CAREER: Multi-Gradient Microtissue Arrays to Analyze Patient-Derived Cancer Cell Modulation by Combinatorial Electrical and Chemical Stimulation

提案#:1652112癌症是美国第二大死亡原因，其中一些结果最差的是高度浸润性脑肿瘤胶质母细胞瘤（GBMs），即使采用手术、放疗和化疗（药物）联合治疗，其5年生存率也不到10%。手术和药物不能消除浸润到中央肿块以外几厘米的恶性细胞，放射疗法对健康组织和恶性组织的损害是一样的。该项目建立在GBM组织工程模型的初步研究基础上，该模型表明，脉冲电场（pef）通过靶向肿瘤细胞改变的电学特性，如表面电荷、膜电容和离子通道表达的变化，诱导肿瘤细胞的选择性破坏。此外，这种针对肿瘤相对不受正常细胞影响的物理特征的效果，通过添加针对肿瘤细胞化学特性改变的药物治疗而增强。该项目的目标是开发一个平台，以有效筛选物理/药物联合治疗对患者来源细胞的影响。这个新颖的平台将扩大对联合方法有益效果的理解，并在新的精准医学范式中巩固基础科学和临床重要进展。教育和推广计划包括：1)通过新的和更新的课程，为研究生和本科生提供研究培训机会，并支持弗吉尼亚州西南部经济困难学生的K-12活动，扩大弗吉尼亚理工大学的“癌症工程”课程，拓展和多样性；2)建立“询问弗吉尼亚理工大学科学家”网络专栏，该专栏将涵盖科学，技术，工程和数学（STEM）领域的广泛主题；当地的K-12学生和教师以及社区参与。有效的精准癌症医学方案最终将取决于多靶向治疗，以治疗高度异质性和适应性强的肿瘤，如GBM。该项目开发的平台将是第一个在正交方向上覆盖多种动态治疗梯度（例如电场振幅和药物浓度）的组织阵列微芯片。该平台将兼容细胞动力学的高分辨率实时测量，并与强大的微流体分子分析相结合，以表征受刺激细胞的表观基因组变化，独特地实现患者来源细胞的组合电/分子治疗的广谱参数分析。该研究计划有三个目标：1)开发与高分辨率共聚焦成像和阻抗测量兼容的多维电/化学梯度微组织阵列芯片；2)开发与微流控染色质免疫沉淀（chip）耦合的高分辨率表观基因组分析；3)对患者GBM细胞进行组合电/化学治疗的广谱参数研究。成功完成这些目标对于以下方面至关重要：1)揭示pef与药物联合可能产生的治疗协同效应，2)以最少的材料（如细胞、试剂）优化组合治疗，以及3)测试与治疗协同效应相关的分子假设。收集的数据将为浸润性肿瘤新的有效治疗策略提供基础科学见解。针对致密岩心和弥漫性浸润区的物理和化学结合方法将有可能改变GBM治疗的模式。

PI: Verbridge, Scott S.Proposal #: 1652112Cancer is the second leading cause of death in the United States, with some of the poorest results being for highly infiltrative brain tumor glioblastomas (GBMs), for which the 5-year survival rate is less than 10% even with combination treatments of surgery, radiation and chemotherapy (drugs). Surgery and drugs fail to eliminate the malignant cells that infiltrate centimeters beyond the bulk central mass and radiotherapies damage healthy and malignant tissues alike. This project builds on preliminary studies in tissue-engineered models of GBM that demonstrate that pulsed electric fields (PEFs) induce selective destruction of tumor cells by targeting their altered electrical properties, e.g., changes in surface charge, membrane capacitance and ion channel expression. Furthermore, this effect, which targets a physical hallmark of the tumor leaving normal cells relatively unaffected, is enhanced by the addition of drug therapies, which target alterations in tumor cell chemical properties. The goal of this project is to develop a platform to enable efficient screening of the effects of combined physical/drug treatments on patient-derived cells. This novel platform will expand understanding of the beneficial effects of the combined approach and underpin basic science and clinically important advances in a new precision medicine paradigm. The education and outreach plan includes: 1) expanding the "Cancer Engineering" curriculum, outreach, and diversity at Virginia Tech through new and updated courses, research training opportunities for graduate and undergraduate students and supporting K-12 activities for economically disadvantaged students in Southwest Virginia and 2) establishing an "Ask a Virginia Tech Scientist" web column, which will cover a wide spectrum of topics in science, technology, engineering and math (STEM), for local K-12 students and teachers and community engagement. Effective precision cancer medicine protocols will ultimately depend on multi-targeted therapies to treat highly heterogeneous and adaptable tumors such as GBM. The platform developed in this project will be the very first tissue array microchip with an overlay of multiple types of dynamic therapeutic gradients (e.g. electric field amplitude and drug concentration) in orthogonal directions. The platform will be compatible with high-resolution live measurement of cellular dynamics, and coupled to powerful microfluidic molecular analysis to characterize the epigenomic alterations in stimulated cells, uniquely enabling broad-spectrum parametric analysis of combinatorial electrical/molecular treatments of patient-derived cells. The research plan is organized under three objectives: 1) develop a multi-dimensional electrical/chemical gradient microtissue array chip compatible with high-resolution confocal imaging and impedance measurement, 2) develop high-resolution epigenomic analysis with novel coupling to microfluidic chromatin-immunoprecipitation (ChIP), and 3) execute broad-spectrum parametric studies of combinatorial electrical/chemical treatments using patient GBM cells. Successful completion of these objectives will be crucial to: 1) reveal therapeutic synergies possible through a combination of PEFs and drugs, 2) optimize combinatorial treatments with minimal input material (e.g. cells, reagents), and 3) test molecular hypotheses related to treatment synergies. The data collected will provide basic science insights for a new and effective treatment strategy for infiltrative tumors. The combined physical and chemical approaches to targeting both the dense core, as well as the diffuse infiltrative zones will potentially altering the paradigm of GBM treatment.