Assessment of Malaria In Vitro Drug Combination Screening and Mixed-Strain Infections Using the Malaria Sybr Green I-Based Fluorescence Assay
Assessment of Malaria In Vitro Drug Combination Screening and Mixed-Strain Infections Using the Malaria Sybr Green I-Based Fluorescence Assay
复制标题
DOI：
10.1128/aac.01370-08
复制
发表时间：
2009-06-01
影响因子：
4.9
通讯作者：
Johnson, Jacob D.
中科院分区：
文献类型：
--
作者：
Co, Edgie-Mark A.;Dennull, Richard A.;Johnson, Jacob D.
一些药物开发战略，包括优化新的抗疟药物组合，已被用来对付疟疾抗药性。我们评估了疟疾Sybr绿色I荧光（MSF）测定法在体外药物组合敏感性测定中的应用。使用MSF测定法测试了先前公布的协同（阿托伐醌和氯胍）、无关（氯喹和阿奇霉素）和拮抗（氯喹和阿托伐醌）抗疟药物相互作用的药物组合对恶性疟原虫菌株D6和W2的影响。计算单个药物和固定比例复方制剂相对于其单个IC（50）的50%抑制浓度（IC（50））。随后的等效线图分析和部分抑制浓度测定证明了每种测试组合的预期药物相互作用模式。此外，我们探索了MSF检测方法检查混合寄生虫种群动态的能力，这在疟疾患者分离株中很常见。具体而言，确定了MSF测定区分单一和混合寄生虫种群的能力。为了模拟体外混合感染，将固定比例的D6和W2菌株与抗疟药物共培养，并使用MSF测定法测定IC（50）。二分浓度曲线表明，敏感和耐药的寄生虫组成的遗传异质性群体是可检测的。进行双相分析以获得亚群IC（50），用于与单独的疟疾株获得的IC（50）进行比较。总之，MSF检测允许可靠的抗疟药物组合筛选，并提供了一个重要的方法来区分同质和异质寄生虫种群。
Several drug development strategies, including optimization of new antimalarial drug combinations, have been used to counter malaria drug resistance. We evaluated the malaria Sybr green I-based fluorescence (MSF) assay for its use in in vitro drug combination sensitivity assays. Drug combinations of previously published synergistic (atovaquone and proguanil), indifferent (chloroquine and azithromycin), and antagonistic (chloroquine and atovaquone) antimalarial drug interactions were tested against Plasmodium falciparum strains D6 and W2 using the MSF assay. Fifty percent inhibitory concentrations (IC(50)s) were calculated for individual drugs and in fixed ratio combinations relative to their individual IC(50)s. Subsequent isobologram analysis and fractional inhibitory concentration determinations demonstrated the expected drug interaction pattern for each combination tested. Furthermore, we explored the ability of the MSF assay to examine mixed parasite population dynamics, which are commonly seen in malaria patient isolates. Specifically, the capacity of the MSF assay to discern between single and mixed parasite populations was determined. To simulate mixed infections in vitro, fixed ratios of D6 and W2 strains were cocultured with antimalarial drugs and IC(50)s were determined using the MSF assay. Dichotomous concentration curves indicated that the sensitive and resistant parasites composing the genetically heterogeneous population were detectable. Biphasic analysis was performed to obtain subpopulation IC(50)s for comparison to those obtained for the individual malaria strains alone. In conclusion, the MSF assay allows for reliable antimalarial drug combination screening and provides an important method to discern between homogenous and heterogeneous parasite populations.