CELLULAR ACCUMULATION OF THE NEW KETOLIDE RU-64004 BY HUMAN NEUTROPHILS - COMPARISON WITH THAT OF AZITHROMYCIN AND ROXITHROMYCIN

We analyzed the uptake of RU 64004 by human neutrophils (polymorphonuc lear leukocytes [PMNs]) relative to those of azithromycin and roxithro mycin, RU 64004 was strongly and rapidly accumulated by PMNs, with a c ellular concentration/extracellular concentration ratio (C/E) of great er than 200 in the first 5 min, and this was followed by a plateau at 120 to 180 min, with a C/E of 461 +/- 14.8 (10 experiments) at 180 min . RU 64004 uptake was moderately sensitive to external pH, and activat ion energy was also moderate (63 +/- 3.8 kJ/mol), RU 64004 was mainly located in PMN granules (about 70%) and egressed slowly from loaded ce lls, owing to avid reuptake, The possibility that PMN uptake of RU 640 04 and other macrolides occurs through a carrier-mediated system was s uggested by three key results. First, there existed a strong interindi vidual variability in uptake kinetics, suggesting variability in the n umbers or activity of a transport protein, Second, macrolide uptake di splayed saturation kinetics characteristic of that of a carrier-mediat ed transport system: RU 64004 had the highest V-max value (3,846 ng/2. 5 x 10(6) PMNs/5 min) and the lowest K-m value (about 28 mu M), indica ting a high affinity for the transporter, Third, as observed previousl y with other erythromycin A derivatives, Ni2+ (a blocker of the Na+/Ca 2+ exchanger which mediates Ca2+ influx in resting neutrophils) impair ed RU 64004 uptake by PMNs, with a 50% inhibitory concentration of abo ut 3.5 mM, In addition, me found that an active process is also involv ed in macrolide efflux, because verapamil significantly potentiated th e release of all three macrolides tested, This effect of verapamil doe s not seem to be related to an inhibition of Ca2+ influx, because neit her EGTA [ethylene glycol-bis (beta-aminoethyl ether)-N,N,N',N'-tetraa cetic acid] nor Ni2+ modified macrolide efflux. The nature and charact eristics of the entry-and efflux-mediating carrier systems are under i nvestigation.