SYNTHESIS OF A THIOSULFONIC ACID BY A STEP OF PERIODATE MEDIATED OXIDATIVE COUPLING OF A THIOSULFONIC ACID WITH AN ANILINE

Abstract

The present invention pertains generally to the field of chemical synthesis, and more particularly to methods for the chemical synthesis of a thiosulfonic acid of Formula (1) by a step of periodate mediated oxidative coupling of a thiosulfonic acid of Formula (2) with an aniline of Formula (3), as described herein. The present invention also relates to such methods which incorporate one or more additional (subsequent and/or preceding) steps, for example, to prepare compounds of Formula (5) from compounds of Formula (1); to prepare compounds of Formula (6) from compounds of Formula (5); and to prepare compounds of Formula (2) from compounds of Formula (4), as described herein.

##STR00001##

Claims

1-126. (canceled)

127. A method of chemical synthesis of a compound of Formula (1): ##STR00039## comprising a step of periodate mediated oxidative coupling, in which a compound of Formula (2): ##STR00040## is reacted with a compound of Formula (3): ##STR00041## and a periodate oxidising agent; to form said compound of Formula (1); wherein: R.sup.1A1 is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; halogenated C.sub.1-4alkyl; C.sub.5-10aryl; halogenated C.sub.5-10aryl; C.sub.5-10aryl-C.sub.1-4alkyl; or halogenated C.sub.5-10aryl-C.sub.1-4alkyl; R.sup.1A2 is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; halogenated C.sub.1-4alkyl; C.sub.5-10aryl; halogenated C.sub.5-10aryl; C.sub.5-10aryl-C.sub.1-4alkyl; or halogenated C.sub.5-10aryl-C.sub.1-4alkyl; R.sup.1B1 is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; halogenated C.sub.1-4alkyl; C.sub.5-10aryl; halogenated C.sub.5-10aryl; C.sub.5-10aryl-C.sub.1-4alkyl; or halogenated C.sub.5-10aryl-C.sub.1-4alkyl; R.sup.1B2 is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; halogenated C.sub.1-4alkyl; C.sub.5-10aryl; halogenated C.sub.5-10aryl; C.sub.5-10aryl-C.sub.1-4alkyl; or halogenated C.sub.5-10aryl-C.sub.1-4alkyl; R.sup.2A is independently H or R.sup.3AA; R.sup.2AA is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; halogenated C.sub.1-4alkyl; C.sub.5-10aryl; halogenated C.sub.5-10aryl; C.sub.5-10aryl-C.sub.1-4alkyl; or halogenated C.sub.5-10aryl-C.sub.1-4alkyl; R.sup.2B is independently H or R.sup.3AA; R.sup.2BB is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; halogenated C.sub.1-4alkyl; C.sub.5-10aryl; halogenated C.sub.5-10aryl; C.sub.5-10aryl-C.sub.1-4alkyl; or halogenated C.sub.5-10aryl-C.sub.1-4alkyl; R.sup.3A is independently H or R.sup.3AA; R.sup.3AA is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; halogenated C.sub.1-4alkyl; C.sub.5-10aryl; halogenated C.sub.5-10aryl; C.sub.5-10aryl-C.sub.1-4alkyl; or halogenated C.sub.5-10aryl-C.sub.1-4alkyl; R.sup.3B is independently H or R.sup.3BB; R.sup.3BB is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; halogenated C.sub.1-4alkyl; C.sub.5-10aryl; halogenated C.sub.5-10aryl; C.sub.5-10aryl-C.sub.1-4alkyl; or halogenated C.sub.5-10aryl-C.sub.1-4alkyl; R.sup.4A is independently H or R.sup.4AA; R.sup.4AA is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; halogenated C.sub.1-4alkyl; C.sub.5-10aryl; halogenated C.sub.5-10aryl; C.sub.5-10aryl-C.sub.1-4alkyl; or halogenated C.sub.5-10aryl-C.sub.1-4alkyl; R.sup.4B is independently H or R.sup.4BB; and R.sup.4BB is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; halogenated C.sub.1-4alkyl; C.sub.5-10aryl; halogenated C.sub.5-10aryl; C.sub.5-10aryl-C.sub.1-4alkyl; or halogenated C.sub.5-10aryl-C.sub.1-4alkyl.

128. A method according to claim 127, wherein each R.sup.1A1is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; or halogenated C.sub.1-4alkyl preferably wherein R.sup.1A1 is independently Me or Et.

129. A method according to claim 127, wherein R.sup.1A2 is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; or halogenated C.sub.1-4alkyl preferably wherein R.sup.1A2 is independently Me or Et.

130. A method according to claim 127, wherein R.sup.1B1 is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; or halogenated C.sub.1-4alkyl preferably wherein R.sup.1B1 is independently Me or Et.

131. A method according to claim 127, wherein R.sup.1B2 is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; or halogenated C.sub.1-4alkyl preferably wherein R.sup.1B2 is independently Me or Et.

132. A method according to claim 127, wherein R.sup.2AA, if present, is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; or halogenated C.sub.1-4alkyl, preferably wherein R.sup.2AA, if present, is independently Me or Et.

133. A method according to claim 127, wherein R.sup.2BB, if present, is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; or halogenated C.sub.1-4alkyl preferably wherein R.sup.2BB, if present, is independently Me or Et.

134. A method according to claim 127, wherein R.sup.3AA, if present, is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; or halogenated C.sub.1-4alkyl preferably wherein R.sup.3AA, if present, is independently Me or Et.

135. A method according to claim 127, wherein R.sup.3BB, if present, is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; or halogenated C.sub.1-4alkyl preferably wherein R.sup.3BB, if present, is independently Me or Et.

136. A method according to claim 127, wherein R.sup.4AA, if present, is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; or halogenated C.sub.1-4alkyl preferably wherein R.sup.4AA, if present, is independently Me or Et.

137. A method according to claim 127, wherein R.sup.4BB, if present, is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; or halogenated C.sub.1-4alkyl preferably wherein R.sup.4BB, if present, is independently Me or Et.

138. A method according to claim 127, wherein the periodate oxidizing agent is iodine peroxide, periodic acid or a periodate salt.

139. A method according to claim 138, wherein the periodate oxidizing agent is an alkali metal periodate salt such as a sodium periodate salt.

140. A method according to claim 127, wherein the ratio, A, of the amount of compound of Formula (2), in equivalents, to the amount of compound of Formula (3), in equivalents, is from about 0.5 to about 3.0.

141. A method according to claims 127, wherein the ratio, B, of the amount of compound of Formula (2), in equivalents, to the amount of compound of periodate oxidizing agent, in equivalents, is from about 0.5 to about 3.0.

142. A method according to claim 127, wherein the reaction is carried out in the presence of an acid such as sulfuric acid.

143. A method according to claim 142, wherein the ratio, C, of the amount of compound of Formula (2), in equivalents, to the amount of H.sup.30 provided by the acid, in equivalents, is from about 0.5 to about 3.0.

144. A method according to claim 127, wherein the reaction is carried out in the presence of water.

145. A method according to claim 127, wherein the method further comprises a preceding step of: converting a compound of Formula (4): ##STR00042## to the corresponding compound of Formula (2): ##STR00043##

146. A method according to claim 127, wherein the method further comprises a subsequent step of: converting the compound of Formula (1): ##STR00044## to the corresponding compound of Formula (5): ##STR00045## wherein X.sup. is one or more anionic counter ions to achieve electrical neutrality preferably wherein X.sup. is independently Cl.sup..

147. A method according to claim 146, wherein the method further comprises a subsequent step of: converting the compound of Formula (5): ##STR00046## to the corresponding compound of Formula (6): ##STR00047## wherein Y.sup. and Z.sup., taken together, are one or more anionic counter ions to achieve electrical neutrality, preferably wherein each of Y.sup. and Z.sup. is independently Cl.sup..

148. A compound of Formula (1) obtained by a method according to claim 127.

149. A compound of Formula (5) obtained by a method according to claim 146.

Description

BRIEF DESCRIPTON OF THE DRAWINGS

[0079] FIG. 1 shows the .sup.1H NMR (300 MHz, DMSO-d.sub.6) spectrum for thiosulphonic acid S-(2-amino-5-dimethyl amino) phenyl ester obtained in Method 1.

[0080] FIG. 2 shows the .sup.1H NMR (300 MHz, DMSO-d.sub.6) spectrum for a mixture of thiosulphonic acid S-(2-amino-5-dimethyl amino) phenyl ester obtained in Method 1 and the reference compound 3-(trimethylsilyl)-1-propanesulfonic acid.

[0081] FIG. 3 shows the .sup.1H NMR (300 MHz, DMSO-d.sub.6) spectrum for (4-(2-(thiosulfate)-4-(dimethylamino)-phenyl-imino)-cyclohex-2,5-dienylidene)-N,N-dimethyl ammonium obtained in Method 2B.

[0082] FIG. 4 shows the .sup.1H NMR (300 MHz, D.sub.2O) spectrum for methylthioninium chloride (MTC) obtained in Method 3.

DETAILED DESCRIPTION OF THE INVENTION

[0083] The present invention relates to methods for chemical synthesis which include the step of preparing a thiosulfonic acid of Formula (1) by a step of periodate mediated oxidative coupling of a thiosulfonic acid of Formula (2) and an aniline of Formula (3).

[0084] The present invention also relates to such methods which incorporate one or more additional (subsequent and/or preceding) steps, for example: to prepare compounds of Formula (5) from compounds of Formula (1); to prepare compounds of Formula (6) from compounds of Formula (5); and to prepare compounds of Formula (2) from compounds of Formula (4).

[0085] These methods, and method steps, are illustrated in the following scheme.

##STR00016##

[0086] Accordingly, one aspect of the invention is a method of chemical synthesis of a compound of Formula (1):

##STR00017##

comprising a step of periodate mediated oxidative coupling, in which a compound of Formula (2):

##STR00018##

is reacted with a compound of Formula (3):

##STR00019##

and a periodate oxidising agent;
to form said compound of Formula (1); wherein:

[0087] R.sup.1A1 is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; halogenated C.sub.1-4alkyl; C.sub.5-10aryl; halogenated C.sub.5-10aryl; C.sub.5-10aryl-C.sub.1-4alkyl; or halogenated C.sub.5-10aryl-C.sub.1-4alkyl;

[0088] R.sup.1A2 is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; halogenated C.sub.1-4alkyl; C.sub.5-10aryl; halogenated C.sub.5-10aryl; C.sub.5-10aryl-C.sub.1-4alkyl; or halogenated C.sub.5-10aryl-C.sub.1-4alkyl;

[0089] R.sup.1B1 is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; halogenated C.sub.1-4alkyl; C.sub.5-10aryl; halogenated C.sub.5-10aryl; C.sub.5-10aryl-C.sub.1-4alkyl; or halogenated C.sub.5-10aryl-C.sub.1-4alkyl;

[0090] R.sup.1B2 is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; halogenated C.sub.1-4alkyl; C.sub.5-10aryl; halogenated C.sub.5-10aryl; C.sub.5-10aryl-C.sub.1-4alkyl; or halogenated C.sub.5-10aryl-C.sub.1-4alkyl;

[0091] R.sup.2A is independently H or R.sup.3AA;

[0092] R.sup.2AA is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; halogenated C.sub.1-4alkyl; C.sub.5-10aryl; halogenated C.sub.5-10aryl; C.sub.5-10aryl-C.sub.1-4alkyl; or halogenated C.sub.5-10aryl-C.sub.1-4alkyl;

[0093] R.sup.2B is independently H or R.sup.3AA;

[0094] R.sup.2BB is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; halogenated C.sub.1-4alkyl; C.sub.5-10aryl; halogenated C.sub.5-10aryl; C.sub.5-10aryl-C.sub.1-4alkyl; or halogenated C.sub.5-10aryl-C.sub.1-4alkyl;

[0095] R.sup.3A is independently H or R.sup.3AA;

[0096] R.sup.3AA is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; halogenated C.sub.1-4alkyl; C.sub.5-10aryl; halogenated C.sub.5-10aryl; C.sub.5-10aryl-C.sub.1-4alkyl; or halogenated C.sub.5-10aryl-C.sub.1-4alkyl;

[0097] R.sup.3B is independently H or R.sup.3BB;

[0098] R.sup.3BB is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; halogenated C.sub.1-4alkyl; C.sub.5-10aryl; halogenated C.sub.5-10aryl; C.sub.5-10aryl-C.sub.1-4alkyl; or halogenated C.sub.5-10aryl-C.sub.1-4alkyl;

[0099] R.sup.4A is independently H or R.sup.4AA;

[0100] R.sup.4AA is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; halogenated C.sub.1-4alkyl; C.sub.5-10aryl; halogenated C.sub.5-10aryl; C.sub.5-10aryl-C.sub.1-4alkyl; or halogenated C.sub.5-10aryl-C.sub.1-4alkyl;

[0101] R.sup.4B is independently H or R.sup.4BB; and

[0102] R.sup.4BB is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; halogenated C.sub.1-4alkyl; C.sub.5-10aryl; halogenated C.sub.5-10aryl; C.sub.5-10aryl-C.sub.1-4alkyl; or halogenated C.sub.5-10aryl-C.sub.1-4alkyl.

[0103] Alkyl Groups

[0104] In one embodiment, the C.sub.1-4alkyl groups are selected from: linear C.sub.1-4alkyl groups, such as Me, Et, nPr, iPr, and nBu; branched C.sub.3-4alkyl groups, such as iPr, iBu, sBu, and tBu; and cyclic C.sub.3-4alkyl groups, such as cPr and cBu.

[0105] Alkenyl Groups

[0106] In one embodiment, the C.sub.2-4alkenyl groups are selected from linear C.sub.1-4alkenyl groups, such as CHCH.sub.2 (vinyl) and CH.sub.2CHCH.sub.2 (allyl).

[0107] Halogenated Alkyl Groups

[0108] In one embodiment, the halogenated C.sub.1-4alkyl groups are selected from: CF.sub.3, CH.sub.2CF.sub.3, CH.sub.2CH.sub.2F, and CF.sub.2CF.sub.3.

[0109] Aryl Groups

[0110] In one embodiment, the C.sub.5-10aryl groups are selected from: C.sub.6-10carboaryl groups, such as phenyl and napthyl; and C.sub.5-10heteroaryl groups, such as thienyl, imidazolyl, pyrazolyl, triazolyl, oxazolyl, isoxazolyl, thiazolyl, isothiazolyl, pyridyl, pyrimidinyl, and quinolinyl.

[0111] Halogenated Aryl Groups

[0112] In one embodiment, the halogenated C.sub.5-10aryl groups are selected from: halogenated C.sub.6-10carboaryl groups, such as 4-fluoro-phenyl, 3-fluoro-phenyl, and 2-fluoro-phenyl, and halogenated C.sub.5-10heteroaryl groups.

[0113] Aryl-Alkyl Groups

[0114] In one embodiment, the C.sub.5-10aryl-C.sub.1-4alkyl groups are selected from: benzyl and phenethyl.

[0115] Halogenated Aryl-Alkyl Groups

[0116] In one embodiment, the halogenated C.sub.5-10aryl-C.sub.1-4alkyl groups are selected from: halogenated C.sub.5-10carboaryl-C.sub.1-4alkyl groups, such as 4-fluoro-benzyl, 3-fluoro-benzyl, and 2-fluoro-benzyl, and halogenated C.sub.5-10heteroaryl-C.sub.1-4alkyl groups.

[0117] The Group R.sup.1A1

[0118] In one embodiment, R.sup.1A1 is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; or halogenated C.sub.1-4alkyl.

[0119] In one embodiment, R.sup.1A1 is independently Me, Et, nPr, nBu, CH.sub.2CHCH.sub.2, or CF.sub.3.

[0120] In one embodiment, R.sup.1A1 is independently C.sub.1-4alkyl; or halogenated C.sub.1-4alkyl.

[0121] In one embodiment, R.sup.1A1 is independently Me, Et, or CF.sub.3.

[0122] In one embodiment, R.sup.1A1 is independently C.sub.1-4alkyl.

[0123] In one embodiment, R.sup.1A1 is independently Me or Et.

[0124] In one embodiment, R.sup.1A1 is independently Me.

[0125] In one embodiment, R.sup.1A1 is independently Et.

[0126] The Group R.sup.1A2

[0127] In one embodiment, R.sup.1A2 is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; or halogenated C.sub.1-4alkyl.

[0128] In one embodiment, R.sup.1A2 is independently Me, Et, nPr, nBu, CH.sub.2CHCH.sub.2, or CF.sub.3.

[0129] In one embodiment, R.sup.1A2 is independently C.sub.1-4alkyl; or halogenated C.sub.1-4alkyl.

[0130] In one embodiment, R.sup.1A2 is independently Me, Et, or CF.sub.3.

[0131] In one embodiment, R.sup.1A2 is independently C.sub.1-4alkyl.

[0132] In one embodiment, R.sup.1A2 is independently Me or Et.

[0133] In one embodiment, R.sup.1A2 is independently Me.

[0134] In one embodiment, R.sup.1A2 is independently Et.

[0135] The Groups R.sup.1A1 and R.sup.1A2

[0136] In one embodiment, R.sup.1A1 and R.sup.1A2 are the same.

[0137] In one embodiment, R.sup.1A1 and R.sup.1A2 are different.

[0138] The Group R.sup.1B1

[0139] In one embodiment, R.sup.1B1 is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; or halogenated C.sub.1-4alkyl.

[0140] In one embodiment, R.sup.1B1 is independently Me, Et, nPr, nBu, CH.sub.2CHCH.sub.2, or CF.sub.3.

[0141] In one embodiment, R.sup.1B1 is independently C.sub.1-4alkyl; or halogenated C.sub.1-4alkyl.

[0142] In one embodiment, R.sup.1B1 is independently Me, Et, or CF.sub.3.

[0143] In one embodiment, R.sup.1B1 is independently C.sub.1-4alkyl.

[0144] In one embodiment, R.sup.1B1 is independently Me or Et.

[0145] In one embodiment, R.sup.1B1 is independently Me.

[0146] In one embodiment, R.sup.1B1 is independently Et.

[0147] The Group R.sup.1B2

[0148] In one embodiment, R.sup.1B2 is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; or halogenated C.sub.1-4alkyl.

[0149] In one embodiment, R.sup.1B2 is independently Me, Et, nPr, nBu, CH.sub.2CHCH.sub.2, or CF.sub.3.

[0150] In one embodiment, R.sup.1B2 is independently C.sub.1-4alkyl; or halogenated C.sub.1-4alkyl.

[0151] In one embodiment, R.sup.1B2 is independently Me, Et, or CF.sub.3.

[0152] In one embodiment, R.sup.1B2 is independently C.sub.1-4alkyl.

[0153] In one embodiment, R.sup.1B2 is independently Me or Et.

[0154] In one embodiment, R.sup.1B2 is independently Me.

[0155] In one embodiment, R.sup.1B2 is independently Et.

[0156] The Groups R.sup.1B1 and R.sup.1B2

[0157] In one embodiment, R.sup.1B1 and R.sup.1B2 are the same.

[0158] In one embodiment, R.sup.1B1 and R.sup.1B2 are different.

[0159] The Groups R.sup.1A1, R.sup.1A2, R.sup.1B1, and R.sup.1B2

[0160] In one embodiment, R.sup.1A1, R.sup.1A2, R.sup.1B1, and R.sup.1B2 are the same.

[0161] The Group R.sup.2A

[0162] In one embodiment, R.sup.2A is independently H.

[0163] In one embodiment, R.sup.2A is independently R.sup.2AA.

[0164] The Group R.sup.2AA

[0165] In one embodiment, R.sup.2AA, if present, is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; or halogenated C.sub.1-4alkyl.

[0166] In one embodiment, R.sup.2AA, if present, is independently C.sub.1-4alkyl; or halogenated C.sub.1-4alkyl.

[0167] In one embodiment, R.sup.2AA, if present, is independently Me, Et, or CF.sub.3.

[0168] In one embodiment, R.sup.2AA, if present, is independently C.sub.1-4alkyl.

[0169] In one embodiment, R.sup.2AA, if present, is independently Me or Et.

[0170] In one embodiment, R.sup.2AA, if present, is independently Me.

[0171] In one embodiment, R.sup.2AA, if present, is independently Et.

[0172] The Group R.sup.2B

[0173] In one embodiment, R.sup.2B is independently H.

[0174] In one embodiment, R.sup.2B is independently R.sup.2BB.

[0175] The Group R.sup.2BB

[0176] In one embodiment, R.sup.2BB, if present, is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; or halogenated C.sub.1-4alkyl.

[0177] In one embodiment, R.sup.2BB, if present, is independently C.sub.1-4alkyl; or halogenated C.sub.1-4alkyl.

[0178] In one embodiment, R.sup.2BB, if present, is independently Me, Et, or CF.sub.3.

[0179] In one embodiment, R.sup.2BB, if present, is independently C.sub.1-4alkyl.

[0180] In one embodiment, R.sup.2BB, if present, is independently Me or Et.

[0181] In one embodiment, R.sup.2BB, if present, is independently Me.

[0182] In one embodiment, R.sup.2BB, if present, is independently Et.

[0183] The Groups R.sup.2A and R.sup.2B

[0184] In one embodiment, R.sup.2A and R.sup.2B are the same.

[0185] In one embodiment, R.sup.2A and R.sup.2B are different.

[0186] The Group R.sup.3A

[0187] In one embodiment, R.sup.3A is independently H.

[0188] In one embodiment, R.sup.3A is independently R.sup.3AA.

[0189] The Group R.sup.3AA

[0190] In one embodiment, R.sup.3AA, if present, is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; or halogenated C.sub.1-4alkyl.

[0191] In one embodiment, R.sup.3AA, if present, is independently C.sub.1-4alkyl; or halogenated C.sub.1-4alkyl.

[0192] In one embodiment, R.sup.3AA, if present, is independently Me, Et, or CF.sub.3.

[0193] In one embodiment, R.sup.3AA, if present, is independently C.sub.1-4alkyl.

[0194] In one embodiment, R.sup.3AA, if present, is independently Me or Et.

[0195] In one embodiment, R.sup.3AA, if present, is independently Me.

[0196] In one embodiment, R.sup.3AA, if present, is independently Et.

[0197] The Group R.sup.3B

[0198] In one embodiment, R.sup.3B is independently H.

[0199] In one embodiment, R.sup.3B is independently R.sup.3BB.

[0200] The Group R.sup.3BB

[0201] In one embodiment, R.sup.3BB, if present, is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; or halogenated C.sub.1-4alkyl.

[0202] In one embodiment, R.sup.3BB, if present, is independently C.sub.1-4alkyl; or halogenated C.sub.1-4alkyl.

[0203] In one embodiment, R.sup.3BB, if present, is independently Me, Et, or CF.sub.3.

[0204] In one embodiment, R.sup.3BB, if present, is independently C.sub.1-4alkyl.

[0205] In one embodiment, R.sup.3BB, if present, is independently Me or Et.

[0206] In one embodiment, R.sup.3BB, if present, is independently Me.

[0207] In one embodiment, R.sup.3BB, if present, is independently Et.

[0208] The Groups R.sup.3A and R.sup.3B

[0209] In one embodiment, R.sup.3A and R.sup.3B are the same.

[0210] In one embodiment, R.sup.3A and R.sup.3B are different.

[0211] The Group R.sup.4A

[0212] In one embodiment, R.sup.4A is independently H.

[0213] In one embodiment, R.sup.4A is independently R.sup.4AA.

[0214] The Group R.sup.4AA

[0215] In one embodiment, R.sup.4AA, if present, is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; or halogenated C.sub.1-4alkyl.

[0216] In one embodiment, R.sup.4AA, if present, is independently C.sub.1-4alkyl; or halogenated C.sub.1-4alkyl.

[0217] In one embodiment, R.sup.4AA, if present, is independently Me, Et, or CF.sub.3.

[0218] In one embodiment, R.sup.4AA, if present, is independently C.sub.1-4alkyl.

[0219] In one embodiment, R.sup.4AA, if present, is independently Me or Et.

[0220] In one embodiment, R.sup.4AA, if present, is independently Me.

[0221] In one embodiment, R.sup.4AA, if present, is independently Et.

[0222] The Group R.sup.4B

[0223] In one embodiment, R.sup.4B is independently H.

[0224] In one embodiment, R.sup.4B is independently R.sup.4BB.

[0225] The Group R.sup.4BB

[0226] In one embodiment, R.sup.4BB, if present, is independently C.sub.1-4alkyl; C.sub.2-4alkenyl; or halogenated C.sub.1-4alkyl.

[0227] In one embodiment, R.sup.4BB, if present, is independently C.sub.1-4alkyl; or halogenated C.sub.1-4alkyl.

[0228] In one embodiment, R.sup.4BB, if present, is independently Me, Et, or CF.sub.3.

[0229] In one embodiment, R.sup.4BB, if present, is independently C.sub.1-4alkyl.

[0230] In one embodiment, R.sup.4BB, if present, is independently Me or Et.

[0231] In one embodiment, R.sup.4BB, if present, is independently Me.

[0232] In one embodiment, R.sup.4BB, if present, is independently Et.

[0233] The Groups R.sup.4A and R.sup.4B

[0234] In one embodiment, R.sup.4A and R.sup.4B are the same.

[0235] In one embodiment, R.sup.4A and R.sup.4B are different.

[0236] Combinations

[0237] It is appreciated that certain features of the invention, which are, for clarity, described in the context of separate embodiments, may also be provided in combination in a single embodiment. Conversely, various features of the invention, which are, for brevity, described in the context of a single embodiment, may also be provided separately or in any suitable sub-combination. All combinations of the embodiments pertaining to the chemical groups represented by variables (e.g., R.sup.1A1, R.sup.1A2, R.sup.1B1, R.sup.1B2, R.sup.2A, R.sup.2AA, R.sup.2B, R.sup.2BB, R.sup.3A, R.sup.3AA, R.sup.3B, R.sup.3BB, R.sup.4A, R.sup.4AA, R.sup.4B, R.sup.4BB, X.sup., Y.sup.,and Z.sup., etc.) are specifically embraced by the present invention and are disclosed herein just as if each and every combination was individually and explicitly disclosed, to the extent that such combinations embrace compounds that are stable compounds (i.e., compounds that can be isolated, characterised, and tested). In addition, all sub-combinations of the chemical groups listed in the embodiments describing such variables are also specifically embraced by the present invention and are disclosed herein just as if each and every such sub-combination of chemical groups was individually and explicitly disclosed herein.

[0238] Periodate Oxidising Agent

[0239] The periodate oxidizing agent is an iodine oxyanion that is able to facilitate periodate mediated oxidative coupling, specifically, the coupling of a compound of Formula (2) and a compound of Formula (3).

[0240] Iodine oxyanions include iodine peroxides, for example, periodic acid and periodate salts.

[0241] Periodic acid may be provided, for example, as HIO.sub.4, H.sub.4I.sub.2O.sub.9 (e.g., 2HIO.sub.4 plus H.sub.2O), or H.sub.5IO.sub.6 (e.g., HIO.sub.4 plus 2H.sub.2O).

[0242] Periodate salts include alkali metal salts, such as sodium salts, such as NaH.sub.4IO.sub.6 (e.g., NaIO.sub.4 plus 2H.sub.2O), Na.sub.2H.sub.3IO.sub.6 (e.g., NaIO.sub.4 plus NaOH plus H.sub.2O), and Na.sub.3H.sub.2IO.sub.6 (e.g., NaIO.sub.4 plus 2NaOH); potassium salts, such as KIO.sub.4; and cesium salts, such as CsIO.sub.4.

[0243] In one embodiment, the periodate oxidizing agent is an iodine peroxide.

[0244] In one embodiment, the periodate oxidizing agent is periodic acid or a periodate salt.

[0245] In one embodiment, the periodate oxidizing agent is periodic acid.

[0246] In one embodiment, the periodate oxidizing agent is a periodate salt.

[0247] In one embodiment, the periodate oxidizing agent is an alkali metal periodate salt.

[0248] In one embodiment, the periodate oxidizing agent is a sodium periodate salt.

[0249] In one embodiment, the periodate oxidizing agent is sodium periodate (NaIO.sub.4).

[0250] The Reaction Step of Periodate Mediated Oxidative Coupling

[0251] The periodate mediated oxidative coupling, specifically, the coupling of a compound of Formula (2) and a compound of Formula (3) using a periodate oxidizing agent, is performed under conditions suitable to achieve coupling to form a compound of Formula (1).

[0252] In one embodiment, the ratio, A, of the amount of compound of Formula (2), in equivalents, to the amount of compound of Formula (3), in equivalents, is from about 0.5 to about 3.0.

[0253] In one embodiment, the range is from about 0.6 to about 2.0.

[0254] In one embodiment, the range is from about 0.7 to about 1.5.

[0255] In one embodiment, the range is from about 0.8 to about 1.2.

[0256] In one embodiment, the range is from about 0.9 to about 1.1.

[0257] In one embodiment, the ratio is about 1.

[0258] As an illustration, in one example of the periodate mediated oxidative coupling, thiosulphonic acid S-(2-amino-5-dimethyl amino) phenyl ester (5.92 g, 248.32 g/mol, 23.8 mmol, 1.0 equivalent) (a compound of Formula (2)) is reacted with N,N-dimethylaniline (C.sub.6H.sub.5N(CH.sub.3).sub.2, 2.89 g, 121.18 g/mol, 23.8 mmol, 1.0 equivalent) (a compound of Formula (3)).

[0259] In another example of the periodate mediated oxidative coupling, thiosulphonic acid S-(2-amino-5-dimethyl amino) phenyl ester (10.0 g, 248.32 g/mol, 40.3 mmol, 1.0 equivalent) (a compound of Formula (2)) is reacted with N,N-dimethylaniline (C.sub.6H.sub.5N(CH.sub.3).sub.2, 4.88 g, 121.18 g/mol, 40.3 mmol, 1.0 equivalent) (a compound of Formula (3)).

[0260] In one embodiment, the ratio, B, of the amount of compound of Formula (2), in equivalents, to the amount of compound of periodate oxidizing agent, in equivalents, is from about 0.5 to about 3.0.

[0261] In one embodiment, the range is from about 1.0 to about 3.0.

[0262] In one embodiment, the range is from about 1.5 to about 2.5.

[0263] In one embodiment, the range is from about 1.8 to about 2.3.

[0264] In one embodiment, the range is from about 1.9 to about 2.2.

[0265] In one embodiment, the range is from about 2.0 to about 2.2.

[0266] In one embodiment, the ratio is about 2.1.

[0267] As an illustration, in one example of the periodate mediated oxidative coupling, thiosulphonic acid S-(2-amino-5-dimethyl amino) phenyl ester (5.92 g, 248.32 g/mol, 23.8 mmol, 1.0 equivalent) (a compound of Formula (2)) is reacted with sodium periodate (NaIO.sub.4, 10.66 g, 213.89 g/mol, 49.8 mmol, 2.09 equivalents) (a periodate oxidizing agent).

[0268] In another example of the periodate mediated oxidative coupling, thiosulphonic acid S-(2-amino-5-dimethyl amino) phenyl ester (10.0 g, 248.32 g/mol, 40.3 mmol, 1.0 equivalent) (a compound of Formula (2)) is reacted with sodium periodate (NaIO.sub.4, 17.67 g, 213.89 g/mol, 82.6 mmol, 2.05 equivalents).

[0269] In one embodiment, the reaction is carried out at a temperature of from about 0 C. to about 30 C.

[0270] In one embodiment, the range is from about 1 C. to about 20 C.

[0271] In one embodiment, the range is from about 1 C. to about 15 C.

[0272] In one embodiment, the range is from about 1 C. to about 10 C.

[0273] In one embodiment, the range is from about 1 C. to about 8 C.

[0274] In one embodiment, the range is from about 1 C. to about 8 C.

[0275] In one embodiment, the range is from about 2 C. to about 20 C.

[0276] In one embodiment, the range is from about 2 C. to about 15 C.

[0277] In one embodiment, the range is from about 2 C. to about 10 C.

[0278] In one embodiment, the range is from about 2 C. to about 8 C.

[0279] In one embodiment, the range is from about 2 C. to about 8 C.

[0280] In one embodiment, the temperature is about 5 C.

[0281] In one embodiment, the reaction time is from about 5 minutes to about 12 hours.

[0282] In one embodiment, the range is from about 10 minutes to about 12 hours.

[0283] In one embodiment, the range is from about 15 minutes to about 6 hours.

[0284] In one embodiment, the range is from about 30 minutes to about 4 hours.

[0285] In one embodiment, the range is from about 1 hour to about 3 hours.

[0286] In one embodiment, the time is about 2 hours.

[0287] In one embodiment, the reaction is carried out in the presence of an acid.

[0288] In one embodiment, the reaction is carried out in the presence of a strong acid.

[0289] In one embodiment, the reaction is carried out in the presence of sulfuric acid.

[0290] In one embodiment, the reaction is carried out in the presence of concentrated sulfuric acid.

[0291] In one embodiment, when the reaction is carried out in the presence of an acid, the ratio, C, of the amount of compound of Formula (2), in equivalents, to the amount of H.sup.+ provided by the acid, in equivalents, is from about 0.5 to about 3.0.

[0292] In one embodiment, the range is from about 0.6 to about 2.0.

[0293] In one embodiment, the range is from about 0.7 to about 1.5.

[0294] In one embodiment, the range is from about 0.8 to about 1.2.

[0295] In one embodiment, the range is from about 0.9 to about 1.1.

[0296] In one embodiment, the ratio is about 1.

[0297] In some embodiments, the acid is a strong diprotic acid, for example sulphuric acid, so that 0.5 equivalents of strong diprotic acid corresponds to 1.0 equivalents of H.sup.+.

[0298] As an illustration, in one example of the periodate mediated oxidative coupling, thiosulphonic acid S-(2-amino-5-dimethyl amino) phenyl ester (5.92 g, 248.32 g/mol, 23.8 mmol, 1.0 equivalent) (a compound of Formula (2)) is used in the presence of sulphuric acid (H.sub.2SO.sub.4, 98%, 1.17 g, 98.08 g/mol, 11.9 mmol, 0.5 equivalents of H.sub.2SO.sub.4, 1.0 equivalents of H.sup.+).

[0299] In another example of the periodate mediated oxidative coupling, thiosulphonic acid S-(2-amino-5-dimethyl amino) phenyl ester (10.0 g, 248.32 g/mol, 40.3 mmol, 1.0 equivalent) (a compound of Formula (2)) is used in the presence of sulphuric acid (H.sub.2SO.sub.4, 98%, 1.97 g, 98.08 g/mol, 20.1 mmol, 0.5 equivalents of H.sub.2SO.sub.4, 1.0 equivalents of H.sup.+).

[0300] In one embodiment, the reaction is carried out in the presence of water.

[0301] In one embodiment, the acid is added to the compound of Formula (3) in water; then the compound of Formula (4) is added; and then the periodate oxidizing agent is added.

[0302] In one embodiment, the acid is added stepwise.

[0303] In one embodiment, the compound of Formula (4) is added in one aliquot.

[0304] In one embodiment, the periodate oxidizing agent is added stepwise.

[0305] Additional Steps

[0306] The present invention also relates to such methods which incorporate one or more additional (subsequent and/or preceding) steps, for example: to prepare compounds of Formula (5) from compounds of Formula (1); to prepare compounds of Formula (6) from compounds of Formula (5); and to prepare compounds of Formula (2) from compounds of Formula (4), as described herein.

[0307] Preceding Step: Thiosulfonic Acid Formation

[0308] In one embodiment, the methods described above further comprise a preceding step of: converting a compound of Formula (4):

##STR00020##

to the corresponding compound of Formula (2):

##STR00021##

wherein each of R.sup.1A1, R.sup.1A2, R.sup.2A, R.sup.3A and R.sup.4A is as defined herein.

[0309] Methods, reagents, and reaction conditions suitable for such a reaction are well known in the art. See, for example, WO 2010/130977 A1 (WisTa Laboratories Ltd., 18 Nov. 2010). In particular, see: Synthesis 1, examples 1 to 6, pages 55 to 59; Synthesis 4, page 61; Synthesis 5, page 62; and Synthesis 6, example 1 page 66-67 and example 2 page 68.

[0310] For example, the compound of Formula (4) may be reacted with aluminium sulphate hexdecahydrate, then sodium thiosulphate, and then potassium persulphate, in water, at a temperature of about 5-15 C., and stirred for about 2 hours; and the precipitate collected, washed, and dried.

[0311] Subsequent Step: Cyclization

[0312] In one embodiment, the methods described above further comprise a subsequent step of: converting the compound of Formula (1):

##STR00022##

to the corresponding compound of Formula (5):

##STR00023##

wherein each of R.sup.1A1, R.sup.1A2, R.sup.1B1, R.sup.1B2, R.sup.2A, R.sup.2B, R.sup.3A, R.sup.3B, R.sup.4A, and R.sup.4B is as defined herein; and
wherein X.sup. is independently one or more anionic counter ions to achieve electrical neutrality.

[0313] In one embodiment, X.sup. is independently a counter anion to achieve electrical neutrality.

[0314] In one embodiment, X.sup. is independently a counter anion shared with one or more other cations (e.g., the cation shown in Formula (5)) to achieve electrical neutrality.

[0315] In one embodiment, X.sup. is independently a halogen anion (i.e., a halide).

[0316] In one embodiment, X.sup. is independently F.sup., Cl.sup., Br.sup., or I.sup..

[0317] In one embodiment, X.sup. is independently Cl.sup., Br.sup., or I.sup..

[0318] In one embodiment, X.sup. is independently Cl.sup..

[0319] In one embodiment, X.sup. is independently NO.sub.3.sup. (nitrate).

[0320] In one embodiment, X.sup. is independently CIO.sub.3.sup. (perchlorate).

[0321] In one embodiment, X.sup. is independently S.sub.2O.sub.8.sup. (persulfate).

[0322] In one embodiment, X.sup. is independently formate, propionate, or benzoate.

[0323] In one embodiment, X.sup. is independently 4-hydroxybenzenesulfonate, p-toluenesulfonate (CH.sub.3C.sub.6H.sub.4S(O).sub.2O.sup.), or methylsulfonate (CH.sub.3S(O).sub.2O.sup.).

[0324] In one embodiment, X.sup. is independently derived from FeCl.sub.3 or ZnCl.sub.2.

[0325] In one embodiment, X.sup. is independently SO.sub.4.sup.2 (sulfate).

[0326] In one embodiment, X.sup. is independently succinate.

[0327] In one embodiment, X.sup. is independently citrate (and, e.g., is shared with one or more other cations (e.g., the cation shown in Formula (6)) to achieve electrical neutrality).

[0328] Methods, reagents, and reaction conditions suitable for such a reaction are well known in the art. See, for example, WO 2010/130977 A1 (WisTa Laboratories Ltd., 18 Nov. 2010), in particular: Synthesis 3, page 60; Synthesis 5, page 63 to 64; Synthesis 6, page 68-69; and Synthesis 7, page 70. Also see, for example WO 2015/052496 A1 (WisTa Laboratories Ltd., 16 Apr. 2015), in particular see examples 1 to 5, pages 47 to 57.

[0329] For example, the compound of Formula (1) may be reacted with copper (II) sulfate, in water, at a temperature of about 85 C. for about 1 hour; the liquid phase collected and reacted with hydrochloric acid and allowed to cool; and the precipitate collected, washed, and dried.

[0330] Subsequent Step: Reduction

[0331] In one embodiment, the methods described above further comprise a subsequent step of: converting the compound of Formula (5):

##STR00024##

to the corresponding compound of Formula (6):

##STR00025##

wherein each of R.sup.1A1, R.sup.1A2, R.sup.1B1, R.sup.1B2, R.sup.2A, R.sup.2B, R.sup.3A, R.sup.3B, R.sup.4A, R.sup.4B, and X.sup. is as defined herein; and
wherein Y.sup. and Z.sup., taken together, are independently one or more anionic counter ions to achieve electrical neutrality.

[0332] In one embodiment, Y.sup. and Z.sup., taken together, are independently two counter anions to achieve electrical neutrality.

[0333] In one embodiment, Y.sup. and Z.sup., taken together, is independently one counter anion to achieve electrical neutrality.

[0334] In one embodiment, Y.sup. and Z.sup., taken together, is independently a counter anion shared with one or more other cations (e.g., the cation shown in Formula (6)) to achieve electrical neutrality.

[0335] In one embodiment, each of Y.sup. and Z.sup. is independently a halogen anion (i.e., a halide).

[0336] In one embodiment, each of Y.sup. and Z.sup. is independently F.sup., Cl.sup., Br.sup., or I.sup..

[0337] In one embodiment, each of Y.sup. and Z.sup. is independently Cl.sup., Br.sup., or I.sup..

[0338] In one embodiment, each of Y.sup. and Z.sup. is independently Cl.sup..

[0339] In one embodiment, each of Y.sup. and Z.sup. is independently NO.sub.3.sup. (nitrate).

[0340] In one embodiment, each of Y.sup. and Z.sup. is independently ClO.sub.3.sup. (perchlorate).

[0341] In one embodiment, each of Y.sup. and Z.sup. is independently S.sub.2O.sub.8.sup. (persulfate).

[0342] In one embodiment, each of Y.sup. and Z.sup. is independently formate, propionate, or benzoate.

[0343] In one embodiment, each of Y.sup. and Z.sup. is independently 4-hydroxybenzenesulfonate, p-toluenesulfonate (CH.sub.3C.sub.6H.sub.4S(O).sub.2O.sup.), methylsulfonate (CH.sub.3S(O).sub.2O.sup.).

[0344] In one embodiment, Y.sup. and Z.sup., taken together, is independently SO.sub.4.sup.2 (sulfate).

[0345] In one embodiment, Y.sup. and Z.sup., taken together, is independently succinate.

[0346] In one embodiment, Y.sup. and Z.sup., taken together, is independently citrate (and, e.g., is shared with one or more other cations (e.g., the cation shown in Formula (6)) to achieve electrical neutrality).

[0347] Methods, reagents, and reaction conditions suitable for such a reaction are well known in the art. See, for example WO 2007/110627 A2 (WisTa Laboratories Ltd., 4 Oct. 2007), in particular see Synthesis 8 and 9, page 57; Synthesis 12 to 18 and 23, pages 59 to 63 and 65; and Synthesis 20 to 22 pages 64 and 65. See also, for example, PCT/EP2016/067302, WO 2017/013137 (WisTa Laboratories Ltd., filed 20 Jul. 2016), in particular: Method 4, part 3, page 97; and Methods 8 to 12, page 111 to 113.

[0348] For example, the compound of Formula (5) may be treated with an acid such as hydrochloric acid in methanol and allowed to stir for 3 hours; the solution is filtered through Celite, washed with methanol and concentrated to provide a compound of Formula (6). Alternatively, for example, the compound of Formula (5) may be treated with an acid such as methane sulfonic acid in methanol and toluene; the mixture is subsequently cooled to 5 C. before ethanol is added such that the product, Formula (6), precipitates and can be collected by filtration.

[0349] For example, the compound of Formula (5) may be treated with an acylating agent, such as acetic anhydride, under basic conditions and stirred for 2 hours at around 90 C. to acylate the aromatic nitrogen. The acylated intermediate may then be treated with an acid, such hydrochloric acid, under heating, for example at 80 C. to give the product, Formula (6).

[0350] Chemical Synthesis

[0351] Methods for the chemical synthesis of compounds of the present invention are described herein. These and/or other well-known methods may be modified and/or adapted in known ways in order to facilitate the synthesis of additional compounds within the scope of the present invention.

[0352] Descriptions of general laboratory methods and procedures, useful for the preparation of the compounds described herein, are provided in Vogel's Textbook of Practical Organic Chemistry, 5th Edition, 1989, (Editors: Furniss, Hannaford, Smith, and Tatchell) (published by Longmann, UK).

[0353] Compositions

[0354] One aspect of the present invention pertains to a composition comprising a compound of Formula (1), Formula (5), or Formula (6) as described herein (for example, which is obtainable, or which is obtained by a method as described herein), and a carrier, diluent, or excipient.

[0355] Another aspect of the present invention pertains to a method of preparing a composition comprising mixing a compound of Formula (1), Formula (5), or Formula (6) as described herein (for example, which is obtainable, or which is obtained by a method as described herein), and a carrier, diluent, or excipient.

[0356] One aspect of the present invention pertains to a pharmaceutical composition comprising a compound of Formula (5) or Formula (6) as described herein (for example, which is obtainable, or which is obtained by a method as described herein), and a pharmaceutically acceptable carrier, diluent, or excipient.

[0357] Another aspect of the present invention pertains to a method of preparing a pharmaceutical composition comprising mixing a compound of Formula (5) or Formula (6) as described herein(for example, which is obtainable, or which is obtained by a method as described herein), and a pharmaceutically acceptable carrier, diluent, or excipient.

[0358] Uses

[0359] The compounds of Formula (5) and Formula (6) as described herein, are useful in medicine (e.g., therapy), for example, in treatment or prophylaxis.

[0360] Use in Methods of Therapy

[0361] One aspect of the present invention pertains to a compound of Formula (5) or Formula (6) as described herein (for example, which is obtainable, or which is obtained by a method as described herein), for use in medicine, for example, for use in treatment or prophylaxis, for example, for use in treatment or prophylaxis a disorder (e.g., a disease), as described herein.

[0362] Use in the Manufacture of Medicaments

[0363] One aspect of the present invention pertains to use of a compound of Formula (1), Formula (5), or Formula (6) as described herein (for example, which is obtainable, or which is obtained by a method as described herein), in the manufacture of a medicament, for example, for use in a method of treatment or prophylaxis, for example, for use in a method of treatment or prophylaxis of a disorder (e.g., a disease), as described herein.

[0364] In one embodiment, the medicament comprises the compound of Formula (5) or Formula (6).

[0365] Methods of Treatment

[0366] One aspect of the present invention pertains to a method of treatment or prophylaxis, for example, a method of treatment or prophylaxis of a disorder (e.g., a disease), as described herein, comprising administering to a subject in need of treatment a therapeutically-effective amount of a compound of Formula (5) or Formula (6) as described herein (for example, which is obtainable, or which is obtained by a method as described herein), preferably in the form of a pharmaceutical composition.

[0367] Disorders Treated

[0368] In one embodiment, the disorder is a disease of protein aggregation.

[0369] In one embodiment, the disorder is a tauopathy.

[0370] In one embodiment, the disorder is Alzheimer's disease (AD), Pick's disease, progressive supranuclear palsy (PSP), frontotemporal dementia (FTD), FTD with parkinsonism linked to chromosome 17 (FTDP 17), frontotemporal lobar degeneration (FTLD) syndromes; disinhibition-dementia-parkinsonism-amyotrophy complex (DDPAC), pallido-ponto-nigral degeneration (PPND), amyotropic lateral sclerosis (ALS), Guam-ALS syndrome, pallido nigro luysian degeneration (PNLD), cortico-basal degeneration (CBD), dementia with argyrophilic grains (AgD), dementia pugilistica (DP) or chronic traumatic encephalopathy (CTE), Down's syndrome (DS), dementia with Lewy bodies (DLB), subacute sclerosing panencephalitis (SSPE), MCl, Niemann-Pick disease, type C (NPC), Sanfilippo syndrome type B (or mucopolysaccharidosis III B (MPS III B)), or myotonic dystrophies (DM), DM1 or DM2.

[0371] In one embodiment, the disorder is Alzheimer's disease.

[0372] In one embodiment, the disorder is Parkinson's disease.

[0373] In one embodiment, the disorder is PSP, ALS, or FTLD.

[0374] In one embodiment, the disorder is Huntington's disease.

[0375] In one embodiment, the disorder is Huntington's disease or another polyglutamine disorder, such as spinal bulbar muscular atrophy (Kennedy disease), dentatorubropallidoluysian atrophy, or spinocerebellar ataxias.

[0376] In one embodiment, the disorder is skin cancer.

[0377] In one embodiment, the disorder is melanoma.

[0378] In one embodiment, the disorder is a bacterial, viral, or protozoal disease condition.

[0379] In one embodiment, the disorder is a viral disease condition.

[0380] In one embodiment, the disorder is Hepatitis C, HIV, or West Nile Virus (WNV) infection.

[0381] In one embodiment, the disorder is a protozoan disease.

[0382] In one embodiment, the disorder is malaria.

[0383] Treatment

[0384] The term treatment, as used herein in the context of treating a disorder, pertains generally to treatment of a human or an animal (e.g., in veterinary applications), in which some desired therapeutic effect is achieved, for example, the inhibition of the progress of the disorder, and includes a reduction in the rate of progress, a halt in the rate of progress, alleviation of symptoms of the disorder, amelioration of the disorder, and cure of the disorder. Treatment as a prophylactic measure (i.e., prophylaxis) is also included. For example, use with patients who have not yet developed the disorder, but who are at risk of developing the disorder, is encompassed by the term treatment.

[0385] The term therapeutically-effective amount, as used herein, pertains to that amount of a compound, or a material, composition or dosage form comprising a compound, which is effective for producing some desired therapeutic effect, commensurate with a reasonable benefit/risk ratio, when administered in accordance with a desired treatment regimen.

[0386] Combination Therapies

[0387] The term treatment includes combination treatments and therapies, in which two or more treatments or therapies are combined, for example, sequentially or simultaneously. For example, the compounds described herein may also be used in combination therapies, e.g., in conjunction with other agents.

[0388] The particular combination would be at the discretion of the physician who would select dosages using his common general knowledge and dosing regimens known to a skilled practitioner.

[0389] The agents (i.e., the compound of Formula (1), Formula (2), Formula (5), or Formula (6) plus one or more other agents) may be administered simultaneously or sequentially, and may be administered in individually varying dose schedules and via different routes.

[0390] The agents (i.e., the compound of Formula (1), Formula (2), Formula (5), or Formula (6) plus one or more other agents) may be formulated together in a single dosage form, or alternatively, the individual agents may be formulated separately and presented together in the form of a kit, optionally with instructions for their use.

[0391] Kits

[0392] One aspect of the invention pertains to a kit comprising (a) a compound of Formula (5) or Formula (6) as described herein (for example, which is obtainable, or which is obtained by a method as described herein), or a composition comprising a compound of Formula (5) or Formula (6) as described herein (for example, which is obtainable, or which is obtained by a method as described herein), e.g., preferably provided in a suitable container and/or with suitable packaging; and (b) instructions for use, e.g., written instructions on how to administer the compound or composition.

[0393] The written instructions may also include a list of indications for which the active ingredient is a suitable treatment.

[0394] Routes of Administration

[0395] The compound of Formula (5) or Formula (6) or pharmaceutical composition comprising the compound, may be administered to a subject by any convenient route of administration. Typically, the compound is administered orally or intravenously.

[0396] The Subject/Patient

[0397] The subject/patient may be a mammal, a placental mammal, a marsupial (e.g., kangaroo, wombat), a rodent (e.g., a guinea pig, a hamster, a rat, a mouse), murine (e.g., a mouse), a lagomorph (e.g., a rabbit), avian (e.g., a bird), canine (e.g., a dog), feline (e.g., a cat), equine (e.g., a horse), porcine (e.g., a pig), ovine (e.g., a sheep), bovine (e.g., a cow), a primate, simian (e.g., a monkey or ape), a monkey (e.g., marmoset, baboon), an ape (e.g., gorilla, chimpanzee, orangutang, gibbon), or a human.

[0398] In one preferred embodiment, the subject/patient is a human.

[0399] Formulations

[0400] While it is possible for a compound of Formula (5) or Formula (6) to be administered alone, it is preferable to present it as a pharmaceutical formulation (e.g., composition, preparation, medicament) comprising at least one compound, as described herein, together with one or more other pharmaceutically acceptable ingredients well-known to those skilled in the art, including pharmaceutically acceptable carriers, diluents, excipients, adjuvants, fillers, buffers, preservatives, anti-oxidants, lubricants, stabilisers, solubilisers, surfactants (e.g., wetting agents), masking agents, colouring agents, flavouring agents, and sweetening agents. If formulated as discrete units (e.g., tablets, etc.), each unit contains a predetermined amount (dosage) of the compound. The formulation may further comprise other active agents, for example, other therapeutic or prophylactic agents.

[0401] Suitable carriers, diluents, excipients, etc. can be found in standard pharmaceutical texts. See, for example, Handbook of Pharmaceutical Additives, 2nd Edition (eds. M. Ash and I. Ash), 2001 (Synapse Information Resources, Inc., Endicott, N.Y., USA), Remington's Pharmaceutical Sciences, 20th edition, pub. Lippincott, Williams & Wilkins, 2000; and Handbook of Pharmaceutical Excipients, 5th edition, 2005.

[0402] The term pharmaceutically acceptable, as used herein, pertains to compounds, ingredients, materials, compositions, dosage forms, etc., which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of the subject in question (e.g., human) without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit/risk ratio. Each carrier, diluent, excipient, etc. must also be acceptable in the sense of being compatible with the other ingredients of the formulation.

[0403] The formulations may be prepared by any methods well known in the art of pharmacy. Such methods include the step of bringing into association the compound with a carrier which constitutes one or more accessory ingredients. In general, the formulations are prepared by uniformly and intimately bringing into association the compound with carriers (e.g., liquid carriers, finely divided solid carrier, etc.), and then shaping the product, if necessary.

[0404] The formulation may be prepared to provide for rapid or slow release; immediate, delayed, timed, or sustained release; or a combination thereof.

[0405] Formulations suitable for oral administration (e.g., by ingestion) include liquids, solutions (e.g., aqueous, non-aqueous), suspensions (e.g., aqueous, non-aqueous), emulsions (e.g., oil-in-water, water-in-oil), elixirs, syrups, electuaries, tablets, granules, powders, capsules, cachets, pills, ampoules, boluses.

[0406] Tablets may be made by conventional means, e.g., compression or moulding, optionally with one or more accessory ingredients. Compressed tablets may be prepared by compressing in a suitable machine the compound in a free-flowing form such as a powder or granules, optionally mixed with one or more binders (e.g., povidone, gelatin, acacia, sorbitol, tragacanth, hydroxypropylmethyl cellulose); fillers or diluents (e.g., lactose, microcrystalline cellulose, calcium hydrogen phosphate); lubricants (e.g., magnesium stearate, talc, silica); disintegrants (e.g., sodium starch glycolate, cross-linked povidone, cross-linked sodium carboxymethyl cellulose); surface-active or dispersing or wetting agents (e.g., sodium lauryl sulfate); preservatives (e.g., methyl p-hydroxybenzoate, propyl p-hydroxybenzoate, sorbic acid); flavours, flavour enhancing agents, and sweeteners. Moulded tablets may be made by moulding in a suitable machine a mixture of the powdered compound moistened with an inert liquid diluent. The tablets may optionally be coated or scored and may be formulated so as to provide slow or controlled release of the compound therein using, for example, hydroxypropylmethyl cellulose in varying proportions to provide the desired release profile. Tablets may optionally be provided with a coating, for example, to affect release, for example an enteric coating, to provide release in parts of the gut other than the stomach.

[0407] Formulations suitable for parenteral administration (e.g., by injection), include aqueous or non-aqueous, isotonic, pyrogen-free, sterile liquids (e.g., solutions, suspensions), in which the compound is dissolved, suspended, or otherwise provided (e.g., in a liposome or other microparticulate). Such liquids may additional contain other pharmaceutically acceptable ingredients, such as anti-oxidants, buffers, preservatives, stabilisers, bacteriostats, suspending agents, thickening agents, and solutes which render the formulation isotonic with the blood (or other relevant bodily fluid) of the intended recipient. Examples of excipients include, for example, water, alcohols, polyols, glycerol, vegetable oils, and the like. Examples of suitable isotonic carriers for use in such formulations include Sodium Chloride Injection, Ringer's Solution, or Lactated Ringer's Injection. Typically, the concentration of the compound in the liquid is from about 1 ng/ml to about 10 g/ml, for example from about 10 ng/ml to about 1 g/ml. The formulations may be presented in unit-dose or multi-dose sealed containers, for example, ampoules and vials, and may be stored in a freeze-dried (lyophilised) condition requiring only the addition of the sterile liquid carrier, for example water for injections, immediately prior to use. Extemporaneous injection solutions and suspensions may be prepared from sterile powders, granules, and tablets.

[0408] Dosage

[0409] It will be appreciated by one of skill in the art that appropriate dosages of the compound of Formula (5) or Formula (6) and compositions comprising the compound can vary from patient to patient. Determining the optimal dosage will generally involve the balancing of the level of therapeutic benefit against any risk or deleterious side effects. The selected dosage level will depend on a variety of factors including the activity of the particular compound, the route of administration, the time of administration, the rate of excretion of the compound, the duration of the treatment, other drugs, compounds, and/or materials used in combination, the severity of the disorder, and the species, sex, age, weight, condition, general health, and prior medical history of the patient. The amount of compound and route of administration will ultimately be at the discretion of the physician, veterinarian, or clinician, although generally the dosage will be selected to achieve local concentrations at the site of action which achieve the desired effect without causing substantial harmful or deleterious side-effects.

[0410] Administration can be effected in one dose, continuously or intermittently (e.g., in divided doses at appropriate intervals) throughout the course of treatment. Methods of determining the most effective means and dosage of administration are well-known to those of skill in the art and will vary with the formulation used for therapy, the purpose of the therapy, the target cell(s) being treated, and the subject being treated. Single or multiple administrations can be carried out with the dose level and pattern being selected by the treating physician, veterinarian, or clinician.

[0411] Examples of Some Preferred Formulations

[0412] A preferred formulation is a dosage unit (e.g., a pharmaceutical tablet or capsule) comprising 20 to 300 mg of a compound of Formula (5) or Formula (6) as described herein; and a pharmaceutically acceptable carrier, diluent, or excipient.

[0413] In some embodiments, the dosage unit is a tablet.

[0414] In some embodiments, the dosage unit is a capsule.

[0415] In some embodiments, said capsules are gelatine capsules.

[0416] In some embodiments, said capsules are HPMC (hydroxypropylmethylcellulose) capsules.

[0417] In some embodiments, the amount is 30 to 200 mg.

[0418] In some embodiments, the amount is about 30 mg.

[0419] In some embodiments, the amount is about 60 mg.

[0420] In some embodiments, the amount is about 100 mg.

[0421] In some embodiments, the amount is about 150 mg.

[0422] In some embodiments, the amount is about 200 mg.

[0423] The dosage amounts as set out above may refer to the amount of the compound itself or may refer to the amount of free base equivalent contained in the dosage unit. Both of these alternatives are specifically and explicitly disclosed by the present disclosure.

[0424] In some embodiments, the pharmaceutically acceptable carrier, diluent, or excipient is or comprises one or both of a glyceride (e.g., Gelucire 44/14; lauroyl macrogol-32 glycerides PhEur, USP) and colloidal silicon dioxide (e.g., 2% Aerosil 200; Colliodal Silicon Dioxide PhEur, USP).

EXAMPLES

[0425] The following examples are provided solely to illustrate the present invention and are not intended to limit the scope of the invention, as described herein.

Method 1

Thiosulphonic acid S-(2-amino-5-dimethyl amino) phenyl ester

[0426] ##STR00026##

[0427] N,N-Dimethyl-p-phenylenediamine (10 g, 136.2 g/mol, 73.4 mmol, 1.0 equivalent) and water (200 mL) was added to a multi-necked round bottom flask.

[0428] The reaction mixture was cooled and maintained at 5 C. using an ice/water cooling bath and stirred for 10 minutes.

[0429] Aluminium sulphate hexadecahydrate (Al.sub.2(SO.sub.4).sub.3. 16H.sub.2O (23.14 g, 630.39 g/mol, 36.7 mmol, 0.5 equivalents) was added to the reaction mixture in one portion.

[0430] After 5 minutes, a solution of sodium thiosulphate (Na.sub.2S.sub.2O.sub.3.5H.sub.2O, 20.04 g, 248.18 g/mol, 80.7 mmol, 1.1 equivalents, dissolved in 20 mL of water) was added to the reaction mixture as a single portion.

[0431] After another 5 minutes, potassium persulphate (K.sub.2S.sub.2O.sub.8, 19.86 g, 270.32 g/mol, 73.5 mmol, 1.0 equivalents) was added to the reaction mixture over a 10 minute period. A rise in temperature from 5 C. to 11 C. was observed.

[0432] The reaction mixture was stirred for another 2 hours, whilst maintained at 5 C. using an ice/water cooling bath.

[0433] The reaction mixture was warmed over a 30 minute period to 20 C.

[0434] The solid product was collected by filtration and washed with 50 C. water (320 mL).

[0435] The filter cake was then washed with ethyl acetate (20 mL) and dried on the filter under suction for 20 minutes.

[0436] The solid was further dried in a vacuum oven (50 C. at <950 mbar) to achieve a constant weight and provide the crude product as a black/purple solid with white flakes (15.791 g).

[0437] The product was characterised using NMR: .sup.1H NMR (300 MHz, D.sub.2O): =7.22 (s, 1H, ArH), 7.07 (m, 2H, ArH), 3.01 (d, 6H, CH.sub.3).

[0438] FIG. 1 shows the .sup.1H NMR (300 MHz, DMSO-d.sub.6) spectrum for thiosulphonic acid S-(2-amino-5-dimethyl amino) phenyl ester obtained in Method 1.

[0439] A DMSO-d.sub.6 solution of equimolar amounts of the crude product and a reference standard (3-(trimethylsilyl)-1-propanesulfonic acid (97%)) was analysed by proton nuclear magnetic resonance spectroscopy (.sup.1H NMR).

[0440] FIG. 2 shows the .sup.1H NMR (300 MHz, DMSO-d.sub.6) spectrum for a mixture of thiosulphonic acid S-(2-amino-5-dimethyl amino) phenyl ester obtained in Method 1 and the reference compound 3-(trimethylsilyl)-1-propanesulfonic acid.

[0441] From the .sup.1H NMR spectrum (FIG. 2), a purity of 87% was calculated. A residual inorganic salt can be seen as the white flakes within the sample. The amount of target compound (thiosulphonic acid S-(2-amino-5-dimethyl amino) phenyl ester) in the crude material was calculated to be 13.74 g (75% yield). The crude material was used in the subsequent reaction step without further purification.

Method 2A

(4-(2-(Thiosulfate)-4-(dimethylamino)-phenyl-imino)-cyclohex-2,5-dienylidene)-N,N-dimethyl ammonium

[0442] ##STR00027##

[0443] N,N-dimethylaniline (C.sub.6H.sub.6N(CH.sub.3).sub.2, 2.89 g, 121.18 g/mol, 23.8 mmol, 1.0 equivalent) and water (79 mL) were added to a multi-necked round bottom flask.

[0444] Sulphuric acid (H.sub.2SO.sub.4, 98%, 1.17 g, 98.08 g/mol, 11.9 mmol, 0.5 equivalents) was added to the reaction mixture drop-wise over 10 minutes.

[0445] Crude thiosulphonic acid S-(2-amino-5-dimethyl amino) phenyl ester (7.89 g which contains 5.92 g, 248.32 g/mol, 23.8 mmol, 1.0 equivalent) was added to the reaction mixture in a single aliquot to form a suspension.

[0446] The reaction mixture was cooled to 5 C. and maintained at 5 C. using an ice/water cooling bath and stirred for over 10 minutes.

[0447] Sodium periodate (NaIO.sub.4, 10.66 g, 213.89 g/mol, 49.8 mmol, 2.09 equivalents) was added to the reaction mixture in aliquots of about 1 g over a 40 minute period while maintained at 5 C. to form a green slurry.

[0448] The reaction mixture was stirred for 2 hours while maintained at 5 C.

[0449] The reaction mixture was filtered and the solid filter cake washed with pre-heated (50 C.) water (220 mL) and dried on the filter under suction for 20 minutes.

[0450] A wet mass of 23 g of crude product was obtained, and was used in the subsequent reaction step without further drying or purification.

Method 2B

(4-(2-(Thiosulfate)-4-(dimethylamino)-phenyl-imino)-cyclohex-2,5-dienylidene)-N,N-dimethyl ammonium

[0451] ##STR00028##

[0452] N,N-dimethylaniline (C.sub.6H.sub.5N(CH.sub.3).sub.2, 4.88 g, 121.18 g/mol, 40.3 mmol, 1.0 equivalent) and water (100 mL) were added to a multi-necked round bottom flask.

[0453] Sulphuric acid (H.sub.2SO.sub.4, 98%, 1.97 g, 98.08 g/mol, 20.1 mmol, 0.5 equivalents) was added to the reaction mixture drop-wise over 10 minutes.

[0454] Thiosulphonic acid S-(2-amino-5-dimethyl amino) phenyl ester (10.0 g, 248.32 g/mol, 40.3 mmol, 1.0 equivalent) was added to the reaction mixture in a single aliquot to form a suspension.

[0455] The reaction mixture was cooled to 5 C. and maintained at 5 C. using an ice/water cooling bath and stirred for 10 minutes.

[0456] Sodium periodate (NaIO.sub.4, 17.67 g, 213.89 g/mol, 82.6 mmol, 2.05 equivalents) was added in aliquots of about 1 g over a 90 minute period while maintained at 5 C. to form a green slurry.

[0457] The reaction mixture was stirred for 2 hours while maintained at 5 C.

[0458] The reaction mixture was filtered and the solid filter cake washed with water (240 mL) and dried on the filter under suction for 20 minutes.

[0459] The filter cake was re-slurried with fresh water (100 mL) and stirred for 20 minutes.

[0460] The slurry was then re-filtered and the solid filter cake washed with water (40 mL) and dried on the filter under suction for 30 minutes.

[0461] The solid was further dried under vacuum (<900 mbar) at 20 C. for 48 hours to provide a green powder (12.78 g).

[0462] The product was characterized, and the resulting data are summarized in the following table.

TABLE-US-00001 TABLE 1 Characterisation of Product of Method 2B Weight loss on drying 7.32% (Karl Fischer) .sup.1H NMR 5 = 3.34 (s, 6H), 3.45 (s, 6H), 7.13-7.37 (m, 6H), (300 MHz, DMSO-d.sub.6) 7.95 (s, 1H) v.sub.max (cm.sup.1) 1594(m), 1411(m), 1360 (m), 1331 (m), 1161 (s), 1015 (s), 869 (m), 627 (s) MS, m/z (ES+) Theoretical: [M + Na].sup.+ 389.0844 (amu) Accurate Mass Measured: [M + Na].sup.+ 389.0837 (amu) HPLC (a/a) 74.74% Accurate yield 59%

[0463] FIG. 3 shows the .sup.1H NMR (300 MHz, DMSO-d.sub.6) spectrum for (4-(2-(thiosulfate)-4-(dimethylamino)-phenyl-imino)-cyclohex-2,5-dienylidene)-N,N-dimethyl ammonium obtained in Method 2B.

Method 3

Methylthioninium Chloride (MTC)

[0464] ##STR00029##

[0465] Crude (4-(2-(thiosulfate)-4-(dimethylamino)-phenyl-imino)-cyclohex-2,5-dienylidene)-N,N-dimethyl ammonium (prepared by Method 2A, approximately 23 g of crude product), water (100 mL) and copper (II) sulphate (CuSO.sub.4.5H.sub.2O, 0.98 g, 249.68 g/mol, 3.93 mmol, 0.165 equivalents based on the thiosulphonic acid used in Method 2A) were added to a multi-necked round bottom flask form a slurry.

[0466] The reaction mixture was heated at 85 C. and stirred for 1 hour during which time an intense blue colour developed.

[0467] The reaction mixture was then filtered while at 85 C., and the solid waste in the filter was washed with pre-heated (50 C.) water (210 mL).

[0468] The combined filtrate from the reaction liquor and washings was then cooled to 35 C. over 30 minutes, during which air was bubbled through the blue solution.

[0469] Hydrochloric acid (HCl, 32%, 12 mL) was added and the reaction mixture was stirred for 14 hours to allow crystallisation.

[0470] The product was collected by filtration and washed with pre-chilled (5 C.) water (210 mL), which had been acidified with hydrochloric acid to pH 1.

[0471] The product was then washed with toluene (10 mL), dried on the filter under suction for 20 minutes, and then dried in a fan-assisted oven at 40 C. for 9 hours to provide (green crystalline needles) (3.93 g).

[0472] The product was characterized, and the resulting data are summarized in the following table.

TABLE-US-00002 TABLE 2 Characterisation of Product of Method 3 Weight loss on drying 11.00% (moisture balance) .sup.1H NMR = 3.03 (d, 12H), 6.73 (d, J = 2.6 Hz, 2H), (300 MHz, D.sub.2O) 6.95 (dd, J = 9.6 Hz, 2H), 7.22 (d, J = 9.7 Hz, 2H) .sup.13C NMR (75 MHz, = 40.49 (4C), 106.00 (2C), 118.32 (2C), 133.75 D.sub.2O) (2C), 134.00 (2C), 136.27 (2C), 1523.15 (2C) v.sub.max (cm.sup.1) 3351 (b, H.sub.2O Solvate), 1592(s), 1486(m), 1391(s), 1334(s), 1176(m), 1137(m), 878(s) MS, m/z (ESI) [M.sup.+] 284 HPLC (w/w) 82.63% Accurate yield 42% of MTC (*) (*) The yield of MTC is reported with respect to the starting material, N,N-dimethylaniline, used in Method 2A. That is, the yield reported is the yield over two steps, Method 2A and Method 3. The yield of MTC over three steps, Method 1, Method 2A and Method 3, with respect to the N,N-Dimethyl-p-phenylenediamine starting material from Method 1 is 28%.

[0473] FIG. 4 shows the .sup.1H NMR (300 MHz, D.sub.2O) spectrum for methylthioninium chloride (MTC) obtained in Method 3.

[0474] The organic purity of the methylthioninium chloride (MTC) product was determined by HPLC analysis and the results are summarised in the following table.

TABLE-US-00003 TABLE 3 HPLC Purity of MTC Product Compound % (a/a) % (w/w) MTC 97.11 82.63 Azure B 2.60 2.20 Azure A 0.17 <0.05(*) Azure C MVB <0.05 (*) 0.14 MVB-CH.sub.3 sDMT <0.05 (*) <0.05 (*) Others 0.12 ND Total 100 84.97 (*) The <0.05 amounts are ignored in the calculation of the total.

[0475] The term others refers to all other compounds that are present, for which a specific value is not reported.

[0476] As used herein, HPLC % (a/a) refers to HPLC percent area by area, and denotes the ratio of the area under the HPLC peak associated with the chemical species to the total area under all of the HPLC peaks observed, expressed as a percent. For example, Azure B % (a/a) denotes the ratio of the area under the HPLC peak associated with Azure B to the total area under all of the HPLC peaks observed, multiplied by 100.

[0477] Similarly, as used herein, HPLC % (w/w) refers to HPLC percent weight by weight, and denotes the ratio of the area under the HPLC peak compared with the area under the HPLC peak of a reference standard, expressed as a percent. For example, Azure B % (w/w) denotes the ratio of the area under the Azure B peak compared against the area under the peak of an Azure B reference standard of known concentration, multiplied by 100.

TABLE-US-00004 TABLE 4 System Parameters for HPLC Purity Analysis of MTC HPLC system Agilent 1200 with DAD and data handling capacity Column Agilent Eclipse XDB-Phenyl, 150 4.6 mm, 3.5 m particle size Column Temperature 50 C. Autosampler Temperature 5 C. Mobile Phase A: 0.1% v/v trifluoroacetic acid B: Acetonitrile Flow Rate 1.5 mL/min Injection volume 50 L Stop time 25.0 min Wavelength 284 nm, slit width 4 nm

TABLE-US-00005 TABLE 5 Solvent Gradient Parameters for HPLC Purity Analysis of MTC Time, min A, % B, % Flow, mL/min 0 90 10 1.5 1 90 10 1.5 13 75 25 1.5 18 40 60 1.5 20 40 60 1.5 20.1 90 10 1.5 25 90 10 1.5

[0478] HPLC standards and samples were prepared as follows: [0479] Fresh MTC reference material always used when preparing MTC stock and lower limit of quantification (LLOQ) standards. Stock and LLOQ standards were used for determination of retention time and quantification. [0480] 25 and 100 mL amber-glass volumetric flasks used to prepare standards and samples. [0481] Concentrated solutions were prepared using 34-38 mg of sample. The sample was dissolved in 50 mL of diluent (90:10, 0.1% TFA: acetonitrile), sonicated for 5 minutes, and then diluted to the graduation mark with diluent. Solutions were then allowed to stand for 1 hour prior to a 1:10 dilution. [0482] For runs, 2 L of 0.1% TFA and 1 L of acetonitrile was used for the eluents.

TABLE-US-00006 TABLE 6 Typical Retention Times for HPLC Purity Analysis of MTC (at 284 nm) Compound Retention time (minutes) Thionine 8.66 MVB-CH.sub.3 10.19 Azure C 10.92 MVB 11.72 Azure A 13.22 sDMT 13.50 Azure B 15.59 MTC 16.58

[0483] For reference, the chemical structures of MTC and the related impurities are shown in the following table.

TABLE-US-00007 TABLE 7 Chemical Structures of MTC and Related Impurities Methylthioninium chloride (MTC) [00030] Azure A [00031] Azure B [00032] Azure C [00033] Methylene Violet Bernthsen (MVB) [00034] 7-(methylamino)- 3H- phenothiazine- 3-one (MVB-CH.sub.3) [00035] 7-amino-3H- phenothiazine- 3-one (MVB-2CH.sub.3) [00036] Thionine [00037] Symmetrical Dimethyl Thionine (sDMT) [00038]

[0484] The foregoing has described the principles, preferred embodiments, and modes of operation of the present invention. However, the invention should not be construed as limited to the particular embodiments discussed. Instead, the above-described embodiments should be regarded as illustrative rather than restrictive. It should be appreciated that variations may be made in those embodiments by workers skilled in the art without departing from the scope of the present invention.

REFERENCES

[0485] A number of publications are cited herein in order to more fully describe and disclose the invention and the state of the art to which the invention pertains. Full citations for these references are provided below. Each of these references is incorporated herein by reference in its entirety into the present disclosure, to the same extent as if each individual reference was specifically and individually indicated to be incorporated by reference.

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