Multifunctional in Vivo EPR Imaging of Tumor Microenvironment

项目总结/摘要 肿瘤微环境（TME）是影响肿瘤恶性程度的化学和生物学背景， 增殖和转移活性，因为混乱的血管生成肿瘤生长变得缺氧 (low pO 2）和更酸性（低pH值）。虽然已知pO 2和pH是肿瘤生长的重要因素， 治疗，间质Pi最近已被确定为在肿瘤发生中重要的新信号分子。 新合成的稳定顺磁分子，自旋探针，允许同时在体内监测 间质pO 2、细胞外pH（pHe）和间质无机磷酸盐（Pi）浓度，使用电子 顺磁共振（EPR）。因为肿瘤是高度异质性的，所以光谱-空间EPR成像是可行的。 需要空间分辨感兴趣的参数。空间分辨率由EPR灵敏度定义。的 体素内的自旋探针分子的量随着空间分辨率的立方而减少，并且可以变为 如果太小则无法检测。灵敏度也直接转化为功能分辨率，即 可以测量pH、pO 2和Pi。PI及其同事关于快速扫描（RS）EPR的建议 在250 MHz频率上探索了该技术，并显示出信噪比提高到2 与标准场调制连续波（CW）方法相比，其数量级更高。额外 对于基于三苯甲基的多功能探针，可以通过增加 EPR频率为700 - 800 MHz（最适合小动物RS EPR研究），扫描频率为50-100 kHz (the当前的数学算法将上扫描频率限制为大约10 kHz）。RS EPR灵敏度 增强将需要硬件（SA 1）和软件（SA 2）的开发。此外，新的4D光谱- 空间算法将进一步发展，使多功能多线EPR成像，因为标准 滤波反投影（FBP）重建不能与具有多功能自旋探针的多功能自旋探针一起工作， 相对较宽的多线EPR谱。一个新开发的多功能EPR成像仪将首先进行测试 在具有已知几何形状、pO 2、pH和Pi模式的标准样品上进行体外试验，然后进行体内概念验证 显像将使用一群自发发展乳腺癌的PyMT转基因小鼠来进行研究。 快速扫描多功能成像和构建TME的空间分辨pO 2、pH和Pi分布， 正常乳腺。我们期望实现生理上显著的体素特异性功能敏感性 约1-2 mmHg的pO 2、0.05 pH单位和0.1 mM的无机磷酸盐。估计空间分辨率将 小于200 μ m。总之，该项目的成功可能会对未来产生重大影响。 在体内功能成像研究TME和超越。

PROJECT SUMMARY/ABSTRACT The tumor microenvironment (TME) is the chemical and biological background that affects tumor malignancy, proliferation and metastatic activity because of chaotic angiogenesis tumors growing to become less oxygenated (low pO2) and more acidic (low pH). While pO2 and pH are known to be important factors for tumor growth and treatment, interstitial Pi has been recently identified as a new signaling molecule of importance in tumorigenesis. Newly synthesized stable paramagnetic molecules, spin probes, permit simultaneous in vivo monitoring of interstitial pO2, extracellular pH (pHe) and concentration of interstitial inorganic phosphate (Pi) using Electron Paramagnetic Resonance (EPR). Because tumors are highly heterogeneous, spectral-spatial EPR imaging is required to spatially resolve the parameters of interest. Spatial resolution is defined by EPR sensitivity. The amount of the spin probe molecules within a voxel decreases as spatial resolution is cubed, and may become undetectable if too small. Sensitivity also directly translates into functional resolution, the accuracy with which pH, pO2, and Pi can be measured. The proposal by the PI and his colleagues for a Rapid Scan (RS) EPR technique has been explored at 250 MHz frequency and showed improvement in signal-to-noise ratio up to two orders of magnitude compared to the standard field-modulated continuous wave (CW) method. An additional order of magnitude signal enhancement can be achieved for trityl-based multifunctional probes by increasing EPR frequency to 700 -800 MHz (optimal for small animal RS EPR studies) and scan frequency to 50-100 kHz (the current mathematical algorithm limits the upper scan frequency to about 10 kHz). RS EPR sensitivity enhancement will require both hardware (SA1) and software (SA2) developments. In addition, novel 4D spectral- spatial algorithm will be further developed to enable multi-functional multi-line EPR imaging, since the standard filtered backprojection (FBP) reconstruction fails to work with multi-functional spin probes that have a comparatively broad multi-line EPR spectra. A newly developed multi-functional EPR imager will first be tested in vitro on standard samples with known geometry, pO2, pH, and Pi patterns, followed by in vivo proof-of-concept imaging. A colony of PyMT transgenic mice will be used that spontaneously-develop breast cancer to perform the rapid scan multifunctional imaging and construct spatially-resolved pO2, pH, and Pi profiles of TME and normal mammary gland. We anticipate to achieve physiologically significant voxel-specific functional sensitivities of about 1-2 mmHg of pO2, 0.05 pH units and 0.1 mM of inorganic phosphate. Estimated spatial resolution will be smaller than 200 m. In summary, the success of this project may have a significant impact on the future of in vivo functional imaging to study TME and beyond.