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- PDB-9e6k: Fully human monoclonal antibody targeting the cysteine-rich subst... -

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Basic information

Entry
Database: PDB / ID: 9e6k
TitleFully human monoclonal antibody targeting the cysteine-rich substrate-interacting region of ADAM17 on cancer cells.
Components
  • Disintegrin and metalloproteinase domain-containing protein 17
  • heavy chain of monoclonal antibody C12
  • light chain monoclonal antibody C12
KeywordsANTITUMOR PROTEIN/IMMUNE SYSTEM / inhibitor / complex / ONCOPROTEIN / ANTITUMOR PROTEIN / ANTITUMOR PROTEIN-IMMUNE SYSTEM complex
Function / homologyDomain of unknown function DUF3850 / Domain of unknown function (DUF3850) / ASCH / ASCH domain / PUA-like superfamily / DUF3850 domain-containing protein
Function and homology information
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.15 Å
AuthorsSaha, N. / De La Cruz, M.J. / Goldgur, Y. / Nikolov, D.B.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Cancer Institute (NIH/NCI) United States
CitationJournal: Biomed Pharmacother / Year: 2024
Title: Fully human monoclonal antibody targeting the cysteine-rich substrate-interacting region of ADAM17 on cancer cells.
Authors: Nayanendu Saha / Sang Gyu Lee / Eeva-Christine Brockmann / M Jason de la Cruz / Yehuda Goldgur / Rachelle P Mendoza / Elisa de Stanchina / Tanzy M Love / Josh Marvald / Yan Xu / Kai Xu / ...Authors: Nayanendu Saha / Sang Gyu Lee / Eeva-Christine Brockmann / M Jason de la Cruz / Yehuda Goldgur / Rachelle P Mendoza / Elisa de Stanchina / Tanzy M Love / Josh Marvald / Yan Xu / Kai Xu / Juha P Himanen / Urpo Lamminmäki / Darren Veach / Dimitar B Nikolov /
Abstract: ADAM17 sheds EGFR/erbB ligands and triggers oncogenic pathways that lead to the progression of solid tumors. We targeted the ADAM17 disintegrin and cysteine rich domain region (D+C) to generate a ...ADAM17 sheds EGFR/erbB ligands and triggers oncogenic pathways that lead to the progression of solid tumors. We targeted the ADAM17 disintegrin and cysteine rich domain region (D+C) to generate a panel of single-chain antibody fragments (scFvs) that selectively bind to the D or C domains of ADAM17, but not of ADAM10 or ADAM19. From the panel, we selected one scFv, referred to as C12, based on its high binding affinity towards the target, and re-formatted it to a full IgG for further studies. High-resolution cryo-electron microscopy studies documented that the mAb binds to the ADAM17 C-domain that in ADAM proteases, notably ADAM10 and ADAM17, is known to impart substrate-specificity. The C12 mAb significantly inhibited EGFR phosphorylation in cancer cell lines by hindering the cleavage of EGFR ligands tethered to the cell surface. This inhibition provides a mechanism for potential anti-tumor effects, and indeed C12 diminished the viability of a variety of EGFR-expressing cancer cell lines. Cell-based ELISA studies revealed that C12 preferentially bound to activated ADAM17 present on tumor cells, as compared to the autoinhibited ADAM17 that is the predominant form on HEK293 and other non-tumor cells. C12 also exhibited tumor growth inhibition in an ovarian cancer xenograft mouse model. Consistent with its selective tumor cell binding in vitro, radioimmuno PET (positron emission tomography) imaging with Zr-DFO-C12 in mouse xenograft models confirmed tumoral accumulation of the C12 mAb.
History
DepositionOct 30, 2024Deposition site: RCSB / Processing site: RCSB
Revision 1.0Nov 20, 2024Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
H: heavy chain of monoclonal antibody C12
L: light chain monoclonal antibody C12
C: Disintegrin and metalloproteinase domain-containing protein 17


Theoretical massNumber of molelcules
Total (without water)59,0493
Polymers59,0493
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

#1: Antibody heavy chain of monoclonal antibody C12


Mass: 23029.830 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Production host: Spodoptera (butterflies/moths)
#2: Antibody light chain monoclonal antibody C12


Mass: 23439.961 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Production host: Spodoptera (butterflies/moths)
#3: Protein Disintegrin and metalloproteinase domain-containing protein 17 / ADAM 17 / Snake venom-like protease / TNF-alpha convertase / TNF-alpha-converting enzyme


Mass: 12579.383 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: ADAM17, CSVP, TACE / Production host: Spodoptera (butterflies/moths) / References: UniProt: P78536, EC: 3.4.24.86
Has protein modificationY

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Complex of ADAM17 disintegrin-cysteine rich domain / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Spodoptera (butterflies/moths)
Buffer solutionpH: 7.4
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: NITROGEN

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: OTHER
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 1000 nm / Nominal defocus min: 500 nm
Image recordingElectron dose: 65.9 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

CTF correctionType: NONE
3D reconstructionResolution: 3.15 Å / Resolution method: OTHER / Num. of particles: 898088 / Symmetry type: POINT
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 82.95 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0034216
ELECTRON MICROSCOPYf_angle_d0.65255720
ELECTRON MICROSCOPYf_chiral_restr0.0422637
ELECTRON MICROSCOPYf_plane_restr0.0055743
ELECTRON MICROSCOPYf_dihedral_angle_d4.6577586

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