Dataset Information

⁸⁹Zr-Labeled Domain II-Specific scFv-Fc ImmunoPET Probe for Imaging Epidermal Growth Factor Receptor In Vivo.

ABSTRACT: Epidermal growth factor receptor I (EGFR) is overexpressed in many cancers. The extracellular domain of EGFR has four binding epitopes (domains I- IV). All clinically approved anti-EGFR antibodies bind to domain III. Imaging agents that bind to domains other than domain III of EGFR are needed for accurate quantification of EGFR, patient selection for anti-EGFR therapeutics and monitoring of response to therapies. We recently developed a domain II-specific antibody fragment 8709. In this study, we have evaluated the in vitro and in vivo properties of ⁸⁹Zr-8709-scFv-Fc (105 kDa). We conjugated 8709-scFv-Fc with the deferoxamine (DFO) chelator and radiolabeled the DFO-8970-scFv with ⁸⁹Zr. We evaluated the binding of ⁸⁹Zr-DFO-8709-scFv-Fc in EGFR positive and negative cell lines DLD-1, MDA-MB-231 and MDA-MB-435, respectively, and in mouse xenograft models. Simultaneously, we have compared the binding of ⁸⁹Zr-8709-scFv-Fc with ¹¹¹In-nimotuzumab, a domain III anti-EGFR antibody. DFO-8709-scFv-Fc displayed similar cell binding specificity as 8709-scFv-Fc. Saturation cell binding assay and immunoreactive fraction showed that radiolabeling did not alter the binding of 8709-scFv-Fc. Biodistribution and microPET showed good uptake of ⁸⁹Zr-8709-scFv-Fc in xenografts after 120 h post injection (p.i). and was domain-specific to EGFR domain II. ⁸⁹Zr-8709-scFv-Fc did not compete for binding in vitro and in vivo with a known domain III binder nimotuzumab. The results show that ⁸⁹Zr-8709-scFv-Fc is specific to domain II of EGFR making it favorable for quantification of EGFR in vivo, hence, patient selection and monitoring of response to treatment with anti-EGFR antibodies.

SUBMITTER: Alizadeh E

PROVIDER: S-EPMC7867132 | biostudies-literature | 2021 Feb

REPOSITORIES: biostudies-literature

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<sup>89</sup>Zr-Labeled Domain II-Specific scFv-Fc ImmunoPET Probe for Imaging Epidermal Growth Factor Receptor In Vivo.

Alizadeh Elahe E Behlol Ayaz Ahmed Khan K Raja Solomon Viswas V Gaja Vijay V Bernhard Wendy W Makhlouf Amal A Gonzalez Carolina C Barreto Kris K Casaco Angel A Geyer C Ronald CR Fonge Humphrey H

Cancers 20210201 3

Epidermal growth factor receptor I (EGFR) is overexpressed in many cancers. The extracellular domain of EGFR has four binding epitopes (domains I- IV). All clinically approved anti-EGFR antibodies bind to domain III. Imaging agents that bind to domains other than domain III of EGFR are needed for accurate quantification of EGFR, patient selection for anti-EGFR therapeutics and monitoring of response to therapies. We recently developed a domain II-specific antibody fragment 8709. In this study, w ...[more]

PMID: 33535661

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Project description:Prostate stem cell antigen (PSCA) is expressed on the cell surface in 83%-100% of local prostate cancers and 87%-100% of prostate cancer bone metastases. In this study, we sought to develop immunoPET agents using (124)I- and (89)Zr-labeled anti-PSCA A11 minibodies (scFv-CH3 dimer, 80 kDa) and evaluate their use for quantitative immunoPET imaging of prostate cancer.A11 anti-PSCA minibody was alternatively labeled with (124)I- or (89)Zr-desferrioxamine and injected into mice bearing either matched 22Rv1 and 22Rv1×PSCA or LAPC-9 xenografts. Small-animal PET data were obtained and quantitated with and without recovery coefficient-based partial-volume correction, and the results were compared with ex vivo biodistribution.Rapid and specific localization to PSCA-positive tumors and high-contrast imaging were observed with both (124)I- and (89)Zr-labeled A11 anti-PSCA minibody. However, the differences in tumor uptake and background uptake of the radiotracers resulted in different levels of imaging contrast. The nonresidualizing (124)I-labeled minibody had lower tumor uptake (3.62 ± 1.18 percentage injected dose per gram [%ID/g] 22Rv1×PSCA, 3.63 ± 0.59 %ID/g LAPC-9) than the residualizing (89)Zr-labeled minibody (7.87 ± 0.52 %ID/g 22Rv1×PSCA, 9.33 ± 0.87 %ID/g LAPC-9, P < 0.0001 for each), but the (124)I-labeled minibody achieved higher imaging contrast because of lower nonspecific uptake and better tumor-to-soft-tissue ratios (22Rv1×PSCA:22Rv1 positive-to-negative tumor, 13.31 ± 5.59 (124)I-A11 and 4.87 ± 0.52 (89)Zr-A11, P = 0.02). Partial-volume correction was found to greatly improve the correspondence between small-animal PET and ex vivo quantification of tumor uptake for immunoPET imaging with both radionuclides.Both (124)I- and (89)Zr-labeled A11 anti-PSCA minibody showed high-contrast imaging of PSCA expression in vivo. However, the (124)I-labeled A11 minibody was found to be the superior imaging agent because of lower nonspecific uptake and higher tumor-to-soft-tissue contrast. Partial-volume correction was found to be essential for robust quantification of immunoPET imaging with both (124)I- and (89)Zr-labeled A11 minibody.

Project description:(89)Zr (half-life, 78.41 h) is a positron-emitting radionuclide that displays excellent potential for use in the design and synthesis of radioimmunoconjugates for immunoPET. In the current study, we report the preparation of (89)Zr-desferrioxamine B (DFO)-J591, a novel (89)Zr-labeled monoclonal antibody (mAb) construct for targeted immunoPET and quantification of prostate-specific membrane antigen (PSMA) expression in vivo.The in vivo behavior of (89)Zr-chloride, (89)Zr-oxalate, and (89)Zr-DFO was studied using PET. High-level computational studies using density functional theory calculations have been used to investigate the electronic structure of (89)Zr-DFO and probe the nature of the complex in aqueous conditions. (89)Zr-DFO-J591 was characterized both in vitro and in vivo. ImmunoPET in male athymic nu/nu mice bearing subcutaneous LNCaP (PSMA-positive) or PC-3 (PSMA-negative) tumors was conducted. The change in (89)Zr-DFO-J591 tissue uptake in response to high- and low-specific-activity formulations in the 2 tumor models was measured using acute biodistribution studies and immunoPET.The basic characterization of 3 important reagents-(89)Zr-chloride, (89)Zr-oxalate, and the complex (89)Zr-DFO-demonstrated that the nature of the (89)Zr species dramatically affects the biodistribution and pharmacokinetics. Density functional theory calculations provide a rationale for the observed high in vivo stability of (89)Zr-DFO-labeled mAbs and suggest that in aqueous conditions, (89)Zr-DFO forms a thermodynamically stable, 8-coordinate complex by coordination of 2 water molecules. (89)Zr-DFO-J591 was produced in high radiochemical yield (>77%) and purity (>99%), with a specific activity of 181.7 +/- 1.1 MBq/mg (4.91 +/- 0.03 mCi/mg). In vitro assays demonstrated that (89)Zr-DFO-J591 had an initial immunoreactive fraction of 0.95 +/- 0.03 and remained active for up to 7 d. In vivo biodistribution experiments revealed high, target-specific uptake of (89)Zr-DFO-J591 in LNCaP tumors after 24, 48, 96, and 144 h (34.4 +/- 3.2 percentage injected dose per gram [%ID/g], 38.0 +/- 6.2 %ID/g, 40.4 +/- 4.8 %ID/g, and 45.8 +/- 3.2 %ID/g, respectively). ImmunoPET studies also showed that (89)Zr-DFO-J591 provides excellent image contrast, with tumor-to-muscle ratios greater than 20, for the delineation of LNCaP xenografts between 48 and 144 h after administration.These studies demonstrate that (89)Zr-DFO-labeled mAbs show exceptional promise as radiotracers for immunoPET of human cancers. (89)Zr-DFO-J591 displays high tumor-to-background tissue contrast in immunoPET and can be used to delineate and quantify PSMA-positive prostate tumors in vivo.

Project description:INTRODUCTION:Chimeric antigen receptor (CAR) T-cells have been recently developed and are producing impressive outcomes in patients with hematologic malignancies. However, there is no standardized method for cell trafficking and in vivo CAR T-cell monitoring. We assessed the feasibility of real-time in vivo 89Zr-p-Isothiocyanatobenzyl-desferrioxamine (Df-Bz-NCS, DFO) labeled CAR T-cell trafficking using positron emission tomography (PET). RESULTS:The 89Zr-DFO radiolabeling efficiency of Jurkat/CAR and human peripheral blood mononuclear cells (hPBMC)/CAR T-cells was 70%-79%, and cell radiolabeling activity was 98.1-103.6 kBq/106 cells. Cell viability after radiolabeling was >95%. Cell proliferation was not significantly different during the early period after radiolabeling, compared with unlabeled cells; however, the proliferative capacity decreased over time (day 7 after labeling). IL-2 or IFN-? secretion was not significantly different between unlabeled and labeled CAR T-cells. PET/magnetic resonance imaging in the xenograft model showed that most of the 89Zr-DFO-labeled Jurkat/CAR T-cells were distributed in the lung (24.4% ± 3.4%ID) and liver (22.9% ± 5.6%ID) by one hour after injection. The cells gradually migrated from the lung to the liver and spleen by day 1, and remained stable in these sites until day 7 (on day 7: lung 3.9% ± 0.3%ID, liver 36.4% ± 2.7%ID, spleen 1.4% ± 0.3%ID). No significant accumulation of labeled cells was identified in tumors. A similar pattern was observed in ex vivo biodistributions on day 7 (lung 3.0% ± 1.0%ID, liver 19.8% ± 2.2%ID, spleen 2.3% ± 1.7%ID). 89Zr-DFO-labeled hPBMC/CAR T-cells showed a similar distribution, compared with Jurkat/CAR T-cells, on serial PET images. CAR T cell distribution was cross-confirmed by flow cytometry, Alu polymerase chain reaction, and immunohistochemistry. CONCLUSION:Real-time in vivo cell trafficking is feasible using PET imaging of 89Zr-DFO-labeled CAR T-cells. This can be used to investigate cellular kinetics, initial in vivo biodistribution, and safety profiles in future CAR T-cell development.

Dataset Information

⁸⁹Zr-Labeled Domain II-Specific scFv-Fc ImmunoPET Probe for Imaging Epidermal Growth Factor Receptor In Vivo.

Publications

<sup>89</sup>Zr-Labeled Domain II-Specific scFv-Fc ImmunoPET Probe for Imaging Epidermal Growth Factor Receptor In Vivo.

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Dataset Information

89Zr-Labeled Domain II-Specific scFv-Fc ImmunoPET Probe for Imaging Epidermal Growth Factor Receptor In Vivo.

Publications

<sup>89</sup>Zr-Labeled Domain II-Specific scFv-Fc ImmunoPET Probe for Imaging Epidermal Growth Factor Receptor In Vivo.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets

⁸⁹Zr-Labeled Domain II-Specific scFv-Fc ImmunoPET Probe for Imaging Epidermal Growth Factor Receptor In Vivo.