Stabilized form of tetrofosmin and its use

Abstract

The present invention provides a stabilized form of Tetrofosmin, which is stable at room temperature as well as in contact with oxygen, and a non-radioactive kit containing a stabilized form of Tetrofosmin for the preparation of a radiopharmaceutical composition in the field of diagnostic radiopharmaceuticals, especially for myocardial perfusion studies in patients with coronary artery diseases and in oncology.

Claims

1. A stabilized form of Tetrofosmin represented by the general formula (I): ##STR00009## wherein R is H and x is a number between 1 and 2.

2. A stabilized form of Tetrofosmin according to claim 1 with the formula (II) ##STR00010## wherein x is 1 or 2.

3. A non-radioactive kit comprising at least one container, wherein one container contains a stabilized form of Tetrofosmin according to claim 1.

4. Non-radioactive kit according to claim 3, wherein the kit additionally comprises one or more of the following: (i) reductant, (ii) transfer ligand, (iii) preservative, (iv) agent for pH adjustment and (v) fillers.

5. Non-radioactive kit according to claim 3, wherein the container is a sealed and sterilized vial, syringe bottle or ampoule.

6. Non-radioactive kit according to claim 3, wherein the storage period of the non-radioactive kit at room temperature is at least 52 weeks.

7. Non-radioactive kit according to claim 4, wherein the reductant is a tin(II) salt.

8. Non-radioactive kit according to claim 4, wherein the transfer ligand is gluconate and/or sulphosalicylate.

9. Non-radioactive kit according to claim 4, wherein the agent for pH adjustment is a hydrogen carbonate and/or a phosphate salt.

10. A non-radioactive kit comprising at least one container, wherein one container contains a stabilized form of Tetrofosmin according to claim 2.

11. Non-radioactive kit according to claim 10, wherein the kit additionally comprises one or more of the following: (i) reductant, (ii) transfer ligand, (iii) preservative, (iv) agent for pH adjustment and (v) fillers.

12. Non-radioactive kit according to claim 10, wherein the container is a sealed and sterilized vial, syringe bottle or ampoule.

13. Non-radioactive kit according to claim 10, wherein the storage period of the non-radioactive kit at room temperature is at least 52 weeks.

14. Non-radioactive kit according to claim 11, wherein the reductant is a tin(II) salt.

15. Non-radioactive kit according to claim 11, wherein the transfer ligand is gluconate and/or sulphosalicylate.

16. Non-radioactive kit according to claim 11, wherein the agent for pH adjustment is a hydrogen carbonate and/or a phosphate salt.

Description

(1) The invention is illustrated by the following examples and figures without representing a restriction of the scope of protection defined in the claims.

(2) Caution: Due to the radiation of the nuclear isotope .sup.99mTc, appropriate protective measures must be taken, when handling .sup.99mTc compounds.

(3) FIG. 1: .sup.1H-NMR spectrum of Tetrofosmin x 2 HBF.sub.4.

(4) FIG. 2: TLC (thin layer chromatography) chromatogram of Tc-99m labeled Tetrofosmin using a non-radioactive kit that comprises one container and D-()-mannitol as filler.

(5) FIG. 3: TLC (thin layer chromatography) chromatogram of Tc-99m labeled Tetrofosmin using a non-radioactive kit that comprises two containers.

(6) FIG. 4: TLC (thin layer chromatography) chromatogram of Tc-99m labeled Tetrofosmin using a non-radioactive kit that comprises two containers and D-()-mannitol as filler.

EXAMPLE 1 SYNTHESIS OF THE TETRAFLUOROBORATE SALT OF TETROFOSMIN (TETROFOSMIN X 2 HBF4)

(7) Under N.sub.2 atmosphere an autoclave container is charged with 1 g (10.5 mmol) 1,2-bisphosphinoethane, 7 mL ethylvinylether (72.3 mmol) and 19.6 mg (0.12 mmol) azobisisobutyronitrile (AlBN). The reaction mixture is stirred in an oil bath at a temperature of 75 C. to 80 C. for 3 hours. The reaction mixture is then transferred in a two-neck flask. The ether is removed in a N.sub.2 stream. Subsequently 30 mL methylene chloride and 3.0 mL 8 N (mol/L) aqueous HBF.sub.4 solution are added under vigorous stirring. The organic phase is separated and the solvent is removed with a rotary evaporator. The remaining clear colorless oil is further dried in vacuum over night. Yield: 5.268 g (89.9%) white solid.

(8) As shown below, the stabilized Tetrofosmin x 2 HBF.sub.4 was analysed by .sup.1H, .sup.31P and .sup.19F NMR as well as by elemental analysis. The .sup.1H-NMR spectrum of Tetrofosmin x 2 HBF.sub.4 is depicted in FIG. 1.

(9) .sup.31P NMR (ppm, CDCl.sub.3): 16.92 (s); 16.93 (s)

(10) .sup.1H NMR (ppm, CDCl.sub.3): 1.17 (t, J=7.2 Hz, 12H, CH.sub.3), 2.76 (m, 12H, PCH.sub.2), 3.53 (m, 8H, OCH.sub.2), 3.77 (m, 8H, OCH.sub.2), 6.23 (d, J=503.6 Hz, 2H, PH)

(11) .sup.19F NMR (ppm, CDCl.sub.3): 150.2 (m)

(12) Elemental analysis: found (calc.): C, 39.0% (38.7%) H, 7.69% (7.59%)

(13) Tetrofosmin x 2 HBF.sub.4 was characterized to have purity over 99% (.sup.1H qNMR with dimethyl-malonic acid as internal standard), which makes it directly suitable for pharmaceutical use without the need of further purification.

EXAMPLE 2 STABILITY OF TETRAFLUOROBORATE SALT OF TETROFOSMIN

(14) Tetrofosmin x 2 HBF.sub.4 was stored at room temperature without protective gas (e. g. argon or nitrogen). In accordance to example 1, the .sup.1H and .sup.31P spectra were unchanged after 12 months.

(15) Moreover, Tetrofosmin x 2 HBF.sub.4 was heated in a normal atmosphere (with oxygen) at 50 C. for 6 hours. The .sup.1H and .sup.31P spectra showed no sign of degradation.

(16) An open vial of Tetrofosmin x 2 HBF.sub.4 was stored at room temperature, normal atmosphere and 50% humidity. After 72 h no change of weight was observed indicating the non-hygroscopic nature of the compound

EXAMPLE 3 ONE-VIAL KIT FORMULATION WITH TETRAFLUOROBORATE SALT OF TETROFOSMIN

(17) Tetrofosmin x 2 HBF.sub.4 is labeled by using a kit comprising the following composition:

(18) TABLE-US-00001 0.34 mg Tetrofosmin 2 HBF.sub.4 0.32 mg disodium sulfosalicylate 1.0 mg sodium D-gluconate 20 mg D-()-mannitol 0.03 mg SnCl.sub.2 2 H.sub.2O

(19) The kit is labeled in approx. 3.5 mL pertechnetate eluate with 300 MBq .sup.99mTc at pH 4-5 and room temperature. The mixture was gently swirled until complete dissolution of the powder. The solution was left for 15 min. Radio-TLC (tin layer chromatography) was performed silica acid (ITLC-SA) using a solvent mixture of acetone and dichloromethane (65:35 v/v). Each component (peak) is characterized by a relative front (R.sub.f) values, which is defined as the ratio of component position [mm] to the total distance traveled by the solvent front.

(20) TLC revealed a radiochemical yield of 95.7% of a .sup.99mTc-Tetrofosmin complex and a purity of 96.1% after 6 hours at room temperature. Only traces of pertechnetate are detected. The corresponding TLC (thin layer chromatography) chromatogram of the .sup.99mTc labeled Tetrofosmin is shown in FIG. 2.

EXAMPLE 4 TWO-VIAL KIT FORMULATION WITH TETRAFLUOROBORATE SALT OF TETROFOSMIN

(21) Tetrofosmin x 2 HBF.sub.4 is labeled by using a kit comprising to vials, which have the following composition:

(22) TABLE-US-00002 Vial I 0.34 mg Tetrofosmin 2 HBF.sub.4 0.32 mg disodium sulfosalicylate 1.0 mg Sodium D-gluconate 0.03 mg SnCl.sub.2 2 H.sub.2O Vial II 3.6 mg/mL Sodium hydrogen carbonate (saline)

(23) The kit was labeled by adding a 0.5 mL aliquot of vial II (1.8 mg NaHCO.sub.3) to vial I with a syringe. The kit was gently swirled until complete dissolution of the powder. Subsequently, 3.5 mL technetium-99m pertechnetate eluate (450 MBq) was added resulting in a pH of 7-8. After swirling again, the solution was left for 15 min. According to example 3, the radio-TLC was performed with ITLC-SN acetone: dichloromethane (65:35).

(24) TLC revealed a radiochemical yield of 93.2% of a .sup.99mTc-Tetrofosmin complex and a purity of 94.6% after 6 hours at room temperature. The corresponding TLC (thin layer chromatography) chromatogram of the .sup.99mTc labeled Tetrofosmin is shown in FIG. 3.

EXAMPLE 5 TWO-VIAL KIT FORMULATION WITH TETRAFLUOROBORATE SALT OF TETROFOSMIN AND FILLER

(25) In one especially preferred kit formulation the kit comprising to vials, which comprises the following composition:

(26) TABLE-US-00003 Vial I 0.34 mg Tetrofosmin 2 HBF.sub.4 0.32 mg disodium sulfosalicylate 1.0 mg Sodium D-gluconate 0.03 mg SnCl.sub.2 2 H.sub.2O 10.0 mg D-()-Mannitol Vial II 9.0 mg/mL Sodium hydrogen carbonate (saline)

(27) The kit was labeled by adding a 0.5 mL aliquot of vial II (4.5 mg NaHCO.sub.3) to vial I with a syringe. The kit was gently swirled until complete dissolution of the powder. Subsequently, 3.5 mL technetium-99m pertechnetate eluate (3 or 10.5 GBq, respectively) was added, resulting in a pH of approximately 8. After swirling again, the solution was left for 15 min. According to example 3, the radio-TLC was performed with ITLC-SN acetone: dichloromethane (65:35).

(28) TLC revealed radiochemical yields of the respective sample forming the .sup.99mTc-Tetrofosmin complex (x=2) after 15 min, 30 min and 6 hours at room temperature.

(29) TABLE-US-00004 Radioactivity Concentration 3 GBq/4 mL 10.5 GBq/4 mL t0 (15 min): 92.7% 93.2% t = 30 min 96.2% 96.2% t = 6 h 97.9% 97.8%

(30) An exemplary TLC (thin layer chromatography) chromatogram of the .sup.99mTc labeled Tetrofosmin is shown in FIG. 4.

EXAMPLE 6 COMPARISON OF TC-99M LABELED KIT FORMULATION CONTAINING TETRAFLUOROBORATE SALT OF TETROFOSMIN WITH A KIT FORMULATION CONTAINING NON-STABILIZED TETROFOSMIN

(31) Biodistribution was determined after technetium-99m labeling of different formulations containing stabilized or non-stabilized Tetrofosmin. Aliquots of each solution were intravenously injected into a tail vein of male Wistar rats. Biodistribution was determined 2 min, 60 min and 24 h p.i. (four animals at each time point). The following tables reveal that biodistribution pattern is essentially the same for all three tested formulations. Hence, the identical technetium-99m Tetrofosmin complex was formed equally either from stabilized or non-stabilized Tetrofosmin.

(32) Technetium-99m labeled kit formulation with non-stabilized TetrofosminMyoview

(33) The commercial available product Myoview (GE Healthcare) was used as a comparative example:

(34) TABLE-US-00005 2 min 60 min 24 h % ID/ % ID/ % ID/ organ organ stdev organ stdev organ stdev blood 0.14 0.131 0.017 0.006 0.001 0.001 brain 0.04 0.034 0.021 0.006 0.003 0.001 pancreas 0.30 0.273 0.301 0.118 0.018 0.010 spleen 0.31 0.269 0.157 0.052 0.007 0.001 kidney 7.58 1.011 2.972 2.053 0.074 0.007 fat 0.03 0.029 0.040 0.017 0.001 0.001 muscle 0.22 0.199 0.227 0.125 0.170 0.038 heart 1.64 0.101 1.044 0.514 0.139 0.022 lung 0.79 0.080 0.241 0.516 0.024 0.008 thyroid 0.09 0.078 0.065 0.018 0.030 0.004 liver 11.44 1.642 2.397 0.432 0.201 0.002 stomach 1.19 1.324 0.723 0.353 5.162 6.451 small 9.49 8.044 26.264 14.279 3.498 1.294 intest large 0.39 0.341 0.478 0.251 0.124 0.016 intest feces 0.42 0.500 0.681 0.402 1.045 0.469 urine 0.20 0.329 5.130 4.219 0.031 0.017

(35) Technetium-99m labeled kit formulation according example 4:

(36) TABLE-US-00006 2 min 60 min 24 h organ % ID/organ stdev % ID/organ stdev % ID/organ stdev blood 0.24 0.092 0.022 0.007 0.002 0.000 brain 0.05 0.005 0.020 0.003 0.004 0.000 pancreas 0.60 0.113 0.352 0.084 0.018 0.010 spleen 0.35 0.093 0.257 0.032 0.014 0.003 kidney 6.22 0.896 1.701 0.091 0.098 0.010 fat 0.07 0.038 0.041 0.011 0.006 0.005 muscle 0.24 0.006 0.230 0.105 0.191 0.105 heart 1.46 0.196 1.302 0.104 0.177 0.069 lung 0.51 0.075 0.285 0.582 0.026 0.013 thyroid 0.10 0.002 0.063 0.024 0.032 0.011 liver 17.81 1.416 6.117 1.727 0.435 0.055 stomach 1.16 0.259 0.983 0.316 0.104 0.025 small 13.75 1.929 28.599 0.310 2.750 0.516 intest large 0.64 0.082 0.512 0.055 0.179 0.034 intest feces 0.40 0.141 0.654 0.131 1.356 1.066 urine 0.61 0.798 11.314 1.863

(37) Technetium-99m labeled kit formulation according example 5:

(38) TABLE-US-00007 2 min 60 min 24 h organ % ID/organ stdev % ID/organ stdev % ID/organ stdev blood 0.14 0.060 0.028 0.018 0.002 0.000 brain 0.05 0.007 0.028 0.007 0.005 0.002 pancreas 0.47 0.105 0.413 0.016 0.025 0.010 spleen 0.40 0.057 0.301 0.083 0.022 0.004 kidney 3.05 1.827 5.816 0.978 0.098 0.003 fat 0.05 0.029 0.041 0.024 0.003 0.001 muscle 0.37 0.167 0.404 0.261 0.245 0.013 heart 1.74 0.146 1.634 0.315 0.279 0.047 lung 0.97 0.547 0.372 0.750 0.024 0.010 thyroid 0.10 0.014 0.089 0.019 0.025 0.011 liver 18.31 5.309 6.320 2.521 0.532 0.035 stomach 1.07 0.028 1.573 0.469 0.104 0.021 small 14.61 1.106 28.227 5.763 2.337 0.305 intest large 0.65 0.047 0.664 0.155 0.183 0.010 intest feces 0.53 0.254 0.414 0.120 0.801 0.631 urine 0.06 0.012 5.752 5.423