Cancer therapy by targeting innate immune cells in vivo using novel virus-like particles - Project Summary: What if we could engineer our own cells to become potent cancer fighters? Here, we will utilize a novel targeted gene therapy approach to specifically engineer distinct immune cell populations (T cells, NK cells and macrophages) to express chimeric antigen receptors (CARs) to enable these immune cells to mediate killing of otherwise refractory cancers. This approach builds off the remarkable success in use of autologous CAR- expressing T cells to better treat and cure B cell malignancies. However, the autologous CAR-T cell strategy requires the cells to be manufactured on a patient-specific basis, making this approach inefficient, expensive, time consuming, and still prone to disease relapse. CAR-expressing NK cells and allogeneic CAR-T cells to provide an off-the-shelf strategy that broadens the range of patients who may be treated and makes this process more efficient, standardized, and affordable. However, there are also concerns about immune responses against the allogeneic cells, and the efficacy of this approach against more common solid tumor malignancies remains unclear. Here, we will engineer endogenous immune cells using a novel virus-like particle (VLP) vector that combines the best aspects of the ease and targeting of lentiviral vectors with use of mRNA expression that is more controlled and regulated compared to genome-integrating lentivirus. Cell lineage-specific DARPins or scFvs expressed by the VLPs will be utilized to separately target T cells, NK cells or macrophages. The immune cells will be engineered to express an anti-mesothelin CAR with signaling domains optimized for either T cells, NK cells or macrophages. We hypothesize these engineered therapeutic vehicles capable of performing guided in vivo cellular engineering of specific immune cells will enable endogenous immune cells to now mediate potent killing of ovarian cancer cells. This approach will first be tested in vitro to determine the efficiency and specificity of DARPin-mediated gene expression in T cell, NK cell and macrophage cell lines and then primary immune cells to express CARs that lead to recognition and killing of ovarian cancer cells. Additional transcriptomic and metabolomic profiling will be done of the VLP- engineered immune cells. Next, we will evaluate and optimize this approach using two different in vivo models. The first in vivo studies will use tumor-bearing immune competent mice that will allow us to test both specificity and efficacy of this approach, as well as safety, persistence, and ability to stimulate a broader immune response against ovarian cancer. Subsequent in vivo studies will use immunodeficient mice engrafted with human immune cells to provide a relevant pre-clinical model of engineering endogenous immune cells to effectively target and kill ovarian cancer cells. Our strategy to not only target T cells, but to separately test efficacy of in vivo engineering of innate immune effector cells (NK cells and macrophages) will enable us to efficiently test a diversity of options for this in vivo approach to ensure success in subsequent clinical translation to better treat and cure otherwise lethal malignancies.
