Improved synthesis of 17ß-hydroxy-16α-iodo-wortmannin, 17ß-hydroxy-16α-iodoPX866, and the [(131)I] analogue as useful PET tracers for PI3-kinase.

An improved method for the synthesis of 17ß-hydroxy-16α-iodo-wortmannin along with the first synthesis of 17ß-hydroxy-16α-iodoPX866 and [(131)I] radiolabeled 17ß-hydroxy-16α-[(131)I]iodo-wortmannin, as potential PET tracers for PI3K was also described. The differences between wortmannin and its iodo analogue were compared by covalently docking each structure to L833 in PI3K.

Actinium-225 Targeted Agents: Where Are We Now?

α-particle targeted radionuclide therapy has shown promise for optimal cancer management, an exciting new era for brachytherapy. Alpha-emitting nuclides can have significant advantages over gamma- and beta-emitters due to their high linear energy transfer (LET). While their limited path length results in more specific tumor 0kill with less damage to surrounding normal tissues, their high LET can produce substantially more lethal double strand DNA breaks per radiation track than beta particles. Over the last decade, the physical and chemical attributes of Actinium-225 (225Ac) including its half-life, decay schemes, path length, and straightforward chelation ability has peaked interest for brachytherapy agent development. However, this has been met with challenges including source availability, accurate modeling for standardized dosimetry for brachytherapy treatment planning, and laboratory space allocation in the hospital setting for on-demand radiopharmaceuticals production. Current evidence suggests that a simple empirical approach based on 225Ac administered radioactivity may lead to inconsistent outcomes and toxicity. In this review article, we highlight the recent advances in 225Ac source production, dosimetry modeling, and current clinical studies.

Development of (99m)Tc-N4-NIM for molecular imaging of tumor hypoxia.

The nitro group of 2-nitroimidazole (NIM) enters the tumor cells and is bioreductively activated and fixed in the hypoxia cells. 1,4,8,11-tetraazacyclotetradecane (N4) has shown to be a stable chelator for (99m)Tc. The present study was aimed to develop (99m)Tc-cyclam-2-nitroimidazole ((99m)Tc-N4-NIM) for tumor hypoxia imaging. N4-NIM precursor was synthesized by reacting N4-oxalate and 1,3-dibromopropane-NIM, yielded 14% (total synthesis). Cell uptake of (99m)Tc-N4-NIM and (99m)Tc-N4 was obtained in 13762 rat mammary tumor cells and mesothelioma cells in 6-well plates. Tissue distribution of (99m)Tc-N4-NIM was evaluated in breast-tumor-bearing rats at 0.5-4 hrs. Tumor oxygen tension was measured using an oxygen probe. Planar imaging was performed in the tumor-bearing rat and rabbit models. Radiochemical purity of (99m)Tc-N4-NIM was >96% by HPLC. Cell uptake of (99m)Tc-N4-NIM was higher than (99m)Tc-N4 in both cell lines. Biodistribution of (99m)Tc-N4-NIM showed increased tumor-to-blood and tumor-to-muscle count density ratios as a function of time. Oxygen tension in tumor tissue was 6-10 mmHg compared to 40-50 mmHg in normal muscle tissue. Planar imaging studies confirmed that the tumors could be visualized clearly with (99m)Tc-N4-NIM in animal models. Efficient synthesis of N4-NIM was achieved. (99m)Tc-N4-NIM is a novel hypoxic probe and may be useful in evaluating cancer therapy.

Development of tyrosine-based radiotracer 99mTc-N4-Tyrosine for breast cancer imaging.

The purpose of this study was to develop an efficient way to synthesize (99m)Tc-O-[3-(1,4,8,11-tetraazabicyclohexadecane)-propyl]-tyrosine ((99m)Tc-N4-Tyrosine), a novel amino acid-based radiotracer, and evaluate its potential in breast cancer gamma imaging. Precursor N4-Tyrosine was synthesized using a 5-step procedure, and its total synthesis yield was 38%. It was successfully labeled with (99m)Tc with high radiochemical purity (>95%). Cellular uptake of (99m)Tc-N4-Tyrosine was much higher than that of (99m)Tc-N4 and the clinical gold standard (18)F-2-deoxy-2-fluoro-glucose ((18)F-FDG) in rat breast tumor cells in vitro. Tissue uptake and dosimetry estimation in normal rats revealed that (99m)Tc-N4-Tyrosine could be safely administered to humans. Evaluation in breast tumor-bearing rats showed that although (99m)Tc-N4-Tyrosine appeared to be inferior to (18)F-FDG in distinguishing breast tumor tissue from chemical-induced inflammatory tissue, it had high tumor-to-muscle uptake ratios and could detect breast tumors clearly by planar scintigraphic imaging. (99m)Tc-N4-Tyrosine could thus be a useful radiotracer for use in breast tumor diagnostic imaging.

Molecular imaging of mesothelioma with (99m)Tc-ECG and (68)Ga-ECG.

We have developed ethylenedicysteine-glucosamine (ECG) as an alternative to (18)F-fluoro-2-deoxy-D-glucose ((18)F-FDG) for cancer imaging. ECG localizes in the nuclear components of cells via the hexosamine biosynthetic pathway. This study was to evaluate the feasibility of imaging mesothelioma with (99m)Tc-ECG and (68)Ga-ECG. ECG was synthesized from thiazolidine-4-carboxylic acid and 1,3,4,6-tetra-O-acetyl-2-amino-D-glucopyranose, followed by reduction in sodium and liquid ammonia to yield ECG (52%). ECG was chelated with (99m)Tc/tin (II) and (68)Ga/(69)Ga chloride for in vitro and in vivo studies in mesothelioma. The highest tumor uptake of (99m)Tc-ECG is 0.47 at 30 min post injection, and declined to 0.08 at 240 min post injection. Tumor uptake (%ID/g), tumor/lung, tumor/blood, and tumor/muscle count density ratios for (99m)Tc-ECG (30-240 min) were 0.47 ± 0.06 to 0.08 ± 0.01; 0.71 ± 0.07 to 0.85 ± 0.04; 0.47 ± 0.03 to 0.51 ± 0.01, and 3.49 ± 0.24 to 5.06 ± 0.25; for (68)Ga-ECG (15-60 min) were 0.70 ± 0.06 to 0.92 ± 0.08; 0.64 ± 0.05 to 1.15 ± 0.08; 0.42 ± 0.03 to 0.67 ± 0.07, and 3.84 ± 0.52 to 7.00 ± 1.42; for (18)F-FDG (30-180 min) were 1.86 ± 0.22 to 1.38 ± 0.35; 3.18 ± 0.44 to 2.92 ± 0.34, 4.19 ± 0.44 to 19.41 ± 2.05 and 5.75 ± 2.55 to 3.33 ± 0.65, respectively. Tumor could be clearly visualized with (99m)Tc-ECG and (68)Ga-ECG in mesothelioma-bearing rats. (99m)Tc-ECG and (68)Ga-ECG showed increased uptake in mesothelioma, suggesting they may be useful in diagnosing mesothelioma and also monitoring therapeutic response.

Chelation-Tamoxifen Conjugates for Imaging of Estrogen Receptors.

Background: The differential diagnosis of estrogen receptor-positive (ER+) pathway-activated systems by using a labeled antiestrogen helps to select the patients for optimal response to endocrine therapy and to discontinue the treatment when resistance occurs. The authors' purpose was to synthesize chelator-tamoxifen conjugates for imaging ER (+) diseases. Materials and Methods: A hydroxypropyl linker was incorporated between either cyclam or cyclam diacetic acid and tamoxifen analog to produce SC-05-L-1 (Z-1-(1,4,8,11-tetraazacyclotetradecan-1-yl)-3-((5-(4-(2-(diethylamino)ethoxy)phenyl)-4,5-diphenylpent-4-en-1-yl)oxy)propan-2-ol) and SC-05-N-1 (Z-2,2'-(4-(3-((5-(4-(2-(diethylamino)ethoxy)phenyl)-4,5-diphenylpent-4-en-1-yl)oxy)-2-hydroxy-propyl)-1,4,8,11-tetraazacyclotetradecane-1,8-diyl)diacetic acid), respectively. In vitro cell uptake and cell/media ratios of 99mTc-SC-05-L-1 and 99mTc- SC-05-N-1 in ER (+) ovarian cancer cells (TOV-112D and OVCAR3) were performed. To ascertain the specificity of cell uptake, the cell uptake was blocked with estrone. In vivo 99mTc-SC-05-L-1 or 99mTc-SC-05-N-1 single-photon emission computed tomography/computed tomography was conducted in tumor-bearing rodents and compared to 18F-fluoro-2-deoxy-d-glucose (18F-FDG) positron emission tomography/magnetic resonance imaging (a reference technology). Results: The radiochemical purities of 99mTc-SC-05-L-1 and 99mTc-SC-05-N-1 were greater than 99% (n = 10). 99mTc-SC-05-L-1 had higher cell/media ratios than 99mTc-SC-05-N-1 in OVCAR-3 ER (+) cells. The cell uptake of 99mTc-SC-05-L-1 was blocked 80% by estrone indicating an ER-mediated process occurred. 99mTc-SC-05-N-1 was further selected for in vivo imaging studies due to higher maximum tolerated dose and superior water solubility than 99mTc-SC-05-L-1. 99mTc-SC-05-N-1 showed higher tumor uptake and tumor/muscle count density ratios than 18F-FDG in tumor-bearing rodents. Conclusion: 99mTc-SC-05-N-1 showed better differential diagnosis of ovarian tumors than 18F-FDG, indicating great promising in chelator-tamoxifen conjugate for ER pathway-directed systems imaging.

Development of 68Ga-glycopeptide as an imaging probe for tumor angiogenesis.

Objective. This study was aimed to study tissue distribution and tumor imaging potential of (68)Ga-glycopeptide (GP) in tumor-bearing rodents by PET. Methods. GP was synthesized by conjugating glutamate peptide and chitosan. GP was labeled with (68)Ga chloride for in vitro and in vivo studies. Computer outlined region of interest (counts per pixel) of the tumor and muscle (at the symmetric site) was used to determine tumor-to-muscle count density ratios. To ascertain the feasibility of (68)Ga-GP in tumor imaging in large animals, PET/CT imaging of (68)Ga-GP and (18)F-FDG were conducted in New Zealand white rabbits bearing VX2 tumors. Standard uptake value of tumors were determined by PET up to 45 min. To determine blood clearance and half-life of (68)Ga-GP, blood samples were collected from 10 seconds to 20 min. Results. Radiochemical purity of (68)Ga-GP determined by instant thin-layer chromatography was >95%. Tumor uptake values (SUV) for (68)Ga-GP and (18)F-FDG in New Zealand white rabbits bearing VX2 tumors were 3.25 versus 7.04. PET images in tumor-bearing rats and rabbits confirmed that (68)Ga-GP could assess tumor uptake. From blood clearance curve, the half-life of (68)Ga-GP was 1.84 hr. Conclusion Our data indicate that it is feasible to use (68)Ga-GP to assess tumor angiogenesis.

Synthesis and evaluation of amino acid-based radiotracer 99mTc-N4-AMT for breast cancer imaging.

PURPOSE: This study was to develop an efficient synthesis of (99m)Tc-O-[3-(1,4,8,11-tetraazabicyclohexadecane)-propyl]-α-methyl tyrosine ((99m)Tc-N4-AMT) and evaluate its potential in cancer imaging. METHODS: N4-AMT was synthesized by reacting N4-oxalate and 3-bromopropyl AMT (N-BOC, ethyl ester). In vitro cellular uptake kinetics of (99m)Tc-N4-AMT was assessed in rat mammary tumor cells. Tissue distribution of the radiotracer was determined in normal rats at 0.5-4 h, while planar imaging was performed in mammary tumor-bearing rats at 30-120 min. RESULTS: The total synthesis yield of N4-AMT was 14%. Cellular uptake of (99m)Tc-N4-AMT was significantly higher than that of (99m)Tc-N4. Planar imaging revealed that (99m)Tc-N4-AMT rendered greater tumor/muscle ratios than (99m)Tc-N4. CONCLUSIONS: N4-AMT could be synthesized with a considerably high yield. Our in vitro and in vivo data suggest that (99m)Tc-N4-AMT, a novel amino acid-based radiotracer, efficiently enters breast cancer cells, effectively distinguishes mammary tumors from normal tissues, and thus holds the promise for breast cancer imaging.

Molecular imaging of Bcr-Abl phosphokinase in a xenograft model.

The purpose of this study was to determine whether the Bcr-Abl tyrosine kinase can be assessed by gamma-imaging using an 111In-labeled anti-phosphotyrosine (APT) antibody, and if the response to treatment with imatinib could be detected using this imaging technique. APT antibody was labeled with 111In using ethylenedicysteine (EC) as a chelator. To determine if 111In-EC-APT could assess a nonreceptor tyrosine kinase, xenografts of the human chronic myelogenous leukemia cell line K562 were used. gamma-Scintigraphy of the tumor-bearing mice, before and after imatinib treatment, was obtained 1, 24, and 48 h after they were given 111In-EC-APT (100 microCi/mouse i.v.). 111In-EC-APT is preferentially taken up by Bcr-Abl-bearing tumor cells when compared with 111In-EC-BSA or 111In-EC-IgG1 controls and comparable with the level of uptake of 111In-EC-Bcr-Abl. Imatinib treatment resulted in decreased expression of phospho-Bcr-Abl by Western blot analysis, which correlated with early (4 days after starting imatinib) kinase down-regulation as assessed by imaging using 111In-EC-APT. The optimal time to imaging was 24 and 48 h after injection of 111In-EC-APT. Although tumor regression was insignificant on day 4 after starting imatinib treatment, it was marked by day 14. 111In-EC-APT can assess intracellular phosphokinase activity, and down-regulation of phosphokinase activity predates tumor regression. This technique may therefore be useful in the clinic to detect the presence of phosphokinase activity and for early prediction of response.

Radiation dosimetry and biodistribution of (99m)Tc-ethylene dicysteine-deoxyglucose in patients with non-small-cell lung cancer.

PURPOSE: To assess the radiation dosimetry and biodistribution of (99m)Tc-labeled ethylene dicysteine deoxyglucose ((99m)Tc-EC-DG) in patients with non-small-cell lung cancer (NSCLC). METHODS: Serial whole-body scans were acquired 0, 2, 4, 6 and 24 h after injection of (99m)Tc-EC-DG (925 MBq) in seven NSCLC patients. Radiation dosimetry, blood clearance and SPECT imaging of the primary tumor were assessed. RESULTS: The critical organ was the bladder wall, with average radiation absorbed dose over all seven patients of 2.47x10(-2) mGy/MBq. The average effective dose equivalent and effective dose were 6.20x10(-3) mSv/MBq (6.89 mSv/1,110 MBq) and 5.90x10(-3) mSv/MBq (6.54 mSv/1,110 MBq), respectively. The primary tumor was visualized with SPECT in six patients. On final pathology, one patient had a granuloma, which did not enhance with (99m)Tc-EC-DG. CONCLUSION: (99m)Tc-EC-DG has acceptable dosimetric and biodistribution properties as a diagnostic tumor-imaging agent. Future studies are planned to evaluate its diagnostic potential.

99m Tc-glycopeptide: synthesis, biodistribution and imaging in breast tumor-bearing rodents.

This study was aimed to develop a glycopeptide (GP) to be used as a carrier for anti-cancer drug delivery. GP was synthesized by conjugating glutamate peptide and chitosan using carbodiimide as a coupling agent. Elemental analysis and capillary electrophoresis confirmed the purity was >95%. GP was labeled with sodium pertechnetate (Na99m TcO4) for in vitro and in vivo studies. Rhenium-GP was synthesized to support the binding site of 99m Tc at the glutamate positions 3-5. In vitro cellular uptake of 99m Tc-GP was performed in breast cancer cells. Cytosol had 60% whereas nucleus had 40% uptake of 99m Tc-GP. When cancer cells were incubated with glutamate or aspartate, followed by 99m Tc-GP, there was decreased uptake in cells treated with glutamate but not aspartate. The findings indicated that cellular uptake of 99m Tc-GP was via glutamate transporters. In addition, 99m Tc-GP was able to measure uptake differences after cells treated with paclitaxel. Biodistribution and planar imaging were conducted in breast tumor-bearing rats. Biodistribution of 99m Tc-GP showed increased tumor-to-tissue ratios as a function of time. Planar images confirmed that 99m Tc-GP could assess tumor uptake changes after paclitaxel treatment. In vitro and in vivo studies indicated that GP could target tumor cells, thus, GP may be a useful carrier for anti-cancer drug delivery.

Imaging and dosimetry of 99mTc EC annexin V: preliminary clinical study targeting apoptosis in breast tumors.

BACKGROUND: Early detection of cellular events is important to predict the outcome of the patients. This study was aimed to use (99m)Tc EC-annexin V to image tumor cells undergoing apoptosis. METHODS: In 10 patients with breast cancer, scintigraphic images and dosimetric estimates were obtained after administering (99m)Tc EC-annexin V. RESULTS: Nine of the 10 cases showed detectable (99m)Tc EC-annexin V uptake in tumor. Higher values of T/N ratios are associated with patient after treatment. CONCLUSIONS: Apoptosis can be quantified using (99m)Tc EC-annexin V.

Efficient Synthesis of Glutamate Peptide-Estradiol Conjugate for Imaging Estrogen Receptor-Positive Diseases.

Molecular imaging of estrogen receptor-positive (ER+) pathway-activated system serves the basis of ER+ disease management such as cancers and endometriosis. ER+ patients have better response to endocrine therapy and survive twice as long as negative ER patients. However, tumor resistance resulting from clinical used aromatase inhibitors and antiestrogens is unpredictable. Radiolabeled ER+ ligand could quantify ER+ tissue uptake which helps to stage and restage of the cancer as well as endometriosis. The differential diagnosis of ER+ lesions by using a labeled ligand helps to select the patients for optimal response to endocrine therapy and to discontinue the treatment when resistance occurs. In addition, radiolabeled ER+ ligand serves as basis for image-guided response follow-up. Glutamate receptors are cell surface receptors which are overexpressed in inflammation and infection. Using glutamate peptide as a drug carrier helps to target intracellular genes via glutamate receptor-mediated process. Reports have shown that polyglutamate is a drug carrier that could alter drug solubility and enhance estrogen receptor-ligand binding pocket. However, polyglutamate was a blend of mixed polymer with a wide range of molecular weight. Thus, the structural confirmation and purity of the conjugates were not optimized. To overcome this problem, the efficient synthesis of glutamate peptide-estradiol (GAP-EDL) conjugate was achieved with high purity. EDL was conjugated site-specific at the first glutamate of GAP. The average cell uptake of 68Ga-GAP-EDL was 5-fold higher than the previous reported synthesis. The efficient synthesis of GAP-EDL has greatly enhanced sensitivity and specificity in cell uptake studies. In vivo PET imaging studies indicated that 68Ga-GAP-EDL could image ER (+) tumors in MCF-7 tumor-bearing mice. Therefore, GAP-EDL makes it possible to image ER-enriched endometriosis and cancer.

Infrared-based module for the synthesis of 68Ga-labeled radiotracers.

INTRODUCTION: Generator-produced positron emission tomography tracers have gained much attention recently due to favorable imaging characteristics, accessibility and affordability. The focus of this study was to design and validate a semiautomated module for 68Ga-labeled chemistry utilizing infrared-based heating for rapid control of thermal cycle. METHODS: A prototype module was built and installed in our laboratory. DOTA (1,4,7,10-tetra-azacyclododecane-1,4,7,10-tetra-acetic acid) was manually labeled (10-1000 nmol) with 68Ga to optimize synthesis conditions. For automation, 250 nmol of DOTA was labeled with 68Ga with reaction times of 5 min (n=5), 10 min (n=5) and 20 min (n=6). A dose calibrator and radio-thin-layer chromatography were used to access the product yield and quality of both manual and automated syntheses. RESULTS: A semiautomated 68Ga synthesis module was developed. The system showed that software control could be used to drive a multistep radiochemical synthesis and to produce 68Ga-DOTA with >95% radiochemical purity, similar to that in manual synthesis. The device also showed that for a short reaction time of 5 min, decay-corrected radioactive yields of >70% could be achieved. The total synthesis was as short as 22 min, including 6-8 min for HCl evaporation. The temperature and pressure profiles of the process were consistent. CONCLUSION: We demonstrated the use of a commercially available 68Ga/68Ge generator with a semiautomated module to successfully label the bifunctional chelator DOTA with 68Ga. Further investigation with different 68Ga-labeled bioconjugates is warranted to demonstrate the usefulness of the module as a tool for tracer development and imaging research.

Functional imaging of estrogen receptors with radiolabeled-GAP-EDL in rabbit endometriosis model.

RATIONALE AND OBJECTIVES: Endometriosis is a common women's health problem. Animal models provide an invaluable tool to study the natural history of endometriosis. We previously have reported that (99m)Tc-labeled glutamate peptide-estradiol ((99m)Tc-GAP-EDL) is a useful agent for imaging functional estrogen receptor (ER) via an ER-mediated process. This study was to evaluate the feasibility of using radiolabeled GAP-EDL to image ER-positive (ER +) endometriosis in nonprimate animal models. MATERIALS AND METHODS: 3-Aminoethyl estradiol (EDL) was conjugated to glutamate peptide (GAP) to yield GAP-EDL. In vitro cellular uptake studies of (99m)Tc and (68)Ga-GAP-EDL inhibition with cold estrone were conducted in 13,762 rat mammary tumor cells. To create a rabbit model with endometriosis, part of uterine tissue was dissected and grafted in the peritoneal wall. Eight weeks after surgery, scintigraphic images were obtained after intravenous injection of (99m)Tc-GAP-EDL (1 mCi/rabbit, intravenous) at 0.5-2.0 hours, and (68)Ga-GAP-EDL at 45 minutes. We also performed (68)Ga-GAP-EDL blocking study in rabbit model by using tamoxifen. The rabbits were sacrificed and the grafts were excised for histologic examination. RESULTS: In vitro uptake study of (99m)Tc- and (68)Ga-GAP-EDL in 13,762 rat breast cancer cells showed gradually increasing uptake of both tracers. Accumulation of (68)Ga-GAP-EDL in 13,762 cells was inhibited with cold estrone in a dose-dependent manner. In the endometriosis model, the grafted uterine tissue could be visualized by (99m)Tc-GAP-EDL. Necropsy was performed at 2.5 hours after injection time. Four follicular endometrial lesions in eight implanted endometrial tissues were detected, and all lesions could be detected by (99m)Tc-GAP-EDL. Planar scintigraphy of uterus, ovary and implants of necropsy specimen revealed an increased uptake of (99m)Tc-GAP-EDL in comparison with surrounding abdominal wall tissue. Microscopic examinations support that (99m)Tc-GAP-EDL was accumulated in the microinvasive endometrial tissue. After blocking with tamoxifen, (68)Ga-GAP-EDL accumulation in the endometrial grafts could not be visualized, and endometrial tissue-to-normal tissue count ratios were statistically higher in a nonblocked image than that in the blocked image. CONCLUSIONS: Endometriosis uptake of radiolabeled GAP-EDL was via an estrogen receptor-mediated process. Radiolabeled-GAP-EDLs are useful agents for imaging endometriosis.

PET and planar imaging of tumor hypoxia with labeled metronidazole.

RATIONALE AND OBJECTIVES: This study was aimed to develop 99mTc- and 68Ga-labeled metronidazole (MN) using ethylenedicysteine (EC) as a chelator and evaluate their potential use to assess tumor hypoxia. MATERIALS AND METHODS: EC-MN was labeled with 99mTc in the presence of tin (II) chloride. Labeling EC-MN with 68Ga was achieved by adding 68GaCl3 (2 mCi with 3.4 microg cold GaCl3). In vitro cellular uptakes of 99mTc- and 68Ga-EC-MN were obtained in various types of tumor cells at 0.5-4 hours. Tissue distribution and PET imaging of 99mTc and 68Ga-EC-MN were evaluated in breast tumor-bearing rats at 0.5-4 hours. Tumor oxygen tension was measured using an oxygen probe. RESULTS: There were similar cellular uptakes (2-10%) between 99mTc- and 68Ga-EC-MN at 0.5-4 hours. In vivo biodistribution of 99mTc- and 68Ga-EC-MN in breast tumor-bearing rats showed increased tumor-to-blood and tumor-to-muscle count density ratios as a function of time. Positron emission tomography images confirmed that the tumors could be visualized clearly with 68Ga-EC-MN. Oxygen tension in tumor tissue was determined to be 6-10 mm Hg compared with 40-50 mm Hg in normal muscle tissue. CONCLUSIONS: The results indicated that it is feasible to use 99mTc- and 68Ga-EC-MN for assessment of tumor hypoxia. These agents may be useful in selecting and evaluating cancer therapy.

Radiolabeled L-lysine for tumor imaging.

RATIONALE AND OBJECTIVES: The aims of this study were to label the versatile amino acid l-lysine with (99m)Tc using 2,3-dimercapto-succinic acid (DMSA) as a chelator, and to assess its tumor imaging feasibility under in vivo and in vitro conditions, and finally to determine the subcellular biodistribution of this radiopharmaceutical. MATERIALS AND METHODS: DMSA-l-lysine was chemically synthesized and labeled with sodium pertechnetate. Nuclear magnetic resonance (NMR) and mass spectral analysis of DMSA-l-lysine were conducted. Radiochemical purity was determined by thin-layer chromatography (TLC) and paper chromatography. Cellular uptake, competition and subcellular localization studies were performed in rat breast cancer cells (13762). In vivo studies of planar imaging and biodistribution studies were performed on female Fischer 344 rats. Medical Internal Radiation Dose (MIRD) dosimetry estimates were calculated. RESULTS: Radiochemical purity (determined by radio-TLC and high-performance liquid chromatography) of these compounds was >95%. (99m)Tc-DMSA-l-lysine showed good uptake in in vitro cell culture assays and uptake was reduced in competition studies. (99m)Tc-DMSA-l-lysine accumulates in the nucleus as much as in the cytoplasm and it was also shown that accumulation of the (99m)Tc-DMSA-l-lysine in the nucleus increases as a function of a time. There was an increase in tumor-to-blood and tumor-to-muscle count density ratios. Tumor/background ratios were 5.75 at 1 hour and 6.87 at 2 hours. In vivo tissue distribution studies revealed that radiation dosimetry of blood-forming organs were within radiation dose limits. CONCLUSION: DMSA-l-lysine kits can be labeled with (99m)Tc easily and efficiently, with high radiochemical purity and cost-effectiveness. In vitro cellular uptake and scintigraphic imaging studies demonstrated the pharmacokinetic distribution and feasibility of using (99m)Tc-DMSA-l-lysine for tumor imaging.