Detecting Attomolar DNA-Damaging Anticancer Drug Activity in Cell Lysates with Electrochemical DNA Devices.
Detecting Attomolar DNA-Damaging Anticancer Drug Activity in Cell Lysates with Electrochemical DNA Devices.
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DOI：
10.1021/acssensors.1c00365
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发表时间：
2021-07-23
期刊：
影响因子：
8.9
通讯作者：
Slinker JD
中科院分区：
文献类型：
--
作者：
Wettasinghe AP;Singh N;Starcher CL;DiTusa CC;Ishak-Boushaki Z;Kahanda D;McMullen R;Motea EA;Slinker JD
在这里，我们利用电化学DNA装置来量化和了解癌症特异性DNA损伤活性的新兴药物在细胞裂解物中的飞摩尔和阿摩尔浓度。制备了一种有效的肿瘤选择性NAD（P）H醌氧化还原酶1（NQO 1）生物活化药物异丁基-脱氧尼波醌（IB-DNQ），并通过Western印迹、NQO 1活性分析、存活测定、耗氧率、细胞外酸化率和过氧化物产生，在高表达NQO 1（NQO 1+）和低表达NQO 1的敲减（NQO 1 −）的癌细胞中进行了生物化学验证。然后将来自这些细胞的裂解物和IB-DNQ药物引入到带有DNA修饰电极阵列的芯片系统中，并通过碱基切除修复引起的DNA介导的电化学变化来定量其DNA损伤活性。使用NQO 1活性的设备级控制和动力学分析来验证和进一步理解IB-DNQ活性。在NQO 1+裂解物中观察到380 aM的IB-DNQ检测限和1.3 fM的损伤中点，这两个指标都比NQO 1 −裂解物低2个数量级，表明IB-DNQ对NQO 1+癌症的高效力和选择性。NQO 1+裂解物中的装置水平损伤中点浓度比细胞存活基准低8个数量级以上，可能是由于IB-DNQ细胞摄取差，表明这些装置可以鉴定需要改善细胞渗透性的有前景的药物。最终，这些结果表明IB-DNQ的显著效力和选择性以及电化学DNA装置分析DNA损伤化疗中涉及的试剂/药物的高灵敏度和精确度。
Here, we utilize electrochemical DNA devices to quantify and understand the cancer-specific DNA-damaging activity of an emerging drug in cellular lysates at femtomolar and attomolar concentrations. Isobutyl-deoxynyboquinone (IB-DNQ), a potent and tumor-selective NAD(P)H quinone oxidoreductase 1 (NQO1) bioactivatable drug, was prepared and biochemically verified in cancer cells highly expressing NQO1 (NQO1+) and knockdowns with low NQO1 expression (NQO1−) by Western blot, NQO1 activity analysis, survival assays, oxygen consumption rate, extracellular acidification rate, and peroxide production. Lysates from these cells and the IB-DNQ drug were then introduced to a chip system bearing an array of DNA-modified electrodes, and their DNA-damaging activity was quantified by changes in DNA-mediated electrochemistry arising from base-excision repair. Device-level controls of NQO1 activity and kinetic analysis were used to verify and further understand the IB-DNQ activity. A 380 aM IB-DNQ limit of detection and a 1.3 fM midpoint of damage were observed in NQO1+ lysates, both metrics 2 orders of magnitude lower than NQO1− lysates, indicating the high IB-DNQ potency and selectivity for NQO1+ cancers. The device-level damage midpoint concentration in NQO1+ lysates was over 8 orders of magnitude lower than cell survival benchmarks, likely due to poor IB-DNQ cellular uptake, demonstrating that these devices can identify promising drugs requiring improved cell permeability. Ultimately, these results indicate the noteworthy potency and selectivity of IB-DNQ and the high sensitivity and precision of electrochemical DNA devices to analyze agents/drugs involved in DNA-damaging chemotherapies.
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影响因子：
4.7
作者：
Gorodetsky, Alon A.;Buzzeo, Marisa C.;Barton, Jacqueline K.
通讯作者：
Barton, Jacqueline K.
影响因子：
50.3
作者：
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通讯作者：
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影响因子：
21.8
作者：
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通讯作者：
Pardee K
影响因子：
7.4
作者：
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通讯作者：
Kelley, Shana O.
影响因子：
46.9
作者：
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通讯作者：
Barton, JK