Active efflux confers intrinsic resistance to multiple antibiotics in Pseudomonas aeruginosa, including old disused molecules. Beside resistance, intracellular survival is another reason for failure to eradicate bacteria with antibiotics. We evaluated the capacity of polyaminoisoprenyl potentiators (designed as efflux pump inhibitors [EPIs]) NV716 and NV731 compared to PAβN to restore the activity of disused antibiotics (doxycycline, chloramphenicol [substrates for efflux], and rifampin [nonsubstrate]) in comparison with ciprofloxacin against intracellular P. aeruginosa (strains with variable efflux levels) in THP-1 monocytes exposed over 24 h to antibiotics alone (0.003 to 100× MIC) or combined with EPIs. Pharmacodynamic parameters (apparent static concentrations [C_s] and maximal relative efficacy [E_max]) were calculated using the Hill equation of concentration-response curves. PAβN and NV731 moderately reduced (0 to 4 doubling dilutions) antibiotic MICs but did not affect their intracellular activity. NV716 markedly reduced (1 to 16 doubling dilutions) the MIC of all antibiotics (substrates or not for efflux; strains expressing efflux or not); it also improved their relative potency and maximal efficacy (i.e., lower C_s; more negative E_max) intracellularly. In parallel, NV716 reduced the persister fraction in stationary cultures when combined with ciprofloxacin. In contrast to PAβN and NV731, which act only as EPIs against extracellular bacteria, NV716 can resensitize P. aeruginosa to antibiotics whether they are substrates or not for efflux, both extracellularly and intracellularly. This suggests a complex mode of action that goes beyond a simple inhibition of efflux to reduce bacterial persistence. NV716 appears to be a useful adjuvant, including to disused antibiotics with low antipseudomonal activity, to improve their activity, including against intracellular P. aeruginosa.

中文翻译：

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聚氨基异戊二烯增强剂 NV716 使旧的废弃抗生素恢复细胞内形式的铜绿假单胞菌感染

主动外排赋予铜绿假单胞菌对多种抗生素的内在抗性，包括旧的废弃分子。除了耐药性之外，细胞内存活是无法用抗生素根除细菌的另一个原因。与环丙沙星相比，我们评估了聚氨基异戊二烯增强剂（设计为外排泵抑制剂 [EPI]）NV716 和 NV731 与 PAβN 相比恢复废弃抗生素（强力霉素、氯霉素 [外排底物] 和利福平 [非底物]）活性的能力在 THP-1 单核细胞中，单独使用抗生素（0.003 至 100 × MIC）或与 EPI 结合超过 24 小时，可对抗细胞内铜绿假单胞菌（具有可变外排水平的菌株）。药效学参数（表观静态浓度 [ C_s ] 和最大相对功效 [ E _max ]) 使用浓度-响应曲线的 Hill 方程计算。PAβN 和 NV731 适度降低（0 到 4 倍稀释）抗生素 MIC，但不影响它们的细胞内活性。NV716 显着降低（1 到 16 倍稀释）所有抗生素的 MIC（底物或不用于外排；菌株是否表达外排）；它还在细胞内提高了它们的相对效力和最大功效（即更低的C _s；更负的E _max）。同时，NV716 与环丙沙星联合使用时，可减少固定培养物中的持久性部分。与 PAβN 和 NV731 仅作为对抗细胞外细菌的 EPI 相比，NV716 可以重新敏感铜绿假单胞菌对抗生素的影响，无论它们是否是细胞外和细胞内外排的底物。这表明了一种复杂的作用模式，它超越了简单的外排抑制以减少细菌的持久性。NV716 似乎是一种有用的佐剂，包括对具有低抗假单胞菌活性的废弃抗生素，以提高其活性，包括对抗细胞内铜绿假单胞菌。