(J-361) Lysyl Oxidase Responsive Polymers for Targeting Myocardial Infarction: Biomarker discovery & material design

Introduction:: More than 800,000 Americans are impacted by acute myocardial infarction (MI) yearly, with a 23% mortality rate, and two-thirds of surviving individuals never fully recover due to negative left ventricular (LV) remodeling. This process is a product of chronic inflammation of the infarct and surrounding tissue, which occurs in the following days to months post-MI. Novel gel-based therapeutic scaffolds have been shown to reduce negative LV remodeling but are difficult to administer to the infarct due to gelation kinetics. Enzyme-responsive nanoparticles have been shown to target the infarct post-intravenous injection at acute time points. However, due to their size, nanoparticles suffer from liver and spleen accumulation, dominating biodistribution patterns. Herein, we describe a class of polymer with tunable molecular weights (5-65 kD), tunable mechanical properties, and discrete molecular structures akin to small globular proteins, resulting in multi-pass pharmacokinetics. Due to the modularity of the approach, these polymers can be designed to respond to enzymes associated with the inflammatory response within the MI. Lysyl oxidase (LOX) is an extracellular matrix (ECM) enzyme that plays a critical role in crosslinking and remodeling the ECM under inflammatory conditions. Through this work, we have demonstrated proof of concept that the development of a LOX responsive polymer results in systemic targeting to the infarct. Through further investigation, we aim to optimize this platform to reduce inflammation in the infarct through therapeutic delivery and biomaterial properties, as well as optimize clearance via the renal system.


Materials and Methods:: Responsive polymers were synthesized via ring opening metastasis polymerization (ROMP). Polymerization kinetics were monitored via ¹H-proton NMR. Mass, size, and diffusion properties were analyzed by SDS PAGE, DLS, and DOSY NMR. In vivo rat studies using an ischemia/reperfusion (I/R) model were carried out. LOX activity and expression during timepoints relevant to acute MI was validated through qPCR and LOX activity assay. To demonstrate targeted localization upon systemic administration, polymers were delivered via tail vein injection and after two days, the heart and satellite organs were harvested for analysis. Systemic and targeted distribution of LOX-responsive polymers was analyzed via fluorescence and confocal microscopy and LICOR infrared imaging.


Results, Conclusions, and Discussions:: Our work has shown that LOX upregulation occurs within 1 day post-MI in rat I/R models (p < 0.01). These results demonstrate for that LOX is a unique and targetable enzymatic biomarker for acute phases of MI. Through fluorescent and infrared imaging, we have demonstrated that LOX-responsive polymers localize to the infarcted myocardium, with nearly negligible trace in the remote myocardium, as well as the kidneys upon systemic administration. These results suggest that LOX responsive polymers accumulate in the inflammatory microenvironment of the infarct through LOX-mediated extracellular crosslinking mechanisms to form a localized scaffold and that bias towards renal clearance. This platform would not only provide targeting to deliver anti-inflammatory therapeutics to the LV via scaffold assembly, but will also allow cellular infiltration into the necrotic tissue, providing opportunities for cellular reprogramming via cell-matrix interactions. Upon further investigation, we aim to optimize scaffold activation and clearance kinetics, as well as develop a therapeutic analog to elucidate an anti-inflammatory response, effectively reducing the progression of negative LV remodeling.


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