Noninvasive imaging of c(RGD)2‐9R as a potential delivery carrier for transfection of siRNA in malignant tumors

The purpose of our study was to develop and evaluate a novel integrin α_vβ₃‐specific delivery carrier for transfection of siRNA in malignant tumors. We adopted arginine‐glycine‐aspartate (RGD) motif as a tissue target for specific recognition of integrin α_νβ₃. A chimaeric peptide was synthesized by adding nonamer arginine residues (9‐arginine [9R]) at the carboxy terminus of cyclic‐RGD dimer, designated as c(RGD)₂‐9R, to enable small interfering RNA (siRNA) binding. To test the applicability of the delivery carrier in vivo, c(RGD)₂‐9R was labeled with radionuclide of technetium‐99m. Biodistribution and γ‐camera imaging studies were performed in HepG2 xenograft‐bearing nude mice. As results, an optimal 10:1 molar ratio of ^99mTc‐c(RGD)₂‐9R to siRNA was indicated by the electrophoresis on agarose gels. ^99mTc‐c(RGD)₂‐9R/siRNA remained stable under a set of conditions in vitro. For in vivo study, tumor radioactivity uptake of ^99mTc‐c(RGD)₂‐9R/siRNA in nude mice bearing HepG2 xenografts was significantly higher than that of control probe (P  < .05). The xenografts were clearly visualized at 4 hours till 6 hours noninvasively after intravenous injection of ^99mTc‐c(RGD)₂‐9R/siRNA, while the xenografts were not visualized at any time after injection of control probe. It was concluded that c(RGD)₂‐9R could be an effective siRNA delivery carrier. Technetium‐99m radiolabeled‐delivery carrier represents a potential imaging strategy for RNAi‐based therapy.     

Preparation via coligand exchange and characterization of [99mTc‐EDDA‐HYNIC‐D‐Phe1,Tyr3]Octreotide (99mTc–EDDA/HYNIC–TOC)

[^99mTc‐EDDA–HYNIC‐D‐Phe¹,Tyr³]octreotide (^99mTc‐EDDA/HYNIC–TOC) is a promising new agent with the potential to replace [¹¹¹In‐DTPA‐D‐Phe¹]‐octreotide in somatostatin receptor scintigraphy. This hydrazinonicotinic acid derivatized somatostatin complex contains ethylenediamine N,N′ diacetic acid (EDDA) as a coligand resulting in a high in vitro and in vivo stability. Since direct ^99mTc‐labelling of HYNIC–TOC with EDDA results in low labelling yields, in this study we describe the preparation of ^99mTc‐EDDA/HYNIC‐TOC via coligand exchange from Tricine for EDDA. Exchange of coligands is achieved at elevated temperature and under optimized conditions of pH, EDDA concentration and stannous ion. High labelling yields (mean 95.9%) were achieved at high specific activities (>37GBq/µmol). Characterization via HPLC, receptor binding and LC–MS of the resulting complex is described. The formulation developed enables rapid and simple labelling of ^99mTc‐EDDA/HYNIC–TOC in a manner suitable for a clinical setting. Copyright © 2003 John Wiley & Sons, Ltd.     

Preparation and comparative evaluation of 99mTc‐HYNIC‐cNGR and 99mTc‐HYNIC‐PEG2‐cNGR as tumor‐targeting molecular imaging probes

The tripeptide sequence asparagine‐glycine‐arginine (NGR) specifically recognizes aminopeptidase N (APN or CD13) receptors highly expressed on tumor cells and vasculature. Thus, NGR peptides can precisely deliver therapeutic and diagnostic compounds to CD13 expressing cancer sites. In this regard, 2 NGR peptide ligands, HYNIC‐c(NGR) and HYNIC‐PEG₂‐c(NGR), were synthesized, radiolabeled with ^99mTc, and evaluated in CD13‐positive human fibrosarcoma HT‐1080 tumor xenografts. The radiotracers, ^99mTc‐HYNIC‐c(NGR) and ^99mTc‐HYNIC‐PEG₂‐c(NGR), could be prepared in approximately 95% radiochemical purity and exhibited excellent in vitro and in vivo stability. The radiotracers were hydrophilic in nature with log P values being ?2.33 ± 0.05 and ?2.61 ± 0.08. The uptake of 2 radiotracers ^99mTc‐HYNIC‐c(NGR) and ^99mTc‐HYNIC‐PEG₂‐c(NGR) was similar in nude mice bearing human fibrosarcoma HT‐1080 tumor xenografts, which was significantly reduced (P < .05) during blocking studies. The 2 radiotracers being hydrophilic cleared rapidly from blood, liver, and intestine and were excreted through renal pathway. The pharmacokinetics of ^99mTc‐labeled HYNIC peptide could not be modulated through introduction of PEG₂ unit, thus posing a challenge for studies with other linkers towards enhanced tumor uptake and retention.     

99mTc‐(EDDA/tricine)‐HYNIC‐GnRH analogue as a potential imaging probe for diagnosis of prostate cancer

Prostate cancer is a serious threat to men's health, so it is necessary to develop the techniques for early detection of this malignancy. Radiolabeled peptides are the useful tools for diagnosis of prostate cancer. In this research, we designed a new HYNIC‐conjugated GnRH analogue and labeled it by ^99mTc with tricine/EDDA as coligands. We used aminohexanoic acid (Ahx) as a hydrocarbon linker to generate ^99mTc‐(tricine/EDDA)‐HYNIC‐Ahx‐[DLys⁶]GnRH. The radiopeptide exhibited high radiochemical purity and stability in solution and serum. Two human prostate cancer cell lines LN‐CaP and DU‐145 were used for cellular experiments. The binding specificity and affinity of radiopeptide for LN‐CaP were superior to DU‐145 cells. The K_d values for LN‐CaP and DU‐145 cells were 41.91 ± 7.03 nM and 55.96 ± 10.56 nM, respectively. High kidney uptake proved that the main excretion route of radiopeptide was through the urinary system. The tumor/muscle ratio of ^99mTc‐HYNIC‐Ahx‐[DLys⁶]GnRH was 4.14 at 1 hr p.i. that decreased to 2.41 at 4 hr p.i. in LN‐CaP tumor‐xenografted nude mice. The blocking experiment revealed that the tumor uptake was receptor‐mediated. The lesion was visualized clearly using ^99mTc‐[DLys⁶]GnRH at 1 hr p.i. Accordingly, this research highlights the capability of ^99mTc‐(tricine/EDDA)‐HYNIC‐Ahx‐[DLys⁶]GnRH peptide as a promising agent for GnRHR‐expressing tumor imaging.     

18F labeled RGD‐A7R peptide for dual integrin and VEGF‐targeted tumor imaging in mice bearing U87MG tumors

The aim of this study is to develop a novel Arg‐Gly‐Asp acid (RGD) and Ala‐Thr‐Trp‐Leu‐Pro‐Pro‐Arg (ATWLPPR A7R) peptide‐containing ligand for ¹⁸F labeling as αvβ3 and vascular endothelial growth factor receptor‐targeted imaging agent. ¹⁸F‐RGD‐A7R was prepared by conjugation with ¹⁸F‐SFB. The final product was purified by high‐performance liquid chromatography and tested in vitro and in vivo. Cell‐binding assays of RGD‐A7R, RGD and RGD‐A7R, A7R were tested in U87MG cells (¹²⁵I‐RGDyK and ¹²⁵I‐A7RY as radioligand, respectively). Preliminary biodistribution of the ¹⁸F‐RGD‐A7R was also evaluated. The RGD‐A7R had good integrin binding affinity (50% inhibitory concentration (IC₅₀) = 21.67 and 23.68 nM, slightly lower than unmodified RGD (40.02 nM) and A7R (50.18 nM)). The radiotracer had receptor‐mediated activity accumulation in U87MG tumor (1.90 ± 0.34 percentage of injected dose per gram (%ID/g) at 0.5 h postinjection), which is known to be integrin positive. After blocking with RGD‐A7R, the tumor uptake was reduced to 0.47 ± 0.06 %ID/g at 0.5 h postinjection. ¹⁸F‐RGD‐A7R exhibited dual receptor targeting properties both in vitro and in vivo. The favorable characterizations of ¹⁸F‐RGD‐A7RY, such as convenient synthesis, high specific activity, and high tumor uptake, warrant its further investigation for clinical cancer imaging.     

99mTc‐HYNIC‐(Ser)3‐J18 peptide: A radiotracer for non‐small‐cell lung cancer targeting

Radiolabeled peptide could be a useful tool for the diagnosis of non‐small‐cell lung cancer (NSCLC). In this study, HYNIC‐(Ser)₃‐J18 peptide was labeled with ^99mTc using EDDA/tricine as coligands. The in vitro and in vivo studies of this radiolabeled peptide were performed for cellular‐specific binding and tumor targeting in A‐549 cells and tumor‐bearing mice, respectively. The high radiochemical purity was obtained and this radiolabeled peptide exhibited high stability in buffer and serum. The radiolabeled peptide showed high affinity for the A‐549 cells with a dissociation constant value (K_D) of 4.4 ± 0.8 nm . The tumor–muscles ratios were 2.7 and 4.4 at 1 and 2 hr after injection of ^99mTc‐(EDDA/tricine)‐HYNIC‐(Ser)₃‐J18 in tumor‐bearing mice. The tumor uptake was decreased after preinjection with non‐labeled peptide for this radiolabeled peptide in blocking experiment. The results of this study showed the ^99mTc‐(EDDA/tricine)‐(Ser)₃‐HYNIC‐J18 peptide might be a promising radiolabeled peptide for NSCLC targeting.     

Biodistribution of 99mTc‐sunitinib as a potential radiotracer for tumor hypoxia imaging

Tyrosine kinases are groups of enzymes, which are over‐expressed in solid tumor cells, representing good targets for different drugs such as sunitinib (N‐[2‐(diethylamino)ethyl]‐5‐{[(3Z)‐5‐fluoro‐2‐oxo‐2,3‐dihydro‐1H‐indol‐3‐ylidene]methyl}‐2,4‐dimethyl‐1H‐pyrrole‐3‐carboxamide). The aim of this work was to design and synthesize ^99mTc‐sunitinib radiotracer and to study its tumor binding specificity as a novel selective radiopharmaceutical for tumor hypoxia imaging. The in vivo biodistribution of ^99mTc‐sunitinib in tumor bearing mice showed high target/non‐target (T/NT) ratio (T/NT ~ 3 at 60 min post injection). This preclinical high biological accumulation in tumor cells suggests that ^99mTc‐sunitinib is ready to go through the clinical trials as a potential selective radiotracer able to image tumor hypoxia.     

Kit with technetium‐99m labelled antimicrobial peptide UBI 29‐41 for specific infection detection

The purpose of this study was to investigate radiochemical and biological characteristics of an instant kit for the preparation of ^99mTc‐labelled UBI 29‐41 for specific detection of infections. The kit is based on ^99mTc‐labelling via HYNIC conjugated to the terminal amine of the peptide, producing a well‐understood labelled compound. One hour after the addition of fresh ^99mTcO to the kit ITLC and HPLC reverse‐phase analysis was performed. Stability of the labelled complex was challenged and the binding to bacterial pellets was assessed. Finally, the biodistribution and accumulation in MRSA‐infected tissues were studied using scintigraphy and ex vivo countings. Data were compared to a non‐kit control method. Radiochemical analysis indicated >96% labelling, stability for 24 h and the preparation was used without purification. In vitro studies showed 41% of radioactivity was bound to bacteria. After injection into mice with a bacterial infection the site of infection was visualized within 30 min. Kit prepared ^99mTc‐HYNIC‐UBI 29‐41 was rapidly (half‐life 113 min) cleared via the kidneys and urinary bladder, essentially slower than control peptide (half‐life 74 min). This slower clearance results in higher activities in blood and other tissues. Nevertheless, ^99mTc‐HYNIC‐UBI 29‐41 shows favourable radiochemical characteristics and deserves further evaluation in a clinical setting. Copyright © 2005 John Wiley & Sons, Ltd.     

Development of a single vial kit formulation of [99mTc]‐labeled doxorubicin for tumor imaging and treatment response assessment‐preclinical evaluation and preliminary human results

The present study describes the successful radiolabeling of [99mTcO^?₄] with doxorubicin, and the resultant product was formulated in to a ready‐to‐label lyophilized single vial kit preparation for convenient use in a routine clinical setting. The radiolabeled preparation of [99mTc]‐doxorubicin exhibited a high radiolabeling efficiency of more than 95.0%, serum stability for up to 24 h, and shelf‐life of lyophilized cold kits was more than 6 months. Animal imaging data in tumor‐bearing mice demonstrated that [99mTc]‐doxorubicin accumulated in the tumor site with high target (tumor) to non‐target (contra‐lateral thigh) ratio (3.2 ± 0.5). The ratio decreased to 1.2 ± 0.6 indicating a good response on follow up imaging performed after 2 weeks of doxorubicin treatment. [99mTc]‐doxorubicin scintigraphic data in human volunteers supported the hepato‐renal excretion of the radiotracer as reflected by the increased accumulation of the radiotracer as a function of time in intestine, kidneys, and urinary bladder. Further, imaging in patients (very limited number) indicated that the technique may be useful in the detection of active sarcoma and post treatment (surgery/chemotherapy) remission or absence of the disease. The technique, however, needs validation through further preclinical evaluation and imaging in a larger number of patients.     

On the 99mTc‐labeling of isoniazid with different 99mTc cores

Recent advances in the chemistry of radiolabeling with ^99mTc such as use of the ^99mTc tricarbonyl and ^99mTc–HYNIC cores have revived interest in ^99mTc‐labeling of small biomolecules and further investigation as potential radiopharmaceuticals. Isoniazid, a drug commonly used for treatment of tuberculosis, has been chosen for the present study. Three distinct strategies were utilized to radiolabel isoniazid with ^99mTc. In the direct labeling protocol, the hydrazino amide functional group of isoniazid was used for ^99mTc‐labeling in the HYNIC sense using tricine and EDDA as co‐ligands. The other strategies of ^99mTc‐labeling involved the derivatization of isoniazid to its N, N‐diacetic acid derivative, which in turn was either used as a tridentate ligand for labeling with the [^99mTc(CO)₃(H₂O)₃]⁺ synthon or directly labeled by the conventional route wherein ^99mTc is in the +3 oxidation state. The complexes prepared in >95% yields were characterized by paper chromatography, thin layer chromatography and HPLC. Comparison of the three approaches showed that maximum specific activity and stability was obtained in the ^99mTc–isoniazid derivative synthesized via the tricarbonyl method. However, in vitro cell‐binding studies indicated that none of the ^99mTc–isoniazid complexes prepared had any appreciable uptake in Mycobacterium tuberculosis. Copyright © 2005 John Wiley & Sons, Ltd.     

Development of kit formulations for 99mTcN‐MPO: a cationic radiotracer for myocardial perfusion imaging

The objective of this study was to develop a kit formulation for [^99mTcN(mpo)(PNP5)]⁺ (MPO = 2‐mercaptopyridine oxide), (^99mTcN‐MPO) to support its clinical evaluations as a SPECT radiotracer. Radiolabeling studies were performed using three different formulations (two‐vial formulation and single‐vial formulations with/without SnCl₂) to explore the factors influencing radiochemical purity (RCP) of ^99mTcN‐MPO. We found that the most important factor affecting the RCP of ^99mTcN‐MPO was the purity of PNP5. ^99mTcN‐MPO was prepared >98% RCP (n = 20) using the two‐vial formulation. For single‐vial formulations with/without SnCl₂, β‐cyclodextrin (β‐CD) is particularly useful as a stabilizer for PNP5. The RCP of ^99mTcN‐MPO was 95–98% using β‐CD, but its RCP was only 90–93% with γ‐cyclodextrin (γ‐CD). It seems that PNP5 fits better into the inner cavity of β‐CD, which forms more stable inclusion complex than γ‐CD in the single‐vial formulations. The results from biodistribution and imaging studies in Sprague–Dawley rats clearly demonstrated biological equivalence of three different formulations. Single photon‐emission computed tomography data suggested that high quality images could be obtained at 0–30‐min post‐injection without significant interference from the liver radioactivity. Considering the ease for ^99mTc‐labeling and high RCP of ^99mTcN‐MPO, the non‐SnCl₂ single‐vial formulation is an attractive choice for future clinical studies.     

99mTc‐labeled NGR‐chlorambucil conjugate, 99mTc‐HYNIC‐CLB‐c(NGR) for targeted chemotherapy and molecular imaging

Targeted delivery of chemotherapeutic drug at the tumor site enhances the efficacy with minimum systemic exposure. Towards this, drugs conjugated with peptides having affinity towards a particular molecular target are recognized as affective agents for targeted chemotherapy. Thus, in the present study, tumor‐homing asparagine‐glycine‐arginine (NGR) peptide ligand was conjugated to DNA alkylating nitrogen mustard, chlorambucil (CLB). The peptide‐drug conjugate (PDC), CLB‐c(NGR), was radiolabeled with ^99mTc‐HYNIC core to trace its pharmacokinetics and biodistribution pattern. In vitro cell‐binding studies of ^99mTc‐HYNIC‐CLB‐c(NGR) were conducted in murine melanoma B16F10 cells. The cytotoxicity studies conducted by incubation of the peptide/drug/PDC with B16F10 cells demonstrated enhanced cytotoxic effect of PDC in comparison to either the peptide or the drug alone. In vivo biodistribution studies in C57BL6 mice bearing melanoma tumor showed maximum tumor uptake at 30 minutes pi (2.45 ± 0.28% ID/g), which reduced to 0.77 ± 0.1% ID /g at 3 hours pi. The radiotracer being hydrophilic cleared rapidly from the heart, lungs, liver, and muscle. The tumor‐to‐blood and tumor‐to‐muscle ratios improved with time. This study opens avenues for conjugation of other targeting peptides with the drug CLB for enhanced toxicity at the diseased site.     

Evaluation of a 188Re‐radiolabeled Arg‐Gly‐Asp peptide for tumor overexpressed αvβ3 receptors

To develop a peptide‐based radiopharmaceutical for the therapy of α_vβ₃ receptors overexpressed tumors, we have prepared a novel Arg‐Gly‐Asp (RGD) peptide (HCRGDCF(D)CRGDC, P12) radiolabeled with ¹⁸⁸Re. With His acid at the end of the peptide containing RGD, the label efficiency was more than 95% within 30 min. The peptide binds to human glioblastoma U87MG cells with high affinity [IC₅₀ = 86.3 nm]. The stability of ¹⁸⁸Re‐P12 in vitro was also investigated. More than 80% of radioactivity was kept in the peptide after 4 h incubation in phosphate buffer solution (pH = 7.4) or calf serum under physiological conditions. Biodistribution of this radiocompound was carried out in mice bearing S180 tumor. Fast clearance of ¹⁸⁸Re‐peptide from blood and specific uptakes by tumors realized higher tumor‐to‐blood ratio (1.80) 4 h post‐injection. Obvious difference was observed between the blocking and unblocking experiments in whole body autoradiography imaging. These results have demonstrated the potential of ¹⁸⁸Re‐labeling RGD as a radiotherapeutic agent.     

Synthesis and evaluation of a 99mTc‐labeled tubulin‐binding agent for tumor imaging

Cholchicine and its derivatives are very potent tubulin‐binding compounds and can be used as a potential tumor targeting agents. In this study, colchicine was labeled with ^99mTc via hydrazinonicotinic acid (HYNIC) and was investigated further. HYNIC/cholchicine was synthesized and labeling with ^99mTc was performed at 95 °C for 15 min and radiochemical analysis included HPLC method. The stability of radiconjugate was checked in the presence of human serum at 37 °C up to 24 h. Biodistribution was studied in breast tumor‐bearing mice. Labeling yield of 95.8 ± 0.54% was obtained corresponding to a specific activity of 54 MBq/µmol. Radioconjugate showed good stability in the presence of human serum. Biodistribution studies in tumor‐bearing mice showed that ^99mTc/HYNIC/colchicine conjugate accumulated in tumor with good uptake (3.17 ± 0.14% g/g at 1 h post‐injection). The radioconjugate was cleared fast from normal organs and showed clearance through urinary and hepatobiliary systems with accumulation of activity in kidneys and intestine. This radioconjugate may be useful to assess the presence of tumor by imaging.     

Synthesis and comparative in vivo evaluation of 99mTc(CO)3‐labeled PEGylated and non‐PEGylated cRGDfK peptide monomers

This work aimed at studying the effect of insertion of medium PEG (PEG₇) on the pharmacokinetic behavior of cRGDfK peptide in comparison with the non‐PEGylated analogue. The cRGDfK peptide has thus been derivatized at ε‐amino group of lysine by conjugation with N₃–PEG₇–COOH/N₃–CH₂–COOH to prepare a PEGylated and a non‐PEGylated analogue of cRGDfK. A tridentate chelator was then incorporated by click chemistry conjugation of the two peptide azides for radiolabeling with [^99mTc(CO)₃(H₂O)₃]⁺ precursor. Comparative in vivo evaluation of the two ^99mTc(CO)₃‐labeled radiotracers, ^99mTc(CO)₃–Pra–Tz–CH₂–cRGDfK 5 and ^99mTc(CO)₃–Pra–Tz–PEG₇–cRGDfK 6 , was carried out in C57BL/6 mice bearing α_vβ₃‐positive melanoma tumors to determine their potential toward targeting integrin α_vβ₃ receptors. The radiotracers exhibited excellent stability in saline as well as in serum. Maximum tumor uptake for the two radiotracers was observed at 30 min p.i. ( 5 : 3.0 ± 0.7% ID/g; 6 : 4.1 ± 0.5% ID/g). The two neutral ^99mTc(CO)₃ radiotracers prepared exhibited receptor‐mediated uptake in melanoma tumor. The increase in the tumor uptake on introduction of PEG₇ unit was accompanied by slower clearance from other organs which resulted in decreased target‐to‐background ratios. The in vivo kinetics of ^99mTc(CO)₃‐labeled radiotracer, ^99mTc(CO)₃–Pra–Tz–CH₂–cRGDfK 5 with only methylene unit as the spacer, was found to be more favorable due to higher tumor/blood, tumor/liver, tumor/kidney, and tumor/lung ratios.     

Synthesis and biological evaluation of 99mTc(CO)3(IDA‐CPT) for tumor imaging

This work reports the synthesis, radiolabeling, and preliminary biodistribution results in tumor‐bearing mice of ^99mTc(CO)₃(IDA‐CPT). The novel camptothecin (CPT) derivate was successfully synthesized by conjugation of iminodiacetic acid (IDA) to camptothecin via a short carbonyl‐methylene linker. Radiolabeling was performed in high yield with [^99mTc(CO)₃]⁺ core to get ^99mTc(CO)₃(IDA‐CPT), which was hydrophilic and stable at room temperature. Biodistribution studies in tumor‐bearing mice showed that ^99mTc(CO)₃(IDA‐CPT) accumulated in the tumor with good uptake and retention. However, its clearance from normal organs was slow, resulting in poor T/NT ratios. Further modification on the linker or/and ^99mTc‐chelate to improve the tumor‐targeting efficacy and in vivo kinetic profiles is currently in progress. Copyright © 2009 John Wiley & Sons, Ltd.     

Initial evaluation of 99mTc‐tricarbonyl‐cyclopentadienyl fatty acids derivatives as SPECT tracers for myocardium

Fatty acids are myocardial metabolic agent for detecting myocardial ischemia and infraction. However, no ^99mTc‐labeled fatty acids had potential use in clinical practice. In this study, ^99mTc‐CpTT‐10‐oxo‐para‐PPA ( 1d ), ^99mTc‐CpTT‐11‐oxo‐para‐PPA ( 2d ), ^99mTc‐CpTT‐12‐oxo‐para‐PPA ( 3d ), ^99mTc‐CpTT‐11‐oxo‐ortho‐PPA ( 4d ), and ^99mTc‐CpTT‐11‐oxo‐meta‐PPA ( 5d ) were synthesized by a double ligand transfer reaction, and their biological behaviors were investigated. Compound 2d achieved good heart to blood ratio (3.39 at 5 min after intravenous), and 2d showed high‐heart uptake of 6.20% ID/g at 5 minutes after injection. Compound 3d displayed a prolonged retention in the myocardium (1.43% ID/g at 60 min after injection). Radioactivity accumulation in the lungs, spleen, and blood was eliminated rapidly. In vivo, metabolite analysis presented that compound 6d may be metabolite of 2d through β‐oxidation in tissue. Unfortunately, the biodistribution studies of 1d , 2d , 3d , 4d , and 5d showed fast heart clearance and poor heart to liver ratios, which suggested that the 5 ^99mTc‐labeled fatty acid analogues cannot be used for diagnosis.     

Synthesis,radiolabeling and in vitro and in vivo characterization of a technetium‐99m‐labeled alpha‐M2 peptide as a tumor imaging agent

Abstract: In an effort to develop a peptide‐based radiopharmaceutical for the detection of breast cancer, we have prepared an analog of αM2 peptide, modified to incorporate an N₃S chelate system. Mercaptoacetyltriglycine (MAG)₃‐derivatized αM2 peptide was prepared by solid‐phase synthesis and radiolabeled with ^99mTc by an exchange method. In vitro cell‐binding on human breast cancer cell lines, MDA‐MB‐231 and MCF‐7, indicated the affinity and specificity of ^99mTc‐MAG₃‐αM2 toward breast cancer cells. Additionally, the radiolabeled peptide showed rapid internalization into human breast cancer cells. In vivo biodistribution in mice showed that the radiolabeled peptide cleared rapidly from the blood and most non‐target tissues and was excreted significantly via the kidneys. Uptake of ^99mTc‐MAG₃‐αM2 in the tumor was moderate. The radiochemical and in vitro and in vivo characterization indicates that the radiolabeled peptide has certain favorable properties and it might be a useful radiopharmaceutical for the detection of breast cancer in vivo.     

99mTc‐tricarbonyl labeling of paclitaxel as an antimicrotubule agent and its evaluation in B16‐F10 melanoma tumor‐bearing mice

In the present study paclitaxel (taxol) was labeled with [^99mTc(CO)₃(H₂O)₃]⁺ core. Labeling was optimized, and radiochemical analysis was determined by thin layer chromatography and high performance liquid chromatography. Radiocomplex was evaluated and verified further as a tumor characterization agent in B16‐F10 melanoma tumor‐bearing mice. The [^99mTc(CO)₃(H₂O)₃]⁺‐paclitaxel complex with high specific activity (0.77 GBq/μmol) and labeling yield (96.8 ± 1.3) was obtained. No decrease in labeling was observed up to 6 hours, and the stability of the radiocomplex was found adequate. Our main achievement was high accumulation of radiolabeled paclitaxel in tumor (4.51 ± 0.65 percentage injected dose per gram [%ID/g] at 2‐h postinjection) followed by significant reduction (1.86 ± 0.27%ID/g) at 4‐hour postinjection. Because paclitaxel is a substrate for multidrug resistance, ^99mTc‐tricarbonyl‐paclitaxel imaging would be useful for tumor characterization rather than tumor detection.     

99mTc‐DTPA‐Amino Acids Conjugate as Specific SPECT Pharmaceuticals for Tumor Imaging

^99mTc‐Diethylene triamine pentaacetic acid‐bis (amide) conjugates have been synthesized and evaluated as a potential radiopharmaceutical for tumor imaging. The compounds were synthesized by the condensation reaction of DTPA bis(anhydride) with different l ‐amino acids (methyl tryptophan, and 5‐hydroxy tryptophan) and were characterized on the basis of IR, NMR, and Mass spectroscopy. ^99mTc‐labeled compounds were found stable for about 24 h under physiological conditions with more than 95% radiolabeling yield. Blood kinetic studies of all these complexes showed a bi‐exponential pattern as well as quick wash out from the blood circulation. The biological t_1/2(F) and t_1/2(S) were found to be 20 ± 0.001 min for DTPA‐(Me‐Trp)₂ and 18 ± 0.001 min for DTPA‐(5HT)₂ and t_1/2 (slow) 5 h 45 min ± 0.001, 5 h 30 ± 0.001 min for DTPA‐(Me‐Trp)₂, and DTPA‐(5HT)₂, respectively. Imaging and biodistribution studies were performed in mice bearing Ehrlich ascites tumor (EAT) tumors in right thigh. Radioconjugate derived from l ‐5‐hydroxytryptophan exhibited remarkable localization at tumor site; whereas radiotracer derived from l ‐methyl tryptophan shows relatively less accumulation at the tumor site. Tumor‐to‐muscles ratios were 5.07 ± 0.001, and 4.2 ± 0.001 at 1 and 4 h for ^99mTc‐DTPA‐(Me trp)₂ and 4.97 ± 0.001 and 5.8 ± 0.001 at 1 and 4 h after postinjection for ^99mTc‐DTPA‐(5HT)₂, respectively. The preliminary results with these amino acid based ligands are encouraging to carrying out further in vivo experiments for targeted tumor imaging.