PET Imaging Of Tumors Research Articles

A New Tool for Extracting Static and Dynamic Parameters from [18F]F-DOPA PET/CT in Pediatric Gliomas

Background/Objectives: PET imaging with [18F]F-DOPA has demonstrated high potential for the evaluation and management of pediatric brain gliomas. Manual extraction of PET parameters is time-consuming, lacks reproducibility, and varies with operator experience. Methods: In this study, we tested whether a semi-automated image processing framework could overcome these limitations. Pediatric patients with available static and/or dynamic [18F]F-DOPA PET studies were evaluated retrospectively. We developed a Python software to automate clinical index calculations, including preprocessing to delineate tumor volumes from structural MRI, accounting for lesions with low [18F]F-DOPA uptake. A total of 73 subjects with treatment-naïve low- and high-grade gliomas, who underwent brain MRI within two weeks of [18F]F-DOPA PET, were included and analyzed. Static analysis was conducted on all subjects, while dynamic analysis was performed on 32 patients. Results: For 68 subjects, the Intraclass Correlation Coefficient for T/S between manual and ground truth segmentation was 0.91. Using our tool, ICC improved to 0.94. Our method demonstrated good reproducibility in extracting static tumor-to-striatum ratio (p = 0.357); however, significant differences were observed in tumor slope (p < 0.05). No significant differences were found in time-to-peak (p = 0.167) and striatum slope (p = 0.36). Conclusions: Our framework aids in analyzing [18F]F-DOPA PET images of pediatric brain tumors by automating clinical score extraction, simplifying segmentation and Time Activity Curve extraction, reducing user variability, and enhancing reproducibility.

Copper-61 is an advantageous alternative to gallium-68 for PET imaging of somatostatin receptor-expressing tumors: a head-to-head comparative preclinical study.

Gallium-68 positron emission tomography (68Ga-PET) with the two registered somatostatin analogs, [68Ga]Ga-DOTA-Tyr3-octreotide ([68Ga]Ga-DOTA-TOC) and [68Ga]Ga-DOTA-Tyr3-octreotate ([68Ga]Ga-DOTA-TATE), where DOTA = 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid, is routinely used for imaging of somatostatin receptor (SST)-expressing tumors. We investigated copper-61 (61Cu) as an alternative radiometal for PET imaging of SST-expressing tumors. Compared to gallium-68, copper-61 (t1/2 = 3.33 h, E β + max = 1.22 MeV) can be produced on a large scale, enables late time point imaging, and has the therapeutic twin copper-67. Herein, DOTA-TOC and 1,4,7-triazacyclononane,1-glutaric acid-4,7-acetic acid (NODAGA)-TOC were labeled with copper-61 and compared with the clinically used [68Ga]Ga-DOTA-TOC. [61Cu]CuCl2 was produced from an irradiated natural nickel target. DOTA-TOC and NODAGA-TOC were labeled with [61Cu]CuCl2 in ammonium acetate buffer so to achieve a reaction pH of 5-6 and a temperature of 95°C for DOTA-TOC or room temperature for NODAGA-TOC. The radioligands were evaluated head-to-head in vitro using human embryonic kidney (HEK)-SST2 cells (affinity, binding sites, cellular uptake, and efflux) and in vivo using HEK-SST2 xenografts [PET/computed tomography (CT) imaging, biodistribution, and pharmacokinetics] and compared with [68Ga]Ga-DOTA-TOC, which was prepared using a standard procedure. Dosimetry estimates were made for [61Cu]Cu-NODAGA-TOC. [61Cu]Cu-DOTA-TOC and [61Cu]Cu-NODAGA-TOC were prepared at an apparent molar activity of 25 MBq/nmol with radiochemical purities of ≥96% and ≥98%, respectively. In vitro, both presented a sub-nanomolar affinity for SST2 (IC50 = 0.23 and 0.34 nM, respectively). They were almost entirely internalized upon binding to SST2-expressing cells and had similar efflux rates at 37°C. In vivo, [61Cu]Cu-DOTA-TOC and [61Cu]Cu-NODAGA-TOC showed the same accumulation in SST2-expressing tumors. However, PET/CT images and biodistribution analyses clearly showed an unfavorable biodistribution for [61Cu]Cu-DOTA-TOC, characterized by accumulation in the liver and the abdomen. [61Cu]Cu-NODAGA-TOC displayed favorable biodistribution, comparable with [68Ga]Ga-DOTA-TOC at 1 h post-injection (p.i.). Notwithstanding, [61Cu]Cu-NODAGA-TOC showed advantages at 4 h p.i., due to the tumor retention and improved tumor-to-non-tumor ratios. The effective dose (2.41 × 10-3 mSv/MBq) of [61Cu]Cu-NODAGA-TOC, but also the dose to the other organs and the kidneys (9.65 × 10-2 mGy/MBq), suggested a favorable safety profile. Somatostatin receptor 61Cu-PET imaging not only matches the performance of 68Ga-PET at 1 h p.i. but has advantages in late-time imaging at 4 h p.i., as it provides improved tumor-to-non-tumor ratios. [61Cu]Cu-NODAGA-TOC is superior to [61Cu]Cu-DOTA-TOC in vivo. The use of the chelator NODAGA allows quantitative labeling with copper-61 at room temperature and enables the straightforward use of a kit formulation for simple manufacturing in medical centers.

Preclinical Evaluation of a Radiolabeled Pan-RAF Inhibitor for RAF-Specific PET/CT Imaging.

Abnormalities in the RAS-RAF signaling pathway occur in many solid tumors, leading to aberrant tumor proliferation, invasion, and metastasis. Due to the elusive pharmacology of RAS, RAF inhibitors have become the main targeted therapeutic drugs. Naporafenib (LXH-254) is a high-affinity pan-RAF inhibitor with FDA Fast Track Qualification. We sought to develop an 18F-labeled molecular probe from LXH-254 for PET imaging of tumors overexpressing RAF to noninvasively screen patients for susceptibility to targeted RAF therapy. To reduce the lipid solubility, LXH-254 was designed with triethylene glycol di(p-toluenesulfonate) (TsO-PEG3-OTs) to obtain the precursor (LXH-254-OTs) and a nucleophilic substitution reaction with 18F to obtain the tracer ([18F]F-LXH-254). [18F]F-LXH-254 exhibited good molar activity (7.16 ± 0.81 GBq/μmol), radiochemical purity (>95%), and stability. Micro-PET imaging revealed distinct radioactivity accumulation of [18F]F-LXH-254 in tumors in the imaging groups, whereas in the blocked group, the tumor radioactivity level was consistent with the background tissue, illustrating the affinity and specificity of [18F]F-LXH-254 in targeting RAF. Overall, [18F]F-LXH-254 is a promising radiotracer for screening and diagnosing patients with RAF-related disease and monitoring their treatment. This is the first attempt at using an 18F-labeled RAF-specific radiotracer.

68Ga]Ga-labeled FAPI Conjugated with Gly-Pro Sequence for PET Imaging of Malignant Tumors.

To improve tumor uptake and prolong tumor retention, a novel fibroblast activation protein (FAP) ligand based on a quinoline-based FAP inhibitor (FAPI) conjugated with the Gly-Pro sequence and 1,4,7,10-tetraazacyclododecane-N,N',N″,N‴-tetraacetic acid (DOTA) was radiolabeled with [68Ga]GaCl3 ([68Ga]Ga-DOTA-GPFAPI-04). Due to the tumor heterogeneity, this study aimed to further validate the preclinical value of [68Ga]Ga-DOTA-GPFAPI-04 PET imaging in tumor mice models with different FAP expression levels. [68Ga]Ga-DOTA-GPFAPI-04 was synthesized and its partition coefficient was measured. The stability of [68Ga]Ga-DOTA-GPFAPI-04 was tested in phosphate-buffered saline (PBS, pH 7.4) and fetal bovine serum (FBS). Small animal PET and semi-quantitative studies were conducted in Panc-1 and A549 xenograft tumor mice models compared with [68Ga]Ga-DOTA-FAPI-04. Immunofluorescent and immunohistochemical staining and western blot assay were performed to confirm FAP expression in xenograft tumors. [68Ga]Ga-DOTA-GPFAPI-04 exhibited a radiochemical purity of > 99% and high stability in PBS and FBS. [68Ga]Ga-DOTA-GPFAPI-04 had higher hydrophilic property than [68Ga]Ga-DOTA-FAPI-04 (-4.09 ± 0.05 vs -3.45 ± 0.05). Small animal PET and semi-quantitative analysis revealed Panc-1 xenograft tumor displayed higher tumor uptake of [68Ga]Ga-DOTA-GPFAPI-04 and tumor-to-background ratios compared to A549 xenograft tumor, consistent with the results of immunofluorescence, immunohistochemistry, and western blot. Moreover, [68Ga]Ga-DOTA-GPFAPI-04 demonstrated higher tumor accumulation and longer tumor retention than [68Ga]Ga-DOTA-FAPI-04 in both Panc-1 and A549 xenograft tumors. Furthermore, the FAP-binding specificity of [68Ga]Ga-DOTA-GPFAPI-04 was confirmed in vivo by co-injection of unlabeled GPFAPI-04. [68Ga]Ga-DOTA-GPFAPI-04 showed more favorable in vivo tumor imaging and longer tumor retention compared to [68Ga]Ga-DOTA-FAPI-04, which has high potential to be a promising PET probe for detecting FAP-positive tumors.

Preclinical studies and absorbed dose estimation of [89Zr]Zr-DFO-Bevacizumab for PET imaging of VEGF-expressing tumors

This study aimed to carry out the preclinical studies of [89Zr]Zr-DFO-Bevacizumab. The radiolabeled compound was prepared with radiochemical purity >99% (ITLC), and a specific activity of 74 GBq/g. Cellular studies indicated the great capability of [89Zr]Zr-DFO-Bevacizumab for binding to SKOV3 cell lines. High accumulation was observed in the tumor. The liver and spleen received the highest absorbed dose with 1.12 and 0.72 mGy/MBq, respectively. This radiopharmaceutical can be considered as a suitable PET agent for VEGF-expressing ovarian cancer imaging.

Synthesis and evaluation of a 68Ga-labeled spermine derivative for tumor PET imaging

BackgroundThe polyamine transporter system (PTS), which renders it a promising target for tumor therapy and imaging applications, facilitates the transmembrane transport of polyamines. We reported a novel derivative of spermine labeled with gallium-68 ([68Ga]Ga-NOTA-Spermine) for the imaging of the PTS in mouse models of tumor. ResultsThe radiochemical yield of [68Ga]Ga-NOTA-Spermine was determined to be 64–69 %, demonstrating exceptional stability and radiochemical purity (>98 %). Cellular uptake experiments revealed that A549 cells exhibited peak uptake of [68Ga]Ga-NOTA-Spermine at 90 min (15.4 % ± 0.68 %). Biodistribution analysis demonstrated significant accumulation of [68Ga]Ga-NOTA-Spermine in kidneys and liver, while exhibiting low uptake levels in muscle, brain, and bones. Furthermore, Micro-PET/CT scans conducted on A549 tumor-bearing mouse models indicated substantial uptake of [68Ga]Ga-NOTA-Spermine, with maximum tumor/muscle (T/M) ratios reaching 3.71. ConclusionThese results suggest that [68Ga]Ga-NOTA-Spermine holds potential as a PET imaging agent for tumors with high levels of PTS.

A PET probe targeting polyamine transport system for precise tumor diagnosis and therapy

Polyamine metabolism dysregulation is a hallmark of many cancers, offering a promising avenue for early tumor theranostics. This study presents the development of a nuclear probe derived from spermidine (SPM) for dual-purpose tumor PET imaging and internal radiation therapy. The probe, radiolabeled with either [68Ga]Ga for diagnostic applications or [177Lu]Lu for therapeutic use, was synthesized with exceptional purity, stability, and specific activity. Extensive testing involving 12 different tumor cell lines revealed remarkable specificity towards B16 melanoma cells, showcasing outstanding tumor localization and target-to-non-target ratio. Mechanistic investigations employing polyamines, non-labeled precursor, and polyamine transport system (PTS) inhibitor, consistently affirmed the probeʼs targetability through recognition of the PTS. Notably, while previous reports indicated PTS upregulation in various tumor types for targeted therapy, this study observed no positive signals, highlighting a concentration-dependent discrepancy between targeting for therapy and diagnosis. Furthermore, when labeled with [177Lu], the probe demonstrated its therapeutic potential by effectively controlling tumor growth and extending mouse survival. Investigations into biodistribution, excretion, and biosafety in healthy humans laid a robust foundation for clinical translation. This study introduces a versatile SPM-based nuclear probe with applications in precise tumor theranostics, offering promising prospects for clinical implementation.

Bispecific radioimmunoconjugates exploit receptor heterogeneity for positron emission tomography of tumors expressing HER2 and/or EGFR

HER2 heterogeneity is a challenge for molecular imaging or treating HER2-positive breast cancer (BC). EGFR is coexpressed in some tumors exhibiting HER2 heterogeneity. Bispecific radioimmunoconjugates (bsRICs) that bind HER2 and EGFR were constructed by linking trastuzumab Fab through polyethyleneglycol (PEG24) to EGF. We established s.c. tumors in NOD-SCID mice that homogeneously or heterogeneously expressed HER2 and/or EGFR by the inoculation of HER2-positive/EGFR-negative SK-OV-3 cells, EGFR-positive/HER2-negative MDA-MB-468 cells or mixtures of these cells. [64Cu]Cu-NOTA-trastuzumab Fab-PEG24-EGF were compared to [64Cu]Cu-NOTA-trastuzumab Fab or [64Cu]Cu-NOTA-EGF for the PET imaging of HER2 and/or EGFR-positive tumors. [64Cu]Cu-NOTA-trastuzumab Fab-PEG24-EGF bsRICs imaged tumors expressing HER2 or EGFR or heterogeneously expressing these receptors, while monospecific agents only imaged HER2-or EGFR-positive tumors. Our results indicate that bsRICs labeled with 64Cu are able to exploit receptor heterogeneity for tumor imaging. PET may select patients for radioimmunotherapy with bsRICs complexed to the β-particle emitter, 177Lu or Auger electron-emitter, 111In in a theranostic approach.

Design, Synthesis, and Evaluation of [18F]BIBD-300 as a Positron Emission Tomography Tracer for Poly(ADP-Ribose) Polymerase-1.

Compounds 8a-j were designed to adjust the mode of interaction and lipophilicity of FTT by scaffold hopping and changing the length of the alkoxy groups. Compounds 8a, 8d, 8g, and BIBD-300 were screened for high-affinity PARP-1 through enzyme inhibition assays and are worthy of further evaluation. PET imaging of MCF-7 subcutaneous tumors with moderate expression of PARP-1 showed that compared to [18F]FTT, [18F]8a, [18F]8d, and [18F]8g exhibited greater nonspecific uptake, a lower target-to-nontarget ratio, and severe defluorination, while [18F]BIBD-300 exhibited lower nonspecific uptake and a greater target-to-nontarget ratio. PET imaging of 22Rv1 subcutaneous tumors, which highly express PARP-1, confirmed that the uptake of [18F]BIBD-300 in normal organs, such as the liver, muscle, and bone, was lower than that of [18F]FTT, and the ratio of tumor-to-muscle and tumor-to-liver [18F]BIBD-300 was greater than that of [18F]FTT. The biodistribution results in mice with MCF-7 and 22Rv1 subcutaneous tumors further validated the results of PET imaging. Unlike [18F]FTT, which mainly relies on hepatobiliary clearance, [18F]BIBD-300, which has lower lipophilicity, undergoes a partial shift from hepatobiliary to renal clearance, providing the possibility for [18F]BIBD-300 to indicate liver cancer. The difference in the PET imaging results for [18F]FTT, [18F]BIBD-300, and [18F]8j in 22Rv1 mice and the corresponding molecular docking results further confirmed that subtle structural modifications in lipophilicity greatly optimize the properties of the tracer. Cell uptake experiments also demonstrated that [18F]BIBD-300 has a high affinity for PARP-1. Metabolized and unmetabolized [18F]FTT and [18F]BIBD-300 were detected in the brain, indicating that they could not accurately quantify the amount of PARP-1 in the brain. However, PET imaging of glioma showed that both [18F]FTT and [18F]BIBD-300 could accurately localize both in situ to C6 and U87MG tumors. Based on its potential advantages in the diagnosis of breast cancer, prostate cancer, and glioma, as well as liver cancer, [18F]BIBD-300 is a new option for an excellent PARP-1 tracer.

Optimization and automation of the radiosynthesis of [18F]Lu-LuFL as a clinically useful PET ligand targeting FAP for tumor imaging

Recently, a novel radiohybrid tracer [18F]Lu-LuFL targeting the fibroblast activation protein (FAP) has been developed for PET imaging of solid tumors. This tracer has shown promising results, prompting us to conduct a first-in-human study to evaluate its efficacy for PET imaging of FAP in human body. In order to facilitate the routine production and clinical application of [18F]Lu-LuFL, a straightforward and efficient automated synthesis is described. The optimum labeling parameters were determined at laboratory scale, and subsequently incorporated into an automated production process. Further studies have demonstrated that clinical doses of [18F]Lu-LuFL can be prepared within 19 min, with excellent radio chemical purity (>99%) and activity yield (23.58% ± 2.20%, non-decay corrected), coupled with solid phase extraction (SPE) purification method. All the quality control results satisfy the required criteria for release. In conclusion, we have successfully synthesized [18F]Lu-LuFL with sufficient radioactivity and superior quality, thereby establishing its potential for further clinical application.

Development of [64Cu]Cu-NODAGA-RGD-BBN as a Novel Radiotracer for Dual Integrin and GRPR-targeted Tumor PET Imaging.

In this study, [64Cu]Cu-NODAGA-RGD-BBN was prepared and its preclinical assessments were evaluated for PET imaging of GRPR overexpressing tumors. NODAGA-RGD-BBN heterodimer peptide was successfully labeled with cyclotronproduced copper-64 at optimized conditions. The radiochemical purity of the radiotracer was checked by HPLC and RTLC methods. The stability of the radiolabeled compound was assessed in PBS (4°C) and in human blood serum (37°C). Binding affinity and internalization of [64Cu]Cu-NODAGA-RGD-BBN were studied on PC3, LNCaP, and CHO cell lines. The biodistribution of the radiotracer was evaluated in normal and tumor-bearing mice. [64Cu]Cu-NODAGA-RGD-BBN was prepared with radiochemical purity >99 ± 0.7% (HPLC/ITLC) and specific activity of 18.5 ± 2.2 TBq/mmol. The radiotracer showed high stability in PBS (95 ± 1.05%) and in human blood serum (96 ± 1.24%) and, high affinity to the GRP expressing tumor cells. [64Cu]Cu-NODAGA-RGD-BBN showed hydrophilic (log p = -1.14) and agonistic nature. The biodistribution and imaging studies demonstrated high uptake at the tumor site at all intervals post-injection and 3-4 h post-injection can be considered an appropriate time of imaging. The results indicated that [64Cu]Cu-NODAGA-RGD-BBN radiolabeled heterodimer peptide can be considered as a high-potential agent for PET imaging of GRPRoverexpressing tumors.

Comparison of IL-2-antibody to IL-2-Fc with or without stereotactic radiation therapy in CEA immunocompetent mice with CEA positive tumors.

The potent immune effects of interleukin-2 (IL-2) for cancer therapy can be increased by genetic fusion of IL-2 to the Fc domain of an antibody (IL-2-Fc) or tumor targeted by genetic fusion to a whole antibody known as an immunocytokine (ICK). An anti-CEA ICK (M5A-IL-2) was compared to an IL-2-Fc fusion protein using tumor therapy and PET imaging in CEA transgenic immunocompetent mice bearing CEA positive colon or breast tumors. Combination with stereotactic radiation therapy (SRT) was performed with either ICK or IL-2-Fc. ICK and IL-2-Fc had comparable antitumor effects in both tumor models, although ICK had higher tumor uptake and slower blood clearance than an IL-2-Fc. Analysis of IFNγ+ /CD8+ and FoxP3+ /CD4+ T cells revealed higher levels of IFNγ-producing CD8+ T cells in ICK treated mice versus more efficient Treg elimination in IL-2-Fc treated mice. No significant or lasting toxicity was detected for either agent. Combination therapies with SRT revealed comparable efficacy and induction of immune memory for both ICK and IL-2-Fc when mice were rechallenged post-therapy. IL-2-Fc had comparable antitumor efficacy to CEA-targeted M5A-IL-2 ICK, while both fusion proteins induced immune memory when combined with SRT. Differences in the therapeutic mechanisms of both agents were observed.

Abstract A016: Novel positron emission tomography imaging targeting cell surface glycans for pancreatic cancer: 18F-labeled rBC2LCN lectin

Abstract Introduction: Advancement in early detection modalities will greatly improve the overall prognoses of pancreatic ductal adenocarcinoma (PDAC). As a molecular imaging technique, [18F]FDG-PET imaging has provided new insight in PDAC diagnosis; however, the utility of FDG for early-stage PDAC detection remains undefined. Therefore, PET radiotracers conjugated to malignancy-specific targeting agents (e.g., antibodies targeting cancer-associated/specific antigens) have been developed to improve specificity toward cancer cells but persist in the developmental stage. One underexplored cancer cell surface-specific target is the glycocalyx. We have previously demonstrated that fucosylated glycans (H type-3 antigens) are expressed in pancreatic cancers and a recombinant lectin, rBC2LCN, binds to these glycans specifically (Shimomura O et al: Mol Cancer Ther 2018). Here, we developed a novel, lectin-based PET probe targeting cell surface glycan for pancreatic cancer. Material and Methods: The rBC2LCN (16kDa) lectin was labeled with N-succinimidyl-4-[18F]fluorobenzoate ([18F]SFB), prepared according to literature (Maeding P, et al: Appl Radiat Isot 2005) in phosphate buffer (pH 8.5). Obtained [18F]FB-rBC2LCN was then purified using gel filtration column (PD-10, GE, Uppsala, Sweden). A PDAC cell line, Capan-1, was used as a model which expresses H type-3 antigens. The biodistribution of the [18F]FB-rBC2LCN was evaluated in athymic nude mice bearing subcutaneous Capan-1 tumors. Static, small-animal PET imaging studies (G4, SOFIE, Dulles, VA) were performed to further evaluate in vivo tumor targeting and specificity. Results: The decay-corrected radiochemical mean yield of [18F]FB-rBC2LCN was 3.6%±0.3% (mean±SD), based on [18F]SFB, and radiochemical purity was above 95%. Cell binding and uptake revealed that [18F]FB-rBC2LCN binds to Capan-1 cells. Mice received 0.32±0.16 MBq (corresponding to 59±21 mg) of product, injected intravenously. Biodistribution studies demonstrated that Capan-1 tumor uptake (6.5±2.0 %ID/g) was high as early as 60 min after injection of [18F]FB-rBC2LCN and the uptake increased over time (9.6±2.2 and 11±3.9 at 150 and 240 min after injection, respectively). Tumor-to-muscle ratio increased with time up to 19±1.8 at 360 min after injection, indicating image contrast. Although kidneys (72±23 %ID/g), liver (7.1±1.7 %ID/g) and lungs (7.4±2.9 %ID/g) showed high uptake at 60 min after injection, the uptake of those organs decreased over time. The blood activity concentration of the tracer was also high at 60 min after injection (12±4.4 %ID/g) and decreased over time. Bone uptake was very low (below 1.8 %ID/g throughout the study), indicating that defluorinating metabolism was negligible. High-contrast PET imaging of tumors was achieved as early as 60 min after injection and the contrast became clearer over 240 min, consistent with the biodistribution result. Conclusions: Our 18F-labeled rBC2LCN lectin warrants further clinical development to improve the accuracy and sensitivity of early-stage PDAC detection. Citation Format: Yukihito Kuroda, Tatsuya Oda, Osamu Shimomura, Bryan J. Mathis, Hiroaki Tateno, Kentaro Hatano. Novel positron emission tomography imaging targeting cell surface glycans for pancreatic cancer: 18F-labeled rBC2LCN lectin [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Pancreatic Cancer; 2023 Sep 27-30; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(2 Suppl):Abstract nr A016.

Site-Specific 68Ga Radiolabeling of Trastuzumab Fab via Methionine for ImmunoPET Imaging.

Bioconjugates of antibodies and their derivatives radiolabeled with β+-emitting radionuclides can be utilized for diagnostic PET imaging. Site-specific attachment of radioactive cargo to antibody delivery vectors provides homogeneous, well-defined immunoconjugates. Recent studies have demonstrated the utility of oxaziridine chemistry for site-specific labeling of methionine residues. Herein, we applied this approach to site-specifically radiolabel trastuzumab-derived Fab immunoconjugates with 68Ga, which can be used for in vivo PET imaging of HER2-positive breast cancer tumors. Initially, a reactive azide was introduced to a single solvent-accessible methionine residue in both the wild-type Fab and an engineered derivative containing methionine residue M74, utilizing the principles of oxaziridine chemistry. Subsequently, these conjugates were functionalized with a modified DFO chelator incorporating dibenzocyclooctyne. The resulting DFO-WT and DFO-M74 conjugates were radiolabeled with generator-produced [68Ga]Ga3+, to yield the novel PET radiotracers, [68Ga]Ga-DFO-WT and [68Ga]Ga-DFO-M74. In vitro and in vivo studies demonstrated that [68Ga]Ga-DFO-M74 exhibited a higher affinity for HER2 receptors. Biodistribution studies in mice bearing orthotopic HER2-positive breast tumors revealed a higher uptake of [68Ga]Ga-DFO-M74 in the tumor tissue, accompanied by rapid renal clearance, enabling clear delineation of tumors using PET imaging. Conversely, [68Ga]Ga-DFO-WT exhibited lower uptake and inferior image contrast compared to [68Ga]Ga-DFO-M74. Overall, the results demonstrate that the highly facile methionine-oxaziridine modification approach can be simply applied to the synthesis of stable and site-specifically modified radiolabeled antibody-chelator conjugates with favorable pharmacokinetics for PET imaging.

Molecular Imaging Investigations of Polymer-Coated Cerium Oxide Nanoparticles as a Radioprotective Therapeutic Candidate.

Cerium oxide nanoparticles (CONPs) have a unique surface redox chemistry that appears to selectively protect normal tissues from radiation induced damage. Our prior research exploring the biocompatibility of polymer-coated CONPs found further study of poly-acrylic acid (PAA)-coated CONPs was warranted due to improved systemic biodistribution and rapid renal clearance. This work further explores PAA-CONPs' radioprotective efficacy and mechanism of action related to tumor microenvironment pH. An ex vivo TUNEL assay was used to measure PAA-CONPs' protection of the irradiated mouse colon in comparison to the established radioprotector amifostine. [18F]FDG PET imaging of spontaneous colon tumors was utilized to determine the effects of PAA-CONPs on tumor radiation response. In vivo MRI and an ex vivo clonogenic assay were used to determine pH effects on PAA-CONPs' radioprotection in irradiated tumor-bearing mice. PAA-CONPs showed excellent radioprotective efficacy in the normal colon that was equivalent to uncoated CONPs and amifostine. [18F]FDG PET imaging showed PAA-CONPs do not affect tumor response to radiation. Normalization of tumor pH allowed some radioprotection of tumors by PAA-CONPs, which may explain their lack of tumor radioprotection in the acidic tumor microenvironment. Overall, PAA-CONPs meet the criteria for clinical application as a radioprotective therapeutic agent and are an excellent candidate for further study.

Integrin αvβ3 and EGFR dual-targeted [64Cu]Cu-NOTA-RGD-GE11 heterodimer for PET imaging in pancreatic cancer mouse model

PurposeRadiolabeled 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 αvβ3 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 αvβ3 and EGFR. Procedures[64Cu]Cu-NOTA-RGD-GE11 was prepared by the conjugation of RGD-PEG4-NOTA-N3 and GE11-PEG4-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. Results[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. ConclusionThe results suggested that the as-developed [64Cu]Cu-NOTA-RGD-GE11 could serve as a potential PET tracer for the noninvasive imaging of integrin αvβ3 and EGFR expression in tumors.

Synthesis and preclinical evaluation of two osimertinib isotopologues labeled with carbon-11 as PET tracers targeting the tyrosine kinase domain of the epidermal growth factor receptor

IntroductionOsimertinib is a third-generation tyrosine kinase inhibitor (TKI) that is able to inhibit the EGFR treatment resistance mutation T790M and primary EGFR mutations Del19 and L858R. The aim of the study was to evaluate the potential of carbon-11 labeled osimertinib to be used as a tracer for the PET imaging of tumors bearing the T790M mutation. MethodsOsimertinib was labeled with carbon-11 at two positions, and the effect of the labeling position on the metabolism and biodistribution was studied in female nu/nu mice. The mutation status specificity of osimertinib was confirmed in vitro in a cell growth inhibition experiment, and the tumor-targeting potential of the carbon-11 isotopologues was evaluated using female nu/nu mice xenografted with NSCLC cell lines; the wild-type EGFR expressing A549, the primary Del19 EGFR mutated HCC827 and the resistance T790M/L858R mutated H1975. One of the osimertinib tracers was selected based on the results acquired and evaluated for tracer specificity and selectivity by assessment of tumor uptake in a PET study where HCC827 tumor-bearing mice were pretreated with osimertinib or afatinib. Results[Methylindole-11C]- and [dimethylamine-11C]osimertinib were synthesized by 11C-methylation of precursors AZ5104 and AZ7550, respectively. Rapid metabolism of both analogs of [11C]osimertinib was observed. Although the tumor uptake and retention of [methylindole-11C]- and [dimethylamine-11C]osimertinib in tumors were similar, the tumor-to-muscle ratios appeared to be higher for [methylindole-11C]osimertinib. The highest uptake, tumor-to-blood, and tumor-to-muscle ratio were observed in the Del19 EGFR mutated HCC827 tumors. However, the specificity and selectivity of [methylindole-11C]osimertinib PET could not be demonstrated in HCC827 tumors. The uptake of [methylindole-11C]osimertinib was not significantly higher in T790M resistance mutated H1975 xenografts compared to the negative control cell line A549. ConclusionsOsimertinib was successfully labeled at two positions with carbon-11, yielding two EGFR PET tracers, [methylindole-11C]osimertinib and [dimethylamine-11C]osimertinib. The preclinical evaluation demonstrated uptake and retention in three NSCLC xenografts; A549, HCC827, and H1975. The highest uptake was observed in the primary Del19 EGFR mutated HCC827. The ability of [methylindole-11C]osimertinib to distinguish between the T790M resistance mutated H1975 xenografts and the wild-type EGFR expressing A549 could not be confirmed in the ex vivo study.

Molecular Design of 68Ga- and 89Zr-Labeled Anticalin Radioligands for PET-Imaging of PSMA-Positive Tumors.

Anticalin proteins directed against the prostate-specific membrane antigen (PSMA), optionally having tailored plasma half-life using PASylation technology, show promise as radioligands for PET-imaging of xenograft tumors in mice. To investigate their suitability, the short-circulating unmodified Anticalin was labeled with 68Ga (τ1/2 = 68 min), using the NODAGA chelator, whereas the half-life extended PASylated Anticalin was labeled with 89Zr (τ1/2 = 78 h), using either the linear chelator deferoxamine (Dfo) or a cyclic derivative, fusarinine C (FsC). Different PSMA targeting Anticalin versions (optionally carrying the PASylation sequence) were produced carrying a single exposed N- or C-terminal Cys residue and site-specifically conjugated with the different radiochelators via maleimide chemistry. These protein conjugates were labeled with radioisotopes having distinct physical half-lives and, subsequently, applied for PET-imaging of subcutaneous LNCaP xenograft tumors in CB17 SCID mice. Uptake of the protein tracers into tumor versus healthy tissues was assessed by segmentation of PET data as well as biodistribution analyses. PET-imaging with both the 68Ga-labeled plain Anticalin and the 89Zr-labeled PASylated Anticalin allowed clear delineation of the xenograft tumor. The radioligand A3A5.1-PAS(200)-FsC·89Zr, having an extended plasma half-life, led to a higher tumor uptake 24 h p.i. compared to the 68Ga·NODAGA-Anticalin imaged 60 min p.i. (2.5% ID/g vs 1.2% ID/g). Pronounced demetallation was observed for the 89Zr·Dfo-labeled PASylated Anticalin, which was ∼50% lower in the case of the cyclic radiochelator FsC (p < 0.0001). Adjusting the plasma half-life of Anticalin radioligands using PASylation technology is a viable approach to increase radioisotope accumulation within the tumor. Furthermore, 89Zr-ImmunoPET-imaging using the FsC radiochelator is superior to that using Dfo. Our strategy for the half-life adjustment of a tumor-targeting Anticalin to match the physical half-life of the applied radioisotope illustrates the potential of small binding proteins as an alternative to antibodies for PET-imaging.

Something old has become new: PET imaging of neural-crest tumors with [18F]-meta-fluorobenzylguanidine

Something old has become new: PET imaging of neural-crest tumors with [18F]-meta-fluorobenzylguanidine

A novel urokinase plasminogen activator receptor-targeted peptide-based probe for in-vivo molecular imaging of glioblastoma.

The urokinase plasminogen activator receptor (uPAR) is a promising biomarker for cancer diagnosis and therapy. We herein fabricated a new type of uPAR-targeted imaging probe Al18F-NOTA-VC and preliminarily evaluated its potential application in PET imaging of the glioma model in vivo. Peptide VC was synthesized and identified by MALDI-TOF-MS. The IC50 between VC/precursor NOTA-VC and uPAR was then determined before the synthesis and purification of Al18F-NOTA-VC, followed by further studies of in-vitro properties of Al18F-NOTA-VC. Meanwhile, the AE105-based probe followed a similar procedure in-vitro test. Finally, the PET imaging properties, including uPAR-targeting ability and the metabolism of Al18F-NOTA-VC, were investigated. The VC and NOTA-VC were obtained successfully and demonstrated a good affinity with uPAR. Followed by Al18F labeling successfully, excellent properties, including the serum stability, water solubility, and specificity of Al18F-NOTA-VC, were obtained in-vitro test compared with AE105 based probe. An excellent tumor uptake and renal excretion data of Al18F-NOTA-VC were acquired from in-vivo U87MG tumor model PET imaging, consistent with the subsequent biodistribution study. In addition to the excellent specificity and high tumor/normal tissue contrast for uPAR-targeted PET imaging of U87MG tumor, Al18F-NOTA-VC possessed promising clearance ability by renal system route. These excellent properties facilitated Al18F-NOTA-VC to be a promising imaging agent for uPAR high-expressing tumors and, thus, provided a paradigm for developing peptide-based probes for uPAR-associated disease diagnosis.