Dataset Information

Immuno-PET imaging of VEGFR-2 expression in prostate cancer with ⁸⁹Zr-labeled ramucirumab.

ABSTRACT: The detection and monitoring of prostate cancer (PrCa) malignancies using most of the conventional strategies is challenging. As an over-expressed biomarker of PrCa, the vascular endothelial growth factor receptor 2 (VEGFR-2) can be delineated by non-invasive imaging to address such issue. Herein, we report the positron emission tomography (PET) of VEGFR-2 expression in a PrCa mice models by composing a novel tracer, [⁸⁹Zr]zirconium-labeled clinical VEGFR-2 antibody (Ramucirumab), i.e. ⁸⁹Zr-Df-R. The VEGFR-2 expression levels among three different PrCa cell lines (PC-3, LNCAP and LAPC-4) were confirmed by flow cytometry. The immuno-PET imaging and bio-distribution (Bio-D) study were conducted in subcutaneous PrCa mice models via the ⁸⁹Zr-Df-R. The regions of interest (ROI) data showed that the uptake of ⁸⁹Zr-Df-R in the positive PC-3 (9.5±3 %ID/g) tumors are obviously higher than those ones in the negative LNCAP (6.0±1.7 %ID/g) or LAPC-4 (4.3±0.7 %ID/g) tumors at 120 hours post-injection, while the accumulation of ⁸⁹Zr-Df-R in PC-3 tumors (4.3±1.2 %ID/g)) could be significantly reduced by the blockade of unlabeled Ramucirumab. These quantitative data coincide with the Bio-D data and proves the specificity. Additionally, the immuno-fluorescent staining results confirmed the expression pattern of VEGFR-2 among various PrCa tumors. Finally, the flow cytometry of PC-3 tumor tissue further proved that the binding of ⁸⁹Zr-Df-R to VEGFR-2 primarily occurs on the PC-3 tumor cells. In summary, the description of the VEGFR-2 expression in PrCa by in-vivo PET with ⁸⁹Zr-Df-R is feasible and it may shed light on the early detection of foci and dynamic monitoring of anti-VEGFR-2 therapy in PrCa.

SUBMITTER: Li M

PROVIDER: S-EPMC6780657 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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Immuno-PET imaging of VEGFR-2 expression in prostate cancer with <sup>89</sup>Zr-labeled ramucirumab.

Li Miao M Jiang Dawei D Barnhart Todd E TE Cao Tianye T Engle Jonathan W JW Chen Weiyu W Cai Weibo W

American journal of cancer research 20190901 9

The detection and monitoring of prostate cancer (PrCa) malignancies using most of the conventional strategies is challenging. As an over-expressed biomarker of PrCa, the vascular endothelial growth factor receptor 2 (VEGFR-2) can be delineated by non-invasive imaging to address such issue. Herein, we report the positron emission tomography (PET) of VEGFR-2 expression in a PrCa mice models by composing a novel tracer, [<sup>89</sup>Zr]zirconium-labeled clinical VEGFR-2 antibody (Ramucirumab), i.e ...[more]

PMID: 31598404

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Project description:Antibody-based PET tracers are exceptionally well-suited for determination of the in vivo biodistribution and quantification of therapeutic antibodies. The continued expansion in antibody-based therapeutics has accordingly driven the development towards more robust conjugation strategies in order to reliably predict the performance of such agents. We therefore aimed to evaluate the effect of site-specific labeling by enzymatic remodeling on the stability, immuno-reactivity and tumor-targeting properties of the monoclonal antibody (mAb) trastuzumab and compare it to conventional, random labeling in a HER2-positive xenograft mouse model. Methods: Trastuzumab was conjugated with the p-SCN-Bn-Desferrioxamine (SCN-Bn-DFO) chelator randomly on lysine residues or site-specifically on enzymatically modified glycans using either β-galactosidase or endoglycosidase S2 prior to 89Zr radiolabeling. 89Zr-DFO-trastuzumab was injected into SK-OV-3 tumor-bearing NMRI nude mice. The antibody dose was titrated with either 100 µg or 500 µg of unlabeled trastuzumab. Mice underwent small animal PET/CT imaging 24, 70 and 120 hours post-injection for longitudinal assessment. Parallel experiments were conducted with an isotype control matched antibody. In vivo imaging was supported by conventional ex vivo biodistribution and HER2 immuno-histochemistry. Furthermore, site-specifically labeled 89Zr-DFO-trastuzumab was evaluated in a panel of subcutaneous patient-derived xenograft (PDX) models. Additionally, the affinity, in vitro stability and immuno-reactivity were assessed for all tracers. Results: Site-specific labeling significantly increased PET tumor uptake (One-way ANOVA, p<0.0001) at all time-points when compared to random labeling. Mean tumor uptakes were 6.7 ± 1.7, 13.9 ± 3.3 and 15.3 ± 3.8 % injected dose per gram tissue (%ID/g) at 70 hours post-injection, for random, β-galactosidase or endoglycosidase S2 labeled probes, respectively. Co-injection with unlabeled trastuzumab increased the circulation time of tracers but did not alter tumor uptake notably. Site-specific probes presented with a superior in vitro stability and immuno-reactivity compared to the randomly labeled probe. Ex vivo biodistribution confirmed the data obtained by in vivo PET imaging, and site-specific 89Zr-DFO-trastuzumab successfully detected HER2-positive tumors in PDX mouse models. Conclusion: 89Zr-DFO-trastuzumab is well-matched for specific immuno-PET imaging of HER2-positive tumors and site-specific labeling of trastuzumab by the SiteClickTM technology minimizes the impact of the DFO chelator on immuno-reactivity, stability and biodistribution. These findings support further development of site-specifically radiolabeled mAbs for immuno-PET.

Project description:PurposeWe present here a Zr-89-labeled inhibitor of prostate-specific membrane antigen (PSMA) as a complement to the already established F-18- or Ga-68-ligands.ProceduresThe precursor PSMA-DFO (ABX) was used for Zr-89-labeling. This is not an antibody, but a peptide analogue of the precursor for the production of [177Lu]Lu-PSMA-617. The ligand [89Zr]Zr-PSMA-DFO was compared with [68Ga]Ga-PSMA-11 and [18F]F-JK-PSMA-7 in vitro by determination of the Kd value, cellular uptake, internalization in LNCaP cells, biodistribution studies with LNCaP prostate tumor xenografts in mice, and in vivo by small-animal PET imaging in LNCaP tumor mouse models. A first-in-human PET was performed with [89Zr]Zr-PSMA-DFO on a patient presenting with a biochemical recurrence after brachytherapy and an ambiguous intraprostatic finding with [18F]F-JK-PSMA-7 but histologically benign cells in a prostate biopsy 7 months previously.Results[89Zr]Zr-PSMA-DFO was prepared with a radiochemical purity ≥ 99.9% and a very high in vitro stability for up to 7 days at 37 °C. All radiotracers showed similar specific cellular binding and internalization, in vitro and comparable tumor uptake in biodistribution experiments during the first 5 h. The [89Zr]Zr-PSMA-DFO achieved significantly higher tumor/background ratios in LNCaP tumor xenografts (tumor/blood: 309 ± 89, tumor/muscle: 450 ± 38) after 24 h than [68Ga]Ga-PSMA-11 (tumor/blood: 112 ± 57, tumor/muscle: 58 ± 36) or [18F]F-JK-PSMA-7 (tumor/blood: 175 ± 30, tumor/muscle: 114 ± 14) after 4 h (p < 0.01). Small-animal PET imaging demonstrated in vivo that tumor visualization with [89Zr]Zr-PSMA-DFO is comparable to [68Ga]Ga-PSMA-11 or [18F]F-JK-PSMA-7 at early time points (1 h p.i.) and that PET scans up to 48 h p.i. clearly visualized the tumor at late time points. A late [89Zr]Zr-PSMA-DFO PET scan on a patient with biochemical recurrence (BCR) had demonstrated intensive tracer accumulation in the right (SUVmax 13.25, 48 h p.i.) and in the left prostate lobe (SUV max 9.47), a repeat biopsy revealed cancer cells on both sides.Conclusion[89Zr]Zr-PSMA-DFO is a promising PSMA PET tracer for detection of tumor areas with lower PSMA expression and thus warrants further clinical evaluation.

Project description:Lung cancer accounts for 17% of cancer-related deaths worldwide, and most patients present with locally advanced or metastatic disease. Novel PET imaging agents for assessing vascular endothelial growth factor receptor-2 (VEGFR-2) expression can be used for detecting VEGFR-2-positive malignancies and subsequent monitoring of therapeutic response to VEGFR-2-targeted therapies. Here, we report the synthesis and characterization of an antibody-based imaging agent for PET imaging of VEGFR-2 expression in vivo.Ramucirumab (named RamAb), a fully humanized IgG1 monoclonal antibody, was conjugated to 2-S-(4-isothiocyanatobenzyl)-1,4,7-triazacyclononane-1,4,7-triacetic acid (p-SCN-Bn-NOTA) and labeled with (64)Cu. Flow cytometry analysis and microscopy studies were performed to compare the VEGFR-2 binding affinity of RamAb and NOTA-RamAb. PET imaging and biodistribution studies were performed in nude mice bearing HCC4006 and A549 xenograft tumors. Ex vivo histopathology was performed to elucidate the expression patterns of VEGFR-2 in different tissues and organs to validate in vivo results.Flow cytometry examination revealed the specific binding capacity of fluorescein isothiocyanate-RamAb to VEGFR-2, and no difference in VEGFR-2 binding affinity was seen between RamAb and NOTA-RamAb. After being labeled with (64)Cu, PET imaging revealed specific and prominent uptake of (64)Cu-NOTA-RamAb in VEGFR-2-positive HCC4006 tumors (9.4 ± 0.5 percentage injected dose per gram at 48 h after injection; n = 4) and significantly lower uptake in VEGFR-2-negative A549 tumors (4.3 ± 0.2 percentage injected dose per gram at 48 h after injection; n = 3). Blocking experiments revealed significantly lower uptake in HCC4006 tumors, along with histology analysis, further confirming the VEGFR-2 specificity of (64)Cu-NOTA-RamAb.This study provides initial evidence that (64)Cu-NOTA-RamAb can function as a PET imaging agent for visualizing VEGFR-2 expression in vivo, which may also find potential applications in monitoring the treatment response of VEGFR-2-targeted cancer therapy.

Project description:Avian pathogenic Escherichia coli (APEC) is a serious systemic infectious disease in poultry infections, causing severe economic losses to the poultry industry. Previous studies have shown that secretion of virulence proteins was required for the pathogenicity of APEC through the secretion system. Outer membrane vesicles (OMVs) are a generalized secretion system of Gram-negative bacteria that play a key role in the long-distance delivery of virulence factors, but whether they are associated with the pathogenic mechanism of APEC has not been determined. In this study, OMVs were purified and characterized from AE17 (O2 serotype) by ultracentrifugation and density gradient centrifugation and their protein cargo was identified using liquid chromatography-tandem mass spectrometry (LC-MS/MS). In addition, 89Zr was labeled after chelating AE17 OMVs by DFO and positron emission tomography PET imaging was used to track 89Zr-DFO-OMVs in chickens and to pathologically analyze the distribution sites. This study showed that AE17 OMVs were membrane vesicles ranging in size from 20 to 200 nm and proteomic analysis revealed the presence of virulence proteins, including adhesion proteins OmpA, OmpC, OmpF, OmpX, FimH, FimC and FigE, and serum resistance proteins OmpT and MliC and immune response regulator proteins (FliC). In addition, in vivo PET imaging to track the biodistribution of AE17 OMVs showed that AE17 OMVs were taken up by the lung region and the gastrointestinal and renal regions but were not detected in other areas. Pathological analysis of the tissue sites where AE17 OMVs were ingested showed inflammatory responses and damage. These findings suggested that AE17 OMVs not only contained a group of virulence proteins associated with AE17 infection but can also deliver these virulence proteins over long distances and caused tissue inflammatory damage. Our study revealed a previously unidentified causative microbial signal in the pathogenesis of APEC that could aid in the development of vaccines and antibiotics effective against APEC.

Dataset Information

Immuno-PET imaging of VEGFR-2 expression in prostate cancer with ⁸⁹Zr-labeled ramucirumab.

Publications

Immuno-PET imaging of VEGFR-2 expression in prostate cancer with <sup>89</sup>Zr-labeled ramucirumab.

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Dataset Information

Immuno-PET imaging of VEGFR-2 expression in prostate cancer with 89Zr-labeled ramucirumab.

Publications

Immuno-PET imaging of VEGFR-2 expression in prostate cancer with <sup>89</sup>Zr-labeled ramucirumab.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

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Immuno-PET imaging of VEGFR-2 expression in prostate cancer with ⁸⁹Zr-labeled ramucirumab.