MXPA01004429A - Manganese compositions and methods for mri - Google Patents

Abstract

An MRI contrast medium of improved safety and efficacy is disclosed. The composition includes a source of calcium ions and a source of manganese ions in a ratio of from 2:1 to 40:1 in a vehicle suitable for parenteral administration. A method of enhancing an MRI signal in a mammalian tissue with the foregoing composition is also provided.

Description

MANGANESE COMPOSITIONS AND METHODS FOR MRI Field of the Invention The invention relates to compositions and methods for improving magnetic resonance imaging of tissues, systems and organs.

Background of the Invention Magnetic resonance imaging allows non-invasive visualization of tissues and organs of the human body. The contrast in the images generated can be increased through the use of an agent that alters the relaxation of the water in the tissues of interest in relation to the volume of water. Species with non-paired electrons, such as the lanthanide metal and paramagnetic transition ions, can be used for this purpose. Manganese chloride was investigated as a contrast agent by Lauterbur and by Wolf in animal models. Both investigators demonstrated significant increase in liver and other organs (but not blood) through the use of manganese chloride, but determined that the potential clinical utility of the agent was limited by acute cardiac toxicity. The development of contrast agents based on other paramagnetic metal ions is similarly constrained by toxicity and solubility. For example, gadolinium chloride, acetate and sulfate demonstrate poor tolerability, including symptoms of heavy metal poisoning and gadolinium accumulation in the liver, vessel and bone.

Praramagnetic transition chelates and lanthanide metals have been used with some success in imaging diagnostics to overcome both toxicity and solubility problems (see U.S. Patent No. 4,647,447). The application of this technology has allowed the development of several gadolinium-based MR contrast agents including Gd-DTPA (Magnevist1 ™ 1, Schering), Gd-DTPA-BMA (Omniscan ™™, Nycomed Amersham PL), Gd- HP-D03A (Prohancemarca, Braceo Diagnostics) and Gd-DOTA (Dotaremarca, Guerbet), as well as the manganese-based contrast agent MnDPDP (Teslascann? Arca, Nycomed Amersham PL). There are, however, disadvantages associated with the use of chelation to solve the problems of toxicity. The metal ions are not maintained irreversibly in the chelate complexes, but are subjected to the balance between the united and free states. In vivo, this balance is further complicated by the balance between the chelator and the endogenous metal ions as well as between the paramagnetic metal ion and the endogenous ligands. Greis, in U.S. Patent No. 5,098,692 and Bosworth, in U.S. Patent No. 5,078,986 describe the use of an excess chelator to minimize the amount of free metal ion in diagnostic compositions based on of chelate. However, the potential for dissociation of the metal ion from the complex remains. The free chelator, either in excess or released, can introduce additional toxicity by itself or through the chelation of those endogenous metal ions that are required as cofactors for essential enzymes or for other biological functions. Unfortunately, the chelates also demonstrate a reduced solution relaxivity in relation to free metal ions. The interaction of the paramagnetic metal ions with the water molecules, which shortens the proton relaxation time in relation to the volume of water and results in an improvement of signal, is obtained by chelation, since the same sites of Metal-water interactions are used to form non-covalent associations between the metal and the chelate. Thus chelation provides security at the price of a reduced imaging efficiency (as compared to free metal ion.) In practice this loss of efficacy can be as high as 60-80%.

Manganese chelate imaging enhancement agents are known, for example MnDPDP, MnDTPA, MnEDTA and its derivatives, Mn porphyrins such as MnTPPS, and fatty acyl DTPA derivatives. These manganese chelates are not known because they agglutinate endogenous macromolecules such as the manganese ion. As a consequence, the increased efficiency seen by the Mn ion after the macromolar association is seen for the Mn chelates only as a function of the rate at which and to the extent to which the manganese ion is dissociated from the complex. This results in the need for an increased dosage of chelates Mn in relation to the free Mn ion. The dose must additionally be increased to recover for losses due to the renal excretion of the chelate during the time of the diagnostic test. In a variation on chelation, Quay (European patent application 308983) has described the use of complex coordination solutions of amino acid manganese. This application also discusses the addition of calcium ions to manganese amino acid solutions at levels up to 0.75 mol equivalents relative to manganese.

About 10 years ago, Schaefer et al. Investigated a mixture of Mn ++ and Ca ++ salts in the form of manganese gluconate and calcium gluconate in a mole ratio of one to one, administered intravenously to dogs, for imaging of cardiac perfusion. Even when the agent normally discriminated ischemic tissue, Schaefer also noticed acute cardiotoxicity similar to that seen with manganese chloride alone. The authors suggested that a possible way around the observed adverse cardiac effects might be to use a chelate rather than a manganese salt. There are no additional studies that use manganese gluconate and calcium gluconate or other salts or complexes that provide Mn ++ and Ca ++ in a higher proportion than the one to one that had subsequently appeared.

U.S. Patent Nos. 5,525,326 and 5,716,598 describe oral manganese formulas for imaging the gastrointestinal tract and for liver imaging; The latter takes advantage of the fact that the blood supply from the gastrointestinal tract passes through the liver, which removes the manganese from the bloodstream before returning the blood to the heart. Additional oral agents have been investigated including manganese polymers, molecular sieves impregnated with manganese, manganese clays, foods with high manganese content, such as cranberry juice. In general, cardiovascular safety is achieved for these agents with the cost of limiting the usefulness of the diagnosis of the agents to MR examination of the gastrointestinal tract and in some cases to the liver. The administration of manganese in nanoparticulate form has been described in U.S. Patent No. 5,401,492. Other particle approaches include the sequestration of Mn compounds into liposomes and metal clusters, such as manganese oxalate and manganese hydroxyapatite. The particulate agents are useful for a limited number of diagnostic applications, such as the gastrointestinal tract and organs, such as the liver and vessel, which are involved in the taking and sequestration of the particles carried by the blood.

Therefore it would be useful to have an agent for the formation of diagnostic images of tissues, systems and organs, particularly in humans, that increase the contrast in an MR image without giving rise to problems of toxicity. It would also be useful to have an agent that could be used for a variety of tissues, systems and organs that are physiologically remote from the gastrointestinal tract.

Synthesis of the Invention This need is satisfied, the limitations of the prior art are overcome and other benefits are realized in accordance with the principles of the present invention, which in one aspect relates to a diagnostic composition comprising a source of a diagnostically effective amount of ion. of Mn ++, from a Ca + + ion source and from a pharmaceutically acceptable carrier for parenteral administration. The Ca ++ ion is present in a molar ratio of 2: 1 to 40: 1 with respect to the Mn ++ ion. Preferred sources of Mn ++ are manganese salts, such as manganese acetate, chloride, gluconate, gluceptate, lactate and sulfate or mixtures thereof. Manganese gluconate or manganese gluceptate are the most preferred sources of Mn ++. The preferred sources of Ca ++ are calcium salts, such as calcium acetate, fluoride, gluconate, glucetate and lactate or mixtures thereof. Calcium gluconate and calcium gluceptate are the most preferred sources of Ca ++. The molar ratio of calcium to manganese is preferably from 4: 1 to 20: 1 and more preferably from 8: 1 to 10: 1. An embodiment of the composition aspect of the invention is a unit dosage form comprising a manganese salt containing from 5 milligrams to 200 milligrams of manganese, a calcium salt containing from 20 milligrams to 3 grams of calcium, and a suitable vehicle for parenteral injection.

In another aspect, the invention relates to a method for improving magnetic resonance imaging of a mammalian tissue, organ or system. The method comprises administering to the mammal a diagnostically effective amount of a source of Mn ++ ion together with from 2 to 200 molar equivalents of a Ca ++ ion source. The preferred sources of Mn ++ and Ca ++ are as before. Manganese and calcium sources can be administered intravenously at 1 μmol of Mn ++ per kilogram of body weight at 100 μmol of Mn ++ per kilogram of body weight and 2 μmol of Ca ++ per kilogram of body weight at 1400 μmol Ca + + per kilogram of body weight. The method is applicable to the visualization of the liver, kidney, pancreas, adrenal glands, heart, brain, salivary glands, gastrointestinal mucosa, uterus, tumors, biliary system, and circulatory system. The source of Mn ++ and the source of Ca ++ can be administered simultaneously or separately, with the administration of Ca preceding the administration of Mn for up to 30 minutes. The method allows one to have access to the metabolic activity of a tissue, distinguishing hyperactive or hypoactive from normal. The method also allows the evaluation of tissue vascularity, blood flow and tissue perfusion.

Brief Description of the Drawings Numerous advantages and features of the present invention will become readily apparent from the following detailed description of the preferred embodiments, the appended claims and the accompanying drawings wherein: Figure 1 is a graph of normalized systolic blood pressure in which the change in petroleum jelly (in percent) is plotted as a function of time (in minutes).

Figure 2 is also a graph of normalized systolic blood pressure in which the change in Vaseline (in percent) is plotted as a function of time (in minutes); Y Figure 3 is a graph of the MRI signal-to-noise ratio as a function of time (in minutes) after administration of the composition of the invention.

Detailed description of the invention Although the preferred embodiments of the invention are described below, it should be understood that the present disclosure should be considered as an example of the principles of this invention and that it is not intended to limit the invention to the illustrated embodiments.

We have found that, in the case of manganese salts, the formulation and / or administration with calcium (II) sources results in a contrast agent with significantly improved safety, provided that the ratio of calcium to manganese is at least 2: 1. Although in principle one can use any ratio higher than that of 2: 1, in practice however, the ratio of Ca ion to Mn ion used for any given dose of Mn ion will be limited by the toxicity Ca For example, assuming LD50 values after a rapid intravenous administration of 138 μmol / kilogram and 1819 μmol / kilogram for Mn ion and Ca ion respectively, for low doses of Mn, one can contemplate the use of very large molar excesses of ion Ca before the toxicity is seen. Therefore at a dose of Mn ion of 1 μmol / kilogram, a ratio of Ca ion to Mn ion as high as 1800: 1 can be considered without exceeding the LD50 of Ca ion more conservatively, a ratio of 180: 1 can be contemplated by approaching l / 10rh of LD50 of Ca ion. However, if the Mn ion dose is increased to 10 μmol / kg, the ratio of Ca to Mn will not exceed 18: 1 if the conservative approach of staying below 1 / 10rh of the LD50 of Ca ion is followed. In practice it is not necessary to approach maximum amounts of aggregate Ca to achieve a significant safety benefit. The particular ratio of Ca ion to Mn ion depends on the indication, of the dose required to achieve the efficacy and the desired safety margin (therapeutic index) for the composition of Mn / Ca. As a consequence of the invention, several advantages are realized. For example, improved utilization of paramagnetic properties of manganese is achieved without reduction in relaxivity or loss of agent through renal secretion associated with chelation. Manganese chelates demonstrate different pharmacokinetics and pharmacodynamics for manganese metal and chelator components, suggesting chelation. In such cases, one can see the toxicity mediated through both the metal and the chelator. In contrast, in the present invention, Mn ion levels that might otherwise not be considered as being physiologically tolerable can surely be achieved.

The compositions and methods of the invention are useful in the imaging of a variety of metabolically active organs, in particular the liver, kidney, pancreas, heart and adrenal glands. The improvement of manganese contrast is also useful for imaging the gastrointestinal mucosa, the uterus, the salivary glands, the brain, the biliary tree, and tumors. Although the applicants do not intend the invention to be restricted to a particular theory, it appears from the results that a macromolecular association between the Mn ion and one or more component (s) / protein (s) of the native body is responsible for the higher Relaxation noticed. It is also known that the Mn ion agglutinates human serum albumin, alpha, 2-macroglobulin, transferin and other blood proteins with a concomitant increase in Mn relaxivity due to the macromolecular association. This increase in relaxivity, coupled with the increased concentration of free manganese in the blood available for the macromolecular association through the use of the compositions of this invention, provides an increase in signal strength sufficient to still allow vascular imaging. at biologically tolerable doses of Mn ++. In fact, on an equimolar basis, the Mn ++ / Ca ++ compositions of the invention are approximately 20 times more effective than the current gold standard gadolinium or chelator to improve the intensity of the MR signal in the blood, without suffering from extravasation. of the contrast agent and of the fast signal to the noise degradation typical of extracellular fluid agents.

The compositions of the present invention can be used in place of the manganese chelates to detect and evaluate myocardial ischemia and reperfusion using the technique described for the chelates in WO 99/01162. After administration, the Mn is taken by mitochondria, and the Mn intake correlates with the dose through the clinically relevant dosing dose. The manganese intake also correlates with the blood flow and metabolic activity of the relevant tissues. As a result, an Mn tissue take allows the use of the compositions of the invention as viability markers (distinguishes viable from non-viable tissue). The vascular increase due to Mn binding to plasma proteins also helps to define blood flow and tissue perfusion (mL of blood flow per gram of tissue). Taken together the vascular and tissue phases of MR imaging with the Mn / Ca compositions of the invention allows the evaluation of the state of a variety of tissues, organs and organ systems.

Of the many alternative diagnostic uses for these compositions, the assessment of myocardial ischemia is particularly noticeable. The compositions of the invention provide diagnostic utility for: (1) characterization of viable tissue against non-viable including myocardium and tumors; (2) the assessment of heart attacks; (3) the characterization of the eschemic tissue or of the tissue at risk of including the myocardium (reversible damage against irreversible); (4) assessment of tissue perfusion; and (5) characterization of vascular lesions close to the areas of ischemia / infarction. With this knowledge, one can establish the need for and planning intervention revascularization procedures (PTCA) anastomosis); one can predict the possibility of success of intervention procedures (salvage tissue from the rest, one can evaluate and follow treated patients (including for reperfusion), and one can assess the effect of therapy, including the new therapy drugs The compositions of the invention are also useful for evaluating organ function before and after organ transplantation, for example, heart, kidney and liver, for tumor characterization (vascularity, metabolic activity, incremental patterns); for the assessment of liver disease and liver abnormalities, such as tumors (particularly to distinguish the heptocellular tumor against the non-hepatocellular tumor); and benign lesions, cirrhosis and angioma; and to characterize tissue vascularity (eg, angiogenesis, tumor, and vascular lesions). One can therefore establish the need for therapy (including but not limited to drugs, genetics, surgery, and revascularization) and evaluate the response to therapy over ischemic tissue, vascular lesions or tumor (including but not limited to surgery, revascularization, transplantation, drug therapy, gene therapy, new drug evaluation, radiation therapy, chemotherapy, tissue ablation therapy).

An improved therapeutic relationship is demonstrated through an increase of LD50 for the Mn ion from 138 μmol / kilogram to 220 μmol / kilogram by administering a 10: 1 composition of Ca + + / Mn ++. It appears that, although unnecessary, the toxicity modifiers can be used with the compositions of the invention. Therefore, one can contemplate, for example, liposomal sequestration, etc., where there is the potential for equilibrium between the bound and free metal ion in vivo. In addition, various administration rates may provide different advantages. For example, by effectively reducing the instantaneous Mn concentration, the lower administration of the agents described in this invention can further improve the therapeutic (diagnostic) rate of the agents or allow the safe administration of larger doses. Therefore, the compositions of the invention can be administered intravenously as a bolus or as an infusion over a period of time. Commonly, although not necessarily, the infusion will be over a period of one minute to 30 minutes. Larger doses improve the imaging of organs, such as the heart, that take manganese less efficiently than the liver does. Similarly, since Mn is effectively and efficiently cleared of blood, primarily by the liver, one can slow the rate of administration to increase the duration of the blood signal intensity improvement without increasing the total dose.

In a preferred embodiment, the gluconate Mn / gluconate Ca (l: 8) is administered parenterally over periods ranging from 10 seconds to 20 minutes. The dosage is related to the target organ of interest and may vary from 1 μmol / kilogram of body weight to 100 μmol / kg body weight of an Mn ++ ion source together with from 2 μmol / kilogram body weight at 14,000 μmol / kilogram body weight of a Ca ++ ion source. Preferably, the source of manganese is administered at 2 μmol / kg body weight at 30 μmol / kg body weight and the calcium source is administered at a body weight of 4 μmol / kg to 400 μmol / kg body weight . More preferably, the manganese source is administered at 3 μmol / kilogram of body weight and the calcium source is administered at 6 μmol / kilogram body weight at 200 μmol / kilogram body weight. The MRI is carried out, according to methods well known to those skilled in the art, from during or immediately after dosing to 24 hours of past dosing (vascular indications expected). The administration rate can be varied to further improve the cardiovascular tolerability of the contrast agent without an adverse effect on the quality of imaging, to increase the duration of the vascular phase of the agent, or to increase the dose without reducing the rate therapeutic agent in order to allow the formation of images of target organs that accumulate manganese less efficiently than the liver does.

The advantages of the invention include: (1) Less than the administered dose is renally excreted (essentially none against 15 or so percent for MnDPDP); this effectively reduces the dose of Mn required for an increase in optimal image formation; (2) The relaxation of solution of manganese salts is maintained; (3) The risk of toxicity associated with the presence of the chelator is avoided. For example, less bioactive metals such as Zn plasma through chelation and renal excision is avoided; (4) The increase of fast tissue is achieved in relation to Mn chelates, allowing the formation of earlier images; (5) additional indications, for example, imaging of tumors as well as vascular and cardiac imaging are possible; and (6) The cost of the composition is lower than that of most chelate compositions and most compositions employing rarer elements, such as gadolinium.

The invention relates to a diagnostic composition comprising a source of a diagnostically effective amount of an Mn ++ ion a Ca + + ion source and a pharmaceutically acceptable carrier for parenteral administration. The term "diagnostically effective" refers to an amount of Mn ion sufficient to increase the signal to signal ratio for MRI of the tissue in question. With the current instrumentation an increase of at least 5% is required to make it diagnostically effective. The term "Mn ++ or Ca ++ ion source" means any chemical species that can supply a measurable concentration of Mn ++ or Ca ++ ion in a normal or normal blood salt solution. Therefore, when the source is a simple salt, soluble at the concentration employed, such as manganese chloride, manganese gluconate or calcium gluconate, the molar concentration of the salt will be the molar concentration of the Mn ++ or Ca ++ ion. When the source is a salt or complex that is not completely dissociated from the concentration employed, the molar concentration of Mn ++ or Ca + + ions will be less than the molar concentration of the salt or complex, but its effective molarity can be calculated radially from the product solubility of the components by methods well known in the art, and the effective molarity of the metal ion can be determined experimentally with a specific ion electrode, as well as similar methods well known in the art. It should be noted that the concentrations of interest of the invention are those of the ions, not the concentrations of the source species, even though the two are the same for the simple soluble salts. A soluble salt, for the purpose of the present invention, refers to a salt which is essentially completely dissociated at the concentration that is being used.

Antioxidants such as ascorbate, and stabilizers, such as saccharate and calcium borate complexes can be added to the compositions of the invention, as can other substances known in the pharmaceutical art to be useful in parenteral formulas. Formulas for parental administration include sterile aqueous and non-aqueous injection solutions which may contain antioxidants, buffers, pH modifiers, bacteriostats and solutes which make the formula isotonic with the blood of the intended recipient. Formulas for parenteral administration also include aqueous and non-aqueous sterile suspensions, which may include suspending agents and thickening agents. The formulas can be presented in unit-dose or multi-dose containers, for example, sealed ampules and sealed containers, and can be stored in a dried and frozen (lyophilized) condition requiring only the addition of a sterile liquid carrier, for example, salt water, water for injection (WFI) or similar, immediately before use. Extemporaneous injection solutions and suspensions can be prepared from sterile powders and granules according to methods well known in the pharmaceutical art. Sterile solutions of the unit dose for injection are preferred. A preferred composition for an embodiment of the invention is a sterile solution of 25 mM manganese salt and 200 mM calcium salt in water for injection adjusted to a pH of 6.0 to 8.2 with sodium hydroxide and / or hydrochloric acid. About 4% to 8%, preferably 6%, by weight of calcium is provided by calcium saccharate and the rest by gluconate or calcium gluceptate.

The manganese source is preferably administered intravenously at 1 μmol / kilogram body weight at 100 μmol / kg body weight, more preferably at 2 μmol / kg body weight at 30 μmol / kg body weight. In many preferred additions, the source of manganese is administered intravenously at 3 μmol / kg of body weight at 15 μmol / kg of body weight. Similarly, the calcium source is preferably administered intravenously at 2 μmol / kilogram of body weight at 1400 μmol / kg body weight, more preferably at 4 μmol / kg body weight at 400 μmol / kg body weight. body. In many preferred embodiments, the calcium source is administered intravenously at 6 μmol / kg body weight at 200 μmol / kg body weight.

The terms "tissues", "organs" and "systems" are used in their normal sense. Thus, "tissue" refers to such biological materials as the gastrointestinal mucosa, the tumor and the like. The "organ" refers to such biological organs as the liver, the kidney, the pancreas, the adrenal glands, the heart, the brain, the salivary glands, and the uterus. The "system" refers to such biological systems as the biliary system, the gastrointestinal system and the cardiovascular or circulatory system.

According to the method of the invention, the source of Mn ++ and the Ca ++ source can be administered separately or as a single composition. As will be apparent to a skilled person, administration of a single parenteral dosage form will usually be the simplest, but the clinician can employ any means known in the art to achieve the desired ratio of Mn ++ to Ca ++ in the individual being treated. treated and to achieve the desired level of Mn ++ in the tissue, organ or system of interest. As will be seen below, the administration of the two ions within 30 minutes of each other will generally be successful but it is necessary to administer calcium first, when administration in sequence is employed.

Example 1; Preparation of Manqanesse Gluconate / Calcium Gluconate 1: X A manganese gluconate delivery solution of 27.9 mM was prepared as described below. The Mn gluconate used contained 5.5% by weight of water per test. Ca gluconate and water for injection were added to the supply to prepare solutions of Mn gluconate and Ca gluconate of fixed molar ratio, as noted below. Without the addition of the solubility enhancers, the concentration of the solutions prepared was limited to approximately 3% by weight of Ca gluconate. The supply solution of 27.9 mM Mn was prepared by adding 6.56 kilograms of Mn glucone. to 500 mL of water for injection. Because the supply solution was 27.9 mM, rather than 30.0 mM, the proportions shown below in the examples, in the tables and in the figures are not exactly 1: 1 etc., but they fall within 10% of the nominal proportion. The conclusions to be drawn are not essentially affected by this discrepancy and the absolute values can be recalculated by the reader if required.

Mn / Ca 1: 1 - To 50 mL of supply solution add 0.646 grams of Ca gluconate.

Mn / Ca 1: 2 - To 50 mL of supply solution add 1,292 grams of gluconate Ca.

Mn / Ca 1: 4 - To 50 mL of supply solution add 50 mL of water for the injection of 2,582 grams of Ca gluconate.

Mn / Ca 1: 6 - To 16.6 mL of supply solution add 33.3 mL of water for the injection of 1.29 grams of gluconate Ca.

Mn / Ca 1: 8 - To 12.5 mL of supply solution add 38.5 mL of water for the injection of 1.29 grams of gluconate Ca.

Alternatively, the desired Mn / Ca compositions can be made by adding the Mn gluconate to the calcium gluconate commercially available for injection, 10% (232 mM) which contains calcium saccharate as a solubility improving agent. For example, Mn / Ca 1: 8 [29 mM in Mn, 272 mM in Ca (II)] Ten milliliters of Calcium Gluconate Injection, 10%, is added to 136 mg of gluconate Mn.

Example 2 Calcium Gluconate Dose Range in Rabbits The contrast medium of the invention, with proportions of Ca to Mn molar from 1: 1 to 8: 1, prepared according to the description of Example 1, were studied for their effects on systolic blood pressure after intravenous administration in white rabbits from New Zealand. The dose of Mn ++ for all compositions was kept constant at around 30 μmol / kg, a dose previously known to induce a minimum of a 20% drop in systolic blood pressure. The changes in blood pressure were monitored through a femoral arterial catheter. Bases of blood pressure were re-established between doses. Table 1 summarizes the findings in a study in which the Ca / Mn ratio was doubled with each successive dose. Table 2 summarizes the findings in a second experiment focused on the most effective range of the proportions studied in the exploratory experiment. The data presented in Tables 1 and 2 together show that the combination of Ca and Mn in 2: 1 or higher ratio exhibits weaker cardiovascular side effects than Mn administered intravenously. The effect becomes related, as measured by both the maximum effect and the area under the curve, which reflects the extension and duration of the Mn response. Of particular note in these data is the result for the composition of 1: 1 Mn / Ca, which was substantially similar to that of Mn alone. This result is consistent with the findings of Schaefer and others, and stands in striking contrast to the results obtained from higher Ca / Mn ratios.

Table 1 'Average Normalized Data (n = 4) Table 2 Figure 1 illustrates the change in normalized systolic blood pressure over time for Mn and for various Mn / Ca compositions of different molar proportions, all administered intravenously in the rabbit at a dose of constant Mn ++ of about 30 μmol / kilogram . The 4: 1 mixture of calcium gluconate and magnesium gluconate depressed the systolic blood pressure only to about half of the extent seen with pure manganese gluconate; the 8: 1 mixture produced very little depression and a short period of systolic blood pressure; 6: 1 was better than 4: 1 and much better than pure manganese gluconate, but not as good as 8: 1 in this test.

Example 3. Conformatory Study of Rabbit Table 3 shows the data collected in a native rabbit for the dose Mn at around 30 μmol / kilogram and Ca at 240 μmol / kilogram, individually as a mixture, following the same protocol used in example 2.

Table 3 The data show that the cardiovascular effects of the Mn / Ca combination are not predictable from the effects of any agent alone. For example, using the area under the curve (AUC) as a measure of both the extent and duration of the change in systolic blood pressure, one could predict an area under load of -0.47 (by adding 0.43 to minus 0.90) for the composition of 1: 8 of Mn / Ca; in fact a significantly smaller area under the curve of -0.135 is seen experimentally. This is clearly visible through the examination of Figure 2, in which the changes in normalized systolic blood pressure for a mixture of 1: 8 mole ratio of Mn gluconate and Ca gluconate, a representative embodiment of this invention the noted effects following intravenous administration of only Mn or Ca are compared. As in the area comparison under the curve, the results for the composition Mn / Ca differ from those predictable through the simple adhesion of the individual Mn and Ca curves.

Example 4 Compositions of Mn / Ca in the rat A 1: 8 molar ratio Mn / Ca contrast agent of this invention, prepared according to the description of example 1, was studied for the effects on mean and systolic blood pressure relative to the gluconate control Mn in the Wistar rats. The agents were intravenously administered a dose of Mn of lOOμmol / kg. Changes in blood pressure were monitored through a femoral artery catheter. The results, shown in table 4, confirm that in the rat as well as in the mouse, the combination of Mn and Ca produces less depression of the hemodynamic parameters than does the Mn alone, thus allowing the safe administration of a Diagnostically effective dose.

Table 4 Example 5 Toxicity as measured by mean lethal dose The acute intravenous toxicity test in a Swiss Webster mouse was carried out on an Mn gluconate, Ca gluconate and a mixture of Mn gluconate and Ca gluconate, 1:10 on a molar basis. All agents were administered through tail vein injection for 30 seconds. For each agent the mean lethal dose was established through Bruce's up and down method (Fundamental and Applied Toxicology 5, 151-157 (1985)). Even when the animals were observed for a period of 24 hours, manganese-related deaths, when they occurred, were noted within minutes of the injection, presumably as the result of cardiovascular collapse. The data is shown in table 5.

Table 5 ** L (MLD / Mn or Ca MLD - 4 X 100 Example 6 Efficacy Studies, conei liver imaging The anterior and posterior axial contrast TI - weight the spin - echo and the gradient of echo images of the liver and were obtained in two rabbits. Posterior contrast images were obtained up to 30 minutes after injection of Mn / Ca (1: 8) (lOμmol / kg Mn). The images were obtained on a GE Sigma imager operating at 1.5 T. The signal intensity and the signal to noise ratios were determined through the region of interest (ROI) analysis. A good improvement of the liver was seen in all cases, as shown in table 6 given below. In addition, the data presented in Figure 3, collected through the ROI analysis of the liver as seen in the vascular images obtained in Example 7, showed that Mn / Ca (1: 8) provides a fairly rapid increase of the liver , with a measured signal to noise greater than 50% of maximum SNR by the injection end of one minute of contrast.

Table 6 Enhanced contrast liver imaging, Mn / Ca (1: 8) Example 7 Efficacy Studies, Rabbit Vascular Imaging Formation The blood famousness of Mn / Ca (1: 8) was examined by MRI in the rabbit. The contrast agent was administered intravenously in the vein of the ear for one minute at a dose of lOμmol / kg Mn. Blood samples (3mL each in heparinized tubes) were taken from the artery of the ear before and immediately after 2,10,20,30 and 60 minutes of contrast administration. The tubes were formed into images in the cross section TI-heavy rotated echo image formation (TR = 300, TE = 15, 4NEX, FOV = 8) on the GE Signa imager 1.5T. The signal intensity data, shown in table 7, demonstrate pharmacokinetics consistent with those found for Mn ++, when measured with a 56Mn tracer (Borg, D.C .; Cotzias, G.C.J. Clin Invest., 37: 1269).

Table 7 Vascular Imaging Increase with Mn / Ca (1: 8) Mn / Ca 1: 8 Evaluation of vascular increment over time Example 8 Exyivo Images Formation - Salt Water vs. Complete Blood The TI shortening seen with Mn / Ca (1: 8) in salt water was compared to that seen in the blood. The saline and blood tubes (3 milliliters each) were stuck with 100%, 50% and 10% intravascular Mn / Ca (1: 8) concentration of what would be expected if administered as a bolus. at a dose lOμmol / kg Mn. For comparison, the Gd DTPA was stuck in 3mL of salt water and blood at 100% of expected intravascular concentration followed by bolus dosing at 100μmol / kg, a dose typically used in the clinic for vascular imaging. The tubes were formed into cross-sectional images through TI-Heavy Rotate echo image formation (TR = 300, TE = 15, 4NET, FOV = 8) on a GE Signa imager 1.5 T. The results they are summarized in table 8. Table 8 It is interesting to note the significant increase in the signal strength seen for Mn / Ca (1: 8) in the blood in relation to salt water. For example, the signal intensity seen for Mn of 10% in the blood is essentially equivalent to that seen for Mn of 100% in salt water. This interaction indicates with the blood components. This increase in efficacy is not seen with Gd DPTA, which shows the increase of reduced signal in the blood in relation to salt water. The signal intensity seen for Mn / Ca (1: 8) at 50% of the expected blood concentration was greater than the view for Gd DTPA at 100% of its expected blood concentration, despite the fact that Gd DPTA was administered at 20 times the dose of Mn / Ca on a molar basis. This corresponds to at least a 40-fold efficacy advantage over Gd DPTA in the blood. An advantage of even greater molar efficacy is expected over an Mn administered as a chelate, since the chelated Mn has a lower relaxivity than does the Gd chelate. For example, MnDPDP, which does not bind to plasma proteins, has a Rl solution of 1.90 s "'mM" 1 at 40 ° C and 20MHz. The GdDTPA has an Rl of 3.84 s ^ mM "1 under similar conditions.

Claims (34)

R E I V I N D I C A C I O N S

1. A diagnostic composition comprising a source of a diagnostically effective amount of Mn ++ ion, a Ca ++ ion source and a pharmaceutically acceptable carrier for parenteral administration, wherein the Ca ++ ion is present in the molar ratio of from 2: 1 to 40: 1 with respect to the Mn ++ ion.

2. A diagnostic composition as claimed in clause 1 wherein said source of Mn ++ is a manganese salt chosen from manganese acetate, chloride, gluconate, gluceptate, lactate- and sulfate or a mixture thereof.

3. A diagnostic composition as claimed in clause 2 characterized in that said source of Mn ++ is manganese gluconate or manganese gluceptate.

4. A diagnostic composition as claimed in clause 1 characterized in that said source of Ca ++ is a calcium salt arising from calcium acetate, chloride, gluconate, gluceptate and lactate or a mixture thereof.

5. A diagnostic composition as claimed in clause 4 characterized in that the source of Ca ++ is calcium gluconate or calcium gluceptate.

6. A diagnostic composition as claimed in clause 1 characterized in that said molar ratio of calcium manganese is from 4: 1 to 20: 1.

7. A diagnostic composition as claimed in clause 6 characterized in that said molar ratio of calcium manganese is from 8: 1 to 10: 1.

8. A diagnostic composition as claimed in clause 1 characterized in that it additionally comprises an antioxidant.

9. A diagnostic composition as claimed in clause 8 characterized in that said antioxidant is ascorbate.

10. A diagnostic composition as claimed in clause 1 characterized in that it additionally comprises a stabilizer.

11. A diagnostic composition as claimed in clause 10 characterized in that said stabilizer is calcium saccharate borate complex.

12. A unit dosage form comprising a manganese salt containing from 5 milligrams to 200 milligrams of manganese, a calcium salt containing from 20 thousand grams to 3 grams of calcium, and a suitable vehicle for parenteral injection, wherein said calcium ion is present in the molar ratio of from 2: 1 to 40: 1 with respect to said manganese ion.

13. A unit dosage form as claimed in clause 12 characterized in that it comprises from 60 milligrams to 1.5 grams of manganese gluconate or manganese gluceptate and from 250 milligrams to 32 grams of gluconate or calcium gluceptate.

14. A unit dose form as claimed in clause 13 characterized in that it is in a water vehicle for injection set at a pH of 6.0 to 8.2 comprising calcium saccharide in an amount such that about 4% to about 8% of the calcium present in the dosage form is in the form of calcium saccharate.

15. A method for increasing a magnetic resonance image of a mammalian tissue, organ or system comprising administering to a mammal a diagnostically effective amount of an Mn ++ ion source together with from 2 to 200 molar equivalents of a Ca ++ ion source , wherein said calcium ion is present in the molar ratio of from 2: 1 to 40: 1 with respect to said manganese ion.

16. A method as claimed in clause 15 characterized in that said source of Mn ++ is a manganese salt chosen from manganese acetate, chloride, gluconate, gluceptate, lactate and sulfate or mixtures thereof and said source of Ca ++ is a salt of calcium chosen from calcium acetate, chloride, gluconate, gluceptate and lactate or a mixture thereof.

17. A method as claimed in clause 16 characterized in that said source of Mn ++ is manganese gluconate or manganese gluceptate and said Ca ++ source is calcium gluconate or calcium gluceptate.

18. A method as claimed in clause 15 characterized in that the molar ratio of calcium to manganese is from 4: 1 to 20: 1.

19. A method as claimed in clause 15 characterized in that the molar ratio of calcium to manganese is from 8: 1 to 10: 1.

20. A method for improving a magnetic resonance image of a mammalian tissue, organ or system comprising administering to the mammal from 1 μmol / kg of body weight to 100 μmol / kg body weight of a Mn ++ ion source together with from 2μmol / kg body weight at 1400μmol / kg body weight of a Ca ++ ion source, wherein said calcium ion is present in the molar ratio of from 2: 1 to 40: 1 with respect to said ion manganese.

21. A method as claimed in clause 20 characterized in that said source of manganese and said source of calcium are administered intravenously.

22. A method as claimed in clause 21 characterized in that said source of manganese is administered at 2μmol / kg by body weight 30μmol / kg by body weight and said source of calcium is administered at 4μmol / kg body weight at 400μmol / kg body weight.

23. A method as claimed in clause 22 characterized in that said source of manganese is administered at 3μmol / kg body weight at 15μmol / kg body weight and said source of calcium is administered at 6μmol / kg body weight 200μmol / kg body weight.

24. A method as claimed in clause 20 characterized in that the tissue, organ or system is chosen from liver, kidney, pancreas, adrenal glands, heart, brain, salivary glands, gastrointestinal mucosa, uterus, biliary system and tumor.

25. A method as claimed in clause 20 characterized in that tissue, the organ or system is the circulatory system.

26. A method as claimed in clause 20 characterized in that the source of Mn ++ is administered separately within 30 minutes after the administration of the Ca ++ source.

27. A method as claimed in clause 20 characterized in that the source of Mn ++ and the source of Ca ++ are administered simultaneously. to

28. A method for assessing the metabolic activity of a tissue comprising administering to a mammal a diagnostically effective amount of a source of Mn ++ ion together with from 2 to 200 molar equivalents of the Ca ++ ion source.

29. A method as claimed in the clause characterized in that the tissue is chosen from cardiac, renal, hepatic, pancreatic and adrenal tissue.

30. A method as claimed in clause 28 characterized in that said tissue is a tumor.

31. A method for evaluating tissue vascularity and tissue perfusion comprising administering to a mammal a diagnostically effective amount of an Mn ++ ion source together with from 2 to 200 molar equivalents of a Ca ++ ion source.

32. A method as claimed in clause 31 characterized in that said tissue is chosen from cardiac, renal, hepatic, pancreatic and adrenal tissue.

33. Such a method and as claimed in clause 31 characterized in that said tissue is a tumor.

34. A method for evaluating relative blood flow in the vasculature comprising administering to a mammal a diagnostically effective amount of a source of Mn ++ ion together with from 2 to 200 molar equivalents of a Ca ++ ion source. SUMMARY An MRI contrast medium is described for improved safety and efficacy. The composition includes a source of calcium ions and a source of magnesium ions in a ratio of from 2: 1 to 40: 1 in a vehicle suitable for parenteral administration. A method for improving the MRI signal in a mammalian tissue with the above composition is also provided.