Human Tissue Culture Bioreactor and Hyperpolarized MR for Biomarker Discovery

项目摘要 通过这个独立之路奖，我希望获得必要的技能，以获得教师 一个独立的研究项目，专注于生物工程和实施新的3D 细胞和组织培养生物反应器，并将该平台与超极化（HP）13 C MR结合使用， 更好地研究癌症代谢。由于前列腺癌的生物学和病理学复杂性， 迫切的临床需要开发更敏感和特异性的成像标记物，用于改善前列腺癌 患者特异性治疗计划和治疗失败的早期评估。一项非凡的新技术 利用超极化（HP）代谢底物具有提供这些MR生物标志物的潜力。最近的HP 在细胞和动物模型中的MR研究表明，HP代谢标志物反映了酶通量， 提供更准确的测量前列腺癌的存在、进展和对治疗的反应。然而，在这方面， 现有的小鼠和细胞培养模型不能可靠地模拟人类疾病，因此我们提出了一种新的 HP 13 C MR和NMR兼容的3D组织培养生物反应器的组合，以研究实时 活的人前列腺组织切片（TSC）的代谢。 本研究的总体目标是设计一种NMR兼容的3D组织培养生物反应器， 与人TSC一起使用，并将其用于鉴定用于改善前列腺癌患者的HP分子成像标志物- 具体的治疗计划和对靶向治疗反应的早期评估。实现这些目标将 需要在原代细胞和组织培养，前列腺生物化学和病理学领域进行额外的培训， HP探针开发、微工程、生物转运和药代动力学。利用这种新的训练， 第一个目的是优化在NMR兼容的3D组织培养物中维持人前列腺TSCs的条件 生物反应器中，并验证随着时间的推移TSC的代谢完整性。连续31 P将用于监测 组织切片在生物反应器中随时间的进展。HP 13 C MR的动态采集将用于 真实的计算与丙酮酸和其它探针的代谢相关的通量。这些数据将被比较 在生物反应器中培养前后进行组织病理学检查，以评估变化。第二个目标是利用这一新的 比较正常和恶性前列腺组织代谢的实验模型，重要的是，确定 HP代谢产物是否与病理分级相关以及它们与代谢和生物转运的关系。的 第三个目的是使用该平台来鉴定对PI 3 K/mTOR抑制剂的治疗应答的HP标志物。 该提案的目标是开发一种工程系统，该系统可以克服当前的限制。 小鼠和细胞培养物模型，并有助于开发相关生物标志物用于临床。 虽然这个独立之路奖的研究重点是前列腺癌， NMR兼容的原代组织培养生物反应器平台结合高灵敏度HP MR探针 将在各种疾病和成像模式中具有广泛的适用性。

Project Summary Through this Pathway to Independence Award, I hope to acquire the skills necessary to obtain a faculty position with an independent research program focused on the bioengineering and implementation of novel 3D cell and tissue culture bioreactors, and the use this platform in conjunction with hyperpolarized (HP) 13C MR to better study cancer metabolism. Due to the biologic and pathologic complexity of prostate cancer, there is an urgent clinical need to develop more sensitive and specific imaging markers for improved prostate cancer patient-specific treatment planning and early assessment of therapeutic failure. An extraordinary new technique utilizing hyperpolarized (HP) metabolic substrates has the potential to provide these MR biomarkers. Recent HP MR studies in cell and animal models suggest that HP metabolic markers reflect enzymatic fluxes and may provide a more accurate measure of prostate cancer presence, progression and response to therapy. However, available murine and cell culture models don't reliably mimic human disease, thus we propose a novel combination of HP 13C MR and NMR-compatible 3D tissue culture bioreactors to study the real-time metabolism of living human prostate tissue slices (TSCs). The overall objective of this research are to engineer an NMR-compatible, 3D Tissue Culture Bioreactor for use with human TSCs and use it to identify HP molecular imaging markers for improved prostate cancer patient- specific treatment planning and early assessment of response to targeted therapy. Accomplishing these aims will require additional training in the areas of primary cell and tissue cultures, prostate biochemistry and pathology, HP probe development, micro-engineering, biotransport, and pharmacokinetics. Utilizing this new training, the first aim is to optimize conditions for maintaining human prostate TSCs in an NMR-compatible, 3D tissue culture bioreactor and to verify the metabolic integrity of TSCs over time. Continuous 31P will be used to monitor the progression of tissue slices in the bioreactor with time. Dynamic acquisitions of HP 13C MR will be used to calculate fluxes associated with metabolism of pyruvate and other probes in real time. This data will be compared to histopathology before and after culture in the bioreactor to assess changes. The second aim is to use this new experimental model to compare normal and malignant prostate tissues metabolism, and importantly, determine whether HP metabolites correlate with pathologic grade and their relationship to metabolism and biotransport. The third aim is to use this platform to identify HP markers of therapeutic response to PI3K/mTOR inhibitors. It is the goal of this proposal to develop an engineered system, which can overcome the limitations of current murine and cell cultures models and aid in the development of relevant biomarkers for translation to the clinic. While the focus of the research in this Pathway to Independence Award is on prostate cancer, the combination of NMR-compatible primary tissue culture bioreactor platform combined with high sensitivity HP MR probes would have wide applicability across a variety of diseases and imaging modalities.