Efficacy of a 10-amino-acid keratin-derived antimicrobial peptide (KAMP-10) against amikacin-resistant Pseudomonas aeruginosa keratitis in mice

Abstract

Purpose : Keratin-derived antimicrobial peptides (KAMPs) are naturally produced by human corneal epithelial cells. Synthetic analogs of KAMPs have potent antibacterial and cytoprotective activity. Recently we reported the novel non-αβ structure of KAMPs and that the central 10-amino-acid hydrophobic strip (KAMP-10) is critical for bacterial membrane permeation. Here, we investigated the efficacy of exogenously administered synthetic KAMP-10 against amikacin-resistant P. aeruginosa keratitis in a mouse model.

Methods : Corneas of anesthetized C57BL/6 mice were scratched and allowed to heal for 4 h before inoculation of 10⁴ CFU (in 5 μl saline) amikacin-resistant P. aeruginosa clinical isolate. KAMP-10 (200 ng) was topically applied once at the time of inoculation as prophylactic treatment. In other experiments, KAMP-10 was subconjunctivally injected (19.2 μg in 8 μl saline, once daily,) and topically applied (5 μg in 5 μl saline, four times daily) for 3 days beginning at 24 h post infection as therapeutic treatment. Vehicle-treated mice served as controls. Cornea pathology was scored daily (0-16 points). At the time of sacrifice, enucleated eyes were homogenized in 1 ml PBS to quantify viable bacteria load. Non-parametric Mann-Whitney U test was used for statistical comparison between two groups.

Results : Control eyes without prophylactic treatment had median [upper, lower quartile] disease scores of 8.5 [10.75, 6.25] and 10.5 [13, 9] at 24 h and 48 h post infection respectively, and bacterial load of 6.6x10⁴ [8.1x10⁴, 3.7x10⁴] CFU/eye. In contrast, eyes treated with KAMP-10 prophylactically were uninfected (P < 0.0005 in each case). Eyes treated therapeutically also had significant reduction in disease scores (2 [3, 2] vs. 8 [8.75, 4.75] at 24 h post treatment) and bacterial load (0 [0, 0] vs. 1.6x10⁴ [2.55x10⁴, 1.05x10⁴] CFU/eye) compared to untreated control (P < 0.001 in each case).

Conclusions : KAMP-10 treatment given prophylactically or therapeutically can effectively prevent or ameliorate P. aeruginosa-induced corneal infection in mice. The data suggest that KAMPs may serve as new treatment options or supplements to curb antibiotic-resistant corneal infection.

This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.