Introduction: Brain tumors remain a significant clinical problem, affecting over 60,000 newly diagnosed Americans each year. During surgery, frozen section diagnoses frequently determine surgical excision versus medical management. However, in some cases, neither gross morphology nor frozen sections can distinguish non-operative versus operative lesions, such as lymphoma versus astrocytoma. In such cases, immunohistochemistry is ordered, which requires 1-2 days to yield a specific diagnosis. Initial diagnostic uncertainty may lead to lengthened hospital admissions and additional surgeries for patients, or even an inappropriate treatment strategy. Improved tools for specific intraoperative brain tumor diagnoses are desperately needed. Here, we develop a novel class of nanoprobes for rapid and specific intraoperative diagnoses.
Methods: Nanoprobes were designed targeting an immunoglobulin on CD20-positive human lymphoma cells, and fluorescence dynamics were evaluated by spectro-fluorometry. Utilizing a one-step staining protocol, labeling affinity was interrogated by flow cytometry and confocal imaging of human cell culture and fresh xenograft brain tumor biopsies harboring human B-cell lymphoma or astrocytoma.
Results: Nanoprobes produced minimal fluorescence in unbound conformations and generated 8-fold increased fluorescence once bound to CD20-positive lymphoma cells. Flow cytometry and confocal imaging demonstrated strong binding to B-cell lymphoma cells within 15 minutes of incubation. Within one hour, specific histopathological differentiation of lymphoma from astrocytoma was routinely possible from biopsies (80.75 ± 2.52% lymphoma cells vs. 8.25 ± 1.51% astrocytoma cells, p< 0.001).
Conclusions: This activatable fluorescent nanoprobe provides rapid and specific labeling of B-cell lymphoma. Our one-step ex vivo labeling approach simplifies tissue staining and drastically reduces time to histopathological diagnoses relative to IHC-based methods. This may improve both speed and accuracy of brain tumor diagnoses, allowing for proper goal-oriented surgery and enrollment in the appropriate treatment strategy.
Patient Care: The nanoprobes developed in this project provide rapid differentiation of operative versus non-operative lesions when frozen section diagnoses are inconclusive. Clinical utilization of these nanoprobes could provide early confidence to proceed with specific treatment plans.
Learning Objectives: By the conclusion of this session, participants should be able to: 1) Describe the advantages of intraoperative fluorescence imaging with novel contrast agents compared to current intraoperative diagnostics, 2) Discuss in small groups the advantages/disadvantages of fluorescence ex vivo imaging compared to frozen section imaging, 3) Identify clinical scenarios which would benefit from rapid and specific intra-operative diagnostics.