American Association for Cancer Research
15357163mct140968t-sup-140633_0_supp_2632577_n2yyyl.doc (3.02 MB)

Supplementary Figures S1-S12 and Supplementary Tables S1-S3 from Novel Glycosylated VEGF Decoy Receptor Fusion Protein, VEGF-Grab, Efficiently Suppresses Tumor Angiogenesis and Progression

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journal contribution
posted on 2023-04-03, 14:48 authored by Jung-Eun Lee, Chan Kim, Hannah Yang, Intae Park, Nuri Oh, Serenus Hua, Haneul Jeong, Hyun Joo An, Sun Chang Kim, Gyun Min Lee, Gou Young Koh, Ho Min Kim

Supplementary Figure S1. Sequence Alignment of VEGF-Grab1, VEGF-Grab3, and VEGF-Trap; Supplementary Figure S2. Analysis of N-linked Glycosylation for VEGF-Grabs and VEGF-Trap; Supplementary Figure S3. Analysis of O-linked Glycosylation in VEGF-Grab1; Supplementary Figure S4. VEGF-Grab3 displays low binding to tumor ECM; Supplementary Figure S5. Pharmacokinetic profile of VEGF-Grab1 and VEGF-Grab3; Supplementary Figure S6. Tissue accumulation of VEGF-Trap and VEGF-Grab3; Supplementary Figure S7. VEGF-Grabs or VEGF-Trap have no effects on VEGFR2 signalling, EC survival, migration, and tube formation in the absence of VEGFA; Supplementary Figure S8. Dose-dependent Inhibition of VEGFR2 Signalling with Anti-VEGF Therapy; Supplementary Figure S9. Histologic Analyses of Vital Organs after Anti-VEGF Therapy; Supplementary Figure S10. VEGF-Grab effectively suppresses the growth of established bulky macroscopic tumors; Supplementary Figure S11. Histological analyses of the kidney and liver after dose dependent anti-VEGF therapy; Supplementary Figure S12. Combination Therapy of VEGF-Grab3 and Cisplatin Enhances Intratumoral Apoptosis; Supplementary Table S1. Primer sequences for site-directed mutagenesis; Supplementary Table S2. Primer sequences for Quantitative Real Time PCR; Supplementary Table S3. Antibodies.



Antiangiogenic therapies targeting VEGFA have been commonly used in clinics to treat cancers over the past decade. However, their clinical efficacy has been limited, with drawbacks including acquisition of resistance and activation of compensatory pathways resulting from elevated circulating VEGFB and placental growth factor (PlGF). To bypass these disadvantages, we developed a novel glycosylated soluble decoy receptor fusion protein, VEGF-Grab, that can neutralize VEGFA, VEGFB, and PlGF. VEGF-Grab has the second and third immunoglobulin (Ig)-like domains of VEGF receptor 1 (VEGFR1) fused to IgG1 Fc, with three potential glycosylation sites introduced into the third Ig-like domain of VEGF-Grab by mutagenesis. Compared with VEGF-Trap, VEGF-Grab showed more potent decoy activity against VEGF and PlGF, mainly attributed to the VEGFR1 backbone. Most importantly, the negatively charged O-glycans attached to the third Ig-like domain of VEGFR1 counterbalanced the originally positively charged VEGFR1 backbone, minimizing nonspecific binding of VEGF-Grab to the extracellular matrix, and resulting in greatly improved pharmacokinetic profile. These advancements led to stronger and more durable antiangiogenic, antitumor, and antimetastatic efficacy in both implanted and spontaneous tumor models as compared with VEGF-Trap, while toxicity profiles were comparable with VEGF-Trap. Collectively, our results highlight VEGF-Grab as a promising therapeutic candidate for further clinical drug development. Mol Cancer Ther; 14(2); 470–9. ©2014 AACR.