Ultra-Low Background NIR Fluorophores for In Vivo Imaging and Image-Guided Surger

描述(由申请人提供)：近红外(NIR)荧光有可能使图像引导手术发生革命性变化。然而，理想的荧光团在体内，并最终在临床上的使用还没有描述。在美国国立卫生研究院生物工程研究伙伴关系(BRP)的资助下，PI的实验室开发了一种外科成像系统，该系统可以同时实时获取两个独立波长的近红外荧光发射图像以及彩色视频图像。该成像系统已经被转化到临床，并正在由NIH资助的三项临床试验中进行正式评估。然而，这项技术未来成功的根本限制是开发在人体内表现最佳的近红外荧光团，并可广泛用于其他学术研究人员。为了在临床上可行，理想的近红外荧光团需要某些光学性质，包括激发和发射H800 nm，以及血清中的高消光系数(5)和量子产率(QY)。然而，现有的近红外荧光团在体内表现如此之差的原因更多地与生物分布和清除有关。静脉注射后，理想的近红外荧光团将在血管内和血管外空间之间迅速平衡，并通过肾滤过有效地清除。到目前为止，文献中描述的每个近红外荧光团都存在两个基本缺陷：1)肝脏清除，导致近红外荧光信号在整个胃肠道持续数小时，和/或2)正常组织中的非特异性背景摄取，通常持续数小时，并导致低信号背景比(SBR)。这笔赠款建立在我们两年前使用近红外荧光量子点(Choi等人，自然生物技术)观察到的基础上。2007年；25：1165-70)。出乎意料的是，由于部分原因，两性离子有机涂层导致极低的非特异性组织摄取率，快速的肾脏清除，并且没有血清蛋白结合。然而，纯阴离子或阳离子涂层的结果正好相反。基于这些数据，我们开始与佐治亚州立大学的Patonay和Strekowski博士合作，他们是近红外荧光团化学领域的领先者，合成了两性离子七甲基吲哚菁近红外荧光团。这里描述的初步结果表明，非靶向和肿瘤靶向的两性离子近红外荧光团都具有显著的光学和体内性质，包括800 nm荧光、高5和QY、快速肾脏清除、无蛋白质结合和超低非特异性组织摄取(即背景)。这笔赠款的具体目标是合成用于活体和外科成像的优化的两性离子近红外荧光团，验证它们作为前列腺癌靶向诊断剂的用途，以及从分析性生产扩大到制备性生产。这些目标的完成将为未来影像引导手术的临床测试奠定基础。重要的是，我们还提出了一种知识产权战略，允许在学术界内免费共享优化的近红外荧光团。 与公共健康相关：近红外光对人眼是不可见的，但可以相对深入地穿透到活组织中。因此，它是图像引导手术的理想选择，因为它为外科医生提供了对癌症等疾病的高灵敏度、高分辨率检测，而不会改变手术领域的外观。虽然使用近红外荧光灯进行图像引导手术的硬件系统已经可用，但优化的荧光团或“灯泡”还没有。这笔赠款的目标是开发一种可以注入血液的新型理想的近红外荧光团。这些荧光团会“附着”在肿瘤和其他病变组织上，但不会附着在正常组织上。

DESCRIPTION (provided by applicant): Near-infrared (NIR) fluorescence has the potential to revolutionize image-guided surgery. However, ideal fluorophores for in vivo, and eventually clinical, use have not yet been described. Under an NIH Bioengineering Research Partnership (BRP) grant, the PI's laboratory has developed a surgical imaging system that simultaneously, and in real-time, acquires two independent wavelengths of NIR fluorescence emission images along with color video images. The imaging system has already been translated to the clinic, and is being formally evaluated in three NIH-funded clinical trials. Nevertheless, the fundamental limitation to the future success of this technology is the development of NIR fluorophores that perform optimally in the body, and which can be made widely available to other academic researchers. To be clinically viable, the ideal NIR fluorophore requires certain optical properties, including excitation and emission H800 nm, and both a high extinction coefficient (5) and quantum yield (QY) in serum. However, the reason why existing NIR fluorophores perform so poorly in vivo has more to do with biodistribution and clearance. After IV injection, the ideal NIR fluorophore would rapidly equilibrate between the intravascular and extra vascular spaces and would be cleared efficiently via renal filtration. To date, every NIR fluorophore described in the literature suffers from two fundamental flaws: 1) hepatic clearance, which results in NIR fluorescence signal throughout the GI tract that persists for hours, and/or 2) non-specific background uptake in normal tissues, which typically persists for hours and results in a low signal-to-background ratio (SBR). This grant builds upon an observation we made two years ago using NIR fluorescent quantum dots (Choi et al., Nature Biotechnol. 2007; 25: 1165-70). Unexpectedly, and for reasons only partially understood, zwitterionic organic coatings resulted in extremely low non-specific tissue uptake, rapid renal clearance, and no serum protein binding. However, purely anionic or cationic coatings gave the opposite results. Based on these data, we began collaborating with Drs. Patonay and Strekowski at Georgia State University, leaders in the field of NIR fluorophore chemistry, to synthesize zwitterionic heptamethine indocyanine NIR fluorophores. The preliminary results, described herein, demonstrate that both non-targeted and tumor-targeted zwitterionic NIR fluorophores have remarkable optical and in vivo properties, including 800 nm fluorescence, high 5 and QY, rapid renal clearance, absence of protein binding, and ultra-low non-specific tissue uptake (i.e., background). The specific aims of this grant are focused on the synthesis of optimized zwitterionic NIR fluorophores for in vivo and surgical imaging, on validating their use as targeted diagnostic agents for prostate cancer, and for scale-up from analytical to preparative production. Completion of these aims will lay the foundation for future clinical testing during image-guided surgery. Importantly, we also present an intellectual property strategy that will permit free sharing of optimized NIR fluorophores within the academic community. PUBLIC HEALTH RELEVANCE: Near-infrared light is invisible to the human eye, but penetrates relatively deeply into living tissue. It is therefore ideal for image-guided surgery, because it provides surgeons with high- sensitivity, high-resolution detection of diseases, such as cancer, without changing the look of the surgical field. Although hardware systems that use near-infrared fluorescent light for image-guided surgery are already available, optimized fluorophores, or "light bulbs" are not. The goal of this grant is to develop a new class of ideal near-infrared fluorophores that can be injected into the bloodstream. These fluorophores would "stick" to tumors and other diseased tissue, but not to normal tissue.