期刊
THERANOSTICS
卷 9, 期 15, 页码 4409-4420出版社
IVYSPRING INT PUBL
DOI: 10.7150/thno.32883
关键词
Positron emission tomography (PET); immuno-PET; site-specific labeling; molecular imaging; HER2
资金
- Novo Nordisk Foundation
- Lundbeck Foundation
- Innovation Fund Denmark
- Svend Andersen Foundation
- Arvid Nilsson Foundation
- John and Birthe Meyer Foundation
- Neye Foundation
- Research Foundation of Rigshospitalet
- Research Foundation of the Capital Region
- Global Excellence Program
- Danish National Research Foundation [126]
- H2020 program
- European Research Council (ERC) [670261]
- European Research Council (ERC) [670261] Funding Source: European Research Council (ERC)
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 beta-galactosidase or endoglycosidase S2 prior to Zr-89 radiolabeling. Zr-89-DFO-trastuzumab was injected into SK-OV-3 tumor-bearing NMRI nude mice. The antibody dose was titrated with either 100 mu g or 500 mu g 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 Zr-89-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, beta-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 Zr-89-DFO-trastuzumab successfully detected HER2-positive tumors in PDX mouse models. Conclusion: Zr-89-DFO-trastuzumab is well-matched for specific immuno-PET imaging of HER2-positive tumors and site-specific labeling of trastuzumab by the SiteClick (TM) 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.
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