Active-site MMP-selective antibody inhibitors discovered from convex paratope synthetic libraries
Dong Hyun Nam, Carlos Rodriguez, Albert G. Remacle, Alex Y. Strongin, Xin Ge
The matrix metalloproteinase (MMP) family members are promising drug targets in diversified pathologies. Clinical trial failures taught us that selective, rather than broad-specificity, inhibitors are required for successful MMP therapies. Achieving target selectivity with small-molecule MMP inhibitors, however, is exceedingly difficult. Because the antigen-binding sites in conventional antibodies are predominantly incompatible with the concave reaction pockets of MMPs, design of inhibitory antibodies, an attractive alternative for selective inhibition, is also challenging. We synthesized human antibody libraries encoding extended convex antigen-binding sites and isolated a panel of inhibitory Fabs that selectively and efficiently inhibited MMP-14, a promising drug target in cancer. The pipeline we established can now be readily applied for the generation of inhibitory antibodies targeting multiple additional enzymes besides MMPs alone.
Functional selection of protease inhibitory antibodies
Tyler Lopez, Zahid Mustafa, Chuan Chen, Ki Baek Lee, Aaron Ramirez, Chris Benitez, Xin Luo, Ru-Rong Ji, Xin Ge
Proteases precisely control a wide variety of physiological processes and thus are important drug targets. Compared with small-molecule inhibitors, monoclonal antibodies (mAbs) are attractive, as they provide required specificity. However, finding inhibitory mAbs is often a bottleneck, largely due to lack of a function-based selection method. We overcame this obstacle and successfully isolated mAbs that effectively inhibited 5 therapeutic targets spanning 4 basic classes of proteases. Our mAb inhibitors are highly selective and deliver desired biochemical and biological actions, including reduction of amyloid beta formation in vitro and pain relief in animal behavioral tests. Thus, the technique described here can be readily applied to many pathophysiologically important proteases for the discovery and engineering of therapeutic mAbs.
Ge X, Lopez T, Ji RR. MMP-9 Antibodies and Methods of Use Thereof. US62/851001. Licensed to Releviate Therapeutics
Ge X, Lee KB. Protease Inhibitory Antibodies and Method of Use Thereof. US11332546. Licensed to Maverick Therapeutics.
Ge X, Nam DH, Lopez T. Inhibitory Antibodies and Methods of Use Thereof. US10975166; WO2018067198A1. Licensed to Releviate Therapeutics.
Reddy ST, Ge X, Boutz D, Ellington AD, Marcotte EM, Georgiou G. Rapid Isolation of Monoclonal Antibodies from Animals. US9090674B2; EP2572203B1; KR20130135028A; CN103003697A; CA2799746A1; WO2011146514A3. Licensed to Cell Signaling Technology Inc.
Ge X, Mazor Y, Georgiou G. Methods for Creating Antibody Libraries. US12868399; EP2470653A1; CA2772298A1; WO2011025826A1.
Chilkoti A, Carlson KT, Christensen T, Filipe CDM, Ge X. Purification of Low-Abundant Recombinant Proteins from Cell Culture. WO2008024311A3. Licensed to PhaseBio Pharmaceuticals Inc.
Nam DH, Lee KB, Kruchowy E, Pham H, Ge X. Protease Inhibition Mechanism of Camelid-Like Synthetic Human Antibodies. Biochemistry (2020) 59:3802-3812. PMID: 32997500
Lee KB, Dunn Z, Lopez T, Mustafa Z, Ge X. Generation of Highly Selective Monoclonal Antibodies Inhibiting a Recalcitrant Protease Using Decoy Designs. Biotechnology and Bioengineering (2020) 117:3664-3676. PMID: 32716053
Li X, Zhao Y, Chen C, Lee H-H, Wang Z, Zhang N, Kolonin MG, An Z, Ge X, Scherer PE, Sun K. Critical Role of Matrix Metalloproteinase 14 in Adipose Tissue Remodeling during Obesity.
Molecular and Cellular Biology (2020) 40:e00564-19. PMID: 31988105
Lee KB, Dunn Z, Ge X. Reducing Proteolytic Liability of a MMP‐14 Inhibitory Antibody by Site‐Saturation Mutagenesis. Protein Sciences (2019) 28:643-653. PMID: 30592555
Lopez T, Ramirez A, Benitez C, Mustafa Z, Pham H, Sanchez R, Ge X. Selectivity Conversion of Protease Inhibitory Antibodies. Antibody Therapeutics (2018) 1:55-63. PMID: 30406213
Lopez T, Chen C, Ramirez A, Chen KE, Lorenson MY, Benitez C, Mustafa Z, Pham H, Sanchez R, Walker AM, Ge X. Epitope Specific Affinity Maturation Improved Stability of Potent Protease Inhibitory Antibodies. Biotechnology and Bioengineering (2018) 115:2673-2682. PMID: 30102763
Chen KE, Chen C, Radecki KC, Bustamante K, Lopez T, Lorenson MY, Ge X, Walker AM. Use of a Novel Camelid-inspired Human Antibody Demonstrates the Importance of MMP-14 to Cancer Stem Cell Function in the Metastatic Process. Oncotarget (2018) 9:29431-29444. PMID: 30034628
Chen L, Holmes M, Schaefer E, Mulchandani A, Ge X. Highly Active Spore Biocatalyst by Self-Assembly of Co-Expressed Anchoring Scaffoldin and Multimeric Enzyme. Biotechnology and Bioengineering (2018) 115:557-564. PMID: 29131302
Rodriguez C, Nam DH, Kruchowy E, Ge X. Efficient Antibody Assembly in E. coli Periplasm by Chaperone Co-expression and Culture Optimization. Applied Biochemistry and Biotechnology (2017) 183:520-529. PMID: 28488120
Lee KB, Nam DH, Nuhn JA, Wang J, Schneider IC, Ge X. Direct Expression of Active Human Tissue Inhibitors of Metalloproteinases by Periplasmic Secretion in Escherichia coli. Microbial Cell Factories (2017) 16:73. PMID: 28454584
Lopez T, Nam DH, Kaihara E, Mustafa Z, Ge X. Identification of Highly Selective MMP-14 Inhibitory Fabs by Deep Sequencing. Biotechnology and Bioengineering (2017) 114:1140-1150. PMID: 28090632
Nam DH, Fang K, Rodriguez C, Lopez T, Ge X. Generation of Inhibitory Monoclonal Antibodies Targeting Membrane-Type 1 Matrix Metalloproteinase by Motif Grafting and CDR Optimization. Protein Engineering Design and Selection (2017) 30:113-118. PMID: 27986919
Remacle AG, Cieplak P, Nam DH, Shiryaev SA, Ge X, Strongin AY. Selective Function-Blocking Monoclonal Human Antibody Highlights the Important Role of Membrane Type-1 Matrix Metalloproteinase (MT1-MMP) in Metastasis. Oncotarget (2017) 8:2781-2799. PMID: 27835863
Li XC, Wang C, Mulchandani A, Ge X. Engineering Soluble Human Paraoxonase 2 for Quorum Quenching. ACS Chemistry Biology (2016) 11:3122-3131. PMID: 27623343
Nam DH, Ge X. Direct Production of Functional Matrix Metalloproteinase-14 Without Refolding or Activation and its Application for In Vitro Inhibition Assays. Biotechnology and Bioengineering (2016) 113:717-723. PMID: 26416249
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