TY - JOUR AU - Schwalm, Martin P AU - Lenz, Christopher AU - Saxena, Krishna AU - Klionsky, Daniel J AU - Proschak, Ewgenij AU - Knapp, Stefan TI - Biochemical investigation of LC3/GABARAP-ligand interaction as an important quality measure for LC3/GABARAP-targeting small molecules: addendum to the guidelines (4th edition). JO - Autophagy VL - 21 IS - 9 SN - 1554-8627 CY - Abingdon, Oxon PB - Taylor & Francis M1 - DKFZ-2025-00952 SP - 2069-2073 PY - 2025 N1 - ISSN 1554-8635 / 2025 Sep;21(9):2069-2073 AB - Targeted protein degradation (TPD) represents a new therapeutic modality that allows the targeting of proteins that are considered undruggable by conventional small molecules. While TPD approaches via the ubiquitin-proteasome system are well established and validated, additional degradation pathways still require rigorous characterization. Here, we focus on macroautophagy/autophagy tethering compounds, a class of small molecules, designed to recruit cargo to LC3/GABARAP proteins for subsequent autophagosome-dependent degradation. We provide guidance for the biophysical and structural characterization of small molecule modulators for studying LC3/GABARAP-ligand interactions. In addition, we discuss potential limitations of autophagy-based TPD systems and emphasize the need for rigorous quality control in the development of LC3/GABARAP-targeting small molecules.Abbreviations: DSF: differential scanning fluorimetry; FP: fluorescence polarization; FRET: Förster/fluorescence resonance energy transfer; HTRF: homogeneous time-resolved fluorescence; ITC: isothermal titration calorimetry; LIR: LC3-interacting region; MGs: molecular glues; NMR: nuclear magnetic resonance; PROTACs: PROteolysis-TArgeting Chimeras; SPR: surface plasmon resonance; TPD: targeted protein degradation; TR-FRET: time-resolved Förster/fluorescence resonance energy transfer; UPS: ubiquitin-proteasome system. KW - ATTECs (Other) KW - AUTACs (Other) KW - Atg8 (Other) KW - GABARAP (Other) KW - LC3 (Other) LB - PUB:(DE-HGF)16 C6 - pmid:40344429 DO - DOI:10.1080/15548627.2025.2498506 UR - https://inrepo02.dkfz.de/record/301267 ER -