WO2008115337A1

WO2008115337A1 - Sulfur-protected mercaptoacetylglycylglycylglycine

Info

Publication number: WO2008115337A1
Application number: PCT/US2008/002455
Authority: WO
Inventors: Dennis L. Nosco; Raghavan Rajagopalan
Original assignee: Mallinckrodt Inc.
Priority date: 2007-03-19
Filing date: 2008-02-25
Publication date: 2008-09-25

Abstract

Novel sulfur-protected mercaptoacetylglyclglycl-amino acid ligands are provided that are useful in synthesizing radiopharmaceuticals that can be utilized as renal imaging agents in nuclear medicine. In one aspect, sulfur-protected mercaptoacetylglyclglyclglycine ligands are disclosed. The sulfur-protecting agents are selected from transition metal cations and organic moieties.

Description

SULFUR-PROTECTED MERCAPTOACETYLGLYCYLGLYCYLGLYCINE

[0001] FIELD OF THE INVENTION

[0002] The present invention generally relates to sulfur-protected mercaptoacetylglycylglycyl-amino acid compositions and to the preparation of radiopharmaceuticals therefrom.

[0003] BACKGROUND OF THE INVENTION

[0004] Mercaptoacetylglycylglycyl-amino acid compounds are ligands that can be complexed with radioisotopes to form radiopharmaceuticals that are used to study the kidneys and other organs. A common mercaptoacetylglycylglycyl-amino acid compound that is used as a ligand in the formation of such radiopharmaceuticals is mercaptoacetylglycylglycylglycine ("MAG3"). MAG3, when complexed with the radioisotope Technetium-99m, is routinely utilized in renal function procedures as a nuclear medicine imaging agent to image a patient's kidneys. Several renal functioning procedures that utilize mercaptoacetylglycylglycyl- amino acid radiopharmaceuticals are known in the art. Examples of several such procedures are described by Fritzberg et al. in U.S. Patent Nos. 4,980,147; 5,322,929; and 5,572,748.

[0005] Technetium-99m ("Tc-99m") is a radioisotope that is commonly used in renal imaging radiopharmaceuticals due to its ideal imaging properties. However, due to its short half-life, radiopharmaceuticals comprising Tc-99m are typically synthesized at a medical facility on the day the radiographic procedure is performed. Kits are often used to synthesize Tc-99m radiopharmaceutical complexes. The kits typically contain a ligand for complexation with Tc-99m, a reducing agent, an intermediate exchange ligand, and other ingredients that can promote the stability of the kit. The kits can be lyophilized to enhance the shelf life of the kits and permit kits to be stored for periods of time until they are used to synthesize a radiopharmaceutical complex. While many radiopharmaceuticals for kidney imaging have been reported in the literature, radiopharmaceuticals comprising Tc-99m-MAG3, in particular, have ideal renal extraction properties that provide superior renal images when conducting renal function diagnostic procedures.

[0006] MAG3 radiopharmaceutical preparation kits typically contain a premeasured amount of benzoyl mercaptoacetylglycylglycylglycine, a reducing agent, an intermediate exchange ligand, and optionally other ingredients to promote the stability of the compound and its reaction with the radioisotope. MAG3 kits can be mixed with Tc-99m radioisotope solution and heated to produce a Tc-99m-MAG3 complex that is administered to a patient.

[0007] SUMMARY OF THE INVENTION

[0008] Briefly, the present invention is directed to protecting groups for thiol groups in nitrogen- sulfur ligands to protect the sulfur atom from oxidation or from other unwanted side reactions.

[0009] In one aspect, the present invention is directed to sulfur-protected MAG3 compounds of Formula 1,

Formula 1

wherein X is selected from the group consisting of -S Mⁿ⁺Ui, and -SR; Mⁿ⁺ is a transition metal cation; R is an organic moiety selected from the group consisting of acyl, alkoxyalkyl, alkoxycarbonyl, 2-pyrrolinyl, 2-pyrrolidinyl, 2-furanyl, 2-pyranyl, and disulfide; Y is -OH₁ or -O; Ui is a Lewis base; and n varies from 1 to 3.

[0010] In another aspect, the invention is directed to a kit for use in preparing a radiopharmaceutical compound, and water-soluble salts thereof, comprising technetium-99m. The kit containing ingredients comprising an amount of the compound of Formula 1 and an amount of a water-soluble reducing agent complexed with an intermediate exchange ligand. The ingredients, when mixed with a water soluble Tc- 99m pertechnetate salt, react to form a diagnostically effective amount of the radiopharmaceutical compound having the formula:

[0011] In another aspect, the invention is directed to a method of preparing a radiopharmaceutical compound, and water-soluble salts thereof, comprising technetium-99m. The method comprising reacting the sulfur-protected compound of Formula 1 with a reducing agent and a water-soluble Tc- 99m pertechnetate salt.

[0012] Other aspects and features of this invention will be in part apparent and in part pointed out hereinafter.

[0013] DETAILED DESCRIPTION OF THE INVENTION

[0014] In accordance with the present invention, sulfur-protected mercaptoacetylglycylglycyl- amino acid compounds are disclosed that can be used in the field of imaging radiopharmaceuticals wherein the sulfur atom is protected from oxidation prior to synthesis reactions that form a radiopharmaceutical complex. Sulfur-protected MAG-3 compounds of the present invention are represented by Formula 1 :

Formula 1

wherein X is -S-M⁰⁺L_n-I, or -S R. Mⁿ⁺ is a transition metal cation. Examples of transition metal cations include, but are not limited to, iron (Fe²⁺), copper (Cu⁺), copper (Cu²⁺), tin (Sn²⁺), zinc (Zn²⁺), nickel (Ni²⁺), manganese (Mn²⁺), and cobalt (Co³⁺). R is an organic moiety such as acyl, alkoxyalkyl, alkoxycarbonyl, 2-pyrrolinyl, 2- pyrrolidinyl, 2-furanyl, 2-pyranyl, disulfide, and the like; and Y is -OH, or -O. L_n-I, a countehon, is a Lewis base, and n varies from 1 to 3. Examples of Lewis bases include, but are not limited to, anions such as halides, hydroxide, oxide, amides, and the like or neutral species such as alcohols, amines, phosphines, and the like.

[0015] In one embodiment, the sulfur protecting group in MAG-3 ligands represented by Formula 1 is a metal cation, Mⁿ⁺. In such cases, the metal ion may exist as a simple thiolate salt of the ligand as or as a chelated species wherein, depending on the valance of the metal ion, the thiol group along with at least one of the amides will coordinate with the metal. The metal cations of the present invention include, but are not limited to, iron (Fe²⁺), copper (Cu⁺), copper (Cu²⁺), tin (Sn²⁺), zinc (Zn²⁺), nickel (Ni²⁺), manganese (Mn²⁺), and cobalt (Co³⁺).

[0016] In another embodiment of the present invention, the sulfur protecting group in MAG3 ligands represented by Formula 1 is an organic moiety, R, wherein R is selected from acyl, alkoxyalkyl, alkoxycarbonyl, 2-pyrrolinyl, 2-pyrrolidinyl, 2-furanyl, 2-pyranyl, disulfide, and the like. Some sulfur-protecting groups of the present invention beneficially cause MAG3 ligands to have improved hydrophilicity. Increased hydrophilicity can result in reduced reaction time for synthesizing Tc-99m radiopharmaceuticals. Advantageously, increased hydrophilicity can also improve the efficiency of the reaction between the ligand and Tc-99m such that greater quantities of mercaptoacetylglyclglycl-amino acid Tc-99m complexes are formed.

[0017] The synthesis reaction in which the sulfur-protected MAG3 compound of Formula 1 is reacted with a radioisotope can be conducted at room temperature or with the application of heat. In one example of the embodiment, the reaction process is carried out at room temperature.

[0018] MAG-3 ligands having transition metal sulfur-protecting groups have beneficial reaction properties in the synthesis of radiopharmaceuticals compared to mercaptoacetylglyclglycl-amino acid ligands of the prior art. Radiopharmaceutical synthesis processes of the prior art include reacting radioisotopes to known mercaptoacetylglyclglycl-amino acid ligands in a reaction mixture in the presence of heat. Beneficially, the MAG- 3 ligands having cation sulfur-protecting groups can react with radioisotopes either at room temperature or in the presence of heat to synthesize radiopharmaceuticals. By conducting the synthesis reaction at room temperature, the number of process steps and the preparation time are reduced in synthesis of radiopharmaceuticals. Significantly, synthesizing radiopharmaceuticals at room temperature can also improve the safety of the radiopharmaceutical kits by minimizing the handing of the radioisotope reactants during the synthesis process, thereby reducing the exposure to radiation for those who prepare the radiopharmaceuticals from mercaptoacetylglyclglycl-amino acid ligands and radioisotopes.

[0019] In one embodiment, R is selected from the group consisting of alkoxycarbonyl, 2-furanyl, 2-pyranyl, -SO3H, and -SOsNa. In one example, R is an alkoxycarbonyl selected from the group consisting of methoxycarbonyl and t-butoxycarbonyl.

[0020] The lower-alkoxyacyl groups form excellent sulfur-protecting groups for the mercaptoacetylglyclglycl-amino acid compounds resulting in a hydrophilic compound that is able to be efficiently labeled with a radioisotope in a nearly quantitative manner. In one example, at least 95% of the mercaptoacetylglyclglycl-amino acid compounds having lower-alkoxyacyl sulfur-protecting groups are labeled with a radioisotope. In another example, at least 98% of the mercaptoacetylglyclglycl-amino acid compounds are labeled with a radioisotope.

[0021] The lower-alkoxyacyl sulfur-protecting groups of the present invention beneficially increase the hydrophilicity of the mercaptoacetylglyclglycl-amino acid compound. As the compound becomes more hydrophilic, it dissolves more rapidly in the aqueous solution in which the labeling reaction occurs, thereby improving the speed and efficiency of the reaction.

[0022] One preferred example, a methoxycarbonyl group is used as a sulfur-protecting group for MAG3 forming a compound of Formula 2:

[0023] In another example, the sulfur-protecting group is a sulfonate group having the general formula:

[0024] wherein M¹ is a hydrogen or an alkali metal.

[0025] In one embodiment, a sulfonate group is used as a sulfur-protecting group for mercaptoacetylglyclglycl-amino acid compounds, wherein Y is defined as provided above, and M¹ is sodium. In one example, a sulfonate group is used as a sulfur-protecting group for MAG3, wherein M¹ is sodium, forming a compound having the Formula 3:

Formula 3

[0026] The sulfonate group, like the methoxycarbonyl groups described above, increases the solubility of the ligand, thereby increasing the speed and efficiency of the radiolabeling reaction. In one example, when combined with Tc-99m pertechnetate and a reducing agent, at least 95% of MAG3 compounds having the sulfonate sulfur-protecting group are labeled with a radioisotope. In another example, at least 98% of the MAG3 compounds are labeled with a radioisotope.

[0027] In particular, the sulfur-protected MAG3 compounds of the present invention can be utilized in synthesizing radiopharmaceuticals for use as renal imaging agents in nuclear medicine. The MAG3 compounds can then be contacted with other agents, examples of which are described below, and a radioisotope to form a radiopharmaceutical complex. The sulfur-protected MAG3 compounds can be formed by chemical synthesis reactions described in further detail below.

[0028] The novel sulfur-protected MAG3 compounds of the present invention, when complexed with a radioisotope, form a radiopharmaceutical that can be administered to a patient by intravenous injection and used in diagnostic scintigraphic procedures. In one embodiment, the radiopharmaceutical is administered to a patient in a diagnostic scintigraphic urography procedure. Once the complex is injected into the patient, images of the patient's kidneys are recorded by means of gamma scintillation cameras.

[0029] Kite

[0030] For convenience, sulfur-protected mercaptoacetylglyclglycl-amino acid ligands of the present invention may be provided to the user in the form of a kit containing some or all of the necessary components. The kit can comprise one or more of the following components: (i) a sulfur-protected mercaptoacetylglyclglycl-amino acid ligand, (ii) a reducing agent, (iii) an intermediate exchange ligand, and (iv) instructions for their combination and use. [0031] Exemplary reducing agents include dithionite, Ce (III), Fe (II), Cu (I), Ti (III), Sb (III), and Sn (II). Of these, Sn (II) is particularly preferred. Although it has been found practical to react a mercaptoacetylglyclglycl-amino acid compound with Tc-99m pertechnetate in the presence of dithionite in the laboratory, both the Tc-99m and the dithionite must be freshly prepared. A more convenient synthesis process utilizes a stannous ion as a reducing agent to induce the reaction of a mercaptoacetylglyclglycl-amino acid with sodium pertechnetate to form a Tc-99m compound. The kit also comprises a suitable intermediate exchange ligand to facilitate the coordination of Tc-99m to the mercaptoacetylglyclglycl-amino acid ligand. Examples of intermediate exchange ligands include acetate, tartrate, malate, lactate, hydroxyisobutyrate, citrate, glucoheptonate, gluconate, pyrophosphate, N-methyl N.N'-bis (2-hydroxyethyl)ethylenediamine, or glycine. The components of the kit may be provided in unit dosage form wherein the reducing agent, intermediate exchange ligand, and mercaptoacetylglyclglycl-amino acid ligand to be attached to the Tc-99m are provided in one vial. In one example, the reducing agent, intermediate exchange ligand, and mercaptoacetylglyclglycl-amino acid ligand to be attached to the Tc-99m are provided in one vial that is capable of long-term storage.

[0032] The kit optionally contains other components frequently intended to improve the ease of synthesis of the radiopharmaceutical by the practicing end user, the ease of manufacturing the kit, the shelf-life of the kit, or the stability and shelf-life of the radiopharmaceutical. Such components of the present invention include lyophilization aids, e.g., mannitol, lactose, sorbitol, dextran, Ficoll (GE Healthcare, Piscataway, NJ), and polyvinylpyyrolidine (PVP); stabilization aids, e.g., ascorbic acid, gentisic acid, and inositol; and bacteriostats, e.g., benzyl alcohol, benzalkonium chloride, chlorbutanol, and methyl, propyl, or butyl paraben. The kit can also be stabilized by filling the container headspace in the kit with an inert gas, for example, nitrogen, argon, other inert gas, and combinations thereof.

[0033] The kit can be stored at the location where radiodiagnostic procedures are conducted. For reasons of enhancing the shelf life and storage properties of the kit, the kit comprising the sulfur-protected mercaptoacetylglyclglycl-amino acid ligand, reducing agent, and intermediate exchange ligand can be provided in a dry, lyophilized state. The lyophilized ingredients can also include excipients. Examples of excipients include any non-toxic, pharmaceutically inert substance used to make up the pharmaceutical composition such as lactose, sucrose, corn starch, starch, gelatin and the like.

[0034] In one embodiment, the lyophilized ingredients are placed in a container that is overiayed with an inert gas for stabilizing the mixture and extending shelf life of the kit. The user may then reconstitute the lyophilized kit by adding a carrier or other solution. Optionally, the reconstitution may also occur at a radiopharmacy and transported to a location where the radiodiagnostic procedures are conducted.

[0035] Tc-99m, a common diagnostic radioisotope, has a half-life of only about six (6) hours. Due do this short half-life, it is not practical to package a Tc-99m-mercaptoacetylglyclglycl-amino acid complex ready for clinical use. Instead, Tc-99m radioisotope is received shortly before the Tc-99m- mercaptoacetylglyclglycl-amino acid complex is to be administered to a patient. Typically, the Tc-99m radioisotope is received as a eluate from a molybdenum-technetium generator, for example from a Ultra- TechneKow generator (Mallinckrodt Inc., St. Louis, Missouri).

[0036] The kit containing the sulfur-protected mercaptoacetylglyclglycl-amino acid is reacted with the Tc-99m radioisotope to form a radiopharmaceutical that is administered to a patient. In one example, the radioisotope is Tc-99m which, when coordinated with a mercaptoacetylglyclglycl-amino acid ligand forms a complex that generally corresponds to Formula 4:

Formula 4

[0037] wherein Z is -H or

^R3 ; and R¹ is -H, -CH₃, or -CH₂CH₃; R² Js -H₁ -CH₂CO₂H, CH₂CONH₂, -CH₂CH₂CO₂H, -CH₂CH₂CONH₂, -CH₃, -CH₂CH₃, -CH₂CH₃, -CH₂C₆H₅, -OCH₂OH, Or -CH₂OH, and R³ is -H, -CO₂H, -CONH₂, -SO₃H, -SO₂NH₂, or CONHCH₂CO₂H.

[0038] In one embodiment, a kit containing the sulfur-protected mercaptoacetylglyclglyclglycine is reacted with Tc-99m, forming a complex that generally corresponds to Formula 5:

Formula 5

[0039] One example of the present invention includes a radiopharmaceutical kit containing ingredients, which, when combined with Tc-99m pertechnetate, forms a diagnostically-effective amount of a Tc- 99m-mercaptoacetylglyclglyclglycine complex. For diagnostic purposes, the amount of Tc-99m utilized in a single radiopharmaceutical dose contains about 1 mCi to about 300 mCi Tc-99m in the form of Tc-99m pertechnetate. In another example, the amount of Tc-99m utilized in a single radiopharmaceutical dose contains about 5 mCi to about 50 mCi Tc-99m in the form of Tc-99m pertechnetate. In still another example, the amount of Tc-99m utilized in a single radiopharmaceutical dose contains about 10 mCi to about 20 mCi Tc-99m in the form of Tc-99m pertechnetate. The kit is formulated to contain diagnostically effective amounts of ingredients such that the Tc-99m-mercaptoacetylglyclglyclglycine complex can be prepared at a laboratory or medical facility shortly preceding its administration to a patient for use as an imaging agent in nuclear medicine.

[0040] In another embodiment, a radiopharmaceutical kit consists of a stannous ion (and intermediate exchange ligand) and sulfur-protected mercaptoacetylglyclglyclglycine of the present invention to be complexed to the Tc-99m radioisotope being provided in one vial, capable of long-term storage. In one example the kit further comprises an excipient.

[0041] Definitions:

[0042] The term "aryl" as used herein alone or as part of another group denote optionally substituted homocyclic aromatic groups, preferably monocyclic or bicyclic groups containing from 6 to 12 carbons in the ring portion, such as phenyl, biphenyl, naphthyl, substituted phenyl, substituted biphenyl or substituted naphthyl. Phenyl and substituted phenyl are the more preferred aryl.

[0043] The term "alkenyl" is a linear or branched radical having at least one carbon-carbon double bond of two to about twenty carbon atoms or, preferably, two to about twelve carbon atoms. More preferred alkyl radicals are "lower alkenyl" radicals having two to about six carbon atoms. Examples of alkenyl radicals include ethenyl, propenyl, allyl, butenyl and 4-methylbutenyl. The terms "alkenyl" and "lower alkenyl" also are radicals having "cis" and "trans" orientations, or alternatively, "E" and "Z" orientations. The term "cycloalkyl" is a saturated carbocyclic radical having three to twelve carbon atoms. More preferred cycloalkyl radicals are "lower cycloalkyl" radicals having three to about eight carbon atoms. Examples of such radicals include cyclopropyl, cyclobutyl, cyclopentyl and cyclohexyl.

[0044] The terms "halide," "halogen," or "halo" as used herein alone or as part of another group refer to chlorine, bromine, fluorine, and iodine.

[0045] The terms "hydrocarbon" and "hydrocarbyl" as used herein describe organic compounds or radicals consisting exclusively of the elements carbon and hydrogen. These moieties include alkyl, alkenyl, alkynyl, and aryl moieties. These moieties also include alkyl, alkenyl, alkynyl, and aryl moieties substituted with other aliphatic or cyclic hydrocarbon groups, such as alkaryl, alkenaryl and alkynaryl. Unless otherwise indicated, these moieties comprise 1 to 18 carbon atoms. They may be straight or branched chain or cyclic and include methyl, ethyl, propyl, isopropyl, allyl, benzyl, hexyl and the like.

[0046] The "substituted hydrocarbyl" moieties described herein are hydrocarbyl moieties which are substituted with at least one atom other than carbon, including moieties in which a carbon chain atom is substituted with a hetero atom such as nitrogen, oxygen, silicon, phosphorous, boron, sulfur, or a halogen atom. These substituents include halogen, heterocyclo, alkoxy, alkenoxy, alkynoxy, aryloxy, hydroxy, keto, acyl, acyloxy, nitro, tertiaryamino, amido, nitro, cyano, ketals, acetals, esters and ethers.

[0047] The following examples are provided in order to more fully illustrate the present invention.

[0048] Example 1 - Synthesis of Benzoyl Mercaptoacetylglvcylglvcylglvcine

[0049] The synthesis of benzoyl mercaptoacetylglycylglycylglycine (benzoyl-MAG3) is accomplished as a multi-step process beginning with dissolving 2.5 grams of glycylglycylglycine in 75 mL of 1.0 N sodium hydroxide in a 500 mL flask at 0° C and under a nitrogen atmosphere.

[0050] A solution of 13.0 grams of chloroacetyl chloride in 100 mL of ether is added dropwise from one addition funnel, while 100 mL of 1.0 N sodium hydroxide is simultaneously added dropwise from a second addition funnel, at the same time continuously stirring the glycylglycylglycine solution. Following dropwise addition of the chloroacetyl chloride and sodium hydroxide, the reaction mixture is maintained at 0° C while being stirred for an additional 1.5 hours.

[0051] The reaction mixture is next acidified to a pH of about 2 by addition of concentrated hydrochloric acid. The reaction mixture is stirred for an additional 30 minutes, warmed to 40⁰C, and concentrated to one-third of its volume under reduced pressure.

[0052] The concentrated mixture is then cooled in an ice bath in order to precipitate out chloroacetylglycylglycylglycine. The precipitate is washed twice with water to obtain chloroacetylglycylglycylglycine.

[0053] One gram of the crude chloroacetylglycylglycylglycine precipitate is next dissolved in 300 mL of anhydrous methanol under a nitrogen atmosphere. 50 mL of a solution containing sodium thiobenzoate (prepared from 175 mL of sodium in methanol to which 1.1 g of thiobenzoic acid is added) is added to the flask, and the reaction mixture is refluxed for 1.5 hours.

[0054] The solvent is next removed under reduced pressure. The resultant solid is isolated by filtration and washed with chloroform. Crystallization from methanol results in the recovery of benzoyl-MAG3.

[0055] Example 2 - Synthesis of Cation-MAG3 Complex

[0056] Cation sulfur-protected ligands of the present invention may be synthesized by first deprotecting the benzoyl-MAG3 compounds of Example 1. This can be performed in one or two steps. For most of the cations described above, a two-step process is carried out to form a cation-MAG3 compound. The two steps are generally: 1) deprotect the benzoyl-MAG3 ligand of Example 1 to permit the formation of a cation- MAG3 compound; and 2) form the cation-MAG3 compound.

[0057] In one example, the benzoyl-MAG3 compound can be deprotected by mixing benzoyl- MAG3 with hydroxide and subjecting the mixture to heat. 440 μL of sodium hydroxide (4N, 2.2 mmoles) is added under argon to a 10 ml solution of benzoyl-MAG3 (370 mg, Immole) in water. The solution is heated at 60° C for 10 minutes. The solution is then acidified to pH 2 with hydrochloric acid (2N, 2.2 mmoles. The resulting solution is rotavapped to form a pale yellow oil.

[0058] A basic salt of a desired cation (e.g., a base and a metal having basic properties, for example, M(OH)_n) is added to the pale yellow oil in at least about equimolar proportions to the deprotected MAG3 to form the cation sulfur-protected MAG3 complex, wherein M is a transition metal having basic properties, such as iron (Fe²⁺), copper (Cu⁺), copper (Cu²⁺), tin (Sn²⁺), zinc (Zn²⁺), nickel (Ni²⁺), manganese (Mn²⁺), and cobalt (Co³⁺), and n is 1 to 3.

[0059] In one embodiment, the proportions of reactants utilized in the deprotection of benzoyl- MAG3 is from about 1 :1 to 2:1 moles cation-hydroxide salt:moles benzoyl-MAG3. The proportions of MAG3:cation to deprotected MAG3 can range from about 1 :1 to about 1 :1.5. The proportions of MAG3:base (e.g., OH) to deprotected MAG3 can range from about 1 :1 to about 1:4.

[0060] Example 3 - Preparation of Cation-MAG3 Labeling Kits

[0061] Under an inert atmosphere of about 0.25 mg to about 25 mg of the cation-MAG3 prepared in Example 2 is added to 25 mL 0.1 molar phosphate buffer at pH 10.5. The mixture is heated for 10 minutes in a water-bath at 100° C. After cooling, 500 mg of lactose monohydrate is added. 2.5 mg SnCl2-H2O is dissolved in 0.5 mL ETOH and the pH is adjusted to pH 12 with NaOH 1N. The solution is diluted to 25 mL with water and dispensed in 1 mL-aliquots in reaction vials. The labeling kits are lyophilized, stoppered under vacuum, and then stored at 4°-8° C.

[0062] Each vial of the lyophilized kit typically contains between about 0.1 mg to about 0.02 mg SnCb, 20 mg lactose, and about 0.05 mg to about 2 mg cation-MAG3 complex.

[0063] Example 4 - Tc-99m-MAG3 Synthesis from Lyophilized Cation-MAG3

[0064] To a labeling kit prepared as described above in Example 3, is added 1 ml to 10 ml Tc-

99m generator eluate, containing 10 mCi to 300 mCi Tc-99m in the form of pertechnetate. The contents are mixed and the vial is incubated for 5 minutes at room temperature.

[0065] Example 5 - Cation-MAG3 Complex Concentrations

[0066] The initial starting amount of cation-MAG3 complex that is mixed with generator eluate containing Tc-99m, described above in Examples 3 and 5, may vary depending on the ionic bond strength formed between the cation and MAG3. Typically, the weaker the complex the less cation-MAG3 ligand will be needed in synthesizing the Tc99m-MAG3 complex. Conversely, stronger complexes (e.g., Cu(ll)-MAG3) will require more cation-MAG3 ligand to synthesize the Tc99m-MAG3 complex.

[0067] Example 6 - MAG3-Thiocarbonate Synthesis [0068] Step i. [0069] A mixture of mercaptoacetic acid (9.2 g, 0.10 mol) and triethylamine (22.0 g, 0.22 mole) in acetonitrile/water (1:1 , 150 mL) is stirred and cooled to 0° C. Thereafter, methylchloroformate (9.3 g, 0.11 mol) is added dropwise at such a rate that the internal temperature of the reaction is maintained between 0° C and 10° C. After the addition is complete, the reaction mixture is stirred at about 0° C for 2 hours. The reaction mixture is treated with 1 M HCI (20 mL) and brine (130 mL), and extracted with ethyl acetate (3 x 100 mL). The organic layer is separated, washed with brine, dried over anhydrous sodium sulfate, filtered, and the filtrate evaporated in vacuo. The crude product is then distilled under high vacuum to give the desired product, S- methoxycarbonylmercaptoacetic acid.

[0070] Step 2.

[0071] A mixture of S-methoxycarbonylmercaptoacetic acid from Step 1(15.0 g, 0.100 mol) and N-hydroxy succinimide (11.7 g, 0.102 mol) in anhydrous acetonitrile (100 mL) is stirred and carefully treated with N,N'-dicyclohexylcarbodiimide (20.8 g, 0.101 mol). After the addition, the entire mixture is stirred at ambient temperature for 16 hours. Dicyclohexylurea is filtered off, and the filtrate is evaporated in vacuo. The crude solid is then purified by recrystallization from ethyl acetate hexanes to give the desired active ester, N-succinimidyl S- methoxycarbonylmercaptoacetate.

[0072] Step 3.

[0073] A solution of triglycine is prepared by dissolving 1.54 g of triglycine in 10 mL of water and 0.8 g of NaOH. A stirring solution of the active ester from Step 2 (4.98 g, 0.02 mol) in acetonitrile (10 mL) is treated with a solution of triglycine (1.54 g, 0.02 mol) that is added in one portion. The entire mixture is stirred at ambient temperature for 2 hours. The solution is then treated with water (20 mL) and 1M HCI (20 mL). The precipitate is collected by filtration, dried, and recrystallized from suitable solvent to give the S-methoxycarbonyl protected MAG3.

[0074] When introducing elements of the present invention or the preferred embodiment(s) thereof, the articles "a," "an," "the," and "said" are intended to mean that there are one or more of the elements. The terms "comprising," "including," and "having" are intended to be inclusive and mean that there may be additional elements other than the listed elements.

[0075] In view of the above, it will be seen that the several objects of the invention are achieved and other advantageous results attained.

[0076] As various changes could be made in the above methods and products without departing from the scope of the invention, it is intended that all matter contained in the above description and shown in any accompanying drawings shall be interpreted as illustrative and not in a limiting sense.

Claims

What is claimed is:

1. A mercaptoacetylglycylglycylglycine compound comprising a mercaptoacetylglycylglycylglycine ligand compound having the formula:

wherein

X is selected from the group consisting of -S Mⁿ⁺L_n-i, and -SR;

Mⁿ⁺ is a transition metal cation;

R is an organic moiety selected from the group consisting of acyl, alkoxyalkyl, alkoxycarbonyl, 2-pyrrolinyl, 2- pyrrolidinyl, 2-furanyl, 2-pyranyl, and disulfide;

Y is -OH. or -0-;

Ln-1 is a Lewis base; and n varies from 1 to 3.

2. The compound of claim 1 , wherein the transition metal cation is selected from the group consisting of Fe²⁺, Cu⁺, Cu²⁺, Sn²⁺, Zn²*, Ni²⁺, Mn²⁺, and Co³⁺.

3. The compound of claim 1 , wherein the Lewis base is selected from the group consisting of halides, hydroxide, oxide, amides, alcohols, amines, and phosphines.

4. The compound of claim 3, wherein the halide is selected from the group consisting of chloride (Cl ), bromide (Br), and iodide (I ) ions.

5. The compound of claim 1 wherein X is -SR and R is selected from the group consisting of alkoxycarbonyl, 2-furanyl, 2-pyranyl, -SOsH₁ and -SOaNa.

6. The compound of claim 5, wherein R is an alkoxycarbonyl selected from the group consisting of methoxycarbonyl and t-butoxycarbonyl.

7. The compound of claim 5, wherein R is a sulfonate group having the formula:

and M¹ is a hydrogen or an alkali metal.

8. The compound of claim 7, wherein M¹ is sodium.

9. A kit for use in preparing a radiopharmaceutical compound comprising technetium-99m, said compound having the general formula:

and water-soluble salts thereof, said kit containing ingredients comprising: an amount of the composition of claim 1 ; and an amount of a water-soluble reducing agent complexed with an intermediate exchange ligand; wherein the ingredients, when mixed with a water soluble Tc-99m pertechnetate salt, react to form a diagnostically effective amount of the radiopharmaceutical compound.

10. The kit of claim 9, wherein the transition metal cation is selected from the group consisting of Fe²⁺, Cu⁺, Cu^2*, Sn²⁺, Zn²⁺, Ni²⁺, Mn²⁺, and Co³⁺.

11. The kit of claim 9, wherein the Lewis base is selected from the group consisting of halides, hydroxide, oxide, amides, alcohols, amines, and phosphines.

12. The kit of claim 11, wherein the halide is selected from the group consisting of chloride (Cl ), bromide (Br), and iodide (I ) ions.

13. The kit of claim 9 wherein X is -SR and R is selected from the group consisting of alkoxycarbonyl, 2-furanyl, 2-pyranyl, -SO3H, and -SO₃Na.

14. The kit of claim 13, wherein R is an alkoxycarbonyl selected from the group consisting of methoxycarbonyl and t-butoxycarbonyl.

15. The kit of claim 13, wherein R is a sulfonate group having the formula:

and M¹ is a hydrogen or an alkali metal.

16. A method of preparing a radiopharmaceutical compound comprising technetium-99m, and water-soluble salts thereof, the compound having the general formula:

The method comprising: reacting the sulfur-protected ligand of claim 1 with a reducing agent and a water-soluble Tc-99m pertechnetate salt.

17. The method of claim 16, wherein the transition metal cation is selected from the group consisting of Fe²⁺, Cu⁺, Cu²⁺, Sn²⁺, Zn²⁺, Ni²⁺, Mn²⁺, and Co³⁺.

18. The method of claim 16, wherein the Lewis base is selected from the group consisting of halides, hydroxide, oxide, amides, alcohols, amines, and phosphines.

19. The method of claim 18, wherein the halide is selected from the group consisting of chloride (Cl ), bromide (Br)₁ and iodide (I ) ions.

20. The method of claim 16, wherein the reaction is performed at room temperature.

21. The method of claim 16 wherein X is -SR and R is selected from the group consisting of alkoxycarbonyl, 2-furanyl, 2-pyranyl, -SO3H, and -SOaNa.

22. The method of claim 21 , wherein R is an alkoxycarbonyl selected from the group consisting of methoxycarbonyl and t-butoxycarbonyl.

23. The method of claim 16, wherein R is a sulfonate group having the formula:

and M¹ is a hydrogen or an alkali metal.

24. The method of claim 23, wherein M¹ is sodium.