Evaluating and enhancing the immune evasion properties of membrane-wrapped nanoparticles

Abstract

Hematopoietic stem and progenitor cells (HSPCs) are crucial for all of the red and white blood cell production. Malfunctions in these cells can lead to severe diseases, particularly monogenic genetic disorders, for which HSPC transplantation is often the only effective treatment. However, these transplants are labor intensive, challenging for patients to endure, and typically require chemotherapy or radiation therapy to eliminate defective HSPCs. Following these treatments, the individual temporarily has no immune system, so they must be contained in controlled environments. There is also a risk that the transplanted cells could recognize the new host as foreign and attack it, which can lead to severe acute and long-term side effects. Consequently, delivering payloads to HSPCs in vivo to repair them in situ is of great interest for a variety of potential applications. Membrane-wrapped nanoparticles (MWNPs) offer an attractive solution as previous research has shown that NPs wrapped with plasma membranes derived from megakaryoblastic CHRF cells can accumulate in bone marrow after intravenous administration to effectively interact with HSPCs. However, a large portion of each dose is sequestered in the liver and spleen, reducing the potential efficacy of any treatment based on this technology. This off-target delivery may be due to the protein corona (PC), which is the buildup of serum proteins on the surface of MWNPs after intravenous injection, leading to their clearance. This dissertation characterizes the impact of the PC on target cell binding and macrophage clearance of unwrapped and wrapped NPs in vitro (Chapter 3) and in vivo (Chapter 4), then develops a strategy to minimize PC impact on MWNPs (Chapter 5). ☐ This dissertation produced several key findings. In the in vitro studies of Chapter 3, MWNPs with a PC exhibited increased target cell uptake and decreased macrophage uptake, whereas unwrapped NPs with a PC exhibited reduced target cell uptake and increased macrophage uptake. This demonstrates that cell membrane wrapping can beneficially influence cellular interactions even in the presence of a PC. Proteomics identified apolipoprotein B and complement component 3 (C3) as the dominant opsonins on both MWNPs and unwrapped NPs after incubation in mouse, bovine, or human serum, with other proteins like immunoglobulins, complement proteins, and apolipoproteins also present. In Chapter 4, in vivo biodistribution studies in wild-type and knockout mice (ApoE-/-, C3-/-, and FcRn-/-) indicated a complex interplay between specific PC components and targeting versus clearance. Specifically, C3 and immunoglobulin G adsorption on MWNPs led to macrophage clearance but also enabled HSPC targeting. Meanwhile, apolipoprotein E adsorption facilitated hepatocyte clearance but reduced immune cell uptake in the liver. In Chapter 5, to reduce C3-mediated clearance, we synthesized MWNPs using cells that were genetically engineered to express CD55, which interferes with the complement cascade. After serum incubation, these CD55-MWNPs exhibited reduced C3 convertase activity compared to unedited MWNPs. Collectively, this research indicates that MWNPs are a promising, next-generation NP platform that can be modified to improve targeting to HSPCs by reducing the effects of clearance mechanisms. In Chapter 6, conclusions and perspectives on future directions for this work are presented.