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Integrin receptor αvβ3 and gastrin-releasing peptide receptor (GRPR) expression of tumors could be detected using PET imaging with radiolabeled Arg-Gly-Asp (RGD) and the antagonistic bombesin analog RM26, respectively. The purpose of this study was to investigate the dual receptor-targeting property of the heterodimer RGD-RM26-03 (denoted as LNC1015), demonstrate the tumor diagnostic value of [68Ga]Ga-LNC1015 in preclinical experiments, and evaluate its preliminary clinical feasibility. Methods: LNC1015 was designed and synthesized by linking cyclic RGD and the RM26 peptide. Preclinical pharmacokinetics were detected in a PC3 xenograft model using microPET and biodistribution studies. The clinical feasibility of [68Ga]Ga-LNC1015 PET/CT was performed in patients with breast cancer, and the results were compared with those of 18F-fluorodeoxyglucose (FDG). Results: [68Ga]Ga-LNC1015 had good stability in saline for at least 2 h, and favorable binding affinity and specificity were demonstrated in vitro and in vivo. The tumor uptake and retention of [68Ga]Ga-LNC1015 during PET imaging were improved compared with its monomeric counterparts [68Ga]Ga-RGD and [68Ga]Ga-RM26 at all the time points examined. In our initial clinical studies, the tumor uptake and tumor-to-background ratio (TBR) of primary and metastatic lesions in [68Ga]Ga-LNC1015 PET/CT were significantly higher than those in [18F]FDG PET/CT, resulting in high lesion detection rate and tumor delineation. Conclusion: The dual targeting radiotracer [68Ga]Ga-LNC1015 showed significantly improved tumor uptake and retention, as well as lower liver uptake than [68Ga]Ga-RGD and [68Ga]Ga-RM26 monomer. The first-in-human study showed high TBRs in patients, suggesting favorable pharmacokinetics and high clinical feasibility for PET/CT imaging of cancer. [ABSTRACT FROM AUTHOR]

Due to the heterogeneity of tumors, strategies to improve the effectiveness of dual-targeting tracers in tumor diagnostics have been intensively practiced. In this study, the radiolabeled [18F]AlF-NOTA-FAPI-RGD (denoted as [18F]AlF-LNC1007), a dual-targeting heterodimer tracer targeting both fibroblast activation protein (FAP) and integrin αvβ3 to enhance specific tumor uptake and retention, was synthesized and evaluated. The tracer was compared with [68Ga]Ga-LNC1007 in preclinical and clinical settings. Methods: The preparation of [18F]AlF- and 68Ga-labeled FAPI-RGD was carried out with an optimized protocol. The stability was tested in PBS and fetal bovine serum (FBS). Cellular uptake and in vivo distribution of the two products were compared and carried out on the U87MG cell line and its xenograft model. The safety and dosimetry of [18F]AlF-LNC1007 PET/CT scan were evaluated in six patients with malignant tumors. Results: Two radiolabeling protocols of [18F]AlF-/[68Ga]Ga-LNC1007 were developed and optimized to give a high yield of tracers with good stability. In vivo microPET images showed that the two tracers exhibited comparable pharmacokinetic characteristics, with high tumor uptake and prolonged tumor retention. In vivo distribution data showed that the target-to-non-target ratios of [18F]AlF-LNC1007 were similar to[68Ga]Ga-LNC1007. A total of six patients underwent [18F]AlF-LNC1007 PET/CT evaluation while two had head-to-head [18F]FDG PET/CT scans. The total body effective dose was 9.94E-03 mSv/MBq. The biodistribution curve showed optimal normal organ uptake with high tumor uptake and long retention of up to 3h p.i., and notably, the tumor-to-background ratio increased over time. Conclusion: We successfully prepared an [18F]AlF-LNC1007 dual-targeting PET probe with comparable performances as [68Ga]Ga-LNC1007. With prolonged tumor retention and tumor specificity, it produced good imaging quality in preclinical and clinical translational studies, indicating that [18F]AlF-LNC1007 is a promising non-invasive tracer for detecting tumors expressing FAP and/or integrin avβ3, with the prospect of clinical implementation. [ABSTRACT FROM AUTHOR]

Purpose: To compare the potential efficiency of [68Ga]Ga-LNC1007 with 2-[18F]FDG/[68Ga]Ga-PSMA PET/CT for detecting renal cell carcinoma (RCC) and to explore parameters derived from [68Ga]Ga-LNC1007 PET/CT for discriminating pathological characteristics in RCC. Methods: Twenty-five RCC patients confirmed by pathology were enrolled in this prospective study. The maximum standardized uptake value (SUVmax), mean SUV (SUVmean), gross tumor volume (GTV) and total lesion-tracer (TL-tracer) of lesions were calculated from the corresponding PET/CT images. Pathological characteristics included World Health Organization/International Society of Urological Pathology (WHO/ISUP) grade and adverse pathological features (tumor necrosis or sarcomatoid or rhabdoid feature). Results: [68Ga]Ga-LNC1007 PET/CT showed a higher detection rate for primary lesions than 2-[18F]FDG and [68Ga]Ga-PSMA (LNC1007 vs. FDG: 13/17 vs. 4/17, P = 0.005; LNC1007 vs. PSMA: 9/11 vs. 6/11, P = 0.361). [68Ga]Ga-LNC1007 PET/CT showed higher SUVmax (6.6 vs. 3.7, P = 0.005), SUVmean (4.1 vs. 2.3, P = 0.001) and TBR (2.6 vs. 1.7, P = 0.011) compared with 2-[18F]FDG PET/CT, and it also showed higher TBR (2.9 vs. 0.5, P = 0.003), TBR-delay (2.8 vs. 0.3, P = 0.003), GTV (84.1 vs. 42.9, P = 0.003) and TL-tracer (442.7 vs. 235.8, P = 0.008) compared with [68Ga]Ga-PSMA PET/CT. SUVmax and TBR derived from [68Ga]Ga-LNC1007 PET/CT could effectively differentiate WHO/ISUP grade (3–4 vs. 1–2) and adverse pathological features (positive vs. negative) (SUVmax: AUC 0.81, P = 0.04; AUC 0.80, P = 0.033; TBR: AUC 0.84, P = 0.026; AUC 0.85, P = 0.014). The SUVmax was positively correlated with the FAP expression, integrin αvβ3 expression and the total expression of FAP and integrin αvβ3 (r = 0.577, P = 0.006, r = 0.701, P < 0.001, and r = 0.702, P < 0.001, respectively). Conclusion: [68Ga]Ga-LNC1007 is a promising tracer for RCC imaging and can effectively identify aggressive pathological characteristics of RCC. [ABSTRACT FROM AUTHOR]

Purpose: The integrin αvβ3 and aminopeptidase N (APN/CD13) play vital roles in the tumor angiogenesis process. They are highly expressed in a variety of tumor cells and proliferating endothelial cells during angiogenesis, which have been considered as highly promising targets for tumor imaging. Arginine-glycine-aspartic (RGD) and asparagine-glycine-arginine (NGR) are two peptides specifically binding to the integrin αvβ3 and CD13, respectively. In this study, we optimized our previously developed probe and preclinically evaluated the new integrin αvβ3 and CD13 dual-targeted probe, NOTA-RGD-NGR (denoted as HX01) radiolabeled with 68Ga, in 10 different subcutaneous and orthotopic tumor models. Methods: The specific activity and radiochemical purity of [68Ga]Ga-HX01 were identified. The dual-receptor targeting ability was confirmed by a series of blocking studies and partly muted tracers using BxPC-3 xenograft model. The dynamic imaging study and dose escalation study were explored to determine the optimal imaging time point and dosage in the BxPC-3 xenograft model. Next, we established a variety of subcutaneous and orthotopic tumor models including pancreas (BxPC-3), breast (MCF-7), gallbladder (NOZ), lung (HCC827), ovary (SK-OV-3), colorectal (HCT-8), liver (HuH-7), stomach (NUGC-4), and glioma (U87) cancers. All models underwent [68Ga]Ga-HX01 PET/CT imaging about 2 weeks post-inoculation, with a subset of them undergoing [18F]FDG PET/CT scan performed concurrently, and their results were compared. In addition, ex vivo biodistribution studies were also performed for verifying the semi-quantitative results of the non-invasive PET images. Results: [68Ga]Ga-HX01 significantly outperformed single target probes in the BxPC-3 xenograft model. All blocking and single target groups exhibited significantly descending tumor uptake. The high tumor uptakes were found in BxPC-3, MCF-7, and NOZ subcutaneous tumors (%ID/g > 1.1), while middle uptakes were observed in HCC827, SK-OV-3, HCT-8, and HuH-7 subcutaneous tumor (%ID/g 0.7–1.0). Due to the low background, the tumor-to-muscle and tumor-to-blood ratios of [68Ga]Ga-HX01 were higher than that of [18F]FDG. Conclusions: [68Ga]Ga-HX01, as a dual target imaging agent, exhibited superior in vivo performance in different subcutaneous and orthotopic mice models of human tumors over [18F]FDG and its respectively mono-receptor targeted agents, which warrants the future clinical translation for tumor imaging. [ABSTRACT FROM AUTHOR]

Background: The primary aims of this study were to compare in patients with esophageal or esophagogastric junction cancers the potential of 68Ga-NODAGA-RGD PET/CT with that of 18F-FDG PET/CT regarding tumoral uptake and distribution, as well as histopathologic examination. Methods: Ten 68Ga-NODAGA-RGD and ten 18F-FDG PET/CT were performed in nine prospectively included participants (1 woman; aged 58 ± 8.4 y, range 40–69 y). Maximum SUV (SUVmax) and metabolic tumor volumes (MTV) were calculated. The Mann–Whitney U test and Spearman correlation analysis (ρ) were used. Results: 68Ga-NODAGA-RGD PET/CT detected positive uptake in 10 primary sites (8 for primary tumors and 2 for local relapse suspicion), 6 lymph nodes and 3 skeletal sites. 18F-FDG PET/CT detected positive uptake in the same sites but also in 16 additional lymph nodes and 1 adrenal gland. On a lesion-based analysis, SUVmax of 18F-FDG was significantly higher than those of 68Ga-NODAGA-RGD (4.9 [3.7–11.3] vs. 3.2 [2.6–4.2] g/mL, p = 0.014). Only one participant showed a higher SUVmax in an osseous metastasis with 68Ga-NODAGA-RGD as compared to 18F-FDG (6.6 vs. 3.9 g/mL). Correlation analysis showed positive correlation between 18F-FDG and 68Ga-NODAGA-RGD PET parameters (ρ = 0.56, p = 0.012 for SUVmax, ρ = 0.78, p < 0.001 for lesion-to-background ratios and ρ = 0.58, p = 0.024 for MTV). We observed that 18F-FDG uptake was homogenous inside all the confirmed primary sites (n = 9). In contrast, 68Ga-NODAGA-RGD PET showed more heterogenous uptake in 6 out of the 9 confirmed primary sites (67%), seen mostly in the periphery of the tumor in 5 out of the 9 confirmed primary sites (56%), and showed slight extensions into perilesional structures in 5 out of the 9 confirmed primary sites (56%). Conclusions: In conclusion, 68Ga-NODAGA-RGD has lower potential in the detection of esophageal or esophagogastric junction malignancies compared to 18F-FDG. However, the results suggest that PET imaging of integrin αvβ3 expression may provide complementary information and could aid in tumor diversity and delineation. Trial registration: Trial registration: NCT02666547. Registered January 28, 2016—Retrospectively registered, https://clinicaltrials.gov/ct2/show/NCT02666547. [ABSTRACT FROM AUTHOR]

To investigate the role of the positron emission tomography (PET) tracer 18F-Alfatide labeled dimerized-RGD (18F-Alfatide-RGD) in non-small cell lung cancer (NSCLC). Forty-eight NSCLC patients were retrospectively enrolled. Evaluation of diagnostic test was used to assess the diagnostic performance of 18F-Alfatide-RGD PET/CT. 18F-Alfatide-RGD PET/CT showed general consistency in diagnosis and pathologic results of primary foci in lung cancer, and high consistency in detection and pathologic results of patients with lymph node-positive and metastatic lymph nodes. 18F-Alfatide-RGD PET/CT in the initial diagnosis of NSCLC has a high sensitivity and specificity, contributing to accurate clinical staging and preoperative evaluation. [ABSTRACT FROM AUTHOR]

Nowadays, pancreatic cancer is still a formidable disease to diagnose. The CXC chemokine receptor 4 (CXCR4) and integrin αvβ3 play important roles in tumor development, progression, invasion, and metastasis, which are overexpressed in many types of human cancers. In this study, we developed a heterodimeric tracer 68Ga-yG5-RGD targeting both CXCR4 and integrin αvβ3, and evaluated its feasibility and utility in PET imaging of pancreatic cancer. The 68Ga-yG5-RGD could accumulate in CXCR4/integrin αvβ3 positive BxPC3 tumors in a high concentration and was much higher than that of 68Ga-yG5 (p < 0.001) and 68Ga-RGD (p < 0.001). No increased uptake of 68Ga-yG5-RGD was found in MX-1 tumors (CXCR4/integrin αvβ3, negative). In addition, the uptake of 68Ga-yG5-RGD in BxPC3 was significantly blocked by excess amounts of AMD3100 (an FDA-approved CXCR4 antagonist) and/or unlabeled RGD (p < 0.001), confirming its dual-receptor targeting properties. The ex vivo biodistribution and immunohistochemical results were consistent with the in vivo imaging results. The dual-receptor targeting strategy achieved improved tumor-targeting efficiency and prolonged tumor retention in BxPC3 tumors, suggesting 68Ga-yG5-RGD is a promising tracer for the noninvasive detection of tumors that express either CXCR4 or integrin αvβ3 or both, and therefore may have good prospects for clinical translation. [ABSTRACT FROM AUTHOR]

The adverse events during antiangiogenic therapy inevitably influence a patient's quality of life. Therefore, biomarkers to identify patients who will experience adverse events would be very valuable in treatment planning. Between September 2016 and December 2019, patients scheduled for single-agent apatinib were prospectively enrolled and underwent 18F-RGD positron emission tomography/computed tomography (PET/CT) pre-treatment. Maximum and mean standard uptake values (SUV max and SUV mean) were obtained from thyroid, liver, gastric cardia, gastric body, gastric pylorus and spleen. Statistical methods included the independent sample t -test, Mann-Whitney U-test, receiver operating characteristic curve analysis and chi-squared test. In total, 60 patients were initially screened and consented for 18F-RGD PET/CT scans. The three most frequent adverse events were fatigue (50%), hypertension (36%) and nausea (36%), accounting for 72% in the 50 patients included in the analysis. SUV max and SUV mean of thyroid and liver were significantly associated with fatigue, whereas SUV max and SUV mean of thyroid and spleen were significantly associated with hypertension and SUV max and SUV mean of thyroid and gastric cardia were significantly associated with nausea (all P < 0.05). The most significant predictors of adverse events were 18F-RGD SUV max-liver for fatigue (area under the curve [AUC] = 0.682), SUV max-spleen for hypertension (AUC = 0.688) and SUV max-gastric cardia for nausea (AUC = 0.698). Classified by the cut-off values for SUV max-liver (4.57), SUV max-spleen (6.77) and SUV max-gastric cardia (2.10), patients with low RGD SUV max in liver, spleen and gastric cardia had statistically higher incidence of fatigue (67.9% versus 27.3%, P = 0.002), hypertension (55.6% versus 13.0%, P = 0.004) and nausea (61.1% versus 21.9%, P = 0.006). Low pre-treatment 18F-RGD uptake in the liver, spleen and gastric cardia were predictive of the adverse events fatigue, hypertension and nausea during apatinib treatment, respectively. • The adverse events during antiangiogenic therapy inevitably influence the quality of life, and no biomarkers were available to identify those high-risk patients. • Compared with the patients with high SUV max , those patients with low RGD in liver, spleen and gastric cardia had statistically higher incidence of fatigue, hypertension and nausea during antiangiogenic therapy. • Those biomarkers to identify patients who will experience adverse events would be useful for high-risk patients to decrease the occurrence of antiangiogenic AEs by early intervention with drugs or physical treatment. [ABSTRACT FROM AUTHOR]

Ovarian cancer has the highest mortality rate of gynecologic malignancy. 18F-FDG positron emission tomography (PET) adds an important superiority over traditional anatomic imaging modalities in oncological imaging but has drawbacks including false negative results at the early stage of ovarian cancer, and false positives when inflammatory comorbidities are present. Aminopeptidase N (APN, also known as CD13) and integrin αvβ3 are two important targets overexpressed on tumor neo-vessels and frequently on ovarian cancerous cells. In this study, we used subcutaneous and metastatic models of ovarian cancer and muscular inflammation models to identify 68Ga-NGR-RGD, a heterodimeric tracer consisting of NGR and RGD peptides targeting CD13 and integrin αvβ3, respectively, and compared it with 18F-FDG. We found that 68Ga-NGR-RGD showed greater contrast in SKOV3 and ES-2 tumors than 18F-FDG. Low accumulation of 68Ga-NGR-RGD but avid uptake of 18F-FDG were observed in inflammatory muscle. In abdominal metastasis models, PET imaging with 68Ga-NGR-RGD allowed for rapid and clear delineation of both peritoneal and liver metastases (3-6 mm), whereas, 18F-FDG could not distinguish the metastasis lesions due to the relatively low metabolic activity in tumors and the interference of intestinal physiological 18F-FDG uptake. Due to the high tumor-targeting efficacy, low inflammatory uptake, and higher tumor-to-background ratios compared to that of 18F-FDG, 68Ga-NGR-RGD presents a promising imaging agent for diagnosis, staging, and follow-up of ovarian tumors. [ABSTRACT FROM AUTHOR]

Introduction: Non-invasive imaging techniques (especially single-photon emission tomography and positron emission tomography) apply several RGD-based imaging ligands developed during a vast number of preclinical and clinical investigations. The RGD (Arg-Gly-Asp) sequence is a binding moiety for a large selection of adhesive extracellular matrix and cell surface proteins. Since the first identification of this sequence as the shortest sequence required for recognition in fibronectin during the 1980s, fundamental research regarding the molecular mechanisms of integrin action have paved the way for development of several pharmaceuticals and radiopharmaceuticals with clinical applications. Ligands recognizing RGD may be developed for use in the monitoring of these interactions (benign or pathological). Although RGD-based molecular imaging has been actively investigated for oncological purposes, their utilization towards non-oncology applications remains relatively under-exploited. Methods and Scope: This review highlights the new non-oncologic applications of RGD-based tracers (with the focus on single-photon emission tomography and positron emission tomography). The focus is on the last 10 years of scientific literature (2009–2020). It is proposed that these imaging agents will be used for off-label indications that may provide options for disease monitoring where there are no approved tracers available, for instance Crohn's disease or osteoporosis. Fundamental science investigations have made progress in elucidating the involvement of integrin in various diseases not pertaining to oncology. Furthermore, RGD-based radiopharmaceuticals have been evaluated extensively for safety during clinical evaluations of various natures. Conclusion: Clinical translation of non-oncological applications for RGD-based radiopharmaceuticals and other imaging tracers without going through time-consuming extensive development is therefore highly plausible. [ABSTRACT FROM AUTHOR]

Introduction: The advent of the Germanium-68 (Ge-68)/Gallium-68 (Ga-68) generator has contributed enormously to a plethora of molecular imaging approaches for in vivo identification of tumor characteristics. The present study compares the effect of 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) and 1,4,7-triazacyclononane,1-gluteric acid-4,7-acetic acid (NODAGA) bifunctional chelators on radiolabeling of arginine-glycine-aspartic acid (RGD) dimer, an antagonist of integrin αvβ3 with Ga-68 and their biodistribution in C57BL/6 mice bearing melanoma and in patients with breast carcinoma. Methods: Radiolabeling parameters for DOTA-(RGD)2 and NODAGA-(RGD)2 with Ga-68 were optimized in-house. After quality control procedures, preclinical studies were done in C57BL/6 mice bearing melanoma. The percent radioactivity associated with per gram of various organs and tumor (% ID/g) was analyzed. Positron emission tomography-computed tomography patient imaging was performed in clinically diagnosed locally advanced breast carcinoma patients (n = 30). The uptake of various organs and lesions for both radiotracers was compared. Results: Radiolabeling yield >95% was obtained by heating 15-20 μg of peptide at 95°C for 5-10 min and 3.5-4.0 reaction pH. NODAGA-(RGD)2 could also be radiolabeled at room temperature, but 40-50 μg peptide was required. Animal biodistribution study revealed the kidney as the major excretory organ for both the radiotracers. Maximum counts were observed in tumor at 45 min. During the clinical study, liver, spleen, bilateral brain ventricles, salivary glands, and intestines were the organs with physiological uptake of both Ga-68-DOTA-(RGD)2 and Ga-68 NODAGA-(RGD)2. The major excretory route was through kidneys. All primary lesions were picked by both the radiotracers. Additionally, in 5 patients, metastatic lesions were also picked up. Conclusion: DOTA- and NODAGA-chelated RGD2 were successfully radiolabeled with Ga-68. Good tumor to background contrast exhibited by Ga-68-DOTA-(RGD)2 and Ga-68 NODAGA-(RGD)2 in both preclinical and clinical studies suggested that both radiotracers can be used as potential molecular tools for imaging angiogenesis. [ABSTRACT FROM AUTHOR]

Purpose: To explore the relationship between metabolic uptake of the 18F-ALF-NOTA-PRGD2 (18F-RGD) tracer on positron emission tomography/computerized tomography (PET/CT) and the antiangiogenic effect of apatinib in patients with solid malignancies. Materials and patients: Patients with measurable lesions scheduled for second- or third-line single-agent therapy with apatinib were eligible for this prospective clinical trial. All patients underwent 18F-RGD PET/CT examination before the start of treatment. Standardized uptake values (SUVs) of contoured tumor lesions were computed and compared using independent sample t-tests or the Mann–Whitney U test. Receiver-operating characteristic (ROC) curve analysis was used to determine accuracy in predicting response. Survival curves were compared using the Kaplan–Meier method. Results: Of 38 patients who consented to study participation, 25 patients with 42 measurable lesions met the criteria for inclusion in this response assessment analysis. The median follow-up time was 3 months (range, 1–10 months), and the median progression-free survival (PFS) was 3 months (95% confidence interval, 1.04–4.96). The SUVpeak and SUVmean were significantly higher in responding tumors than in non-responding tumors (4.98 ± 2.34 vs 3.59 ± 1.44, p = 0.048; 3.71 ± 1.15 vs 2.95 ± 0.49, P = 0.036). SUVmax did not differ between responding tumors and non-responding tumors (6.58 ± 3.33 vs 4.74 ± 1.83, P = 0.078). An exploratory ROC curve analysis indicated that SUVmean [area under the ROC curve (AUC) = 0.700] was a better parameter than SUVpeak (AUC = 0.689) for predicting response. Using a threshold value of 3.82, high SUVmean at baseline was associated with improved PFS (5.0 vs. 3.4 months, log-rank P = 0.036). Conclusion: 18F-RGD uptake on PET/CT imaging pretreatment may predict the response to antiangiogenic therapy, with higher 18F-RGD uptake in tumors predicting a better response to apatinib therapy. [ABSTRACT FROM AUTHOR]

Radiolabeled heterodimeric peptide has emerged as a highly promising targeting strategy for PET imaging due to their superior properties. RGD and GE11 are two peptides binding to receptor integrin α v β 3 and EGFR, respectively, which both overexpress in many different types of tumors. This study focuses on the synthesis and evaluation of a RGD and GE11-containing heterodimeric radiotracer [64Cu]Cu-NOTA-RGD-GE11 for PET imaging of tumors that simultaneously overexpress integrin α v β 3 and EGFR. [64Cu]Cu-NOTA-RGD-GE11 was prepared by the conjugation of RGD-PEG 4 -NOTA-N 3 and GE11-PEG 4 -BCN via metal-free click chemistry, followed by radiolabeling with 64Cu. Cell uptake and efflux studies, saturation binding assay, the animal PET/CT and biodistribution studies were conducted to characterize the biological properties of [64Cu]Cu-NOTA-RGD-GE11. [64Cu]Cu-NOTA-RGD-GE11 was synthesized with a radiochemical purity of >97 % and molar activity of 23 GBq/μmol at the end of synthesis. [64Cu]Cu-NOTA-RGD-GE11 showed moderate hydrophilicity, good stability in mouse serum and high specific uptake by the human pancreatic cancer cell line (BxPC3) in the in vitro studies. Compared to the two monomeric counterparts [64Cu]Cu-NOTA-RGD and [64Cu]Cu-NOTA-GE11, [64Cu]Cu-NOTA-RGD-GE11 demonstrated significantly improved tumor uptakes (e.g. 4.63 ± 0.25 %ID/g vs 1.24 ± 0.18 %ID/g and 0.77 ± 0.13 %ID/g, 2 h after injection, p < 0.05) in the subsequent in vivo evaluation in mice bearing BxPC3 xenograft. Tumor uptake could be blocked in the presence of both non-radioactive c(RGDyK) and GE11 peptides, indicating good tumor specificity of [64Cu]Cu-NOTA-RGD-GE11 in vivo. The results suggested that the as-developed [64Cu]Cu-NOTA-RGD-GE11 could serve as a potential PET tracer for the noninvasive imaging of integrin α v β 3 and EGFR expression in tumors. A RGD and GE11-containing heterodimeric radiotracer [64Cu]Cu-NOTA-RGD-GE11 was successfully designed and synthesized in this study. [64Cu]Cu-NOTA-RGD-GE11 showed moderate hydrophilicity, good stability in mouse serum and high specific uptake by the human pancreatic cancer cell line (BxPC3) in the in vitro studies. Compared to the corresponding monomeric counterparts ([64Cu]Cu-NOTA-RGD and [64Cu]Cu-NOTA-GE11), [64Cu]Cu-NOTA-RGD-GE11 demonstrated improved tumor uptakes in the following in vivo evaluation in mice bearing BxPC3 xenograft. The tumor uptake could be blocked in the presence of both non-radioactive c(RGDyK) and GE11 peptides, indicating good tumor specificity of [64Cu]Cu-NOTA-RGD-GE11 in vivo. The results revealed that [64Cu]Cu-NOTA-RGD-GE11 could serve as a potential PET tracer for the noninvasive imaging of integrin α v β 3 and EGFR expression in tumors. [Display omitted] [ABSTRACT FROM AUTHOR]

Activation of the vitronectin receptor αvβ3 and the phosphorylation of extracellular signal-regulated kinase (ERK)1/2 are critical events during tumor development and progression. The aim of the present study was to investigate the effects of WD-3, a formula used in traditional Chinese medicine, on integrin αvβ3 and ERK1/2 expression in vivo using a nude mouse-human gastric cancer xenograft model combined with non-invasive, real-time 18F-Arg-Gly-Asp (RGD) positron emission tomography (PET)/computerized tomography (CT) imaging methods. SGC-7901 human gastric cancer cells were subcutaneously injected into BALB/c nude mice. Following tumor development, animals were randomly assigned into the following 4 groups (n=6 mice/group): Control group (CG), Chinese medicine group (CMG), Western medicine group (WMG) and Chinese and Western medicine combination group (CMG + WMG). Mice in the CG and CMG received daily intragastric injections of 0.5 ml saline and 0.5 ml WD-3, respectively. Mice in the WMG received an intravenous injection of albumin-bound paclitaxel (25 mg/kg) on days 0, 2 and 4 Mice in the CMG + WMG received combination therapy of WD-3 and albumin-bound paclitaxel. Tumor growth was monitored using standard caliper technique and via PET imaging. 18F-RGD PET/CT analysis was performed on days 3, 7, 18 and 24 following drug administration. Radioactivity uptake was measured and expressed as the percentage of injected dose (ID) per tissue weight (%ID/g) and the standardized uptake value (SUV). Animals were sacrificed at 30 days following treatment and tumor weight was measured. Immunohistochemistry was used to detect the expression of phosphorylated (p)-ERK1/2 protein in tumor tissue samples. No statistically significant differences were observed in %ID/g and SUV among the various groups prior to treatment. At the end of treatment, mice in the CMG, WMG and CMG + WMG exhibited significantly reduced tumor mass when compared with mice in the CG. In addition, mice in the CMG and CMG + WMG demonstrated reduced %ID/g and SUV when compared with mice in the CG. Conversely, mice in the WMG exhibited no significant difference in %ID/g and SUV compared with the CG. Furthermore, p-ERK1/2 expression was significantly reduced in mice from all treatment groups when compared with those in the CG. The results of the present study suggest that the traditional Chinese formula WD-3 may inhibit gastric tumor growth, potentially via the downregulation of integrin αvβ3 and the inhibition of ERK1/2 phosphorylation in vivo. [ABSTRACT FROM AUTHOR]

The present article describes the development of robust lyophilized kit for convenient formulation of [68Ga]Ga-DOTA-E−[c(RGDfK)] 2 (E = glutamic acid, R = arginine, G = glycine, D = aspartic acid, f = phenylalanine, K = lysine) radiopharmaceutical for clinical use in non-invasive monitoring of malignancies overexpressing integrin α v β 3 receptors. Five batches of the kit were prepared with optimized kit contents, all of which showed high 68Ga-radiolabeling yield (>98%). Pre-clinical evaluation of the [68Ga]Ga-radiotracer in SCID mice bearing FTC133 tumour exhibited significant accumulation in the tumor xenograft. Preliminary human clinical investigation carried out in a 60 year old male patient with metastatic lung cancer revealed high radiotracer uptake in the tumor along with satisfactory target to non-target contrast. The developed kit formulation also showed a long shelf-life of at least 12 months on storage at 0 °C. All these results point towards the promising attributes of the developed kit formulation for convenient preparation of [68Ga]Ga-DOTA-E−[c(RGDfK)] 2 for routine clinical use. • A lyophilized kit of DOTA-E−[c(RGDfK)] 2 developed. • It enabled convenient formulation of [68Ga]Ga-DOTA-E−[c(RGDfK)] 2 for routine clinical use. • Pre-clinical PET-CT imaging showed significant accumulation of the radiotracer in tumor. • Preliminary human clinical investigations showed good uptake of the radiotracer in cancer lesions and satisfactory image contrast. [ABSTRACT FROM AUTHOR]

Purpose: The study aims to investigate the role of F-alfatide positron emission tomography/computed tomography (PET/CT) in predicting the short-term outcome of concurrent chemoradiotherapy (CCRT) in patients with advanced non-small cell lung cancer (NSCLC). Methods: Eighteen patients with advanced NSCLC had undergone F-alfatide PET/CT scans before CCRT and PET/CT parameters including maximum and mean standard uptake values (SUV/SUV), peak standard uptake values (SUV) and tumor volume (TV and TV) were obtained. The SUV of tumor and normal tissues (lung, blood pool and muscle) were measured, and their ratios were denoted as T/NT (T/NT, T/NT and T/NT). Statistical methods included the Two-example t test, Wilcoxon rank-sum test, Receiver-operating characteristic (ROC) curve analysis and logistic regression analyses. Results: We found that SUV, SUV, T/NT, T/NT and T/NT were higher in non-responders than in responders ( P = 0.0024, P = 0.016, P < 0.001, P = 0.003, P = 0.004). According to ROC curve analysis, the thresholds of SUV, SUV, T/NT, T/NT and T/NT were 5.65, 4.46, 7.11, 5.41, and 11.75, respectively. The five parameters had high sensitivity, specificity and accuracy in distinguishing non-responders and responders. Multivariate logistic regression analyses showed that T/NT was an independent predictor of the short-term outcome of CCRT in patients with advanced NSCLC ( P = 0.032). Conclusions: F-alfatide PET/CT may be useful in predicting the short-term outcome of CCRT in patients with advanced NSCLC. [ABSTRACT FROM AUTHOR]

Background/Aim: We prospectively investigated the potential usefulness of PET using a new tracer targeting integrin αvβ3 (termed RGD-K5) in patients with head and neck cancer (HNC) undergoing definitive concurrent chemoradiotherapy (CCRT). Patients and Methods: Newly diagnosed patients with locally advanced HNC scheduled for definitive CCRT were eligible. RDG-K5 PET and FDG PET scans were performed at three different time points (baseline, 2 weeks, and 3 months post-treatment). Results: Nine patients completed all of the three scans, whereas two patients withdrew after two scans only. Uptake of both RGD-K5 and FDG generally decreased following CCRT. However, the observed decrease did not differ significantly between complete responders and non-responders. At 3 months post-treatment, the uptake of both RGD-K5 and FDG at the main tumors was significantly lower in those who achieved complete responses than in those with residual tumors. Conclusion: RGD-K5 PET has the potential to identify patients with incomplete responses to CCRT. [ABSTRACT FROM AUTHOR]

Radiolabeled Arg-Gly-Asp (RGD) peptide analogs have been extensively studied for αvβ3 integrin-targeted angiogenesis imaging. According to recently presented evidence, the dodecapeptide GE11 has high affinity to the epidermal growth factor receptor (EGFR), which is overexpressed in many types of cancer. Dual-receptor molecular imaging probes with two different heterodimeric peptides exhibit improved cancer targeting efficacy. In the present study, the design and synthesis of a new RGD-GE11 peptide heterodimer for dual αvβ3 integrin/EGFR-targeted cancer imaging are described. The RGD-GE11 heterodimer was linked with 6-aminohexanoic acid (6-Ahx) and cysteine and conjugated with 1,4,7-triazacyclononane-N,N′,N″-triacetic acid (NOTA) to form NOTA-RGD-cys-6-Ahx-GE11. The monomeric peptides, NOTA-cys-6-Ahx-GE11 and c(RGDyK), were formed by a peptide synthesizer. The peptide heterodimer NOTA-RGD-GE11 was obtained by NOTA-cys-6-Ahx-GE11 and maleimidopropyl-c(RGDyK) conjugation with a thioether linkage. The NOTA peptide conjugate was labeled with freshly eluted 68Ga and purified using reversed-phase high-performance liquid chromatography. The 68Ga-NOTA-RGD-cys-6-Ahx-GE11 was successfully prepared, in this study, with a radiochemical yield of 85% and a radiochemical purity of >98%. These results warrant further investigation of this heterodimeric peptide's binding affinity to the receptors. [ABSTRACT FROM AUTHOR]

It is well recognized that carbon chirality plays a critical role in the design of drug molecules. However, very little information is available regarding the effect of stereoisomerism of macrocyclic bifunctional chelators (BFC) on biological behaviors of the corresponding radiopharmaceuticals. To evaluate such effects, three enantiopure stereoisomers of a copper radiopharmaceutical BFC bearing two chiral carbon atoms were synthesized in forms of R,R-, S,S-, and R,S-. Their corresponding peptide conjugates were prepared by coupling with a model peptide sequence, c(RGDyK), which targets the αvβ3 integrin for in vitro and in vivo evaluation of their biological behaviors as compared to the racemic conjugate. Despite the chirality differences, all the conjugates showed a similar in vitro binding affinity profile to the αvβ3 integrin (106, 108, 85 and 100 nM for rac-H2-1, RR-H2-1, SS-H2-1, and RS-H2-1 respectively with all p values > 0.05) and a similar level of in vivo tumor uptake (2.72 ± 0.45, 2.60 ± 0.52, 2.45 ± 0.48 and 2.88 ± 0.59 for rac- 64Cu-1, RR-64Cu-1, SS-64Cu-1, and RS-64Cu-1 at 1 h p.i. respectively). Furthermore, they demonstrated a nearly identical biodistribution pattern in major organs (e.g. 2.07 ± 0.21, 2.13 ± 0.58, 1.70 ± 0.20 and 1.90 ± 0.46 %ID/g at 24 h p.i. in liver for rac- 64Cu-1, RR-64Cu-1, SS-64Cu-1, and RS-64Cu-1 respectively; 1.80 ± 0.46, 2.30 ± 1.49, 1.73 ± 0.31 and 2.23 ± 0.71 at 24 h p.i. in kidneys for rac- 64Cu-1, RR-64Cu-1, SS-64Cu-1, and RS-64Cu-1 respectively). Therefore we conclude that the chirality of BFC plays a negligible role in αvβ3-targeted copper radiopharmaceuticals. However, we believe it is still worthwhile to consider the chirality effects of BFCs on other targeted imaging or therapeutic agents. [ABSTRACT FROM AUTHOR]

Abstract: Introduction: The aim of this study was to synthesize and perform a side-by-side comparison of two new tumor-angiogenesis PET tracers 68Ga-NODAGA-E[c(RGDyK)]2 and 64Cu-NODAGA-E[c(RGDyK)]2 in vivo using human xenograft tumors in mice. Human radiation burden was estimated to evaluate potential for future use as clinical PET tracers for imaging of neo-angiogenesis. Methods: A 68Ge/68Ga generator was used for the synthesis of 68Ga-NODAGA-E[c(RGDyK)]2. 68Ga and 64Cu labeled NODAGA-E[c(RGDyK)]2 tracers were administrated in nude mice bearing either human glioblastoma (U87MG) or human neuroendocrine (H727) xenograft tumors. PET/CT scans at 3 time points were used for calculating the tracer uptake in tumors (%ID/g), integrin αVβ3 target specificity was shown by blocking with cold NODAGA-E[c(RGDyK)]2, and biodistribution in normal organs were also examined. From biodistribution data in mice human radiation-absorbed doses were estimated using OLINDA/EXM software. Results: 68Ga-NODAGA-E[c(RGDyK)]2 was synthesized with a radiochemical purity of 89%–99% and a specific activity (SA) of 16–153MBq/nmol. 64Cu-NODAGA-E[c(RGDyK)]2 had a purity of 92%–99% and an SA of 64–78MBq/nmol. Both tracers showed similar uptake in xenograft tumors 1h after injection (U87MG: 2.23 vs. 2.31%ID/g; H727: 1.53 vs. 1.48%ID/g). Both RGD dimers showed similar tracer uptake in non-tumoral tissues and a human radiation burden of less than 10mSv with an administered dose of 200MBq was estimated. Conclusion: 68Ga-NODAGA-E[c(RGDyK)]2 and 64Cu-NODAGA-E[c(RGDyK)]2 can be easily synthesized and are both promising candidates for PET imaging of integrin αVβ3 positive tumor cells. 68Ga-NODAGA-E[c(RGDyK)]2 showed slightly more stable tumor retention. With the advantage of in-house commercially 68Ge/68Ga generators, 68Ga-NODAGA-E[c(RGDyK)]2 may be the best choice for future clinical PET imaging in humans. [Copyright &y& Elsevier]

RGD peptides, radiolabeled with F, have been used in the clinic for PET imaging of tumor angiogenesis in cancer patients. RGD peptides are typically labeled using a prosthetic group such as N-succinimidyl 4-[F]-fluorobenzoate ([F]SFB) or 4-nitrophenyl 2-[F]-fluoropropionate ([F]NPFP). However, the complex radiosynthetic procedures have impeded their broad application in clinical studies. We previously radiolabeled proteins and peptides with the prosthetic group, N-succinimidyl 4-[F]-fluoromethylbenzoate ([F]SFMB), which was prepared in a simple one-step procedure. In this study, we labeled a PEGylated cyclic RGD peptide dimer, PEG-E[c(RGDyK)] (PRGD2), using [F]SFMB and evaluated for imaging tumor αvβ3 integrin expression with positron emission tomography (PET). [F]SFMB was prepared in one step using [F]fluoride displacement of a nitrobenzenesulfonate leaving group under mild reaction conditions followed by HPLC purification. The F-labeled peptide, [F]FMBPRGD2 was prepared by coupling PRGD2 with [F]SFMB in pH 8.6 borate buffer and purified with HPLC. The direct labeling on BMBPRGD2 was also attempted. A Siemens Inveon PET was used to image the uptake of the [F]FMBPRGD2 into a U87MG xenograft mouse model. [F]FMBPRGD2, was prepared with a 15% overall radiochemical yield (uncorrected) in a total synthesis time of 90 min, which was considerably shorter than the preparation of [F]SFB- and [F]NPFP-labeled RGD peptides. The direct labeling, however, was not successful. High quality microPET images using [F]FMBPRGD2 clearly visualized tumors by 15 min with good target to background ratio. Early tracer accumulation in the bladder suggests fast renal clearance. No obvious bone uptake can be detected even at 4-h time point indicating that fluorine attachment is stable in mice. In conclusion, N-succinimidyl 4-[F]-fluoromethylbenzoate ([F]SFMB) prosthetic group can be a good alternative for labeling RGD peptides to image αvβ3 integrin expression and for labeling other peptides. [ABSTRACT FROM AUTHOR]

Purpose: One of the major obstacles of the clinical translation of F-labeled arginine-glycine-aspartic acid (RGD) peptides has been the laborious multistep radiosynthesis. In order to facilitate the application of RGD-based positron emission tomography (PET) probes in the clinical setting we investigated in this study the feasibility of using the chelation reaction between AlF and a macrocyclic chelator-conjugated dimeric RGD peptide as a simple one-step F labeling strategy for development of a PET probe for tumor angiogenesis imaging. Methods: Dimeric cyclic peptide E[ c(RGDyK)] (RGD) was first conjugated with a macrocyclic chelator, 1,4,7-triazacyclononane-1,4,7-triacetic acid (NOTA), and the resulting bioconjugate NOTA-RGD was then radiofluorinated via AlF intermediate to synthesize F-AlF-NOTA-RGD. Integrin binding affinities of the peptides were assessed by a U87MG cell-based receptor binding assay using I-echistatin as the radioligand. The tumor targeting efficacy and in vivo profile of F-AlF-NOTA-RGD were further evaluated in a subcutaneous U87MG glioblastoma xenograft model by microPET and biodistribution. Results: NOTA-RGD was successfully F-fluorinated with good yield within 40 min using the AlF intermediate. The IC of F-AlF-NOTA-RGD was determined to be 46 ± 4.4 nM. Quantitative microPET studies demonstrated that F-AlF-NOTA-RGD showed high tumor uptake, fast clearance from the body, and good tumor to normal organ ratios. Conclusion: NOTA-RGD bioconjugate has been successfully prepared and labeled with AlF in one single step of radiosynthesis. The favorable in vivo performance and the short radiosynthetic route of F-AlF-NOTA-RGD warrant further optimization of the probe and the radiofluorination strategy to accelerate the clinical translation of F-labeled RGD peptides. [ABSTRACT FROM AUTHOR]

Abstract: Labeling biomolecules with 18F is usually done through coupling with prosthetic groups, which generally requires several time-consuming radiosynthetic steps resulting in low labeling yield. Recently, the tetrazine-trans-cyclooctene ligation has been introduced as a method of bioconjugation that proceeds with fast reaction rates without need for catalysis. Herein, we report the development of an extremely fast and efficient method for generating 18F labeled probes based on the tetrazine-trans-cyclooctene ligation. Starting with only 30μg (78μM) of a tetrazine-RGD conjugate and 2mCi (5μM) of 18F-trans-cyclooctene, the 18F labeled RGD peptide could be obtained in more than 90% yield within five minutes. The 18F labeled RGD peptide demonstrated prominent tumor uptake in vivo. The receptor specificity was confirmed by blocking experiments. These results successfully demonstrate that the tetrazine-trans-cyclooctene ligation serves as an efficient labeling method for PET probe construction. [Copyright &y& Elsevier]

peptide heterodimer comprises two different receptor-targeting peptide ligands. Molecular imaging probes based on dual-receptor targeting peptide heterodimers exhibit improved tumor targeting efficacy for multi-receptor expressing tumors compared with their parent single-receptor targeting peptide monomers. Previously we have developed bombesin (BBN)-RGD (Arg-Gly-Asp) peptide heterodimers, in which BBN and RGD are covalently connected with an asymmetric glutamate linker (J Med Chem 52:425-432, 2009). Although F-labeled heterodimers showed significantly better microPET imaging quality than F-labeled RGD and BBN monomers in a PC-3 xenograft model which co-expresses gastrin-releasing peptide receptor (GRPR) and integrin αvβ3, tedious heterodimer synthesis due to the asymmetric nature of glutamate linker restricts their clinical applications. In this study, we report the use of a symmetric linker AEADP [AEADP = 3,3′-(2-aminoethylazanediyl)dipropanoic acid] for the synthesis of BBN-RGD peptide heterodimer. The F-labeled heterodimer (F-FB-AEADP-BBN-RGD) showed comparable microPET imaging results with glutamate linked BBN-RGD heterodimers, indicating that the replacement of glutamate linker with AEADP linker did not affect the biological activities of BBN-RGD heterodimer. The heterodimer synthesis is rather easy and straightforward. Because tumors often co-express multiple receptors, the use of a symmetric linker provides a general method of fast assembly of various peptide heterodimers for imaging multi-receptor expressing tumors. [ABSTRACT FROM AUTHOR]

Purpose: In vivo imaging of α(v)β(3) has important diagnostic and therapeutic applications. (18)F-Galacto-arginine-glycine-aspartic acid (RGD) has been developed for positron emission tomography (PET) imaging of integrin α(v)β(3) expression and is now being tested on humans. Dimerization and multimerization of cyclic RGD peptides have been reported to improve the integrin α(v)β(3)-binding affinity due to the polyvalency effect. Here, we compared a number of new dimeric RGD peptide tracers with the clinically used (18)F-galacto-RGD.Procedures: RGD monomers and dimers were coupled with galacto or PEG(3) linkers, and labeled with (18)F using 4-nitrophenyl 2-(18)F-fluoropropionate ((18)F-NFP) or N-succinimidyl 4-(18)F-fluorobenzoate as a prosthetic group. The newly developed tracers were evaluated by cell-based receptor-binding assay, biodistribution, and small-animal PET studies in a subcutaneous U87MG glioblastoma xenograft model.Results: Starting with (18)F-F(-), the total reaction time for (18)F-FP-SRGD2 and (18)F-FP-PRGD2 is about 120 min. The decay-corrected radiochemical yields for (18)F-FP-SRGD2 and (18)F-FP-PRGD2 are 52 ± 9% and 80 ± 7% calculated from (18)F-NFP. Noninvasive small-animal PET and direct tissue sampling experiments demonstrated that the dimeric RGD peptides had significantly higher tumor uptake as compared to (18)F-galacto-RGD.Conclusion: Dimeric RGD peptide tracers with relatively high tumor integrin-specific accumulation and favorable in vivo kinetics may have the potential to be translated into clinic for integrin α(v)β(3) imaging. [ABSTRACT FROM AUTHOR]

Radiolabeled Arg-Gly-Asp (RGD) and bombesin (BBN) peptide analogs have been extensively investigated for the imaging of tumor integrin αvβ3 and gastrin-releasing peptide receptor (GRPR) expression, respectively. Recently, we designed and synthesized a RGD-BBN heterodimeric peptide from c(RGDyK) and BBN(7–14) through a glutamate linker. The goal of this study was to investigate the dual receptor-targeting property and tumor diagnostic value of RGD-BBN heterodimeric peptide labeled with generator-eluted 68Ga (t1/2 68 min, β+ 89% and EC 11%), 68Ga-NOTA-RGD-BBN. RGD-BBN heterodimer was conjugated with 1,4,7-triazacyclononanetriacetic acid (NOTA) and labeled with 68Ga. The dual receptor binding affinity was investigated by a radioligand competition binding assay. The in vitro and in vivo dual receptor targeting of 68Ga-NOTA-RGD-BBN was evaluated and compared with that of 68Ga-NOTA-RGD and 68Ga-NOTA-BBN. NOTA-RGD-BBN had integrin αvβ3 and GRPR binding affinities comparable to those of the monomeric RGD and BBN, respectively. The dual receptor targeting property of 68Ga-NOTA-RGD-BBN was validated by blocking studies in a PC-3 tumor model. 68Ga-NOTA-RGD-BBN showed higher tumor uptake than 68Ga-NOTA-RGD and 68Ga-NOTA-BBN. 68Ga-NOTA-RGD-BBN can also image tumors with either integrin or GRPR expression. 68Ga-NOTA-RGD-BBN exhibited dual receptor targeting properties both in vitro and in vivo. The favorable characterizations of 68Ga-NOTA-RGD-BBN such as convenient synthesis, high specific activity, and high tumor uptake, warrant its further investigation for clinical cancer imaging. [ABSTRACT FROM AUTHOR]

Radiolabeled cyclic RGD (Arg-Gly-Asp) peptides have great potential for the early tumor detection and noninvasive monitoring of tumor metastasis and therapeutic response. 18F-labeled RGD analogs ([18F]-AH111585 and [18F]Galacto-RGD) have been investigated in clinical trials for positron emission tomography (PET) imaging of integrin expression in cancer patients. To develop new RGD radiotracers with higher tumor accumulation, improved in vivo kinetics, easy availability and low cost, we developed two new RGD peptides and labeled them with generator-eluted 68Ga (t1/2 = 68 min) for PET imaging of integrin αvβ3 expression in tumor xenograft models. The two new cyclic RGD dimers, E[PEG4-c(RGDfK)]2 (P4-RGD2, PEG4 = 15-amino-4,7,10,13-tetraoxapentadecanoic acid) and E[Gly3-c(RGDfK)]2 (G3-RGD2, G3 = Gly-Gly-Gly) were designed, synthesized and conjugated with 1,4,7-triazacyclononanetriacetic acid (NOTA) for 68Ga labeling. The microPET imaging and biodistribution of the 68Ga labeled RGD tracers were investigated in integrin αvβ3-positive tumor xenografts. The new RGD dimers with the Gly3 and PEG4 linkers showed higher integrin αvβ3 binding affinity than no-linker RGD dimer (RGD2). NOTA-G3-RGD2 and NOTA-P4-RGD2 could be labeled with 68Ga within 30 min with higher purity (>98%) and specific activity (8.88–11.84 MBq/nmol). Both 68Ga-NOTA-P4-RGD2 and 68Ga-NOTA-G3-RGD2 exhibited significantly higher tumor uptake and tumor-to-normal tissue ratios than 68Ga-NOTA-RGD2. Because of their high affinity, high specificity and excellent pharmacokinetic properties, further investigation of the two novel RGD dimers for clinical PET imaging of integrin αvβ3 expression in cancer patients is warranted. [ABSTRACT FROM AUTHOR]

We and others have reported that 18F- and 64Cu-labeled arginine–glycine–aspartate (RGD) peptides allow positron emission tomography (PET) quantification of integrin αvβ3 expression in vivo. However, clinical translation of these radiotracers is partially hindered by the necessity of cyclotron facility to produce the PET isotopes. Generator-based PET isotope 68Ga, with a half-life of 68 min and 89% positron emission, deserves special attention because of its independence of an onsite cyclotron. The goal of this study was to investigate the feasibility of 68Ga-labeled RGD peptides for tumor imaging. Three cyclic RGD peptides, c(RGDyK) (RGD1), E[c(RGDyK)]2 (RGD2), and E{E[c(RGDyK)]2}2 (RGD4), were conjugated with macrocyclic chelator 1,4,7-triazacyclononane-1,4,7-triacetic acid (NOTA) and labeled with 68Ga. Integrin affinity and specificity of the peptide conjugates were assessed by cell-based receptor binding assay, and the tumor targeting efficacy of 68Ga-labeled RGD peptides was evaluated in a subcutaneous U87MG glioblastoma xenograft model. U87MG cell-based receptor binding assay using 125I-echistatin as radioligand showed that integrin affinity followed the order of NOTA–RGD4 > NOTA–RGD2 > NOTA–RGD1. All three NOTA conjugates allowed nearly quantitative 68Ga-labeling within 10 min (12–17 MBq/nmol). Quantitative microPET imaging studies showed that 68Ga-NOTA–RGD4 had the highest tumor uptake but also prominent activity accumulation in the kidneys. 68Ga-NOTA–RGD2 had higher tumor uptake (e.g., 2.8 ± 0.1%ID/g at 1 h postinjection) and similar pharmacokinetics (4.4 ± 0.4 tumor/muscle ratio, 2.0 ± 0.1 tumor/liver ratio, and 1.1 ± 0.1 tumor/kidney ratio) compared with 68Ga-NOTA–RGD1. The dimeric RGD peptide tracer 68Ga-NOTA–RGD2 with good tumor uptake and favorable pharmacokinetics warrants further investigation for potential clinical translation to image integrin αvβ3. [ABSTRACT FROM AUTHOR]

In this study, we evaluated the 12- O-tetradecanoyl-phorbol-13-acetate (TPA)-induced acute and chronic inflammation in living mice by PET imaging of TNF-α and integrin αvβ3 expression. TPA was topically applied to the right ear of BALB/c mice every other day to create the inflammation model. 64Cu-DOTA-etanercept and 64Cu-DOTA-E{E[c(RGDyK)]2}2 were used for PET imaging of TNF-α and integrin αvβ3 expression in both acute and chronic inflammation. Hematoxylin and eosin staining, ex vivo autoradiography, direct tissue sampling, and immunofluorescence staining were also performed to confirm the non-invasive PET imaging results. The ear thickness increased significantly and the TNF-α level more than tripled after a single TPA challenge. MicroPET imaging using 64Cu-DOTA-etanercept revealed high activity accumulation in the inflamed ear, reaching 11.1 ± 1.3, 13.0 ± 2.0, 10.9 ± 1.4, 10.2 ± 2.2%ID/g at 1, 4, 16, and 24 h post injection, respectively ( n = 3). Repeated TPA challenges caused TPA-specific chronic inflammation and reduced 64Cu-DOTA-etanercept uptake due to lowered TNF-α expression. 64Cu-DOTA-E{E[c(RGDyK)]2}2 uptake in the chronically inflamed ears (after four and eight TPA challenges) was significantly higher than in the control ears and those after one TPA challenge. Immunofluorescence staining revealed increased integrin β3 expression, consistent with the non-invasive PET imaging results using 64Cu-DOTA-E{E[c(RGDyK)]2}2 as an integrin αvβ3-specific radiotracer. Biodistribution and autoradiography studies further confirmed the quantification capability of microPET imaging. Successful PET imaging of TNF-α expression in acute inflammation and integrin αvβ3 expression in chronic inflammation provides the rationale for multiple target evaluation over time to fully understand the inflammation processes. [ABSTRACT FROM AUTHOR]

We have previously reported that 18F-FB-E[c(RGDyK)]2 (18F-FRGD2) allows quantitative PET imaging of integrin αvβ3 expression. However, the potential clinical translation was hampered by the relatively low radiochemical yield. The goal of this study was to improve the radiolabeling yield, without compromising the tumor targeting efficiency and in vivo kinetics, by incorporating a hydrophilic bifunctional mini-PEG spacer. 18F-FB-mini-PEG-E[c(RGDyK)]2 (18F-FPRGD2) was synthesized by coupling N-succinimidyl-4-18F-fluorobenzoate (18F-SFB) with NH2-mini-PEG-E[c(RGDyK)]2 (denoted as PRGD2). In vitro receptor binding affinity, metabolic stability, and integrin αvβ3 specificity of the new tracer 18F-FPRGD2 were assessed. The diagnostic value of 18F-FPRGD2 was evaluated in subcutaneous U87MG glioblastoma xenografted mice and in c- neu transgenic mice by quantitative microPET imaging studies. The decay-corrected radiochemical yield based on 18F-SFB was more than 60% with radiochemical purity of >99%. 18F-FPRGD2 had high receptor binding affinity, metabolic stability, and integrin αvβ3-specific tumor uptake in the U87MG glioma xenograft model comparable to those of 18F-FRGD2. The kidney uptake was appreciably lower for 18F-FPRGD2 compared with 18F-FRGD2 [2.0 ± 0.2%ID/g for 18F-FPRGD2 vs 3.0 ± 0.2%ID/g for 18F-FRGD2 at 1 h post injection (p.i.)]. The uptake in all the other organs except the urinary bladder was at background level. 18F-FPRGD2 also exhibited excellent tumor uptake in c- neu oncomice (3.6 ± 0.1%ID/g at 30 min p.i.). Incorporation of a mini-PEG spacer significantly improved the overall radiolabeling yield of 18F-FPRGD2. 18F-FPRGD2 also had reduced renal uptake and similar tumor targeting efficacy as compared with 18F-FRGD2. Further testing and clinical translation of 18F-FPRGD2 are warranted. [ABSTRACT FROM AUTHOR]

Multimeric ligands consisting of multiple pharmacophores connected to a single backbone have been widely investigated for diagnostic and therapeutic applications. In this review, we summarize recent developments regarding multimeric radioligands targeting integrin αvβ3 receptors on cancer cells for molecular imaging and diagnostic applications using positron emission tomography (PET). Integrin αvβ3 receptors are glycoproteins expressed on the cell surface, which have a significant role in tumor angiogenesis. They act as receptors for several extracellular matrix proteins exposing the tripeptide sequence arginine-glycine-aspartic (RGD). Cyclic RDG peptidic ligands c(RGD) have been developed for integrin αvβ3 tumor-targeting positron emission tomography (PET) diagnosis. Several c(RGD) pharmacophores, connected with the linker and conjugated to a chelator or precursor for radiolabeling with different PET radionuclides (18F, 64Cu, and 68Ga), have resulted in multimeric ligands superior to c(RGD) monomers. The binding avidity, pharmacodynamic, and PET imaging properties of these multimeric c(RGD) radioligands, in relation to their structural characteristics are analyzed and discussed. Furthermore, specific examples from preclinical studies and clinical investigations are included. [ABSTRACT FROM AUTHOR]

Background: Activated hepatic stellate cells (HSCs), which express integrin αvβ3, are a major fibrogenic factor in NASH pathophysiology. 18F-labeled cyclic arginine-glycine-aspartic acid penta-peptide (18F-FPP-RGD2) has been used as a PET probe for tumors expressing integrin αvβ3. The aim of this study was to assess the potential of PET with 18F-FPP-RGD2 to detect hepatic integrin αvβ3 expression in non-alcoholic steatohepatitis (NASH) model mice.Results: Thirty-two male C57BL/6 mice aged 6 weeks were fed a choline-deficient, L-amino acid-defined, high-fat diet (CDAHFD) for 3 and 8 weeks. 18F-FPP-RGD2 PET imaging of the liver was performed at 3 and 8 weeks after CDAHFD feeding. After PET scanning, levels of hepatic integrin αvβ, 3α-smooth muscle actin (α-SMA), and collagen type 1 alpha 1(col1a1) were measured. Histopathological analysis of hepatic steatosis, inflammation, and fibrosis, as well as blood biochemistry analysis, was also performed. CDAHFD for 3 and 8 weeks produced a moderate-to-severe steatosis and inflammation of the liver in mice. NAFLD activity score (NAS) in mice fed the CDAHFD for 3 and 8 weeks were more than 4 indicating NASH or borderline NASH pathology. Fibrosis was observed only in mice fed the CDAHFD for 8 weeks. PET imaging showed that the hepatic standardized uptake value, SUV80-90 min, was increased with prolonged CDAHFD feeding compared with the respective controls (CDAHFD 3 weeks 0.32 ± 0.06 vs 0.48 ± 0.05, p < 0.01; CDAHFD 8 weeks 0.35 ± 0.04 vs 0.75 ± 0.07, p < 0.01, respectively). Prolonged CDAHFD feeding increased hepatic mRNA and protein levels of integrin αv and β3 at 3 and 8 weeks. Hepatic 18F-FPP-RGD2 uptake and amount of integrin αv and β3 protein were well correlated (r = 0.593, p < 0.05 and r = 0.835, p < 0.001, respectively). Hepatic 18F-FPP-RGD2 uptake also showed a positive correlation with Sirius red-positive area.Conclusions: The hepatic uptake of 18F-FPP-RGD2 correlated well with integrin αv and β3 expression and histological fibrosis in a mouse model of NASH, suggesting the predictability of fibrosis in NASH pathology. [ABSTRACT FROM AUTHOR]