Application of a new 68Ga-labeled cNGR dimer peptide to microPET imaging of ovarian cancer

Abstract Introduction: Peptides containing the asparagine-glycine-arginine (NGR) sequence have been found to specifically bind to cluster of differentiation 13 (CD13) (aminopeptidase N), a tumor neovascular biomarker that is overexpressed on the surface of angiogenic blood vessels and various tumor cells and plays an important role in angiogenesis and tumor progression. The aim of this study was to evaluate the efficacy of a gallium-68 ( 68 Ga)-labeled dimeric cyclic NGR (cNGR) peptide as a new molecular probe that binds to CD13 in vitro and in vivo .Materials and Methods: A dimeric cNGR peptide conjugated with 1,4,7,10-tetraazacyclododecane-N,N',N'',N'''-tetraacetic acid (DOTA) and DOTA-c(NGR) 2 was synthesized and labeled with 68 Ga. In vitro uptake and binding analysis was performed in two ovarian tumor cell lines, ES2 and SKOV3, each of which have different expression levels of CD13. An in vivo biodistribution study was performed in normal mice, and micro positron emission tomography (PET) imaging was performed in nude mice xenografts with ES2 and SKOV3 tumors. Results : 68 Ga-DOTA-c(NGR) 2 with high radiochemical purity (>95%) was obtained and found to be stable at room temperature and when incubated with bovine serum at 37°C for 3 h. In vitro studies showed that uptake of 68 Ga-DOTA-c(NGR) 2 in ES2 cells increased over time, was higher than that in SKOV3 cells at all time points, and could be blocked by cold DOTA-c(NGR) 2 . Biodistribution studies demonstrated that 68 Ga-DOTA-c(NGR) 2 was mainly excreted from the kidney and rapidly cleared from blood. MicroPET imaging of ES2 tumor xenografts showed that focal uptake in tumors was distinctly observed from 1 to 1.5 h post-injection of 68 Ga-DOTA-c(NGR) 2 . Clear and high-contrast tumor visualization occurred at 1 h, which corresponded to the highest tumor/background ratio of 10.30±0.26. Moreover, accumulation of the probe in ES2 tumors apparently declined with pretreatment of unlabeled peptide, which further proved the specificity of 68 Ga-DOTA-c(NGR) 2 . In SKOV3 tumor models, the tumor was not obviously displayed under the same imaging protocols. Conclusion : We conclude that 68 Ga-DOTA-c(NGR) 2 might be a potential molecular probe for evaluating the expression levels of CD13 in different tumors, thereby providing a basis for targeting angiogenesis in cancer therapy.

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Effect of ethanol and cocaine on [11C]MPC-6827 uptake in SH-SY5Y cells

Molecular Biology Reports ◽

10.1007/s11033-021-06336-7 ◽

2021 ◽

Author(s):

Naresh Damuka ◽

Miranda Orr ◽

Paul W. Czoty ◽

Jeffrey L. Weiner ◽

Thomas J. Martin ◽

...

Keyword(s):

Mechanism Of Action ◽

Ex Vivo ◽

Neuroblastoma Cells ◽

Proper Function ◽

Brain Functions ◽

Biodistribution Studies ◽

Positron Emission ◽

Vitro Binding

AbstractMicrotubules (MTs) are structural units in the cytoskeleton. In brain cells they are responsible for axonal transport, information processing, and signaling mechanisms. Proper function of these processes is critical for healthy brain functions. Alcohol and substance use disorders (AUD/SUDs) affects the function and organization of MTs in the brain, making them a potential neuroimaging marker to study the resulting impairment of overall neurobehavioral and cognitive processes. Our lab reported the first brain-penetrant MT-tracking Positron Emission Tomography (PET) ligand [11C]MPC-6827 and demonstrated its in vivo utility in rodents and non-human primates. To further explore the in vivo imaging potential of [11C]MPC-6827, we need to investigate its mechanism of action. Here, we report preliminary in vitro binding results in SH-SY5Y neuroblastoma cells exposed to ethanol (EtOH) or cocaine in combination with multiple agents that alter MT stability. EtOH and cocaine treatments increased MT stability and decreased free tubulin monomers. Our initial cell-binding assay demonstrated that [11C]MPC-6827 may have high affinity to free/unbound tubulin units. Consistent with this mechanism of action, we observed lower [11C]MPC-6827 uptake in SH-SY5Y cells after EtOH and cocaine treatments (e.g., fewer free tubulin units). We are currently performing in vivo PET imaging and ex vivo biodistribution studies in rodent and nonhuman primate models of AUD and SUDs and Alzheimer's disease.

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Gold Nanoparticle-Based Fluorescent Theranostics for Real-Time Image-Guided Assessment of DNA Damage and Repair

International Journal of Molecular Sciences ◽

10.3390/ijms20030471 ◽

2019 ◽

Vol 20 (3) ◽

pp. 471 ◽

Cited By ~ 2

Author(s):

Shriya S. Srinivasan ◽

Rajesh Seenivasan ◽

Allison Condie ◽

Stanton L. Gerson ◽

Yanming Wang ◽

...

Keyword(s):

Molecular Imaging ◽

Real Time ◽

Targeted Delivery ◽

Specific Binding ◽

Imaging Techniques ◽

Cancer Cell Line ◽

Molecular Probe ◽

Biodistribution Studies

Chemotherapeutic dosing, is largely based on the tolerance levels of toxicity today. Molecular imaging strategies can be leveraged to quantify DNA cytotoxicity and thereby serve as a theranostic tool to improve the efficacy of treatments. Methoxyamine-modified cyanine-7 (Cy7MX) is a molecular probe which binds to apurinic/apyrimidinic (AP)-sites, inhibiting DNA-repair mechanisms implicated by cytotoxic chemotherapies. Herein, we loaded (Cy7MX) onto polyethylene glycol-coated gold nanoparticles (AuNP) to selectively and stably deliver the molecular probe intravenously to tumors. We optimized the properties of Cy7MX-loaded AuNPs using optical spectroscopy and tested the delivery mechanism and binding affinity using the DLD1 colon cancer cell line in vitro. A 10:1 ratio of Cy7MX-AuNPs demonstrated a strong AP site-specific binding and the cumulative release profile demonstrated 97% release within 12 min from a polar to a nonpolar environment. We further demonstrated targeted delivery using imaging and biodistribution studies in vivo in an xenografted mouse model. This work lays a foundation for the development of real-time molecular imaging techniques that are poised to yield quantitative measures of the efficacy and temporal profile of cytotoxic chemotherapies.

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New 55Co-labeled Albumin-Binding Folate Derivatives as Potential PET Agents for Folate Receptor Imaging

Pharmaceuticals ◽

10.3390/ph12040166 ◽

2019 ◽

Vol 12 (4) ◽

pp. 166 ◽

Cited By ~ 4

Author(s):

Lauren L. Radford ◽

Solana Fernandez ◽

Rebecca Beacham ◽

Retta El Sayed ◽

Renata Farkas ◽

...

Keyword(s):

Folate Receptor ◽

Small Animal ◽

Small Animal Pet ◽

Receptor Imaging ◽

Albumin Binding ◽

Liver Uptake ◽

Biodistribution Studies ◽

Positron Emission

Overexpression of folate receptors (FRs) on different tumor types (e.g., ovarian, lung) make FRs attractive in vivo targets for directed diagnostic/therapeutic agents. Currently, no diagnostic agent suitable for positron emission tomography (PET) has been adopted for clinical FR imaging. In this work, two 55Co-labeled albumin-binding folate derivatives-[55Co]Co-cm10 and [55Co]Co-rf42-with characteristics suitable for PET imaging have been developed and evaluated. High radiochemical yields (≥95%) and in vitro stabilities (≥93%) were achieved for both compounds, and cell assays demonstrated FR-mediated uptake. Both 55Co-labeled folate conjugates demonstrated high tumor uptake of 17% injected activity per gram of tissue (IA/g) at 4 h in biodistribution studies performed in KB tumor-bearing mice. Renal uptake was similar to other albumin-binding folate derivatives, and liver uptake was lower than that of previously reported [64Cu]Cu-rf42. Small animal PET/CT images confirmed the biodistribution results and showed the clear delineation of FR-expressing tumors.

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124I-Radiolabeling of a Au(III)-NHC Complex for in Vivo Biodistribution Studies

10.26434/chemrxiv.11996970 ◽

2020 ◽

Author(s):

Federica Guarra ◽

Alessio Terenzi ◽

Christine Pirker ◽

Rossana Passannante ◽

Dina Baier ◽

...

Keyword(s):

Anticancer Agents ◽

Direct Evidence ◽

Heterocyclic Carbenes ◽

Emission Tomography ◽

Direct Oxidation ◽

In Vivo Biodistribution ◽

Biodistribution Studies ◽

Positron Emission

Au(III) complexes with N-Heterocyclic Carbenes (NHCs) ligands have shown remarkable potential as anticancer agents, yet their fate in vivo has not been thoroughly examined and understood. Herein we report on the synthesis of new Au(III)-NHC complexes via direct oxidation with radioactive [124I]I2 as a valuable strategy to monitor the in vivo biodistribution of this class of compounds using positron emission tomography (PET) and, in combination with in vitro analyses, to provide direct evidence of the importance of Au(III)-to-Au(I) reduction for achieving full anticancer activity.

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[68Ga]Ga-DFO-c(RGDyK): Synthesis and Evaluation of Its Potential for Tumor Imaging in Mice

International Journal of Molecular Sciences ◽

10.3390/ijms22147391 ◽

2021 ◽

Vol 22 (14) ◽

pp. 7391

Author(s):

Sona Krajcovicova ◽

Andrea Daniskova ◽

Katerina Bendova ◽

Zbynek Novy ◽

Miroslav Soural ◽

...

Keyword(s):

Tumor Cell ◽

Tumor Imaging ◽

Ex Vivo ◽

Cell Uptake ◽

Αvβ3 Integrin ◽

In Vitro Characterization ◽

Positron Emission ◽

Gallium 68

Angiogenesis has a pivotal role in tumor growth and the metastatic process. Molecular imaging was shown to be useful for imaging of tumor-induced angiogenesis. A great variety of radiolabeled peptides have been developed to target αvβ3 integrin, a target structure involved in the tumor-induced angiogenic process. The presented study aimed to synthesize deferoxamine (DFO)-based c(RGD) peptide conjugate for radiolabeling with gallium-68 and perform its basic preclinical characterization including testing of its tumor-imaging potential. DFO-c(RGDyK) was labeled with gallium-68 with high radiochemical purity. In vitro characterization including stability, partition coefficient, protein binding determination, tumor cell uptake assays, and ex vivo biodistribution as well as PET/CT imaging was performed. [68Ga]Ga-DFO-c(RGDyK) showed hydrophilic properties, high stability in PBS and human serum, and specific uptake in U-87 MG and M21 tumor cell lines in vitro and in vivo. We have shown here that [68Ga]Ga-DFO-c(RGDyK) can be used for αvβ3 integrin targeting, allowing imaging of tumor-induced angiogenesis by positron emission tomography.

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Preliminary Study of New Gallium-68 Radiolabeled Peptide Targeting NRP-1 to Detect Brain Metastases by Positron Emission Tomography

Molecules ◽

10.3390/molecules26237273 ◽

2021 ◽

Vol 26 (23) ◽

pp. 7273

Author(s):

Albert Moussaron ◽

Valérie Jouan-Hureaux ◽

Charlotte Collet ◽

Julien Pierson ◽

Noémie Thomas ◽

...

Keyword(s):

Positron Emission Tomography ◽

Water Solubility ◽

Fatal Outcome ◽

Emission Tomography ◽

Diagnostic Tools ◽

In Vitro Toxicity ◽

Positron Emission ◽

Gallium 68

Due to their very poor prognosis and a fatal outcome, secondary brain tumors are one of the biggest challenges in oncology today. From the point of view of the early diagnosis of these brain micro- and macro-tumors, the sensitivity and specificity of the diagnostic tools constitute an obstacle. Molecular imaging, such as Positron Emission Tomography (PET), is a promising technique but remains limited in the search for cerebral localizations, given the commercially available radiotracers. Indeed, the [18F]FDG PET remains constrained by the physiological fixation of the cerebral cortex, which hinders the visualization of cerebral metastases. Tumor angiogenesis is recognized as a crucial phenomenon in the progression of malignant tumors and is correlated with overexpression of the neuropilin-1 (NRP-1) receptor. Here, we describe the synthesis and the photophysical properties of the new gallium-68 radiolabeled peptide to target NRP-1. The KDKPPR peptide was coupled with gallium-68 anchored into a bifunctional NODAGA chelating agent, as well as Cy5 for fluorescence detection. The Cy5 absorbance spectra did not change, whereas the molar extinction coefficient (ε) decreased drastically. An enhancement of the fluorescence quantum yield (ΦF) could be observed due to the better water solubility of Cy5. [68Ga]Ga-NODAGA-K(Cy5)DKPPR was radiosynthesized efficiently, presented hydrophilic properties (log D = −1.86), and had high in vitro stability (>120 min). The molecular affinity and the cytotoxicity of this new chelated radiotracer were evaluated in vitro on endothelial cells (HUVEC) and MDA-MB-231 cancer cells (hormone-independent and triple-negative line) and in vivo on a brain model of metastasis in a nude rat using the MDA-MB-231 cell line. No in vitro toxicity has been observed. The in vivo preliminary experiments showed promising results, with a high contrast between the healthy brain and metastatic foci for [68Ga]Ga-NODAGA-K(Cy5)DKPPR.

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124I-Radiolabeling of a Au(III)-NHC Complex for in Vivo Biodistribution Studies

10.26434/chemrxiv.11996970.v2 ◽

2020 ◽

Author(s):

Federica Guarra ◽

Alessio Terenzi ◽

Christine Pirker ◽

Rossana Passannante ◽

Dina Baier ◽

...

Keyword(s):

Anticancer Agents ◽

Direct Evidence ◽

Heterocyclic Carbenes ◽

Emission Tomography ◽

Direct Oxidation ◽

In Vivo Biodistribution ◽

Biodistribution Studies ◽

Positron Emission

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Microwave-assisted radiolabelling of 52Mn-porphyrins

10.26434/chemrxiv-2021-v8wm5 ◽

2021 ◽

Author(s):

Peter J. Gawne ◽

Sara M. A. Pinto ◽

Karin M. Nielsen ◽

Mariette M. Pereira ◽

Rafael T. M. de Rosales

Keyword(s):

Lipid Bilayers ◽

Mri Contrast Agents ◽

Mri Contrast ◽

Manganese Porphyrins ◽

Biodistribution Studies ◽

Positron Emission ◽

Radiochemical Yields

Manganese porphyrins have several therapeutic/imaging applications; including their use as radioprotectants (in clinical trials), and as paramagnetic MRI contrast agents. The affinity of porphyrins for lipid bilayers also makes them candidates for cell/liposome labelling. We hypothesised that metalation with the positron emission tomography (PET) radionuclide 52Mn (t1/2 = 5.6 d) would allow long-term in vivo biodistribution studies of Mn-porphyrins as well as a method to label and track cells/liposomes, but methods for fast and efficient radiolabelling are lacking. Several porphyrins were produced and radiolabelled by addition to neutralised [52Mn]MnCl2 and heated at 165 oC for 1 h using a microwave (MW) synthesiser at a ligand concentration of 0.6 – 0.7 mM. These conditions were compared with non-MW heating at 70oC. MW radiosynthesis allowed >95 % radiochemical yields (RCY) in just 1 h. Conversely, non-MW heating at 70 oC for 1 h resulted in low RCY (0 – 25 % RCY) and most porphyrins did not reach completion after 24h. Formation of the 52Mn-complexes were confirmed with radio-HPLC by comparison with their non-radioactive 55Mn counterparts. Following this, several 52Mn-porphyrins were used to radiolabel liposomes by incubation at 50 oC for 30 min resulting in 75 – 86 % labelling efficiency (LE). Two lead 52Mn-porphyrins were taken forward to label MDA-MB-231 cancer cells in vitro, achieving ca. 11 % LE. After 24 h, 32 – 45 % of the 52Mn-porphyrin was retained in cells. In contrast to standard methods, MW heating allows fast synthesis of 52Mn-porphyrins with >95% radiochemical yields that avoid purification. 52Mn-porphyrins also show promising cell/liposome labelling properties. This technique can potentially be exploited for the in vivo imaging of Mn-porphyrin therapeutics, as well as for the accurate in vivo quantification of Mn-porphyrin MRI agents.

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Exploiting the Concept of Multivalency with 68Ga- and 89Zr-Labelled Fusarinine C-Minigastrin Bioconjugates for Targeting CCK2R Expression

Contrast Media & Molecular Imaging ◽

10.1155/2018/3171794 ◽

2018 ◽

Vol 2018 ◽

pp. 1-12 ◽

Cited By ~ 5

Author(s):

Dominik Summer ◽

Christine Rangger ◽

Maximilian Klingler ◽

Peter Laverman ◽

Gerben M. Franssen ◽

...

Keyword(s):

Metabolic Stability ◽

Cell Uptake ◽

Late Time ◽

Biodistribution Studies ◽

Imaging Probes ◽

Gallium 68 ◽

Late Time Point ◽

Time Point

Cholecystokinin-2 receptors (CCK2R) are overexpressed in a variety of malignant diseases and therefore have gained certain attention for peptide receptor radionuclide imaging. Among extensive approaches to improve pharmacokinetics and metabolic stability of minigastrin (MG) based radioligands, the concept of multivalency for enhanced tumour targeting has not been investigated extensively. We therefore utilized fusarinine C (FSC) as chelating scaffold for novel mono-, di-, and trimeric bioconjugates for targeting CCK2R expression. FSC-based imaging probes were radiolabelled with positron emitting radionuclides (gallium-68 and zirconium-89) and characterized in vitro (log⁡D, IC50, and cell uptake) and in vivo (metabolic stability in BALB/c mice, biodistribution profile, and microPET/CT imaging in A431-CCK2R/A431-mock tumour xenografted BALB/c nude mice). Improved targeting did not fully correlate with the grade of multimerization. The divalent probe showed higher receptor affinity and increased CCK2R mediated cell uptake while the trimer remained comparable to the monomer. In vivo biodistribution studies 1 h after administration of the 68Ga-labelled radioligands confirmed this trend, but imaging at late time point (24 h) with 89Zr-labelled counterparts showed a clearly enhanced imaging contrast of the trimeric probe compared to the mono- and dimer. Furthermore, in vivo stability studies showed a higher metabolic stability for multimeric probes compared to the monomeric bioconjugate. In summary, we could show that FSC can be utilized as suitable scaffold for novel mono- and multivalent imaging probes for CCK2R-related malignancies with partly improved targeting properties for multivalent conjugates. The increased tumour accumulation of the trimer 24 h postinjection (p.i.) can be explained by slower clearance and increased metabolic stability of multimeric conjugates.

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Immuno-PET Imaging of the Programmed Cell Death-1 Ligand (PD-L1) Using a Zirconium-89 Labeled Therapeutic Antibody, Avelumab

Molecular Imaging ◽

10.1177/1536012119829986 ◽

2019 ◽

Vol 18 ◽

pp. 153601211982998 ◽

Cited By ~ 9

Author(s):

Elaine M. Jagoda ◽

Olga Vasalatiy ◽

Falguni Basuli ◽

Ana Christina L. Opina ◽

Mark R. Williams ◽

...

Keyword(s):

Target Tissue ◽

Dependent Manner ◽

Binding Studies ◽

Expression Levels ◽

High Affinity ◽

Biodistribution Studies ◽

Programmed Cell Death 1 ◽

Spleen And Lymph Nodes

Objective: The goal is to evaluate avelumab, an anti-PD-L1 monoclonal immunoglobulin G antibody labeled with zirconium-89 in human PD-L1-expressing cancer cells and mouse xenografts for clinical translation. Methods: [89Zr]Zr-DFO-PD-L1 monoclonal antibody (mAb) was synthesized using avelumab conjugated to desferrioxamine. In vitro binding studies and biodistribution studies were performed with PD-L1+MDA-MB231 cells and MDA-MB231 xenograft mouse models, respectively. Biodistributions were determined at 1, 2, 3, 5, and 7 days post coinjection of [89Zr]Zr-DFO-PD-L1 mAb without or with unlabeled avelumab (10, 20, 40, and 400 µg). Results: [89Zr]Zr-DFO-PD-L1 mAb exhibited high affinity (Kd ∼ 0.3 nM) and detected moderate PD-L1 expression levels in MDA-MB231 cells. The spleen and lymph nodes exhibited the highest [89Zr]Zr-DFO-PD-L1 mAb uptakes in all time points, while MDA-MB231 tumor uptakes were lower but highly retained. In the unlabeled avelumab dose escalation studies, spleen tissue–muscle ratios decreased in a dose-dependent manner indicating specific [89Zr]Zr-DFO-PD-L1 mAb binding to PD-L1. In contrast, lymph node and tumor tissue–muscle ratios increased 4- to 5-fold at 20 and 40 µg avelumab doses. Conclusions: [89Zr]Zr-DFO-PD-L1 mAb exhibited specific and high affinity for PD-L1 in vitro and had target tissue uptakes correlating with PD-L1 expression levels in vivo. [89Zr]Zr-DFO-PD-L1 mAb uptake in PD-L1+tumors increased with escalating doses of avelumab.

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