The antibacterial and cytotoxic activity of the human cathelicidin peptide LL-37 is inhibited by plasma. Because LL-37 does not undergo rapid degradation in human plasma, we postulated that this inhibition results from binding of LL-37 to unidentified proteins. An LL-37 binding plasma protein has now been isolated by affinity chromatography. SDS-polyacrylamide gel electrophoresis of proteins that bound to an LL-37 column revealed one band with a molecular mass of about 26 kDa, and amino acid sequence analysis identified the protein as apolipoprotein A-I (apoA-I). Biomolecular interaction analysis using surface plasmon resonance showed that LL-37 and isolated apoA-I bind with an apparent K d in the low micromolar range. 50 M of apoA-I inhibits the antibacterial activity of 50 M LL-37 by about 50% of the inhibition exhibited by plasma. In addition, anti-apoA-I IgG completely blocks the plasma inhibition of LL-37 antibacterial activity up to a peptide concentration of 25 M and blocks most of the plasma inhibition at higher LL-37 concentrations. These results indicate that apoA-I is the main LL-37 binding protein in human plasma and may work as a scavenger of LL-37, thus suggesting a novel mechanism involved in the regulation of a cathelicidin peptide.

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Aberrant signal transduction by mutant or overexpressed protein kinases has emerged as a promising target for treatment of acute myeloid leukemia (AML). We here present a novel low

Aberrantly expressed tyrosine kinases have emerged as promising targets for drug development in acute myeloid leukemia (AML). We report that AKN-028, a novel tyrosine kinase inhibitor (TKI), is a potent FMS-like receptor tyrosine kinase 3 (FLT3) inhibitor (IC50=6 nℳ), causing dose-dependent inhibition of FLT3 autophosphorylation. Inhibition of KIT autophosphorylation was shown in a human megakaryoblastic leukemia cell line overexpressing KIT. In a panel of 17 cell lines, AKN-028 showed cytotoxic activity in all five AML cell lines included. AKN-028 triggered apoptosis in MV4-11 by activation of caspase 3. In primary AML samples (n=15), AKN-028 induced a clear dose-dependent cytotoxic response (mean IC50 1 μℳ). However, no correlation between antileukemic activity and FLT3 mutation status, or to the quantitative expression of FLT3, was observed. Combination studies showed synergistic activity when cytarabine or daunorubicin was added simultaneously or 24 h before AKN-028. In mice, AKN-028 demonstrated high oral bioavailability and antileukemic effect in primary AML and MV4-11 cells, with no major toxicity observed in the experiment. In conclusion, AKN-028 is a novel TKI with significant preclinical antileukemic activity in AML. Possible sequence-dependent synergy with standard AML drugs and good oral bioavailability has made it a candidate drug for clinical trials (ongoing).