Accurate detection of brain tumor boundaries is crucial for successful tumor removal and better patient outcomes. A novel method using label-free Fluorescence Lifetime Imaging (iFLIm) is presented in this study. The approach involved developing an optimized classification model based on tumor enhancement status, utilizing multispectral FLIm. The method was evaluated on 52 patients with adult-type diffuse glioma, demonstrating promising results with 87% sensitivity, 92% specificity, and an AUC of 0.90. This FLIm-based model has the potential to offer a non-invasive and real-time technique to assist neurosurgeons in accurately identifying tumor infiltrates, potentially improving tumor resection and patient outcomes.
Stereotactic needle biopsy is a time-consuming and invasive procedure that often cannot accurately distinguish recurrent tumors from treatment effect in gliomas. We report an intraoperative multispectral fluorescence lifetime imaging (FLIm) system coupled with a custom-made fiber optic probe integrated with the stealth biopsy needle as an optical biopsy tool. FLIm parameters collected from 3 suspected recurrent glioma patients changed over the biopsy trajectory as the needle passed different brain areas. An SVM classifier validated using a leave-one-patient-out validation scheme could identify the lesions from the normal surrounding tissue with sensitivity=0.99, specificity=0.91, and accuracy=0.95.
In-situ identification of glioma subtype can enable modifications of clinical and surgical strategies. Particularly, astrocytoma benefit from more aggressive resection than oligodendroglioma, which have a more favorable response to post-surgical chemotherapy. Preoperative MRI and intraoperative histology cannot accurately determine glioma subtype. There is a need for real-time identification of adult-type diffuse glioma subtypes to aid the neurosurgeon’s decision-making during resection surgery. Fluorescence lifetime imaging (FLIm) where tissue autofluorescence can be used as an indicator to distinguish among brain tumor tissue types in real-time could aid this process. Here, we report the use of label-free FLIm in distinguishing IDH-mutant glioma subtypes (astrocytoma and oligodendroglioma). The FLIm system (excitation: 355 nm; emission bands: 390/40 nm, 470/28 nm, 542/50 nm) was used to scan brain tissue from the resection margins of glioma patients during tumor resection. Fluorescence lifetimes were extracted and analyzed by constrained least-squares deconvolution with the Laguerre expansion method. FLIm data was validated with histopathology of collected biopsies. Current results show that FLIm provides optical contrast between tumor and healthy white matter, and between IDH-mutant astrocytoma (N=7 patients) and oligodendroglioma (N=5 patients). Tumors showed shorter lifetime values (470-nm: 3.6±0.6ns; 542-nm: 3.3±0.7ns) than healthy white matter (470-nm: 4.6±0.4ns; 542-nm: 4.3±0.5ns, p<0.01). Oligodendroglioma had shorter lifetimes in the 470-nm (3.3±0.1ns) and 542-nm (2.8±0.2ns) channels, which are associated with NAD(P)H and FAD fluorescence respectively, when compared with IDH-mutant astrocytoma (470-nm: 4.1±0.1ns; 542-nm: 3.9±0.2ns, p<0.01). Together, these results demonstrate the feasibility of using FLIm as an intraoperative tool in glioma diagnosis.
The standard treatment for infiltrative gliomas is surgery to remove as much tumor tissue as possible without compromising neurological functions. Thus, real-time identification of infiltrative tumor tissue is necessary. Here a fluorescence lifetime imaging (FLIm) was used to distinguish between healthy brain and areas with different degrees of tumor cellularity as defined by histopathology. We conducted FLIm measurements and collected microbiopsies from tumor resection margins to identify the FLIm characteristics of tumor edges in cortex and white matter of low- and high-grade gliomas. Results from a 13-patient cohort indicate that FLIm identifies infiltrations of up to moderate tumor cellularity.
5-ALA induced PpIX is increasingly used for guiding brain tumor resection surgery. The current intensity-based approach fails at detecting lower concentrations of PpIX found in low-grade gliomas or the infiltrative edge of glioblastomas. Here, we report the first results in patients of real-time, PpIX fluorescence lifetime measurements using a hand-held fiber probe. Fluorescence from different spectral channels (390/40 nm (Collagen), 470/28 nm (NADH), 629/53 nm (PPIX)), is augmented onto a video stream of the surgical field-of-view to provide intraoperative tumor visualization in real-time. In-vivo data reveals strong contrast between regions of high PpIX accumulation associated with tumor (>8 ns) and healthy brain tissue (<4 ns).
Significance: 5-aminolevulinic acid (5-ALA)-induced protoporphyrin IX (PpIX) fluorescence is currently used for image-guided glioma resection. Typically, this widefield imaging method highlights the bulk of high-grade gliomas, but it underperforms at the infiltrating edge where PpIX fluorescence is not visible to the eyes. Fluorescence lifetime imaging (FLIm) has the potential to detect PpIX fluorescence below the visible detection threshold. Moreover, simultaneous acquisition of time-resolved nicotinamide adenine (phosphate) dinucleotide [NAD(P)H] fluorescence may provide metabolic information from the tumor environment to further improve overall tumor detection.
Aim: We investigate the ability of pulse sampling, fiber-based FLIm to simultaneously image PpIX and NAD(P)H fluorescence of glioma infiltrative margins in patients.
Approach: A mesoscopic fiber-based point-scanning FLIm device (355 nm pulses) was used to simultaneously resolve the fluorescence decay of PpIX (629/53 nm) and NAD(P)H (470/28 nm). The FLIm device enabled data acquisition at room light and rapid (<33 ms) augmentation of FLIm parameters on the surgical field-of-view. FLIm measurements from superficial tumors and tissue areas around the resection margins were performed on three glioblastoma patients in vivo following inspection of PpIX visible fluorescence with a conventional neurosurgical microscope. Microbiopsies were collected from FLIm imaged areas for histopathological evaluation.
Results: The average lifetime from PpIX and NAD(P)H fluorescence distinguished between tumor and surrounding tissue. FLIm measurements of resection margins presented a range of PpIX and NAD(P)H lifetime values (τPpIX ∼ 3 to 14 ns, τNAD(P)H = 3 to 6 ns) associated with unaffected tissue and areas of low-density tumor infiltration.
Conclusions: Intraoperative FLIm could simultaneously detect the emission of PpIX and NAD(P)H from patients in vivo during craniotomy procedures. This approach doubles as a clinical tool to identify tumor areas while performing tissue resection and as a research tool to study tumor microenvironmental changes in vivo. Intraoperative FLIm of 5-ALA-induced PpIX and tissue autofluorescence makes a promising surgical adjunct to guide tumor resection surgery.
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