PHOTOACOUSTIC MICROSCOPY OF NEOVASCULARIZATION IN RENAL CANCER GROWTH AND THERAPY

抽象的。靶向肿瘤新生血管的前景仍未实现。因此，对 微血管功能如何影响肿瘤生物学以及肿瘤或内皮细胞信号通路如何调节 新生血管是必要的。我们开发了一种新的非侵入性成像技术，光声 显微镜(PAM)。PAM使用激光激发血红蛋白(Hb)来确定新生血管构筑，Hb 毛细血管水平的浓度(红细胞压积)、血氧饱和度(SO2)和每个肿瘤微血管的流量 分辨率不需要外在的对比度，也不需要构造窗口。这些数据唯一地使 肿瘤氧耗代谢率(MRO2)的微区测定。我们将结合PAM 通过生物学、药理学和遗传操作来验证肿瘤新生血管的假设 血管内皮细胞生长因子和PI3K信号调控肿瘤细胞的结构和功能 内皮细胞。我们将研究肾癌，因为它是由于缺氧过度表达而导致的血管丰富- 诱导因子(HIF)-2和-1上调血管内皮生长因子和其他血管生成因子。我们将使用人类786-O (VHL和PTEN阴性)免疫缺陷小鼠异种移植以询问相同的血管网络 在所有肿瘤中由相同的动静脉对提供。我们将用这些具体目标来检验我们的假设：1.0。 研制一种集成的无标记光声显微镜，可对血管进行纵向成像。 切面、红细胞压积、二氧化硫、血流量和MRO2。目前我们使用两个PAM仪器来成像 分别测定红细胞压积(ChB)/二氧化硫(SO2)和血管横截面积/血流。两个系统的量化 由于重新定位和异步性，MRO2容易出错。2.0版本。确定新生血管功能， 786-O肾癌移植瘤生长过程中的肿瘤代谢和细胞生物学。我们将使用纵向 PAM成像以阐明微血管功能，肿瘤MRO2，肿瘤和内皮细胞增殖，生存， 血管生成和PI3K信号通路在肿瘤生长过程中相互交错。3.0。抑制血管内皮生长因子信号转导 测定新生血管和肾癌细胞的功能反应。我们将使用一种反 以人和鼠血管内皮生长因子为靶点的血管内皮生长因子抗体，并检测每个PAM参数的归一化， 血管内皮细胞和肿瘤细胞增殖和存活的减少，以及逃避血管生成信号 上调。4.1.药理学测定肾脏中mTORC1或同时检测mTORC1和-2的功能 癌细胞和肿瘤相关的内皮细胞。我们将使用Rapalog(Everolimus)或Dual MTORC1/2抑制物(Pp242)及其对新生血管功能和肿瘤细胞分化机制的检测 生物敏感性。4.2.测定肾癌内皮细胞中TORC2的功能。我们会 有条件地删除成人受体内皮中必要的mTORC2组件Rictor，检测 新生血管功能、MRO2和肿瘤细胞存活及增殖信号的正常化。网络的影响 这项拟议的研究将提高肾癌和其他实体恶性肿瘤患者的存活率。

Abstract. The promise of targeting tumor neovascularization remains unrealized. Therefore new insights into how microvessel function impacts tumor biology and how tumor or endothelial signaling pathways regulate neovascularization are necessary. We developed a novel noninvasive imaging technique, photoacoustic microscopy (PAM). PAM uses laser excitation of hemoglobin (Hb) to determine neovascular architecture, Hb concentration (hematocrit), oxygen saturation (SO2), and flow in each tumor microvessel at capillary level resolution without exogenous contrast or tisue window construction. These data uniquely enable microregional determination of tumor metabolic rate of oxygen consumption (MRO2). We will combine PAM with biological, pharmacological, and genetic manipulations to test the hypothesis that tumor neovascular architecture and function regulate, and are regulated by, VEGF and PI3K signaling in tumor or in endothelial cells. We will study renal cancer because it is hypervascular due to overexpression of hypoxia- inducible factors (HIF)-2 and -1 that upregulate VEGF and other angiogenic factors. We will use human 786-O (VHL and PTEN negative) xenografts in immunodeficient mice to interrogate the same vascular network supplied by the same arteriovenous pair in all tumors. We will test our hypothesis with these Specific Aims: 1.0. Develop an integrated label-free photoacoustic microscope that longitudinally images vessel cross- section, hematocrit, SO2, blood flow, and MRO2. Currently we use two PAM instruments to image separately hematocrit (CHb)/SO2 and vessel cross section/flow vessel-by-vessel. Two systems quantifying MRO2 are prone to eror due to repositioning and asynchronicity. 2.0. Determine neovascular function, tumor metabolism, and cell biology during 786-O renal cancer xenograft growth. We will use longitudinal PAM imaging to elucidate how microvessel function, tumor MRO2, tumor and endothelial proliferative, survival, angiogenic, and PI3K signaling pathways are interlaced during tumor growth. 3.0. Inhibit VEGF signaling and determine the functional response of the neovasculature and renal cancer cells. We will use an anti- VEGF antibody, targeting human and mouse VEGF, and test for normalization of each PAM parameter, diminutions in endothelial and tumor cell proliferation and survival, and evasive angiogenic signaling upregulation. 4.1. Pharmacologically determine mTORC1 or both mTORC1 and -2 function in renal carcinoma cels and tumor-associated endothelium. We will use a rapalog (everolimus) or a dual mTORC1/2 inhibitor (PP242) and test for mechanisms of differential neovascular functional and cancer cell biological sensitivity. 4.2. Determine TORC2 function in the endothelial cells of renal carcinomas. We will conditionally delete the necesary mTORC2 component, Rictor, in adult recipient endothelium, testing for normalization of neovascular function, MRO2 and tumor cell survival and proliferative signaling. The impact of this proposed study will be to improve survival of patients with renal cancer and other solid malignancies.