ASTE1 (asteroid structure-specific endonuclease 1) is a structure-specific DNA endonuclease that specifically cleaves single-stranded DNA and 3' overhang DNA structures 1. As a downstream effector of the shieldin complex, ASTE1 is recruited to DNA double-strand breaks (DSBs) in a SHLD2-dependent manner and functions to promote non-homologous end-joining (NHEJ) while antagonizing homologous recombination (HR) 1. Mechanistically, ASTE1 cleaves 3' exposed DNA ends, thereby inhibiting DNA end resection—a process necessary for HR—and promoting DNA end protection required for NHEJ 1. Loss of ASTE1 impairs NHEJ, causes hyper-resection, and leads to defective immunoglobulin class switch recombination 1. In BRCA1-deficient cells, ASTE1 deficiency restores homologous recombination capacity and confers resistance to poly(ADP-ribose) polymerase inhibitors 1. Clinically, ASTE1 is frequently mutated in microsatellite-instable (MSI) cancers, particularly colorectal cancer, where frameshift mutations in ASTE1 occur in ~80% of MSI tumors and generate immunogenic neopeptides recognized by CD8+ T cells 234. These ASTE1-derived frameshift peptide neoantigens correlate with tumor-infiltrating lymphocyte density and have been evaluated in phase I/IIa neoantigen vaccine trials for MMR-deficient cancers 4.