Uncompetitive Phospholipase A2 Inhibition by CHEC Sequences Including Oral Treatment of Experimental Autoimmune Myeloencephalitis

This is the authors' abstract. We don't add key points for this paper.

Catalytic sites of metal dependent phosphatases and phospholipases are characterized by a catalytic histidine (H) and adjacent acidic metal binding residue E (or D). For the secreted phospholipases A2 (sPLA2s), the active sites are stabilized by nearby flanking cystines. CHEC-9 (CHEASAAQC), which displays this organization, is a sPLA2 inhibitor that is an effective treatment for models of traumatic and autoimmune neurodegenerative disorders. We screened modifications of this sequence and found that elimination of the two central alanines followed by cyclization results in another potent sPLA2 inhibitor, CHEC-7 (CHEASQC). CHEC-7, like CHEC-9, had uncompetitive kinetic properties making it ideal for in vivo applications. Subcutaneous injections of CHEC-7 into restrained rats inhibited the transient rise in plasma sPLA2 activity suggesting that this peptide was also effective in vivo. Oral delivery of the peptide attenuated the LPSinduced increases in sPLA2 activity. Thin layer chromatography and mass spectroscopy of representative LPS-treated rats suggested that the peptide also reduced levels of lysophosphatidylcholine, a hydrolytic product of the enzyme reaction. In order to test the efficacy of CHEC-7 for neuroimmune therapies, rats were immunized with spinal cord homogenates to induce experimental autoimmune encephalomyelitis (EAE), followed by daily treatment with oral (1.5 mg/kg) or subcutaneous (0.1, 0.5, and1.5mg/kg) CHEC-7 and CHEC-9 (n=10 for each different condition). The rats were scored for severity of disease applying a standard EAE scale. Control rats all developed disease over the 23-day course of the experiments. Both CHEC-9 and CHEC-7 produced highly significant reductions in disease severity including via oral delivery. The performance of oral CHEC-7 was notable in that this treatment completely prevented disease in half the rats. The results suggested that continued study of modifications of CHEC sequences would provide stable therapeutic peptides for a variety of inflammatory and autoimmune disorders.