Patent Publication Number: US-2007104646-A1

Title: Stabilized and lyophilized radiopharmaceutical agents

Description:
This is a continuation-in-part of a pending U.S. utility application Ser. No. 10/904,099 entitled Stabilized and Lyophilized Radiopharmaceutical Agents which is a continuation-in-part of provisional application No. 60/580,455 entitled Stabilized and Lyophilized Radiopharmaceutical Agents filed on Jun. 17, 2004 and a provisional application No. 60/608,060 of that name filed on Sep. 8, 2004, and a provisional application No. 60/522,619 filed on Oct. 20, 2004, and a continuation-in-part of U.S. application Ser. No. 11/611,862 filed Dec. 16, 2006 and Ser. No. 11/570,852 filed Dec. 18, 2006 which are US national stage entries of PCT/US2005/21847 which claim priority from the above 2004 applications and other applications as more fully set forth in PCT/US2005/21847. 
    
    
     FIELD OF THE INVENTION  
      The present invention relates to the method of preparation and stabilization of a diagnostic or therapeutic radiopharmaceutical useful, for example, in mammalian imaging and cancer detection, and resulting composition. In particular, the present invention relates to the novel method of preparation of radioactive diagnostic radiopharmaceutical in a stable, shippable, lyophilized form by an apparatus designed to rapidly flash freeze and dehydrate a radiopharmaceutical composition to minimize auto radiolysis, the novelty centering on rapid cooling and removal of ambient vapor, and then ultra cold removal when the potential of explosive liquid oxygen is eliminated. The radioactive diagnostic radiopharmaceutical requires no further cold or refrigerated storage, including with respect to shipping, subsequent to stabilization. The preferred composition can be reconstituted “on site” by the addition of a suitable diluent to bring the radiopharmaceutical complex into solution at a desired concentration at the time of administration to the patient in need of a therapeutic or diagnostic radiopharmaceutical. Heading  
     SUMMARY OF THE INVENTION  
      The present invention is directed to a stable radioactive diagnostic radiopharmaceutical composition that may be formed without stabilization additives and to a method of preparing such a composition. Stabilization additives may be added. Traditional techniques for freeze-drying (lyophilization) are subject to the lengthy crystal formation time of water. The composition is formed by avoiding that lengthy crystal formation time and the concurrent loss of diagnostic specificity due to autoradiolysis of the radiopharmaceutical. The length of traditional freeze-drying techniques and loss of diagnostic specificity due to autoradiolysis interfere with the technical accuracy necessary for nuclear medicine.  
      The novel technique of the inventors involves utilization of flash freeze techniques along with increasing the cold-exposed surface area and then rapidly decreasing the vapor pressure as well as super cold freeze drying of the radiopharmaceutical composition, the combination of which results in extremely rapid freeze-drying/lyophilization, enabling use of higher concentrations of radionuclides in the small scale amounts used in radiopharmaceutical imaging without damaging the ligands. The radiopharmaceutical composition can be reconstituted immediately prior to administration with confidence of little or no ligand damage, or little or no damage to the non-radioactive bonds and chemical structure of the composition.  
      The preferred composition results from forming a complex between a gamma emitting radionuclide and a ligand in a suitable solvent, generally an aqueous solution and then lyophilizing the solution by use of small quantities in large surface area vessels at vacuum pressure in conjunction with rapid sub-zero cooling. The radioactive diagnostic radiopharmaceutical in this invention requires no further cold or refrigerated storage, including with respect to shipping, subsequent to stabilization. The lyophilized radiopharmaceutical composition is shipped and stored and is often reconstituted “on site” by the addition of a suitable diluent to bring the radiopharmaceutical complex into solution at the time of administration to the patient in need of a therapeutic or diagnostic radiopharmaceutical. The present invention further is directed to stable radioactive diagnostic radiopharmaceutical compositions prepared by this method.  
     BACKGROUND OF THE INVENTION  
      With the invention of the Gamma Camera, and, just as importantly, with the invention of better high-speed imaging machines, pharmaceutical substances with radioactive “tags” have become extremely important in medical imaging and treatment. The concept is that a compound, or just as often, a part of a compound, called a ligand, sometimes referred to as an “agent” or which bonds to some other substance, is designed to target a particular area of a mammal&#39;s body or a particular type of tissue or molecule in that body. The compound, ligand or agent will be referred to as a ligand for convenience sake. The mammal this is most often used on is the human body, and references in this invention to a human are equally applicable to any mammal, or for that matter to any animal or plant.  
      For instance, certain ligands tend to concentrate in heart muscle tissue. The concept behind radiopharmaceutical imaging is to “tag” that ligand with a radioactive substance, i.e. radioactively mark a substance to create an “imaging agent,” so that a health care provider can find out where the ligand exists or is concentrating. By administering the radioactively tagged ligand, and placing the patient in an imaging machine, a health care provider can “look inside” a patient&#39;s body to assist in therapy or diagnosis. If a person has poor heart circulation, the radionuclide tagged ligand, such as Tc 99m TIBI, will not be well-circulated to areas of the heart muscle which have compromised blood flow, enabling evaluation of a person&#39;s “heart condition.” Importantly, the health care provider can often “look inside” without having to actually cut open or invade the body (non-invasive technique), or can minimize bodily invasion. Obviously, the continued presence of radioactive substances is not desirable, so substances are selected with a short “half-life.” The half-life is a time defined as the time in which the radioactive emission declines by one-half. The diminution of radioactivity is referred to as radioactive decay. Between the body washing out the radiopharmaceutical substances used in conjunction with this invention, and the use of substances with a short half-life, the amount of a patient&#39;s radioactive exposure is minimized.  
      Radioactive pharmaceuticals are in common use in imaging studies to aid in the diagnosis of a wide variety of illnesses including cardiac, renal and neoplastic diseases. These pharmaceuticals, known in the art as “imaging agents,” typically are based on a gamma-emitting radionuclide attached to a carrier molecule or “ligand.” Gamma-emitting radionuclides are the radionuclides of choice for conducting diagnostic imaging studies because, while gamma emitting radiation is detectable with appropriate imaging equipment, it is substantially less-ionizing than beta or alpha radiation. Thus, gamma emitting radiation causes minimal damage to targeted or surrounding tissues.  
      Radioactive pharmaceuticals now are finding increased use as diagnostic agents for finding neoplastic disorders, especially tumors. Diagnostic radiopharmaceuticals generally incorporate a gamma emitting radionuclide, the radiation emission being useful in the detection of certain neoplastic disorders.  
      The radioactive marking or tagging is often done by complexing the radioactive substance inside a group of ligands, that is surrounding it by a complex of ligands, so that the desired chemical characteristics are expressed toward the exterior of the complex with the tag shielded by the outer complex and simply carried along as a marker. The entire complex with the radioactive element, also called a radionuclide, functions as a radioactive marker, and can be more generally referred to as a radiopharmaceutical.  
      The use of small quantities of drugs used for such activities is desirable for cost reasons, and it is desirable to minimize the amount of radioactive substance used.  
      While the efficacy of radioactive diagnostic and therapeutic agents is established, it is also well known that the emitted radiation can cause substantial chemical damage or destabilization to various components in radiopharmaceutical preparations, referred to as autoradiolysis. Emitted radiation causes the generation of free radicals in water solutions, which free radicals are generally peroxides and superoxides. Such free radicals can precipitate proteins present in the preparations, and can cause chemical damage to other substances present in the preparations. Free radicals are molecules with unbonded electrons that often result because the emissions from the radioactive element can damage molecules by knocking apart water molecules forming hydroxyl radicals and hydrogen radicals, leaving an element or compound with a shell of charged electrons which seek to bond with other molecules and atoms and destabilize or change those molecules and atoms. The degradation and destabilization of proteins and other components caused by the radiation is especially problematic in aqueous preparations. Under the present art, the radiolysis causes the aqueous stored ligand and radioactive isotope bonded to the ligand to degenerate and destroys the complex which renders it useless for imaging because the biological characteristics that localize the complex to a tissue are gone. The degradation or destabilization lowers or destroys the effectiveness of radiopharmaceutical preparations, and has posed a serious problem in the art. Wahl, et al, Journal of Nuclear Medicine, Vol 31, Issue 1 84-89, discuss the fact that freezing radiolabeled antibodies at −70 degrees C. stabilizes the molecule for an indefinite period but 80 to 90% of the immunoreactivity is lost in as little as 24 hours when stored at 4 degrees C.  
      If the ligands are permitted to reside with the radioactive elements for an extended period, particularly in an aqueous (water-based) solution, the radiolysis is increased. Thus, any process to reduce the compounds to dried form has to be rapid and yield predictable result. Further, to avoid the higher concentrations and protect the ligands, presently the radiopharmaceutical solution is diluted, but that in itself only slows the drying time and complicates the problem and increases the unpredictability of the non-radioisotope portion of the radiopharmaceutical because of radiolysis. Heating the radiopharmaceutical in solution to accelerate the drying and removal of water has the undesirable effect of potentially damaging the ligand since chemical activity normally increases upon heating or injection of energy and therefore the effects of radiolysis are also increased during this prolonged drying period with heating. Most proteins are badly damaged upon heating. Certain ligands, such as isonitrile, simply evaporate and disappear upon heating. Further, minimization of localized heating at an atomic scale is important to preserve both the small quantities needed and to yield a specific concentration of desired product.  
      Wolfangel, U.S. Pat. No. 5,219,556, Jun. 15, 1993, entitled stabilized therapeutic radiopharmaceutical complexes, expressed his concern as follows: “The isotopes which are most useable with this process are determined by practical considerations. Again, Tc-99m would be a poor candidate for use since its six-hour half-life makes lyophilization impractical, as the lyophilization step itself generally takes about 24 hours to perform.” 
      Facially, the &#39;556 invention seemed to identify a useful process and resulting composition, but the lyophilization step in &#39;556 invention, as the application stated, took about 24 hours. The &#39;556 invention stated: “The lyophilization is carried out by pre-freezing the product, and then subjecting the frozen product to a high vacuum to effect essentially complete removal of water through the process of sublimation. The resultant pellet contains the complex in an anhydrous form which generally can be stored indefinitely, with practical consideration being given to the half-life of the radionuclide. The intended period of storage for radiopharmaceutical products is thus practically limited by the half-life of the radionuclides. In the case of Re-186, for example, the desired period of storage would range from 7 to about 30 days. Thus, this pellet can be shipped to the end users of the product and reconstituted with a diluent at the time of administration to the patient with very little effort on the part of the health care professional and/or nuclear pharmacist.” 
      Because the procedures in &#39;556 did not rapidly lyophilize the product, and contemplated a 24 hour period for lyophilization, the claims of &#39;556 invention were necessarily limited to utilization of a “therapeutic amount of an alpha- or beta-emitting radionuclide.” Wolfangel had observed that compounds with a half-life of at least 12 hours are preferred. By contrast, the use of Tc-99m, which also emits gamma rays, with a half-life of only six hours, or the use of other similarly short-lived radioisotopes, becomes impractical.  
      In a recent comprehensive text on the subject of lyophilization, a pre-eminent authority in the field made the following observations:  
      “Lyophilization is a multistage operation in which quite obviously each step is critical. The main actors of this scenario are all well known and should be under strict control to achieve a successful operation.  
      The product, . . .  
      The surrounding “medium” . . .  
      The equipment, . . .  
      The process, which has to be adapted to individual cases according to the specific requirements and low-temperature behavior of the different products under treatment.  
      The final conditioning and storage parameters of the finished product, which will vary not only from one substance to another one but in relationship with its “expected therapeutic life” and marketing conditions (i.e., vaccines for remote tropical countries, international biological standards, etc.). In other words a freeze-dryer is not a conventional balance; it does not perform in the same way with different products. There is no universal recipe for a successful freeze-drying operation and the repetitive claim that “this material cannot be freeze-dried” has no meaning until each successive step of the process has been duly challenged with the product in a systematic and professional way and not by the all-too-common “trial-and-error” game.  
      The freeze-drying cycle. It is now well established that a freeze-drying operation includes:  
     
         
         
           
              The ad hoc preparation of the material (solid, liquid, paste emulsion) to be processed taking great care not to impede its fundamental properties.  
              The freezing step during which the material is hardened by low temperatures. During this very critical period, all fluids present become solid bodies, either crystalline, amorphous, or glass. Most often water gives rise to a complex ice network but it might also be imbedded in glassy structures or remain more or less firmly bound within the interstitial structures. Solutes do concentrate and might finally crystallize out. At the same time, the volumetric expansion of the system might induce powerful mechanical stresses that combine with the osmotic shock given by the increasing concentration of interstitial fluids.  
              The sublimation phase or primary drying will follow when the frozen material, placed under vacuum, is progressively heated to deliver enough energy for the ice to sublimate. During this very critical period a correct balance has to be adjusted between heat input (heat transfer) and water sublimation (mass transfer) so that drying can proceed without inducing adverse reactions in the frozen material such as back melting, puffing, or collapse. A continuous and precise adjustment of the operating pressure is then compulsory in order to link the heat input to the “evaporative possibilities” of the frozen material.  
              The desorption phase or secondary drying starts when ice is being distilled away and a higher vacuum allows the progressive extraction of bound water at above zero temperatures. This again is not an easy task since overdrying might be as bad as underdrying. For each product, an appropriate residual moisture has to be reached under given temperatures and pressures.  
              Final conditioning and storage begins with the extraction of the product from the equipment. During this operation great care has to be taken not to lose the refined qualities that have been achieved during the preceding steps. Thus, for vials, stoppering under vacuum or neutral gas within the chamber is of current practice. For products in bulk or in ampoules, extraction might be done in a tight gas chamber by remote operation. Water, oxygen, light, and contaminants are all important threats and must be monitored and controlled. 
            Ultimate storage has to be carried according to the specific “sensitivities” of the products (at room temperature, +4 C −20 C). Again uncontrolled exposures to water vapor, oxygen (air), light, excess heat, or nonsterile environment are major factors to be considered. This obviously includes the composition and qualify of the container itself, i.e., glass elastomers of the stoppers, plastic or organic membranes.    
         
              At the end, we find the reconstitution phase. This can be done in many different ways with water, balanced salt solutions, or solvents either to restore the concentration of the initial product or to reach a more concentrated or diluted product. For surgical grafts or wound dressing, special procedures might be requested. It is also possible to use the product as such, in its dry state, in a subsequent solvent extraction process when very dilute biochemicals have to be isolated from a large hydrated mass, as is the case for marine invertebrates. [emphasis added as underlined material; italics in original]” 
           
         
       
    
      Rey, Louis and May, Joan C., editors, Freeze-Drying/Lyophilization of Pharmaceutical and Biological Products, pp. (Marcel Dekker, Inc., New York, Basel 1999 (Nat&#39;l Library of Medicine Call no. WI DR893B v. 96 1999)).  
      Professor Rey states in his introduction that he and two others led the first conference in 1958 on cryobiology, including freeze-drying of pharmaceuticals. Id. at 2.  
      Juxtaposing the most important underlined material from the above excerpt from Professor Rey&#39;s commentary based on his life-long experience, the following important principles are stated to be the art as of 1999: 
          a) “Lyophilization is a multistage operation in which, quite obviously, each step is critical. The main actors of this scenario are all well known and should be under strict control to achieve a successful operation.”    b) In other words, a freeze-dryer is not a conventional balance; it does not perform in the same way with different products. There is no universal recipe for a successful freeze-drying operation and the repetitive claim that “this material cannot be freeze-dried” has no meaning until each successive step of the process has been duly challenged with the product in a systematic and professional way and not by the all-too-common “trial-and-error” game.     c) The freeze-drying cycle. It is now well established that a freeze-drying operation includes: . . . 
            1). preparation of the material     2) The freezing step     3) The sublimation phase or primary drying will follow when the frozen material, placed under vacuum, is progressively heated to deliver enough energy for the ice to sublimate. During this very critical period a correct balance has to be adjusted between heat input (heat transfer) and water sublimation (mass transfer) so that drying can proceed without inducing adverse reactions in the frozen material such as back melting, puffing, or collapse. A continuous and precise adjustment of the operating pressure is then compulsory in order to link the heat input to the “evaporative possibilities” of the frozen material.     4) The desorption phase or secondary drying starts when ice is being distilled away and a higher vacuum allows the progressive extraction of bound water at above zero temperatures. This again is not an easy task since overdrying might be as bad as underdrying. For each product, an appropriate residual moisture has to be reached under given temperatures and pressures.     5) Final conditioning and storage . . .     6) Ultimate storage . . . . . . reconstitution phase . . . [emphasis added as underlined material; italics in original]”   
               

      The current patent art corroborates Professor Rey&#39;s assertion of the need for progressive heating, particularly the Wolfangel &#39;556 art, and Corbo et al art, U.S. Pat. No. 6,024,938, Feb. 15, 2000. Wolfangel &#39;556 contains the heating step that Professor Rey asserts is well-settled, while the present invention omits that step while achieving a superior art, and sets out a procedure that is not isolated to a particular product, but as useful for all radiopharmaceuticals. Further, the present invention presents an advantage of rapidity of process not seen in the prior art.  
      Notably, Wolfangel not only specifically also includes a heating step, but simultaneously and specifically states his invention is not applicable to short-half-life radionuclides. Corbo &#39;938 also contains the heating step, as does DeRosch, U.S. Pat. No. 6,428,768, Aug. 6, 2002, and all except Wolfangel take upwards of 24 hours. Thus, the later developing art is in fact moving to longer periods of time, notwithstanding the possible aspiration to a shorter time.  
      This invention thus defies the conventional wisdom by omitting the heating step, but lyophilizing, and dehydrating, and thereby stabilizing a radiopharmaceutical capable of storage at room temperature by a different technique, thereby achieving a superior result as demonstrated by the comparative experimental results discussed momentarily.  
      Wolfangel &#39;556 proposed in his example 1 to first lyophilize certain compounds, add the radionuclide complex, sparge with gas, seal the vial and then heat it. Unfortunately, the heating to 100 degree C. renders the procedure useless in conjunction with most proteins or peptides, and many commonly used complexes. Further, the proposal was to use 1 ml of sodium perrhenate Re-186 containing 1 mg of rhenium, with water added to produce 3 ml. The quantities contemplated were substantial and exposed the workers to substantial amounts of radiation. In example 3, it was proposed that the complex be frozen to −30 degree C. or colder and then apply a vacuum, but it was proposed to apply shelf heat at 6 degree per hour until a product temperature of 30 degree C. was reached, at which time the temperature would be held for two hours. That would require 12 hours. The procedure suffered from the infirmity of not quickly removing water and therefore not preventing radiolysis of the water and not preventing the generation of free radicals which damage the complexes. The second example 2 followed the first, but used smaller quantities, and proposed heating. Example 3 proposed heating to 85 degree C. for 30 minutes which would destroy most proteins and thereafter freezing and lyophilizing the sealed vials.  
      For diagnostic imaging purposes, radiopharmaceuticals based on a coordination complex comprised of a gamma-emitting radionuclide and a chelate have been used to provide both negative and positive images of body organs, skeletal images and the like. The Tc-99m skeletal imaging agents are well-known examples of such complexes. One drawback to the use of these radioactive complexes is that while they are administered to the patient in the form of a solution, neither the complexes per se nor the solutions prepared from them are overly stable. Consequently, the coordination complex and solution to be administered commonly are prepared “on site,” that is, they are prepared by a nuclear pharmacist or health care technician just prior to conducting the study. The preparation of appropriate radiopharmaceutical compositions is complicated by the fact that several steps may be involved, during each of which the health care worker must be shielded from the radionuclide.  
      The preparation of stable radiopharmaceutical diagnostic agents, due to the type of radioactivity, presents even greater problems. These agents typically are based on a relatively energetic gamma emitting radionuclide complexed with a chelate. Frequently, the radionuclide/chelate complex is in turn bound to a carrier molecule which bears a site-specific receptor. Thus, it is known that a gamma emitting radionuclide attached to a tumor-specific antibody or antibody fragment can destroy targeted neoplastic or otherwise diseased cells via exposure to the emitted ionizing radiation. Bi-functional chelates useful for attaching a diagnostic radionuclide to a carrier molecule such as an antibody are known in the art. See e.g. Meares et al., Anal. Biochem. 142:68-78 (1984).  
      For most imaging and diagnostic applications of radiopharmaceutical complexes of the types mentioned above, the nonradioactive portion(s) of the complex is prepared and stored until time for administration to the patient, at which time the radioactive portion of the complex is added to form the radiopharmaceutical of interest. For example, attempts to prepare radionuclide-antibody complexes have resulted in complexes which must be administered to the patient just after preparation because, as a result of radiolysis, immunoreactivity may decrease considerably after addition of the radionuclide to the antibody. In Mather et al., J. Nucl. Med., 28:1034-1036 (1987), a technique for labeling monoclonal antibodies with large activities of radio iodine using the reagent N-bromosuccinimide is described. The authors suggest that the antibodies labeled in this manner be administered to the patient immediately after preparation to avoid losses of immunoreactivity. Other examples of the preparation of the nonradioactive portion of the complex followed by on-site addition of the radioactive portion are disclosed in U.S. Pat. No. 4,652,440 (1987). Further, in many situations, the radioactive component of the complex must be generated and/or purified at the time the radiopharmaceutical is prepared for administration to the patient. U.S. Pat. No. 4,778,672 (1988) describes, for example, a method for purifying pertechnetate and perrhenate for use in a radiopharmaceutical.  
      According to Wolfangel &#39;556, EP 250,966 (1988) describes a method for obtaining a sterile, purified, complexed radioactive perrhenate from a mixture which includes, in addition to the ligand-complexed radioactive perrhenate, uncomplexed ligand, uncomplexed perrhenate, rhenium dioxide and various other compounds. Specifically, the application teaches a method for purifying a complex of rhenium-186 and 1-hydroxyethylidene diphosphonate (HEDP) chelate from a crude solution. Because of the instability of the complex, purification of the rhenium-HEDP complex by a low pressure or gravity flow chromatographic procedure is required. The purification procedure involves the aseptic collection of several fractions, followed by a determination of which fractions should be combined. After combining the appropriate fractions, the fractions are sterile-filtered and diluted prior to injection into the patient. The purified rhenium-HEDP complex should be injected into the patient within one hour of preparation to avoid the possibility of degradation. The rhenium complex may have to be purified twice before use, causing inconvenience and greater possibilities for radiation exposure to the health-care technician.  
      While the lyophilization process has been applied to various types of pharmaceutical preparations in the past, the notion of lyophilizing short lived gamma emitting radiopharmaceutical preparations has not been addressed. In part, this is believed to be due to skepticism of those skilled in the art that such a procedure could be safely carried out. U.S. Pat. No. 4,489,053 (Azuma et al.; Dec. 18, 1984) relates to Tc-99m-based diagnostic imaging agents. The patentee notes that the non-radioactive agents may be prepared in lyophilized form and that stabilizers are required to prevent radiolysis once the Tc-99m is added.  
      Thus, there is a need in the art for a method of centrally preparing and purifying a stabilized diagnostic radiopharmaceutical for shipment to the site of use in a form ready for simple reconstitution prior to its administration in diagnostic applications without the necessity of additional stabilizers. Because of the length of the Wolfangel process, many of the protein combinations with radionuclides are impractical because of the sensitivity of the protein in combination to any free radical attack caused by radioactive decay, and thus the present invention is a novel means to enable practical commercial use of radionuclide labelled proteins and peptides. The length also effectively prohibits the use of shorter half life radionuclides because in order to use them with the Wolfangel process, the concentrations of the radionuclides have to be increased to account for the several half lives during the 24 hours lyophilization and the time for shipment, which concentration exposes workers to higher concentrations of radioactivity and which time exposes the ligands to radiolysis which decreases their predictability of use in the patient, if they are effective at all. If, in order to avoid the higher concentrations, more dilute amounts are used, then the quantity of liquid involved jeopardizes the efficacy of lyophilization. There is a particular need in the art for a method of centrally preparing and purifying radionuclide-labeled antibodies and antibody fragments, owing to their relatively unstable immunoreactivities once in aqueous solution. Most particularly, this invention enables the use of short-half-life radionuclides with ligands potentially subject to radiolysis that are stable with useful shelf life at room temperatures that can be shipped in a commercially cheaper manner, and easily reconstituted.  
     OBJECTIVES OF THE INVENTION  
      An object of the invention is to accelerate the removal of water to minimize the peroxidation-related effects of radiolysis because of the accelerated removal of water which facilitates stabilization and predictability of concentration of a ligand or non-radioactive portion of a radiopharmaceutical because of reduced radiolysis.  
      An object of the invention is to use the minimization of peroxidation-related effects to improve the preservation of the chemical substituent complexes typically surrounding a radionuclide.  
      An object of the invention is to use small quantities at concentrations which enable accelerated lyophilization, longer predictable storage and overnight shipment, and increase worker safety. Corollary to this objective is the elimination of need for cold storage and refrigeration.  
      An object of the invention is to use vials with an expanded surface area, extremely cold temperatures and very low level pressures in combination to accelerate lyophilization.  
      An object of the invention is to use a two stage system to accelerate lyophilization by not only lowering vacuum pressure, but also, after initial removal of oxidizing agents, to extract vapor more rapidly by supercooling gas being evacuated.  
      An object of the invention is to create a stable vehicle for delivering selectively toxic radionuclides to target tissues. 
    
    
     DETAILED DESCRIPTION OF THE INVENTION  
      In contrast to the Wolfangel &#39;556 invention which stated: “the lyophilization step itself generally takes about 24 hours to perform,” the present invention proposes to produce a stable radiopharmaceutical complex by a lyophilization process which “freeze-dries” the complex in five hours or less, normally 2-4 hours, and then requires no further refrigeration.  
      The preferred mode of the invention is utilized in conjunction with Iodine-123 (“I-123” (123 being the sum of the protons and neutrons)) radionuclides. The following illustrates the compositions and processes of this invention, but is not meant to limit the scope of the invention in any way.  
      An I-123 labelled compound such as meta-iodo-benzyl-guanidine (“MIBG”) is prepared. The concentration is increased so that ultimately one-half milliliter or less will equal one dose. For example the usual does of I-123 MIBG for a typical patient would be 10 mCi (millicuries). Because the half life is 12 hours, in order to allow for normal radioactive decay in shipment so that the dose is 10 mCi upon administration, 36 mCi would be mixed on the prior day anticipating overnight shipment.  
      The condensing system is heavily insulated.  
      A hose runs from the top or side of the stainless steel pot of the primary condenser to the vacuum pump.  
      A vacuum pump capable of producing a vacuum of at least 10-4 Torr would be used to evacuate the chamber. An appropriate vacuum pump is model RV-12 available from BOCEdwards, an international company, of Wilmington, Massachusets, which can be contacted through the internet.  
      In order to achieve the composition contemplated in this invention, the primary condensing coil is readied at or below −40 deg. C. Promptly after mixing the radiopharmaceutical composition, the vial containing the radiopharmaceutical composition, in the preferred mode the 0.36 ml. of aqueous I-123MIBG, is stoppered with the lyophilization stopper, with the lyophilization stopper in a position to permit passage of vapour. The vial and stopper will be fully sealed at the end of the process.  
      The vial(s) is (are) placed into the tray and a sufficient amount of liquid nitrogen is poured onto the tray in order to flash freeze the vials by the heat transfer from the aqueous I-123MIBG through the sides of the vial. Because of the small quantity which is used and the high surface area of the vial, the freezing occurs virtually instantaneously. The tray is placed into a stoppering frame in the chamber with the inner tube connected and installed so that at the end of the procedure, before the vacuum is broken, the port to the inner tube can be opened and the tube will inflate and force the stoppers fully into the vials in order to seal them.  
      As the liquid nitrogen evaporates off, a thermistor on one of the vials is connected to the electrical connector on the rubber stopper which connects to an outside temperature monitoring device. The liquid nitrogen is allowed to evaporate, all the while maintaining the temperature of the vial at or below −10 degrees C.  
      The top of the chamber is installed and forms a seal with the cylindrical side of the chamber. After evaporation of the liquid nitrogen, the gas valve on top of the chamber is closed, and the rubber stopper is installed.  
      After the tray containing the flash-frozen vials is placed into the chamber, and the chamber has been sealed, the vacuum pump is turned on. A vacuum pressure is first felt in the primary condenser and any vapor in the chamber begins to flow out through the secondary condenser and freezes in the primary condenser which is kept at a temperature above the boiling point of oxygen, meaning preferably kept at about −40 degrees C. A reasonably skilled practitioner in the art would recognize that at 10-2 Torr and −40 degree C. the amount of oxygen present would be sufficiently low that the danger of oxygen oxidation damage from liquid oxygen if the temperature is lowered below −40 degree C. is eliminated. The preferable level for activating the secondary condenser is 10-3 Torr. When the vacuum pump gauge shows the preferred level of 10-3 Torr, usually after about 20 minutes, liquid Nitrogen at −196 degrees C. is allowed to flow through the secondary condenser and cool the stainless steel tube contained in the secondary condenser through which gas evacuated from the chamber is flowing. The very cold liquid Nitrogen in the secondary compressor is used to increase the temperature difference between the secondary condenser and the vial contents to accelerate the lyophilization. The secondary condenser is placed in series with the primary condenser and the evacuated chamber containing the tray of vials. The secondary condenser takes over as the larger and faster heat sink to capture the vaporized water. A reasonably skilled practitioner would understand that the vacuum pump continues to run to the end of the procedure, and the pressure continues to drop to the rated capacity of the vacuum pump. A reasonably skilled practitioner would know that the pump referenced, the model RV-12 available from BOCEdwards, has a rated capacity of approximately 10-6. Thus, after the system has been sealed and the pump is turned on, the pressure drops through the 10-2 Torr and 10-3 Torr levels to the rated capacity of the vacuum pump.  
      Because the acrylic chamber has no refrigeration, the temperature of the vial and the vial contents tend to rise above 0 degrees C. after all of the water is removed. This signals the completion of the cycle. The thermistor probe connected through the rubber stopper to the outside monitoring device enables the monitoring of the vial temperature. The vials would then be sealed in partial pressure of pharmaceutically inert gas that is fully dehydrated or “dry,” meaning gas that is non-reactive with the pharmaceutical composition, the gas preferably being argon or nitrogen. An inner tube will have been placed in the chamber to be inflated to force the stoppers into the vial to seal them. An auxiliary cylinder of gas that is chemically inert relative to the lyophilized radionuclide is used to gradually inflate the inner tube through the valve to force the stoppers into the vials. The vacuum is broken. The vial stoppers further secured with an aluminum seal. At the end of the process upon warming, the water which was frozen and subsequently melted will be drained from the primary condenser.  
      The vials are ready to be shipped with predictable half lives for the radionuclide and a stabilized ligand in powdered form.  
      If it is desired to accelerate the lyophilization process, inert gas may be admitted through the gas valve into the chamber to displace any oxygen and enable the secondary condenser to be turned on sooner. The displacement is necessary to prevent accumulation of liquid oxygen in the secondary condenser. In the ordinary procedure, if the secondary condenser is activated before the 10-3 level is reached, there is a risk of collecting liquid oxygen which is potentially explosive.  
      The secondary condenser is in series with the primary condenser, and could be located subsequent to the primary condenser in the evacuation and condensing system.  
      The speed of the lyophilization process is positively influenced by the lowering of the vapor pressure external to the material being dried. Secondly, the speed is positively influenced by the greater temperature difference between product being cooled and the temperature of the condenser where the water is being collected.  
      The radioactive diagnostic radiopharmaceutical in this invention requires no further cold or refrigerated storage, including with respect to shipping, subsequent to stabilization. The lyophilized radiopharmaceutical composition is reconstituted “on site” for administration to patients by the addition of a suitable diluent to bring the radiopharmaceutical complex into solution at the time of administration to the patient.  
      For administration, the I-123 labelled MIBG in the vial must be reconstituted. Because of the minute quantity of material, the vial of radionuclide complex, in the preferred mode the I-123 labelled MIBG will appear empty. The MIBG ligand is stable for several days because of the absence of water which is the primary substance from which free radicals are generated by gamma ray collisions with water molecules. The gamma rays are being emitted by the radionuclide, that is the I-123. The health care provider would add up to 2 ml. of sterile normal saline. The desired dose would be withdrawn and measured in a dose calibrator of a type manufactured by Capintec of Montville, N.J. If the glass vial is measured in the dose calibrator, the person measuring the dose must recognize that the glass vial will decrease the apparent activity. Upon calibration of the desired dose, the I-123 MIBG now re-dissolved in the solution is promptly administered to the patient.  
      The advantages are that the flash freezing and lowering of vapor pressure result in quick formation and evaporation or sublimation (evaporation from ice to water vapour (a gas)) of water from the I-123 MIBG. The I-123 MIBG need not be shipped frozen in dry ice nor need it be shipped for overnight delivery. Shipping in dry ice over a weekend is generally not commercially practical. The I-123 MIBG can be shipped over the weekend and be used on Monday while simply maintaining it at room temperature or below.  
      In order to establish the advantages of the novel process and resulting composition, a series of tests were run utilizing meta iodo benzyl guanidine (MIBG) in which the radionuclide I-131 was the iodine in the MIBG.  
      The MIBG was prepared as follows: eight vials were prepared of MIBG in solution with a radioactive concentration of MIBG of 1 mCi per vial. The MIBG in six of those vials were then stabilized and lyophilized according to the process described in this invention. One vial was frozen and maintained at a temperature of −10 degrees, and another vial was simply refrigerated at approximately 5 degrees.  
      Six vials were prepared according to the process in this invention in order to enable several to be reconstituted from the lyophilized state and their activity tallied.  
      The radioactive concentration of MIBG per vial was 1 mCi per vial.  
      The results showing the percent of iodine remaining bound to the MIBG are set forth in table I. One each of the vials was reconstituted after 24, 48, 72 and 168 hours respectively.  
                                                               0   24                       hours   hours   48 hours   72 hours   168 hours (1 wk.)                                                            Lyophilized   96.3%   97%   96.6%     96.2%     95.9%         and       stabilized       per       invention       stored at       room temp.       Frozen −10°   96.3%   94%   91%   84%   72%       Refrigerated   96.3%   92%   85%   77%   55%       ˜+5°                  
 
      In sum, the radiolysis damage was virtually eliminated from the composition stabilized and lyophilized under this invention while, as the prior art suggests, MIBG that was not so stabilized and lyophilized per this invention deteriorated sharply in activity.  
      As another example, I-131 Hippuran was prepared. The I-131 Hippuran was prepared as follows: 9 vials were prepared of I-131 Hippuran in solution with a radioactive concentration of MIBG of 1 mCi per vial. Each vial had 4 cc. The I-131 in seven of those vials was then stabilized and lyophilized according to the process described in this invention. One vial was frozen and maintained at a temperature of −10 degrees, and another vial was maintained room temperature. Room temperature was selected because Hippuran is thought to be stable at room temperature even in conjunction with a radioisotope.  
      The results showing the percent of Hippuran remaining bound to the I-123 are set forth in table 2. One each of the vials was reconstituted after 24, 48, 72 and 168 hours respectively.  
                                           TABLE 2                                   0 hours   24 hrs   48 hrs   72 hrs   96 hrs   120 hrs                                                                Lyophilized   98%   98.4%   98.6%     98%   98.4%   98.5%         and stabilized       per invention       stored at room       temp.       Frozen −10°   98%   97.8%   97%   94%   92.5%   91       Room Temp.   98%     96%   95%   94.5%       92%   90%                  
 
      In sum, the radiolysis damage was virtually eliminated from the composition stabilized and lyophilized under this invention.  
      If one desires to ship product, maintaining a product reliably frozen even at −10 degrees is difficult and expensive as a practical matter; this invention makes such shipment practical over the techniques of the prior art. One reference has suggested that storage at −70° C. can limit autoradiolysis damage, but even in that article, the percent free iodine, e.g. unbonded iodine, rose from what appears to be 1.6% to 4.3% in 24 hours. Wahl, Inhibition of Autoradiolysis of Radiolabeled monoclonal Antibodies by Cryopreservation, 31(1) J. Nucl. Med. 84-89 (January 1990). Conversely, putting those results in a form analogous to Table I, the percentage of free iodine in the Wahl article commenced at 98.4% and fell in 24 hours to 95.7% in Wahl&#39;s Table 1. The contrast between that fall in bonded iodine in 24 hours of some 3.7% in the Wahl reference versus a fall of 0.4% during a week for the composition stabilized and lyophilized per this invention illustrates the sharp advantage of the present method and resulting composition. In addition, it is not practical in real-world conditions to replenish the cooling fluid to maintain −70° C. much less to ship it cost-effectively.  
      The micro quantities involved for radionuclide complexes such as I-123 MIBG substantially reduce the exposure of production workers and health care providers because minute quantities are involved.  
      More generally, the preferred mode will use compounds that have a half life of one hour to a maximum of 12 hours. Longer half lives are less used because of slower radioactive decay exposing the body to increased radiation. It is generally preferable to apply the flash-freezing first because application of the reduced pressure may cause the solution to boil out of the vial.  
      Applying the invention more generally, the intent is to utilize the invention to produce stabilized radiopharmaceutical compositions. Such stabilized radiopharmaceutical compositions include radionuclides which are combined with ligand useful for diagnosis or diagnostic treatment or therapy to form radiopharmaceutical complexes in solution or suspension. These complexes then are lyophilized in accord with the above procedure according to the desired radioactivity level for the selected radionuclide. The form of radiopharmaceutical composition lyophilized according to this invention can be stored until needed for use. This invention allows for the central preparation, purification and shipment of a stabilized form of a radiopharmaceutical complex which merely is reconstituted prior to use. Thus, complicated or tedious formulation procedures, as well as unnecessary risk of exposure to radiation, at the site of use are avoided.  
      The radioactive diagnostic radiopharmaceutical in this invention requires no further cold or refrigerated storage, including with respect to shipping, subsequent to stabilization.  
      The term “radiopharmaceutical composition” includes any chemical composition including a radionuclide. Such term “radionuclide” includes cyclotron-produced radionuclides including those referenced in Table 1 on page 7 of M. Welch and C. Redvanly, Handbook of Radiopharmaceuticals: Radiochemistry and Applications (John Wiley &amp; Sons, Ltd, Chichester, West Sussex, England 2003) (hereafter “Handbook of Radiopharmaceuticals”), Table III on p. 77 of the Handbook of Radiopharmaceuticals, and throughout chapters 1 and 2 of the Handbook of Radiopharmaceuticals. Such term “radionuclide” includes reactor-produced radionuclides including those referenced in Table 2 on page 98 of the Handbook of Radiopharmaceuticals and throughout chapter 3 of the Handbook of Radiopharmaceuticals. Radionuclide also includes radioactive isotopes of any element referenced in the Table 1 and Table 2 referenced in this paragraph, and includes Cu64 (which has traditionally not been recognized as useful), Fe, including Fe52 and 5959 and Fe3+ radioisotopes, Yt, and Bi. Details of Gallium, Indium, and Copper radionuclides included are referenced in Tables 1 on page 264, Table 4 on page 374, and Table 1 on page 402 of the Handbook of Radiopharmaceuticals, respectively. Other useful radionuclides, which sometimes overlap those of Table 1 and Table 2 just referenced can be found for iodine radionuclides at p. 424 of the Handbook of Radiopharmaceuticals, and bromine radionuclides at p. 442 of the Handbook of Radiopharmaceuticals. The Technetium radionuclides and technetium radiopharmaceutical compositions are included. The term radiopharmaceutical composition is intended to be comprehensive because of the utility of the invention to radiopharmaceuticals and their longer-term preservation. Therefore, the term is defined to include the ligands bonded with radionuclides, compounds in which the radionuclide is integral to the ligand or compound, and compounds or mixtures in which the radionuclide is complexed. Accordingly, further amplification of the comprehensive scope of radiopharmaceutical composition is given herein.  
      The term “radiopharmaceutical composition” includes isotopes that are beta particle emitters, including those listed in Table 2 on page 773 of the Handbook of Radiopharmaceuticals, and Fe52, Cu64, Cu67, Ga68, Br77 and 1124.  
      The term “radiopharmaceutical composition” includes radionuclides bonded to a ligand. For the purposes of this application, the term “ligand” is taken to mean a bio-compatible vehicle, typically a molecule, capable of binding a radionuclide and rendering the radionuclide appropriate for administration to a patient. Thus, by way of illustration and not limitation, the term ligand encompasses both chelating agents capable of sequestering the radionuclide (usually a chemically-reduced form of the radionuclide) as well as carrier molecules, such as lipophilic cations with radioisotope labeling, antibodies, antibody fragments, fatty acids, amino acids or other peptides or proteins. The term radiopharmaceutical composition includes receptor specific agents, tumor agents, tumor associated antigen, antithrombotic GPIIb/IIa receptor antagonists, agents for neuroreceptors/transporters and amyhloid plaque, BZM, and monoclonal or polyclonal antibodies, particularly in Tc radiopharmaceuticals where preservation of the ligand is important (a general summary of which is on p. 349 of the Handbook of Radiopharmaceuticals). The application of the invention to compounds for assessment of multi-drug resistance status is contemplated. Chelating agents can include bifunctional and multifunctional chelates. A non-exhaustive list of chelating agents is referenced on pages 366 and page 376 of the Handbook of Radiopharmaceuticals. Included in the term ligand are antibodies bound via a chelate. Such antibodies may include monoclonal antibodies or polyclonal antibodies. Other ligands contemplated include neuroreceptor imaging agents, and receptor imaging agents, and myocardial sympathetic nerve imaging agents, many of which are referenced in Handbook of Radiopharmaceuticals. The carrier molecules often are specifically targeted at a tumor cell or tumor-specific antigen, an organ or a system of interest for observational and consequent diagnostic purposes, or in need of therapy. Carrier molecules may be directly labeled with the radionuclide, in which case any pharmaceutically acceptable counter-ion for the therapy or diagnostic intended may be used. The radionuclide may be bound to a carrier molecule via a chelate or other binding functionality. The term “complex” is taken to mean, broadly, the union of the radionuclide and the ligand to which it is attached. The chemical and physical nature of this union varies with the nature of the ligand. The invention includes compounds in the Handbook of Radiopharmaceuticals seeking receptors, including so-called antagonists which fit receptors, a partial, but fairly complete list of which is found on pages 452-457 and 717 of the Handbook of Radiopharmaceuticals.  
      The term “radiopharmaceutical composition” refers to a composition including the radionuclide-ligand complex as well as suitable stabilizers, preservatives and/or excipients appropriate for use in the preparation of an administrable pharmaceutical. The invention contemplates that for certain large proteins susceptible to breaking from the freezing process, such large protein structures would be supported by a lyophilization aid known to reasonably skilled practitioners in the art of pharmacy such as lactose, dextrose, albumin, gelatin or sodium chloride.  
      The term “radiopharmaceutical composition”, includes, for therapeutic purposes, therapeutic radionuclides, including Auger electron emitters such as those described on pages 772 and 776 of the Handbook of Radiopharmaceuticals. Auger electron emitters can be useful because they can result in additional deposition of energy in tissue as to which radiopharmaceutical damage is desired. Such damage is generally desired to be minimized in diagnostic uses.  
      The general method of this invention, and the composition contemplated to be created can be implemented on a general basis as follows: after a radiopharmaceutical composition is prepared by known methods appropriate to the composition, aliquots of the radioactive complex are aseptically dispensed into sterile vials consistent with the procedure outlined and the radioactive product is lyophilized according to the procedure of this invention to produce the stable lyophilized powder. The virtually complete absence of water results in a substantial improvement in the stability of the preparation, from both radio chemical purity and chemical purity standpoints, versus prior preparations. The stabilized complex can be prepared several days in advance, shipped and stored until needed for use. The preferred mode of the invention is focused on radionuclides that are gamma emitters of diagnostic value and with a half-life sufficiently long to make the preparation, lyophilization and shipment of the compounds practical, but the invention is useful for alpha- and beta-emitting radionuclides.  
      As an example of an additional preferred mode of invention, Cu64 can be complexed with zinc isonitrile and Cu64 isonitrile can be used for PET (Positron Emission Tomography) imaging. Without the use of the process and composition of Cu64 isonitrile described herein, the half-life of Cu64 is such that its use as an imaging agent is relatively impractical. For cardiac imaging, the use of an I123 or I124 isotope in combination with a fatty acid is useful on a broader patient base than the current commonly used FDG imaging. In order to use 2-deoxy-2-[18F]fluoro-D-glucose [18FDG] for imaging the heart, the heart must be converted from fatty acid metabolism to glucose metabolism which is accomplished by feeding the patient high levels of glucose, usually three or four candy bars and waiting for approximately an hour. This is unhealthy for diabetics. This invention enables the use of shorter half-life compounds and in particular the I123 or I-124 fatty acid radiopharmaceuticals and eliminates the necessity of conversion of the heart from fatty acid metabolism to glucose metabolism. This process and the composition of the invention present a novel opportunity to use radioisotopes of shorter half-lives. I-124 radionuclides generally, and I-124 fatty acid radiopharmaceuticals can be used in conjunction with PET imaging.  
      Another preferred mode of invention is to use I124 MIBG for neuroendocrine imaging and I124 fatty acids both stabilized by the lyophilization process in this invention. Once again, only with the invention is the use of I124 practical to sufficiently concentrate the I124 while preserving the integrity of the overall I124 radiopharmaceutical composition. The use of I123 radionuclides is also made more practical by this invention, particularly in conjunction with fatty acid labeling.  
      At the point of use, the radiopharmaceutical compositions of the present invention are prepared for administration to a patient. Such preparation advantageously merely involves reconstitution with an appropriate diluent to bring the complex into solution. This diluent may be sterile water for injection (SWFI), dextrose and sodium chloride injection or sodium chloride (physiological saline) injection, for example. The preferred diluent is water for injection or physiological saline (9 mg/ml) which conforms to the requirements listed in the U.S. Pharmacopeia.  
      The present invention is particularly well suited for the preparation of stable, pre-labeled antibodies for use in the diagnosis and treatment of cancer and other diseases. For example, antibodies expressing affinity for specific tumors or tumor-associated antigens are labeled with a diagnostic radionuclide, either directly or via a bi-functional chelate, and the labeled antibodies are stabilized through lyophilization. Where a bi-functional chelate is used, it generally is covalently attached to the antibody. The antibodies used can be polyclonal or monoclonal, and the radionuclide-labeled antibodies can be prepared according to methods known in the art. The method of preparation will depend upon the type of radionuclide and antibody used. The stable, lyophilized, radio labeled antibody merely is reconstituted with suitable diluent at the time of intended use, thus greatly simplifying the on site preparation process. The process of this invention can be applied to stabilize many types of pre-labeled antibodies, including, but not limited to, polyclonal and monoclonal antibodies to tumors associated with melanoma, colon cancer, breast cancer, prostate cancer, etc. Such antibodies are known in the art and are readily available. Other ligands with specific affinities to sites in need of radiotherapy are known in the art and will continue to be discovered.  
      The radiopharmaceutical composition which results from the method of this invention may be further purified after reconstitution, if desired. One method of purification is described in EP 250966, noted above. Other methods are known to those skilled in the art.  
      The radiopharmaceutical composition can include other components, if desired. Useful additional components include chemical stabilizers, lyophilization aids and microbial preservatives. Such chemical stabilizers include ascorbic acid, gentisic acid, reductic acid, para-amino benzoic acid, and erythorbic acid among others. In some cases, these agents are beneficial in protecting the oxidation state of the radionuclide by preferential reaction with oxygen or by direct effect. The term lyophilization aids includes those substances known to facilitate good lyophilization of the product. These aids are used to provide bulk and stability to the dried pellet and include lactose, dextrose, albumin, gelatin, sodium chloride, mannitol, dextran and pharmaceutically-acceptable carriers, among others. Antimicrobial preservatives inhibit the growth of or kill microbial contaminants which are accidentally added to the product during preparation. The term antimicrobial preservatives includes methylparaben, propylparaben and sodium benzoate. These components generally are added to the composition after the complex has been formed between the ligand and the radionuclide but prior to lyophilization. Bacteriastatic agents, for example, methyl and propyl-paraben may be added. Also contemplated are the addition of solubilizing agents such as polyethylene glycol to enhance the solubility of fatty acid compounds tagged with radionuclides in normal saline solution or other water based solutions.  
      The above process, apparatus and resulting composition is adaptable to the stabilization and preservation of virtually all radionuclides whatever the solvent used for initial composition. Some preferred applications include stabilization of radiolabeled peptides, [18 F] deoxyglucose, radiolabelled annexin, 99 mTc-annexin, radiolabelled monocyte chemoattractant protein. i.e. 125-I-(MCP-1), radiolabelled Dopamine transporter agents, (S)-N-(1-ethylpyrrolidin-2-ylmethyl)-2-hydroxy-3-iodo-6-methoxybenzamide (3-IBZM)(More generally “BZM,), (S)-N-(1-ethylpyrrolidin-2-ylmethyl)-2-hydroxy-5-iodo-6-methoxybenzamide (5-IBZM), 1-123-2-beta-carbomethoxy-3-beta(4-iodophenyl) N-(3-fluro propyl) nortropane (“CIT” or “beta-CIT”) and various tropane derivatives, I-123 fatty acids, particularly for cardiovascular imaging, radiolabelled octreotide or radiolabelled depreotide, HEDP (diagnostic skeletal imaging or treatment of metastatic bone pain), radiolabelled antibodies, both polyclonal and monoclonal, with selective affinities for tumor-associated antigens diagnosis or in situ radiotherapy of malignant tumors such as melanomas), and ligands with selective affinity for the hepatobiliary system (the liver-kidney system), including 2,6-dimethylacetanilideiminodiacetic acid and the family of other imidoacetic acid group-containing analogs thereof (collectively referred to herein as “HIDA agents”), mono-, di- and polyphosphoric acids and their pharmaceutically-acceptable salts including polyphosphates, pyrophosphates, phosphonates, diphosphonates and imidophosphonates. Preferred ligands are 1-hydroxyethylidene diphosphonate, methylene diphosphonate, (dimethylamino)methyl diphosphonate, methanehydroxydiphosphonate, and imidodiphosphonate (for bone-scanning and alleviation of pain); strontium 89 ethylene diamine tetramethylene phosphate, samarium 153-ethylene diamine tetramethylene phosphate, radiolabelled monoclonal antibodies, 99m-Tc HMPAO (hexamethylproplyene amine oxime), yttrium 90-labeled ibritumomab tiuxetan (Zevalin® Registered Trademark of Biogen Idec, Inc.), and meta-iodo-benzyl guanidine. Ethylene diamine tetramethylene phosphate and ethylene diamine tetramethylene phosphoric acid and the pharmaceutically related mono-, di- and polyphosphoric acids and their pharmaceutically-acceptable salts including polyphosphates, pyrophosphates, phosphonates, diphosphonates and imidophosphonates are collectively called EDTMP.  
      Suitable radionuclides which are well-known to those skilled in the art include radioisotopes of copper, technetium-99m, rhenium-186, rhenium-188, antimony-127, lutetium-177, lanthanum-140, samarium-153, radioisotopes of iodine, indium-111, gallium-67 and -68, chromium-51, strontium-89, radon-222, radium-224, actinium-225, californium-246 and bismuth-210. Other suitable radionuclides include F-18, C-11, Y-90, Co-55, Zn-62, Fe-52, Br-77, Sr-89, Zr-89, Sm-153, Ho-166, and Tl-201.  
      The invention is not meant to be limited to the disclosures, including best mode of invention herein, and contemplates all equivalents to the invention and similar embodiments to the invention for humans, mammals and plant science. Equivalents include combinations with or without stabilizing agents and adjuncts that assist in reservation, and their pharmacologically active racemic mixtures, diastereomers and enantiomers and their pharmacologically acceptable salts in combination with suitable pharmaceutical carriers.