Synthesis and ligand binding of nortropane derivatives: N-substituted 2beta-carbomethoxy-3beta-(4'-iodophenyl)nortropane and N-(3-iodoprop-(2E)-enyl)-2beta-carbomethoxy-3beta-(3',4'-disubstituted phenyl)nortropane. New high-affinity and selective compounds for the dopamine transporter.

Two novel series of iodinated N-substituted analogs of 2beta-carbomethoxy-3beta-(4'-iodophenyl)tropane (beta-CIT) and N-(3-iodoprop-(2E)-enyl)-2beta-carbomethoxy-3beta-(3',4'-dis ubstituted phenyl)nortropane were synthesized. They were evaluated for their inhibitory properties on dopamine (DA(T)), serotonin (5-HT(T)), and norepinephrine (NE(T)) transporters in rat brain homogenates using [3H]GBR-12935, [3H]paroxetine, and [3H]nisoxetine as specific ligands. All new N-substituted analogs of beta-CIT exhibited higher DAT selectivity over both 5-HT(T) and NE(T) than beta-CIT. Moreover compounds with the N-substituents propynyl (6), crotyl (4), 2-bromoprop-(2E)-enyl (5), and 3-iodoprop-(2E)-enyl (3d) showed similar to higher DA(T) affinities than beta-CIT (respectively 14, 15, 30, and 30 nM vs 27 nM). Compound 3d was found to be the most selective DA(T) agent of this series (5-HTT/DA(T) = 32.0 vs 0.1 for beta-CIT). The N-(3-iodoprop-(2E)-enyl) chain linked to the tropane nitrogen was therefore maintained on the tropane structure, and phenyl substitution was carried out in order to improve DA(T) affinity. K(i) values of N-(3-iodoprop-(2E)-enyl)-2beta-carbomethoxy-3beta-(3',4'-dis ubstituted phenyl)nortropanes revealed that phenyl, 4'-isopropyl, and 4'-n-propyl derivatives weakly inhibited specific binding to DA(T), whereas phenyl substitution with 4'-methyl (3c), 3',4'-dichloro (3b), and 4'-iodo (3d) yielded high-DA(T) reuptake agents with increased DA(T) selectivity compared to beta-CIT. These results demonstrate that the combination of a nitrogen and a phenyl substitution yields compounds with high affinity and selectivity for the dopamine transporter which are usable as SPECT markers for DA neurons.

Br/Rb ratio obtained by XRF analysis of kidneys of normal and tumor-bearing mice treated with cis-DDP.

The Br/Rb ratio in kidneys of normal and tumor-bearing mice was studied and a significant difference was found between the two groups. The mice were treated with cis-DDP with and without selenite, with selenite alone, and compared with controls. The animals were sacrificed 7d, 14d, and 21-28d after treatment. The kidneys were removed and subjected to Br and Rb determination by a novel X-ray fluorescence (XRF) method. The results are presented in terms of estimated net count rates under the BrK alpha and RbK alpha peaks. No significant differences were found in Br and Rb levels or in the Br/Rb ratio in kidneys of treated and untreated mice belonging to the same group-normals or lung carcinoma inoculated. However, there was a significant difference in the Br/Rb ratio between the two tested groups (normals and inoculated). Preliminary results on the Br/Rb ratio obtained from normal and malignant tissues of colorectal cancer (CRC) patients and from mice inoculated with melanoma, show the same trend as those from the group of mice with lung carcinoma. In all three groups tested the Br/Rb ratio was 2-4 times higher in normal than in malignant tissue.

The distribution of BrK alpha/RbK alpha peak intensity ratio in human whole blood samples. Comparison between normal and colorectal cancer cases.

A preliminary study was conducted to compare the BrK alpha/RbK alpha peak intensity ratio in colorectal cancer (CRC) patients and healthy subjects (controls) in Israel. Blood samples were obtained from 61 CRC patients and 124 controls. The controls represent a normal population from different areas in Israel. Three hundred microliters of wet whole blood samples were analyzed by the EDXRF method. A weighted mean of 2.45 +/- 0.38 Br/Rb ratio was obtained for CRC patients, as compared with 3.28 +/- 0.40 Br/Rb ratio for controls. The mean value for controls represents only 112 subjects, since 12 individuals of the control group suffered from some other diseases and therefore were not included for the mean value calculation in this group. The results indicate that the Br/Rb ratio in CRC patients is significantly lower (p < 0.05) than in controls. No significant difference was found in Br/Rb ratio correlated to age and sex. It was not possible to draw final conclusion concerning the relationship between the Br/Rb ratio and the malignancy stage, since there was a relatively small number of cases in each stage under investigation.

An XRF study of trace elements accumulation in kidneys of tumor-bearing mice after treatment with cis-DDP with and without selenite and selenocistamine.

The effect of cis-DDP treatment with and without selenite and selenocistamine was studied on kidneys of tumor-bearing mice. The amounts of cis-DDP, selenite, and selenocistamine injected were chosen so as to be compatible with the treatment of humans. The animals were sacrificed at 7, 14, and 28 d after treatment. The kidneys were removed and subjected to trace element analysis by a novel X-ray fluorescence (XRF) method and for pathological assessment. The results show that following treatment with cis-DDP, K, Fe, Cu, and Zn reach a maximum level after 7 d; K, Fe, and Cu levels were significantly reduced by the addition of selenite. The level of Zn was reduced only in groups treated with selenite whereas that of K and Cu was reduced also in groups treated with selenocistamine with and without cis-DDP. The greatest increase in Pt and Se levels was reached 1 wk after injection with cis-DDP, with and without selenocompounds, and in the case of Pt was partly reduced by addition of selenite. Se returned to control values 2 wk after injection, although Pt was still high in all groups 2 and 4 wk after injection. The results corroborate the findings of our previous studies. The effect of selenocistamine in cis-DDP treated mice was partly insufficient. The pathological examination of the kidneys did not show any differences in the effect of various additives during the study.

Assessment of the purity of d,l HM-PAO from diastereomeric mixtures using NMR techniques.

We have shown that 1H NMR can serve as a suitable analytical tool for the assessment of relative amounts of d,l and meso HM-PAO isomers in a diastereomeric mixture and for the determination of the purity of either one of the two diastereomers alone. Therefore, we believe that this method will be useful for quality control in both academic and government regulatory laboratories as well as for radiopharmaceutical manufacturers. In addition, this analytical tool can provide rapid screening of possible suitable stereospecific reducing agents for the preferential formation of the d,l HM-PAO diastereomer without the necessity of completely working up the reduction reaction mixture.

Labeling of sarcoma associated monoclonal antibody with 111In, 67Ga and 125I.

Labeling of human sarcoma-associated murine monoclonal antibody (MAb) 23H7 with 67Ga and 111In by the bifunctional ligand method is reported. 67Ga was chelated to the MAb via desferrioxamine B and 111In via the cyclic anhydride of DTPA. Higher specific activity was obtained with 67Ga (4-5 microCi/micrograms) as compared with 111In (2 microCi/micrograms). The binding capacity of the MAb was confirmed by repeated indirect immuno-fluorescence assays performed before and after labeling. A fast blood clearance was observed: 33% recovered dose (R.D.) blood level 3 h post-injection as compared with 56% after injection of control polyclonal IgG. Preliminary results on chemically induced sarcoma bearing mice showed a relatively high tumor uptake of the labeled antibody.

Human sarcoma-associated murine monoclonal antibody labeled with indium-111, gallium-67, and iodine-125.

Monoclonal antibody (MAb) 23H7 is a hybridoma-derived IgG that is generated following fusion of mouse myeloma cell line P3U1 and spleen cells from BALB/c mice hyperimmunized with a human fibrosarcoma. It detects a mesenchymal antigen of 23KD expressed on human sarcoma tissues and other neoplasms, including myeloid leukemias, but it rarely binds to normal tissues. The MAb 23H7 was labeled with 67Ga and 111In using the bifunctional ligand method. The 67Ga was chelated to the MAb via desferrioxamine B, while 111In was chelated via the cyclic anhydride of diethylenetriamine pentaacetic acid. Higher specific activity was obtained with 67Ga than with 111In (4.5 and 2 muCi/microgram, respectively); both gave stable complexes. When 23H7 was labeled with 125I, considerable breakdown was observed. This, together with the physical shortcomings of this isotope, emphasizes the advantages of labeling with 111In and 67Ga. The rapid blood clearance of the labeled sarcoma-associated MAb may be beneficial for early tumor uptake and for imaging shortly after injection.

Radioprotection of sheep erythrocytes by polyvinylpyrrolidone.

The protective effect of polyvinylpyrrolidone of mol. wt 10,000 and 40,000, on sheep erythrocytes irradiated up to 13 kGy, and stored for one month at 4 degrees C was studied. Addition of these compounds at a level of 10-15% reduced hemolysis from 90% in irradiated erythrocyte solution to approximately 10%. The high efficiency of this protective agent is discussed.