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89Zr-Onartuzumab PET imaging of c-MET receptor dynamics.

ABSTRACT: PURPOSE:c-MET and its ligand hepatocyte growth factor are often dysregulated in human cancers. Dynamic changes in c-MET expression occur and might predict drug efficacy or emergence of resistance. Noninvasive visualization of c-MET dynamics could therefore potentially guide c-MET-directed therapies. We investigated the feasibility of 89Zr-labelled one-armed c-MET antibody onartuzumab PET for detecting relevant changes in c-MET levels induced by c-MET-mediated epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor erlotinib resistance or heat shock protein-90 (HSP90) inhibitor NVP-AUY-922 treatment in human non-small-cell lung cancer (NSCLC) xenografts. METHODS:In vitro membrane c-MET levels were determined by flow cytometry. HCC827ErlRes, an erlotinib-resistant clone with c-MET upregulation, was generated from the exon-19 EGFR-mutant human NSCLC cell line HCC827. Mice bearing HCC827 and HCC827ErlRes tumours in opposite flanks underwent 89Zr-onartuzumab PET scans. The HCC827-xenografted mice underwent 89Zr-onartuzumab PET scans before treatment and while receiving biweekly intraperitoneal injections of 100 mg/kg NVP-AUY-922 or vehicle. Ex vivo, tumour c-MET immunohistochemistry was correlated with the imaging results. RESULTS:In vitro, membrane c-MET was upregulated in HCC827ErlRes tumours by 213?±?44% in relation to the level in HCC827 tumours, while c-MET was downregulated by 69?±?9% in HCC827 tumours following treatment with NVP-AUY-922. In vivo, 89Zr-onartuzumab uptake was 26% higher (P?

SUBMITTER: Pool M

PROVIDER: S-EPMC5486818 | biostudies-literature | 2017 Aug

REPOSITORIES: biostudies-literature

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<sup>89</sup>Zr-Onartuzumab PET imaging of c-MET receptor dynamics.

Pool Martin M Terwisscha van Scheltinga Anton G T AGT Kol Arjan A Giesen Danique D de Vries Elisabeth G E EGE Lub-de Hooge Marjolijn N MN

European journal of nuclear medicine and molecular imaging 20170319 8

<h4>Purpose</h4>c-MET and its ligand hepatocyte growth factor are often dysregulated in human cancers. Dynamic changes in c-MET expression occur and might predict drug efficacy or emergence of resistance. Noninvasive visualization of c-MET dynamics could therefore potentially guide c-MET-directed therapies. We investigated the feasibility of <sup>89</sup>Zr-labelled one-armed c-MET antibody onartuzumab PET for detecting relevant changes in c-MET levels induced by c-MET-mediated epidermal growth ...[more]

PMID: 28315949

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Project description:Methods that provide rapid access to radiolabeled antibodies are vital in the development of diagnostic and radiotherapeutic agents for PET or radioimmunotherapy. The human hepatocyte growth factor receptor (c-MET) signaling pathway is dysregulated in several malignancies, including gastric cancer, and is an important biomarker in drug discovery. Here, we used a photoradiochemical approach to produce 89Zr-radiolabeled onartuzumab (a monovalent, antihuman c-MET antibody), starting directly from the fully formulated drug (MetMAb). Methods: Simultaneous 89Zr-radiolabeling and protein conjugation was performed in one-pot reactions containing 89Zr-oxalate, the photoactive chelate desferrioxamine B (DFO)-aryl azide (DFO-ArN3), and MetMAb to give 89Zr-DFO-azepin-onartuzumab. As a control, 89Zr-DFO-benzyl Bn-isothiocyanate Bn-NCS-onartuzumab was prepared via a conventional two-step process using prepurified onartuzumab and DFO-Bn-NCS. Radiotracers were purified by using size-exclusion methods and evaluated by radiochromatography. Radiochemical stability was studied in human serum, and immunoreactivity was determined by cellular binding assays using MKN-45 gastric carcinoma cells. PET imaging at multiple time points (0-72 h) was performed on female athymic nude mice bearing subcutaneous MKN-45 xenografts. Biodistribution experiments were performed after the final image was obtained. The tumor specificity of 89Zr-DFO-azepin-onartuzumab was assessed in vivo by competitive inhibition (blocking) studies. Results: Initial photoradiosynthesis experiments produced 89Zr-DFO-azepin-onartuzumab in less than 15 min, with an isolated decay-corrected radiochemical yield (RCY) of 24.8%, a radiochemical purity of approximately 90%, and a molar activity of approximately 1.5 MBq nmol-1 Reaction optimization improved the radiochemical conversion of 89Zr-DFO-azepin-onartuzumab to 56.9% ± 4.1% (n = 3), with isolated RCYs of 41.2% ± 10.6% (n = 3) and radiochemical purity of more than 90%. Conventional methods produced 89Zr-DFO-Bn-NCS-onartuzumab with an isolated RCY of more than 97%, radiochemical purity of more than 97% and molar activity of approximately 14.0 MBq nmol-1 Both radiotracers were immunoreactive and stable in human serum. PET imaging and biodistribution studies showed high tumor uptake for both radiotracers. By 72 h, tumor and liver uptake (percentage injected dose [%ID]) reached 15.37 ± 5.21 %ID g-1 and 6.56 ± 4.03 %ID g-1, respectively, for 89Zr-DFO-azepin-onartuzumab (n = 4) and 21.38 ± 11.57 %ID g-1 and 18.84 ± 6.03 %ID g-1, respectively, for 89Zr-DFO-Bn-NCS-onartuzumab (n = 4). Blocking experiments gave a statistically significant reduction in tumor uptake (6.34 ± 0.47 %ID g-1) of 89Zr-DFO-azepin-onartuzumab (n = 4). Conclusion: The experiments demonstrated that photoradiosynthesis is a viable alternative approach for producing 89Zr-radiolabeled antibodies directly in protein formulation buffer, reducing protein aggregation and liver uptake.

Project description:BackgroundAmivantamab is a novel bispecific antibody that simultaneously targets the epidermal growth factor receptor (EGFR) and the hepatocyte growth factor receptor (HGFR/c-MET) that are overexpressed in several types of cancer including triple-negative breast cancer (TNBC). Targeting both receptors simultaneously can overcome resistance to mono-targeted therapy. The purpose of this study is to develop 89Zr-labeled amivantamab as a potential companion diagnostic imaging agent to amivantamab therapy using various preclinical models of TNBC for evaluation.MethodsAmivantamab was conjugated to desferrioxamine (DFO) and radiolabeled with 89Zr to obtain [89Zr]ZrDFO-amivantamab. Binding of the bispecific [89Zr]ZrDFO-amivantamab as well as its mono-specific "single-arm" antibody controls were determined in vitro and in vivo. Biodistribution studies of [89Zr]ZrDFO-amivantamab were performed in MDA-MB-468 xenografts to determine the optimal imaging time point. PET/CT imaging with [89Zr]ZrDFO-amivantamab or its isotype control was performed in a panel of TNBC xenografts with varying levels of EGFR and c-MET expression.Results[89Zr]ZrDFO-amivantamab was synthesized with a specific activity of 148 MBq/mg and radiochemical yield of ≥ 95%. Radioligand binding studies and western blot confirmed the order of EGFR and c-MET expression levels: HCC827 lung cancer cell (positive control) > MDA-MB-468 > MDA-MB-231 > MDA-MB-453. [89Zr]ZrDFO-amivantamab demonstrated bispecific binding in cell lines co-expressed with EGFR and c-MET. PET/CT imaging with [89Zr]ZrDFO-amivantamab in TNBC xenografted mice showed standard uptake value (SUVmean) of 6.0 ± 1.1 in MDA-MB-468, 4.2 ± 1.4 in MDA-MB-231, and 1.5 ± 1.4 in MDA-MB-453 tumors, which are consistent with their receptors' expression levels on the cell surface.ConclusionWe have successfully prepared a radiolabeled bispecific antibody, [89Zr]ZrDFO-amivantamab, and evaluated its pharmacologic and imaging properties in comparison with its single-arm antibodies and non-specific isotype controls. [89Zr]ZrDFO-amivantamab demonstrated the greatest uptake in tumors co-expressing EGFR and c-MET.

Project description:PurposeProgrammed cell death-1 receptor (PD-1) and its ligand (PD-L1) are the targets for immunotherapy in many cancer types. Although PD-1 blockade has therapeutic effects, the efficacy differs between patients. Factors contributing to this variability are PD-L1 expression levels and immune cells present in tumors. However, it is not well understood how PD-1 expression in the tumor microenvironment impacts immunotherapy response. Thus, imaging of PD-1-expressing immune cells is of interest. This study aims to evaluate the biodistribution of Zirconium-89 (89Zr)-labeled pembrolizumab, a humanized IgG4 kappa monoclonal antibody targeting PD-1, in healthy cynomolgus monkeys as a translational model of tracking PD-1-positive immune cells.ProceduresPembrolizumab was conjugated with the tetrafluorophenol-N-succinyl desferal-Fe(III) ester (TFP-N-sucDf) and subsequently radiolabeled with 89Zr. Four cynomolgus monkeys with no previous exposure to humanized monoclonal antibodies received tracer only or tracer co-injected with pembrolizumab intravenously over 5 min. Thereafter, a static whole-body positron emission tomography (PET) scan was acquired with 10 min per bed position on days 0, 2, 5, and 7. Image-derived standardized uptake values (SUVmean) were quantified by region of interest (ROI) analysis.Results89Zr-N-sucDf-pembrolizumab was synthesized with high radiochemical purity (> 99 %) and acceptable molar activity (> 7 MBq/nmol). In animals dosed with tracer only, 89Zr-N-sucDf-pembrolizumab distribution in lymphoid tissues such as mesenteric lymph nodes, spleen, and tonsils increased over time. Except for the liver, low radiotracer distribution was observed in all non-lymphoid tissue including the lung, muscle, brain, heart, and kidney. When a large excess of pembrolizumab was co-administered with a radiotracer, accumulation in the lymph nodes, spleen, and tonsils was reduced, suggestive of target-mediated accumulation.Conclusions89Zr-N-sucDf-pembrolizumab shows preferential uptake in the lymphoid tissues including the lymph nodes, spleen, and tonsils. 89Zr-N-sucDf-pembrolizumab may be useful in tracking the distribution of a subset of immune cells in non-human primates and humans.Trial registrationClinicalTrials.gov Identifier: NCT02760225.

Project description:BackgroundIn vivo monitoring of cell biodistribution using positron emission tomography (PET) provides a quantitative non-invasive method to further optimize cell therapies and related new developments in the field. Our group has earlier optimized and evaluated the in vitro properties of two radiotracers,[89Zr]Zr-(oxinate)4 and [89Zr]Zr-DFO-NCS, for the radiolabelling of different cell types. Here, we performed a microPET study to assess the in vivo biodistribution of cells in rats using these two radiotracers. Human decidual stromal cells (hDSC) and rat macrophages (rMac) were radiolabelled with [89Zr]Zr-(oxinate)4 or [89Zr]Zr-DFO-NCS. Rats were intravenously injected with radiolabelled cells, and the in vivo biodistribution was monitored with microPET/CT imaging for up to day 7. Organ uptake was evaluated and presented as a percentage of injected activity per gram tissue (%IA/g) and total absorbed organ doses (mSv/MBq).ResultsThe biodistribution in vivo showed an immediate uptake in the lungs. Thereafter, [89Zr]Zr-(oxinate)4 labelled cells migrated to the liver, while the signal from [89Zr]Zr-DFO-NCS labelled cells lingered in the lungs. The differences in the in vivo behaviour for the same cell type appeared related to the radiotracer labelling. After 24 h, [89Zr]Zr-(oxinate)4 labelled cells had over 70% higher liver uptake for both hDSC and rMac compared to [89Zr]Zr-DFO-NCS labelled cells, whereas [89Zr]Zr-DFO-NCS labelled cells showed over 60% higher uptake in the lungs compared to [89Zr]Zr-(oxinate)4 labelled cells. This difference in both lung and liver uptake continued until day 7. Dosimetry calculations showed a higher effective dose (mSv/MBq) for [89Zr]Zr-DFO-NCS compared to [89Zr]Zr-(oxinate)4, for both cell types. Although the bone uptake was higher for [89Zr]Zr-(oxinate)4 labelled cells, the prolonged uptake in the lungs contributed to a significant crossfire to bone marrow resulting in a higher bone dose.ConclusionThe [89Zr]Zr-DFO-NCS labelled cells suggest a prolonged accumulation in the lungs, while [89Zr]Zr-(oxinate)4 suggests quicker clearance of the lungs followed by accumulation in the liver. Accumulation of radiolabelled cells in the liver corresponds to other cell-tracking methods. Further studies are required to determine the actual location of the [89Zr]Zr-DFO-NCS labelled cell.

Project description:The encapsulation of Glucocorticoids (GCs) into long-circulating liposomes (LCLs) is a proven strategy to reduce the side effects of glucocorticoids and improve the treatment of inflammatory diseases, such as rheumatoid arthritis (RA). With the aim of supporting the development of GC-loaded LCLs, and potentially predict patient response to therapy clinically, we evaluated a direct PET imaging radiolabelling approach for preformed GC-LCLs in an animal model of human inflammatory arthritis. Methods: A preformed PEGylated liposomal methylprednisolone hemisuccinate (NSSL-MPS) nanomedicine was radiolabelled using [89Zr]Zr(oxinate)4 (89Zr-oxine), characterised and tracked in vivo using PET imaging in a K/BxN serum-transfer arthritis (STA) mouse model of inflammatory arthritis and non-inflamed controls. Histology and joint size measurements were used to confirm inflammation. The biodistribution of 89Zr-NSSL-MPS was compared to that of free 89Zr in the same model. A therapeutic study using NSSL-MPS using the same time points as the PET/CT imaging was carried out. Results: The radiolabelling efficiency of NSSL-MPS with [89Zr]Zr(oxinate)4 was 69 ± 8 %. PET/CT imaging of 89Zr-NSSL-MPS showed high uptake (3.6 ± 1.5 % ID; 17.4 ± 9.3 % ID/mL) at inflamed joints, with low activity present in non-inflamed joints (0.5 ± 0.1 % ID; 2.7 ± 1.1 % ID/mL). Importantly, a clear correlation between joint swelling and high 89Zr-NSSL-MPS uptake was observed, which was not observed with free 89Zr. STA mice receiving a therapeutic dose of NSSL-MPS showed a reduction in inflammation at the time points used for the PET/CT imaging compared with the control group. Conclusions: PET imaging was used for the first time to track a liposomal glucocorticoid, showing high uptake at visible and occult inflamed sites and a good correlation with the degree of inflammation. A subsequent therapeutic response matching imaging time points in the same model demonstrated the potential of this radiolabeling method as a theranostic tool for the prediction of therapeutic response - with NSSL-MPS and similar nanomedicines - in the treatment of inflammatory diseases.

Dataset Information

89Zr-Onartuzumab PET imaging of c-MET receptor dynamics.

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<sup>89</sup>Zr-Onartuzumab PET imaging of c-MET receptor dynamics.

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