Intraoperative confocal microscopy for quantitative delineation of low-grade glio

描述(申请人提供)：近年来，由5-氨基乙酰丙酸(5-ALA)诱导的原卟啉IX(PpIX)提供对比的荧光图像引导手术(FICS)已被证明可以改善胶质瘤患者的手术结果。然而，这种宽视场(低分辨率)成像方法有其局限性。具体地说，根据胶质瘤荧光强度的细微变化很难准确识别手术边缘，因为胶质瘤是弥漫的，缺乏从肿瘤到正常组织的明显过渡。此外，宽视野(低分辨率)方法，如无花果和MRI，提供的像素强度代表许多细胞的平均值，导致检测胶质瘤外围稀疏肿瘤细胞群的能力减弱。这一问题在低级别胶质瘤中加剧，在低级别胶质瘤中，5-ALA诱导的PpIX荧光仅由罕见的增殖细胞群产生，通常无法通过广野无花果检测到。最近，该项目的合作者Nader Sanai博士展示了术中细胞分辨率共聚焦显微镜可以用来显示接受5-ALA治疗的低级别胶质瘤患者中这些稀疏的荧光细胞。这一发现的意义是非常重要的，因为有报道说 对于低级别胶质瘤，大体全切除率(GTR)一直是次佳的(14%至46%)，这表明需要改进图像引导技术。在这里，我们建议开发一种手持式术中共聚焦显微镜来观察5-ALA诱导的PPIX在低级别胶质瘤组织中的表达。临床标本的体外成像研究将证明，这种实时在体成像技术1)与金标准组织病理学相比，能够准确地量化PPIX的表达，2)具有基于T2加权磁共振成像的灵敏度，能够识别超出传统放射边缘的肿瘤浸润性，3)与目前唯一用于神经外科的光学切片显微镜--蔡司Optiscan(R)共聚焦显微镜相比，提供更好的图像质量。最后，提出了一项首次人体研究，以证明将我们的设备整合到手术工作流程中的可行性，提供实时定量的“光学活检”，补充现有的神经外科广域成像技术，并帮助在肿瘤切除的最后阶段校准手术决策。临床前和临床研究将利用巴罗神经研究所(BNI)的Nader Sanai博士的现有专业知识，该研究所是美国规模最大的胶质瘤切除手术中心，也是唯一一家同时使用广野无花果和术中共焦显微镜的机构。这些技术发展和翻译研究的结果将证明，未来的临床试验将定量研究术中共聚焦显微镜作为一种手术策略，以最大限度地扩大切除范围，最大限度地减少手术发病率，并提高低级别胶质瘤患者的总体生存。

DESCRIPTION (provided by applicant): In recent years, fluorescence image-guided surgery (FIGS) with contrast provided by 5-aminolevulinic acid (5- ALA)-induced protoporphyrin IX (PpIX) has been demonstrated to improve surgical outcomes for glioma patients. However, there are limitations to this wide-field (low-resolution) imaging method. Specifically, it is diffiult to accurately identify a surgical margin based on subtle variations of fluorescence intensity in gliomas, which are diffuse and lack a distinct transition from tumor to normal tissue. Furthermore, wide-field (low- resolution) approaches, such as FIGS and MRI, provide pixel intensities that represent an average value from many cells, resulting in a diminished ability to detect the sparse tumor cell populations at the glioma periphery. This problem is exacerbated in low-grade gliomas, where 5-ALA-induced PpIX fluorescence is only generated by rare proliferating cell populations and is typically undetectable via wide-field FIGS. Recently, Dr. Nader Sanai, a collaborator on this project, has shown that intraoperative cellular-resolution confocal microscopy can be used to visualize these sparse fluorescent cells in low-grade glioma patients treated with 5-ALA. The implications of this finding are highly significant since reported rates of gross-total resection (GTR) have been suboptimal for low-grade gliomas (14% to 46%), suggesting a need for improved image-guidance techniques. Here, we propose to develop a hand-held intraoperative confocal microscope to visualize 5-ALA-induced PpIX expression in low-grade glioma tissues. Ex vivo imaging studies with clinical specimens will demonstrate that this real-time in vivo imaging technique 1) quantifies PpIX expression accurately compared to gold-standard histopathology, 2) has the sensitivity to identify tumor infiltration beyond conventional radiographic margins based on T2-weighted MRI, and 3) provides superior image quality compared to the only existing optical- sectioning microscope in neurosurgical use, the Zeiss Optiscan(R) confocal microscope. Finally, a first-in-human study is proposed to demonstrate the feasibility of incorporating our device into the operative workflow, providing a real-time quantitative "optical biopsy" that complements existing wide-field imaging techniques in neurosurgery and helps to calibrate surgical decision-making at the final stages of tumor resection. Preclinical and clinical studies will leverage the existing expertise of Dr. Nader Sanai at the Barrow Neurological Institute (BNI), which is the highest-volume operative center for glioma resection in the United States and the only institution with extensive experience using both wide-field FIGS and intraoperative confocal microscopy. The results of these technological developments and translational studies will justify future clinical trials investigating quantitatie intraoperative confocal microscopy as a surgical strategy to maximize extent of resection, minimize operative morbidity, and improve overall survival for low-grade glioma patients.