Near-IR Bioluminescence Imaging

描述(申请人提供)：萤火虫荧光素酶的生物发光成像(BLI)作为一种有效、廉价和非侵入性的方法已被广泛接受，用于监测整个生物体的基因表达、酶活性、蛋白质-蛋白质相互作用和蛋白质降解。相对于其他成像手段，如PET和MRI，生物发光成像具有成本低、速度快、灵敏度高、高通量和易于非专业人员使用等优点。这些优势使BLI成为快速评估肿瘤进展和对潜在治疗方法反应的首选方法。BLI的主要限制是可见光对组织的穿透性差。被照亮的组织对近红外光(650-900 nm)最透明，在近红外光中，自体荧光、散射和被血红蛋白吸收的可见光最少。尽管花了相当大的努力来分离和诱变发光蛋白，但并没有最大发光波长&gt；650 nm的荧光素酶。我们提出了两种将萤火虫荧光素酶的光输出转移到近红外的协同方法：1)将共振能量转移到目标近红外荧光团，2)合成最大限度地发射更长波长的新型荧光素底物。这些目标与体内癌症成像相关，因为它们将提高生物发光成像的速度、检测极限和深度穿透。将表达发光蛋白的癌细胞引入老鼠体内，使研究人员能够监测活体动物的癌症进展和对药物治疗的反应。然而，发射的光被血液强烈吸收，因此不会通过老鼠传播很远。我们正在改变发射光的波长，以避免被血液吸收，并允许光线更深地穿透老鼠，从而提高我们检测小肿瘤或深部器官肿瘤的能力。

DESCRIPTION (provided by applicant): Bioluminescent imaging (BLI) with firefly luciferase has gained widespread acceptance as a powerful, inexpensive, and non-invasive method to monitor gene expression, enzymatic activity, protein-protein interactions and protein degradation in the context of the whole organism. Relative to other imaging modalities such as PET and MRI, bioluminescence imaging has the advantages of low cost, speed, sensitivity, high throughput and ease of use by non-specialists. These advantages make BLI the method of choice for rapidly assessing tumor progression and response to potential therapeutics. The major limitation of BLI is the poor penetration of visible light through tissue. Illuminated tissue is most transparent to near-IR light (650-900 nm), where autofluorescence, scattering, and the absorption of visible light by hemoglobin is minimal. Despite considerable effort spent isolating and mutagenizing luminescent proteins, there is no luciferase that maximally emits light >650 nm. We propose two synergistic approaches to shift the light output of firefly luciferase to the near-IR: 1) Resonance energy transfer to a targetable near-IR fluorophore, and 2) Synthesis of novel luciferin substrates that maximally emit light at longer wavelengths. These aims are relevant to in vivo cancer imaging because they will improve the speed, detection limit, and depth penetration of bioluminescence imaging. Introduction of cancer cells that express light-emitting proteins into mice has allowed researchers to monitor cancer progression and response to drug treatment in living animals. However, the emitted light is strongly absorbed by blood, and thus does not travel far through the mouse. We are changing the wavelength of the emitted light to avoid absorption by blood and allow deeper penetration of the light through the mouse, improving our ability to detect small tumors or tumors in deep organs.