Enabling targeted cargo delivery to hematologic cells with biomimetic membrane-wrapped nanoparticles
Date
2022
Authors
Journal Title
Journal ISSN
Volume Title
Publisher
University of Delaware
Abstract
An estimated 1.2 million cases of blood cancer occur each year, an increase of 40% over the past decade.1–3 Beyond these cancers, hematologic disorders affect millions more. The common factor linking all these diseases is the dysfunctionality of hematopoietic stem and progenitor cells (HSPCs). HSPCs, which reside in the bone marrow (BM), are the cells responsible for the differentiation of all blood cell types including red blood cells, white blood cells, megakaryocytes (Mks), and platelets. Therefore, the ability to provide targeted cargo delivery to HSPCs is a challenging problem whose solution could aid in the treatment of many diseases. For many of these, the only curative treatment is a stem cell transplant (SCT), where patients receive high-dose chemotherapy or radiation to remove diseased HSPCs and are then infused with healthy HSPCs to repopulate the BM. While transplant results are promising, they carry risk of infection, graft rejection, and secondary cancers developing. This work proposes the use of an alternative technology: cell membrane-wrapped, biomimetic nanoparticles (NPs) that will provide targeted delivery of therapeutic cargo to HSPCs in vitro and in vivo. ☐ Biomimicry has recently emerged as an exciting new era of investigation in nanomedicine and the development of membrane-wrapped nanoparticles (MWNPs) is at the forefront of this field. Historically in NP development, researchers have added poly(ethylene glycol) (PEG) or targeting moieties to NPs. While these strategies can extend circulation time and provide site-specific accumulation, new studies have shown that the body can develop an anti-PEG immune response and that NPs with active targeting agents may still have off-target effects. This has led researchers to begin designing NPs to “mimic” cells within the body by wrapping the NPs in isolated cell membranes. This effectively hides them from the immune system, prolonging their circulation time and enhancing cargo delivery. ☐ Mk microparticles are naturally-occurring transport systems that bud off Mk cells to deliver cargo to HSPCs. Since their membranes provide innate homing to HSPCs, they are ideal to use as a cloaking system for delivery of nanotherapeutics to HSPCs. This dissertation develops an Mk-based targeting system, examines the in vitro and in vivo capabilities, and establishes the potential of the system through proof-of-concept cargo delivery. ☐ First, methods were developed to wrap poly(lactic-co-glycolic) acid NPs in Mk membranes, creating MWNPs. Membrane wrapping was validated through several particle characterization techniques, including nanoparticle tracking analysis and transmission electron microscopy. Next, HSPCs and control cell lines were dosed with MWNPs to examine their preferential targeting abilities. MWNP uptake was observed to occur more rapidly in HSPCs than other cell lines, indicating the membrane functionality remained. Taking advantage of this targeting mechanism, small interfering RNA was loaded into the particles, delivered to HSPCs, and measured to reduce gene expression by 15.6%. After proving the in vitro potential of the system, MWNPs were intravenously-administered into mice to assess their in vivo targeting abilities. Through biodistribution studies and cellular analysis, it was confirmed that they were able to reach HSPCs in mice at levels more than double their PEG-coated counterparts. ☐ Based on these promising data, a second set of studies focused on using MWNPs for the delivery of doxorubicin (DOX), a chemotherapeutic traditionally used in preparation for a SCT. Clinical advances have been made to decrease the systemic side effects of chemotherapies, including the use of liposome-encapsulated drugs. However, these treatments lack targeting abilities, making DOX-loaded MWNPs an ideal candidate to provide specific targeting of chemotherapeutics and improve the patient experience. Here, experiments assessed the ability of MWNPs to encapsulate and release DOX in both storage and physiologically-relevant conditions, as well as induce apoptotic cell death in a manner that retained the mechanistic functionality of the DOX molecule. Notably, MWNPs successfully induced up to 80% cell death, compared to the rates of 12-17% seen in the control systems. This research establishes the promising capabilities of drug-loaded MWNPs that can be used for future in vivo HSPC-targeting experiments. ☐ The final set of studies tested the abilities of a biomanufactured, Mk-derived extracellular vesicle (MkEV) to reach HSPCs in vivo. This natural carrier system has been genetically developed to be the perfect cell messenger system, which may be advantageous over the semi-synthetic MWNPs. As with the previous biodistribution studies, systemically-injected MkEVs localized to the BM, specifically reaching the most primitive hematopoietic stem cells. This system was tested at two timepoints, informing the development of future treatment plans using cargo-loaded MkEVs to treat a range of BM disorders. ☐ In conclusion, this work establishes a new therapeutic pathway for targeted treatment of hematologic cancers and related diseases. The efficacy of MWNPs and MkEVs has been demonstrated through in vitro studies of gene regulation and chemotherapeutic delivery as well as in vivo biodistribution experiments examining the specific cells reached by the particles. Given the promising data presented in this work, the development of this novel technology could have a profound and lasting impact on the HSPC field, MWNP advancement, and the treatment options available to patients.
Description
Keywords
Biomimicry, Blood cancer, Chemotherapy, Extracellular vesicles, Bone marrow