Targeting the ATG12–ATG3 protein-protein interaction: From structural insights to therapeutic opportunities in autophagy modulation

Autophagy sustains cellular metabolism, shapes immune signaling and, when dysregulated, contributes to cancer progression and cytokine-storm syndromes. A crucial catalytic step is conjugation of microtubule-associated protein 1 light chain 3 (LC3) to phosphatidylethanolamine, driven by direct binding of the E2-like enzyme autophagy-related protein 3 (ATG3) to the ubiquitin-like protein autophagy-related protein 12 (ATG12). Disrupting this ATG12–ATG3 protein–protein interaction (PPI) could silence both the degradative and secretory arms of autophagy with high pathway selectivity. Here we review the rapid evolution of ATG12–ATG3 inhibition from structural insight to drug-like chemical matter. High-resolution crystallography pinpointed a hydrophobic pocket around ATG12 Trp73 that accommodates ATG3 Met157, revealing an “anchor-and-latch”—a motif in which one residue (‘anchor’) buries deeply while flanking residues (‘latch’) secure the complex— topology ideal for small-molecule competition. A split Gaussia luciferase screen of more than 40 000 compounds, guided by in-silico pocket bias, yielded 17 micromolar disruptors; systematic structure–activity-relationship (SAR) exploration transformed an off-target casein kinase 2 (CK2) hit into naphthalene lead compound 189, which binds ATG12 directly (dissociation constant, K_D ≈ 5 µM). This lead collapses autophagic flux at single-digit micromolar concentrations, arrests autophagy-addicted tumor cells and suppresses interleukin-1β (IL-1β) secretion from macrophages—all without kinase or lysosomal liabilities. Medicinal-chemistry principles distilled from more than 150 analogues define the hydrophobic “plug,” polar “claw,” and polarity-tuning handles that govern potency and selectivity. An integrated assay toolbox—spanning surface plasmon resonance (SPR), dual-color LC3 flux reporters and disease-relevant phenotypes—now drives nanomolar optimization and safety profiling. We conclude by mapping future directions: covalent-reversible chemotypes, proteolysis-targeting chimera (PROTAC) degraders, targeted-delivery platforms and combination regimens poised to translate ATG12–ATG3 disruption into first-in-class therapeutics for oncology, immunology and infectious disease.