MicroSPECT and MicroPET Imaging of Small Animals for Drug Development

The process of drug discovery and development requires substantial resources and time. The drug industry has tried to reduce costs by conducting appropriate animal studies together with molecular biological and genetic analyses. Basic science research has been limited to in vitro studies of cellular processes and ex vivo tissue examination using suitable animal models of disease. However, in the past two decades new technologies have been developed that permit the imaging of live animals using radiotracer emission, X-rays, magnetic resonance signals, fluorescence, and bioluminescence. The main objective of this review is to provide an overview of small animal molecular imaging, with a focus on nuclear imaging (single photon emission computed tomography and positron emission tomography). These technologies permit visualization of toxicodynamics as well as toxicity to specific organs by directly monitoring drug accumulation and assessing physiological and/or molecular alterations. Nuclear imaging technology has great potential for improving the efficiency of the drug development process.

Biodistribution of 99mTc Labeled Integrin Antagonist

The selective targeting of an integrin alphavbeta3 receptor using radioligands may enable the assessment of angiogenesis and integrin alphavbeta3 receptor status in tumors. The aim of this research was to label a peptidomimetic integrin alphavbeta3 antagonist (PIA) with 99mTc(CO)3 and to test its receptor targeting properties in nude mice bearing receptor-positive tumors. PIA was reacted with tris-succinimidyl aminotriacetate (TSAT) (20 mM) as a PIA per TSAT. The product, PIA-aminodiacetic acid (ADA), was radiolabeled with [99mTc(CO)3(H2O)3](+1), and purified sequentially on a Sep-Pak C-18 cartridge followed by a Sep-Pak QMA anion exchange cartridge. Using gradient C-18 reverse-phase HPLC, the radiochemical purity of 99mTc(CO)3-ADA-PIA (retention time, 10.5 min) was confirmed to be > 95%. Biodistribution analysis was performed in nude mice (n = 5 per time point) bearing receptor-positive M21 human melanoma xenografts. The mice were administered 99mTc(CO)3-ADA-PIA intravenously. The animals were euthanized at 0.33, 1, and 2 hr after injection for the biodistribution study. A separate group of mice were also co-injected with 200 microg of PIA and euthanized at 1 hr to quantify tumor uptake. 99mTc(CO)3-ADA-PIA was stable in phosphate buffer for 21 hr, but at 3 and 6 hr, 7.9 and 11.5% of the radioactivity was lost as histidine, respectively. In tumor bearing mice, 99mTc(CO)3-ADA-PIA accumulated rapidly in a receptor-positive tumor with a peak uptake at 20 min, and rapid clearance from blood occurring primarily through the hepatobiliary system. At 20 min, the tumor-to-blood ratio was 1.8. At 1 hr, the tumor uptake was 0.47% injected dose (ID)/g, but decreased to 0.12% ID/g when co-injected with an excess amount of PIA, indicating that accumulation was receptor mediated. These results demonstrate successful 99mTc labeling of a peptidomimetic integrin antagonist that accumulated in a tumor via receptor-specific binding. However, tumor uptake was very low because of low blood concentrations that likely resulted from rapid uptake of the agent into the hepatobiliary system. This study suggests that for 99mTc(CO)3-ADA-PIA to be useful as a tumor detection agent, it will be necessary to improve receptor binding affinity and increase the hydrophilicity of the product to minimize rapid hepatobiliary uptake.

Biodistribution of 99mTc Tricarbonyl Glycine Oligomers

99mTc tricarbonyl glycine monomers, trimers, and pentamers were synthesized and evaluated for their radiolabeling and in vivo distribution characteristics. We synthesized a 99mTc-tricarbonyl precursor with a low oxidation state (I). 99mTc(CO)3(H2O)3 + was then made to react with monomeric and oligomeric glycine for the development of bifunctional chelating sequences for biomolecules. Labeling yields of 99mTc-tricarbonyl glycine monomers and oligomers were checked by high-performance liquid chromatography. The labeling yields of 99mTc-tricarbonyl glycine and glycine oligomers were more than 95%. We evaluated the characteristics of 99mTc-tricarbonyl glycine oligomers by carrying out a lipophilicity test and an imaging study. The octanol-water partition coefficient of 99mTc tricarbonyl glycine oligomers indicated hydrophilic properties. Single-photon emission computed tomography imaging of 99mTc-tricarbonyl glycine oligomers showed rapid renal excretion through the kidneys with a low uptake in the liver, especially of 99mTc tricarbonyl triglycine. Furthermore, we verified that the addition of triglycine to prototype biomolecules (AGRGDS and RRPYIL) results in the improvement of radiolabeling yield. From these results, we conclude that triglycine has good characteristics for use as a bifunctional chelating sequence for a 99mTc-tricarbonyl-based biomolecular imaging probe.

Single-Dose Oral Toxicity of Fermented Scutellariae Radix Extract in Rats and Dogs

The aim of this study was to investigate the acute oral toxicity of fermented Scutellariae Radix (JKTMHGu-100) in rats and dogs. JKTM-HGu-100 was orally administered at a dose of 2,000 mg/kg in Sprague-Dawley rats. An escalating single-dose oral toxicity test in beagle dogs was performed at doses of 500, 1000, and 2000 mg/kg with 4-day intervals. Clinical signs, changes in body weight, mortality, and necropsy findings were examined for 2 weeks following oral administration. No toxicological changes related to the test substance nor mortality was observed after administration of a single oral dose of JKTM-HGu-100 in rats or dogs. Therefore, the approximate lethal dose (LD) for oral administration of JKTMHGu-100 in rats was considered to be over 2,000 mg/kg, and the maximum tolerance doses (MTDs) in rats and dogs were also estimated to be over 2,000 mg/kg. These results indicate that JKTM-HGu-100 shows no toxicity in rodents or non-rodents at doses of 2,000 mg/kg or less.