Date

12-19-2024

Document Type

Thesis

Abstract

Doxorubicin (Doxo) is a historically significant anti-cancer drug that has been used to treat multiple types of cancers for half a century. The drug targets DNA of cancer cells through a binding mode called intercalation, acting as roadblock during DNA replication to stop tumor formation. While Doxo has been extensively studied through traditional methods, only a handful of research has utilized single molecule methods, which can provide new insights at the molecular level. In this study, we used dual beam optical tweezers to isolate single DNA molecules and mechanically manipulate the DNA in the presence of Doxo. We conducted two types of experiments: slow stretching experiments and constant force experiments. In the slow stretching experiments, we allowed time for Doxo to reach binding equilibrium by pausing after each step while stretching the DNA. In the constant force experiments, we held the DNA at constant tension and measured the extension as a function of time. The equilibrium extensions of the DNA upon binding to Doxo were used, along with a one-dimensional binding model, to find how strongly the drug binds to DNA (the binding affinity) at various forces. Although we performed only one set of constant force experiments at 20pN, we validated that the equilibrium extensions upon binding to Doxo and the binding affinity from both experiments matched within uncertainty. Quantifying the results provided us with an equilibrium extension of DNA upon an intercalation event to be 0.20±0.03 nm. The optical tweezers experiment revealed a nano molar binding affinity of 57±12 nM in contrast to micromolar affinitiesreported in the literature. These results could be very beneficial to not only reduce the dosage and the side effects of Doxo but also to the development of better anti-cancer drugs in the future.

Department

Physics

Thesis Committee

Dr. Thayaparan Paramanathan, Thesis Advisor
Dr. Edward Deveney, Committee Member
Dr. Thomas Kling, Committee Member

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