A covalent molecular glue hijacks the E3 ligase MKRN2 to degrade the ribosomal protein RPS7 and induce synthetic lethality in p53‐deficient NSCLC cells

BACKGROUND AND PURPOSE: Functional inactivation of the tumour suppressor p53 drives therapeutic resistance in non-small cell lung cancer (NSCLC). While targeted protein degradation offers a potential solution, the repertoire of ligandable E3 ligases remains limited. This study aimed to identify and characterise 2,6-didehydroxyspeperomin B (DPB) as a novel covalent molecular glue that targets p53-deficient NSCLC and to elucidate its mechanism of action involving the E3 ligase MKRN2. EXPERIMENTAL APPROACH: Cytotoxicity of DPB was assessed in p53 wild-type and p53-deficient NSCLC cells and orthotopic mouse models. Direct cellular targets were identified using quantitative thiol-reactivity proteomics (QTRP) and validated via biophysical assays and site-directed mutagenesis. The molecular glue mechanism was mapped using co-immunoprecipitation-mass spectrometry and genetic knockout/rescue studies. Downstream effects on ribosome biogenesis and protein synthesis were quantified. KEY RESULTS: DPB exhibited potent, selective cytotoxicity against p53-deficient NSCLC cells and suppressed tumour growth in vivo, outperforming standard chemotherapy. The E3 ubiquitin ligase MKRN2 was the direct target, with DPB covalently modifying the Cys335 residue. Mechanistically, DPB acted as a molecular glue, creating a neo-interface recruiting the ribosomal protein RPS7 to MKRN2. This induced ubiquitination and proteasomal degradation of RPS7, triggering acute nucleolar stress and apoptosis. The synthetic lethal effect was entirely dependent on a functional MKRN2-RPS7 axis. CONCLUSION AND IMPLICATIONS: Our findings identify DPB as a covalent molecular glue that harnesses the previously unexploited E3 ligase MKRN2 to eliminate p53-deficient cancer cells. This work validates the MKRN2-RPS7 axis as a synthetic lethal vulnerability and expands the E3 ligase toolbox for targeted protein degradation, providing a novel therapeutic strategy for drug-resistant NSCLC.