Sarah Kurtz's GOx Aptamer Selection Project (2019)

Therapeutic Aptamer Targeting Glucose Oxidase to Suppress Tumor Growth through the Creation of Artificial Super Neutrophils

Introduction and Background

In 2019 alone, more than 1.7 million people are expected to be diagnosed with cancer (Street, 2019). Additionally, more than 1,600 Americans are expected to die from cancer everyday (Street, 2019). Cancer is not only deadly, but costly as well. In 2015 alone, cancer medical costs were over $80 billion for Americans last year, placing a financial strain on the families of those affected (Street, 2019). Glucose oxidase (GOx) is an enzyme that has been shown to be effective in cancer diagnosis and treatment and is more cost effective.

Glucose oxidase (GOx) is an enzyme that catalyzes beta-D glucose into H₂O₂ and gluconic acid (Bankar, Bule, Singhal, & Ananthanarayan, 2009) as shown in Figure 1 (Afjeh, M. E. A., Pourahmad, R., Akbari-Adergani, B., & Azin, M, 2019).

Glucose oxidase is effective in cancer therapy by depleting the tumor’s access to oxygen through catalyzing glucose into hydrogen peroxide and gluconic acid. By combining glucose oxidase with other cancer therapies, tumors can be effectively killed, and the cancer eradicated (Fu et al., n.d.).

Aptamers can be used to target glucose oxidase. Aptamers are short single-stranded RNA or DNA that can bind with affinity and specificity to certain targets. Aptamers can be used in diagnosis, therapy, and drug delivery. Selection of aptamers is done through the SELEX process, which includes a pool-binding reaction, washes, reverse transcription, PCR, transcription, PAGE, RNA quantification, and binding assay and sequencing. Multiple rounds of selections are performed before an aptamer can be selected in order to get an aptamer with a good level of stringency. Aptamers are considered to be analogues of antibodies, but they perform even more effectively and efficiently. Aptamers are more cost effective and easier to produce than antibodies (Lakhin, Tarantul, & Gening, 2013). Aptamers have been used in the diagnosis and treatment for colorectal cancer (Chen, Zhou, Cai, & Tang, 2017) and in the targeting of cancer stem cells for cancers such as leukemia, lung cancer, and breast cancer (Zhou et al., 2017).

In this selection, the aptamer will be used for a therapeutic application. Neutrophils are a form of leukocytes that are present in the body. Neutrophils fight against tumors, cancer growth, and infections. Despite this, there is often not enough neutrophils, or the neutrophils are not strong enough to fight the tumors. Recently, artificial “super neutrophils” have been invented to work better against tumor growth than natural neutrophils. The artificial neutrophils were made using a zinc compound structure (zeolitic imidazolate framework-8), GOx, and chloroperoxidase (CPO) as shown in Figure 2 (Zhang et al., 2019.).

In this study, artificial neutrophils were shown to produce more reactive HClO, which is considered to be antitumor, than the natural neutrophils (Zhang et al., 2019). The artificial neutrophils were also less damaging to the tissues surrounding the tumors and less inflammation (Zhang et al., 2019). The aptamer in this selection will be used instead of the zeolitic imidazolate framework-8 as the structural support for an artificial super neutrophil. An aptamer would be cheaper and more cost effective than the zeolitic imidazolate framework-8. This aptamer will be stabilized and used in conjunction with DNA origami, which are strands of DNA that are folded into shapes and dimensions that can be used as scaffolding and support in cancer therapy (Rajagopalan, n.d.). The DNA origami scaffolding can be used in conjunction with the aptamer to provide the framework for the artificial super neutrophil that would suppress and decrease tumor growth. This will be done by binding the aptamer to the glucose oxidase in conjunction with the CPO, which would increase production of the HClO and have stronger antitumor effects than a naturally occurring neutrophil. This aptamer would relieve patients suffering from cancer by decreasing the size of their tumors and by providing another step in the process toward remission that can be used in conjunction with already existing cancer treatments. This possible treatment option could also have less side effects than traditional cancer treatment. Many people suffer from cancer, and hopefully, this can help to ease or even end their suffering.

An aptamer selection is underway and will begin this week with filter-based in vivo selection. The research objective at the moment is to finish my second round of filter-based selection by the next progress report. The aptamer to address the problem of cancer with artificial neutrophils will hopefully be discovered and selected by numerous rounds of selection. This aptamer will help reduce deaths due to cancer in upcoming years.

Click here for Final Report

References

Aptamer Stream. (2019). 2019 Spring Target List and Brief Descriptions. Retrieved from https://utexas.instructure.com/courses/1239466/files/folder/Target%20information?preview=48966343.

Aptamer Stream (2019). Glucose oxidase-biotin. [PDF]. Retrieved from https://utexas.instructure.com/courses/1239466/files/folder/Target%20information?preview=48049115.

Afjeh, M. E. A., Pourahmad, R., Akbari-Adergani, B., & Azin, M. (2019). Use of Glucose Oxidase Immobilized on Magnetic Chitosan Nanoparticles in Probiotic Drinking Yogurt. Food Science of Animal Resources, 39(1), 73–83. https://doi.org/10.5851/kosfa.2019.e5

Bankar, S. B., Bule, M. V., Singhal, R. S., & Ananthanarayan, L. (2009). Glucose oxidase — An overview. Biotechnology Advances, 27(4), 489–501. https://doi.org/10.1016/j.biotechadv.2009.04.003

Chen, C., Zhou, S., Cai, Y., & Tang, F. (2017). Nucleic acid aptamer application in diagnosis and therapy of colorectal cancer based on cell-SELEX technology. Npj Precision Oncology, 1(1), 37. https://doi.org/10.1038/s41698-017-0041-y

Fu, L.-H., Qi, C., Lin, J., & Huang, P. (2018). Catalytic chemistry of glucose oxidase in cancer diagnosis and treatment. Chemical Society Reviews, 47(17), 6454–6472. https://doi.org/10.1039/C7CS00891K

Fu, L.-H., Qi, C., Hu, Y.-R., Lin, J., & Huang, P. (n.d.). Glucose Oxidase-Instructed Multimodal Synergistic Cancer Therapy. Advanced Materials, 0(0), 1808325. https://doi.org/10.1002/adma.201808325

Fu, L.-H., Qi, C., Lin, J., & Huang, P. (2018). Catalytic chemistry of glucose oxidase in cancer diagnosis and treatment. Chemical Society Reviews, 47(17), 6454–6472. https://doi.org/10.1039/C7CS00891K

Lakhin, A. V., Tarantul, V. Z., & Gening, L. V. (2013). Aptamers: Problems, Solutions and Prospects. Acta Naturae, 5(4), 34–43.

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Sitaula, S., Branch, S. D., & Ali, M. F. (2012). GOx signaling triggered by aptamer-based ATP detection. Chemical Communications (Cambridge, England), 48(74), 9284–9286. https://doi.org/10.1039/c2cc34279k

Street, W. (2019). Cancer Facts & Figures 2019. 76.

Zhang, C., Zhang, L., Wu, W., Gao, F., Li, R.-Q., Song, W., … Zhang, X.-Z. (n.d.). Artificial Super Neutrophils for Inflammation Targeting and HClO Generation against Tumors and Infections. Advanced Materials, 0(0), 1901179. https://doi.org/10.1002/adma.201901179

Zhou, G., Latchoumanin, O., Bagdesar, M., Hebbard, L., Duan, W., Liddle, C., … Qiao, L. (2017). Aptamer-Based Therapeutic Approaches to Target Cancer Stem Cells. Theranostics, 7(16), 3948–3961. https://doi.org/10.7150/thno.20725