title: Therapeutic antibody targeting of individual Notch receptors.
abstract: The four receptors of the Notch family are widely expressed transmembrane proteins that function as key conduits through which mammalian cells communicate to regulate cell fate and growth. Ligand binding triggers a conformational change in the receptor negative regulatory region (NRR) that enables ADAM protease cleavage at a juxtamembrane site that otherwise lies buried within the quiescent NRR. Subsequent intramembrane proteolysis catalysed by the gamma-secretase complex liberates the intracellular domain (ICD) to initiate the downstream Notch transcriptional program. Aberrant signalling through each receptor has been linked to numerous diseases, particularly cancer, making the Notch pathway a compelling target for new drugs. Although gamma-secretase inhibitors (GSIs) have progressed into the clinic, GSIs fail to distinguish individual Notch receptors, inhibit other signalling pathways and cause intestinal toxicity, attributed to dual inhibition of Notch1 and 2 (ref. 11). To elucidate the discrete functions of Notch1 and Notch2 and develop clinically relevant inhibitors that reduce intestinal toxicity, we used phage display technology to generate highly specialized antibodies that specifically antagonize each receptor paralogue and yet cross-react with the human and mouse sequences, enabling the discrimination of Notch1 versus Notch2 function in human patients and rodent models. Our co-crystal structure shows that the inhibitory mechanism relies on stabilizing NRR quiescence. Selective blocking of Notch1 inhibits tumour growth in pre-clinical models through two mechanisms: inhibition of cancer cell growth and deregulation of angiogenesis. Whereas inhibition of Notch1 plus Notch2 causes severe intestinal toxicity, inhibition of either receptor alone reduces or avoids this effect, demonstrating a clear advantage over pan-Notch inhibitors. Our studies emphasize the value of paralogue-specific antagonists in dissecting the contributions of distinct Notch receptors to differentiation and disease and reveal the therapeutic promise in targeting Notch1 and Notch2 independently.
authors.affiliation: Department of Antibody Engineering, Genentech, Inc., 1 DNA Way, South San Francisco, California 94080, USA.
authors.forename: Yan
authors.initials: Y
authors.lastname: Wu
authors.forename: Carol
authors.initials: C
authors.lastname: Cain-Hom
authors.forename: Lisa
authors.initials: L
authors.lastname: Choy
authors.forename: Thijs J
authors.initials: TJ
authors.lastname: Hagenbeek
authors.forename: Gladys P
authors.initials: GP
authors.lastname: de Leon
authors.forename: Yongmei
authors.initials: Y
authors.lastname: Chen
authors.forename: David
authors.initials: D
authors.lastname: Finkle
authors.forename: Rayna
authors.initials: R
authors.lastname: Venook
authors.forename: Xiumin
authors.initials: X
authors.lastname: Wu
authors.forename: John
authors.initials: J
authors.lastname: Ridgway
authors.forename: Dorreyah
authors.initials: D
authors.lastname: Schahin-Reed
authors.forename: Graham J
authors.initials: GJ
authors.lastname: Dow
authors.forename: Amy
authors.initials: A
authors.lastname: Shelton
authors.forename: Scott
authors.initials: S
authors.lastname: Stawicki
authors.forename: Ryan J
authors.initials: RJ
authors.lastname: Watts
authors.forename: Jeff
authors.initials: J
authors.lastname: Zhang
authors.forename: Robert
authors.initials: R
authors.lastname: Choy
authors.forename: Peter
authors.initials: P
authors.lastname: Howard
authors.forename: Lisa
authors.initials: L
authors.lastname: Kadyk
authors.forename: Minhong
authors.initials: M
authors.lastname: Yan
authors.forename: Jiping
authors.initials: J
authors.lastname: Zha
authors.forename: Christopher A
authors.initials: CA
authors.lastname: Callahan
authors.forename: Sarah G
authors.initials: SG
authors.lastname: Hymowitz
authors.forename: Christian W
authors.initials: CW
authors.lastname: Siebel
mesh_terms: D020533:Angiogenesis Inhibitors / Q000276:immunology / Q000494:pharmacology / Q000627:therapeutic use; D000818:Animals; D000906:Antibodies / Q000009:adverse effects / Q000276:immunology / Q000494:pharmacology* / Q000627:therapeutic use*; D000918:Antibody Specificity / Q000276:immunology; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D020397:Goblet Cells / Q000187:drug effects / Q000473:pathology; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D041681:NIH 3T3 Cells; D009369:Neoplasms / Q000098:blood supply / Q000188:drug therapy* / Q000378:metabolism* / Q000473:pathology; D009389:Neovascularization, Pathologic / Q000188:drug therapy; D019151:Peptide Library; D054218:Precursor T-Cell Lymphoblastic Leukemia-Lymphoma / Q000188:drug therapy / Q000378:metabolism / Q000473:pathology; D051881:Receptor, Notch1 / Q000037:antagonists & inhibitors / Q000276:immunology; D051883:Receptor, Notch2 / Q000037:antagonists & inhibitors / Q000276:immunology; D051880:Receptors, Notch / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects
chemical_list: D020533:Angiogenesis Inhibitors; D000906:Antibodies; C497761:NOTCH1 protein, human; C497764:NOTCH2 protein, human; C497762:Notch1 protein, mouse; C497765:Notch2 protein, mouse; D019151:Peptide Library; D051881:Receptor, Notch1; D051883:Receptor, Notch2; D051880:Receptors, Notch
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