Abstract
Tumor-specific promoters and cis-regulatory genetic elements are used for transcriptional control of therapeutic transgene expression in cancer gene therapy. HRE (hypoxia response element) and ARE (antioxidant response element) cis-regulatory elements are targets for HIF1 and Nrf2 transcriptional factors, respectively, and mediate activation of gene transcription in a response to hypoxia and oxidative stress, characteristic of most solid tumors. Due to these features HREs and AREs are used in genetic constructs for cancer gene therapy to provide tumor-specific therapeutic transgene expression or replication of oncolytic adenoviruses. In this work on the basis of the tumor-specific promoter hTERT we have constructed hybrid promoters carrying combinations of HRE and ARE. We showed that upon imitation of hypoxia in human lung cancer cell lines the activity of the hybrid promoter HRE-ARE-hTERT is substantially higher compared to promoters carrying only ARE or HRE. Using in vitro suicide cancer gene therapy with the CD: UPRT/5-FC (cytosine deaminase: uracil phosphoribosyl transferase/5-fluorocytosine) enzyme-prodrug system as a model we showed an enhancement of the cytotoxic effect on human lung cancer cells upon imitation of hypoxia when cytosine deaminase: uracil phosphoribosyl transferase was expressed under the control of the HRE-ARE-hTERT promoter compared to HRE-hTERT and ARE-hTERT promoters. The novel hybrid promoter HRE-ARE-hTERT could be used for transcriptional targeting of therapeutic transgene expression or oncolytic adenovirus replication upon development of novel anti-cancer gene therapeutics.
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ACKNOWLEDGMENTS
The work used the infrastructure of the Core Facility Center of the Institute of Gene Biology, Russian Academy of Sciences.
Funding
This work was supported by grant no. 075-15-2019-1661 of the Ministry of Science and Higher Education of the Russian Federation.
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The work does not use animals or biological materials obtained from humans. The authors declare no conflict of interest.
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Abbreviations: ARE, antioxidant response element; HRE, hypoxia response element; hTERT, human telomerase reverse transcriptase; CD:UPRT, cytosine deaminase: uracil phosphoribosyl transferase fusion protein; 5FC, 5-fluorocytosine; RLU, relative luminescence units; SD, standard deviation; 3′‑UTR, 3′-untranslated region.
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Kalinichenko, S.V., Korobko, I.V. & Shepelev, M.V. Combination of ARE and HRE cis-Regulatory Elements Elevates the Activity of Tumor-Specific hTERT Promoter. Mol Biol 55, 555–564 (2021). https://doi.org/10.1134/S0026893321030055
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DOI: https://doi.org/10.1134/S0026893321030055