Matrix Metalloproteinase-9 (MMP-9) Activatable Gold Nanoparticles for In Situ Zymography and Diagnostics of Neurofibromatosis Type 2 (NF2) Tumors

Neurofibromatosis type 2-related schwannomatosis (NF2-SWN) is a devastating genetic tumor-disposition syndrome characterized by multiple nervous system neoplasms. A hallmark of NF2-SWN is bilateral vestibular schwannomas (VSs) that cause hearing loss, vertigo, and life-threatening brainstem compression. Current imaging methods detect NF2-associated VS often late in their development and could lead to delayed therapeutic interventions. Matrix metalloproteinase-9 (MMP-9) is a key protease involved in extracellular matrix remodeling and tumor progression in NF2-associated VS, making it an attractive molecular target for activity-based sensing. Here, we develop MMP-9 activatable gold nanoparticles (AuNPs) incorporating a protease-cleavable peptide to enable sensitive reporting of protease activity in VS tissue. Through systematic tuning of polyethylene glycol (PEG) linker length and peptide valency, we establish an AuNP configuration that exhibits both efficient enzymatic cleavage and selectivity for MMP-9 over other VS-associated proteases. In schwannoma tissue, nanoparticles distinguish tumor from healthy nerve with >95% accuracy and detect 2 mm tumorscorresponding, based on published VS growth rates, to detection approximately 23 months earlier than conventional MRIenabling substantially earlier intervention. Longitudinal measurement of nanoparticle activation further resolves changes in MMP-9 activity within the tumor in response to therapeutic intervention, illustrating the platform's capacity in monitoring treatment response. Together, these findings establish MMP-9 activatable AuNP as a sensitive, spatially resolved diagnostic tool for NF2-SWN that complements imaging by directly quantifying protease activity in tumors that are inaccessible to biopsy.