WO2000058267A1 - Enhanced selectivity in the synthesis of dtpa esters fom diethylenetriamine - Google Patents

Abstract

The synthesis of compounds of formula (XI) is disclosed. These compounds are useful intermediates in the synthesis of compounds of formula (X). These compounds are useful in the preparation of nuclear pharmaceuticals such as those used for tumor imaging or tumor therapy.

Description

ENHANCED SELECTIVITY IN THE SYNTHESIS OF DTPA ESTERS FROM DIETHYLENETRIAMINE

BACKGROUND OF THE INVENTION This invention relates to the synthesis of esters of diethylenetriaminepentaacetic acid (DTPA) and of intermediates useful in the synthesis of such esters. These esters are well known, and have the general formula:

(I)

wherein R, and R₂ are H, and R₃, R₄, and R₅ are t-butyl (compound (II)) or a similar protecting group. Such compounds are useful in the preparation of nuclear pharmaceuticals where they serve as a metal chelate and a link between a peptide and a radionuclide. Activation of the free dicarboxylic acid rapidly forms intramolecular acid anhydride which then reacts with the amino group on a peptide to form the DTPA-peptide conjugate. Acid mediated cleavage of the esters gives free tetra-carboxylic acid which readily forms stable metal complexes with the radionuclide of choice.

Using conventional techniques, these compounds are prepared by the following sequence (t-Bu = t-butyl):

(III) (IV)

Compound (IV) is commercially available. Compound (III) was prepared by the method taught in Rapoport, J. Org. Chem. 1993, 58, 1151-1158 (incorporated herein by reference). This reaction yields two compounds in about a 1 :4 ratio. The major product is the penta ester (compound (V)), (i.e.: compound (I) in which all of R_rR₅ = t-butyl). This product is useless and must be disposed of. The minor product is:

(VI)

Compound (VI) is reacted with the compound (Bz = benzyl):

(VII)

This compound is not commercially available, but may be synthesized by literature method as described by Rapoport (see the reference for compound (III) above).

The reaction product of (VI) and (VII) is:

(VIII)

This compound is subjected to catalytic hydrogenation at room temperature to yield

(II) (i.e.: compound (I) where Rj and R₂ are H and R₃, R₄, and R₅ are t-butyl).

Although these compounds are useful, their syntheses are often complicated and expensive. Further, it would be useful to have other compounds available with different binding affinities.

US 5,618,513 (Mallinckrodt: A. Srinivasan) teaches a general method for using DTPA compounds in the preparation of radiopharmaceuticals. This reference also teaches the preparation of such compounds as outlined above.

US 4,479,930 (Univ. of Massachusetts: D. Hnatowich) teaches the use of a dicyclic dianhydride compound to couple polypeptides. Although useful, this method results in diaddition products which are not medically useful.

S. Ram and L. Spicer, Rapid Debenzylation of N-Benzylamino Derivatives to Amino-Derivatives Using Ammonium Formate as Catalytic Hydrogen Transfer Agent, Tetrahedron Letters, Vol. 28, No. 5, pp 515-516, 1987, teaches the deprotection of various N-benzyl compounds using ammonium formate as the hydrogen source.

S. Ram and L. Spicer, Debenzylation of N-Benzylamino Derivatives by Catalytic Transfer Hydrogenation with Ammonium Formate, Synthetic Communication, 17(4), 415-418 ( 1987), is similar to the first Ram and Spicer reference. C. Grote, D. Kim, and H. Rapoport, Stereo controlled Synthesis of DTPA Analogues Branched in the Ethylene Unit, J. Org. Chem., 1995, 60, 6987-6997, teaches a synthesis similar to that outlined above, with the addition of stereo control of the reaction.

M. Brechbiel and O. Gansow, Backbone-Substituted DTPA Ligands for ⁹⁰Y Radioimmunotherapy, Bioconjugate Chem. 1991, 2, 187-194, teaches the synthesis of new bifunctional DTPA ligands.

US 5,514,810 (Schering: J. Platzek et al), disclose some of the compounds which may be synthesized by this invention. However, this reference teaches a different method of synthesis.

WO 98/05626 (Bracco: P. L. Anelli, et al.) teaches compounds that are similar to the compounds made by the instant invention.

SUMMARY OF THE INVENTION

Briefly, the invention comprises a method of synthesis of compounds of the formula

(XI) and

(X) DETAILED DESCRIPTION OF THE INVENTION

In this specification and claims, numerical values and ranges are not critical unless otherwise stated. That is, the numerical values and ranges may be read as if they were prefaced with the word "about" or "substantially".

The invention includes a method for synthesizing compounds of the formula:

(X) wherein R, and R₈ are each a linking moiety having 1 to 10, desirably 1 to 6, preferably 1 to 4, and most preferably 2 carbon atoms; each R₄ is a removable protecting group, generally (a) an alkyl group having 1 to 15, desirably 2 to 10, more desirably 2 to 8, preferably 3 to 6, and more preferably 4 carbon atoms, and most preferably being t-butyl or (b) benzyl or a benzyl derivative such as methoxy benzyl or nitrobenzyl, preferably benzyl. If R₄ is t-butyl or a similar group, the compound (X) is more useful in fluorenylmethoxycarbonyl (Fmoc) peptide synthesis, and if R₄ is benzyl or a similar group, the compound (X) is more useful in acid labile t-butoxycarbonyl (Boc) peptide synthesis; and R₉ is hydrogen or a to C₅₀ alkyl moiety such as that taught by US 5,514,810 (incorporated herein by reference), preferably hydrogen.

The synthesis begins with the reaction of:

H H₂N R₆ N R₈ NH₂

wherein R₆ and R₈ are as defined above, with a compound Z which is any protecting group that will react with primary amines but not (under the same conditions) with secondary amines. Z is desirably trifluoroacetyl in the presence of crown ethers, 2-acetyldimedone (Dde) , or phthaloyl (Pth), preferably Pth for Fmoc procedures and preferably Dde for Boc procedures. Compound (XII) is commercially available.

The reaction yields:

H ZN R₆ N R₈ NZ

(XIII) Compound (XIII) can be reacted with the compound:

(XIV) wherein R₉ is hydrogen or a C, to C₅₀ alkyl moiety, desirably such as that taught by US 5,514,810 (incorporated herein by reference), preferably hydrogen; and R₅ is a removable protecting group different from and removable separately from R₄(defmed below), generally (a) t-butyl, allyl, or chlorotrityl, preferably t-butyl or (b) allyl, benzyl, or a benzyl derivative such as methoxy benzyl or nitrobenzyl, preferably benzyl or methoxy benzyl, and most preferably benzyl; and X is a group that will react with the amine of compound (XIII), desirably a halide, a mesylate, or a triflate, more desirably a halide, preferably Cl or Br, and most preferably Br. If R₄ is t-butyl or a similar group, R₅ is preferably benzyl or a benzyl derivative, and if R₄ is benzyl or a similar group, R₅ is preferably t-butyl or allyl. The reaction product is:

R₉ - OORg

ZN R₆ N R₈ NZ

(XV) Compound (XV) is then subjected to selective removal of the Z groups, for instance by hydrolysis. The result is:

(XVI)

Compound (XVI) is then reacted with a compound of the formula:

X^' COOR₄ (XVII) wherein X is a group that will react with the amines of compound (XVI), desirably a halide, a mesylate, or a triflate, more desirably a halide, preferably Cl or Br, and most preferably Br; and R₄ is as defined above. The resulting compound is:

(XI)

Selective removal of R₅, yields:

(X)

The invention is further illustrated in the following examples.

EXAMPLE 1

Synthesis of bis-N,N-N",N"-(phthalimido)diethylenetriamine N,N,N",N"-bis(2- acetyldimedone)diethylenetriamine (Compound (XIII) where R₆ and R₈ = ethylene and Z = phthaloyl). A mixture of phthalic anhydride (lOg, 67.51 mmol) and diethylenetriamine (3.17 g, 30.72 mmol) in 100 ml of glacial acetic acid was refluxed for 2 hours. The mixture was allowed to cool to room temperature and hexane (100 ml) was added to the reaction mixture and the hexane-acetic acid mixture was evaporated en vacuo as an azeotrope. More hexane was added and the mixture evaporated. This procedure was repeated until most of the acetic acid was removed. The orange residue obtained was dissolved in dichloromethane (100 ml) and washed with saturated aqueous sodium bicarbonate until the aqueous phase became alkaline (pH indicator paper). The organic phase was then washed with brine (100 ml) and dried over magnesium sulfate. Evaporation of the solvent gave 13 g of the crude product which was heated in methanol until a fine solid dispersion was obtained. The solid powder was filtered and washed with methanol (100 ml). It was dried under vacuum to give the pure product as a pale yellow powder (9 g, 81%).

EXAMPLE 2

Synthesis of N'-(benzyloxycarbonylmethyl)-bis-N,N-N",N"-(phthalimido) diethylenetriamine (Compound (XV) where R₆ and R₈ = ethylene, Z = phthaloyl, R₉ = H and Rs = benzyl). A mixture of N,N-N",N"-bis( phthalimido) diethylenetriamine (4 g, 11 mmol), benzyl bromoacetate (3 g, 13.22 mmol), and diisopropylethylamine (2 g, 15.42 mmol) in 60 ml of acetonitrile was refluxed for 24 hours. The solvent was evaporated and dichloromethane (50 ml) was added to the residue. The solution obtained was washed with water (2 x 50 ml) and saturated sodium bicarbonate until a clear aqueous phase was obtained. The organic layer was finally washed with brine (2 x 50 ml) and dried over magnesium sulfate. Evaporation of the solvent gave a viscous liquid which precipitated at room temperature as a yellow solid (5.2 g, 92%).

EXAMPLE 3

Synthesis of N,N,N",N"-bis(2-acetyIdimedone)diethylenetriamine (Compound

(XIII) where R₆ and R₈ = ethylene and Z = 2-Acetyldimedone). Reflux a mixture of 2-Acetyldimedone (49.2 g, 270 mmol), diethylenetriamine (4.6 g, 45 mmol) and triethylamine (27.3 g, 270 mmol) in anhydrous ethanol (300 ml) for 12 hours. Evaporate the solvent and redissolve the residue in ethyl acetate (200 ml). Wash the organic phase with 5% aqueous sodium bicarbonate (3 x 50 ml) and dry the ethyl acetate layer over magnesium sulfate. Evaporate the solvent to obtain a viscous oil which can be crystallized with hexane/ether.

EXAMPLE 4

Preparation of N'-benzyloxycarbonylmethyl-N,N,N",N"-bis(2- acetyldimedone) diethylenetriamine (Compound (XV) where R₆ and R₈ = ethylene, R₉ = H, R₅ = benzyl and Z = 2-Acetyldimedone). Reflux a mixture of benzyloxycarbonylmethyl-N,N,N",N"-bis(2-acetyldimedone)diethylenetriamine (15.4 g, 35.7 mmol), benzyl bromoacetate (12 g, 53.6 mmol) and diisopropylethylamine (10 g, 78.6 mmol) in 300 ml of acetonitrile for 16 hours. Evaporate the solvent and purify the residue on a silica gel column by dry flash chromatography. Elute the pure compound with 30% of diethyl ether in hexane.

EXAMPLE 5

Synthesis of N'-benzyloxycarbonylmethyldiethylenetriamine (Compound (XVI) where 1^ and R₈ = ethylene, R₉ = H, and R₅ = benzyl). Prepare a solution of 2% (w/v) hydrazine monohydrate in dimethylformamide (50 ml). Add N'- benzyloxycarbonylmethyl-N,N,N",N"-bis(2-acetyldimedone)diethylenetriamine (5 g, 7.65 mmol) to the hydrazine solution at room temperature. Stir the mixture for 60 minutes and concentrate the residue to dryness. Dissolve the residue in 0.1 M aqueous HC1 and wash the solution with diethyl ether. Carefully adjust the pH of the aqueous solution to pH 8 with 0.5 M aqueous sodium hydroxide. Wash this solution with diethyl ether and evaporate the solvent. Add ethanol to precipitate inorganic salts and filter off the solid residue. Dry the filtrate over potassium hydroxide pellets and evaporate the ethanol to give the desired diamine. EXAMPLE 6

Synthesis of N'-benzyloxycarbonylmethyl-N,N,N",N"-tetrakis(t- butyloxycarbonylmethyl)diethylenetriamine (Compound (XI) where R₆ and R₈ = ethylene, R₉ = H, R₄ = t-Butyl and R₅ = benzyl). A solution of t-butyl bromoacetate (1.4 g, 7.1 mmol) in 10 ml of anhydrous dimethylformamide was stirred in a 50 ml three-neck flask. Solid potassium bicarbonate (0.8 g, 7.8 mmol) was added. The flask was purged with argon and cooled to 0°C in an ice bath. To the stirring mixture was added dropwise a solution of N'- benzyloxycarbonylmethyldiethylenetriamine (1.0 g, 1.4 mmol) in 1 ml of dimethylformamide. After the addition was complete, the mixture was stirred for 20 minutes at 0°C. The ice bath was removed and the mixture stirred at room temperature for 6 hours. The reaction mixture was partitioned between 10 ml of methylene chloride and 10 ml of saturated sodium bicarbonate solution. The layers were separated and the methylene chloride layer was again washed with 5 ml of saturated sodium bicarbonate solution. The combined aqueous layers was extracted twice with 5 ml of methylene chloride. The combined methylene chloride layers was washed with 10 ml of brine, and dried over magnesium sulfate. The methylene chloride was removed with aspirator vacuum at ca. 35°C, and the remaining dimethylformamide was removed under vacuum at about 45°C. The crude product obtained was purified by dry flash chromatography on silica gel column and the pure compound was eluted with 30% ethyl acetate in hexane.

EXAMPLE 7 Preparation of N'-acetic acid- N,N,N",N"-tetrakis(t-butyloxycarbonylmethyl) diethylenetriamine (Compound (X) where R₆ and R₈ = ethylene, R₄ = t-Butyl,

R₉ and R₅ = H). A mixture of 10% palladium on carbon (0.1 g) and N'- (benzyloxycarbonylmethyl)-N,N,N",N"-tetrakis(t-butyloxycarbonylmethyl) diethylenetriamine (0.6 g, 0.85 mmol) in 30 ml of methanol was hydrogenolyzed at 45 psi for 2 hours. The mixture was filtered over celite and the residue was washed with methanol (50 ml). The solvent was evaporated to give the pure mono- carboxylic acid (0.5 g, 96%) as a viscous pale yellow oil.

EXAMPLE 8

Synthesis of DTPA-Octreotate derivative. The DTPA-Octreotate conjugate was prepared by solid phase synthesis using pre-loaded fluorenemethoxycarbonyl- threonine (Fmoc-Thr) Wang resin on 0.025 mmol scale. Commercially available automated peptide synthesizer from Applied Biosystems (Model 432A SYNERGY Peptide Synthesizer) was used. Cartridges containing Fmoc-protected amino acids were used in the solid phase synthesis. Cysteines were protected with acetamidomethyl group. Coupling reaction was carried out with 0.075 mmol of the protected amino acid and 2-(lH-benzotriazole-lyl)-l,l,3,3-tetramethyluronium hexafluorophosphate (HBTU)/N-hydroxybenzotriazole (HOBT). The amino acids and tetra-t-butyl DTPA (compound X) cartridges were placed on the peptide synthesizer and the product was synthesized from the C-terminal to the N-terminal position. After the synthesis was completed, the product was cleaved from the solid support with a cleavage mixture containing trifluoroacetic acid (85%):water (5%):phenol (5%):thioanisole (5%>) for 6 hours. Note that the t-butyl esters of tetra- t-butyl DTPA were also cleaved to give the free tetra-carboxylic acid. The DTPA- peptide conjugate was precipitated with t-butyl methyl ether and lyophilized with water : acetonitrile (2/3) mixture. Mass spectral analysis indicated that only the mono peptide-DTPA conjugate was obtained in accordance with the following sequence: DTPA-D-Phe-Cys(Acm)-Tyr-D-T -Lys-The-Cys(Acm)-Thr.

Claims

What is claimed is:

1. A method of synthesizing a compound of the formula

(XI) wherein each R₄ is a removable protecting group; R₅ is a removable protecting group different from and removable separately from R₄; each R and R₈ is a linking moiety having 1 to 10 carbon atoms; and R₉ is hydrogen or a Ci to C₅₀ alkyl moiety; comprising reacting together compounds of the formulas

(XVI) and

X^{' ^}COOR₄ (XVII) wherein X is a group that will react with the amines of compound (XVI)

2. The method of claim 1 wherein each R₄ is (a) an alkyl group having 1 to 15 carbon atoms or (b) benzyl or a benzyl derivative; if R₄ is an alkyl group then R₅ is allyl, benzyl, or a benzyl derivative, and if R₄ is benzyl or a benzyl derivative, then R₅ is t-butyl, chlorotrityl, or allyl; each Rg has 1 to 6 carbon atoms; R₉ is hydrogen; and X is a halide, a mesylate, or a triflate.

3. The method of claim 2 wherein each R₄ is (a) an alkyl group having 3 to 6 carbon atoms or (b) benzyl or a benzyl derivative; if R₄ is an alkyl group then R₅ is allyl, benzyl, or a benzyl derivative, and if R₄ is benzyl or a benzyl derivative, then R₅ is t-butyl, chlorotrityl, or allyl; each Rg has 1 to 4 carbon atoms; and X is a halogen.

4. The method of claim 3 wherein each R₄ is t-butyl; R₅ is benzyl, Rg is ethyl; and X is Br.

5. The method of claim 3 wherein each R₄ is benzyl; R₅ is t-butyl, Rg is ethyl; and X is Br.

6. A method of synthesizing a compound of the formula

(X) wherein each R₄ is a removable protecting group; each Rg and R₈ is a linking moiety having 1 to 10 carbon atoms; and R₉ is hydrogen or a C, to C₅₀ alkyl moiety comprising, reacting a compound of the formula

(XI) wherein R₅ is a removable protecting group different from and removable separately from R₄; under conditions such that R₅ is replaced by a hydrogen.

7. The method of claim 6 wherein the conditions such that R₅ is replaced by a hydrogen comprise a hydrogenation reaction.

8. A method of synthesizing a compound of the formula

(XVI) wherein R₅ is a removable protecting group; each Rg and R₈ is a linking moiety having 1 to 10 carbon atoms; and R₉ is hydrogen or a to C₅₀ alkyl moiety; comprising reacting together compounds of the formula

(XIII) and

wherein X is a group that will react with the amine of compound (XIII); to obtain the product

(XV) and selectively removing the Z groups to obtain compound (XVI).