Labeling Yield Research Articles

Trastuzumab cotreatment improves survival of mice with PC-3 prostate cancer xenografts treated with the GRPR antagonist 177 Lu-DOTAGA-PEG2 -RM26.

Gastrin‐releasing peptide receptors (GRPRs) are overexpressed in prostate cancer and are suitable for targeted radionuclide therapy (TRT). We optimized the bombesin‐derived GRPR‐antagonist PEG2‐RM26 for labeling with 177Lu and further determined the effect of treatment with 177Lu‐labeled peptide alone or in combination with the anti‐HER2 antibody trastuzumab in a murine model. The PEG2‐RM26 analog was coupled to NOTA, NODAGA, DOTA and DOTAGA chelators. The peptide‐chelator conjugates were labeled with 177Lu and characterized in vitro and in vivo. A preclinical therapeutic study was performed in PC‐3 xenografted mice. Mice were treated with intravenous injections (6 cycles) of (A) PBS, (B) DOTAGA‐PEG2‐RM26, (C) 177Lu‐DOTAGA‐PEG2‐RM26, (D) trastuzumab or (E) 177Lu‐DOTAGA‐PEG2‐RM26 in combination with trastuzumab. 177Lu‐DOTAGA‐PEG2‐RM26 demonstrated quantitative labeling yield at high molar activity (450 GBq/μmol), high in vivo stability (5 min pi >98% of radioligand remained when coinjected with phosphoramidon), high affinity to GRPR (K D = 0.4 ± 0.2 nM), and favorable biodistribution (1 hr pi tumor uptake was higher than in healthy tissues, including the kidneys). Therapy with 177Lu‐DOTAGA‐PEG2‐RM26 induced a significant inhibition of tumor growth. The median survival for control groups was significantly shorter than for treated groups (Group C 66 days, Group E 74 days). Trastuzumab together with radionuclide therapy significantly improved survival. No treatment‐related toxicity was observed. In conclusion, based on in vitro and in vivo characterization of the four 177Lu‐labeled PEG2‐RM26 analogs, we concluded that 177Lu‐DOTAGA‐PEG2‐RM26 was the most promising analog for TRT. Radiotherapy using 177Lu‐DOTAGA‐PEG2‐RM26 effectively inhibited tumor growth in vivo in a murine prostate cancer model. Anti‐HER2 therapy additionally improved survival.

Quantitative Single-Residue Bioorthogonal Labeling of G Protein-Coupled Receptors in Live Cells.

High-end microscopy studies of G protein-coupled receptors (GPCRs) require installing onto the receptors bright and photostable dyes. Labeling must occur in quantitative yields, to allow stoichiometric data analysis, and in a minimally invasive fashion, to avoid perturbing GPCR function. We demonstrate here that the genetic incorporation of trans-cyclooct-2-ene lysine (TCO*) allows achieving quantitative single-residue labeling of the extracellular loops of the β2-adrenergic and the muscarinic M2 class A GPCRs, as well as of the corticotropin releasing factor class B GPCR. Labeling occurs within a few minutes by reaction with dye-tetrazine conjugates on the surface of live cells and preserves the functionality of the receptors. To precisely quantify the labeling yields, we devise a method based on fluorescence fluctuation microscopy that extracts the number of labeling sites at the single-cell level. Further, we show that single-residue labeling is better suited for studies of GPCR diffusion than fluorescent-protein tags, since the latter can affect the mobility of the receptor. Finally, by performing dual-color competitive labeling on a single TCO* site, we devise a method to estimate the oligomerization state of a GPCR without the need for a biological monomeric reference, which facilitates the application of fluorescence methods to oligomerization studies. As TCO* and the dye-tetrazines used in this study are commercially available and the described microscopy techniques can be performed on a commercial microscope, we expect our approach to be widely applicable to fluorescence microscopy studies of membrane proteins in general.

Trastuzumab-Modified Gold Nanoparticles Labeled with 211At as a Prospective Tool for Local Treatment of HER2-Positive Breast Cancer.

Highly localized radiotherapy with radionuclides is a commonly used treatment modality for patients with unresectable solid tumors. Herein, we propose a novel α-nanobrachytherapy approach for selective therapy of human epidermal growth factor receptor 2 (HER2)-positive breast cancer. This uses local intratumoral injection of 5-nm-diameter gold nanoparticles (AuNPs) labeled with an α-emitter (211At), modified with polyethylene glycol (PEG) chains and attached to HER2-specific monoclonal antibody (trastuzumab). The size, shape, morphology, and zeta potential of the 5 nm synthesized AuNPs were characterized by TEM (Transmission Electron Microscopy) and DLS (Dynamic Light Scattering) techniques. The gold nanoparticle surface was modified by PEG and subsequently used for antibody immobilization. Utilizing the high affinity of gold for heavy halogens, the bioconjugate was labelled with 211At obtained by α irradiation of the bismuth target. The labeling yield of 211At was greater than 99%. 211At bioconjugates were stable in human serum. Additionally, in vitro biological studies indicated that 211At-AuNP-PEG-trastuzumab exhibited higher affinity and cytotoxicity towards the HER2-overexpressing human ovarian SKOV-3 cell line than unmodified nanoparticles. Confocal and dark field microscopy studies revealed that 211At-AuNP-PEG-trastuzumab was effectively internalized and deposited near the nucleus. These findings show promising potential for the 211At-AuNP-PEG-trastuzumab radiobioconjugate as a perspective therapeutic agent in the treatment of unresectable solid cancers expressing HER2 receptors.

Radiosynthesis, radiochemical, and biological characterization of 177 Lu-labeled diethylenetriamine penta-acetic acid.

Diethylenetriamine penta-acetic acid (DTPA), when complexed with a gamma (γ)-emitter radioisotope like 99m Tc, is used for renal function diagnosis and many other diagnostic applications. The main aim of this study was to develop a novel and versatile single-step methodology for the synthesis of a new 177 Lu-labeled radiopharmaceutical with high radiochemical yield, which can be used for diagnostic purposes and therapeutic purposes also. The single and well-defined 177 Lu-DTPA complex was radiochemically characterized by paper chromatography, thin-layer chromatography, high-performance liquid chromatography, and electrophoresis techniques. Dependence of the labeling yield of 177 Lu-DTPA complex on different factors was studied in detail. Biological evaluation was also performed in a normal rabbit by developing images under a γ camera at various time intervals. More than 99% labeling yield was obtained by reacting DTPA with 177 Lu at specific conditions (pH 7.0, 15 minutes reaction time at 100 °C). 177 Lu-DTPA complex showed high stability both at room temperature and in vitro. Biodistribution studies in normal mice indicated the fractional renal uptake of intravenously administered 177 Lu-DTPA complex, which reached in the kidneys within 2-3 minutes. Scintigraphy showed rapid clearance from the body. Based on these results, we propose that 177 Lu-DTPA complex might be used as an ideal candidate for functional evaluation of kidneys and the urinary tract, especially when needed to be transported to long-range consumer sites, because of its suitable half-life.

Synthesis of a Potential Tumor Imaging Agent by Oxidative Radioiodination of Aspirin and Its Preclinical Study

Aspirin, one of nonsteroidal anti-inflammatory analgesic drugs, was labeled with 125I with a labeling yield of 85.5% under the following conditions: pH 9, 100 mg of the substrate, 50 µg of Chloramine-T, 5–15 min, room temperature. The radiochemical yield of 125I-aspirin was determined by paper chromatography and electrophoresis, and the radiochemical purity was determined by HPLC.125I-aspirin was stable for at least 3 h. Biodistribution of 125I-aspirin in normal and tumor-bearing mice was studied. The uptake of 125I-aspirin in tumor sites makes it promising as a new radiopharmaceutical for tumor imaging.

Synthesis, characterization and evaluation of a Cu-labeled macrocyclic-porphyrin as a potential chelator for 64Cu-based radiopharmaceuticals

Synthesis of 5-(4-aminophenyl)-10,15,20-triphenyl porphyrin (B) was performed using the Lindsey method and characterized by 1H NMR. Cu-porphyrin complexation conditions were tested using natCu and UV–Vis analysis confirmed metallation. High specific activity 64Cu was produced via cyclotron using the 64Ni(p,n) reaction. For radiolabeling, 64CuCl2 dissolved in acetate buffer (pH 5.6) was mixed with compound (B) in DMSO, and incubated for 60 min at 37 °C, obtaining a labeling yield > 95% at 60 min. The labeled complex [64Cu]-5-(4-aminophenyl)-10,15,20-triphenyl porphyrin (C) showed good stability (> 95%) for up 48 h when incubated in human serum. MicroPET images were acquired in rats and mice using 64Cu-porphyrin (C) and 64CuCl2 in order to compare their biodistribution. Results suggest that (C) is an excellent candidate to prepare stable metal complexes suitable for the development of novel 64Cu-based radiopharmaceuticals using biological moieties to target specific receptors.

Intranasal drug delivery of iodo-haloperidol as a radiopharmaceutical brain imaging agent

Haloperidol (HP) is a dopamine blocking agent. HP was radiolabeled with 125I using direct electrophilic substitution. Different HP formulations such as intravenous solution (VS), intranasal solution (NS) and intranasal microemulsion (NME) have been prepared. Optimum labeling yield of 125I-HP was obtained 93.42 ± 1.53%. Isopropylmyristate oil (10%), Tween 80 (30%), propylene glycol (15%) and aqueous phase (45%) were used for NME preparation. Using Swiss Albino mice, the % brain uptake were 1.4 ± 0.2, 4.3 ± 0.2 and 5.7 ± 0.4% for VS, NS and NME, respectively. 125I-HP in NME formula showed superiority as drug delivery system for brain imaging.

Application of 188Re-labeled BaGdF5-PEG nanoparticles on the growth inhibition of hepatoma cells and the SPECT imaging in rabbit models

Objective To investigate the inhibitory effect of 188Re-labeled BaGdF5-poly(ethylene glycol)(PEG) nanoparticles (NPs) on hepatoma cells, and explore the application of the radiolabeled NPs for SPECT imaging. Methods BaGdF5-PEG NPs were synthesized by hydrothermal method, and were further radiolabeled with 188Re using diethylene triamine pentaacetic acid (DTPA) as a coupling agent. The human hepatoma cells SMCC 7721 were treated with different concentrations of BaGdF5-PEG NPs, 188ReO4- or 188Re-DTPA-BaGdF5 NPs (14.8, 74.0, 370.0×104 Bq/ml) for 24 h, and then the cell proliferation rates were measured by 3-(4, 5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2-H-tetrazolium bromide (MTT) assay. 188ReO4- and 188Re-DTPA-BaGdF5 NPs were administrated into normal rabbits via the ear vein, respectively. For the former, static SPECT/CT imaging were performed at 30, 60 min post-injection, and for the latter, dynamic SPECT images were captured within 10 min, and static SPECT/CT images at 30, 60, 120 min post-injection. The rabbit VX2 tumor model was established, and a microcatheter was inserted into hepatic artery via the rabbit femoral artery, and then the mixture of 188Re-DTPA-BaGdF5 NPs and lipiodol was injected into the tumor region. SPECT/CT imaging for VX2 tumor was performed at 30 min later. Data were analyzed by two-sample t test. Results The BaGdF5-PEG NPs were nearly square and the particle size was about 10 nm. The labeling yield of 188Re-DTPA-BaGdF5 was 94.1% at the optimum conditions. Moreover, it showed high stability in vitro and in vivo. In vitro, BaGdF5-PEG NPs did not exhibit obvious cytotoxicity even at a high concentration. Both 188ReO4- and 188Re-DTPA-BaGdF5 could inhibit the proliferation of SMCC 7721 cells, but 188Re-DTPA-BaGdF5 showed a significantly stronger inhibitory effect at the doses of 74.0 and 370.0×104 Bq/ml (t values: 4.21, 4.09, both P<0.01). In vivo,188ReO4- was absorbed by maxillary glands and was quickly eliminated from blood via the kidneys. The 188Re-DTPA-BaGdF5 NPs mainly accumulated in the liver and spleen. In addition, retention and accumulation of 188Re-DTPA-BaGdF5 NPs in the liver tumor could be achieved by using transarterial intervention technique for drug delivery. Conclusion 188Re-DTPA-BaGdF5 NPs have certain killing effects on hepatoma cells in vitro, and with the help of transarterial intervention technique, the NPs can be aggregated within liver tumor, where they not only can be used for SPECT imaging, but also have potential therapeutic effects. Key words: Liver neoplasms; Tumor cells, cultured; Nanoparticles; Rhenium; Tomography, emission-computed, single-photon; Tomography, X-ray computed; Rabbits

Sorption of 223Ra and 211Pb on modified nanodiamonds for potential application in radiotherapy

223Ra and 211Pb/211Bi radionuclides with radiochemical purity were obtained from 227Ac/223Ra generator on DGA resin. Covalent grafting of organic complexons (ethylenediaminetetraacetic acid—EDTA and derivatives of amino acids—L1 and L2 to aminated nanodiamonds (ND) was performed by carbodiimide method. Adsorption behavior of Ra and Pb in plasma volume expander on modified nanodiamonds was studied in detail. Effective adsorption have been found for 211Pb in physiologically acceptable conditions. Sorption behavior of generator radionuclides 223Ra and 211Pb on the surface of ND-L1 and ND-L2 in aqueous solution (pH = 6.3) was studied. It was shown that the labeling yield of 211Pb from the solution was about 94%. The sorption of 223Ra under the same conditions was weak. The sorption/desorption kinetics of 211Pb on the surface of ND-EDTA in the plasma volume expander was studied. The desorption rate of 211Pb did not exceed 2.5% within 60 min (pH 7.2–7.4).

An Efficient Method for Labeling Single Domain Antibody Fragments with 18F Using Tetrazine- Trans-Cyclooctene Ligation and a Renal Brush Border Enzyme-Cleavable Linker.

Single domain antibody fragments (sdAbs) labeled with 18F have shown promise for assessing the status of oncological targets such as the human epidermal growth factor receptor 2 (HER2) by positron emission tomography (PET). Earlier, we evaluated two residualizing prosthetic agents for 18F-labeling of anti-HER2 sdAbs; however, these methods resulted in poor labeling yields and high uptake of 18F activity in the kidneys. To potentially mitigate these limitations, we have now developed an 18F labeling method that utilizes the trans-cyclooctene (TCO)-tetrazine (Tz)-based inverse-electron demand Diels-Alder reaction (IEDDAR) in tandem with a renal brush border enzyme-cleavable glycine-lysine (GK) linker in the prosthetic moiety. The HER2-targeted sdAb 2Rs15d was derivatized with TCO-GK-PEG4-NHS or TCO-PEG4-NHS, which lacks the cleavable linker. As an additional control, the non HER2-specific sdAb R3B23 was derivatized with TCO-GK-PEG4-NHS. The resultant sdAb conjugates were labeled with 18F by IEDDAR using [18F]AlF-NOTA-PEG4-methyltetrazine. As a positive control, the 2Rs15d sdAb was radioiodinated using the well-characterized residualizing prosthetic agent, N-succinimidyl 4-guanidinomethyl-3-[125I]iodobenzoate ([125I]SGMIB). Synthesis of [18F]AlF-NOTA-Tz-TCO-GK-2Rs15d was achieved with an overall radiochemical yield (RCY) of 17.8 ± 1.5% ( n = 5) in 90 min, a significant improvement over prior methods (3-4% in 2-3 h). In vitro assays indicated that [18F]AlF-NOTA-Tz-TCO-GK-2Rs15d bound with high affinity and immunoreactivity to HER2. In normal mice, when normalized to coinjected [125I]SGMIB-2Rs15d, the kidney uptake of [18F]AlF-NOTA-Tz-TCO-GK-2Rs15d was 15- and 28-fold lower ( P < 0.001) than that seen for the noncleavable control ([18F]AlF-NOTA-Tz-TCO-2Rs15d) at 1 and 3 h, respectively. Uptake of [18F]AlF-NOTA-Tz-TCO-GK-2Rs15d in HER2-expressing SKOV-3 ovarian carcinoma xenografts implanted in athymic mice was about 80% of that seen for coinjected [125I]SGMIB-2Rs15d. On the other hand, kidney uptake was 5-6-fold lower, and as a result, tumor-to-kidney ratios were 4-fold higher for [18F]AlF-NOTA-Tz-TCO-GK-2Rs15d than those for [125I]SGMIB-2Rs15d. SKOV-3 xenografts were clearly delineated even at 1 h after administration of [18F]AlF-NOTA-Tz-TCO-GK-2Rs15d by Micro-PET/CT imaging with even higher contrast observed thereafter. In conclusion, this strategy warrants further evaluation for labeling small proteins such as sdAbs because it offers the benefits of good radiochemical yields and enhanced tumor-to-normal tissue ratios, particularly in the kidney.

Preclinical Evaluation of Radioiodinated Quinoxaline Derivative as a Possible Brain Imaging Agent

4-{(4-Methylphenyl)glycyl}-3,4-dihydroquinoxalin-2(1H)-one (QIN) was prepared from 4-(2-chloroacetyl)- 3,4-dihydroquinoxalin-1(1H)-one). 125I-QIN was prepared by electrophilic substitution using Chloramine- T as an oxidizing agent. The highest labeling yield, 94%, was obtained under the following conditions: pH 7, 15 min, 100 μg of QIN, 50 μg of Chloramine-T, and carrier-free Na125I. The labeled compound was stable for up to 12 h post labeling. A biodistribution study of 125I-QIN in mice showed that its brain uptake was about 5.1% at 30 min post injection and remained on this level up to 1 h. As compared to commercially used radiopharmaceuticals for brain imaging, 125I-QIN is more stable and shows higher brain uptake.

Technetium-99m radiolabeling and biological study of epirubicin for in vivo imaging of multi-drug-resistant Staphylococcus aureus infections via single photon emission computed tomography.

The development of functional imaging is a promising strategy for diagnosis and treatment of infectious and cancerous diseases. In this study, epirubicin was developed as a [99m Tc]-labeled radiopharmaceutical for the imaging of multi-drug-resistant Staphylococcus aureus infections. The labeling was carried out using sodium pertechnetate (Na99m TcO4 ; ~370MBq). The other parameters such as amount of ligand, reducing agent (SnCl2 .2H2 O), and pH were optimized. The highest labeling yield ≥96.98% was achieved when 0.3mg epirubicin, 13μg SnCl2 .2H2 O, and ~370MBq Na99m TcO4 were incubated at pH 7 for 15min in the presence of ascorbic acid at room temperature. Radiochemical purity, stability, charge, and glomerular filtration rate were studied to evaluate the biological compatibility for in vivo administration. Biodistribution investigations showed radiotracer uptake (13.89±1.56%ID/gm organ) by liver and 7.79±0.38%ID/gm organ by kidneys at 30min post-injection which promisingly wash out at 24hr post-injection. Scintigraphy study showed selective uptake in S.aureus-infected tissues in contrast to turpentine oil-induced inflamed tissues. Target-to-non-target ratio (6.7±0.05) was calculated at 1hr post-injection using SPECT gamma camera. The results of this study reveal that the [99m Tc]-epirubicin can be a choice of imaging and monitoring the treatment process of multi-drug resistant S.aureus bacterial infections.

Leveraging PET to image folate receptor \u03b1 therapy of an antibody-drug conjugate

BackgroundThe folate receptor α (FRα)-targeting antibody-drug conjugate (ADC), IMGN853, shows great antitumor activity against FRα-expressing tumors in vivo, but patient selection and consequently therapy outcome are based on immunohistochemistry. The aim of this study is to develop an antibody-derived immuno-PET imaging agent strategy for targeting FRα in ovarian cancer as a predictor of treatment success.MethodsWe developed [89Zr]Zr-DFO-M9346A, a humanized antibody-based radiotracer targeting tumor-associated FRα in the preclinical setting. [89Zr]Zr-DFO-M9346A’s binding ability was tested in an in vitro uptake assay using cell lines with varying FRα expression levels. The diagnostic potential of [89Zr]Zr-M9346A was evaluated in KB and OV90 subcutaneous xenografts. Following intravenous injection of [89Zr]Zr-DFO-M9346A (~90 μCi, 50 μg), PET imaging and biodistribution studies were performed. We determined the blood half-life of [89Zr]Zr-DFO-M9346A and compared it to the therapeutic, radioiodinated ADC [131I]-IMGN853. Finally, in vivo studies using IMG853 as a therapeutic, paired with [89Zr]Zr-DFO-M9346A as a companion diagnostic were performed using OV90 xenografts.ResultsDFO-M9346A was labeled with Zr-89 at 37 °C within 60 min and isolated in labeling yields of 85.7 ± 5.7%, radiochemical purities of 98.0 ± 0.7%, and specific activities of 3.08 ± 0.43 mCi/mg. We observed high specificity for binding FRα positive cells in vitro. For PET and biodistribution studies, [89Zr]Zr-M9346A displayed remarkable in vivo performance in terms of excellent tumor uptake for KB and OV xenografts (45.8 ± 29.0 %IA/g and 26.1 ± 7.2 %IA/g), with low non-target tissue uptake in other organs such as kidneys (4.5 ± 1.2 %IA/g and 4.3 ± 0.7 %IA/g). A direct comparison of the blood half life of [89Zr]Zr-M9346A and [131I]-IMGN853 corroborated the equivalency of the radiopharmaceutical and the ADC, paving the way for a companion PET imaging study.ConclusionsWe developed a new folate receptor-targeted 89Zr-labeled PET imaging agent with excellent pharmacokinetics in vivo. Good tumor uptake in subcutaneous KB and OV90 xenografts were obtained, and ADC therapy studies were performed with the precision predictor.

Preclinical evaluation of 99mTc direct labeling ZHER2:V2 for HER2 positive tumors imaging.

The present study aimed to label ZHER2:V2 with technetium-99m (99mTc) using a simple method and to evaluate its clinical potential as a diagnostic probe for human epidermal growth factor receptor type 2 (HER2)-positive tumors. The ZHER2:V2 (Affibody molecule of ZHER2:2395-C, which is based on the ZHER2:342 binding sequence with C-terminal engineered cysteine) with C-terminal chelating sequence GGGC was designed and labeled with 99mTc. The 99mTc-ZHER2:V2 labeling efficiency was analyzed. The cellular uptake, retention and binding affinity, and the stability of the probe were examined in vitro. 99mTc-ZHER2:V2 biodistribution analysis and imaging were performed in BALB/c nude mice bearing SKOV3 (HER2-overexpression) xenografts. Furthermore, imaging of the probe was performed in MCF-7 (HER2 low-expression) xenografts. The 99mTc-ZHER2:V2 labeling efficiency was identified as 98.99±0.99% (n=6), and was stable in physiological saline and fresh human serum at 37°C in vitro. The cellular uptake peak of SKOV3 cells at 24 h was 6.15±0.18%, the cellular retention ratio of the probe was 48.58±4.52% at 6 h following interrupted incubation, and ~70% of 99mTc-ZHER2:V2 was membrane bound following 24 h. 99mTc-ZHER2:V2 was blocked by excess amounts of unlabeled ZHER2:V2 in SKOV3 cells. 99mTc-ZHER2:V2 exhibited high distribution (10.07% ID/g) in SKOV3 ×enografts at 6 h following injection. The single photon emission computed tomography (SPECT) imaging revealed clear localization of 99mTc-ZHER2:V2 in the SKOV3 ×enografts at 4 h. However, there was low uptake in MCF-7 tumors on the SPECT images. The SKOV3 ×enograft imaging could be blocked by excess amounts unlabelled ZHER2:V2. 99mTc-ZHER2:V2 is an easy and quick labeling method, with high labeling yields, and radiochemical purity. 99mTc-ZHER2:V2 is a promising probe for the diagnosis of HER2-overexpression tumors and the monitoring of therapy response.

Impact of elution impurities on DOTA and NOTA labeling with two commercial 68Ge/68Ga generators

Different amounts of commercial p-SCN-Bn-DOTA and p-SCN-Bn-NOTA (Macrocyclics, USA) were labeled with 68Ga from IGG-100 purified and unpurified elutions and ITG unpurified elution at different incubation times and temperature. Significant differences in labeling yield were found. Using the ITG generator’s direct elution up to 560 GBq/µmol molar activity can be obtained, while a maximum of 185 GBq/µmol can be obtained with direct elution of the IGG-100 generator. Purification of the elution of the IGG-100 generator increases the labeling yields significantly. Both generators perform per manufacturer specifications and are routinely used for the preparation of 68Ga imaging agents in the clinic.

Noninvasive PET Imaging of a Ga-68-Radiolabeled RRL-Derived Peptide in Hepatocarcinoma Murine Models.

Tc-99m- and I-131-labeled arginine-arginine-leucine (RRL) peptides have shown the feasibility of tumor imaging in our previous studies. However, there have been no reports using RRL peptide for positron emission tomography (PET) imaging. In this study, RRL was radiolabeled with Ga-68 under optimized reaction conditions to develop a better specific and effective tumor imaging agent. RRL was synthesized and conjugated to a bifunctional chelating agent (DOTA-NHS), then radiolabeled with Ga-68. Labeling yield was optimized by varying pH, temperature, and reaction time and the stability was evaluated in human fresh serum. Cellular uptakes of [68Ga]DOTA-RRL and FITC-conjugated RRL in HepG2 cells were evaluated using a gamma counter, confocal microscopy, and flow cytometry. PET images and biodistribution were performed in HepG2 tumor-bearing mice after injection of [68Ga]DOTA-RRL or [68Ga]GaCl3 at different time points. Further, blocking study was investigated using cold RRL. The labeling yield of [68Ga]DOTA-RRL was 80.6 ± 3.9% with a pH of 3.5-4.5 at 100°C for 15min. The cellular uptake of [68Ga]DOTA-RRL in HepG2 cells was significantly higher than that of [68Ga]GaCl3 (P < 0.05). Moreover, the high fluorescent affinity of FITC-conjugated RRL in HepG2 cells was shown using confocal microscopy and flow cytometry. After injection of [68Ga]DOTA-RRL in HepG2 tumor-bearing mice, tumor regions exhibited high radioactive accumulation over 120min and the highest uptake at 30min. After blocked with cold RRL, HepG2 tumors could not be visualized. [68Ga]GaCl3 was unable to show tumor images at any time point. The biodistribution results confirmed the PET imaging and blocking results. Our study successfully prepared [68Ga]DOTA-RRL with a high labeling yield under the optimized reaction conditions and demonstrated its potential role as a PET imaging agent for tumor-targeted diagnosis.

Improved Targeting and Tumor Retention of a Newly Synthesized Antineoplaston A10 Derivative by Intratumoral Administration: Molecular Docking, Technetium 99m Radiolabeling, and In Vivo Biodistribution Studies.

Recently, the direct intratumoral (i.t.) injection of anticancer agents has been investigated. A newly synthesized Antineoplaston A10 analog 3-(4-methoxybenzoylamino)-2,6-piperidinedione (MPD) showed an antitumor activity in human breast cancer cell line. Unfortunately, MPD suffered from poor water solubility. Pseudoternary phase diagram of oil (isopropyl myristate), surfactant (Tween 80), cosurfactant (ethanol), and water was plotted. MPD microemulsion (MPDME) was developed and characterized for particle size (PS), polydispersity index (PDI), zeta potential (ZP), and morphology (transmission electron microscopy). MPDME and MPD solution (MPDS) were radiolabeled with technetium 99m (99mTc) using stannous chloride dihydrate (SnCl2.2H2O). Molecular docking of MPD and 99mTc-MPD was performed to study the interaction with DNA. The impacts of intravenous (i.v.) and i.t. injections of 99mTc-MPDME and 99mTc-MPDS on biodistribution were studied. The developed MPDME showed spherical droplets with mean PS (74.00 ± 5.69 nm), PDI (0.25 ± 0.03), and ZP (33.90 ± 0.90 mV). Labeling yield of 99mTc-MPDME and 99mTc-MPDS was 97.00% ± 0.60% and 92.02% ± 0.45%, respectively. MPD and 99mTc-MPD showed almost same binding affinity with DNA binding site. Biodistribution results showed that i.t. injection of 99mTc-MPDME significantly enhanced tumor retention compared to i.v. route. Herein, the authors concluded that microemulsion could be used as i.t. injectable delivery vehicle to improve targeting and tumor retention of MPD.

Radioiodination and biodistribution of newly synthesized 3-benzyl-2-([3-methoxybenzyl]thio)benzo[g]quinazolin-4-(3H)-one in tumor bearing mice

3-Benzyl-2-((3-methoxybenzyl)thio)benzo[g]quinazolin-4(3H)-one was previously synthesized and proved by physicochemical analyses (HRMS, 1H and 13C NMR). The target compound was examined for its radioactivity and the results showed that benzo[g]quinazoline was successfully labeled with radioactive iodine using NBS via an electrophilic substitution reaction. The reaction parameters that affected the labeling yield such as concentration, pH and time were studied to optimize the labeling conditions. The radiochemical yield was 91.2 ± 1.22% and the in vitro studies showed that the target compound was stable for up to 24 h. The thyroid was among the other organs in which the uptake of 125I-benzoquinazoline has increased significantly over the time up to 4.1%. The tumor uptake was 6.95%. Radiochemical and metabolic stability of the benzoquinazoline in vivo/in vitro and biodistribution studies provide some insights about the requirements for developing more potent radiopharmaceutical for targeting the tumor cells.

Preparation and evaluation of 188 Re sulfide colloidal nanoparticles loaded biodegradable poly (L-lactic acid) microspheres for radioembolization therapy.

Radioembolization with radioactive microspheres has been an effective method for the treatment of liver lesions. The aim of this study was to prepare carrier-free 188 Re loaded poly (L-lactic acid) (PLLA) microspheres through 188 Re sulfide colloidal nanoparticles (188 Re-SC nanoparticles). The formation of 188 Re-SC nanoparticles was confirmed by ultraviolet-visible spectrophotometry. The labeling yield of 188 Re-SC nanoparticles was verified using the RTLC method. Effects of synthesis parameters on morphology and size of prepared 188 Re-sulfide colloidal-PLLA microspheres (188 Re-SC-PLLA microspheres) were studied by scanning electron microscopy. In vitro stability of 188 Re-SC-PLLA microspheres was investigated in normal saline at room temperature and in human serum at 37°C. In vivo distribution studies and gamma camera imaging were performed in healthy BALB/c mice. The microspheres could be prepared with sizes between 13 and 48μm (modal value 29μm) and radiolabeling efficiency >99%. After incubation, the microspheres were found stable in vitro up to 72hours. The biodistribution after intravenous injection in healthy BALB/c mice showed high accumulation in lung as a first capture pathway organ for microsphere followed by great retention over 48hours for these microspheres. These data show that 188 Re-SC-PLLA microspheres are suitable candidate for clinical studies.

Intrinsically Zirconium-89-Labeled Manganese Oxide Nanoparticles for In Vivo Dual-Modality Positron Emission Tomography and Magnetic Resonance Imaging.

Manganese-based nanoparticles (NPs) have recently attracted much attention in the field of biomedical imaging due to their impressive enhanced T1 contrast ability. Although the reported manganese-based NPs have exhibited good imaging capabilities as contrast agents, it is still urgent to develop novel multifunctional manganese-based imaging probes for future biomedical imaging, especially PET/MRI probes. Herein, we present chelator-free zirconium-89 (89Zr, t1/2: 78.4 h) labeling of manganese oxide NPs (Mn3O4@PEG) with ∼78% labeling yield and good stability. Serial positron emission tomography (PET) and magnetic resonance imaging (MRI) studies non-invasively assessed the biodistribution patterns of the NPs and the feasibility of in vivo dual-modality imaging and lymph-node mapping. Since Mn3O4 NPs exhibited desirable properties for enhanced T1 imaging and the simplicity of chelator-free radiolabeling, [89Zr]Mn3O4@PEG NPs offer a novel, simple, safe and accurate nanoplatforms for future precise cancer imaging and diagnosis.