Construction of Graphene Oxide-DNA Nanoprobe for Adenosine 5-Triphosphate Detection and Drug Delivery

doi:10.19894/j.issn.1000-0518.230012

Chinese Journal of Applied Chemistry ›› 2024, Vol. 41 ›› Issue (1): 118-127.DOI: 10.19894/j.issn.1000-0518.230012

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Construction of Graphene Oxide-DNA Nanoprobe for Adenosine 5-Triphosphate Detection and Drug Delivery

Yue ZHANG, Rui LIANG, Can-Nan ZHAO, Chun-Mei LI()

College of Pharmaceutical Sciences，Southwest University，Chongqing 400715，China

Received:2023-01-19 Accepted:2023-04-05 Published:2024-01-01 Online:2024-01-30
Contact: Chun-Mei LI
About author:licm1024@swu.edu.cn
Supported by:
the National Natural Science Foundation of China （No.?22074124）?, the Natural Science Foundation of Chongqing （No.?CSTB2022NSCQ-MSX0521） and Chongqing Municipal Training Program of Innovation and Entrepreneurship for Undergraduates(S202210635349)

Abstract

Abstract:

The abnormal concentration of adenosine 5-triphosphate （ATP） in cancer cells is closely related to the process of tumorigenesis and development. Therefore， rapid and accurate detection of ATP level in and out of cells is of great significance. Graphene oxide （GO） is widely used owing to its advantages of low toxicity， large specific surface area， easy functionalization， efficient and stable loading of DNA nanoprobes into cells. However， ssDNA bound to GO by physical adsorption is easily displaced by biomolecules in complex biological environment， leading to false positive signals. Based on this， a novel GO-DNA nanoprobe is proposed and applied to the detection of ATP and targeted delivery of the anticancer drug doxorubicin hydrochloride （DOX）. Double stranded DNA （dsDNA） is formed by hybridization of aptamer of ATP with complementary strand， and then DOX is loaded by G-C base pair to form dsDNA-DOX. dsDNA-DOX is adsorbed on GO surface through polyA sequence extended by complementary strand to construct GO-dsDNA-DOX nanoprobes， which could greatly reduce false positive signal caused by the physical adsorption of biomolecules in complex environment. The specific binding of ATP to aptamer can lead to the release of DOX， and quantitative analysis of ATP is achieved according to its fluorescence “off-on”. DOX fluorescence intensity shows a good linear relationship with the ATP content within the range of 0.08~8.0 mmol/L， and the linear equation is I_F=3.0897c+129.08. The detection limit （3σ/k） is 0.059 mmol/L， and the method has good selectivity and anti-interference ability. Cytotoxicity and fluorescence imaging results show that the DOX-loaded nanoprobes have significant drug release in MCF-7 cells and significantly induce apoptosis. This study establishes a label-free， simple and rapid method for ATP content detection and realizes targeted drug delivery by the high content of ATP in cancer cells， which reduces the toxic and side effects on normal cells and provides a new idea for cancer treatment.

Key words: Adenosine 5-triphosphate, DNA nanoprobe, Graphene oxide, Fluorescence detection, Doxorubicin hydrochloride, Drug delivery

CLC Number:

O655

Yue ZHANG, Rui LIANG, Can-Nan ZHAO, Chun-Mei LI. Construction of Graphene Oxide-DNA Nanoprobe for Adenosine 5-Triphosphate Detection and Drug Delivery[J]. Chinese Journal of Applied Chemistry, 2024, 41(1): 118-127.

Figures/Tables 6

Fig.1 Schematic illustration for the construction of GO-dsDNA-DOX nanoprobe for ATP detection and targeted DOX delivery

Fig.2 Investigation on the ATP response and salt stability of GO-dsDNA-DOX nanoprobe. （A） Fluorescence spectra of GO-dsDNA-DOX before or after reaction with ATP；（B） Zeta potential analysis of GO-dsDNA-DOX before or after reaction with ATP；（C） Salt stability of GO-dsDNA-DOX

Fig.3 Optimization of experiment conditions. （A） DOX loading temperature；（B） Concentration of GO；（C） Adsorption temperature of GO；（D） Adsorption time of GO；（E） ATP reaction time；（F） pH of buffer （Error bars represented standard deviations of three repetitive experiments）

Fig.4 Detection， specificity and anti-interference of GO-dsDNA-DOX probe for ATP assay. （A） Fluorescence response of the GO-dsDNA-DOX to different ATP concentrations；（B） The linear relationship between fluorescence intensity and ATP concentrations；（C） Specificity for the detection of ATP；（D） Anti-interference of probe （Error bars represented standard deviations of three repetitive experiments）

Fig.5 Analysis of nuclease stability and cytotoxicity of GO-dsDNA-DOX probe. （A） Investigation on stability of probe against nuclease；（B） Cytotoxicity of MCF-7 cells incubated with different concentrations of probes （Error bars represented standard deviations of three repetitive experiments）

Fig.6 Fluorescence images of the GO-dsDNA-DOX in MCF-7 cells. Line 1： control cells； Line 2： cells incubated with DOX； Line 3： cells incubated with GO-dsDNA-DOX； Line 4： cells incubated with GO-dsDNA-DOX-oligomycin

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Construction of Graphene Oxide-DNA Nanoprobe for Adenosine 5-Triphosphate Detection and Drug Delivery

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