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A high-affinity, high-stability photoacoustic agent for imaging gastrin-releasing peptide receptor in prostate cancer.

ABSTRACT: To evaluate the utility of targeted photoacoustic imaging (PAI) in providing molecular information to complement intrinsic functional and anatomical details of the vasculature within prostate lesion.We developed a PAI agent, AA3G-740, that targets gastrin-releasing peptide receptor (GRPR), found to be highly overexpressed in prostate cancer. The binding specificity of the agent was evaluated in human prostate cancer cell lines, PC3 and LNCaP, and antagonist properties determined by cell internalization and intracellular calcium mobilization studies. The imaging sensitivity was assessed for the agent itself and for the PC3 cells labeled with agent. The in vivo stability of the agent was determined in human plasma and in the blood of living mice. The in vivo binding of the agent was evaluated in PC3 prostate tumor models in mice, and was validated ex vivo by optical imaging.AA3G-740 demonstrated strong and specific binding to GRPR. The sensitivity of detection in vitro indicated suitability of the agent to image very small lesions. In mice, the agent was able to bind to GRPR even in poorly vascularized tumors leading to nearly 2-fold difference in photoacoustic signal relative to the control agent.The ability to image both vasculature and molecular profile outside the blood vessels gives molecular PAI a unique advantage over currently used imaging techniques. The imaging method presented here can find application both in diagnosis and in image-guided biopsy.

SUBMITTER: Levi J

PROVIDER: S-EPMC4121111 | biostudies-literature | 2014 Jul

REPOSITORIES: biostudies-literature

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A high-affinity, high-stability photoacoustic agent for imaging gastrin-releasing peptide receptor in prostate cancer.

Levi Jelena J Sathirachinda Ataya A Gambhir Sanjiv S SS

Clinical cancer research : an official journal of the American Association for Cancer Research 20140521 14

<h4>Purpose</h4>To evaluate the utility of targeted photoacoustic imaging (PAI) in providing molecular information to complement intrinsic functional and anatomical details of the vasculature within prostate lesion.<h4>Experimental design</h4>We developed a PAI agent, AA3G-740, that targets gastrin-releasing peptide receptor (GRPR), found to be highly overexpressed in prostate cancer. The binding specificity of the agent was evaluated in human prostate cancer cell lines, PC3 and LNCaP, and antag ...[more]

PMID: 24850845

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Project description:The prostate-specific membrane antigen (PSMA) and gastrin-releasing peptide receptor (GRPR) are identified as important targets on prostate cancer. Receptor-targeting radiolabeled imaging pharmaceuticals with high affinity and specificity are useful in studying and monitoring biological processes and responses. Two potential imaging pharmaceuticals, AMBA agonist (where AMBA = DO3A-CH2CO-G-[4-aminobenzyl]- Gln-Trp-Ala-Val-Gly-His-Leu-Met-NH2) and RM1 antagonist (where RM1 = DO3A-CH2CO-G-[4-aminobenzyl]-D-Phe-Gln-Trp-Ala-Val-Gly-His-Sta-Leu-NH2), have demonstrated high binding affinity (IC50) to GRP receptors and high tumor uptake. Antagonists, despite the poor tumor cell internalization properties, can show clearer images and pharmacokinetic profiles by virtue of their higher tumor uptake in animal models compared to agonists. For characterization, development, and translation of a potential imaging pharmaceutical into the clinic, it must be evaluated in a series of tests, including in vitro cell binding assays, in vitro buffer and serum stability studies, the biodistribution of the radiolabeled material, and finally imaging studies in preclinical animal models. Data related to acetate buffer, mouse, canine, and human sera stability of 177Lu-labeled RM1 are presented here and compared with the acetate buffer and sera stability data of AMBA agonist. The samples of 177Lu-labeled RM1 with a high radioconcentration degrade faster than low-radioconcentration samples upon storage at 2-8 °C. Addition of stabilizers, ascorbic acid and gentisic acid, improve the stability of 177Lu-labeled RM1 significantly with gentisic acid being more efficient than ascorbic acid as a stabilizer. The degradation kinetics of 177Lu-labeled AMBA and RM1 in sera follow the order (fastest to slowest): mouse > canine > human sera. Finally, 177Lu-labeled RM1 antagonist is slower to degrade in mouse, canine, and human sera than 177Lu-labeled AMBA agonist, further suggesting that an antagonist is a more promising candidate than agonist for the positron emission tomography (PET) imaging and therapy of prostate cancer patients.

Project description:Gastrin-releasing peptide (GRP) receptors (GRPr) are frequently overexpressed in human prostate cancer, and radiolabeled GRPr affinity ligands have shown promise for in vivo imaging of prostate cancer with PET. The goal of this study was to develop a dual-modality imaging probe that can be used for noninvasive PET imaging and optical imaging of prostate cancer.MethodsWe designed and synthesized an IRDye 650 and DOTA-conjugated GRPr antagonist, HZ220 (DOTA-Lys(IRDye 650)-PEG4-[D-Phe6, Sta13]-BN(6-14)NH2), by reacting DOTA-Lys-PEG4-[D-Phe6, Sta13]-BN(6-14)NH2 (HZ219) with IRDye 650 N-hydroxysuccinimide (NHS) ester. Receptor-specific binding of gallium-labeled HZ220 was characterized in PC-3 prostate cancer cells (PC-3), and tumor uptake in mice was imaged with PET/CT and fluorescence imaging. Receptor binding affinity, in vivo tumor uptake, and biodistribution were compared with the GRPr antagonists HZ219, DOTA-PEG4-[D-Phe6, Sta13]-BN(6-14)NH2 (DOTA-AR), and DOTA-(4-amino-1-carboxymethyl-piperidine)-[D-Phe6, Sta13]-BN(6-14)NH2 (DOTA-RM2).ResultsAfter hydrophilic-lipophilic balance cartridge purification, 68Ga-HZ220 was obtained with a radiochemical yield of 56% ± 8% (non-decay-corrected), and the radiochemical purity was greater than 95%. Ga-HZ220 had a lower affinity for GRPr (inhibitory concentration of 50% [IC50], 21.4 ± 7.4 nM) than Ga-DOTA-AR (IC50, 0.48 ± 0.18 nM) or Ga-HZ219 (IC50, 0.69 ± 0.18 nM). Nevertheless, 68Ga-HZ220 had an in vivo tumor accumulation similar to 68Ga-DOTA-AR (4.63 ± 0.31 vs. 4.07 ± 0.29 percentage injected activity per mL [%IA/mL] at 1 h after injection) but lower than that of 68Ga-DOTA-RM2 (10.4 ± 0.4 %IA/mL). The tumor uptake of 68Ga-HZ220 was blocked significantly with an excessive amount of GRP antagonists. IVIS spectrum imaging also visualized PC-3 xenografts in vivo and ex vivo with a high-contrast ratio. Autoradiography and fluorescent-based microscopic imaging with 68Ga-HZ220 consistently colocated the expression of GRPr. 68Ga-HZ220 displayed a higher kidney uptake than both 68Ga-DOTA-AR and 68Ga-DOTA-RM2 (16.9 ± 6.5 vs. 4.48 ± 1.63 vs. 5.01 ± 2.29 %IA/mL).Conclusion68Ga-HZ220 is a promising bimodal ligand for noninvasive PET imaging and intraoperative optical imaging of GRPr-expressing malignancies. Bimodal nuclear/fluorescence imaging may not only improve cancer detection and guide surgical resections, but also improve our understanding of the uptake of GRPr ligands on the cellular level.

Project description:The gastrin-releasing peptide receptor (GRPR), a G protein-coupled receptor, is overexpressed in solid malignancies and particularly in prostate cancer. We synthesized a novel bombesin derivative, [68Ga]Ga-ProBOMB1, evaluated its pharmacokinetics and potential to image GRPR expression with positron emission tomography (PET), and compared it with [68Ga]Ga-NeoBOMB1. ProBOMB1 (DOTA-pABzA-DIG-d-Phe-Gln-Trp-Ala-Val-Gly-His-Leu-ψ(CH2N)-Pro-NH2) was synthesized by solid-phase peptide synthesis. The polyaminocarboxylate chelator 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) was coupled to the N-terminal and separated from the GRPR-targeting sequence by a p-aminomethylaniline-diglycolic acid (pABzA-DIG) linker. The binding affinity to GRPR was determined using a cell-based competition assay, whereas the agonist/antagonist property was determined with a calcium efflux assay. ProBOMB1 was radiolabeled with 68GaCl3. PET imaging and biodistribution studies were performed in male immunocompromised mice bearing PC-3 prostate cancer xenografts. Blocking experiments were performed with coinjection of [d-Phe6,Leu-NHEt13,des-Met14]bombesin(6-14). Dosimetry calculations were performed with OLINDA software. ProBOMB1 and the nonradioactive Ga-ProBOMB were obtained in 1.1 and 67% yield, respectively. The K i value of Ga-ProBOMB1 for GRPR was 3.97 ± 0.76 nM. Ga-ProBOMB1 behaved as an antagonist for GRPR. [68Ga]Ga-ProBOMB1 was obtained in 48.2 ± 10.9% decay-corrected radiochemical yield with 121 ± 46.9 GBq/μmol molar activity and >95% radiochemical purity. Imaging/biodistribution studies showed that the excretion of [68Ga]Ga-ProBOMB1 was primarily through the renal pathway. At 1 h postinjection (p.i.), PC-3 tumor xenografts were clearly delineated in PET images with excellent contrast. The tumor uptake for [68Ga]Ga-ProBOMB1 was 8.17 ± 2.57 percent injected dose per gram (% ID/g) and 9.83 ± 1.48% ID/g for [68Ga]Ga-NeoBOMB1, based on biodistribution studies at 1 h p.i. This corresponded to tumor-to-blood and tumor-to-muscle uptake ratios of 20.6 ± 6.79 and 106 ± 57.7 for [68Ga]Ga-ProBOMB1 and 8.38 ± 0.78 and 39.0 ± 12.6 for [68Ga]Ga-NeoBOMB1, respectively. Blockade with [d-Phe6,Leu-NHEt13,des-Met14]bombesin(6-14) significantly reduced the average uptake of [68Ga]Ga-ProBOMB1 in tumors by 62%. The total absorbed dose was lower for [68Ga]Ga-ProBOMB1 in all organs except for bladder compared with [68Ga]Ga-NeoBOMB1. Our data suggest that [68Ga]Ga-ProBOMB1 is an excellent radiotracer for imaging GRPR expression with PET. [68Ga]Ga-ProBOMB1 achieved a similar uptake as [68Ga]Ga-NeoBOMB1 in tumors, with enhanced contrast and lower whole-body absorbed dose.

Project description:(90)Y has been used to label various new therapeutic radiopharmaceuticals. However, measuring the radiation dose delivered by (90)Y is challenging because of the absence of suitable ? emissions and its low abundance of positron emissions. For the treatment of prostate cancer, radiolabeled gastrin-releasing peptide receptor (GRPr) antagonists have yielded promising results in mouse models. In this study, we evaluated whether Cerenkov luminescence imaging (CLI) could be used to determine radiation doses of a (90)Y-labeled GRPr antagonist in nude mice.Mice bearing subcutaneous prostate cancer xenografts were injected with 0.74-18.5 MBq of the (90)Y-labeled GRPr antagonist DOTA-AR and underwent in vivo and ex vivo CLI at 1-48 h after injection. After imaging, animals were sacrificed, their tumors and organs were harvested, and the activity concentration was measured by liquid scintillation counting. In a second set of experiments, Cerenkov photon counts for tumor and kidney on in vivo CLI were converted to activity concentrations using conversion factors determined from the first set of experiments.(90)Y-DOTA-AR concentration in the 3 tumor models ranged from 0.5% to 4.8% of the injected activity per gram at 1 h after injection and decreased to 0.05%-0.15 injected activity per gram by 48 h after injection. A positive correlation was found between tumor activity concentrations and in vivo CLI signal (r(2) = 0.94). A similar correlation was found for the renal activity concentration and in vivo Cerenkov luminescence (r(2) = 0.98). Other organs were not distinctly visualized on the in vivo images, but ex vivo CLI was also correlated with the radioactivity concentration (r(2) = 0.35-0.94). Using the time-activity curves from the second experiment, we calculated radiation doses to tumor and kidney of 0.33 ± 0.12 (range, 0.21-0.66) and 0.06 ± 0.01 (range, 0.05-0.08) Gy/MBq, respectively.CLI is a promising, low-cost modality to measure individual radiation doses of (90)Y-labeled compounds noninvasively. The use of Cerenkov imaging is expected to facilitate the development and comparison of (90)Y-labeled compounds for targeted radiotherapy.

Dataset Information

A high-affinity, high-stability photoacoustic agent for imaging gastrin-releasing peptide receptor in prostate cancer.

Publications

A high-affinity, high-stability photoacoustic agent for imaging gastrin-releasing peptide receptor in prostate cancer.

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