PLGA 纳米粒中的联合抗逆转录病毒药物治疗 HIV-1。
Combination antiretroviral drugs in PLGA nanoparticle for HIV-1.
机构信息
Department of Pharmacy Practice, Creighton University School of Pharmacy & Health Professions, Omaha, NE, USA.
出版信息
BMC Infect Dis. 2009 Dec 9;9:198. doi: 10.1186/1471-2334-9-198.
BACKGROUND
Combination antiretroviral (AR) therapy continues to be the mainstay for HIV treatment. However, antiretroviral drug nonadherence can lead to the development of resistance and treatment failure. We have designed nanoparticles (NP) that contain three AR drugs and characterized the size, shape, and surface charge. Additionally, we investigated the in vitro release of the AR drugs from the NP using peripheral blood mononuclear cells (PBMCs).
METHODS
Poly-(lactic-co-glycolic acid) (PLGA) nanoparticles (NPs) containing ritonavir (RTV), lopinavir (LPV), and efavirenz (EFV) were fabricated using multiple emulsion-solvent evaporation procedure. The nanoparticles were characterized by electron microscopy and zeta potential for size, shape, and charge. The intracellular concentration of AR drugs was determined over 28 days from NPs incubated with PBMCs. Macrophages were imaged by fluorescent microscopy and flow cytometry after incubation with fluorescent NPs. Finally, macrophage cytotoxicity was determined by MTT assay.
RESULTS
Nanoparticle size averaged 262 +/- 83.9 nm and zeta potential -11.4 +/- 2.4. AR loading averaged 4% (w/v). Antiretroviral drug levels were determined in PBMCs after 100 microg of NP in 75 microL PBS was added to media. Intracellular peak AR levels from NPs (day 4) were RTV 2.5 +/- 1.1; LPV 4.1 +/- 2.0; and EFV 10.6 +/- 2.7 microg and continued until day 28 (all AR >or= 0.9 microg). Free drugs (25 microg of each drug in 25 microL ethanol) added to PBMCs served as control were eliminated by 2 days. Fluorescence microscopy and flow cytometry demonstrated phagocytosis of NP into monocytes-derived macrophages (MDMs). Cellular MTT assay performed on MDMs demonstrated that NPs are not significantly cytotoxic.
CONCLUSION
These results demonstrated AR NPs could be fabricated containing three antiretroviral drugs (RTV, LPV, EFV). Sustained release of AR from PLGA NP show high drug levels in PBMCs until day 28 without cytotoxicity.
背景
联合抗逆转录病毒（AR）治疗仍是 HIV 治疗的主要方法。然而，抗逆转录病毒药物不依从可能导致耐药性的产生和治疗失败。我们设计了含有三种 AR 药物的纳米颗粒（NP），并对其大小、形状和表面电荷进行了表征。此外，我们还研究了 NP 在外周血单核细胞（PBMC）中的 AR 药物体外释放情况。
方法
采用复乳-溶剂蒸发法制备含利托那韦（RTV）、洛匹那韦（LPV）和依非韦伦（EFV）的聚（乳酸-共-乙醇酸）（PLGA）纳米颗粒（NP）。通过电子显微镜和 Zeta 电位对 NP 的大小、形状和电荷进行了表征。将 NP 与 PBMC 孵育 28 天后，测定 AR 药物的细胞内浓度。用荧光 NP 孵育巨噬细胞后，通过荧光显微镜和流式细胞术对巨噬细胞进行成像。最后，通过 MTT 测定法测定巨噬细胞的细胞毒性。
结果
NP 的平均粒径为 262 +/- 83.9nm，Zeta 电位为-11.4 +/- 2.4。AR 载药量平均为 4%（w/v）。将 75μL PBS 中的 100μg NP 加入培养基中，测定 PBMC 中的 AR 药物水平。NP 细胞内峰值 AR 水平（第 4 天）为 RTV 2.5 +/- 1.1；LPV 4.1 +/- 2.0；EFV 10.6 +/- 2.7μg，持续至第 28 天（所有 AR >或= 0.9μg）。加入 PBMC 的游离药物（每种药物 25μg 在 25μL 乙醇中）作为对照，2 天内即被消除。荧光显微镜和流式细胞术显示 NP 被单核细胞衍生的巨噬细胞（MDMs）吞噬。对 MDMs 进行的细胞 MTT 测定表明，NP 没有明显的细胞毒性。
结论
这些结果表明，可制备含有三种抗逆转录病毒药物（RTV、LPV、EFV）的 AR NP。PLGA NP 中 AR 的持续释放显示，在没有细胞毒性的情况下，PBMC 中的药物水平可维持至第 28 天。
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