Engineer Biomimetic Microfluidic Models to Investigate and Reprogram Tumor Associated Neutrophils for Cancer Therapy

Project Summary Neutrophils play critical roles during different stages of tumor development. In mice, systemic depletion of neutrophils results in decreased tumor growth in glioblastoma (GBM) and lung cancer, but promote tumor growth in pre-metastatic lung and other solid tumors, indicating their stage-specific and tissue-dependent functions in tumor progression. Neutrophils could also facilitate cancer cell resistance to chemotherapy, radiotherapy, and immunotherapy in different tumors by releasing various cytokines. Despite these preclinical and animal studies on tumor-associated neutrophils (TANs), a knowledge gap remains in our mechanistic understanding of how human neutrophils regulate cancer progression and therapeutic resistance in GBM, due to the short life and resistance to gene editing of neutrophils as well as technical hurdles in isolating stage-specific TANs. To address this gap, we propose to harness the power of microfluidics and human induced pluripotent stem cells (hiPSCs) to interrogate the diversity and plasticity of neutrophils in human GBM development. Elucidating the underlying mechanism will also enable the much-needed development and evaluation of neutrophil-targeted cancer therapy. Our central hypothesis here is that the microfluidic model will recapitulate the different stages of human tumor progression, providing a platform for phenotypic and mechanistic understanding of the roles of neutrophils in GBM development. To test this hypothesis, we will implement a novel interstitial tumor-microenvironment-on- chip (iT-MOC), and interrogate neutrophil-mediated tumor progression and therapeutic resistance at different GBM growth stages in Aim 1. Then in Aim 2, we will determine the morphology, polarization, life-span and antitumor cytotoxicity of GBM-infiltrating neutrophils. In Aim 3, we will reprogram tumor-associated neutrophils towards antitumor effector cells via genetic engineering of hiPSCs with chimeric antigen receptors (CARs) and microRNAs (miRNAs). This is a novel approach as human neutrophils cannot be genetically modified. Successful completion of these aims will offer an innovative platform to study the diversity and plasticity of TANs, and provide insights into reprograming them towards antitumor effector cells and the proof-of-concept for CAR-neutrophils in targeted cancer therapy.