PT  - JOURNAL ARTICLE
AU  - David Gonzalez-Perez
AU  - Satyajit Das
AU  - Elliot Medina
AU  - Daniel Antfolk
AU  - Emily D. Egan
AU  - Stephen C. Blacklow
AU  - Paulo C. Rodriguez
AU  - Vincent C. Luca
TI  - An affinity-matured DLL4 ligand for broad-spectrum activation and inhibition of Notch signaling
AID  - 10.1101/2022.03.07.483330
DP  - 2022 Jan 01
TA  - bioRxiv
PG  - 2022.03.07.483330
4099  - http://biorxiv.org/content/early/2022/03/07/2022.03.07.483330.short
4100  - http://biorxiv.org/content/early/2022/03/07/2022.03.07.483330.full
AB  - The Notch pathway regulates cell fate decisions and is an emerging target for regenerative and cancer therapies. Recombinant Notch ligands are attractive candidates for modulating Notch signaling; however, their intrinsically low receptor-binding affinity restricts their utility in biomedical applications. To overcome this limitation, we evolved variants of the ligand Delta-like 4 (DLL4) with enhanced affinity and cross-reactivity. A consensus variant with maximized binding affinity, DeltaMAX, engages human and murine Notch receptors with 500- to 1000-fold increased affinity compared to wild-type human DLL4. DeltaMAX also potently activates human Notch in plate-bound, bead-bound, and cellular formats. When administered as a soluble decoy, DeltaMAX inhibits Notch activation in response to either Delta-like (DLL) or Jagged (Jag) ligands, highlighting its utility as both an agonist and antagonist. Finally, we demonstrate that DeltaMAX stimulates increased proliferation and expression of effector mediators in primary activated human T cells. Taken together, our data defines DeltaMAX as a versatile biotechnological tool for broad-spectrum activation or inhibition of Notch signaling.Competing Interest StatementV.C.L., P.C.R., and D.G.P., have filed provisional patents describing the engineering and applications of DeltaMAX (application numbers PCT/US2020/041765 and PCT/US2020/030977). V.C.L. is a consultant on an unrelated project for Cellestia Biotech. S.C.B. is on the SAB and receives funding from ERASCA, Inc. for an unrelated project, and is a consultant on unrelated projects for Scorpion Therapeutics, Odyssey Therapeutics, Ayala Pharmaceuticals, MPM Capital, and Droia Ventures.