Peptide boronic acids useful in making salts thereof
Tripeptide boronic acids of (R,S,R) configuration, for example Cbz-(R)-Phe-(S)-Pro-(R)-Mpg-B(OH) 2 , and their use to make base addition salts of such acids. The salts are formulated into anti-thrombotic pharmaceutical formulations.
1 . An isolated compound selected from boronic acids of formula (IIIa):
where:
X is H (to form NH 2 ) or an amino-protecting group;
aa 1 is an amino acid having a hydrocarbyl side chain containing no more than 20 carbon atoms and comprising at least one cyclic group having up to 13 carbon atoms;
aa 2 is an imino acid having from 4 to 6 ring members;
R 9 is a straight chain alkyl group interrupted by one or more ether linkages and in which the total number of oxygen and carbon atoms is 3, 4, 5 or 6 or R 9 is —(CH 2 ) m —W where m is from 2, 3, 4 or 5 and W is —OH or halogen.
2 . A compound of claim 1 wherein aa 1 is selected from Dpa, Phe, Dcha and Cha.
3 . A compound of claim 1 wherein aa 1 is Phe or Dpa.
4 . A compound of claim 2 wherein aa 2 is a residue of an imino acid of formula (IV)
where R 11 is —CH 2 —, —CH 2 —CH 2 —, —S—CH 2 —, —S—C(CH 3 ) 2 — or —CH 2 —CH 2 —CH 2 —, which group, when the ring is 5- or 6-membered, is optionally substituted at one or more —CH 2 — groups by from 1 to 3 C 1 -C 3 alkyl groups.
5 . A compound of claim 4 wherein R 1 is a group of the formula —(CH 2 ) s -Z, where s is 2, 3 or 4 and Z is —OH, —OMe, —OEt or halogen.
6 . A compound of claim 5 wherein the boronic acid is of formula (VIII):
X-(R)-Phe-(S)-Pro-(R)-Mpg-B(OH) 2 (VIII).
7 . A compound of claim 5 where X is R 6 —(CH 2 ) p —C(O)—, R 6 —(CH 2 ) p —S(O) 2 —, R 6 —(CH 2 ) p —NH—C(O)— or R 6 —(CH 2 ) p —O—C(O)— wherein p is 0, 1, 2, 3, 4, 5 or 6 and R 6 is H or a 5 to 13-membered cyclic group optionally substituted by 1, 2 or 3 substituents selected from halogen, amino, nitro, hydroxy, a C 5 -C 6 cyclic group, a C 1 -C 4 alkyl group, a C 1 -C 4 alkyl group containing an in-chain O, a C 1 -C 4 alkyl group containing an in-chain O and linked to the cyclic group through an in-chain O and a C 1 -C 4 alkyl group linked to the cyclic group through an in-chain O, the aforesaid alkyl groups optionally being substituted by a substituent selected from halogen, amino, nitro, hydroxy and a C 5 -C 6 cyclic group, and optionally wherein said 5 to 13-membered cyclic group is aromatic or heteroaromatic.
8 . A compound of claim 5 wherein X is R 6 —(CH 2 ) p —C(O)— or R 6 —(CH 2 ) p —O—C(O)— and p is 0 or 1.
9 . A compound of claim 1 wherein the boronic acid is Cbz-(R)-Phe-(S)-Pro-(R)-Mpg-B(OH) 2 .
10 . A particulate composition comprising an isolated compound selected from boronic acids of formula (IIIa):
where:
X is H (to form NH 2 ) or an amino-protecting group;
aa 1 is an amino acid having a hydrocarbyl side chain containing no more than 20 carbon atoms and comprising at least one cyclic group having up to 13 carbon atoms;
aa 2 is an imino acid having from 4 to 6 ring members;
R 9 is a straight chain alkyl group interrupted by one or more ether linkages and in which the total number of oxygen and carbon atoms is 3, 4, 5 or 6 or R 9 is —(CH 2 ) m —W where m is from 2, 3, 4 or 5 and W is —OH or halogen.
11 . A compound selected from boronic acids of formula (IIIa):
where:
X is H (to form NH 2 ) or an amino-protecting group;
aa 1 is an amino acid having a hydrocarbyl side chain containing no more than 20 carbon atoms and comprising at least one cyclic group having up to 13 carbon atoms;
aa 2 is an imino acid having from 4 to 6 ring members;
R 9 is a straight chain alkyl group interrupted by one or more ether linkages and in which the total number of oxygen and carbon atoms is 3, 4, 5 or 6 or R 9 is —(CH 2 ) m —W where m is from 2, 3, 4 or 5 and W is —OH or halogen,
the compound being in a diastereomeric excess of at least about 98% over its (R,S,S) diastereomer.
12 . A compound of claim 11 which has a decrease in weight upon drying of less than 0.5% of its initial weight when dried in a vacuum drier at 40° C. at 100 mbar for 2 hours.
13 . A compound of claim 11 wherein the boronic acid is of formula (VIII):
X-(R)-Phe-(S)-Pro-(R)-Mpg-B(OH) 2 (VIII).
14 . A compound of claim 11 wherein
X is R 6 —(CH 2 ) p —C(O)—, R 6 —(CH 2 ) p —S(O) 2 —, R 6 —(CH 2 ) p —NH—C(O)— or R 6 —(CH 2 ) p —O—C(O)— wherein p is 0, 1, 2, 3, 4, 5 or 6 and R 6 is H or a 5 to 13-membered cyclic group optionally substituted by 1, 2 or 3 substituents selected from halogen, amino, nitro, hydroxy, a C 5 -C 6 cyclic group, a C 1 -C 4 alkyl group, a C 1 -C 4 alkyl group containing an in-chain O, a C 1 -C 4 alkyl group containing an in-chain O and linked to the cyclic group through an in-chain O and a C 1 -C 4 alkyl group linked to the cyclic group through an in-chain 0, the aforesaid alkyl groups optionally being substituted by a substituent selected from halogen, amino, nitro, hydroxy and a C 5 -C 6 cyclic group, and optionally said 5 to 13-membered cyclic group is aromatic or heteroaromatic;
aa 1 is selected from Phe, Dpa and wholly or partially hydrogenated analogues thereof;
aa 2 is a residue of an imino acid of formula (IV)
where R 11 is —CH 2 —, —CH 2 —CH 2 —, —S—CH 2 —, —S—C(CH 3 ) 2 — or —CH 2 —CH 2 —CH 2 —, which group, when the ring is 5- or 6-membered, is optionally substituted at one or more —CH 2 — groups by from 1 to 3 C 1 -C 3 alkyl groups;
R 1 is 2-bromoethyl, 2-chloroethyl, 2-methoxyethyl, 3-bromopropyl, 3-chloropropyl or 3-methoxypropyl and wherein the (R,S,R) isomer is in a diastereomeric excess of 99% or more over the (R,S,S) isomer.
15 . A compound of claim 14 wherein the boronic acid is a compound of claim 1 wherein the boronic acid is Cbz-(R)-Phe-(S)-Pro-(R)-Mpg-B(OH) 2 .
16 . A compound of claim 11 which is substantially free of degradation product derived from cleavage of the C—B bond thereof.
17 . A compound of claim 15 which has a purity measured as HPLC peak area of at least 97.5%, the % peak area being determined by the method of Example 43.
18 . A compound of claim 17 wherein the purity is at least 99%.
19 . Cbz-(R)-Phe-(S)-Pro-(R)-Mpg-B(OH) 2 when substantially free of the compound:
20 . A compound of claim 5 in which R 1 is a group of the formula —(CH 2 ) s -Z wherein Z is —OMe or —OEt and which is free of any compound which is of the same structure except for replacement of the R 1 group by a group of the formula —(CH 2 ) s —H.
21 . Cbz-(R)-Phe-(S)-Pro-(R)-Mpg-B(OH) 2 when substantially free of the compound:
22 . Cbz-(R)-Phe-(S)-Pro-(R)-Mpg-B(OH) 2 when in a diastereomeric excess of at least 99% over the (R,S,S) diastereomer, substantially free of the compound:
and free of the compound:
23 . A product comprising isolated, dry Cbz-(R)-Phe-(S)-Pro-(R)-Mpg-B(OH) 2 in a diastereomeric excess of 99% or more over its (R,S,S) diastereomer.
24 . A product of claim 23 which is sterile and otherwise adapted to be pharmaceutically acceptable.
25 . A process for making a pharmaceutically acceptable base addition salt of a boronic acid of formula (IIIa):
where:
X is H (to form NH 2 ) or an amino-protecting group;
aa 1 is an amino acid having a hydrocarbyl side chain containing no more than 20 carbon atoms and comprising at least one cyclic group having up to 13 carbon atoms;
aa 2 is an imino acid having from 4 to 6 ring members;
R 9 is a straight chain alkyl group interrupted by one or more ether linkages and in which the total number of oxygen and carbon atoms is 3, 4, 5 or 6 or R 9 is —(CH 2 ) m —W where m is from 2, 3, 4 or 5 and W is —OH or halogen selected from F, Cl, Br or I.
the process comprising contacting a boronic acid of formula (IIIa) which is a diastereomeric excess of at least about 98% over the (R,S,S) isomer with a base capable of making such a salt.
26 . The process of claim 25 wherein the salt is a metal salt
27 . The process of claim 25 wherein the salt is a salt of an alkali metal, an alkaline earth metal or zinc.
28 . The process of claim 25 wherein the base is an organic base having a pKb of 7 or more.
29 . A process for making a pharmaceutically acceptable base addition salt of an organoboronic acid inhibitor of thrombin having a neutral thrombin S1-binding moiety linked through a peptide linkage to a hydrophobic thrombin S2/S3-binding moiety, comprising combining a solution of the organoboronic add in a water-miscible organic solvent with an aqueous solution or suspension of the base, causing or allowing the acid and the base to react, and recovering the salt.
30 . A process of claim 29 in which the base comprises a cation of valency n and is used in a stoichiometry (boronic acid:base) of about n:1.
31 . A process of claim 30 in which the base is a metal base.
32 . A process of claim 31 in which the base is an alkali metal or alkaline earth metal base, or a zinc base.
33 . A process of claim 30 in which the base is an organic nitrogen-containing compound having a pKb of about 7 or more.
34 . A process of claim 29 wherein the solvent is acetonitrile.
35 . A process of claim 34 wherein the recovery of the salt comprises evaporating to dryness to obtain an evaporation residue and the process further comprises:
redissolving the evaporation residue in acetonitrile and evaporating the resulting solution to dryness; and
repeating the redissolution and evaporation steps as often as necessary to obtain a substantially dry evaporation residue.
36 . A process of claim 35 which further comprises:
dissolving the dry evaporation residue in acetonitrile or tetrahydrofuran to form a solution;
adding, at a rate sufficiently slow to avoid lump formation, said solution to a 3:1 to 1:3 v/v mixture of diethylether and an aliphatic or cycloaliphatic solvent to form a precipitate, said solution being added to the diethylether/(cyclo)aliphatic solvent mixture in a ratio (solution:mixture) of from 1:5 to 1:15 v/v;
recovering the precipitate; and
removing solvent from the recovered precipitate under reduced pressure whilst maintaining the temperature at no more than 35° C.
37 . A process of claim 36 in which the aliphatic or cycloaliphatic solvent is n-heptane.
38 . A process of claim 34 in which the base is sodium hydroxide.
39 . A process of claim 36 in which the base is sodium hydroxide and the dry evaporation residue is dissolved in acetonitrile.
40 . A process of claim 34 in which the base is calcium hydroxide.
41 . A process of claim 36 in which the base is calcium hydroxide and the dry evaporation residue is dissolved in tetrahydrofuran.
42 . A process of claim 29 wherein the solvent is tetrahydrofuran or an alcohol.
43 . A process claim 29 wherein the solvent is dry.
44 . A process of claim 29 wherein the boronic acid is of formula (I):
wherein
Y comprises a hydrophobic moiety which, together with the aminoboronic acid residue —NHCH(R 9 )—B(OH) 2 , has affinity for the substrate binding site of thrombin; and
R 9 is a straight chain alkyl group interrupted by one or more ether linkages and in which the total number of oxygen and carbon atoms is 3, 4, 5 or 6 or R 9 is —(CH 2 ) m —W where m is from 2, 3, 4 or 5 and W is —OH or halogen selected from F, Cl, Br or I.
45 . A process of claim 44 wherein R 9 is an alkoxyalkyl group and Y is an optionally N-terminally protected dipeptide which binds to the S3 and S2 binding sites of thrombin and the peptide linkages in the acid are optionally and independently N-substituted by a C 1 -C 13 hydrocarbyl optionally containing in-chain or in-ring nitrogen, oxygen or sulfur and optionally substituted by a substituent selected from halo, hydroxy and trifluoromethyl, and optionally wherein said dipeptide is N-terminally protected, all the peptide linkages in the acid are unsubstituted or the dipeptide is N-terminally protected and all the peptide linkages in the acid are unsubstituted.
46 . A process of claim 45 wherein the S3-binding amino acid residue is of (R)-configuration, the S2-binding residue is of (S)-configuration, and the fragment —NHCH(R 9 )—B(OH) is of (R)-configuration.
47 . A process of claim 29 wherein the boronic acid is of formula (II):
where:
X is H (to form NH 2 ) or an amino-protecting group;
aa 1 is an amino acid having a hydrocarbyl side chain containing no more than 20 carbon atoms and comprising at least one cyclic group having up to 13 carbon atoms;
aa 2 is an imino acid having from 4 to 6 ring members;
R 1 is a group of the formula —(CH 2 ) s -Z, where s is 2, 3 or 4 and Z is —OH, —OMe, —OEt or halogen selected from F, Cl, Br or I.
48 . A process of claim 47 wherein
X is R 6 —(CH 2 ) p —C(O)—, R 6 —(CH 2 ) p —S(O) 2 —, R 6 —(CH 2 ) p —NH—C(O)— or R 6 —(CH 2 ) p —O—C(O)— wherein p is 0, 1, 2, 3, 4, 5 or 6 and R 6 is H or a 5 to 13-membered cyclic group optionally substituted by 1, 2 or 3 substituents selected from halogen, amino, nitro, hydroxy, a C 5 -C 6 cyclic group, a C 1 -C 4 alkyl group, a C 1 -C 4 alkyl group containing an in-chain 0, a C 1 -C 4 alkyl group containing an in-chain 0 and linked to the cyclic group through an in-chain 0 and a C 1 -C 4 alkyl group linked to the cyclic group through an in-chain O, the aforesaid alkyl groups optionally being substituted by a substituent selected from halogen, amino, nitro, hydroxy and a C 5 -C 6 cyclic group, and optionally said 5 to 13-membered cyclic group is aromatic or heteroaromatic, e.g. is phenyl or a 6-membered heteroaromatic group, for example X is benzyloxycarbonyl;
aa 1 is selected from Phe, Dpa and wholly or partially hydrogenated analogues thereof, and optionally is selected from Dpa, Phe, Dcha and Cha;
aa 2 is a residue of an imino acid of formula (IV)
where R 11 is —CH 2 —, —CH 2 —CH 2 —, —S—CH 2 —, —S—C(CH 3 ) 2 — or —CH 2 —CH 2 —CH 2 —, which group, when the ring is 5- or 6-membered, is optionally substituted at one or more —CH 2 — groups by from 1 to 3 C 1 -C 3 alkyl groups;
R 1 is 2-bromoethyl, 2-chloroethyl, 2-methoxyethyl, 3-bromopropyl, 3-chloropropyl or 3-methoxypropyl; and
aa 1 is of (R)-configuration, aa 2 is of (S)-configuration and the fragment —NH—CH(R 1 )— B(OH) 2 is of (R)-configuration.
49 . A process of claim 29 wherein the boronic acid is of formula (VIII):
X-(R)-Phe-(S)-Pro-(R)-Mpg-B(OH) 2 (VIII).
50 . A solution whose solvent is a water-miscible organic solvent and which contains species selected from an organoboronic add inhibitor of thrombin having a neutral thrombin S1-binding moiety linked through a peptide linkage to a hydrophobic thrombin S2/S3-binding moiety, and equilibrium forms of the organoboronic acid.
51 . A solution of claim 50 wherein the solvent is acetonitrile.
52 . A solution of claim 50 wherein the solvent is an alcohol or tetrahydrofuran.
53 . A solution of claim 50 having the characteristics of a solution obtained by dissolving the free organoboronic acid in the solvent.
54 . A solution of claim 50 which consists essentially of the solvent, the organoboronic acid dissolved in the solvent and any species in equilibrium with the organoboronic acid.
55 . A solution of claim 50 wherein the organoboronic acid is of formula (I):
wherein
Y comprises a hydrophobic moiety which, together with the aminoboronic acid residue —NHCH(R 9 )—B(OH) 2 , has affinity for the substrate binding site of thrombin; and
R 9 is a straight chain alkyl group interrupted by one or more ether linkages and in which the total number of oxygen and carbon atoms is 3, 4, 5 or 6 or R 9 is —(CH 2 ) m —W where m is from 2, 3, 4 or 5 and W is —OH or halogen selected from F, Cl, Br or I.
56 . A solution of claim 50 wherein the boronic acid is of formula (II):
where:
X is H (to form NH 2 ) or an amino-protecting group;
aa 1 is an amino acid having a hydrocarbyl side chain containing no more than 20 carbon atoms and comprising at least one cyclic group having up to 13 carbon atoms;
aa 2 is an imino acid having from 4 to 6 ring members;
R 1 is a group of the formula —(CH 2 ) s -Z, where s is 2, 3 or 4 and Z is —OH, —OMe, —OEt or halogen selected from F, Cl, Br or I.
57 . A solution of claim 50 wherein the acid is Cbz-(R)-Phe-(S)-Pro-(R)-Mpg-B(OH) 2 and the solvent is acetonitrile.
58 . A solution of claim 50 wherein the acid is Cbz-(R)-Phe-(S)-Pro-(R)-Mpg-B(OH) 2 and the solvent is tetrahydrofuran or an alcohol.
59 . A process for making a pharmaceutical formulation, comprising performing the process of claim 25 and formulating the product salt into a pharmaceutical formulation.