Identification and potency of cyclodextrin-lipid inhibitors of Staphylococcus aureus α-toxin.

Purpose: Staphylococcus aureus, a leading cause of bacterial keratitis, secretes α-toxin, a cytotoxin active on the corneal epithelium. This study describes the production and testing of chemical inhibitors of α-toxin action.

Methods: Purified α-toxin was titered by its ability to lyse rabbit erythrocytes in buffered saline (PBS). To prepare potential toxin inhibitors, each of 18 lipids was incorporated into a complex with methyl-β-cyclodextrin (MβCD) or hydroxypropyl-β-cyclodextrin (HPβCD). Serial dilutions of each lipid-cyclodextrin (CD-lipid) complex were mixed with α-toxin prior to the addition of rabbit erythrocytes. Select CD-lipid complexes were mixed with 12 hemolytic units (HU) α-toxin and injected into the rabbit corneal stroma so the resulting corneal erosions could be measured at 4 and 8 hours post-injection (PI). Eyes injected with toxin alone, MβCD, or HPβCD alone served as controls.

Results: Neither form of CD alone inhibited α-toxin. Of the 36 complexes prepared, 6 lipid-CD complexes were found to inhibit >100 HU of α-toxin. Four lipid complexes able to inhibit >200 HU of α-toxin were tested in toxin-injected corneas; at 4 and 8 hours PI, the complexes of cholesterol or lanosterol with MβCD and squalene or desmosterol with HPβCD caused a significant reduction in the corneal erosion size as compared to eyes injected with α-toxin alone (P ≤ 0.05).

Conclusions: Specific lipid inclusion complexes with either MβCD or HPβCD demonstrated a significant inhibition of α-toxin in both in vitro and in vivo assays. Changes in either the cyclodextrin or lipid of a complex affected the inhibitory activity.