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Structural and functional characterization of C0021158, a high-affinity monoclonal antibody that inhibits Arginase 2 function via a novel non-competitive mechanism of action

DOI: 10.1080/19420862.2020.1801230 DOI Help

Authors: Mark Austin (Cancer Research UK AstraZeneca Antibody Alliance Laboratory; AstraZeneca) , Daniel Burschowsky (University of Leicester) , Denice T. Y. Chan (Cancer Research UK AstraZeneca Antibody Alliance Laboratory) , Lesley Jenkinson (Cancer Research UK AstraZeneca Antibody Alliance Laboratory) , Stuart Haynes (Cancer Research UK AstraZeneca Antibody Alliance Laboratory) , Agata Diamandakis (Cancer Research UK AstraZeneca Antibody Alliance Laboratory) , Chitra Seewooruthun (University of Leicester) , Alexandra Addyman (Cancer Research UK AstraZeneca Antibody Alliance Laboratory) , Sebastian Fiedler (Cancer Research UK AstraZeneca Antibody Alliance Laboratory) , Stephanie Ryman (Cancer Research UK AstraZeneca Antibody Alliance Laboratory) , Jessica Whitehouse (Cancer Research UK AstraZeneca Antibody Alliance Laboratory) , Louise H. Slater (Cancer Research UK AstraZeneca Antibody Alliance Laboratory) , Andreas V. Hadjinicolaou (Weatherall Institute of Molecular Medicine, University of Oxford) , Uzi Gileadi (Weatherall Institute of Molecular Medicine, University of Oxford) , Ellen Gowans (Cancer Research UK AstraZeneca Antibody Alliance Laboratory) , Yoko Shibata (Cancer Research UK AstraZeneca Antibody Alliance Laboratory) , Michelle Barnard (Cancer Research UK AstraZeneca Antibody Alliance Laboratory) , Teresa Kaserer (Cancer Research UK) , Pooja Sharma (Cancer Research UK AstraZeneca Antibody Alliance Laboratory) , Nadia M. Luheshi (AstraZeneca) , Robert W. Wilkinson (AstraZeneca) , Tristan J. Vaughan (AstraZeneca) , Sarah V. Holt (Cancer Research UK AstraZeneca Antibody Alliance Laboratory) , Vincenzo Cerundolo (Weatherall Institute of Molecular Medicine, University of Oxford) , Mark D. Carr (University of Leicester) , Maria A. T. Groves (Cancer Research UK AstraZeneca Antibody Alliance Laboratory; AstraZeneca)
Co-authored by industrial partner: Yes

Type: Journal Paper
Journal: Mabs , VOL 12

State: Published (Approved)
Published: January 2020
Diamond Proposal Number(s): 19880

Open Access Open Access

Abstract: Arginase 2 (ARG2) is a binuclear manganese metalloenzyme that catalyzes the hydrolysis of L-arginine. The dysregulated expression of ARG2 within specific tumor microenvironments generates an immunosuppressive niche that effectively renders the tumor ‘invisible’ to the host’s immune system. Increased ARG2 expression leads to a concomitant depletion of local L-arginine levels, which in turn leads to suppression of anti-tumor T-cell-mediated immune responses. Here we describe the isolation and characterization of a high affinity antibody (C0021158) that inhibits ARG2 enzymatic function completely, effectively restoring T-cell proliferation in vitro. Enzyme kinetic studies confirmed that C0021158 exhibits a noncompetitive mechanism of action, inhibiting ARG2 independently of L-arginine concentrations. To elucidate C0021158’s inhibitory mechanism at a structural level, the co-crystal structure of the Fab in complex with trimeric ARG2 was solved. C0021158’s epitope was consequently mapped to an area some distance from the enzyme’s substrate binding cleft, indicating an allosteric mechanism was being employed. Following C0021158 binding, distinct regions of ARG2 undergo major conformational changes. Notably, the backbone structure of a surface-exposed loop is completely rearranged, leading to the formation of a new short helix structure at the Fab-ARG2 interface. Moreover, this large-scale structural remodeling at ARG2’s epitope translates into more subtle changes within the enzyme’s active site. An arginine residue at position 39 is reoriented inwards, sterically impeding the binding of L-arginine. Arg39 is also predicted to alter the pK A of a key catalytic histidine residue at position 160, further attenuating ARG2’s enzymatic function. In silico molecular docking simulations predict that L-arginine is unable to bind effectively when antibody is bound, a prediction supported by isothermal calorimetry experiments using an L-arginine mimetic. Specifically, targeting ARG2 in the tumor microenvironment through the application of C0021158, potentially in combination with standard chemotherapy regimens or alternate immunotherapies, represents a potential new strategy to target immune cold tumors.

Journal Keywords: ARG2; Arginase 2; antibody co-crystal structure; phage display selections; cancer therapeutics; ARG2 epitope; human monoclonal antibody; ARG2 neutralization

Diamond Keywords: Enzymes

Subject Areas: Biology and Bio-materials, Chemistry, Medicine


Instruments: I04-1-Macromolecular Crystallography (fixed wavelength) , I04-Macromolecular Crystallography

Other Facilities: ID23-1 at ESRF; HZB at BESSY II

Added On: 09/09/2020 13:52

Documents:
Structural and functional characterization of C0021158 a high affinity monoclonal antibody that inhibits Arginase 2 function via a novel non.pdf

Discipline Tags:

Catalysis Life Sciences & Biotech Health & Wellbeing Cancer Drug Discovery Non-Communicable Diseases Structural biology Chemistry Biochemistry

Technical Tags:

Diffraction Macromolecular Crystallography (MX)