Preparation of 177Lu‐labeled Nimotuzumab for radioimmunotherapy of EGFR‐positive cancers: Comparison of DOTA and CHX‐A″‐DTPA as bifunctional chelators

This study was aimed at evaluating the role of bifunctional chelators DOTA‐NCS and CHX‐A″‐DTPA‐NCS used for conjugating ¹⁷⁷Lu with Nimotuzumab on the radiochemical yields, purity, in vitro stability, and specificity of the radioimmunoconjugates to EGFR. Two immunoconjugates were prepared wherein Nimotuzumab was conjugated with the acyclic ligand p‐NCS‐Bn‐CHX‐A″‐DTPA and macrocyclic ligand p‐NCS‐Bn‐DOTA. These were radiolabeled with ¹⁷⁷Lu, purified on PD‐10 column, and characterized by SE‐HPLC. In vitro stability was determined up to 4 days post preparation. Specificity of the radioimmunoconjugates was ascertained by in vitro studies in A431 cells while the biodistribution patterns were studied in normal Swiss mice up to 96 hours post injection. Four to five molecules of CHX‐A″‐DTPA/DOTA were attached to one molecule of Nimotuzumab. Radiochemical purity of both ¹⁷⁷Lu‐CHX‐A″‐DTPA‐Nimotuzumab and ¹⁷⁷Lu‐DOTA‐Nimotuzumab was determined to be greater than 98%. Both the radioimmunoconjugates exhibited good in vitro stability at 37°C up to 4 days post preparation in saline, and their clearance was largely by the hepatobiliary route. The DOTA‐ and CHX‐A″‐DTPA‐based radioimmunoconjugates could be prepared with good radiochemical purity, in vitro stability, and specificity to EGFR. Further studies in EGFR‐positive cancers would pave way for them for use in the clinics.     

Preclinical evaluation of new GnRH‐I receptor radionuclide therapy with 177Lu‐peptide tracer

The purpose of this study was to develop preclinical evaluation of a novel radiolabeled gonadotropin‐releasing hormone (GnRH) receptor targeting peptide for prostate cancer therapy. The new antiproliferative agent of GnRH‐I analogue was developed on the basis of the D‐Trp⁶‐GnRH‐I scaffold, and in vivo pharmacokinetics and receptor binding affinity were enhanced by the substitution of Gly‐NHNH₂ for Gly‐NH₂ at position 10 in D‐Trp⁶‐GnRH‐I. To evaluate ¹⁷⁷Lu‐DOTA‐triptorelin‐hydrazide as radionuclide therapy of tumor, the quality control tests and preclinical stage assessment were carried out. Solid‐phase method was used to synthesize new peptide. Characterization and purity of peptide were done by mass spectroscopy and high‐performance liquid chromatography (HPLC). In order to be utilized in targeted therapy, the new GnRH‐I agonist was coupled with pSCN‐Bn‐DOTA. The precipitate crude of DOTA‐triptorelin‐hydrazide was then purified via preparative HPLC. At optimal conditions of time, temperature, ligand amount, and lutetium content, DOTA‐triptorelin‐hydrazide was labeled with ¹⁷⁷Lu (specific activity not less than 925 GBq/mg). Investigation of the in vivo biodistribution and in vitro studies for ¹⁷⁷Lu‐DOTA‐TRPHYD was performed in three different ways, and the binding of radiopeptide to GnRH receptors was expressed on the human cell lines using ¹²⁵I‐labeled D‐TRP⁶GnRH‐I as a tracer, respectively. Synthesized novel GnRH‐I was obtained with purity greater than 98%. Paper chromatography was found to be the most suitable with R_f of the complex and observed radiochemical purity of RTLC and HPLC greater than 97%. For in vivo studies, ¹⁷⁷Lu‐DOTA‐triptorelin‐hydrazide showed promising results with fast clearance from the blood and resulted in good T/NT ratios at 1, 4, and 24 hours postinjection and satisfactory biodistribution with no significant activity seen in normal tissue. The values of internalization efficiency and receptor affinity of new radiopeptide binding were IC₅₀ = 0.47 ± 0.06 vs 0.13 ± 0.01 nM for triptorelin and cellular uptake: 3.4 ± 0.7% at 1 hour and 6.8 ± 1.17% at 4 hours of the internal reference. The results showed a good stability and radiochemical purity of the obtained radioconjugate. For in vivo and in vitro studies, new radiopeptide showed a high uptake of ¹⁷⁷Lu conjugate in tumor and rapid clearance from the blood stream almost entirely via the renal/urinary pathway and binding to the GnRH receptors with high specificity and affinity, respectively.     

177Lu‐labeled monomeric,dimeric and multimeric RGD peptides for the therapy of tumors expressing α(ν)β(3) integrins

The conjugation of peptides to gold nanoparticles (AuNPs) produces biocompatible and stable multimeric systems with target‐specific molecular recognition. Peptides based on the cyclic Arg‐Gly‐Asp (RGD) sequence have been reported as high‐affinity agents for the α(ν)β(3) integrin. The aim of this research was to prepare a multimeric system of ¹⁷⁷Lu‐labeled gold nanoparticles conjugated to c(RGDfK)C (cyclo(Arg‐Gly‐Asp‐Phe‐Lys)Cys) and to compare the radiation‐absorbed dose with that of ¹⁷⁷Lu‐labeled monomeric and dimeric RGD peptides to α(ν)β(3) integrin‐positive U87MG tumors in mice. DOTA‐GGC (1,4,7,10‐tetraazacyclododecane‐N‐N′,N″,N?‐tetraacetic acid‐Gly‐Gly‐Cys) and c(RGDfK)C peptides were synthesized and conjugated to AuNPs by a spontaneous reaction of the thiol groups. Transmission electron microscopy, ultraviolet–visible, X‐ray photoelectron spectroscopy, Raman and far‐infrared spectroscopy techniques demonstrated that AuNPs were functionalized with the peptides. For the ¹⁷⁷Lu‐AuNP‐c(RGDfK)C to be obtained, the ¹⁷⁷Lu‐DOTA‐GGC radiopeptide was first prepared and added to a solution of AuNPs followed by c(RGDfK)C (25 µl, 5 µ m ) at 18 °C for 15 min. ¹⁷⁷Lu‐DOTA‐GGC, ¹⁷⁷Lu‐DOTA‐cRGDfK and ¹⁷⁷Lu‐DOTA‐E‐c(RGDfK)₂ were prepared by adding ¹⁷⁷LuCl₃ (370 MBq) to 5 µl (1 mg/ml) of the DOTA derivative diluted with 50 µl of 1 m acetate buffer pH 5. The mixture was incubated at 90 °C in a block heater for 30 min. Radiochemical purity was determined by ultrafiltration and HPLC analyses. Biokinetic studies were accomplished in athymic mice with U87MG‐induced tumors. The radiochemical purity for all ¹⁷⁷Lu‐RGD derivatives was 96 ± 2%. ¹⁷⁷Lu‐absorbed doses per injected activity delivered to U87MG tumors were 0.357 ± 0.052 Gy/MBq (multimer), 0.252 ± 0.027 Gy/MBq (dimer) and 0.102 ± 0.018 Gy/MBq (monomer). ¹⁷⁷Lu‐labeled dimeric and multimeric RGD peptides demonstrated properties suitable for targeted radionuclide therapy of tumors expressing α(ν)β(3) integrins. Copyright © 2012 John Wiley & Sons, Ltd.     

Single vial kit formulation of DOTATATE for preparation of 177Lu‐labeled therapeutic radiopharmaceutical at hospital radiopharmacy

The clinical applications of radiolabeled somatostatin analogue ¹⁷⁷Lu‐DOTA‐Tyr³‐Thr⁸‐Octreotide (¹⁷⁷Lu‐DOTATATE) constitute a promising treatment option for patients with disseminated and inoperable neuroendocrine (NET) tumors. Formulation of ¹⁷⁷Lu‐DOTATATE in hospital radiopharmacy under aseptic conditions in a safe and reliable manner is a major constraint for its extensive use. The present work was intended to develop a kit for the safe preparation of the therapeutic radiopharmaceutical, viz. ¹⁷⁷Lu‐DOTATATE of high quality that can be easily adapted at conventional hospital radiopharmacies. Single vial kits of DOTATATE were formulated and evaluated for suitability for radiolabeling as well as stability on its storage. Patient dose of ¹⁷⁷Lu‐DOTATATE (7.4 GBq) could be successfully prepared using semi‐automated in‐house setup that assures safe handling and high yields of product of pharmaceutical purity suitable for clinical use. Fast clearance of activity via renal route was observed in preclinical biodistribution studies of ¹⁷⁷Lu‐DOTATATE carried out in normal Swiss mice. Deployment of in‐house produced ¹⁷⁷LuCl₃, cold kits and easy adaptability of synthesis setup at hospital radiopharmacy for preparation is likely to expand applications of peptide receptor radionuclide therapy.     

Multifunctional targeted therapy system based on 99mTc/177Lu‐labeled gold nanoparticles‐Tat(49–57)‐Lys3‐bombesin internalized in nuclei of prostate cancer cells

Radiolabeled gold nanoparticles may function simultaneously as radiotherapy and thermal ablation systems. The gastrin‐releasing peptide receptor (GRP‐r) is overexpressed in prostate cancer, and Lys³‐bombesin is a peptide that binds with high affinity to the GRP‐r. HIV Tat(49–57) is a cell‐penetrating peptide that reaches the DNA. In cancer cells, ¹⁷⁷Lu shows efficient crossfire effect, whereas ^99mTc that is internalized in the cancer cell nuclei acts as an effective system of targeted radiotherapy because of the biological Auger effect. The aim of this research was to evaluate the in vitro potential of ^99mTc‐labeled and ¹⁷⁷Lu‐labeled gold nanoparticles conjugated to Tat(49–57)‐Lys³‐bombesin peptides (^99mTc/¹⁷⁷Lu‐AuNP‐Tat‐BN) as a plasmonic photothermal therapy and targeted radiotherapy system in PC3 prostate cancer cells. Peptides were conjugated to AuNPs (5 nm) by spontaneous reaction with the thiol group of cysteine (Cys). The effect on PC3 cell viability after laser heating of the AuNP‐Tat‐BN incubated with the cancer cells was conducted using an Nd:YAG laser pulsed for 5 ns at 532 nm (0.65 W/cm²). For the ^99mTc/¹⁷⁷Lu‐AuNP‐Tat‐BN to be obtained, the ¹⁷⁷Lu‐DOTA‐Gly‐Gly‐Cys and ^99mTc‐HYNIC‐octreotide radiopeptides were first prepared and added simultaneously to a solution of AuNP‐Tat‐BN. ^99mTc/¹⁷⁷Lu‐AuNP‐Tat‐BN (20 Bq/cell) was incubated with PC3 cells, and the effect on the cell proliferation was evaluated after 3 days. Fluorescence images of ^99mTc/¹⁷⁷Lu‐AuNP‐Tat‐BN internalized in nuclei of PC3 were also obtained. After laser irradiation, the presence of AuNP‐Tat‐BN caused a significant increase in the temperature of the medium (46.4 vs 39.5 °C of that without AuNP) resulting in a significant decrease in PC3 cell viability down to 1.3%. After treatment with ^99mTc/¹⁷⁷Lu‐AuNP‐Tat‐BN, the PC3 cell proliferation was inhibited. The nanosystem exhibited properties suitable for plasmonic photothermal therapy and targeted radiotherapy in the treatment of prostate cancer.     

Preparation and preliminary biological evaluation of 177Lu‐labeled GluDTPA‐cyclo(RGDfK) for integrin ανβ3 receptor‐positive tumor targeting

Integrin α_νβ₃ is a receptor and is highly expressed on activated and proliferating endothelial cells during the growth and metastasis of solid tumors but not on resting endothelial cells and normal organs. Because RGD peptide binds to integrin α_νβ₃ receptor, a variety of radiolabeled RGD peptides have been evaluated for non‐invasive imaging of integrin α_νβ₃‐positive tumors. In an attempt to develop RGD‐based radiopharmaceuticals, a novel GluDTPA‐cyclo arginine‐glycine‐aspartic acid‐d ‐phenylalanine‐lysine (GluDTPA‐cycloRGDfK) was simply synthesized and radiolabeled with ¹⁷⁷Lu. Also, tumor targeting and retention of the radiolabeled complex were evaluated in U87MG glioma‐bearing mice. The ¹⁷⁷Lu‐labeled GluDTPA‐cyclo(RGDfK) was formulated with a high radiolabeling yield (>98%) under mild condition, and the radiochemical purity was sustained in both saline and serum for over 4 days at 37°C. The radiolabeled compounds were rapidly cleared from the blood pool and non‐target tissue. Tumor‐to‐blood ratio was 12.09 at 2 h post injection and increased to 134.67 at 24 h, while tumor to liver ratio was 2.01 at 24 h similar to that of 2 h. Though it is inappropriate for targeted therapy due to its low uptake in tumor (~ 1 %ID/g), the acceptable results on radiochemistry and biodistribution propose to take a further assessment for non‐invasive imaging and detection of integrin α_νβ₃‐positive tumors by applying diagnostic radionuclides. Copyright © 2011 John Wiley & Sons, Ltd.     

Synthesis and biodistribution studiesof 177Lu‐trastuzumab as a therapeutic agent in the breast cancer mice model

Trastuzumab is a humanized monoclonal antibody against the HER2 that has the potential to be used as radioimmunotherapy (RIT) agent in treatment of breast cancer. Lutetium‐177 has beta energy suitable for therapy and gamma photons for imaging. We labeled trastuzumab with lutetium‐177 via DOTA as chelator and performed some necessary tests for the first stage in using complex as a RIT agent. Radiochemical purity, immunoreactivity and stability of complex were determined. The biodistribution and imaging studies were determined in mice bearing breast tumor. The radiochemical purity was 94±0.9%. Lutetium‐Trastuzumab showed a good stability at biological condition. The tumor to blood ratio was calculated 3.29(±0.09) after 7 days. The good tumor uptake in biodistribution studies was agreed with gamma camera images after 7 days. The results showed that the new complex could be considered for further evaluation in animals and possibly in humans as a new radiopharmaceutical for use in RIT against breast cancer. Copyright © 2010 John Wiley & Sons, Ltd.     

Dual‐labeled prostate‐specific membrane antigen (PSMA)‐targeting agent for preoperative molecular imaging and fluorescence‐guided surgery for prostate cancer

The objective of this study was to report the synthesis and characteristics of a dual modality imaging agent, Tc‐99m GRFLTGGTGRLLRIS‐GHEG‐ECG‐K(‐5‐carboxy‐X‐rhodamine)‐NH₂ (GRFLT‐ECG‐ROX), and to verify its feasibility as both molecular imaging and intraoperative guidance agent. GRFLT‐ECG‐ROX was synthesized using Fmoc solid‐phase peptide synthesis. Radiolabeling of GRFLT‐ECG‐ROX with Tc‐99m was accomplished using ligand exchange via tartrate. Binding affinity and in vitro cellular uptake studies were performed. Gamma camera imaging, biodistribution, and ex vivo imaging studies were performed using LNCaP and PC‐3 tumor‐bearing murine models. Surgical removal of tumor nodules in murine models with peritoneal carcinomatosis was performed under a fluorescence imaging system. After radiolabeling procedures with Tc‐99m, Tc‐99m GRFLT‐ECG‐ROX complexes were prepared in high yield (>96%). The binding affinity value (K_d) of Tc‐99m GRFLT‐ECG‐ROX for LNCaP cells was estimated to be 9.5 ± 1.3 nM. In gamma camera imaging, the tumor to normal muscle uptake ratios of Tc‐99m GRFLT‐ECG‐ROX increased with time (3.1 ± 0.2, 4.0 ± 0.4, and 6.3 ± 0.9 at 1, 2, and 3 h, respectively). Under real‐time optical imaging, the removal of visible nodules was successfully performed. Thus, Tc‐99m GRFLT‐ECG‐ROX could provide both preoperative molecular imaging and fluorescence imaging guidance for tumor removal.     

Preclinical assessment of 68Ga‐PSMA‐617 entrapped in a microemulsion delivery system for applications in prostate cancer PET/CT imaging

It has in recent years been reported that microemulsion (ME) delivery systems provide an opportunity to improve the efficacy of a therapeutic agent whilst minimising side effects and also offer the advantage of favourable treatment regimens. The prostate‐specific membrane antigen (PSMA) targeting agents PSMA‐11 and PSMA‐617, which accumulate in prostate tumours, allow for [⁶⁸Ga]Ga³⁺‐radiolabelling and positron emission tomography/computed tomography (PET) imaging of PSMA expression in vivo. We herein report the formulation of [⁶⁸Ga]Ga‐PSMA‐617 into a ME ≤40 nm including its evaluation for improved cellular toxicity and in vivo biodistribution. The [⁶⁸Ga]Ga‐PSMA‐617‐ME was tested in vitro for its cytotoxicity to HEK293 and PC3 cells. [⁶⁸Ga]Ga‐PSMA‐617‐ME was administered intravenously in BALB/c mice followed by microPET/computed tomography (CT) imaging and ex vivo biodistribution determination. [⁶⁸Ga]Ga‐PSMA‐617‐ME indicated negligible cellular toxicity at different concentrations. A statistically higher tolerance towards the [⁶⁸Ga]Ga‐PSMA‐617‐ME occurred at 0.125 mg/mL by HEK293 cells compared with PC3 cells. The biodistribution in wild‐type BALB/C mice showed the highest amounts of radioactivity (%ID/g) presented in the kidneys (31%) followed by the small intestine (10%) and stomach (9%); the lowest uptake was seen in the brain (0.5%). The incorporation of [⁶⁸Ga]Ga‐PSMA‐617 into ME was successfully demonstrated and resulted in a stable nontoxic formulation as evaluated by in vitro and in vivo means.     

A “mix‐and‐use” approach for formulation of human clinical doses of 177Lu‐DOTMP at hospital radiopharmacy for management of pain arising from skeletal metastases

Use of bone‐seeking radiopharmaceuticals is an established modality in the palliative care of pain due to skeletal metastases. ¹⁷⁷Lu‐DOTMP is a promising radiopharmaceutical for this application owing to the ideally suited decay properties of ¹⁷⁷Lu and excellent thermodynamic stability and kinetic rigidity of the macrocyclic complex. The aim of the present study is to develop a robust and easily adaptable protocol for formulation of clinical doses of ¹⁷⁷Lu‐DOTMP at hospital radiopharmacy. After extensive radiochemical studies, an optimized strategy for formulation of clinical doses of ¹⁷⁷Lu‐DOTMP was developed, which involves simple mixing of approximately 3.7 GBq of ¹⁷⁷Lu activity as ¹⁷⁷LuCl₃ solution to an aqueous solution containing 5 mg of DOTMP and 8 mg of NaHCO₃. The proposed protocol yielded ¹⁷⁷Lu‐DOTMP with >98% radiochemical purity, and the resultant formulation showed excellent in vitro stability and desired pharmacokinetic properties in animal model. Preliminary clinical investigations in 5 patients showed specific skeletal accumulation with preferential localization in the osteoblastic lesion sites and almost no uptake in soft tissue or any other major nontarget organ. The developed “mix‐and‐use” strategy would be useful for large number of nuclear medicine centers having access to ¹⁷⁷Lu activity and would thereby accelerate the clinical translation of ¹⁷⁷Lu‐DOTMP.     

Radiolabeled nanobodies as theranostic tools in targeted radionuclide therapy of cancer

Introduction: The integration of diagnostic testing for the presence of a molecular target is of interest to predict successful targeted radionuclide therapy (TRNT). This so-called ‘theranostic’ approach aims to improve personalized treatment based on the molecular characteristics of cancer cells. Moreover, it offers new insights in predicting adverse effects and provides appropriate tools to monitor therapy responses. Recent findings using nanobodies emphasize their potential as theranostic tools in cancer treatment. Nanobodies are recombinant, small antigen-binding fragments that are derived from camelid heavy-chain-only antibodies.

Areas covered: We review the current status of theranostic approaches in TRNT, with a focus on antibodies, peptides, scaffold proteins and emerging nanobodies. In recent years, nanobodies have been evaluated intensively for molecular imaging. In addition, novel data on TRNT using radiolabeled nanobodies for carcinomas and multiple myeloma highlight their promising opportunities in cancer treatment.

Expert opinion: We trust that radiolabeled nanobodies will have a future potential as theranostic tools in cancer therapy, both for diagnosis as well as for TRNT.     

Bispecific radioligands targeting prostate‐specific membrane antigen and gastrin‐releasing peptide receptors on the surface of prostate cancer cells

Metastases of prostate cancer usually show highly heterogeneous or partly lost prostate‐specific membrane antigen (PSMA) expression. In order to image and treat both PSMA positive and negative tissues, the extension of PSMA tracer specificity to other receptors, also highly expressed on the surface of prostate cancer cells, has been suggested. Prostate cancer cells usually express both PSMA and gastrin‐releasing peptide (GRP) receptors; thus, bispecific heterodimeric molecules, addressing both targets at the same time, may significantly improve prostate cancer imaging and therapy. This article summarizes preclinical data regarding low‐molecular weight molecules targeting PSMA and GRP receptors.     

Spacer length impacts the efficacy of targeted docetaxel conjugates in prostate-specific membrane antigen expressing prostate cancer

Abstract

Combination of targeted delivery and controlled release is a powerful technique for cancer treatment. In this paper, we describe the design, synthesis, structure validation and biological properties of targeted and non-targeted N-(2-hydroxypropyl)methacrylamide (HPMA) copolymer-docetaxel conjugates. Docetaxel (DTX) was conjugated to HPMA copolymer via a tetrapeptide spacer (–GFLG-). 3-(1,3-dicarboxypropyl)-ureido]pentanedioic acid (DUPA) was used as the targeting moiety to actively deliver DTX for treatment of Prostate-Specific Membrane Antigen (PSMA) expressing prostate cancer. Short and long spacer DUPA monomers were prepared, and four HPMA copolymer – DTX conjugates (non-targeted, two targeted with short spacer of different molecular weight and targeted with long spacer) were prepared via Reversible Addition-Fragmentation Chain Transfer (RAFT) copolymerization. Following confirmation of PSMA expression on C4-2 cell line, the DTX conjugates’ in vitro cytotoxicity was tested against C4-2 tumor cells and their anticancer efficacies were assessed in nude mice bearing s.c. human prostate adenocarcinoma C4-2 xenografts. The in vivo results show that the spacer length between targeting moieties and HPMA copolymer backbone can significantly affect the treatment efficacy of DTX conjugates against C4-2 tumor bearing nu/nu mice. Moreover, histological analysis indicated that the DUPA-targeted DTX conjugate with longer spacer had no toxicity in major organs of treated mice.     

High specific activity is not optimal: 18F‐fluoroestradio positron emission tomography‐computed tomography results in a breast cancer xenograft

The purpose of the preliminary study was to investigate whether high specific activity (SA) of ¹⁸F‐fluoroestradiol was optimal in breast cancer diagnosis. Imaging at variable SA was conducted in a ZR‐75‐1 xenograft model of estrogen‐receptor positive human breast cancer in 6 mice. The region of interest was manually drawn, and the percent of injected dose per gram of the tumor and muscle in the regions of interest were recorded. Tumor‐to‐muscle ratio (T/M) was calculated and compared in each group with different SAs. In addition, the correlation between blood estradiol and sex hormone‐binding globulin and the value of T/M were also analyzed. The value of T/M increased initially with the rise of SA and it reached the peak at SA of 1.6 Ci/μmol. After that, the value fell down sharply and remained stable from SA of 3.1 Ci/μmol. The value of T/M was highest at SA of 1.6 Ci/μmol (P < .001). Additionally, the blood levels of estradiol and sex hormone‐binding globulin showed no correlation with the value of T/M (P > .05). High SA of ¹⁸F‐fluoroestradiol leads to low T/M results in breast cancer xenograft models. We should control SA in a reasonable range to obtain high‐quality images.     

Preparation,evaluation, and first clinical use of 177Lu‐labeled hydroxyapatite (HA) particles in the treatment of rheumatoid arthritis: utility of cold kits for convenient dose formulation at hospital radiopharmacy

While radiation synovectomy (RSV) constitutes a successful paradigm for the treatment of arthritis, a major cornerstone of its success resides in the selection of appropriate radiolabeled agent. Among the radionuclide used for RSV, the scope of using ¹⁷⁷Lu [T_1/2 = 6.65 d, E_β(max) = 497 keV, E_γ = 113 KeV (6.4%), 208 KeV (11%)] seemed to be attractive owing to its suitable decay characteristics, easy availability, and cost‐effective production route. The present article describes a formulation of ¹⁷⁷Lu‐labeled hydroxyapatite (HA) using ready‐to‐use kits of HA particles of 1–10 µm size range. The developed kits enable convenient one‐step preparation of ¹⁷⁷Lu‐HA (400 ± 30 MBq doses) in high radiochemical purity (>99%) and stability at hospital radiopharmacy. The preparation showed promising results in pre‐clinical studies carried out in Wistar rats bearing arthritis in knee joints. In preliminary clinical investigation, significant improvement in the disease conditions was reported in 10 patients with rheumatoid arthritis of knee joints treated with 333 ± 46 MBq doses of ¹⁷⁷Lu‐HA. The studies reveal that while ¹⁷⁷Lu labeled HA particles holds considerable promise as a cost‐effective agent for RSV, the adopted strategy of using HA kits could be a potential step toward wider clinical utilization of radiolanthanide‐labeled HA particles. Copyright © 2014 John Wiley & Sons, Ltd.     

The roles of endoplasmic reticulum stress and mitochondrial apoptotic signaling pathway in quercetin‐mediated cell death of human prostate cancer PC‐3 cells

Prostate cancer has its highest incidence and is becoming a major concern. Many studies have shown that traditional Chinese medicine exhibited antitumor responses. Quercetin, a natural polyphenolic compound, has been shown to induce apoptosis in many human cancer cell lines. Although numerous evidences show multiple possible signaling pathways of quercetin in apoptosis, there is no report to address the role of endoplasmic reticulum (ER) stress in quercetin‐induced apoptosis in PC‐3 cells. The purpose of this study was to investigate the effects of quercetin on the induction of the apoptotic pathway in human prostate cancer PC‐3 cells. Cells were treated with quercetin for 24 and 48 h and at various doses (50–200 μM), and cell morphology and viability decreased significantly in dose‐dependent manners. Flow cytometric assay indicated that quercetin at 150 μM caused G0/G1 phase arrest (31.4–49.7%) and sub‐G1 phase cells (19.77%) for 36 h treatment and this effect is a time‐dependent manner. Western blotting analysis indicated that quercetin induces the G0/G1 phase arrest via decreasing the levels of CDK2, cyclins E, and D proteins. Quercetin also stimulated the protein expression of ATF, GRP78, and GADD153 which is a hall marker of ER stress. Furthermore, PC‐3 cells after incubation with quercetin for 48 h showed an apoptotic cell death and DNA damage which are confirmed by DAPI and Comet assays, leading to decrease the antiapoptotic Bcl‐2 protein and level of ΔΨ_m, and increase the proapoptotic Bax protein and the activations of caspase‐3, ‐8, and ‐9. Moreover, quercetin promoted the trafficking of AIF protein released from mitochondria to nuclei. These data suggest that quercetin may induce apoptosis by direct activation of caspase cascade through mitochondrial pathway and ER stress in PC‐3 cells. © 2012 Wiley Periodicals, Inc. Environ Toxicol 29: 428–439, 2014.     

Comparative evaluation of the new GRPR‐antagonist 111In‐SB9 and 111In‐AMBA in prostate cancer models: Implications of in vivo stability

Gastrin‐releasing peptide receptors (GRPRs) are overexpressed in prostate cancer, representing attractive targets for diagnosis and therapy with bombesin (BBN)‐like radioligands. GRPR‐antagonists have lately attracted much attention owing to inherent biosafety and favorable pharmacokinetics. We herein present the GRPR‐antagonist SB9 structurally resembling the known BBN‐based agonist AMBA (SB9 = [Leu¹³NHEt‐desMet¹⁴]AMBA). The profiles of ¹¹¹In‐SB9 and ¹¹¹In‐AMBA were directly compared in PC‐3 cells and tumor‐bearing mice. SB9 and AMBA displayed high GRPR affinities. ¹¹¹In‐AMBA strongly internalized in PC‐3 cells, while ¹¹¹In‐SB9 remained bound on the cell surface showing a typical GRPR‐radioantagonist profile. ¹¹¹In‐SB9 was more stable than ¹¹¹In‐AMBA, but coinjection of the neprilysin (NEP) inhibitor phosphoramidon (PA) stabilized both in vivo. The radioligands displayed high tumor uptake (20.23 ± 3.41 %ID/g and 18.53 ± 1.54 %ID/g, respectively, at 4 hours pi), but ¹¹¹In‐SB9 washed faster from background. PA coinjection led to significant increase of tumor uptake, combined with better clearance for ¹¹¹In‐SB9. In short, this study has revealed superior pharmacokinetics and higher stability for the GRPR‐antagonist ¹¹¹In‐SB9 vs the corresponding agonist ¹¹¹In‐AMBA consolidating previous evidence that GRPR antagonists are preferable to agonists for tumor imaging and therapy. It has also demonstrated that further pharmacokinetic improvements were feasible by in situ metabolic radioligand stabilization using PA.