Richard T. Dean

Coupling agents for joining radionuclide metal ions with biologically useful proteins

This invention relates to bifunctional coupling agents useful in forming conjugates with biologically useful molecules, such as antibodies. These conjugates can be complexed with radionuclide metal ions to provide materials useful for in vivo diagnostic and therapeutic applications.

External Imaging of Atherosclerosis in Rabbits Using an 123I‐Labeled Synthetic Peptide Fragment

The oligopeptide fragment of apolipoprotein B, SP‐4, has demonstrated pronounced uptake in the healing edges of balloon‐injured rabbit aortic endothelium. To assess 123I‐labeled SP‐4 for identification of atherosclerotic plaques by gamma camera imaging, 14 Watanabe heritable hyperlipidemic (WHHL) and 5 normal rabbits were imaged 5 minutes and 12 and 24 hours after intravenous injection of123I‐SP‐4. In addition, two WHHL and two normal rabbits were injected with 125I‐SP‐4 for autoradiography. Twelve of the 14 WHHL, but none of the normal, rabbits had visually apparent focal radioiodine accumulation in the region of the aorta. Focus‐to‐lung and focus‐to‐heart count ratios were 2.4 ± 1.3 and 1.0 ± 0.4, respectively. Five of the visually positive WHHL rabbits were reimaged 4 and 8 weeks later with 123I‐NaI and 123I‐SP‐2 (an apo E peptide), respectively, as negative controls. Perceptible, but faint, aortic localization of 123I‐NaI and of 123I‐SP‐2 was seen in only one animal each. The distributions of atherosclerotic lesions on photographs of the opened WHHL aortas and of film blackening on 125I‐SP‐4 autoradiograms were identical. In contrast, the two normal rabbit aortas did not exhibit plaques on photographs or film blackening on autoradiograms. Thus, in an animal model closely simulating human atherosclerotic disease, SP‐4 localizes specifically in aortic atherosclerotic lesions.

Synthesis and characterization of technetium complexes with phosphoruscontaining ligands. The homoleptic trimethylphosphite, dimethylmethylphosphonite and methyldiethylphosphinite technetium(I) cations

Abstract Technetium(I) complexes of trimethylphosphite (tmp), dimethylmethylphosphonite (dmmp) and methyldiethylphosphinite (mdep) have been synthesized through direct interaction of the ligand with sodium pertechnetate. The homoeleptic, hexakis (ligand), cationic complexes can be isolated as tetraphenylborate salts. Characterization using multinuclear NMR of 99 Tc (−422 and −248 ppm versus TcO 4 − for the tmp and dmmp complexes, respectively) and 31 P (158.5 and 248 ppm versus 85% H 3 PO 4 for the tmp and dmmp complexes, respectively) gives chemical shift values which compare favorably with previous data on Tc-phosphine and transition-metal-phosphite complexes. Mass spectra (fast atom bombardment and plasma desorption) of the tmp and dmmp complexes exhibit the expected parent peaks and show series of peaks attributed to stepwise loss of ligand molecules or possible decompositon products. X-ray photoelectron spectra of the tmp and dmmp complexes are consistent with formulation as the Tc(I) oxidation state when referenced to well-characterized complexes in known oxidation states.

Biodistribution and autoradiographic localization of I-125--labeled synthetic Peptide in aortic atherosclerosis in cholesterol-fed rabbits.

I-125-labeled SP4 is a synthetic oligopeptide derived from apolipoprotein B of low-density lipoprotein that has been shown to localize in atherosclerotic plaques in experimental animals. However, its biodistribution and mechanism of localization need to be further elucidated. Twenty-four cholesterol-fed (CF) and 20 normal (NL) New Zealand White rabbits were injected with I-125-SP4 and killed 15 to 30 min (6 NL; 6 CF) or 2 h (14 NL; 18 CF) later. We obtained aortic autoradiograms and activity concentrations (% injected dose/gin) in aortic segments and other tissues. The uptake of I-125-SP4 was higher in CF than in NL rabbits in all aortic segments (p < 0.05). I-125-SP4 was cleared rapidly in both CF and NL rabbits with 60 to 70% of the injected dose cleared from the blood by 1 h. No statistically significant differences in radiotracer biodistribution were observed between NL and CF rabbits although activity tended to be higher in the liver, gallbladder, and intestine in NL rabbits and in the kidney and spleen in CF rabbits. Silver grains were distributed mainly on foam cells of the fatty streaks in aortic microautoradiograms from two additional rabbits that had been injected with I-125-SP4. There were 23,518 ± 15,878 (SD) grains/mm2 in fatty plaques but only 14,669 ± 11,035 grains/mm2 in media muscle (p< 0.0001 [9 sections, 17 areas evaluated] in an atherosclerotic animal) in injected animals and 13,439 ± 5,565 grains/mm2 in media muscle (two sections, four areas) in the normal control animals (NS versus media of atherosclerotic animal). I-125-SP4 specifically localizes in aortic atherosclerotic plaques in CF rabbits. There is no significant difference in tissue distribution between normal and CF rabbits except in the aorta. Preliminarily, it appears that the site of tracer uptake is on foam cells and this suggests the possibility of relative specificity for fatty plaque.

Localization of (99m)Tc-Labeled ApoB Synthetic Peptide in Arterial Lesions of an Experimental Model of Spontaneous Atherosclerosis.

We have previously shown that after administration of 123[-SP-4 (a synthetic ApoB peptide fragment) to Watanabe heritable hyperlipidemic (WHHL) rabbits that foci of tracer uptake can be identified by external gamma camera imaging which correspond to regions of the aortas found to contain abundant atherosclerotic lesions at postmortem evaluation. Because 99mTc is preferred over 123I for scintigraphic imaging, we prepared a 99mTc-labeled form of the SP-4 peptide, designated 99mTc-P199. To assess the feasibility of detecting atherosclerotic lesions using 99mTc-P199 and to compare the relative uptake of the 99mTc-labeled and radioiodinated peptides by such lesions, an admixture of 99mTc-P199 and 125I-SP-4 was administered to 11 WHHL and 2 normal rabbits. These animals were imaged for up to 3 h and were sacrificed 3–4 h after injection. The extent of aortic lesion involvement and radiotracer uptake were quantitatively compared by planimetric analysis of photographs of the endothelial surface, 99mTc-P199 ex vivo images and 125I-SP-4 autoradiograms of the excised aortas. Pair wise correlation coefficients for planimetric analysis were as follows: photographs versus ex vivo images, r = 0.83, p = 0.003; photographs versus autoradiograms, r = 0.87, p – 0.001; ex vivo images versus autoradiograms, r = 0.83, p = 0.003. 99mTc-P199 in vivo gamma camera images revealed relatively weak focal aortic uptake in 8 of 11 WHHL rabbits manifesting aortic lesions, and focal carotid artery uptake in 4 of 6 WHHL rabbits manifesting carotid lesions. Neither aortic nor carotid foci were visualized in the normal rabbits. We conclude that 99mTc-P199 localizes specifically in atherosclerotic lesions and may be useful for external imaging of atherosclerosis.