Primate Monocytes - CD14, CD16 - Ziegler-Heitbrock

Effect of formyl peptide-toxin conjugates on myeloid cancer cell lines in vitro

Abstract

We examined whether the receptor (R) for the chemotactic formyl peptide (fMLP) on monocytic cells (U937 and Mono Mac 6) could be exploited to deliver cytotoxic drugs into cells. A dose-response for binding and uptake of fML [H-3]P in unstimulated and stimulated U937 cells was determined. The toxin melphalan or the A chain of ricinus communis was cross-linked to fMLPK and fMLPKK, respectively U937 or Mono Mac 6 cells were cultured overnight with either the conjugate, the toxin or fMLPK/fMLPKK and pulsed with H-3-leucine. Cell-associated radioactivity was acid-precipitated measured in a beta-counter and calculated as a percentage of H-3-leucine uptake by the control cells. incorporation of 3H-leucine by U937 cells treated with fMLPKK-ricin A feel sharply at the nM level, which is approximately the ED(50) for the ligand (1). Ricin A alone was 10-100-fold less effective than the conjugate, while fMLPKK alone had no effect. FMLPK-melphalan conjugates at 10(-7) M had only a little stronger, but at 10(-6) M statistically significant, inhibitory effect on H-3-leucine incorporation in U937 cells than melphalan alone. Experiments with stimulated Mono Mac 6 cells suggested that fMLPK-melphalan (at > 10(-7) M was more cytotoxic for these sells than melphalan alone. Trypan blue exclusion tests suggested that cells incubated with melphalan conjugate were less viable than cells kept with melphalan alone. Our results suggest that fMLP-toxin conjugates are internalized into myeloid cells via fMLP-receptors and are more cytotoxic for the cells in vitro than are the toxins alone, but only at or above the ED(50) concentration for ligand binding to the receptor.