. Then, azircycloheptane (600 mL, 5.36 mmol) was added to the reaction mixture at RT. After stirring for 3 hours, the volatiles were removed under reduced pressure, and the crude product was purified by FCC (Sfäramino, 0–100% EtOAc / MeOH (9 / 1 v / v), in cyclohexane) to provide the title compound (intermediate 50: 140 mg, 0.452 mmol, 51% yield).

[1234] LC-MS Method 2: tR = 0.82 min, MS (ESI) m / z = 310.2 [M+H]+

[1235] Step 5 - 1-(2-(((2-carbamoyl-4-methylthiophene-3-yl)methyl)amino)-2-oxoethyl)-1-(2-(isoxazo-3-ylamino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion) (Example 37)

[1236] Similar to Example 39, Example 37 was prepared as follows: starting from a suitable amine and bromide.

[1237] Intermediate 50: 40 mg, 0.13 mmol (1.0 equivalent)

[1238] Intermediate 2: 30 mg, 0.15 mmol (1.1 equivalent)

[1239] Purified by FCC (Sfär amino, 0-100% EtOAc / MeOH (9 / 1 v / v), in cyclohexane)

[1240] Amount / yield: 10 mg, 0.023 mmol, 18% yield.

[1241] LC‑MS Method 2: tR = 0.59 min, MS (ESI) m / z = 434.3 [M]+

[1242] 1H NMR (500 MHz, DMSO‑d6) δppm 9.23 (br t, J=5.0 Hz, 1 H), 8.33 (d, J=1 .5 Hz, 1 H), 7.90 (br s, 1 H), 7.48 (br s, 1 H), 7.26 (d, J=0.7 Hz, 1 H), 6.75 (d, J=1.6 Hz, 1 H), 4.57 (s, 2 H), 4.50 (d, J=5.1 Hz, 2 H), 4.04 (s, 2 H), 3.61 - 3.79 (m, 4 H), 2.16 (s, 3 H), 1.85 (br s, 4 H), 1.58 (br s, 4 H).

[1243] Example 40: (R)-1-(2-(((2-carbamoyl-4-methylthiophene-3-yl)methyl)amino)-2-oxoethyl)-1-(2-oxo-2-((1-phenylethyl)amino)ethyl)azacycloheptane-1-onium bromide

[1244] Similar to Example 39, Example 40 was prepared as follows: starting from a suitable amine and bromide.

[1245] Intermediate 50: 50 mg, 0.16 mmol (1.0 equivalent)

[1246] Intermediate 3: 44 mg, 0.18 mmol (1.1 equivalent)

[1247] Purified by FCC (Sfär-C18, 0-50% ACN + 0.1% HCOOH / H2O + 0.1% HCOOH).

[1248] Amount / Yield: 7.5 mg, 0.014 mmol, 8% yield.

[1249] LC-MS Method 1: tR = 0.65 min, MS (ESI) m / z = 471.32 [M]+

[1250] 1H NMR (500 MHz, DMSO-d6) δppm 9.11 (d, J=7.7 Hz, 1 H), 8.76 (t, J=5.4 Hz, 1 H), 7.39–7.95 (m, 2 H), 7.30–7.37 (m, 4 H), 7.29 (s, 1 H), 7.22–7.27 (m, 1 H), 4.92 (quin, J=7.1 Hz, 1 H), 4.46–4.57 (m, 2 H), 4.44 (s, 2 H), 4.37 (s, 2 H), 3.55–3.78 (m, 4 H), 2.15 (s, 3 H), 1.86 (br s, 4 H), 1.58 (br s, 4 H), 1.37 (d, J = 7.1 Hz, 3 H).

[1251] The conversion of the zwitterionic form of the compounds of the present invention to obtain the corresponding salt form can be carried out as described in the following examples (chloride salts).

[1252] Example 1, as a chloride salt: 2-(benzylamino)-N,N-dimethyl-N-(2-((4-methyl-2-(pyrrolidine-1-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)-2-oxoethyl-1-ammonium chloride

[1253] Example 1 (zwitterionic) (33 mg, 0.070 mmol) was dissolved in DCM (1.0 mL) and HCl was added dropwise in a 2 M Et2O solution (50 µL, 0.11 mmol) at RT. After 5 minutes, the volatiles were removed by depressurization, yielding the title compound (Example 1, as a chloride salt: 7 mg, 0.015 mmol, 21% yield), which is a white powder.

[1254] LC-MS Method 1: tR = 0.63 min, m / z = 443.4 [M]+

[1255] 1H NMR (400 MHz, DMSO-d6) δppm 10.37 (s, 1H), 9.06 (t, J = 5.8 Hz, 1H), 7.40 - 7.21 (m, 6H), 4.56 (s, 2H), 4.40 (s, 2H), 4.36 (d, J = 5.8 Hz, 2H), 3.40 (s, 6H), 3.48 - 3.35 (m, 4H), 2.07 (d, J = 1.0 Hz, 3H), 1.81 (br s, 4H)

[1256] Similar to the procedure described in Example 1 as a chloride salt, it begins with a suitable zwitterion as an amphoteric ion.Example: The following embodiments have been prepared.

[1257]

[1258] Specification 96 / 106 pages 104 CN 122180674 A

[1259] Example 78: 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-methyl-4-(morpholino-4-carbonyl)thiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium bromide

[1260] Step 1 - 4-(2-bromoacetamido)-5-methylthiophene-3-carboxylic acid methyl ester (intermediate 103)

[1261]

[1262] Similar to intermediate 1, intermediate 103 is prepared as follows: starting from a suitable amine:

[1263] 4-amino-5-methylthiophene-3-carboxylic acid methyl ester hydrochloride: 1.50 g, 7.22 mmol (1.0 equivalent)

[1264] Amount / Yield: 1.57 g, 5.36 mmol, 74% yield.

[1265] UPLC Method 1: rt 0.79 min, MS (ESI) m / z = 292.0 / 294.0 [M+H]+

[1266] Step 2 - 4-(2-(azacycloheptane-1-yl)acetamido)-5-methylthiophene-3-carboxylic acid methyl ester (intermediate 104)

[1267]

[1268] Similar to intermediate 11, intermediate 104 was prepared as follows: starting from a suitable bromide:

[1269] Intermediate 103: 1.57 g, 5.36 mmol (1.0 equivalent)

[1270] Amount / Yield: 1.52 g, 4.90 mmol, 91% yield.

[1271] UPLC method 1: rt 0.46 min, MS (ESI) m / z = 311.1 [M+H]+

[1272] Step 3 - 4-(2-(azacycloheptane-1-yl)acetamido)-5-methylthiophene-3-carboxylic acid sodium chloride (intermediate 105)

[1273]

[1274] Similar to intermediate 7, intermediate 105 was prepared as follows: starting from a suitable ester:

[1275] Intermediate 104: 1.52 g, 4.90 mmol (1.0 equivalent)

[1276] Amount / yield: 1.97 g, 88% wt, quantitative yield.

[1277] UPLC Method 1: rt 0.41 min, MS (ESI) m / z = 297.1 [M+H]+

[1278] Step 4 - 2-(azacycloheptane-1-yl)-N-(2-methyl-4-(morpholino-4-carbonyl)thiophene-3-yl)acetamide (intermediate 106) Specification 97 / 106 pages 105 CN 122180674 A

[1279]

[1280] Similar to intermediate 8, intermediate 106 is prepared as follows: starting from a suitable acid and amine:

[1281] Intermediate 105: 350 mg, 0.97 mmol (1.0 equivalent)

[1282] Morpholine: 0.17 mL, 1.97 mmol (2.0 equivalent)

[1283] Amount / yield: 211 mg, 0.57 mmol, 59% yield.

[1284] UPLC Method 1: rt 0.44 min, MS (ESI) m / z = 366.2 [M+H]+

[1285] Step 5 - 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-methyl-4-(morpholino-4-carbonyl)thiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium bromide (Example 78)

[1286] Similar to Example 3, Example N1568-18-1 CHD-074186 was prepared as follows: starting from suitable amines and bromides:

[1287] Intermediate 106: 110 mg, 0.30 mmol (1.0 equivalent)

[1288] N-benzyl-2-bromoacetamide: 76 mg, 0.33 mmol (1.1 equivalent)

[1289] Dosage / Yield: 121 mg, 0.20 mmol, 68% yield.

[1290] UPLC method 1: rt 0.64 min, MS (ESI) m / z = 513.2 [M]+

[1291] 1H NMR (500 MHz, DMSO-d6) δ = 10.55–10.16 (m, 1H), 9.05 (br s, 1H), 7.56–7.06 (m, 6H), 4.36 (d, J = 5.1 Hz, 2H), 4.61–4.21 (m, 4H), 3.89–3.63 (m, 4H), 3.61–3.55 (m, 4H), 3.53–3.39 (m, 4H), 2.23 (br s, 3H), 2.01–1.86 (m, 4H), 1 .72 - 1.57 (m, 4H)

[1292] Example 79: 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-methyl-4-(pyrrolidine-1-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium bromide

[1293] Step 1 - 2-(azacycloheptane-1-yl)-N-(2-methyl-4-(pyrrolidine-1-carbonyl)thiophen-3-yl)acetamide (intermediate 107)

[1294]

[1295] Similar to intermediate 8, intermediate 107 is prepared as follows: starting from a suitable acid and amine:

[1296] Intermediate 104: 350 mg, 0.97 mmol (1.0 equivalent)

[1297] Pyrrolidine: 0.16 mL, 1.97 mmol(2.0 equivalent)

[1298] Amount / yield: 204 mg, 0.58 mmol, 59% yield.

[1299] UPLC Method 1: rt 0.50 min, MS (ESI) m / z = 350.2 [M+H]+

[1300] Step 2 - 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-methyl-4-(pyrrolidine-1-carbonyl)thiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium bromide (Example 79)

[1301] Similar to Example 3, Example 79 was prepared as follows: starting from a suitable amine and bromide:

[1302] Intermediate 107: 110 mg, 0.32 mmol (1.0 equivalent)

[1303] N-benzyl-2-bromoacetamide: 79 mg, 0.35 mmol (1.1 equivalent)

[1304] Amount / Yield: 96 mg, 0.17 mmol, 53% yield.

[1305] UPLC method 1: rt 0.68 min, MS (ESI) m / z = 497.3 [M]+

[1306] 1H NMR (500 MHz, DMSO-d6) δ = 10.28 (br s, 1H), 9.78–8.64 (m, 1H), 7.64–7.40 (m, 1H), 7.38–7.18 (m, 5H), 4.64–4.20 (m, 6H), 3.90–3.66 (m, 4H), 3.43 (br t, J = 6.3 Hz, 2H), 3.33–3.25 (m, 2H), 2.24 (br s, 3H), 2.00–1.87 (m, 4H), 1.85 - 1.71 (m, 4H), 1.69 - 1.59 (m, 4H)

[1307] Example 80: 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-((2-methoxyethyl)(methyl)carbamoyl)-2-methylthiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium bromide

[1308] Step 1 - 4-(2-(azacycloheptane-1-yl)acetamido)-N-(2-methoxyethyl)-N,5-dimethylthiophene-3-carboxamide (intermediate N1568-17-1)

[1309]

[1310] Similar to intermediate 8, intermediate N1568-17-1 was prepared as follows: starting from a suitable acid and amine:

[1311] Intermediate 101: 350 mg, 0.97 mmol (1.0 equivalent)

[1312] 2-Methoxy-N-methylethyl-1-amine: 0.20mL, 1.97 mmol (2.0 equivalent)

[1313] Amount / yield: 200 mg, 0.54 mmol, 55% yield.

[1314] UPLC Method 1: rt 0.47 min, MS (ESI) m / z = 368.2 [M+H]+

[1315] Step 2 - 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-((2-methoxyethyl)(methyl)carbamoyl)-2-methylthiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium bromide (Example 80)

[1316] Similar to Example 3, Example 80 was prepared as follows: starting from suitable amines and bromides:

[1317] Intermediate N1568-17-1: 110 mg, 0.30 mmol (1.0 equivalent)

[1318] N-benzyl-2-bromoacetamide: 75 mg, 0.33 mmol (1.1 equivalent)

[1319] Dosage / Yield: 97 mg, 0.16 mmol, 54% yield.

[1320] UPLC method 1: rt 0.66 min, MS (ESI) m / z = 515.3 [M]+

[1321] 1H NMR (500 MHz, DMSO-d6) δ = 10.22 (br s, 1H), 9.48–8.63 (m, 1H), 7.49–7.37 (m, 1H), 7.37–7.22 (m, 5H), 4.65–4.39 (m, 4H), 4.35 (br d, J = 5.1 Hz, 2H), 3.89–3.65 (m, 4H), 3.55–3.38 (m, 4H), 3.27–3.18 (m, 3H), 3.06–2.83 (m, 3H), 2.24 (br s, 3H), 1.99–1.85 (m, 4H), 1.73–1.57 (m, 4H)

[1322] Compounds C1 and C2 were prepared as described below.

[1323] C1 is characterized by having a phenyl ring instead of the amide-substituted tiophene or benzothiophene ring of the compounds of the present invention. Specification 99 / 106 pages 107 CN 122180674 A

[1324] C2 is characterized by having a different heteroaryl ring instead of the amide-substituted tiophene or benzothiophene ring of the compounds of the present invention.

[1325] Compound C1: N,N-di(phenylcarbamoylmethyl)dimethylammonium chloride (i.e., carbaryl) was prepared as described on page 6 of WO99 / 63985.

[1326] Compound C2: 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((5-(methoxycarbonyl)-3-methylisoxazo-4-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion)

[1327] Step 1: 4-(2-bromoacetamido)-3-methylisoxazo-5-carboxylic acid methyl ester (intermediate 52)

[1328]

[1329] 2-bromoacetyl bromide (0.31 mL, 3.52 mmol) was added dropwise to a DCM (20 mL) suspension of potassium carbonate (1.10 g, 8.01 mmol) and 4-amino-3-methylisoxazo-5-carboxylic acid methyl ester (500 mg, 3.20 mmol). After 3 h, water was added and the organic phase was separated, dried and concentrated under reduced pressure on a phase separator. The residue was purified by FCC (KP Sfär silica, 0–40% cyclohexane / EtOAc) to provide the title compound (intermediate 52,800 mg, 2.89 mmol, 90% yield) as a creamy white powder.

[1330] LC-MS Method 1: tR = 0.66 min, MS (ESI) m / z = 277.1 / 279.0 [M+H]+

[1331] Step 2 - 2-(azacycloheptane-1-yl)-N-(isoxazo-3-yl)acetamide (intermediate 54)

[1332]

[1333] Azacycloheptane (0.28 mL, 2.49 mmol) and potassium carbonate (361 mg, 2.62 mmol) were added to a stirred solution of intermediate 53 (200 mg, 1.25 mmol) in ACN (5 mL), and the mixture was stirred at RT for 2 h. The evaporation was evaporated, and the residue was partitioned between water and EtOAc. The organic phase was separated, and the aqueous phase was extracted with EtOAc. The collected organic layer was washed with brine and concentrated under reduced pressure to provide the title compound (intermediate 54: 270 mg, 1.21 mmol, 97% yield), an orange oil.

[1334] LC-MS Method 2: tR = 0.78 min, MS (ESI) m / z = 224.4 [M]+

[1335] Step 3 - 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((5-(methoxycarbonyl)-3-methylisoxazo-4-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (zwitterion) (compound C2)

[1336] Intermediate 52 (50 mg, 0.180 mmol) was added to a stirred solution of intermediate 54 (40 mg, 0.180 mmol) in ACN (1.2 mL). The mixture was stirred at RT for 24 h. The mixture was then concentrated under reduced pressure, and the residue was passed through an FCC (Sfär amino, 0-5%) filter.MeOH / DCM purification yielded the title compound (compound C2: 57 mg, 0.114 mmol, 75% yield).

[1337] LC-MS Method 2: tR = 0.52 min, MS (ESI) m / z = 420.1 [M]+

[1338] 1H NMR (500 MHz, DMSO-d6) δppm 12.25 - 14.42 (m, 1 H), 8.76 (d, J=1.6 Hz, 1 H), 6.92 (d, J=1.6 Hz, 1 H), 4.61 (s, 2 H), 4.14 (s, 2 H), 3.74 (s, 3 H), 3.68 - 3.85 (m, 4 H), 2.11 (s, 3 H), 1.93 (br s, 4 H), 1.63 (br s, 4 H)

[1339] Pharmacological activity of the compounds of the present invention

[1340] hNaV 1.7 cell assay protocol

[1341] Intracellular and extracellular inhibition of NaV 1.7 by representative embodiments of the present invention were tested in an automated patch-clamp assay described herein.

[1342] Cell culture

[1343] CHO cells stably expressing the human NaV 1.7 channel were obtained from B'SYS GmbH, Switzerland. Cells were cultured in F12 Nutmix (Gibco, Carlsbad CA) supplemented with 10% fetal bovine serum (Invitrogen, Waltham MA), 1% penicillin-streptomycin (Gibco, Carlsbad CA) and 200 µg / ml hygromycin B (Invitrogen, Waltham MA). Cells were grown and maintained at 37 °C in a humidified environment containing 5% CO2 / air. Cells were removed from the culture flasks for passage and harvesting using TrypLE Express (Sigma-Aldrich, St. Louis, MO). Experimental flasks were prepared using 2–4 day old cells without antibiotics and used at ~80% confluence. Before the experiment, the cells were washed with Ca2+ and Mg2+-free PBS and desorbed with pre-warmed TrypLE Express. After a brief centrifugation step, the cells were resuspended in CHO-S-serum-free medium II (Life Technologies, Carlsbad CA). The cells were counted, and the final concentration was set at 3–4 million cells per mL.

[1344] Patch-clamp solution & drugs

[1345] The intracellular solution (or internal solution) contained the following: 140 mM CsF, 10 mM NaCl, 1 mM EGTA and 10 mMHEPES, adjusted to pH 7.2 with CsOH, and the volumetric osmolarity was adjusted to 325 with sucrose. The extracellular solution (or external solution) contained the following: 40 mM NMDG, 100 mM NaCl, 4 mM KCl, 1 mM MgCl2, 2 mM CaCl2, 10 mM HEPES, 10 mM glucose, adjusted to pH 7.4 with HCl and the volumetric osmolarity was adjusted to 310 with sucrose. All chemicals were from Sigma-Aldrich, St. Louis, MO.

[1346] For both intracellular and extracellular applications of the test compound, the spotted compound was dissolved in intracellular and extracellular solutions, respectively, and tested up to 60 μM, which resulted in a pIC50 value of not less than 4.2.

[1347] Automated Patch-Clamp Test Protocol

[1348] The automated patch-clamp was performed as follows: The temperature was set to 22°C on a Qube 384 (Sophion Bioscience A / S, Ballerup, Denmark) equipped with multi-well QChips. A whole-cell configuration was formed using the default Qube seal and break-in parameters. The membrane potential was maintained at -100 mV before the voltage program began. The voltage program consisted of 20 pulses, with 30 scans for intracellular applications and 55 scans for extracellular applications, both before and after compound application. The cells were maintained at -100 mV, and a depolarization pulse to -20 mV was applied for 10 ms. The scan interval was set to 9 s while the holding potential was -100 mV.

[1349] Intracellular Inhibition of the Test Compound

[1350] For intracellular application of the compound, the QChip was removed from the recording chamber. The intracellular solution was replaced with a solution containing the test compound. After intracellular solution exchange, the QChip was returned to the recording chamber and a voltage pulse was applied.

[1351] Data filtering was performed using Sophion analyzer software. Only wells with a minimum sealing resistance of 40 MΩ and a pre-compound application current of 1 nA were used for further analysis using Aplus software. The average peak current amplitude of the 20th pulse of the last three programs applied in the pre-compound application and post-compound application groups was calculated. For each well, the post-compound application / pre-compound application ratio was calculated.

[1352] The results were then normalized as follows: 0.3% DMSO control was considered 0%, and complete blockade (100 µM N-(2,6-dimethylaminocarbamoylmethyl)triethylammonium chloride, i.e., QX314, for internal application) was considered 100% inhibition. % activity was used as the percentage of activity.Figures were plotted, and concentration-response curves were fitted using a 4-parameter logarithmic Hill formula. From the fitted curves, IC50 values ​​were estimated and intracellular pIC50 values ​​were calculated.

[1353] External blocking effect of the test compound (extracellular inhibition)

[1354] For extracellular administration of the compound, the extracellular solution was replaced with a solution containing the test compound. After the extracellular solution exchange, a voltage pulse was applied.

[1355] Data filtering was performed using Sophion analyzer software. Only wells with a minimum sealing resistance of 50 MΩ and a pre-compound application current of 1 nA were used for further analysis using Aplus software. The average peak current amplitude of the 20th pulse of the last three procedures applied in the pre-compound application and post-compound application groups was calculated. For each well, the post-compound application / pre-compound application ratio was calculated.

[1356] The results were then normalized as follows: 0.3% DMSO control was considered 0%, and complete blocking (300 µM tetracaine, for external application) was considered 100% inhibition. Plotting % activity values ​​and fitting concentration-response curves using a 4-parameter logarithmic Hill formula. From the fitted curves, IC50 values ​​were estimated and extracellular pIC50 values ​​were calculated.

[1357] A pIC50 of ≤4.2 was obtained at the maximum compound concentration of 60 μM, where inhibition was less than or equal to 50%.

[1358] Intracellular and extracellular inhibition of representative examples are reported in Table 2.

[1359] Table 2 - Intracellular and extracellular inhibition of representative examples Specification 102 / 106 pages 110 CN 122180674 A

[1360] Specification 103 / 106 pages 111 CN 122180674 A

[1361]

[1362] As can be appreciated, the compounds in Table 2 (i.e., the compounds according to the invention) exhibit high intracellular inhibitory activity against the NaV 1.7 receptor. Conversely, the compounds in Table 2 exhibit lower extracellular inhibitory activity against the NaV 1.7 receptor.

[1363] Table 3 shows the inhibition data of the selected embodiments tested in both zwitterionic and salt (chloride) forms. As can be appreciated, both forms exhibit high intracellular inhibitory activity against the NaV 1.7 receptor. Negligible variations were observed in some cases between the respective zwitterionic and salt forms, which can be considered within the range of experimental error.

[1364] Table 3 - Comparison of intracellular and extracellular inhibition of representative embodiments in zwitterionic or salt forms, hNav 1.7

[1365]

[1366] Accordingly, the compounds of the present invention in both zwitterionic and salt forms can be used to treat respiratory diseases such as cough, asthma, idiopathic pulmonary fibrosis (IPF) and chronic obstructive pulmonary disease (COPD), and especially chronic cough.

[1367] Comparison of compounds

[1368] Compounds C1 and C2 were tested in the same binding test described above and their intracellular and extracellular pIC50 values ​​are reported in Table 4.

[1369] Table 4 - Intracellular and extracellular inhibition of comparative compounds Specification 104 / 106 pages 112 CN 122180674 A

[1370]

[1371] As shown in Tables 2, 3 and 4 (Example 7), the compounds of formula (I) of the present invention exhibit intracellular inhibitory activity against NaV 1.7, expressed as pIC50 values ​​above 5, 5 to 5.5, above 5.5, or 5.5 to 6.

[1372] These data show that, unlike comparative compound C1, the presence of an amide-substituted thiophene or benzothiophene ring in the compounds of the present invention results in an unexpectedly and notably significant increase in intracellular inhibitory activity against NaV 1.7 and increases the difference in intracellular and extracellular inhibition of NaV 1.7 (reported as DELTA in Table 4).

[1373] Additionally, as noted in the same section of the Experimental section, the reported data show that, in contrast to the comparative compound C2, the presence of an amide-substituted thiophene ring replacing a different heteroaryl ring in the compounds of the present invention, particularly in Example 7, unexpectedly and notably determined a significant increase in intracellular inhibitory activity against the NaV 1.7 receptor and increased the difference in intracellular and extracellular inhibition of NaV 1.7 (DELTA).

[1374] hERG Assay Protocol

[1375] The effect of the test substance on the tail current of the human ether-à-go-go-related gene (hERG) was evaluated using a QPatch device (Sophion Bioscience A / S, Ballerup, Denmark) in an automated patch-clamp protocol.

[1376] Cell Culture

[1377] hERG was expressed in HEK293 cells under tetracycline induction.

[1378] Cells were cultured in the following: minimum essential medium (MEM) supplemented with 10% heat-inactivated fetal bovine serum, 1% non-essential amino acids, 1% sodium pyruvate, 2 mM L-glutamine, 1% penicillin-streptomycin, 15 µg / ml isoprothiolane, and 100 µg / ml hygromycin. hERG cells were induced with 10 µg / ml tetracycline for 24 to 72 hours prior to recording.

[1379] Prior to the experiment, cells were washed with calcium- and magnesium-free Dulbecco phosphate-buffered saline (DPBS), removed with TrypLE (pages 105 / 106, CN 122180674 A), and then resuspended and cell aggregates were broken up in serum-free medium containing 25 mM Hepes and soybean trypsin inhibitor. Cells were counted and the final concentration was set at 4–6 million cells per mL. All chemicals used for cell culture were from Life Technologies Italia.Srl, Monza, Italy.

[1380] Patch-clamp solution & drugs

[1381] The intracellular solution contained the following (mM): KCl 130, MgCl2 1, ethylene glycol di(β-aminoethyl ether)-N,N,N′,N′-tetraacetic acid (EGTA) 5, MgATP 5, N-2-hydroxyethylpiperazine-N′-2-ethanesulfonic acid (HEPES) 10, adjusted to pH 7.2 with 1 M KOH.

[1382] The external solution contained the following (mM): NaCl 137; KCl 4; CaCl2 1.8; MgCl2 1; d-glucose 10; HEPES 10; adjusted to pH 7.4 with NaOH.

[1383] All chemicals used in the patch-clamp solution were from Merck Life Science Srl, Milano, Italy.

[1384] To evaluate the degree of external blockade, the compound was first dissolved in DMSO and then diluted in an extracellular buffer to achieve final experimental concentrations of 0.6 μM, 6 μM, and 60 μM in 0.3% DMSO.

[1385] Voltage Program

[1386] A single-well QPlate chip (Sophion Bioscience, Denmark) was used for voltage clamp experiments. The membrane potential was held at -80 mV before the voltage program began. The voltage program consisted of the following steps in sequence: -50 mV for 200 ms, +20 mV for 4.8 s, -50 mV for 5 s, and then back to the holding potential. The hERG tail current was measured during the repolarization step from +20 mV to -50 mV. Each voltage program scan lasted 15 s and was continuously repeated at a frequency of 0.066 Hz during the experiment.

[1387] Liquid Procedure

[1388] The external solution and then the medium solution (equivalent to the external solution containing 0.3% DMSO) were each applied for 3 minutes and 45 seconds to achieve stable current recording. Then, three test compounds at increased concentrations (0.6, 6, and 60 μM, in 0.3% DMSO) were each applied for 5 minutes.

[1389] Data Analysis

[1390] The mean tail current was measured from the last four voltage scans during each compound application procedure for each concentration in each cell using Sophion Analyzer software (Sophion Bioscience, Denmark) and used to calculate % inhibition by comparison with the medium pretreatment group. The IC50 value was estimated using the Hill fitting formula in the Sophion Analyzer software. Manual 106 / 106 pages 114 CN 122180674 A

Claims

1. Compound of formula (I) (I) At least one of Y and Z is S, and the other is CR4; R1 and R2 are independently -(C1-C6)alkyl or fused to form -(C3-C7)heteroalkyl, wherein the -(C3-C7)heteroalkyl is optionally substituted by one or more groups selected from halogens, -(C1-C6)alkyl and -(C1-C6)alkyl-OR8; R3 and R4 are independently H or -(C1-C6)alkyl, or, in the case that Y is CR4, R3 and R4 are fused to form an aryl group; R5 and R6 are independently H or selected from -(C1-C6)alkyl, -(C1-C6)aminoalkyl, -(C1-C6)alkyl-OR8, -(C1-C6)alkyl-NR7R8, -(C1-C4)alkyl-NR7-C(O)OR8, -(C1-C4)alkyl-(C3-C7)heterocyclicalkyl and -(C3-C7)heterocyclicalkyl, or fused to form -(C3-C7)heterocyclicalkyl, wherein the -(C1-C4)alkyl-(C3-C7)heterocyclicalkyl, -(C3-C7)heterocyclicalkyl, -(C1-C6)aminoalkyl or -(C1-C6)alkyl-NR7R8 is optionally substituted by one or more groups selected from -(C1-C6)alkyl, -C(O)OR8 and -SO2R7; R7 and R8 are independently H or selected from -(C1-C6)alkyl, -(C3-C6)cycloalkyl and -(C1-C6)haloalkyl, or fused to form -(C3-C7)heteroalkyl; L1 is a bond or selected from -(C1-C6)alkylene- and -(C1-C6)cycloalkylene-; and A is a ring selected from aryl and heteroaryl, wherein any aryl or heteroaryl is optionally substituted by one or more groups selected from -OR8, -C(O)OR8, -C(O)NR7R8 and -(C1-C6)alkyl; provided that any aryl is not substituted by -(C1-C6)alkyl when L1 is a bond. It exists in zwitterionic or pharmaceutically acceptable salt form.

2. The compound of formula (I) according to claim 1, wherein Z, Y, R1, R2, R3, R4, R5, R6, R7 and R8 are as defined in claim 1; L1 is selected from -(C1-C6)alkylene- and -(C1-C6)cycloalkylene-; and A is a ring selected from aryl and heteroaryl, wherein any of the aryl or heteroaryl groups are optionally substituted by one or more groups selected from -OR8, -C(O)OR8, -C(O)NR7R8 and -(C1-C4)alkyl; It exists in zwitterionic or pharmaceutically acceptable salt form.

3. The compound of formula (I) according to claim 1, wherein L1 is a bond, represented by formula (Ia). (Ia) in Z, Y, R1, R2, R3, R4, R5, R6, R7, and R8 as defined in claim 1; and A is a ring selected from aryl and heteroaryl, wherein any of the aryl groups are optionally substituted by one or more groups selected from -OR8, -C(O)OR8 and -C(O)NR7R8, and wherein any of the heteroaryl groups are optionally substituted by one or more groups selected from -OR8, -C(O)OR8, -C(O)NR7R8 and -(C1-C6)alkyl; It exists in zwitterionic or pharmaceutically acceptable salt form.

4. The compound according to any one of the preceding claims, wherein R1 and R2 are independently -(C1-C4)alkyl or fused to -(C3-C7)heteroalkyl, wherein the -(C3-C7)heteroalkyl is optionally substituted by one or more groups selected from halogens, -(C1-C4)alkyl and -(C1-C4)alkyl-OR8; R3 and R4 are independently H or -(C1-C6)alkyl, or, in the case that Y is CR4, R3 and R4 are fused to form an aryl group; R5 and R6 are independently H or selected from -(C1-C4)alkyl, -(C1-C4)aminoalkyl, -(C1-C4)alkyl-OR8, -(C1-C4)alkyl-NR7R8, -(C1-C4)alkyl-NR7-C(O)OR8, -(C1-C4)alkyl-(C3-C7)heterocyclicalkyl, -(C3-C7)heterocyclicalkyl, or fused to form -(C3-C7)heterocyclicalkyl, wherein the -(C1-C4)alkyl-(C3-C7)heterocyclicalkyl, -(C3-C7)heterocyclicalkyl, -(C1-C4)aminoalkyl or -(C1-C4)alkyl-NR7R8 is optionally substituted by one or more groups selected from -(C1-C4)alkyl, -C(O)OR8 and -SO2R7; and R7 and R8 are independently H or selected from -(C1-C4)alkyl, -(C3-C6)cycloalkyl and -(C1-C4)haloalkyl, or fused to form -(C6-C7)heteroalkyl; It exists in zwitterionic or pharmaceutically acceptable salt form.

5. The compound according to any one of the preceding claims, wherein R1 and R2 are fused together to form a -(C6-C7)heterocyclic alkyl group, wherein the -(C6-C7)heterocyclic alkyl group is optionally substituted by one or more groups selected from -OR8, -(C1-C4)alkyl and -(C3-C7)heterocyclic alkyl; It exists in zwitterionic or pharmaceutically acceptable salt form.

6. The compound according to any one of the preceding claims, wherein R3 and R4 are independently H or -(C1-C4) alkyl; It exists in zwitterionic or pharmaceutically acceptable salt form.

7. A compound according to any one of the preceding claims, wherein Z is S and Y is CH, and R3 is H or methyl; It exists in zwitterionic or pharmaceutically acceptable salt form.

8. The compound according to any one of the preceding claims, wherein A is a heteroaryl group, optionally substituted by one or more groups selected from -OR8, -C(O)OR8, -C(O)NR7R8 and -(C1-C6)alkyl; It exists in zwitterionic or pharmaceutically acceptable salt form.

9. The compound according to any claim 1, selected from: 2-(benzylamino)-N,N-dimethyl-N-(2-((4-methyl-2-(pyrrolidine-1-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)-2-oxoethyl-1-ammonium (Example 1); 2-(benzylamino)-N-(2-((2-(dimethylcarbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-N,N-dimethyl-2-oxoethyl-1-ammonium (Example 2); 2-(benzylamino)-N-(2-((2-(tert-butylcarbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)-N,N-dimethyl-2-oxoethyl-1-ammonium (Example 3); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(morpholin-4-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 4); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(methylcarbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 5); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(pyrrolidine-1-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 6); 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(pyrrolidine-1-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 7); 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(morpholin-4-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 8); 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(methylcarbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium, (Example 9); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-(dimethylcarbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 10); 1-(2-((2-(dimethylcarbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-(isoxazo-3-ylamino)-2-oxoethyl)azacycloheptane-1-onium (Example 11); 1-(2-((4-methyl-2-(methylcarbamoyl)thiophen-3-yl)amino)-2-oxoethyl)-1-(2-((4-methylisoxazol-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 12); 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((2-((2-methoxyethyl)carbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)piperidine-1-onium (Example 13); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-(dimethylcarbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)piperidine-1-onium (Example 14); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-((2-((tert-butoxycarbonyl)(methyl)amino)ethyl)carbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 15); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-((2-methoxyethyl)carbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 16); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-((2-methoxyethyl)carbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)piperidine-1-onium (Example 17); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-((2-hydroxyethyl)carbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 18); 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((4-methyl-2-((2-(methylamino)ethyl)carbamoyl)thiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 19); 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((2-((2-methoxyethyl)carbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 20); 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(oxetane-3-ylcarbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 21); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-((2-(methylamino)ethyl)carbamoyl)thiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 22); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(oxecyclobutane-3-ylcarbamoyl)thiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 23); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-((2-hydroxyethyl)carbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)piperidine-1-onium (Example 24); 1-(2-((2-((2-hydroxyethyl)carbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)-1-(2-(isoxazo-3-ylamino)-2-oxoethyl)azacycloheptane-1-onium (Example 25); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-((2-(methylamino)ethyl)carbamoyl)thiophene-3-yl)amino)-2-oxoethyl)piperidine-1-onium (Example 26); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(4-methylpiperazin-1-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 27); 1-(2-((2-(dimethylcarbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((4-methylisoxazol-3-yl)amino)-2-oxoethyl)piperidine-1-onium (Example 28); 1-(2-((2-((2-methoxyethyl)carbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)-1-(2-((4-methylisoxazol-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 29); 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((4-methyl-2-((2-morpholinoethyl)carbamoyl)thiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 30); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-(4-(tert-butoxycarbonyl)piperazine-1-carbonyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 31); 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(4-(methanesulfonyl)piperazine-1-carbonyl)thiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 32); 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(piperazin-1-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 33); 1-(2-((2-(4-(tert-butoxycarbonyl)piperazine-1-carbonyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)-1-(2-(isoxazo-3-ylamino)-2-oxoethyl)azacycloheptane-1-onium (Example 34); 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(4-methylpiperazin-1-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 35); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(4-(methanesulfonyl)piperazin-1-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 36); 1-(2-(((2-carbamoyl-4-methylthiophen-3-yl)methyl)amino)-2-oxoethyl)-1-(2-(isoxazo-3-ylamino)-2-oxoethyl)azacycloheptane-1-onium (Example 37); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(piperazin-1-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 38); 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((2-(morpholin-4-carbonyl)benzo[b]thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 39); (R)-1-(2-(((2-carbamoyl-4-methylthiophen-3-yl)methyl)amino)-2-oxoethyl)-1-(2-oxo-2-((1-phenylethyl)amino)ethyl)azacycloheptane-1-onium (Example 40); 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((4-methyl-2-((2-(trimethylammonium)ethyl)carbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 41); 1-(2-((2-(dimethylcarbamoyl)benzo[b]thiophen-3-yl)amino)-2-oxoethyl)-1-(2-(isoxazo-3-ylamino)-2-oxoethyl)azacycloheptane-1-onium (Example 42); 1,1-Di(2-((2-(dimethylcarbamoyl)benzo[b]thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 43); 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((2-((2-methoxyethyl)(methyl)carbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 44); 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((4-methyl-2-((3-(methylamino)propyl)carbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 45); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-((2-(trimethylammonyl)ethyl)carbamoyl)thiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 46); 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((4-methyl-2-((2-(4-methylpiperazin-1-yl)ethyl)carbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 47); 1-(2-(isoxazo-3-ylamino)-2-oxoethyl)-1-(2-((4-methyl-2-((2-(piperazin-1-yl)ethyl)carbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 48); 1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((4-methyl-2-(4-methylpiperazin-1-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 49); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-((2-methoxyethyl)(methyl)carbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 50); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-((2-(piperazin-1-yl)ethyl)carbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 51); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-((2-(4-methylpiperazin-1-yl)ethyl)carbamoyl)thiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 52); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-((3-(methylamino)propyl)carbamoyl)thiophene-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 53); 1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((4-methyl-2-(morpholin-4-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 54); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-((2-hydroxyethyl)(methyl)carbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 55); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-(methyl(2-(methylamino)ethyl)carbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 56); 1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((4-methyl-2-(piperazin-1-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 57); (S)-1-(2-((2-((6-amino-1-methoxy-1-oxohexane-2-yl)carbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)-1-(2-(benzylamino)-2-oxoethyl)azacycloheptane-1-onium (Example 58); 1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((4-methyl-2-((2-(methylamino)ethyl)carbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 59); 4-Fluoro-1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-4-methyl-1-(2-((4-methyl-2-((2-(methylamino)ethyl)carbamoyl)thiophen-3-yl)amino)-2-oxoethyl)piperidine-1-onium (Example 60); 1-(2-((4-methyl-2-((2-(methylamino)ethyl)carbamoyl)thiophene-3-yl)amino)-2-oxoethyl)-1-(2-oxo-2-((2-(trifluoromethoxy)benzyl)amino)ethyl)azacycloheptane-1-onium (Example 61); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-methyl-2-((piperidin-4-ylmethyl)carbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 62); 1-(2-((2-((2-(cyclopropylamino)ethyl)carbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 63); 1-(2-((2-(di(2-hydroxyethyl)carbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 64); 1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((4-methyl-2-(piperidin-4-ylcarbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 65); 1-(2-((2-(dimethylcarbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 66); 1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-4,4-dimethyl-1-(2-((4-methyl-2-(methylcarbamoyl)thiophen-3-yl)amino)-2-oxoethyl)piperidine-1-onium (Example 67); 1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((4-methyl-2-(methylcarbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 68); 1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((4-methyl-2-(methylcarbamoyl)thiophen-3-yl)amino)-2-oxoethyl)piperidine-1-onium (Example 69); 1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((4-methyl-2-(oxetane-3-ylcarbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 70); 1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-4,4-dimethyl-1-(2-((4-methyl-2-((2-(methylamino)ethyl)carbamoyl)thiophen-3-yl)amino)-2-oxoethyl)piperidine-1-onium (Example 71); 1-(2-((2-(ethoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-4,4-dimethyl-1-(2-((4-methyl-2-((3-(methylamino)propyl)carbamoyl)thiophen-3-yl)amino)-2-oxoethyl)piperidine-1-onium (Example 72); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-(dimethylcarbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-4,4-di(hydroxymethyl)piperidine-1-onium (Example 73); (2-(4,4-dimethyl-1-(2-((4-methyl-2-(methylcarbamoyl)thiophen-3-yl)amino)-2-oxoethyl)piperidin-1-onthiol-1-yl)acetyl)(isoxazo-3-yl)amide (Example 74); 1-(2-((2-(dimethylcarbamoyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-4,4-di(hydroxymethyl)-1-(2-oxo-2-((1-phenylcyclopropyl)amino)ethyl)piperidine-1-onium (Example 75); 1-(2-((2-(methoxycarbonyl)-4-methylthiophen-3-yl)amino)-2-oxoethyl)-1-(2-((4-methyl-2-((piperidin-4-ylmethyl)carbamoyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 76); and (R)-1-(2-((2-((6-amino-1-methoxy-1-oxohexane-2-yl)carbamoyl)-4-methylthiophene-3-yl)amino)-2-oxoethyl)-1-(2-(benzylamino)-2-oxoethyl)azacycloheptane-1-onium (Example 77); 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-methyl-4-(morpholin-4-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 78) 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((2-methyl-4-(pyrrolidine-1-carbonyl)thiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 79) 1-(2-(benzylamino)-2-oxoethyl)-1-(2-((4-((2-methoxyethyl)(methyl)carbamoyl)-2-methylthiophen-3-yl)amino)-2-oxoethyl)azacycloheptane-1-onium (Example 80) It exists in zwitterionic or pharmaceutically acceptable salt form.

10. Use of an intermediate compound selected from compounds (V), (VIII), (X), (XI), (XV), and (XVI) for the preparation of a compound of formula (I) according to any one of claims 1 to 9. Wherein R1, R2, R3, R4, R5, L1 and A are as defined in claims 1 to 9, X2 is a halogen, preferably chlorine or bromine, and PG is a -(C1-C6) alkyl group.

11. A pharmaceutical composition comprising a compound according to any one of claims 1 to 9 and one or more pharmaceutically acceptable carriers or excipients mixed therewith.

12. The pharmaceutical composition of claim 11, formulated for administration by inhalation.

13. The compound of formula (I) according to any one of claims 1-9 or the pharmaceutical composition according to claim 11 or 12, used as a medicine.

14. The compound or pharmaceutical composition of formula (I) according to claim 13, for the prevention and / or treatment of diseases, disorders or conditions related to sodium channel receptor mechanisms.

15. The compound or pharmaceutical composition of formula (I) according to claim 13 or 14, for the prevention and / or treatment of respiratory diseases selected from the following: cough, subacute or chronic cough, treatment-resistant cough, intractable chronic cough, idiopathic chronic cough, post-viral cough, iatrogenic cough, asthma, idiopathic pulmonary fibrosis (IPF), chronic obstructive pulmonary disease (COPD), and cough associated with respiratory diseases such as COPD, asthma, and bronchospasm.

16. The compound or pharmaceutical composition of formula (I) according to claim 15, for the prevention and / or treatment of chronic cough.