ORIGINAL ARTICLE 
Annals of Nuclear Medicine Vol. 13, No.2, 83-88, 1999 
Relationship between the biodistributions of radioactive metal nuclides in tumor tissue and the physicochemical properties of these metal ions 
Atsushi ANDO,* Itsuko ANDO,* Shigeru SANADA,* Tatsunosuke HIRAKI,* Tetsuo TAKEUCHl,** Kinichi HISADA** and Norihisa TONAMI** 
*School of Health Sciences and **School of Medicine, Faculty of Medicine, Kanazawa University 
This study was undertaken to elucidate the relationship between the biodistribution of radioactive metal nuclides in tumor tissue and its physicochemical properties. Potassium analogs (86Rb, 134Cs, 201Tl) were taken up into viable tumor tissue, although 22Na concentrated in necrotic tumor tissue 67Ga and 111In were more predominant in inflammatory tissue than in the viable and necrotic tumor tissue. 169Yb and 167Tm accumulated in viable tumor tissue and tissue containing viable and necrotic tumor tissue 67Ga, 111In, 169Yb and 167Tm were bound to the acid mucopolysaccharide with a mol. wt. of about 10,000 daltons in the tumor tissue 46Sc 51Cr, 95Zr 181Hf, 95Nb, 182Ta, and 103Ru were highly concentrated in inflammatory tissue and were bound to the acid mucopolysaccharides with a mol, wt. exceeding 40,000 daltons 65Zn and 103pd concentrated in viable tumor tissue and were bound to the protein in the tissue. The results suggest that the difference in intra-tumor distribution of these elements is caused by a difference in the binding substances (or status) of these elements in the tissues, and the binding substance is determined by physicochemical properties of the elements. We therefore conclude that the biodistribution of radioactive metal ions in tumor tissue is determined by its own physicochemical properties. 
Key words: biodistribution, macroautoradiogram, radioactive metal nuclides, physicochemical properties, tumor tissue 
INTRODUCTION 
SOME RADIOISOTOPES injected intravenously produce a higher concentration in tumor tissue than in normal tissues. 67Ga-citrate and 201Tl-chloride have been used for detection and location by scintiscanning. 
Since the discovery of tumor accumulation of radioactive bismuth by Kahn1 in 1930, a number of radioactive inorganic compounds have been carefully studied by many investigators. These reports have been summarized by Paterson et al.,2 by Larson 3 by Lopez-Majano and Alvarez-Cervera.4 
In 1968 we began to quantitatively determine the 
Received June 17, 1998, revision accepted December 24, 1998. 
For reprint contact: Atsushi Ando, Ph.D.. School of Health Sciences. Faculty of Medicine, Kanazawa University, 5-11-80, Kodatsuno, Kanazawa 920-0942, JAPAN. 
biodistribution of radioactive inorganic compounds in tumor bearing animals. We summarized the biodistribution of 54 elements and 66 compounds in tumor bearing animals as already reported.5,6 on the other hand, we have been investigating the biodistribution of radioactive metal nuclides in tumor tissue by macroautoradiography.7,8,13-17 
In this study, metal organic compounds such as 67Ga-citrate and 95Zr-oxalate, were classified as inorganic compounds because these metal nuclides were dissociated from these metal organic compounds in the body after intravenous (IV) or intraperitoneal (IP) administration. To date we have investigated the biodistributions of 17 elements in tumor tissue and examined the relationship between the accumulation in tumor tissue and the properties of these metal compounds. We summarized relationship among the biodistribution, the binding substance (or status) of these nuclides in tumor tissue, and the physicochemical properties of these metal ions. 
MATERIALS AND METHODS 
Animals and experimental tumors 
Male Donryu rats weighing 150-200 g underwent subcutaneous implantation of Yoshida sarcoma (1 x 108 cells/ 0.25 ml) in the right thigh. Five to six days later these animals were used in these experiments.7,17 Male ddY mice weighing 25-30 g were implanted with Ehrlich tumor (5 x 107 cells/0.1 ml) in the right thigh and used in these experiments 8-1O days later.7,8,13-17 
Radioactive compounds 
67Ga-citrate solution (1 ml containing about 7.4 MBq of carrier free 67Ga) was prepared by diluting 67Ga-citrate solution (Philips-Duphar Cyclotron and Isotopes Laboratory, Holland) with 0.08 M sodium citrate solution.7 
111In-citrate solution (1 ml containing about 7.4 MBq of carrier free 111In) was prepared from 111In-chloride (Nihon Mediphysics Co. Ltd., Japan) and 0.08 M sodium citrate solution.7 
169Yb-citrate solution (1 ml containing about 2 MBq and 0.1 ug of Yb) was prepared from 169Yb-chloride solution (The Radiochemical Centre, England) and 0.08 M sodium citrate solution.7 
167Tm-citrate solution ( 1 ml containing about 0.37-3.7 MBq of carrier free 167Tm) was prepared from 167Tm-chloride solution8 and 0.08 M sodium citrate solution.8 
201Tl(I)-chloride solution (1 ml containing about 7.4 MBq of carrier free 201Tl) was prepared from 201Tl-chloride (Daiichi Radioisotope Labs., Ltd., Japan) and 0.9% NaCl solution.7 
22Na-chloride solution (1 ml containing about 1.1 MBq of carrier free 22Na) was prepared by neutralizing 22Na-chloride in 5 M HCl solution (New England Nuclear, Corp., U.S.A.) with 0.1 M NaOH solution and diluted with 0.9% NaCl solution.7 86Rb-chloride ( 1 ml containing about 2.6 MBq and 40 ug of Rb) was prepared by neutralizing 86Rb-chloride in 0.5 M HCl solution (New England Nuclear, Corp., U.S.A.) with 0.1 M NaOH solution and diluted with O.9% NaCl solution.7 
134Cs-chloride (1 ml containing about 1.1 MBq and 0.3 ,ug of Cs) was prepared by neutralizing 134Cs-chloride in 0.5 M HCl solution (New England Nuclear. Corp., U.S.A.) with 0.1 M NaOH solution and diluted with 0.9% NaCl solution.7 
46Sc-citrate solution (1 ml containing about 2 MBq and 1.5 ug of Sc) was prepared by the following procedure: 0.08 M sodium citrate solution (10 ml) was added to 2.0 ml of 46Sc-chloride in 2 M HCl solution (New England Nuclear, Corp., U.S.A.) and adjusted to pH 8.0 with sodium bicarbonate. After that, it was heated at about 100deg.C for 10 min.13 
51Cr(III)-chloride solution (1 ml containing about 2.8 MBq and 1 ug of Cr) was prepared by diluting 51Cr(III)-chloride in 0.1 M HCl solution (Commissariat a L'Energie Atomique, France) with 0.9% NaCl solution and adjusting to pH 2.0 with 0.1 M HCl.13 
95Zr-oxalate solution (1 ml containing about 0.26-1.5 MBq of carrier free 95Zr) was prepared from 95Zr-oxalate in 0.7 M oxalic acid solution (Oak Ridge National Laboratory, U.S.A.) and 0.9% NaCl solution.15 
181Hf-chloride in 0.05 M HCI solution ( 1 ml containing about 0.74 MBq and 4 ug of Hf) was prepared from 181Hf in 2 M HCl solution (The Radiochemical Centre, En-gland) and 0.9% NaCl solution.15 
103Ru-chloride in 0.05 M HCl solution ( 1 ml containing 0.2-0.74 MBq and 5O-200 ug of Ru) was prepared from l03Ru-chloride in 4 M HCl solution (The Radiochemical Centre, England) and 0.9% NaCl solution.14 
95Nb-oxalate solution (1 ml containing about 0.45 MBq of carrier free 95Nb) was prepared by diluting 95Nb-oxalate in 0.5% oxalic acid solution (The Radiochemical Centre, England) with 0.15 M sodium oxalate solution.16 
182Ta-oxalate solution ( 1 ml containing about 0.4 MBq and about 2.3 ug of Ta) was prepared by diluting 182Ta-oxalate in 1.2% oxalic acid solution (The Radiochemical Centre, England) with 0.15 M sodium oxalate solution,16 
65Zn-chloride solution ( 1 ml containing about 0.4 MBq of carrier free 65Zn) was prepared from 65Zn-chloride in 0.5 M HCl solution (New England Nuclear, Corp., U.S.A.) and 0.9% NaCl solution.17 
l03pd-chloride solution (1 ml containing about 0.4 MBq of carrier free l03pd) was prepared from l03pd-chloride in 1 M HCl solution (The Radiochemical Centre, England) and 0.9% NaCl solution.17 
Methods7,8,13-17 
Each preparation was injected intravenously through the tail vein of rats and injected intraperitoneally into the mice. These animals were killed and tumor tissue was excised at three, twenty-four and forty-eight hours after the injection of the radioactive compounds. These tissues were embedded in two percent carboxymethyl cellulose sodium salts and frozen with dry ice-acetone (-70deg.C) immediately after excision. Following this, the frozen tissues were cut into thin serial sections (10 um) in a cryostat (-20deg.C). One of these sections was then placed on X-ray film which was developed after an exposure of several days, a second section was fixed in ethanol, and then stained with hematoxylin-eosin. 
RESULTS 
Biodistribution in tumor tissue 
Hematoxylin-eosin stained sections were divided into the following four categories: 1 ) viable tumor tissue, 2) tissue containing viable and necrotic tumor tissue, 3) necrotic tumor tissue, 4) connective tissue which contained inflammatory tissue. The relationship between morphological specimens of tumor tissue and accumulation of nuclides was determined from the autoradiogram and hematoxylin-eosin stained sections. 
22Na: Figure 1 shows a typical autoradiogram, hematoxylin-eosin staining tissue and its sketch illustration of Yoshida sarcoma treated with 22NaCl. The accumulation of 22Na was seen in the necrotic tumor tissue and was not seen in the viable tumor tissue or connective tissue.7 
86Rb, 134Cs and 201Tl: Figure 1 shows the result for 86Rb. A large amount of 86Rb accumulated in the viable tumor tissue and was not seen in the necrotic tumor tissue. A small amount of this nuclide accumulated in the connective tissue and tissue containing viable and necrotic tumor tissue. Biodistributions of 134Cs and 201Tl were very similar to that of 86Rb.7 
67Ga and 111In: Figure 1 shows the result for 67Ga. The accumulation of 67Ga was predominant in connective tissue (especially inflammatory tissue) rather than in the viable tumor tissue, and was hardly seen in necrotic tumor tissue. Distribution of 111In was very similar to that of 67Ga.7 
169Yb and l67Tm: Figure 1 shows the result for 169Yb. A large amount of this nuclide accumulated in viable tumor tissue and the tissue containing viable and necrotic tumor tissue. Fairly large amounts of this nuclide accumulated in the connective tissue which contained inflammatory tissue. But 169Yb was not seen in the necrotic tumor tissue regardless of the species of malignant tumor.7 In the case of 167Tm, very similar results were obtained.8 Biodistributions of 169Yb and 167Tm in tumor tissue differed only slightly from those of 67Ga and 111In. 
46Sc,5lCr,l03Ru,95Zr,181Hf,95Nb and 182Ta: Figure 1 shows the result for 46Sc. 46Sc accumulated much more avidly in connective tissue (especially inflammatory tissue) than in viable tumor tissue and necrotic tumor tissue, regardless of the species of malignant tumor.13 Figure 1 shows the results for 95Zr,95Nb and 103Ru. The accumulation of these nuclides in tumor tissue was very similar to that of 46Sc.14-16 In the case of 51Cr, 181Hf and 182Ta, very similar results were obtained.13,15,16 These elements became extremely concentrated in inflammatory tissue. 
65Zn and l03pd : Figure 1 shows the result for 65zn and l03pd. These nuclides accumulated in viable tumor tissue, and were not seen in the necrotic tumor tissue.17 
Classlfication of metal elements and accumulated area in tumor tissue 
Biodistributions of these radioactive nuclides in tumor tissue are summarized in Table 1 . 22Na concentrated in necrotic tumor tissue 86Rb, 134Cs 201Tl, 65Zn and l03pd accumulated in viable tumor tissue and were not seen in necrotic tumor tissue 67Ga and 111In were predominant in connective tissue (especially inflammatory tissue) rather than in viable tumor tissue and were hardly seen in necrotic tumor tissue. 
169Yb and 167Tm accumulated in viable tumor tissue and tissue containing viable and necrotic tumor tissue. The accumulations of these nuclides in tumor tissue differed only slightly from those 67Ga and 111In. 
The accumulations of 46Sc,51Cr,103Ru,95zr,181Hf,95Nb and 182Ta in connective tissue (especially inflammatory tissue) were much more dominant than those in viable and necrotic tumor tissue. 
Relation between metal ions and binding substances (or status) in tumor tissue 
We reported previously that monovalent cations (Na+, Rb+, Cs+, Tl+) existed as free ions in the tissue fluids, that Ga3+,In3+,Yb3+,Tm3+ were bound to the acid mucopolysaccharide with a molecular weight of about ten thousands daltons in the soft tissues, and that Cr3+, Sc3+ Ru3+, Zr4+, Hf4+, Nb5+ and Ta5+ were bound to the acid mucopolysaccharides with molecular weights exceeding forty thousand daltons in tissues such as tumor and inflammatory tissues.8,11-16 It is known that Zn2+ and Pd2+ are bound to the SH-radicals in the protein.18,19 The binding substances (or status) of these metal ions in the tissues are summarized in Table 2. 
DISCUSSION 
Relation between the physicochemical properties of metal ions and their chemical bonds 
It is well known that potassium ion (ionic radius 0.133 nm) exists in the intracellular fluid in tissue, sodium ion (0.079 nm) existing in extracellular fluid. We reported previously that monovalent cations (Rb+, Cs+, Tl+) whose ionic radii exceed 0.133 nanometers behave like the potassium ion.9 Potassium and potassium analogs (rubidium, cesium, thallium) are avidly taken up into viable tumor tissue whose Na+,K+-ATPase activity is increased, although sodium is more dominant in necrotic tumor tissue than in viable tumor tissue. 
Regarding the chemical bond of metal compounds, the following rules apply.10 A number of Lewis acids of diverse types are classified as hard acids and soft acids. Hard acids prefer to bind to hard bases. Soft acids prefer to bind to soft bases. Among the ions of the above metals, Ga3+,In3+,Yb3+,Tm3+,Cr3+,Sc3+,Zr4+,Hf4+,Nb5+ and Ga3+,In3+,Yb3+,Tm3+,Cr3+,Sc3+,Zr4+,Hf4+,Nb5+ and Ta5+ are hard acids, Pd2+ is a soft acid, and Ru3+, Zn2+ are borderline acids. 
As is shown in Table 2, Ga and In become trivalent hard acids by losing electrons from the s-shell and the p-shell. Yb and Tm become trivalent hard acids by losing electrons from the f-shell and the s-shell. Ga and In are typical elements. Yb and Tm are contained in the transition elements and called the f-block elements. Sc, Cr, Zr. Hf, Nb and Ta become hard acids of tri-, tetra- and penta-valence which are formed by losing electrons from the d-shell and the s-shell. Ru becomes a borderline acid of trivalence which is formed by losing electrons from the d-shell and the s-shell. These elements are contained in the transition elements and called the d-block elements. Zn becomes a borderline acid of bivalence which is formed by losing electrons from the s-shell. Pd becomes a soft bivalent acid which is formed by losing electrons from the d-shell. 
Among the radicals contained in body constituents, R-SO3-, R-PO32- and R-COO- are hard bases, and R-S- and R-SH are soft bases. It is reasonable to assume that hard acids such as Sc3+,Cr3+,Ga3+,In3+,Yb3+,Tm3+,Zr4+,Hf4+,Nb5+ and Ta5+ would bind to hard bases such as R-SO3-, R-P032- and R-COO- in tissue. It is also reasonable to assume that metallothioneins in tissue are composed of R-SH radicals in protein and soft acids such as Pd2+. In fact, it is known that metallothioneins in the body are composed of a specific protein and Pd2+. 
It is a well known fact that fibroblasts produce a large amount of acid mucopolysaccharides (the carbohydrate component of proteoglycan) as the inter-cellular substance in inflammatory tissue, and that these mucopolysaccharides have many carboxy radicals and sulphonic groups in their structure. There is convincing evidence that 67Ga3+ binds to the acid mucopolysaccharide.11 These ten elements, which become hard acids of tri-, tetra- and penta-valence, behave like 67Ga3+ in tissue. 
These results indicate that the difference in intra-tumor distribution of these elements is caused by a difference in the binding substances (or status) of these elements in tissue. 
Relation among the location of elements in the Thomsen-Bohr-type periodic table, binding substances (or status) and area of accumulation of these elements in tumor tissue 
Binding substances (or status) and the area of accumulation of metal ions are shown below each element in the Thomsen-Bohr type periodic table (Fig. 2). For transition elements of the first transition series, the 3d-orbital is filled with electrons from Sc to Ni. In the second transition series, the 4d-orbital is filled with electrons from Y to Rh. In the third transition series, the 5d-orbital is filled with electrons from Lu to Pt. Sc and Cr have incomplete 3d-shells, Zr, Nb and Ru have incomplete 4d-shells, and Hf and Ta have incomplete 5d-shells. These seven elements, which have incomplete d-shells, were bound to acid mucopolysaccharides whose molecular masses exceed 40.000 daltons. And trivalent hard acids (Ga3- +, In3+, Yb3 + and Tm3+), which have complete d-shells, were bound to the acid mucopolysaccharide wlth a molecular mass of about 10,000 daltons. 
These results clearly suggest that trivalent hard acids which have complete d-shells are essentially bound to the acid mucopolysaccharide with a molecular mass of about 10.000 daltons in soft tissues. In addition, most hard acids of tri-, tetra-, and pentavalent and some borderline acids (Ru3+, etc.) which have incomplete d-shells are essentially bound to the acid mucopolysaccharides whose mo-lecular weights exceed 40,000 daltons. It is certain that soft acids (except for some monovalent soft acids) and some borderline acids (Zn2+, etc.) are bound to the SH-radicals in protein and exist as components of metallo-thionein in tissues.18,19 Most of the 22Na 86Rb and 134Cs (alkaline metals), and 201Tl exists in the free form in tumor tissue. Therefore, the binding substance (or status) of metal ion in the tissue is determined by its physicochemical properties. 
In conclusion, the biodistribution of radioactive metal ions in tumor tissue is determined by its own physicochemical properties. 
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