CA1068068A - Stannous-phosphate complex - Google Patents

Abstract

ABSTRACT OF THE DISCLOSURE
A kit is provided for forming a bone seeking complex with technetium-99m, which comprises a stannous-phosphate com-plex sealed in a sterile, non-pyrogenic container, the phosphate moiety of the complex being a ring phosphate having the formula PnO3n-n and having a molecular weight less than 300, the weight ration of stannous to phosphate moiety ranging from 10-3 to 0.5;
the kit can be employed for the generation of the complex with technitium 99m as required, this latter complex is character-ized by improved uptake into the bone marrow and thus represents an improved diagnostic aid.

Description

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he present invention relates to a stannous phosphate complex arld its preparation, which complex is useful in the manufacture of a bone seeking technetium 99m complex, the invention further relates to a kit containing such a stannous phosphate complex.
This application is a divisional application of Canadian Patent Application Ser. No. 179,092, filed August 17, 1973.
It has been known for some time that phosphates, including long chain linear polyphosphates, when introduced into the blood stream of mammals will selectively seek out and collect in the bone or skeletal structure. Pro. Soc. Exp, Biol. Med.
,~ Volume 100, pages 53-55 (1959), Journal of Labelled Compounds, , April-June 1970, Vol. VI, No. 2, pages 166-173; Journal of Nuclear Medicine, Vol. 11, No. 6, pages 380-381, 1970, Journal of Nuclear Medicine, Vol. 1, No. 1, Janaury 1960, pages 1-13.
In these cases a phosphorous atom or atoms of the phosphate are ; radioactive, i.e., 3 P.
It has alsc been known for some time that technetium-99m (99mTc) is a preferred radionuclide for radioactively scan-ning organs because of its short half life and because it radiates gamma rays which can be easily measured, compared, for example, to beta rays. See Radiology, Vol. 99, April 1971, pages 192-196.
It has also been known for some time to use divalent stannous tin (Sn+ ) in the form of stannous chloride, or di-; valent iron (Fe ) or reduced zirconium to bind radioactive 99mTc to carriers, such as chelating agents, red blood cells, albumln and other proteins, which selectively seek out certain organs of the body, in order to carry the 99mTc with them tosuch organs of the body where it is concentrated, whereby such :` .

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1C)~80~8 6, organ can be radioactively scanned or imaged for diagnostic or other purposes, e.g. radioactive treatment of a pathological condition. See Journal of Nuclear Medicine, Vol. 11, No. 12, 1970, page 761, Journal of Nuclear Medicine, Vol. 12, No. 1, 1971, pages 22-24, Journal of Nuclear Medicine, Vol. 13, No. 2, 1972, pages 180-181, Journal of Nuclear Medicine, Vol. 12, No. 5, May 1971, pages 204-211, Radiology, Vol. 102, January 1972, pages 185-196, Journal of Nuclear Medicine, Vol. 13, No. 1, 1972, pages 58-65.
Also it has been suggested to label a stannous compound with 99mTc for radioactively imaging bone marrow, Journal of Nuclear Medicine, Vol. 11, 1970, pages 365-366.
` It has also been known for some time that the stannous ion Sn++ forms soluble complexes with long chain polyphosphates, Journal Inorganic Nuc. Chem., Vol. 28, 1966, pages 493-502.
It has been suggested to employ the aforesaid 99mTc for radioactively scanning the skeletal bone structure of -~
mammals by complexing or binding it to tripolyphosphate carrier : :.
by use of the aforesaid stannous ion as a binding agent in order ~, for such phosphate to selectively carry the 99mTc to, and con- ~
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centrate it in, the skeletal bone structure upon in vivo ,~ intravenous administration for subsequent radioactive scanning :; . . .
; or imaging the skeletal structure. Radiology, Vol. 99, April 1971, pages 192-196. The use of 99mTc in this manner is ;
alleged to have certain advantages over the use of strontium, e.g. 85Sr, as the radioactive label which has been used for radioactive bone scanning in the past. These advantages are those which are inherent in 99mTc, i.e. short half life and lower energy gamma rays. However, the bone uptake (the percent of the total dosage which becomes concentrated in the skeletal structure within a certain time after in vivo intravenous ~ ;',' '

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1068~8 administration) of such 99mTc by the other organs of the body (the higher these ratios the better), i.e. radioactive contrast, are not nearly as high as with radioactive strontium.
It has been discovered that if, in the aforesaid 99mTc-stannous-phosphate complex the phosphate moiety comprises a cyclic or ring (meta) phosphate of formula PnO3n n, prefer-ably having a molecular weight of less than 300, rather than a polyphosphate, which is a linear straight or branched chain phosphate having the general formula PnO3n+l( n ), bone up-take of the 99mTc and the ratios of bone uptake to uptake of 99mTc by other organs, i.e. the liver, blood, kidneys and gastrointestinal system (G.I.) are substantially increased. `It has also been discovered that optimum results are achieved if at least 15 to 25% by weight, preferably at least 30 to 40%
(between 80 and 90 to 100% is more preferred) of such phosphate moiety is made up of such ring phosphate and if such phosphate moiety contains no more than about 15 to 20% or 25%, preferably ~-no more than 5 to 10% and more preferably no more than 5% ~
(less than 5% is the most preferred), by weight of such poly- ~-phosphate of molecular weight greater than that of pyrophos-` phate.
The term "phosphate moiety" as used herein refers to the phosphorus and oxygen atoms only of the phosphàte.
` The presence of polyphosphates of formula PnO3 +1( n+2) and molecular weight greater than 300, more particularly greater than that of pyrophosphate, seems to reduce bone take-up and the aforesaid ratios, as compared to complexes without such higher molecular weight polyphosphates. However, as aforesaid, --some of such higher molecular weight polyphosphates can be ~
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tolerated, preferably not more than about 15% to 20% or 25%, more preferably no more than 5% to 10% and still more preferably -'~

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not more than 5% (less than 5% i5 the most preferred), by weight of the total phosphate moiety.
The rest of the phosphate moiety of the complex, where the ring phosphate does not constitute 100% of the phos-phate moiety, is preferably ortho (P04 3) and pyrophosphate (phosphatemoietymolecular weight of less than 300) and more preferably pyrophosphate only.
A highly preferred ring phosphate is trimetaphosphate (P309 3-molecular weight of 237) of the formula:

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The complex is made from water soluble alkali metal (preferably sodium) or ammonium salt or acid salt of the ring phosphate, e.g. sodium trimetaphosphate.
~; Preferably the sodium trimetaphosphate is admixed with a stannous salt, e.g. SnC12 (the stannous salts of other acids which are pharmaceutically acceptable, i.e., safely intra- -venously administered, can be used) to form the stannous-tri-metaphosphate complex, the pH of which is adjusted to 3-8, preferably 5-8, by a pharmaceutically acceptable base, such '''f 20 as NaOH or Na2C03 or NaHC03, followed by admixing with the ` stannous-trimetaphosphate complex, an aqueous saline solution of radioactive sodium pertechnetate (99mTc) to form the 99mTc-stannous-trimetaphosphate complex at the time it is desired to intravenously administer the 99mTc complex. The stannous-tri-metaphosphate complex may be sealed in a sterile non-pyrogenic container~or vial as a solution or a dry lyophilized solid and shipped as a kit with the freshly generated sterile and non-~ .

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pyrogenic 99 ~ c being added aseptically at the situs just prior to use.
According to the invention there is provided a method of making stannous-phosphate complex comprising admixing a ~`, solution of a phosphate, the phosphate moiety of which comprises a ring phosphate of formula PnO3n n and molecular weight less than 300 with a stannous compound to form the complex.
The stannous compound is suitably a solid stannous compound, for example, a lyophilized stannous compound.
In an aspect of the invention there is provided a stannous-phosphate complex.
According to another aspect of the invention there is provided a kit for forming a bone seeking complex with technetium-99m, comprising a stannous-phosphate complex cealed in a sterile, non-pyrogenic container, the phosphate moiety of said complex comprising a ring phosphate having the formula PnO3n n and a molecular weight less than 300.
Suitably the weight ratio of stannous to phosphate moiety ranges from 10-3 to 0.5.
According to yet another aspect of the invention there is provided a method of making a kit for forming a bone seeking complex, comprising admixing with a stannous compound, a phosphate, the phosphate moiety of which comprises a ring phosphate having the formula PnO3n n and a molecular weight less than 300, to form a stannous-phosphate-complex, sterilizing the complex and sealing it in a sterile non-pyrogenic con-tainer; the weight ratio of stannous to phosphate moi-ety suitably ranges from 10 to 0.5.
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. The following compositions were prepared:

Sample No. Descri.ption 1 A commercial sodium polyphosphate sold by FMC
Corporation under the trade name FMC Glass H :;
(average chain length of 21 and average M.W. about 2100).
1-1 A first high molecular weight fraction of the FMC ; -Glass E~ of Sample 1 obtained by fractionating an : aqueous solution of Sample 1 with acetone accord- -:: ing to the technique described in Van Wazer, Phos-; phorous And Its Compounds, Interscience Publishers, .`
Inc. 1961 (pages 744-747) to precipitate out of the aqueous solution of the FMC Glass H, as an oil, the highest molecular weight fraction of poly-:~ phosphates (composition given in TABLE 2).

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Sample No. Description 1-2 A second acetone fraction of FMC Glass H achieved by adding more acetone to precipitate out of the remaining supernatant of 1-1, as 'an oil, the next !~ higher molecular weight polyphosphates tcomposi-tion given in TABLE 2). The acetone decreases the solubility of the polyphosphates in the water;
~- the higher the molecular weight of the polyphos-,- 10 phate the less soluble it is so that the highestmolecular weights are forced out of solution first.
1-3 A third acetone fraction of FMC Glass H containing the next higher molecular weight polyphosphates is precipitated out of the remaining supernatant solution of 1-2, as an oil, upon addition of fur-,. - . :
ther amounts of acetone (composition given in TABLE 2).
1-4 A fourth acetone fraction of FMC Glass H containing ` the next higher molecular weight polyphosphates (composition given in TABLE 2) is precipitated out of the supernatant solution of 1-3, as an oil, by adding more acetone.
1-5 A fifth acetone fraction of the FMC Glass H (eon-taining the next higher molecular weight polyphos-phates) (composition given in TABLE 2) is precipi-tated out of the remaining supernatant solution of 1-4, as an oil, by adding more acetone.
1-6 A sixth acetone fraction of FMC Glass H (composi-tion given in TABLE 2) is precipitated out of the remaining supernatant solution of 1~5, as a solid precipitate of the next higher molecular weight polyphosphates by adding more acetone.

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10~80~8 TABLE 1 (Cont'd) Sample No. Description 1-7 A seventh acetone fraction of FMC Glass H (Composi-tion given in TABLE 2) is precipitated out of the remaining supernatant solution of 1-6, as a solid precipitate of the next higher molecular weight polyphosphates by adding more acetone. ' '' 1-8 The residue fraction in the supernatant liquid left , `~
after removal of the 1-7 fraction (composition given in TABLE 2) is recovered by evaporating off '', the supernatant liquid. ' ' 2 An acetone end fraction of sample 1 after 90/O by - ~
weight had been previously fractionated off and 'i ' ,, leaving by removal of such end fraction 3% by weight ~'',' , in the supernatant (composition given in TABLE 2).
~'~, 4 A mixture of 86% sodium trimetaphosphate (Na3P30 3% sodium orthophosphate (Na3PO4)(molecular weight ,~ of phosphate moiety-95) and 10% sodium pyrophos- ",- ~
'', phate (Na4P207)(linear polyphosphate -molecular ~ , , 20 weight of phosphate moiety-174) obtained by acetone ", fractionation of sodium trimetaphosphate obtained , `, from Monsanto. Sodium trimetaphosphate as afore- ,'' , , said, is one of a plurality of cyclic phosphates , having the general formula PnO3n n . Sodium ortho- ~ , -phosphate is a phosphate monomer. Sodium pyro-`~ phosphate is a dipolyphosphate.
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'l 5 An acetone end fraction of a food grade polyphos- ' ,j phate sold by FMC under the name FMC FG (composi-.~ ,,, tion given in TABLE 2).
6 A commercial cyclic trimetaphosphate sold by ,, , Stauffer,Chemical, (composition given in TABLE 2).
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7 Sodium orthophosphate.

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10~80f~i8 TAsLE 1 (Cont'd) Sample No. Descri~tion 8 Sodium pyrophosphate.
9 Sodium tripolyphosphate.
Sodium tetrapolyphosphate - Na6P4013 - a poly-phosphate - phosphate moiety having a M.W. of 348.
It, together with the pyrophosphate and tripoly-phosphate, fall in the class of linear chain poly-phosphates having the general formula Pn03n+l (n+2)-An aqueous solution of each of the phosphate composition samples 1 through 10 (40 mg. phosphate/l ml. solution) were made with distilled water in which the dissolved oxygen content was reduced in a conventional manner by bubbling through such water gaseous nitrogen for a period of two hours. The water and phos-phates were mixed to form the solutions in a nitrogen atmosphere , and in a nitrogen flushed container. The reason for this is to -; reduce oxidation of the divalent Sn++ to be subsequently admixed with each solution sample. However, it is not essential (but highly preferred) to use nitrogen-treated water or a nitrogen at-mosphere or a nitrogen-flushed container. Other known pharmaceu-` tically acceptable conditions, which will inhibit oxidation of the Sn++ upon subsequent mixing thereof with the phosphate solution, can be used, including the use of conventional pharmaceutically acceptable reducing agents and anti-oxidants in the products used.
Each of these solutions, samples 1 through 10, in an amount :
equal to 100 ml, was mixed with 0.16g of solid SnC12 2H20 under a nitrogen atmosphere. The SnC12 2H20 was made by adding to 84.5mg.
of metallic tin, sufficient concentrated HCl with mixing until all the tin has dissolved followed by removing excess acid and water ;
by lyophilization (this operation also being carried out in a vacu-`- um or in a nitrogen atmosphere and in a nitrogen flushed container to prevent oxidation of stannous to stannic). Anti-oxidants, which can be administered intravenously, may also be used. A stannous . ~
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lOfi8068 (Sn+~)-phosphate complex or mixture of some kind was formed in each case, the phosphate moiety of each sample corresponding to the phosphate moieties of tne phosphates set forth in TABLE 2.
Thus, in the case of sample 1-7, sixty percent of the phosphate moiety was trimetaphosphate whereas in sample 1, 96.5% of the phosphate moiety constitutes long chain linear polyphosphates of ~ -

5 or more phosphorous atoms.
Sufficient aqueous solution of 3N NaOH (sodium carbonate or bicarbonate can also be used), in the case of all samples ex-cept 8, and 3N HCl, in the case of sample 8, is then added to each sample to give a pH of 6.0 to achieve a pH suitable for sub-sequent intravenous in vivo administration into the body of a mammal, in this case adult mice. The pH adjustement is preferably -done under a nitrogen atmosphere also.
After thorough mixing, the solutions are sterilized by passing them through a Millipore*biological filter of 0.22 micron ;
pore size under a nitrogen atmosphere. Thereafter milliliter portions of each of the sterile solutions are poured into indivi-dual sterile and non-pyrogenic storage glass vials under aseptic conditions and the vials are aseptically sealed so that the in-terior and contents of each sealed vial is sterile and non-pyro-genic and under a nitrogen atmosphere.
In the case of each sample, vials are lyophilized by con-ventional freeze drying equipment under aseptic conditions to ~-remove water. This provides a solid stannous-phosphate complex which aids in shipping and which is more stable than the complex in solution.
Each vial contains 1.35 mg. SnC12 and 40 mg. of the phos-phate.
The vials can be sealed and stored until needed sub-sequently to form the technetium-99m-stannous-phosphate complex at the use situs.
* trademark 10t;8~68 To prepare the technetiusn-99m complex, 3 to 7 (S) ml.
of fresh sodium pertechnetate, removed as a sterile non-pyrogenic eluate from a sterile NEN Tc Generator (any other source of pharmaceutically acceptable 99mTc can be used, including 99mTc generators manufactured by others than NEN), in a 0.0% saline solution is aseptically added to each vial containing the sterile and non-pyrogenic stannous-phosphate complex and the vial is swirled until a solution is obtained. In each case a technetium-99m-stannous-phosphate complex or mixture of some kind is formed 10 in aqueous solution (9 mg. per ml. solution when 5ml of per- ~`
technetate are used), the phosphate moiety of which corresponds to the phosphate moieties of the phosphate compounds of each -sample set forth in TABLE 2.
Aseptic techniques and sterile, non-pyrogenic ingredients and containers were used at all steps, such procedures being standard to those skilled in the art.
Each of the technetium-99m-stannous-phosphate complex-containing solutions is aseptically intravenously injected in vivo into a vein in the tail of adult mice (average weight 0.040 ;
kgs) in an amount equal to between 1 and 3 mCi and a volume of 0.12 ml (8 mg. of phosphate per ml solution in samples 1 through 10) .
Three hours after intravenous administration, some of the mice to which each sample was admnistered were sacrificed and -the various organs of their bodies (skeletal, liver, G.I., blood, kidneys) were counted by conventional gamma ray counting techni-ques to determine uptake of 99mTc by each organ and thereby deter-mine contrast of bone uptake as compared to uptake by other organs. -As aforesaid, it is not only important to have a high bone uptake (based on total technetium-99m dosage) but it is also important that the ratio of uptake by the bone to uptake by the other organs ~f be high.

, - 11 - , The results are set forth in TABLE 2 below, in which the uptakes (the bone uptake figures represent the average bone uptake for the skeletal system) are in terms of percent of the total tech~etium-99m activity injected (corrected for radioactive decay) which has collected in the various organs indicated three hours after in vivo intravenous injection, in which the ratio amounts are computed from the uptake amounts, in which "Percent Having Phosphate Moiety M.W. Less Than 300" refers to weight percent of the phosphate moiety based on the total phosphate moiety of the sample identified in the first horizontal column, in which the percents referred to under Phosphate Composition are weight per-cents of the whole phosphate moiety of the sample (as aforesaid, phosphate moiety as used herein is limited to that part of the compound or complex made up of phosphate phosphorus and oxygen atoms), in which Ortho Pl refers to the phosphate moiety of sodium orthophosphate, Pyro P2 refers to the phosphate moiety of sodium pyrophosphate. Tripoly P3 refers to the phosphate moiety of sodium tripolyphosphate, Tetrapoly P4 refers to the phosphate moiety of sodium tetrapolyphosphate, Trimeta R3 refers to the phosphate moiety of sodium trimetaphosphate, Tetrameta R4 refers to the phosphate moiety of sodium tetrametaphosphate, both trimeta and tetrametaphosphates falling within the class of cyclic or ring phosphates having the formula P303n n, in which "Pentapoly And Longer Linear Chains" refers to the phosphate moiety of sodium pentapolyphosphate and longer linear (linear as used herein in-cludes straight and branched linear phosphate chains) polyphospha-tes of formula Pn03n+l (n+2), in which "Average M.W." refers to the average molecular weight of the phosphate moiety of the sample and in which "Fraction In Raw Stock" with reference to samples 1-1, 1-2, 1-3, 1-4, 1-6, 1-7 and 1-8 refers to the normalized per-cent by weight of each of these samples in sample 1, which is the raw stock which is fractionated.

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10~80~8 Conventional gamma counting techniques for measuring technetium 99m take-up in the organs are conventional gamma ray-excitable scintillation counters for radioassaying multiple samples of the organs of the sacrificed mice.
Also, conventional scanning by radioactive imaging using a gamma ray-excited scintillation or gamma camera and a dual crystal rectilinear scanner was used in vivo. In vivo scintiphotos of the total body using the Anger camera were obtained as well as rectilinear total body scans.
The figures given in TABLE 2 are average figures achieved by the aforesaid conventional counting techniques, each sample having been intravenously administered to mice followed by radio-active counting.
Followin~ intravenous administration, the technetium 99m- ;~
stannous-ring phosphate complexes of the invention are rapidly -cleared from the blood by deposition in bone and excretion into urine. Thus, the technetium-99m-stannous-ring phosphate complexes are metabolizable. The deposition of the 99mTc-stannous-ring phos-phate complexes of the invention appears to be primarily a function 20 of the bone blood flow as well as being related to the efficiency -¢
of the bone in extracting the complex from the blood which perfuses the bones.
It was observed that the deposition of the Tc in the skeleton is bilaterally symmetrical with increased accumulations being present in the axial skeleton as compared to the appendicular skeleton. There is also increased deposition in the distal aspect of long bones.
Localized areas of abnormal accumulation of the radio-pharmaceutical may be seen in primary malignancies of the bone, metastatic malignancies of the bone, acute or chronic osteo-myelitis, arthritides, recent fractures, areas of ectopic calci-fication, Paget~s disease, regional migratory osteoporosis, areas ,.. ,. -, ~(~68068 of aseptic necrosis and in general any pathological situation `
involving bone in which there is increased osteogenic activity or localized increased osseous blood perfusion.
The acute toxici-ty level in mice (LD50/30) for Sample No~ 2 has been determined to be 150 mg/Kg body weight and for Sample No. 6 it is 800 mg/Kg and for Sample No. 8 it is 70 mg/~g.
Subacute toxicity studies in mice of Sample 2 have shown no signs of toxicity after 15 daily injections at dose levels as high as 63 mg/Kg body weight/day. A similar subacute study in dogs indi-cates no signs of toxicity at a dose level of 3.6 mg/Kg body weight/day.
It was found that samples 4 and 6 were only one-fourth as toxic to mice as sample 2 and one-eighth as toxic to mice as sam-ple 1 The complexes of the invention have been used success-fully as a skeletal imaging or scanning agent to visualize areas `~
of altered blood flow to the bone and altered osteogenic activity, including suspected bone lesions not shown on X-ray, bone survey performed as part of a work-up in patients with known or suspected malignancy, to follow the response of metastatic or primary bone lesions to radiation therapy, metabolic bone disease, to diagnose arthritis and osteomyelitis, and to diagnose and determine healing rate of bone fractures.
Seven clinical studies by seven doctors on humans using sample 2 and involving 91 patients showed no adverse reactions and were deemed to be highly successful and clinically useful for skeletal diagnostic purposes in the case of 90 patients.
The technetium-99m (99mTc) labeling reactions involved in preparing the 99m~c stannous-phosphate complexes of the invention depend on maintaining the tin in the reduced or stannous (Sn+2) state. Oxidants present in the pertechnetate supply may adversely affect quality.

1068~ 8 The radioactive dosage of the 99mTc complex of the inven-tion may vary from 1 to 25 mCi (millicuries) but preferably is from 10 to 15 mCi. The dosage in terms of the 99mTc complex may vary over a wide range, i.e. from 0.001 to 30 mg per kilogram body weight of mammal.
The concentration of ring phosphate moiety in the final solution is preferably between~l and 40, more preferably between 2 and 20 mgs per ml of solution. --An advantage of a complex containing a relatively large amount of ring phosphate is that the ring phosphate, in addition to providing excellent up-take and bone-to-other-organ ratios, has a low toxicity. Where the phosphate moiety contains phosphate other than ring phosphate it is advantageous for such other phos-phate to be pyrophosphate because of its high bone upta~e.
Scanning may be commenced as early as one hour after intravenous administration and may be as long after injection as clinically useful amounts of Tc remain in the organ~
Another manner of making the complex of the invention is to weigh 4 mg. of SnC12 2H20 and 100 mg of sodium trimetaphosphate into a flask (the flask is sterile and non-pyrogenic and is flush-ed with nitrogen before weighing and is kept under nitrogen during this step and for the next step). Add, under aseptic conditions, 12 ml of sterile, non-pyrogenic sodium pertechnetate in 0.9% saline solution. Shake the mixture until a solution is obtained followed by intravenous injection (preferably the pH of the mixture is asep-tically adjusted to pH 4-8 before intravenous injection).
Also, the sterile stannous chloride can first be aseptical-ly mixed with the sterile 99mTc saline solution to form a 99mTc-stannous complex, followed by adding the sterile sodium ring phos-phate under aseptic conditions to form the 99mTc-stannous-ring phosphate, adjusting the pH to 4-8, followed by intravenous injec-tion.

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It can be seen from TABLE 2 that the Tc-stannous-phosphate complexes, the phosphate moiety of which is cyclic (in the form of a ring) and has a molecular weight of less than 300, e.g. samples 1-7, 1-8, 2, 4, 5 and 6, provide surprising and markedly higher bone uptake of 99mTc and higher ratios of bone uptake to other organs, as compared to those complexes, the phos-phate moiety of which is in the form of linear long chains of molecular weight above that of pyrophosphate, e.g. samples, 1, ~ -1-1, 1-2, 1-3, 1-4, 1-6, 9 and 10.
In accordance with the invention, the ring phosphate moiety of the 99mTc-stannous-phosphate complex should be at least -~
15% or 20%, preferably at least 30% to 40%, more preferably more than 50% or 60% and most preferably 80% to 90% or more, by weight of the total phosphate moiety of the complex.
Trimetaphosphate is a highly preferred ring phosphate.
Although the stannous (Sn++) ion is by far preferred, the divalent ferrous (Fe++) ion in the form of ferrous ascorbate, ;
and reduced zirconium can also be used but without as good results.
All these metals can exist in a plurality of redox states.
The phosphate may be added to the solid SnC12 as an aque-ous solution, or it may be added to a solution of the SnC12 to form the Sn++-phosphate complex followed by adding the 99mTc solu-tion.
Very little Sn++ need be used to form the complex of the invention, e.g. less than 7 to 10% of the phosphate based on molecular weights.
The weight ratio of Sn++ ion to the ring phosphate moiety may vary over a wide range, i.e. from 10 3 to 0.50, preferably 0.01 to 0.4. The maximum ratio is dictated by the amount beyond solubility of the Sn++. The minimum amount required that amount necessary to bind a sufficient amount of 99mTc to the ring phos-phate to achieve good bone uptake and contrast. This can be ,~

determined by routine experiment.
The p~ of the stannous-phosphate complex should be be-tween 3 and 8.
The water used for making the complexes of the invention is distilled and is at an elevated temperature of 200F during removal of dissolved oxygen and reduction of oxidants by bubbling the nitrogen gas therethrough.
The maximum amount of 99mTc is that beyond the capacity of the Sn++-ring phosphate complex to bind the 99mTc. This can be determined by routine thin layer radiochromatography to deter-mine the percent of free or unbound 9mTc in the complex. The minimum amount is dictated by that amount below which there is an insufficient amount to give good scanning of bone uptake and contrast, which also can be determined by routine experiment.
Generally, the amount of 99mTc added to the Sn++-ring phosphate `~
complex should be sufficient to achieve the counting rate desired by the doctor or laboratory personnel for the volume to be inject-ed; ordinarily, as aforesaid, the activity dosage varies from 5 to 25 millicuries.
Although sodium ring (meta) phosphates are preferred, any alkali metal, such as potassium and lithium, or ammonium can be used as the cation so long as it i9 pharmaceutically acceptable so that it can be safely administered intravenously. Also, the acid pyrophosphates of such cations can be used.
Although in the examples given above saline water was used as the vehicle, any other vehicle which is pharmaceutically acceptable for intravenous administration can be used.
It is not intended that the invention be limited to any theory which may have been given above or to the specific examples set forth above but only by the claims appended hereto and their equivalents.

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Claims (44)

The embodiments of the invention in which an exclusive property or privilege is claimed are defined as follows:-

1. A method of making stannous-phosphate complex com-prising admixing a solution of a phosphate, the phosphate moiety of which comprises a ring phosphate of formula PnO3n-n and molecular weight less than 300 with a solid stannous compound to form the complex.

2. A method according to claim 1, wherein said phosphate moiety comprises at least 15% to 20% by weight of said ring phosphate.

3. A method according to claim 2, wherein said phosphate moiety contains no more than 20% by weight of linear poly-phosphates of formula PnO3n+1-(n+2) having a polyphosphate moiety of molecular weight greater than pyrophosphate.

4. A method according to claim 3, wherein said phosphate moiety is substantially free from said linear polyphosphates.

5. A method according to claim 3, wherein at least 30%
to 40% by weight of said phosphate moiety being said ring phosphate.

6. A method according to claim 3, wherein at least a major portion of any remaining phosphate moiety is selected from the group consisting of ortho and pyrophosphates and combinations thereof.

7. A method according to claim 2, 3 or 6, wherein n is equal to 3.

8. A method according to claim 3, wherein more than 50%
by weight of said phosphate moiety is said ring phosphate.

9. A method according to claim 3, wherein said phosphate moiety comprises pyrophosphate, a ring phosphate of formula PnO3n-n and orthophosphate.

10. A method according to claim 8 or 9, wherein n is 3.

11. A method according to claim 2, 3 or 6, wherein sub-stantially any remaining phosphate moiety comprises one or more phosphates having the formula PnO(3n+1)-(n+2) where n is 3 or less.

12. A method according to claim 2, 3 or 6, wherein sub-stantially any remaining phosphate moiety is one or more phos-phates of formula PnO3n+1-(n+2) of which not more than 20% by weight has an n value greater than 2.

13. A method according to claim 1, wherein said phosphate is admixed with said stannous compound under non-oxidizing conditions.

14. A method according to claim 13, wherein said solid stan-nous compound is a lyophilized solid, lyophilized under non-oxidizing conditions.

15. A complex, whenever prepared by the method of claim 1, 13 or 14, or by an obvious chemical equivalent.

16. A complex, whenever prepared by the method of claim 2, 3 or 4, or by an obvious chemical equivalent.

17. A complex, whenever prepared by the method of claim 5 or 6, or by an obvious chemical equivalent.

18. A complex, whenever prepared by the method of claim 8 or 9, or by an obvious chemical equivalent.

19. A method of making a kit for forming a bone seeking complex, comprising admixing with a stannous compound, a phosphate, the phosphate moiety of which is a ring phosphate having the formula PnO3n-n and a molecular weight less than 300, to form a stannous-phosphate complex, sterilizing said complex and sealing it in a sterile non-pyrogenic container, the weight ratio of stannous to phosphate moiety ranging from 10-3 to 0.5.

20. A method of making a kit for forming a bone seeking complex, comprising admixing with a stannous compound, a phosphate, at least 15% to 20% by weight of the phosphate moiety of which comprises a ring phosphate having the formula PnO3n-n and a molecular weight less than 300, to form a stannous phosphate complex, sterilizing said complex and sealing it in a sterile non-pyrogenic container.

21. A method according to claim 20, wherein said phosphate moiety contains no more than 25% by weight of linear poly-phosphates of formula PnO3n+1-(n+2) having a molecular weight greater than pyrophosphate.

22. A method according to claim 20, wherein at least 30 to 40% by weight of said phosphate moiety is said ring phosphate.

23. A method according to claim 20, wherein at least the major portion of any phosphate in said phosphate moiety other than said ring phosphate is selected from the group consisting of pyrophosphate, orthophosphate and combinations thereof.

24. A method according to claim 20, wherein substantially the remaining phosphate moiety being one or more phosphates of formula PnO3n+1-(n+2) of which not more than 20% by weight has an n value greater than 2.

25. A method according to claim 21, at least 30-40% by weight of said phosphate moiety being said ring phosphate and at least the major portion of any phosphate in said phosphate moiety other than said ring phosphate being selected from the group consisting of pyrophosphate and orthophosphate and combinations thereof, the weight ratio of stannous to phosphate moiety ranging from 10-3 to 0.5.

26. A method according to claim 20, wherein the n of said ring phosphate is 3.

27. A method according to claim 26, wherein said phosphate moiety consists of pyrophosphate, said ring phosphate of formula PnO3n n and orthophosphate.

28. A method according to claim 25, substantially the remaining phosphate moiety being one or more phosphates of formula PnO3n+1-(n+2) of which not more than 20% by weight has an n value greater than 2.

29. A method according to claim 19, 20, or 25, wherein said sterilized complex is lyophilized and sealed in said container in a lyophilized state.

30. A method according to claim 25, wherein said phosphate is admixed with lyophilized solid stannous compound.

31. A method according to claim 30, wherein said phosphate admixed with said lyophilized stannous compound is in the form of a solution and said complex is sterilized and sealed in a non-oxidizing atmosphere.

32. A method of making a stannous phosphate complex for forming a bone seeking complex, comprising admixing with a stannous compound, a phosphate, at least 15% to 20% by weight of the phosphate moiety of which comprises a ring phosphate having the formula PnO3n-n and a molecular weight less than 300 to form a stannous phosphate complex.

33. A method according to claim 32, wherein said phosphate moiety contains no more than 25% by weight of linear poly-phosphates of formula PnO3n+1-(n+2) having a molecular weight greater than pyrophosphate.

34. A method according to claim 33, wherein at least the major portion of phosphate in said phosphate moiety other than said ring phosphate is selected from the group consisting of pyrophosphate and orthophosphate and combinations thereof.

35. A method according to claim 34, wherein the n of said ring phosphate is 3 and the weight ratio of stannous to phos-phate moiety ranges from 10-3 to 0.5.

36. A kit made according to any one of claims 19, 20 and 21.

37. A kit made according to any one of claims 22, 23 and 24.

38. A kit made according to any one of claims 25, 26 and 27.

39. A kit made according to any one of claims 28, 30 and 31.

40. A stannous phosphate complex made according to the method of any one of claims 32, 33 and 34.

41. A stannous phosphate complex made according to the method of claim 35.

42. A method of making stannous-phosphate complex com-prising admixing a solution of a phosphate, the phosphate moiety of which comprises a ring phosphate of formula PnO3n-n and molecular weight less than 300 with a stannous compound to form the complex.

43. A stannous-phosphate complex, whenever prepared by the method of claim 42, or an obvious chemical equivalent.

44. A method according to claim 19 or 32, wherein said stannous compound is in solid form.