WO2000058264A1

WO2000058264A1 - Method for the synthesis of non-symmetrical dtpa compounds

Info

Publication number: WO2000058264A1
Application number: PCT/US2000/007461
Authority: WO
Inventors: Samuel I. Achilefu; Ananthachari Srinivasan
Original assignee: Mallinckrodt Inc.
Priority date: 1999-03-26
Filing date: 2000-03-21
Publication date: 2000-10-05
Also published as: AU3766000A

Abstract

The synthesis of compounds of formula (XIV) wherein each R4 is an alkyl group having 1 to 15 carbon atoms, and R6 is a linking moiety having 1 to 10 carbon atoms, 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

METHOD FOR THE SYNTHESIS OF NON-SYMMETRICAL DTPA COMPOUNDS

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 have the general formula:

(I)

wherein R, and R₂ are H, and R₃, R,,, 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, 1 151-1 158 (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:

(vπi)

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

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

The synthesis described above suffers from several problems, the primary being that the major product of the first reaction is a waste product that must be disposed of. Another problem is that exhaustive chromatographic purification is required to isolate the minor product. This process further lowers the final yield of compound (VI) as a result of the inherent instability of compound (VI) on silica gel columns. The conventional method requires the preparation of several reaction intermediates which is time-consuming and may not be amenable to large-scale production.

Accordingly it would be desirable to have an alternative synthesis of these compounds.

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, Stereocontrolled 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.

SUMMARY OF THE INVENTION

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

(XIV)

In another respect the invention comprises a method of synthesis of compounds of the formula

(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 Rg and R₈ are linking moieties each having 1 to 10, desirably 1 to 6, preferably 1 to 4, and most preferably 2 carbon atoms; and each R₄ is 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.

The synthesis begins with the reaction of: H₂N R₆ NH ^τ X^' "COOR₄ ^κ9

(XI) (XII) wherein R₄, R^, are as defined above; W^ is a removable protecting group, desirably benzyl, trifluoroacetyl, methoxy benzyl, chlorotrityl, or nitrobenzyl, preferably benzyl, trifluoroacetyl, or methoxy benzyl, and most preferably benzyl; and X is a group that will react with the amines of compound (XI), desirably a halogen or a mesylate or a triflate, more desirably a halogen, preferably Cl or Br, and most preferably Br. This reaction produces:

(XIII)

When subjected to selective removal of the Rg group (for instance, by hydrogenation), the compound becomes:

(XIV)

Compound (XIV) is known from the conventional synthesis described in the

Background of the Invention, above. However, in this reaction it is produced as essentially the sole product, rather than as the minor product. From this point on, the synthesis is conventional. Compound (XIV) can be reacted with:

(XV) wherein R₈ is as defined above; both of R, are protecting groups that can be removed to leave a hydrogen, under conditions that do not remove any of R₄, desirably benzyl, allyl, methoxybenzyl, nitrobenzyl, or chlortrityl, more preferably benzyl allyl, methoxybenzyl, or nitrobenzyl, preferably benzyl or allyl, and most preferably benzyl; and Z is a group that will react with the secondary amine of compound (XIV), desirably a halogen or a mesylate or a triflate, preferably Cl or Br, and most preferably Br; to produce:

(XVI) This compound can then be subjected to hydrogenation to replace both R_{ moieties with hydrogens, to yield compound (X).

This new procedure has several advantages over the prior art. Some of these include the use of fewer reaction steps, cheaper starting materials and high yield of the critical secondary amine (compound XIV). This method is also amenable to large-scale production and it is applicable to the production of a variety of DTPA derivatives.

EXAMPLE 1

Synthesis of 2-[Bis-(benzyloxycarbonylmethyl)amino]ethyl bromide (Compound (VII) or Compound (XV) where Z = Br, R₈ = ethyl, and Rj = benzyl). A solution of 50 ml of dimethylformamide and benzyl bromoacetate (16.0 g, 70 mmol) was stirred in a 100 ml three-neck flask. Solid potassium bicarbonate (7.8 g, 78 mmol) was added. The flask was purged with argon and cooled to 0°C with an ice bath. To the stirring mixture was added dropwise a solution of ethanolamine (1.9 g, 31 mmol) and 4 ml of dimethylformamide over 5 minutes. After the addition was complete the mixture was stirred for 1 hour at 0°C. The ice bath was removed and the mixture stirred at room temperature over night. The reaction mixture was partitioned between 100 ml of methylene chloride and 100 ml of saturated sodium bicarbonate solution. The layers were separated and the methylene chloride layer was again washed with 100 ml of saturated sodium bicarbonate solution. The combined aqueous layers were extracted twice with 25 ml of methylene chloride. The combined methylene chloride layers were washed with 100 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 with vacuum at about 45°C. The crude material was left on a vacuum line over night at room temperature. The crude material from above was dissolved in 100 ml of methylene chloride at room temperature. Triphenylphosphine (8.91 g, 34 mmol) was added and dissolved with stirring. An argon purge was started and the mixture cooled to 0°C with an ice bath. The N-bromosuccinimide (6.05 g, 34 mmol) was added portionwise over 2 minutes. The mixture was stirred for 1.5 hours at 0°C. The methylene chloride was removed with vacuum and gave a purple oil. This oil was triturated with 200 ml of ether with constant manual stirring. During this time the oil became very thick. The ether solution was decanted and the oil was triturated with 100 ml of ether. The ether solution was decanted and the oil was again triturated with a 100 ml portion of ether. The ether was decanted and the combined ether solutions allowed to stand for about 2 hours to allow the triphenylphosphine oxide to crystallize. The ether solution was decanted from the crystals and the solid washed with 100 ml of ether. The volume of the combined ether abstracts was reduced with vacuum until a volume of about 25 ml was obtained. This was allowed to stand over night at 0°C. Ether (10 ml) was added to the cold mixture which was mixed to suspend the solid. The mixture was percolated through a column of 45 g of silica gel and eluted with ether, 75 ml fractions were collected. The fractions that contained product by TLC were pooled and the ether removed with vacuum. This gave 10.1 g of crude product. The material was flash chromatographed on silica gel with hexane, changing to 9: 1 hexane:ether. The product-containing fractions were pooled and the solvents removed with vacuum. This gave 7.4 g (57% yield) of pure product.

EXAMPLE 2 Synthesis of N-benzyl-N,N',N'-tris(t-butyloxycarbonylmethyl) ethylenediamine (Compound (XIII) where R₄ = t-butyl, R^ = ethyl, and R, = benzyl). N-Benzylethylenediamine (Compound (XI) where R^ = ethyl and Rg = benzyl) (5g, 33.28 mmol) and potassium bicarbonate (19.3g, 139.7 mmol) were added to 200 ml of anhydrous acetonitrile and stirred vigorously under argon, t- Butyl bromoacetate (Compound (XII) where X = Br and R₄ = t-butyl) was diluted in 30 ml of anhydrous acetonitrile and the solution was added dropwise to the reaction mixture over 90 minutes. The progress of the reaction was monitored by thin-layer chromatography and was essentially complete in about 4 hours but was stirred at room temperature for about 12 hours in order to assure complete alkylation of the amine. The insoluble residue was filtered and washed with acetonitrile. The filtrate was evaporated to give 20g of a yellow liquid. Hexane (100 ml) was added to the crude mixture and stirred vigorously until white precipitate formed. The precipitate was filtered and the filtrate was evaporated to give a yellow liquid. The pure compound was obtained by washing the crude product over dry flash chromatographic column and the desired compound was eluted with 10% diethyl ether in hexane (12.5 g, 80% yield).

EXAMPLE 3

Synthesis of N,N',N'-tris(t-butyloxycarbonylmethyI)ethylenediamine (Compound (VI) or Compound (XIV) where R₄ = t-butyl and Rg = ethyl). N- benzyl-N,N'N'-tris(t-butyloxycarbonylmethyl) ethylenediamine (from Example 2) (6 g, 12 mmol) was added to a heterogeneous mixture of 10% palladium on carbon (6 g, 1 weight equivalent) in 100 ml of methanol. Anhydrous ammonium formate (3.8 g, 60.26 mmol) was added to the reaction mixture in one bulk. The mixture was stirred at room temperature for 2 hours. The mixture was filtered over celite and the residue was washed with chloroform. The filtrate was evaporated until white precipitates began to form. The residue was triturated in chloroform and the insoluble formate was filtered. Evaporation of the filtrate gave a pale yellow liquid (4.6 g, 96% yield) which was identified as the pure compound by NMR analysis.

EXAMPLE 4

Synthesis of N,N',N'-tris(t-butyloxycarbonylmethyl)-N",N"- bis(benzyloxycarbonylmethyl)diethylenetriamine (Compound (VIII) or

Compound (XVI) where Rj = benzyl, R₄ = t-butyl, and R₆ = ethyl). A mixture of N,N'N'-tris(t-butyloxycarbonylmethyl) ethylenediamine (4.4 g, 9.82 mmol) and 2-[Bis-(benzyloxycarbonylmethyl)amino]ethyl bromide (5.3 g, 12.76 mmol) was added to a solution of ethydiisopropylamine (3.8 g, 29.45 mmol) in 100 ml acetonitrile. The mixture was stirred at reflux for 24 hours under nitrogen. After the reaction was complete, the solvent was evaporated and the residue was partitioned between dichloromethane (100 ml) and distilled water (100 ml). The organic layer was washed with 100 ml of water and 100 ml of brine. It was dried over magnesium sulfate and the solvent was evaporated to give about 10 g of the crude product. The product was purified by dry flash chromatography and the pure compound was eluted with 40% of diethyl ether in hexane as a pale yellow liquid (6.5 g, 90% yield).

EXAMPLE 5

Preparation of N,N',N'-tris(t-butyloxycarbonylmethyl)-N",N"-bis(acetic acid)diethylenetriamine (II). A mixture of 10% palladium on carbon (0.21 g) and a solution of N,N',N'-tris(t-butyloxycarbonylmethyl)-N",N"- bis(benzyloxycarbonylmethyl)diethylenetriamine (3.3 g, 4.45 mmol) in 50 ml of methanol was hydrogenolyzed at 40 psi for 2 hours. The mixture was filtered over celite and the residue was washed with methanol. The solvent was evaporated to give an off-white powder which was shown by mass spectral analysis, HPLC and NMR to be the pure compound (2.4 g, 96% yield).

EXAMPLE 6

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 tri-t-butyl DTPA (compound II) 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 tri-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: DTP A-D-Phe-Cys(Acm)-Tyr-D-T -Lys-The-Cys(Acm)-Thr.

-π-

Claims

What is claimed is:

1. A method of synthesizing a compound of the formula:

(XIV) wherein each R₄ is an alkyl group having 1 to 15 carbon atoms, and R^ is a linking moiety having 1 to 10 carbon atoms, comprising

(a) reacting together compounds of the formulas:

H₂N R₆ NH ⁺ X COOR₄ ^R9

(XI) (XII) wherein R^ and Rg, are as defined above, R₉ is a removable protecting group, and X is a group that will react with the amines of compound (XI), to produce a compound of the formula:

(XIII) and;

(b) selectively removing Re, to produce compound (XIV).

2. The method of claim 1 wherein each R₄ has 2 to 8 carbon atoms, R has 1 to 6 carbon atoms, R₉ is benzyl, trifluoroacetyl, methoxy benzyl, chlorotrityl, or nitrobenzyl, and X is a halogen, a mesylate, or a triflate.

3. The method of claim 2 wherein each R₄ has 3 to 6 carbon atoms, R^ has 1 to 4 carbon atoms, Re, is benzyl, trifluoroacetyl, or methoxy benzyl, and X is a halogen.

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

5. A method of preparing a compound of the formula

(X) wherein each R₄ is an alkyl group having 1 to 15 carbon atoms, and Rg and R₈ are each linking moieties having 1 to 10 carbon atoms, comprising (a) reacting together compounds of the formulas:

H₂N R₆ NH

\ + X^' "COOR₄

R₉

(XI) (XII) wherein R^ and Rg, are as defined above, Rς, is a removable protecting group, and X is a group that will react with the amines of compound (XI), to produce a compound of the formula:

(XIII);

(b) selectively removing Rg to produce a compound of the formula:

(XIV);

(c) reacting compound (XIV) with a compound of the formula:

(XV) wherein R₈ is as defined above; both of R_{ are protecting groups that can be removed to leave a hydrogen, under conditions that do not remove any of R₄; and Z is a group that will react with the secondary amine of compound (XIV), to produce a compound of the formula:

(XVI) and; (d) selectively replacing both R moieties with hydrogens, to yield compound

(X).

6. The method of claim 5 wherein R, is benzyl, allyl, methoxybenzyl, nitrobenzyl, or chlortrityl,, each R₄ has 2 to 8 carbon atoms, R_$ and R₈ each have 1 to 6 carbon atoms, Rg is benzyl, trifluoroacetyl, methoxy benzyl, chlorotrityl, or nitrobenzyl, and X is a halogen, a mesylate, or a triflate.

7. The method of claim 6 wherein R, is benzyl, or allyl, each R₄ has 3 to 6 carbon atoms, R-g and R₈ each have 1 to 4 carbon atoms, R^ is benzyl, trifluoroacetyl, or methoxy benzyl, and X is a halogen.

8. The method of claim 7 wherein R, is benzyl, each R₄ is t-butyl, Rg and R₈ are each ethyl, Rg is benzyl, and X is Br.

9. The method of claim 5 wherein Rj is an alkyl group having 2 to 8 carbon atoms and R₄ is benzyl, trifluoroacetyl, methoxy benzyl, chlorotrityl, or nitrobenzyl.

10. The method of claim 5 wherein R] is t-butyl and R₄ is benzyl.