Optimization of Renal Delivery of an MMP2‐inhibitory Peptide Delivered Using the Elastin‐like Polypeptide Carrier

Abstract

Chronic kidney disease (CKD) is a global health issue associated with the development of renal fibrosis. Renal fibrosis is caused by the excessive deposition of extracellular matrix (ECM) leading to organ failure and is the final common pathway in CKD. ECM homeostasis and degradation are mediated by matrix metalloproteinases (MMPs). Imbalances in MMP activity exacerbate renal damage and fibrosis. While increased expression of some MMPs can be anti‐fibrotic, other MMPs, such as MMP2, activate pro‐inflammatory and fibrotic chemokines and cytokines. Our group has developed an MMP2 inhibitory biologic by fusing an MMP2 inhibitory peptide to a protein biopolymer (ELP‐MMP2i). We hypothesize that ELP‐MMP2i will accumulate in the kidney after systemic administration and reduce or reverse the effects of renal fibrosis. ELP‐MMP2i has potent MMP2 inhibitory activity in vitro (IC50 = 1.1 mM). A pilot study utilizing continuous infusion of ELP‐MMP2i demonstrated reduced proteinuria and albuminuria in a hypertension induced glomerulosclerosis rat model (Dahl Salt Sensitive Rat). The aim of the current study was to optimize the renal delivery of ELP‐MMP2i. Three ELP‐MMP2i proteins using ELP domains of varying molecular weights (26 kDa, 50 kDa, and 99 kDa) were constructed. Pharmacokinetics and biodistribution of these proteins were determined following iv or sc injections (600 nmol/kg) in Sprague Dawley Rats (n= 4/group). The plasma clearance following an iv injection was inversely proportional to the molecular weight, and half‐lives were 0.4, 0.6, and 1.1 hours for the 26, 50, and 99 kDa proteins respectively (Figure A3). Similar results were observed with sc administration, with the smallest protein exhibiting the fastest absorption and the highest peak plasma levels (400 nM, 240 nM, and 180 nM peak plasma, respectively; Figure B2). Bioavailability was 55.3, 25.1, and 16.7% for 26, 50, and 99 kDa proteins respectively following sc administration. Both iv and sc injections exhibited significant deposition in the kidney (Figures A1 & B1), and an inverse correlation between protein size and renal tissue concentrations of 6.2 ± 1.9, 3.6 ± 0.2, and 2.5 ± 0.4 mM renal concentrations for the 26, 50, and 99 kDa proteins, respectively, following iv administration (two‐way ANOVA p<0.0001 Figure A2), and 2.0 ± 0.7, 0.6 ± 0.1, and 0.5 ± 0.1 mM renal concentrations following sc injections (two‐way ANOVA p<0.0001, Figure B2). To further elucidate the pharmacokinetic properties of the protein in a model with renal injury and to test the effects of various doses, Dahl Salt Sensitive rats on an 8% NaCl diet were given a daily sc injections of 300 nmol/kg, 600 nmol/kg or 1,200 nmol/kg of the smallest ELP‐MMP2i peptide (n=3/group) for 7 days. ELP‐MMP2i accumulated almost exclusively in the kidneys with renal concentrations of 7.1 ± 0.3, 12.9 ± 1.2, and 25.0 ± 2.8 mM (p<0.0001, two‐way ANOVA) following doses of 300, 600, and 1,200 nmol/kg/day respectively). These studies revealed that the smallest 26 kDa ELP‐MMP2i protein achieved the best balance of bioavailability and renal deposition. The results indicate that therapeutic doses of ELP‐MMP2i above the IC50can be achieved by daily sc administration.