Patent Publication Number: US-2005136003-A1

Title: Novel chemical agents comprising a cardiotonic moiety and an imaging moiety and methods of their use

Description:
CROSS REFERENCE TO RELATED APPLICATION  
      This non-provisional application claims priority from provisional application U.S. Ser. No. 60/516,566, filed Oct. 31, 2003. 
    
    
     FIELD OF THE INVENTION  
      The present invention relates to novel chemical agents comprising a cardiotonic moiety and an imaging moiety; and their use for diagnosing certain disorders in a subject. The present invention further relates to a kit for myocardial perfusion imaging.  
     BACKGROUND OF THE INVENTION  
      Cardiotonics are agents that have a strengthening effect on the heart or that can increase cardiac output. They may be cardiac glycosides, sympathomimetics, or other drugs. They are used after myocardial infarct, cardiac surgical procedures, in shock, or in congestive heart failure (heart failure, congestive). They are known to be highly specific for myocardial tissue. However, to the best of the applicants knowledge, the use of cardiotonics have previously been limited to therapeutic use.  
     SUMMARY OF THE INVENTION  
      The present invention relates to novel chemical agents comprising an cardiotonic moiety and an imaging moiety. In one embodiment, the novel chemical agent is a chemical compound comprising a cardiotonic moiety covalently linked to an imaging moiety, either directly or indirectly via a linker moiety. In another embodiment, the novel chemical agent is a chemical complex comprising a cardiotonic moiety linked to an imaging moiety via a non-covalent force. Non-limiting examples of the non-covalent force include ionic, hydrogen bonding, and van der Waals force.  
      The present invention is also directed to a method of imaging myocardial perfusion. Such method comprises administering to a subject an imaging agent which comprises a cardiotonic moiety and an imaging moiety; and scanning the subject using diagnostic imaging techniques. In one embodiment, the imaging agent is a chemical compound comprising a cardiotonic moiety covalently linked to an imaging moiety, either directly or indirectly via a linker moiety. In another embodiment, the novel chemical agent is a chemical complex comprising a cardiotonic moiety linked to an imaging moiety via a non-covalent force. Non-limiting examples of the non-covalent force include ionic, hydrogen bonding, and van der Waals force. Such agent are useful as imaging agents.  
      The present invention is further directed to a kit for myocardial perfusion imaging. Such kit comprises (1) a compound A which comprises a cardiotonic moiety and (2) a compound B which comprises an imaging moiety. Compounds A and B can be reacted to each other and form an imaging agent which comprises a cardiotonic moiety and an imaging moiety. In one embodiment, the imaging agent is a chemical compound comprising a cardiotonic moiety covalently linked to an imaging moiety, either directly or indirectly via a linker moiety. In another embodiment, the novel chemical agent is a chemical complex comprising a cardiotonic moiety linked to an imaging moiety via a non-covalent force. Non-limiting examples of the non-covalent force include ionic, hydrogen bonding, and van der Waals force.  
      The cardiotonic moiety can be selected from the group consisting of cardenolides, bufolides, diazoxide, and 2H-pyridinones and analogs thereof. 
    
    
     DETAILED DESCRIPTION OF ILLUSTRATIVE EMBODIMENTS  
      The present invention utilizes cardiotonic&#39;s affinity for myocardium and combines one or more cardiotonic moieties with one or more imaging moieties to form a novel agent suitable for imaging myocardial perfusion.  
      The present invention relates to novel chemical agents comprising an cardiotonic moiety and an imaging moiety. In one embodiment, the novel chemical agent is a chemical compound comprising a cardiotonic moiety covalently linked to an imaging moiety, either directly or indirectly via a linker moiety. In another embodiment, the novel chemical agent is a chemical complex comprising a cardiotonic moiety linked to an imaging moiety via a non-covalent force. Non-limiting examples of the non-covalent force include ionic, hydrogen bonding, and van der Waals force.  
      The present invention is also directed to a method of imaging myocardial perfusion. Such method comprises administering to a subject an imaging agent which comprises a cardiotonic moiety and an imaging moiety; and scanning the subject using diagnostic imaging techniques. In one embodiment, the imaging agent is a chemical compound comprising a cardiotonic moiety covalently linked to an imaging moiety, either directly or indirectly via a linker moiety. In another embodiment, the novel chemical agent is a chemical complex comprising a cardiotonic moiety linked to an imaging moiety via a non-covalent force. Non-limiting examples of the non-covalent force include ionic, hydrogen bonding, and van der Waals force.  
      The present invention is further directed to a kit for myocardial perfusion imaging. Such kit comprises (1) a compound A which comprises a cardiotonic moiety and (2) a compound B which comprises an imaging moiety. Compounds A and B can be reacted to each other and form an imaging agent which comprises a cardiotonic moiety and an imaging moiety. In one embodiment, the imaging agent is a chemical compound comprising a cardiotonic moiety covalently linked to an imaging moiety, either directly or indirectly via a linker moiety. In another embodiment, the novel chemical agent is a chemical complex comprising a cardiotonic moiety linked to an imaging moiety via a non-covalent force. Non-limiting examples of the non-covalent force include ionic, hydrogen bonding, and van der Waals force.  
      The cardiotonic moiety can be selected from groups consisting of cardenolides, bufolides, diazoxide, and 2H-pyridinones and analogs thereof.  
      Cardenolides may include, for example, digitoxin, digoxin, convallitoxin, cymarin, neriifolin, and oleandrin.  
      Bufolides may include for example, bufalin and resibufogenin.  
      2H-pyridinones may include, for example, amrinone, milrinone, loprinone, and vesnarinone.  
      In a first embodiment, the imaging agent is described by Formula (I):  
                 
 
 wherein: 
      A is H, C 1 -C 6  alkyl, or C(═O)H;     B is H or OH;     C is  
                 
 
 wherein R 2  is an imaging moiety or H; 
    D is an imaging moiety, H or OC(═O)R′, wherein R′ is C1-C6 alkyl;     E is OH;     G is an imaging moiety, H, or a bond wherein G and E, together with the atoms to which they are attached, form an epoxide ring; and     J is an imaging moiety, OH or  
                 
        wherein j is 0, 1, or 2;     R 3  is an imaging moiety, H, or OH;     R 4  is an imaging moiety, OH, or OCH 3 ; and     R 5  is an imaging moiety or OH, 
 
 with the proviso that at least one of R 2 , D, G, J, R 3 , R 4 , and R 5  is an imaging moiety. 
   
       

      In a preferred embodiment of Formula (I), the imaging agent is:  
                 
 
      In a second embodiment, the imaging agent is described by Formula (II):  
                 
 
 wherein: 
      J′ is an imaging moiety or H;     K is an imaging moiety or H; and     L is an imaging moiety or H, 
 
 with the proviso that at least one of J′, K, and L is an imaging moiety. 
   

      In a preferred embodiment of Formula (II), the imaging agent is:  
                 
 
      In a third embodiment, the imaging agent is described by Formula (III):  
                 
 
 wherein: 
      M is CN or NH 2 ;     Q is H or an imaging moiety;     T is  
                 
        wherein R 6 , R 7 , R 8 , R 9 , R 10 , R 11 , and R 12 , are each independently selected from H and an imaging moiety; and    
        U is H, CH 3 , or CH 2  attached to an imaging moiety, 
 
 with the proviso that at least one of Q, R 6 , R 7 , R 8 , R 9 , R 10 , R 11 , R 12  and U is an imaging moiety. 
   

      In a preferred embodiment of Formula (III), the imaging agent is:  
                 
 
      In a fourth embodiment, the imaging agent is described by Formula (IV):  
                 
 
 wherein R 13 , R 14 , R 15 , R 16 , R 17 , R 18 , R 19 , and R 20  are each independently selected from H and an imaging moiety, 
 
 with the proviso that at least one of R 13 , R 14 , R 15 , R 16 , R 17 , R 18 , R 19 , and R 20  is an imaging moiety. 
 
      In a preferred embodiment of Formula (IV), the imaging agent is:  
                 
 
 Imaging Moieties 
 
      Imaging moieties include those that are well known to those skilled in the art, and include those moieties that may be useful in the generation of diagnostic images by diagnostic techniques well known to the ordinarily skilled artisan. An imaging moiety is sometimes also referred to as a contrast moiety.  
      In one embodiment, the imaging moiety may be a radioisotope for nuclear medicine imaging, a radioisotope for X-ray CT imaging, a paramagnetic species for use in MRI imaging, an echogenic entity for use in ultrasound imaging, a fluorescent entity for use in fluorescence imaging, or an a light-active entity for use in optical imaging.  
      Nuclear medicine imaging moiety of the present invention include  11 C,  13 N,  18 F,  123 I,  125 I,  99m Tc,  95 Tc,  111 In,  62 Cu,  64 Cu,  67 Ga, and  68 Ga.  11 C-Palmitate has been used to probe fatty acid oxidation and  11 C-acetate has been used to assess oxidative metabolism in the myocardium (Brown, M., Marshall, D. R., Sobel, B. E., Bergmann, S. R. Circulation, 1987, 76, 687-696).  13 N-Ammonia has been used widely to image myocardial perfusion (Krivokapich J; Smith G T; Huang S C; Hoffman E J; Ratib O; Phelps M E; Schelbert H R. Circulation, 1989, 80, 1328-37). Compounds based on  18 F have been used for imaging purpose for hypoxia and cancer (Pauwels, E. K. J., A. A. van der Klaau w., Corporaal, T., Stokkel, M. P. M. Drugs of the Future, 2002, 27, 655-667). 15-(p-( 123 I)-iodophenyl)-pentadecanoic acid and 15-(p-( 123 I)-iodophenyl)-3(R, S)-methylpentadecanoic acid are two iodinated agents that have been used for imaging myocardial metabolism. In one embodiment, the imaging moiety employed in the present imaging agents is  18 F. Further imaging agents of the present invention may be comprised of one or more cardiotonic moieties attached to one or more X-ray absorbing or “heavy” atoms of atomic number 20 or greater, further comprising an optional linking moiety, L, between the one or more cardiotonic moieties and the X-ray absorbing atoms. A frequently used heavy atom in X-ray imaging agents is iodine. Recently, X-ray imaging agents comprised of metal chelates (Wallace, R., U.S. Pat. No. 5,417,959) and polychelates comprised of a plurality of metal ions (Love, D., U.S. Pat. No. 5,679,810) have been disclosed. More recently, multinuclear cluster complexes have been disclosed as X-ray imaging agents (U.S. Pat. No. 5,804,161, PCT WO91/14460, and PCT WO 92/17215). The disclosures of each of the foregoing documents are hereby incorporated herein by reference in their entireties. Preferred metals include Re, Sm, Ho, Lu, Pm, Y, Bi, Pd, Gd, La, Au, Au, Yb, Dy, Cu, Rh, Ag, and Ir.  
      MRI imaging agents of the present invention may be comprised of one or more cardiotonic moieties attached to one or more paramagnetic metal ions, further comprising an optional linking moiety, L, between the one or more cardiotonic moieties and the paramagnetic metal ions. The paramagnetic metal ions may be present in the form of metal chelates or complexes or metal oxide particles. U.S. Pat. Nos. 5,412,148, and 5,760,191, describe examples of chelators for paramagnetic metal ions for use in MRI imaging agents. U.S. Pat. No. 5,801,228, U.S. Pat. No. 5,567,411, and U.S. Pat. No. 5,281,704, describe examples of polychelants useful for complexing more than one paramagnetic metal ion for use in MRI imaging agents. U.S. Pat. No. 5,520,904, describes particulate compositions comprised of paramagnetic metal ions for use as MRI imaging agents. The disclosures of each of the foregoing documents are hereby incorporated herein by reference in their entireties. Preferred metals include Gd 3+ , Fe 3+ , In 3+ , and Mn 2+ .  
      The ultrasound imaging agents of the present invention may comprise one or more cardiotonic moieties attached to or incorporated into a microbubble of a biocompatible gas, a liquid carrier, and a surfactant microsphere, further comprising an optional linking moiety, L, between the one or more cardiotonic moieties and the microbubble. In this context, the term “liquid carrier” means aqueous solution and the term “surfactant” means any amphiphilic material which may produce a reduction in interfacial tension in a solution. A list of suitable surfactants for forming surfactant microspheres is disclosed, for example, in EP0727225A2, the disclosure of which is hereby incorporated herein by reference in its entirety. The term “surfactant microsphere” includes microspheres, nanospheres, liposomes, vesicles and the like. The biocompatible gas can be any physiologically accepted gas, including, for example, air, or a fluorocarbon, such as a C 3 -C 5  perfluoroalkane, which provides the difference in echogenicity and thus the contrast in ultrasound imaging. The gas may be encapsulated, contained, or otherwise constrained in or by the microsphere to which is attached the analog moiety, optionally via a linking group. The attachment can be covalent, ionic or by van der Waals forces. Specific examples of suitable imaging moieties include, for example, lipid encapsulated perfluorocarbons with a plurality of tumor neovasculature receptor binding peptides, polypeptides or peptidomimetics. Examples of gas filled imaging moieties include those found in U.S. patent application Ser. No. 09/931,317, filed Aug. 16, 2001, and U.S. Pat. Nos. 5,088,499, 5,547,656, 5,228,446, 5,585,112, and 5,846,517, the disclosures of which are hereby incorporated herein by reference in their entireties. 
          For imaging agents of Formula (I), the imaging moiety is preferably  18 F.     For imaging agents of Formula (II), the imaging moiety is preferably  18 F.     For imaging agents of Formula (III), the imaging moiety is preferably  18 F.     For imaging agents of Formula (IV), the imaging moiety is preferably  18 F. 
 
 Methods of Making 
       

      The novel agents of the present invention can be made using techniques which would be readily apparent to one of ordinary skill in the art, once armed with the teachings in the present application. Preferred reaction solvents include, for example, DMF, NMP, DMSO, THF, EtOAc, DCM, and chloroform. The reaction solution may be kept neutral or basic by the addition of an amine such as triethylamine or DIEA. Reactions may be carried out at ambient temperatures and protected from oxygen and water with a nitrogen atmosphere.  
      Temporary protecting groups may be used to prevent other reactive functionality, such as amines, thiols, alcohols, phenols, and carboxylic acids, from participating in the reaction. Preferred amine protecting groups include, for example, t-butoxycarbonyl and trityl (removed under mild acidic conditions), Fmoc (removed by the use of secondary amines such as piperidine), and benzyloxycarbonyl (removed by strong acid or by catalytic hydrogenolysis). The trityl group may also used for the protection of thiols, phenols, and alcohols. Preferred carboxylic acid protecting groups include, for example, t-Butyl ester (removed by mild acid), benzyl ester (usually removed by catalytic hydrogenolysis), and alkyl esters such as methyl or ethyl (usually removed by mild base). All protecting groups may be removed at the conclusion of synthesis using the conditions described above for the individual protecting groups, and the final product may be purified by techniques which would be readily apparent to one of ordinary skill in the art, once armed with the present disclosure.  
      Use  
      The imaging agents of the present invention may be used in a method of imaging, including methods of imaging in a subject (e.g., a human patient or an animal) comprising administering the imaging agent to the subject by injection, infusion, or any other known method, and imaging the area of the subject wherein the event of interest is located.  
      The useful dosage to be administered and the particular mode of administration will vary depending upon such factors as age, weight, and particular region to be treated, as well as the particular contrast agent used, the diagnostic use contemplated, and the form of the formulation, for example, suspension, emulsion, microsphere, liposome, or the like, as will be readily apparent to those skilled in the art.  
      Typically, dosage is administered at lower levels and increased until the desirable diagnostic effect is achieved. In one embodiment, the above-described imaging agents may be administered by intravenous injection, usually in saline solution, at a dose of about 0.1 to about 100 mCi per 70 kg body weight (and all combinations and subcombinations of dosage ranges and specific dosages therein), or preferably at a dose of about 0.5 to about 50 mCi. Imaging is performed using techniques well known to the ordinarily skilled artisan.  
      For use as nuclear medicine imaging agents, the compositions of the present invention, dosages, administered by intravenous injection, will typically range from about 0.5 μmol/kg to about 1.5 mmol/kg (and all combinations and subcombinations of dosage ranges and specific dosages therein), preferably about 0.8 μmol/kg to about 1.2 mmol/kg.  
      For use as MRI imaging agents, the compositions of the present invention may be used in a similar manner as other MRI agents as described in U.S. Pat. No. 5,155,215; U.S. Pat. No. 5,087,440; Margerstadt et al., Magn. Reson. Med., 1986, 3, 808; Runge et al., Radiology, 1988, 166, 835; and Bousquet et al., Radiology, 1988, 166, 693. The disclosures of each of the foregoing documents are hereby incorporated herein by reference in their entireties. Generally, sterile aqueous solutions of the contrast agents may be administered to a patient intravenously in dosages ranging from about 0.01 to about 1.0 mmoles per kg body weight (and all combinations and subcombinations of dosage ranges and specific dosages therein).  
      The ultrasound imaging agents of the present invention may be administered by intravenous injection in an amount from about 10 to about 30 μL (and all combinations and subcombinations of dosage ranges and specific dosages therein) of the echogenic gas per kg body weight or by infusion at a rate of approximately 3 μL/kg/min.  
      Buffers can optionally be included. Buffers useful in the preparation of imaging agents and kits include, for example, phosphate, citrate, sulfosalicylate, and acetate buffers. A more complete list can be found in the United States Pharmacopoeia, the disclosure of which is hereby incorporated herein by reference in its entirety.  
      Lyophilization aids can optionally be included. Lyophilization aids useful in the preparation of imaging agents and kits include, for example, mannitol, lactose, sorbitol, dextran, FICOLL® polymer, and polyvinylpyrrolidine (PVP).  
      Stabilization aids can optionally be included. Stabilization aids useful in the preparation of imaging agents and kits include, for example, ascorbic acid, cysteine, monothioglycerol, sodium bisulfite, sodium metabisulfite, gentisic acid, and inositol.  
      Solubilization aids can optionally be included. Solubilization aids useful in the preparation of imaging agents and kits include, for example, ethanol, glycerin, polyethylene glycol, propylene glycol, polyoxyethylene sorbitan monooleate, sorbitan monoloeate, polysorbates, poly(oxyethylene)-poly(oxypropylene)-poly(oxyethylene) block copolymers (“Pluronics”) and lecithin. Preferred solubilizing aids are polyethylene glycol and Pluronics.  
      Bacteriostats can be optionally included. Bacteriostats useful in the preparation of imaging agents and kits include, for example, benzyl alcohol, benzalkonium chloride, chlorbutanol, and methyl, propyl, or butyl paraben.  
      A component in a diagnostic kit can also serve more than one function. For example, a reducing agent for a radionuclide can also serve as a stabilization aid, or a buffer can also serve as a transfer ligand, or a lyophilization aid can also serve as a transfer, ancillary, or co-ligand.  
      The compounds herein described may have asymmetric centers. Unless otherwise indicated, all chiral, diastereomeric and racemic forms are included in the present invention. Many geometric isomers of olefins, C═N double bonds, and the like can also be present in the compounds described herein, and all such stable isomers are contemplated in the present invention. It will be appreciated that compounds of the present invention contain asymmetrically substituted carbon atoms, and may be isolated in optically active or racemic forms. It is well known in the art how to prepare optically active forms, such as by resolution of racemic forms or by synthesis from optically active starting materials. Two distinct isomers (cis and trans) of the peptide bond are known to occur; both can also be present in the compounds described herein, and all such stable isomers are contemplated in the present invention. The D and L-isomers of a particular amino acid are designated herein using the conventional 3-letter abbreviation of the amino acid, as indicated by the following examples: D-Leu, or L-Leu.  
      When any variable occurs more than one time in any substituent or in any formula, its definition in each occurrence is independent of its definition at every other occurrence. Thus, for example, if a group, or plurality of groups, is shown to be substituted with 0-2 R, then said group(s) may optionally be substituted with up to two R, and R at each occurrence in each group is selected independently from the defined list of possible R. Also, by way of example, for the group —N(R′) 2 , each of the two R′ substituents on N is independently selected from the defined list of possible R′. Combinations of substituents and/or variables are permissible only if such combinations result in stable compounds. When a bond to a substituent is shown to cross the bond connecting two atoms in a ring, then such substituent may be bonded to any atom on the ring.  
      Definitions  
      As used herein, “alkyl” is intended to include both branched and straight-chain saturated aliphatic hydrocarbon groups having the specified number of carbon atoms, examples of which include, but are not limited to, methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, sec-butyl, t-butyl, pentyl, hexyl, heptyl, octyl, nonyl, and decyl; cycloalkyl including saturated and partially unsaturated ring groups, including mono-, bi- or poly-cyclic ring systems, such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl and adamantyl; bicycloalkyl including saturated bicyclic ring groups such as [3.3.0]bicyclooctane, [4.3.0]bicyclononane, [4.4.0]bicyclodecane (decalin), [2.2.2]bicyclooctane, and so forth.  
      The term “alkoxy” means an alkyl-CO— group wherein alkyl is as previously described. Exemplary groups include methoxy, ethoxy, and so forth.  
      As used herein, the term “alkene” or “alkenyl” is intended to include hydrocarbon chains having the specified number of carbon atoms of either a straight or branched configuration and one or more unsaturated carbon-carbon bonds which may occur in any stable point along the chain, such as ethenyl, propenyl, and the like.  
      As used herein, the term “alkyne” or “alkynyl” is intended to include hydrocarbon chains having the specified number of carbon atoms of either a straight or branched configuration and one or more unsaturated carbon-carbon triple bonds which may occur in any stable point along the chain, such as propargyl, and the like.  
      As used herein, the term alkyl ether is intended to include hydrocarbon chains having the specified number of carbon atoms of either a straight or branched configuration and a noncarbon atom, such as S or O, with two points of attachment (i.e., is a diradical) in the chain.  
      As used herein, “aryl” or “aromatic residue” is intended to mean phenyl or naphthyl, which when substituted, the substitution can be at any position.  
      The term “aryloxy” means an aryl-CO— group wherein aryl is as previously described. Exemplary groups include phenoxy and naphthoxy.  
      As used herein, the term “alkaryl” means an aryl group bearing an alkyl group of 1-10 carbon atoms; the term “aralkyl” means an alkyl group of 1-10 carbon atoms bearing an aryl group; the term “arylalkaryl” means an aryl group bearing an alkyl group of 1-10 carbon atoms bearing an aryl group; the term “heteroaralkyl” means an alkyl group of 1-10 carbon atoms bearing an aryl group and a heteroatom; and the term “heterocycloalkyl” means an alkyl group of 1-10 carbon atoms bearing a heterocycle.  
      “Ancillary” or “co-ligands” are ligands that may be incorporated into a radiopharmaceutical during its synthesis. They may serve to complete the coordination sphere of the radionuclide together with the chelator or radionuclide bonding unit of the reagent. For radiopharmaceuticals comprised of a binary ligand system, the radionuclide coordination sphere may be composed of one or more chelators or bonding units from one or more reagents and one or more ancillary or co-ligands, provided that there are a total of two types of ligands, chelators or bonding units. For example, a radiopharmaceutical comprised of one chelator or bonding unit from one reagent and two of the same ancillary or co-ligands and a radiopharmaceutical comprised of two chelators or bonding units from one or two reagents and one ancillary or co-ligand are both considered to be comprised of binary ligand systems. For radiopharmaceuticals comprised of a ternary ligand system, the radionuclide coordination sphere may be composed of one or more chelators or bonding units from one or more reagents and one or more of two different types of ancillary or co-ligands, provided that there are a total of three types of ligands, chelators or bonding units. For example, a radiopharmaceutical comprised of one chelator or bonding unit from one reagent and two different ancillary or co-ligands is considered to be comprised of a ternary ligand system. Ancillary or co-ligands useful in the preparation of radiopharmaceuticals and in diagnostic kits useful for the preparation of said radiopharmaceuticals may be comprised of one or more oxygen, nitrogen, carbon, sulfur, phosphorus, arsenic, selenium, and tellurium donor atoms. A ligand can be a transfer ligand in the synthesis of a radiopharmaceutical and also serve as an ancillary or co-ligand in another radiopharmaceutical. Whether a ligand is termed a transfer or ancillary or co-ligand depends on whether the ligand remains in the radionuclide coordination sphere in the radiopharmaceutical, which is determined by the coordination chemistry of the radionuclide and the chelator or bonding unit of the reagent or reagents.  
      A “bacteriostat” is a component that inhibits the growth of bacteria in a formulation either during its storage before use of after a diagnostic kit is used to synthesize a radiopharmaceutical.  
      The term “bond”, as used herein, means either a single or double bond.  
      The term “bubbles” or “microbubbles,” as used herein, refers to vesicles which are generally characterized by the presence of one or more membranes or walls surrounding an internal void that is filled with a gas or precursor thereto. Exemplary bubbles or microbubbles include, for example, liposomes, micelles and the like.  
      A “carbohydrate” is a polyhydroxy aldehyde, ketone, alcohol or acid, or derivatives thereof, including polymers thereof having polymeric linkages of the acetal type.  
      A “chelator” or “bonding unit” is the moiety or group on a reagent that binds to a metal ion through the formation of chemical bonds with one or more donor atoms. Preferred chelators of the present invention are described in U.S. Pat. No. 6,511,648, the disclosure of which is hereby incorporated herein by reference in its entirety.  
      A “cyclodextrin” is a cyclic oligosaccharide. Examples of cyclodextrins include, but are not limited to, α-cyclodextrin, hydroxyethyl-α-cyclodextrin, hydroxypropyl-α-cyclodextrin, β-cyclodextrin, hydroxypropyl-β-cyclodextrin, carboxymethyl-β-cyclodextrin, dihydroxypropyl-β-cyclodextrin, hydroxyethyl-β-cyclodextrin, 2,6 di-O-methyl-β-cyclodextrin, sulfated-β-cyclodextrin, γ-cyclodextrin, hydroxypropyl-γ-cyclodextrin, dihydroxypropyl-γ-cyclodextrin, hydroxyethyl-γ-cyclodextrin, and sulfated γ-cyclodextrin.  
      A “diagnostic kit” or “kit” comprises a collection of components, termed the formulation, in one or more vials which are used by the practicing end user in a clinical or pharmacy setting to synthesize diagnostic radiopharmaceuticals. The kit preferably provides all the requisite components to synthesize and use the diagnostic pharmaceutical except those that are commonly available to the practicing end user, such as water or saline for injection, a solution of the radionuclide, equipment for heating the kit during the synthesis of the radiopharmaceutical, if required, equipment necessary for administering the radiopharmaceutical to the patient such as syringes, shielding, imaging equipment, and the like. Contrast agents are provided to the end user in their final form in a formulation contained typically in one vial, as either a lyophilized solid or an aqueous solution. The end user typically reconstitutes the lyophilized material with water or saline and withdraws the patient dose or just withdraws the dose from the aqueous solution formulation as provided.  
      The term “donor atom” refers to the atom directly attached to a metal by a chemical bond.  
      The suffix “ene” when used with the hydrocarbons defined above, indicates that the group has two points of attachment (i.e., is a diradical). For example, a saturated aliphatic hydrocarbon group disposed between two other moieties would be referred to herein as “alkylene.” 
      As used herein, the term “heterocycle” or “heterocyclic system” is intended to mean a stable 5- to 7-membered monocyclic or bicyclic or 7- to 10-membered bicyclic heterocyclic ring which is saturated, partially unsaturated, or unsaturated (aromatic), and which consists of carbon atoms and from 1 to 4 heteroatoms independently selected from the group consisting of N, O and S and including any bicyclic group in which any of the above-defined heterocyclic rings is fused to a benzene ring. The nitrogen and sulfur heteroatoms may optionally be oxidized. The heterocyclic ring may be attached to its pendant group at any heteroatom or carbon atom which results in a stable structure. The heterocyclic rings described herein may be substituted on carbon or on a nitrogen atom if the resulting compound is stable. If specifically noted, a nitrogen in the heterocycle may optionally be quaternized. It is preferred that when the total number of S and O atoms in the heterocycle exceeds 1, then these heteroatoms are not adjacent to one another. It is preferred that the total number of S and O atoms in the heterocycle is not more than 1. As used herein, the term “aromatic heterocyclic system” or “heteroaryl” is intended to mean a stable 5- to 7-membered monocyclic or bicyclic or 7- to 10-membered bicyclic heterocyclic aromatic ring which consists of carbon atoms and from 1 to 4 heteroatoms independently selected from the group consisting of N, O and S. It is preferred that the total number of S and O atoms in the aromatic heterocycle is not more than 1. Examples of heterocycles include, but are not limited to, 1H-indazole, 2-pyrrolidonyl, 2H,6H-1,5,2-dithiazinyl, 2H-pyrrolyl, 3H-indolyl, 4-piperidonyl, 4aH-carbazole, 4H-quinolizinyl, 6H-1,2,5-thiadiazinyl, acridinyl, azocinyl, benzimidazolyl, benzofuranyl, benzothiofuranyl, benzothiophenyl, benzoxazolyl, benzthiazolyl, benztriazolyl, benztetrazolyl, benzisoxazolyl, benzisothiazolyl, benzimidazalonyl, carbazolyl, 4aH-carbazolyl, carbolinyl, chromanyl, chromenyl, cinnolinyl, decahydroquinolinyl, 2H,6H-1,5,2-dithiazinyl, dihydrofuro[2,3-b]tetrahydrofuran, furanyl, furazanyl, imidazolidinyl, imidazolinyl, imidazolyl, 1H-indazolyl, indolenyl, indolinyl, indolizinyl, indolyl, isobenzofuranyl, isochromanyl, isoindazolyl, isoindolinyl, isoindolyl, isoquinolinyl, isothiazolyl, isoxazolyl, morpholinyl, naphthyridinyl, octahydroisoquinolinyl, oxadiazolyl, 1,2,3-oxadiazolyl, 1,2,4-oxadiazolyl, 1,2,5-oxadiazolyl, 1,3,4-oxadiazolyl, oxazolidinyl., oxazolyl, oxazolidinylperimidinyl, phenanthridinyl, phenanthrolinyl, phenarsazinyl, phenazinyl, phenothiazinyl, phenoxathiinyl, phenoxazinyl, phthalazinyl, piperazinyl, piperidinyl, pteridinyl, piperidonyl, 4-piperidonyl, pteridinyl, purinyl, pyranyl, pyrazinyl, pyrazolidinyl, pyrazolinyl, pyrazolyl, pyridazinyl, pyridooxazole, pyridoimidazole, pyridothiazole, pyridinyl, pyridyl, pyrimidinyl, pyrrolidinyl, pyrrolinyl, pyrrolyl, quinazolinyl, quinolinyl, 4H-quinolizinyl, quinoxalinyl, quinuclidinyl, carbolinyl, tetrahydrofuranyl, tetrahydroisoquinolinyl, tetrahydroquinolinyl, 6H-1,2,5-thiadiazinyl, 1,2,3-thiadiazolyl, 1,2,4-thiadiazolyl, 1,2,5-thiadiazolyl, 1,3,4-thiadiazolyl, thianthrenyl, thiazolyl, thienyl, thienothiazolyl, thienooxazolyl, thienoimidazolyl, thiophenyl, triazinyl, 1,2,3-triazolyl, 1,2,4-triazolyl, 1,2,5-triazolyl, 1,3,4-triazolyl, xanthenyl. Preferred heterocycles include, but are not limited to, pyridinyl, furanyl, thienyl, pyrrolyl, pyrazolyl, imidazolyl, indolyl, benzimidazolyl, 1H-indazolyl, oxazolidinyl, benzotriazolyl, benzisoxazolyl, oxindolyl, benzoxazolinyl, or isatinoyl. Also included are fused ring and spiro compounds containing, for example, the above heterocycles.  
      As used herein, the term “lipid” refers to a synthetic or naturally-occurring amphipathic compound which comprises a hydrophilic component and a hydrophobic component. Lipids include, for example, fatty acids, neutral fats, phosphatides, glycolipids, aliphatic alcohols and waxes, terpenes and steroids. Exemplary compositions which comprise a lipid compound include suspensions, emulsions and vesicular compositions.  
      “Liposome” refers to a generally spherical cluster or aggregate of amphipathic compounds, including lipid compounds, typically in the form of one or more concentric layers, for example, bilayers. They may also be referred to herein as lipid vesicles.  
      A “lyophilization aid” is a component that has favorable physical properties for lyophilization, such as the glass transition temperature, and is generally added to the formulation to improve the physical properties of the combination of all the components of the formulation for lyophilization.  
      “Metallopharmaceutical” means a pharmaceutical comprising a metal. The metal is the cause of the imageable signal in diagnostic applications. Radiopharmaceuticals are metallopharmaceuticals in which the metal is a radioisotope.  
      The phrase “pharmaceutically acceptable” is employed herein to refer to those compounds, materials, compositions, and/or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit/risk ratio.  
      As used herein, “pharmaceutically acceptable salts” refer to derivatives of the disclosed compounds modified by making acid or base salts. Examples of pharmaceutically acceptable salts include, but are not limited to, mineral or organic acid salts of basic residues such as amines; alkali or organic salts of acidic residues such as carboxylic acids; and the like. The pharmaceutically acceptable salts include the conventional non-toxic salts or the quaternary ammonium salts of the parent compound formed, for example, from non-toxic inorganic or organic acids. For example, such conventional non-toxic salts include those derived from inorganic acids such as hydrochloric, hydrobromic, sulfuric, sulfamic, phosphoric, nitric and the like; and the salts prepared from organic acids such as acetic, propionic, succinic, glycolic, stearic, lactic, tartaric, citric, ascorbic, pamoic, maleic, hydroxymaleic, phenylacetic, glutamic, benzoic, salicylic, sulfanilic, 2-acetoxybenzoic, fumaric, toluenesulfonic, methanesulfonic, ethane disulfonic, oxalic, isethionic, and the like. The pharmaceutically acceptable salts of the present invention can be synthesized from the parent compound which contains a basic or acidic moiety by conventional chemical methods. Generally, such salts can be prepared by reacting the free acid or base forms of these compounds with a stoichiometric amount of the appropriate base or acid in water or in an organic solvent, or in a mixture of the two; generally, nonaqueous media like ether, ethyl acetate, ethanol, isopropanol, or acetonitrile are preferred. Lists of suitable salts are found in  Remington&#39;s Pharmaceutical Sciences,  17th ed., Mack Publishing Company, Easton, Pa., 1985, p. 1418, the disclosure of which is hereby incorporated by reference.  
      A “polyalkylene glycol” is a polyethylene glycol, polypropylene glycol, polybutylene glycol, or similar glycol having a molecular weight of less than about 5000, terminating in either a hydroxy or alkyl ether moiety.  
      As used herein, the term “polycarboxyalkyl” means an alkyl group having from about two and about 100 carbon atoms and a plurality of carboxyl substituents; and the term “polyazaalkyl” means a linear or branched alkyl group having from about two and about 100 carbon atoms, interrupted by or substituted with a plurality of amine groups.  
      By “reagent” is meant a compound of this invention capable of direct transformation into a metallopharmaceutical of this invention. Reagents may be utilized directly for the preparation of the metallopharmaceuticals of this invention or may be a component in a kit of this invention.  
      A “reducing agent” is a compound that reacts with a radionuclide, which is typically obtained as a relatively unreactive, high oxidation state compound, to lower its oxidation state by transferring electron(s) to the radionuclide, thereby making it more reactive. Reducing agents useful in the preparation of radiopharmaceuticals and in diagnostic kits useful for the preparation of said radiopharmaceuticals include, for example, stannous chloride, stannous fluoride, formamidine sulfinic acid, ascorbic acid, cysteine, phosphines, and cuprous or ferrous salts. Other reducing agents are described, for example, in Brodack et. al., PCT Application 94/22496, the disclosure of which is incorporated herein by reference in its entirety.  
      The term “salt”, as used herein, is used as defined in the CRC Handbook of Chemistry and Physics, 65th Edition, CRC Press, Boca Raton, Fla., 1984, as any substance which yields ions, other than hydrogen or hydroxyl ions.  
      A “stabilization aid” is a component that is typically added to the metallopharmaceutical or to the diagnostic kit either to stabilize the metallopharmaceutical or to prolong the shelf-life of the kit before it must be used. Stabilization aids can be antioxidants, reducing agents or radical scavengers and can provide improved stability by reacting preferentially with species that degrade other components or the metallopharmaceutical.  
      By “stable compound” or “stable structure” is meant herein a compound that is sufficiently robust to survive isolation to a useful degree of purity from a reaction mixture, and formulation into an efficacious pharmaceutical agent.  
      A “solubilization aid” is a component that improves the solubility of one or more other components in the medium required for the formulation.  
      The term “substituted”, as used herein, means that one or more hydrogens on the designated atom or group is replaced with a selection from the indicated group, provided that the designated atom&#39;s or group&#39;s normal valency is not exceeded, and that the substitution results in a stable compound. When a substituent is keto (i.e., ═O), then 2 hydrogens on the atom are replaced.  
      A “transfer ligand” is a ligand that forms an intermediate complex with a metal ion that is stable enough to prevent unwanted side-reactions but labile enough to be converted to a metallopharmaceutical. The formation of the intermediate complex is kinetically favored while the formation of the metallopharmaceutical is thermodynamically favored. Transfer ligands useful in the preparation of metallopharmaceuticals and in diagnostic kits useful for the preparation of diagnostic radiopharmaceuticals include, for example, gluconate, glucoheptonate, mannitol, glucarate, N,N,N′,N′-ethylenediaminetetraacetic acid, pyrophosphate and methylenediphosphonate. In general, transfer ligands are comprised of oxygen or nitrogen donor atoms.  
      As used herein, the term “vesicle” refers to a spherical entity which is characterized by the presence of an internal void. Preferred vesicles are formulated from lipids, including the various lipids described herein. In any given vesicle, the lipids may be in the form of a monolayer or bilayer, and the mono- or bilayer lipids may be used to form one of more mono- or bilayers. In the case of more than one mono- or bilayer, the mono- or bilayers are generally concentric. The lipid vesicles described herein include such entities commonly referred to as liposomes, micelles, bubbles, microbubbles, microspheres and the like. Thus, the lipids may be used to form a unilamellar vesicle (comprised of one monolayer or bilayer), an oligolamellar vesicle (comprised of about two or about three monolayers or bilayers) or a multilamellar vesicle (comprised of more than about three monolayers or bilayers). The internal void of the vesicles may be filled with a liquid, including, for example, an aqueous liquid, a gas, a gaseous precursor, and/or a solid or solute material, including, for example, a bioactive agent, as desired.  
      As used herein, the term “vesicular composition” refers to a composition which is formulate from lipids and which comprises vesicles.  
      As used herein, the term “vesicle formulation” refers to a composition which comprises vesicles and a bioactive agent.  
      The following abbreviations are used herein: 
      Acm acetamidomethyl     b-Ala, beta-Ala or bAla 3-aminopropionic acid     ATA 2-aminothiazole-5-acetic acid or 2-aminothiazole-5-acetyl group     Boc t-butyloxycarbonyl     CBZ, Cbz or Z Carbobenzyloxy     Cit citrulline     Dap 2,3-diaminopropionic acid     DCC dicyclohexylcarbodiimide     DIEA diisopropylethylamine     DMAP 4-dimethylaminopyridine     DMSO dimethyl sulfoxide     DMF N,N-dimethylformamide     EtOAc Ethyl Acetate     EOE ethoxyethyl     Fmoc 9-fluorenylmethoxycarbonyl     HBTU 2-(1H-Benzotriazol-1-yl)-1,1,3,3-tetramethyluronium hexafluorophosphate     hynic boc-hydrazinonicotinyl group or 2-[[[5-[carbonyl]-2-pyridinyl]hydrazono]methyl]-benzenesulfonic acid,     NMeArg or MeArg a-N-methyl arginine     NMeAsp a-N-methyl aspartic acid     NMM N-methylmorpholine     NMP N-Methyl-2-pyrrolidone     OcHex O-cyclohexyl     OBzl O-benzyl     oSu O-succinimidyl     TBTU 2-(1H-Benzotriazol-1-yl)-1,1,3,3-tetramethyluronium tetrafluoroborate     THF tetrahydrofuranyl     THP tetrahydropyranyl     Tos tosyl     Tr or Trt trityl    

      The present invention is further described in the following non-limiting examples.  
     EXAMPLES  
     Example 1  
     Digoxin Triflate  
      A suspension of digoxin (78 mg) in dimethylformamide (DMF, 1 mL) is cooled to 0 degrees centigrade and a slurry of sodium hydride (5.0 mg) in THF is added. The resultant mixture is stirred at 0 degrees for 10 minutes, and trifluoromethanesulfonic anhydride (54 mg) is added as a DMF (200 uL) solution. The mixture is stirred at room temperature for one hour, the poured into dilute aqueous HCl (5%, 50 mL). The mixture is extracted exhaustively with chloroform, and the combined organics are dried (sat&#39;d NaCl, MgSO 4 ) and concentrated to afford the crude triflate. Chromatography via HPLC provides the desired pure triflate.  
     Example 2  
     Deoxy- 18 F-digoxin  
      A solution of the above prepared triflate (93 mg) from Example 1 and kryptofix 2.2.2 in water-ethanol (2:1) is treated with potassium  18 F-fluoride (30 uCi), and the resultant mixture is heated for 25 minutes. The reaction mixture is passed through successive columns of alumina and silica gel (RP-C18). Concentration of the eluant affords the desired compound in good chemical and radiochemical purity.  
     Example 3  
     7-chloro-3-hydroxymethyl-2H-1,2,4-benzothiadiazine 1,1-dioxide  
      To a cooled (0 degrees C.) solution of 2-amino-5-chlorobenzenesulfonamide and triethylamine in dimethyl acetamide is added one equivalent of 2-trimethylsilyloxyacetyl chloride. The resultant mixture is stirred at 0 degrees for one hour and room temperature for one hour. The mixture is poured into a mixture of water and methylene chloride and the layers separated. The aqueous phase is separated and extracted with additional methylene chloride. The combined organics are washed repeatedly with water, dried (sat&#39;d aq NaCl, MgSO 4 ) and concentrated to afford an oil, which is used directly in the following step.  
      The oil from the above procedure is dissolved in ethanol and one equivalent of potassium hydroxide is added. The mixture is heated at reflux for one hour. The mixture is cooled, poured into water, and extracted with methylene chloride. The organic layer is separated, and washed (sat&#39;d aq NaHCO 3 ), dried (sat&#39;d aq NaCl) and concentrated to afford the desired product, 7-chloro-3-hydroxymethyl-2H-1,2,4-benzothiadiazine 1,1-dioxide.  
     Example 4  
     7-chloro-3-trifluoromethanesulfonatomethyl-2H-1,2,4-benzothiadiazine 1,1-dioxide  
      To a cooled (0 degrees C.) solution of 7-chloro-3-hydroxymethyl-2H-1,2,4-benzothiadiazine 1,1-dioxide (from Example 3) in THF is added a suspension of sodium hydride in THF. The resultant solution is stirred for 30 minutes at 0 degrees, then trifluoromethanesulfonic anhydride is added. The mixture is allowed to warm to ambient temperature and stirred overnight. The solution is then poured in water, and extracted twice with methylene chloride. The organic layers are combined and dried (sat&#39;d aq NaCl, MgSO 4 ), and concentrated to afford an oil. Chromatography affords pure material.  
     Example 5  
     7-chloro-3- 18 F-fluoromethyl-2H-1,2,4-benzothiadiazine 1,1-dioxide  
      The above prepared triflate from Example 4 is dissolved in acetonitrile (1 mL), and added to a dry solution of K 18 F(50 mCi, via repeated azeotropic distillation of acetonitrile), kryptofix (2 mg) and potassium carbonate (50 ug). The vial is sealed and heated at 110 degrees C. for 15 minutes. The mixture is then cooled via active airflow, diluted with dichloromethane (4 mL) and passed through a Sep-Pak (silica gel, 230-400 mesh). The Sep Pak is further eluted with dichloromethane (4 mL) and the combined organics are concentrated. The resultant residue is dissolved in acetonitrile and purified via reverse-phase HPLC (C-18, acetonitrile:water 20% to 100% gradient). Concentration of the fractions containing radiolabel affords pure 7-chloro-3- 18 F-fluoromethyl-2H-1,2,4-benzothiadiazine 1,1-dioxide.  
     Example 6  
     4-ethoxy-1-hydroxy-3-(4-pyridinyl)-3-buten-2-one  
      1-hydroxy-3-(4-pyridinyl)-2-propanone (1 mole) is added to a cooled (0 degrees C.) solution of triethylorthoformate (1.05 mole), and acetic anhydride (1.05 mole) in acetic acid (2 mL) such that the exotherm does not exceed 45 degrees C. The mixture is allowed to cool to room temperature and stirred overnight, then concentrated in vacuo (&lt;75 degrees C.) to afford a crude oil, 4-ethoxy-1-hydroxy-3-(4-pyridinyl)-3-buten-2-one.  
     Example 7  
     1,2-dihydro-6-hydroxymethyl-2-oxo-5-(4-pyridinyl)-nicotinonitrile  
      The oil obtained in Example 6 is used directly and is dissolved in ethanol (1 l) and malononitrile (1 mole) is added. The mixture is heated at reflux for seven hours, during which time a precipitate forms. The mixture is cooled to room temperature and the resultant solid is filtered to afford the desired product, 1,2-dihydro-6-hydroxymethyl-2-oxo-5-(4-pyridinyl)-nicotinonitrile. Concentration of the filtrate affords a second crop of material of sufficient purity for use.  
     Example 8  
     1,2-dihydro-6-trifluoromethanesulfonatomethyl-2-oxo-5-(4-pyridinyl)-nicotinonitrile  
      A solution of 1,2-dihydro-6-hydroxymethyl-2-oxo-5-(4-pyridinyl)-nicotinonitrile (from Example 7) (10 mmole) and diisopropylethylamine (10.5 mmole) in DMF (20 mL) is cooled at 0 degrees C. while a solution of trifluoromethanesulfonic anhydride (10.5 mmole) in DMF (10 mL) is added. The reaction is stirred at 0 degrees C. for thirty minutes, then at room temperature for one hour. The solvent is removed in vacuo, and the residue is recrystallized to afford pure 1,2-dihydro-6-trifluoromethanesulfonatomethyl-2-oxo-5-(4-pyridinyl)-nicotinonitrile.  
     Example 9  
     1,2-dihydro-6- 18 F-fluoromethyl-2-oxo-5-(4-pyridinyl)-nicotinonitrile  
      A solution of 6-trifluoromethanesulfonatomethyl-2-oxo-5-(4-pyridinyl)-nicotinonitrile (from Example 8) in acetonitrile (1 mL), is added to a dry solution of K 18 F (50 mCi, via repeated azeotropic distillation of acetonitrile), kryptofix (2 mg) and potassium carbonate (50 ug). The vial is sealed and heated at 110 degrees C. for 15 minutes. The mixture is then cooled via active airflow, diluted with dichloromethane (4 mL) and passed through a Sep-Pak (silica gel, 230-400 mesh). The Sep Pak is further eluted with dichloromethane (4 mL) and the combined organics are concentrated. The resultant residue is dissolved in acetonitrile and purified via reverse-phase HPLC (C-18, acetonitrile:water 20% to 100% gradient). Concentration of the fractions containing radiolabel affords 1,2-dihydro-6- 18 F-fluoromethyl-2-oxo-5-(4-pyridinyl)-nicotinonitrile.  
      The disclosures of each patent, patent application, and publication cited or described in this document are hereby incorporated herein by reference, in their entireties.  
      Various modifications of the invention, in addition to those described herein, will be apparent to those skilled in the art from the foregoing description. Such modifications are also intended to fall within the scope of the appended claims.