LETTER TO THE EDITOR. Journal of Radioanalytical and Nuclear Chemistry, Vol. 240, No. 1 (1999) 349-352. Quality control and clinical application of two bone.
LETTER TO THE EDITOR Journal of Radioanalytical and Nuclear Chemistry, Vol. 240, No. 1 (1999) 349-352
Quality control and clinical application of two bone imaging 99mTc radiopharmaceuticals M. Ayranov,* A. Strezov,* A. Jordanova,* E. Piperkova,** S. Sergieva** * Institute for Nuclear Research and Nuclear Energy, Sofia, Bulgaria ** National Oncological Centre, Sofia, Bulgaria (Received March 30, 1998)
The radiochemical purity of MDP and HEDP has been determined by means of gel chromatography on Sephadex and thin layer chromatography on plastic foil silica gel. The comparison of the two radiopharmaceuticals shows equal level of complex formation with 99mTc. The biodistribution demonstrated that the application of HEDP allows earlier scanning than MDP. MDP and HEDP show equal effectivity during the clinical investigations. There is no significant difference in the radiochemical purity within six hours after the reconstitution of the freeze-dried kits. HEDP kit demonstrates shorter period of accumulation and equivalent complex formation levels, so it can be used in routine nuclear medicine diagnostics together with MDP kit.
Introduction The radionuclide 99mTc has nearly ideal physical characteristics for nuclear medicine imaging. Consequently, most of the major organ systems have been studied with the aid of this radionuclide. The MDP and HEDP labelled by 99mTc find intensive application for skeletal imaging in the routine clinical practice. The purpose of this work is to compare the performance of the two bone imaging agents as well as their quality control. In the present paper the results of gel chromatography (GC) on Sephadex G 25M, instant thin layer chromatography (ITLC) on silica gel 60 and clinical result comparison of 99mTc(Sn)-MDP and 99mTc(Sn)-HEDP are reported. The bench life on labeled radiopharmaceuticals was studied. The clinical surveys were carried out at the National Oncological Centre - Sofia.
Experimental Methylene diphosphonic acid (MDP, Sigma), Hydroxy ethylene diphosphonic acid (HEDP, Sigma), stannous fluoride (SnF 2, Sigma), stannous chloride dihydrate (SnC12.2H20, Merck) sodium hydroxide (NaOH, Fluka) and sodium p-aminobenzoate (Sigma) are commercially purchased. The 99Mo/99mTc Drygen-Biomedica or UltraTechneKow FM Mallinckrodt generators are used for the experiments. The studied kits are regularly produced from Radiochemistry and Radiation Chemistry Group at Institute for Nuclear Research and Nuclear Energy Sofia. The kits are freeze-dried, sterile and sealed under nitrogen atmosphere. The studied kits content are shown in Table 1.
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Table 1. Content of MDP and HEDP kits MDP, mg Methylene-diphosphonic acid Hydroxy-ethylene diphosphonic acid Stannous fluoride Stannous chloride dihydrate Sodium hydroxide Sodium para-aminobenzoate
6.20 0.35 2.20 2.00
HEDP, mg 15.00 1.00 6.55 -
The kits are labelled by addition 5 ml of 99mTcNaO4 (175-740 MBq 99mTc) with incubation period of twenty minutes. This routine procedure is close to the standard labelling one at the hospitals. For GC, prepackaged columns (10mm inner diameter and 300 mm length) are used. The column matrix (Sephadex G 25M, Sigma) is preequilibrated with the eluent (physiological saline 0.9% NaC1) before column packaging. The column void volume is determined using Blue Dextran (Fluka). On the top of the column 0.1 ml of 99mTc(Sn)-complex is introduced. After the void volume elution (13-14 ml) the column is sealed and prepared for scanning. 1,2 For ITLC, plastic foil silica gel 60 (Merck) is used, cut into 10xl00 mm strips. On the start of each strip, 50 ~1 of labeled radiopharmaceutical is dropped. As mobile phases methyl ethyl ketone and physiological saline solution are used, The strips are placed in 50 ml graduated cylinder, to which 4 ml of eluent is added and developed until the solvent front migrates to the top. 3 The Rf values for methyl ethyl ketone are: (99mTcO2+99mTc(Sn)-complex: Rj=0; 99mTcO4-: Rj=I) while Rf values for physiological saline are: (99mTcO2: Rf=0; 9 9 m T c O 4 - + 9 9 m T c ( S n ) - c o m p l e x : R f = l )
Elsevier Science B. V., Amsterdam Akadgmiai Kiad6, Budapest
M. AYRANOV et al.: QUALITY CONTROLAND CLINICAL APPLICATIONOF TWO BONE IMAGING99mTc RADIOPHARMACEUTICALS
Biodistribution 94.8%
0
2
4
6
8 10 12 14 16 18 20 22 24 26 28 30 Column length, cm
Fig. 1. Sephadexchromatogramprofilefor 99mTc(Sn)-MDP
Both columns and chromatogram's strips are scanned per centimeter by means of a NaI scintillation counter (SAM-2, Eberline) using appropriate shielding and collimators. Sephadex chromatogram profile for 99mTc(Sn)-MDP is presented in Fig. 1. The three peaks correspond to 99mTcO2-2.0%, 99mTcO4--3.2% and 99mTc(Sn)-MDP-94.8%, respectively.
The 99mTc(Sn)-HEDP biodistribution is conducted, on male rats Wistar strain according US Pharmacopoeia XXI on 56 animals divided in groups of seven. The HEDP kit is labelled with 5 ml NaO499mTc and activity 185-300 MBq. The animals were injected with 0.2 ml 0.05+0.1 MBq per gram weight in the tail vein. No anaesthetics are used to avoid influence on the biodistribution. The animals are sacrificed by decapitation in period of 5, 15, 30, 60, 120, 180 min, 6 and 24 hour after radiopharmaceutical's application. The biodistribution is estimated as a percentage of injected doze per gram organ weight. 4
Clinical survey
Whole body bone scintigraphy is performed during the postoperative follow up of 196 women with histologically confLrmed breast cancer, treated at the National Oncological Centre. The average age of the patients was 53.2 years, ranging between 29 and 72 years.
Fig. 2. Wholebody bone scintigraphywith 99rnTc(Sn)-MDPin a patient with breast carcinomawho had
bone scan evidenceof widespreadmetastaticinvolvementof the skeleton
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M. AYRANOVet al.: QUALITYCONTROLAND CLINICALAPPLICATIONOF TWO BONE IMAGING99mTc RADIOPHARMACEUTICALS
They passed surgical treatment, locoregional radiotherapy and/or chemotherapy according to the TNM stage of the disease. M D P and HEDP, labelled with 99mTc, activity 7.5-9.5 MBq/kg, is administered intravenously. The bone imaging is carried out on a rotating gamma camera ( D I A C A M II, SIEMENS) 2 - 4 hours post administration of the radiopharmaceuticals is presented at Fig. 2.
Results and discussion Radiochemical purity The GC and ITLC analyses showed that under standard labelling conditions the radiochemical purity was high. The chelate fractions for six years' analyses are shown in Table 2. The contents of the labeled complexes are above 94%. There was no deviation between M D P and HEDP chelate level. The lower complex formation level in case of GC analyses can be explained with dissociation of 99mTc(Sn)-complexes during the gel chromatography. 5,6 The stability o f 99mTc(Sn)-MDP and 99mTc(Sn)HEDP radiopharmaceuticals up to the sixth hour after reconstitution are presented in Table 3. The labeled kits maintain their stability within six hours after formulation.
injection. The observed distribution of 2.41% activity in bone, 0.55% in kidneys, 0.05% in liver and 0.86% in the blood, shows the high imaging quality of the radiopharmaceutical.
Clinical survey The tracer's uptake in percentages is compared using symmetrical regions o f interest (ROI) for quantitative evaluation o f the bone metastases. Solitary bone metastases are found in 21 of the patients, with metastases and multi focal metastatic lesions are visualized in 43 of the patients. Usually bone metastases appeared in cubic bones, lumbar and thoracic vertebras or/and hip joints. Quite often metastatic lesions involved the skull and the ribs.
Table 2. Radiochemical purity of MDP and HEDP kits on Sephadex G 25 M (GC) and silica gel plastic foil (ITLC)
Radiopharmaceuticals
MDP Chelate fraction, %
HEDP
GC ITLC
94.8+0.94 98.8+0.98
95.1+_0.95 98.9-2_0.98
Table 3. The 99mTc(Sn)-MDPand 99mTc(Sn)-HEDP radiopharmaceuticals stability
Biod~tnbu~on Kit The 99mTc(Sn)-HEDP biodistribution results are presented in Table 4. The 99mTc(Sn)-HEDP as bone imaging agent shows excellent biodistribution. Maximal inclusion in bones can be expected 30 minutes to 6 hours after intravenous application. Appropriate period for bone imaging is the second hour after intravenous
20 min
60 min
Chelate fraction obtained by GC MDP 94.8 94.8 HEDP 95.1 95.1 Chelate fraction obtained by ITLC MDP 98.8 98.8 HEDP 98.9 98.9
120 min
180 min
360 min
94.1 95.0
93.7 95.0
93.4 94.8
98~2 98.5
98.0 98.2
97.8 98.0
Table 4. Biodistribution of 99mTc(Sn)-HEDP, percent of injected doze per grain organ weight
Organ Blood Lung Heart Liver Spleen Brain Kidney Stomach Large intestine Small intestine Femur
5th min 6.24 0.88 0.43 0.23 0.17 0.04 2.15 0.32 0.27 0.23 0.60
15th min 4.75 0.49 0.15 0.11 0.23 0.02 1.21 0.28 0.18 0.18 1.83
30th min 3.53 0.35 0.12 0.08 0.07 0.01 1.04 0.12 0.09 0.11 2.02
60th min 2.04 0.13 0.06 0.05 0.04 0.01 0.49 0.08 0.06 0.06 2.28
120th min
180th min
6th hour
24th hour
0.86 0.09 0.04 0.05 0.03 0.003 0.55 0.15 0.04 0.13 2.41
0.52 0.04 0.02 0.05 0.03 0.004 0.64 0.03 0.02 0.02 2.24
0.33 0.04 0.01 0.04 0.02 0.004 0.61 0.06 0.03 0.02 2.50
0.19 0.02 0.006 0.02 0.02 0.002 0.42 0.01 0.02 0.02 1.56
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M. AYRANOVet al.: QUALITYCONTROLAND CLINICALAPPLICATIONOF TWO BONE IMAGING99mTc RAD1OPHARMACEUTICALS
Conclusions
Application of two radiochromatographic methods (GC and ITLC) and different mobile phases, allows precise estimation of 99mTc(Sn)-complex, 99mTcO4- and 99mTcO2 fractions. The labeled kits are stable and the reducing of chelate fraction within six hours after reconstitution is negligible. The 99mTc(Sn)-HEDP biodistribution confirms that bone images can be obtained two hours after intravenous injection of the kik The biodistribution of MDP and HEDP in the normal tissues and their uptake in the metastatic bone lesions is found to be of equal high quality. MDP kit application
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allows visualization 3-4 hours after intravenous injection, while investigation with HEDP can be achieved at the second hour post injection. References 1. Y. M. I-IY1GEN, T. G. TJI, W. J. GELSEMA, DE L1GNY, Intern. J. Appl. Radiation Isotopes, 39 (1988) 25. 2. T. A. MINEV, M. I. MIKHAJLOV, S. A. MAIKAROV, Z. D. STOINOV, 1. A. PENEV, Roentgenol. Radiol., (Sofia) 7, (1988). 3. A. M. Z1MMER, D. G. PAVEL, J. Nucl. Med., 18 (1977) 1230. 4. United States Pharmacopoeia XXI, 1985, p. 1012. 5. J. L. VUCINA, J. Radioanal. Nucl. Chem., 199 (1995) 347. 6. P. RICHARDS, J. STEIGMAN, Applied Chemistry of Technetium, 1976, p. 23.
