Nanodelivery of microRNA Based Therapeutics to Macrophages
Creators: name: Nally, Frances
Publisher: name: Royal College of Surgeons in Ireland
Year: 2024
Subjects: classificationCode: 320499
subject: Immunology not elsewhere classified
classificationCode: 320604
subject: Nanomedicine
classificationCode: 320601
subject: Gene and molecular therapy
classificationCode: 310803
subject: Plant cell and molecular biology
classificationCode: 310902
subject: Animal cell and molecular biology

description: Macrophages are key effector cells with a multifaceted role in the innate immune response, capable of both potentiating inflammation, an ‘M1’ like response, or suppressing it and promoting tissue repair in an ‘M2’ like state. Manipulation of this polarisation capacity has the potential to be clinically useful in diseases where inflammatory macrophages are key drivers of pathogenicity, including but not limited to Sepsis, Multiple Sclerosis, and Rheumatoid Arthritis. MicroRNAs have become increasingly established as key regulators of cellular processes, including pathways that drive inflammation. Of particular interest is miR-155, whose expression is high in the ‘M1’ and supressed in an IL-10 driven ‘M2’ phenotype. Furthermore, previous work in the group has identified miR-155 target Arginase-2 (Arg2) as a key effector of IL-10 driven pro-resolution pathways.
Here we investigate the therapeutic potential of direct miR-155 inhibition using an anti-miRNA oligonucleotide (AMO-155), as well as the impact of disrupting the miR-155-Arg2 axis using a Target Site Blocker (TSB-Arg2) on limiting the macrophage inflammatory response. We show that while both AMO-155 and TSB-Arg2 demonstrate a similar Arg2 boosting capacity <em>in vitro</em>, TSB-Arg2 demonstrates a superior capacity to modulation macrophage phenotype as determined by cell surface marker expression and metabolic profiling. In keeping with this, TSB-Arg2 demonstrates a superior inflammation limiting capacity in an <em>in vivo</em> model of acute inflammation, thus for the first time comparing AMO vs TSB based strategies and outlining the therapeutic potential of TSB mediated miRNA manipulation.
While nucleic acid therapeutics are of rapidly increasing interest, successful intracellular delivery remains a key challenge to their success. Bioinspired star polymers with Poly-L-Lysine arms (Star-PLLs) designed in RCSI represent a promising delivery vector, and were herein investigated for their ability to deliver small single stranded nucleic acids to macrophages. We demonstrate that previous lead vector G5-Star-PLL forms polyplexes with AMO-155 and TSB-Arg2, which were well tolerated and demonstrate functional delivery of therapeutic cargo to bone marrow derived macrophages (BMDM) <em>in vitro</em>. Moreover, cell specific targeting of nanomedicines holds promise in increasing desired cargo effectiveness. This concept is explored in this thesis via surface modification of G5-Star-PLL with a CD206 targeted mannose ligand. Ultimately, mannosylated G5 (mG5-TSB-Arg2) polyplexes demonstrate increased intracellular delivery capacity of therapeutic cargo in BMDM over their unmodified counterparts, but do not mediate macrophage specific enhancement of uptake.
While objectives of macrophage specific uptake enhancement were not met by mannose conjugation of Star-PLL G5, results from this study highlight the importance of the physicochemical properties of nanoparticles in determining nanoparticle uptake and intracellular cargo release and provides insight that may inform further development of Star-PLLs. Collectively, this thesis describes a novel potential therapeutic, TSB-Arg2, and presents Star-PLLs as an option for the delivery of single stranded miRNA related cargoes to facilitate their clinical translation.