Fluorescence Guided Surgery using Near Infrared Nerve-specific Probes for Cranial Nerve Preservation

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PROJECT SUMMARY Iatrogenic nerve injury presents one of the most feared surgical complications and a major source of morbidity across all surgical specialties. While crucial to maintain their vital functions, preservation of cranial nerves re- mains a major challenge in skull base surgeries. Additionally, with the increased use of minimally invasive sur- gical approaches, difficulty with cranial nerve identification and visualization is amplified. Cranial nerves are often intimately associated with tumors of the skull base, where surgery can be curative, but must be balanced against injury risk. Unlike other critical tissues (e.g., blood vessels), nerve repair produces negligible, unreliable func- tional improvement. Permanent motor or sensory disabilities and chronic neuropathies result, limiting patient quality of life, employability and participation in daily activities. The only known method to avoid nerve injury is to prevent its occurrence. However, no clinically approved technology sufficiently enhances intraoperative nerve visualization, where a combination of neuroanatomical knowledge, white light visualization and neurophysiolog- ical monitoring are currently used. This work will directly address this unmet clinical need. Fluorescence Guided Surgery (FGS) has successfully integrated into clinical medicine with only a few FDA-approved fluorophores (i.e., fluorescein, Aminolevulinic/Protoporphyin IX [ALA/PpIX], methylene blue and indocyanine green [ICG]). While FGS systems for imaging outside the skull operate almost exclusively in the near infrared (NIR, 650-900 nm), imaging system for neurosurgery are commonly equipped with the ability to image in the visible. However, all clinical FGS systems have an “800 nm” channel designed to image ICG, including neurosurgical microscopes and wide-field exoscopes. To facilitate clinical translation and utility for neurosurgery, the overall goal herein is to generate a nerve-specific small molecule fluorophore with spectral properties matched to ICG. This novel probe would enable cranial nerve visualization that is spectrally distinct from the visible fluorescein and ALA/PpIX that are commonly used for tumor enhancement, while enabling future clinical translation using existing neuro- surgical FGS infrastructure. Development of a NIR nerve-specific probe has presented a synthetic challenge as molecules must be small enough to cross the tight blood nerve and/or blood brain barrier(s) (BNB and/or BBB, respectively), but with a sufficient degree of conjugation to reach NIR wavelengths. This is a particular challenge in neurosurgical applications where identification and visualization of structures at the interface of the peripheral and central nervous system (PNS and CNS, respectively) are required for successful surgical outcomes. In pre- liminary work, our group has designed and developed NIR oxazine-based probes with exquisite PNS specificity and recently discovered that a new library of oxazine probes provide BBB-cross and CNS specificity, however an agent suitable for clinical translation does not yet exist. Herein, we will develop NIR nerve-specific fluoro- phore(s) for identification and visualization of the cranial nerves with and without tumor counterstain (i.e., fluo- rescein or ALA/PpIX) in animal models of meningioma and schwannoma.