Oral Presentation 29th Annual Lorne Proteomics Symposium 2024

Attenuation of CXCR4 signal transduction by pharmacological and nanobody inhibition of CXCR4 receptor dimerization. (#40)

Wei Wu 1
  1. ASTAR-SIgN, Singapore, SINGAPORE

G-protein coupled receptors (GPCRs) form the largest class of plasma membrane targets that are theoretically druggable to inhibit cancer proliferation and metastasis. Interference in GPCR signaling could target any of the processes along the signal transduction path, from ligand binding to receptor dimerization, and to various steps of intracellular signal transduction. Here we report detailed mechanisms of (i) CXCR4 inverse agonism by a small molecule inhibitor, and (ii) inhibition of receptor dimerization by an engineered nanobody, leading to attenuation of CXCR4 intracellular signal relay. Using biophysical approaches such as Bioluminescence Resonance Energy Transfer (BRET) and dSTORM single-molecule imaging, we demonstrate the impact of these modes of inhibition on CXCR4 receptor oligomerisation. Using deep phosphoproteomics with temporal resolution, we identified the inhibited molecular paths and elucidated priming phosphorylation changes in leukemic cells that sensitise them to apoptosis. Finally, we hypothesized, tested and validated that interfering with CXCR4 receptor dimerization can sensitise cells to apoptosis via the said mechanism, and synergise with Venetoclax, a clinically prescribed drug to treat chronic lymphocytic leukemia. With these, we demonstrate the feasibility of targeting, not ligand binding, but receptor dimerization to attenuate CXCR4 oncogenic signaling. This unlocks a new strategy to treat receptor over-activation in cancer, that is less likely to fuel drug resistance. The work presented here showcase a generalisable roadmap to investigate this new class of drugs and the associated novel mechanisms.