Dissertation
Synthesis, stability, and metal chelator interactions of triphenylphosphonium conjugates
University of Iowa
Doctor of Philosophy (PhD), University of Iowa
Autumn 2022
DOI: 10.25820/etd.006767
Abstract
Mitochondria are organelles in eukaryotic cells that play an essential role in various processes including those regulating cellular life and death. One of the main functions of the mitochondria is to generate ATP through oxidative phosphorylation (OXPHOS), which is the process of oxidizing nutrients to release chemical energy. Dysfunction of the mitochondria can lead to several disease states and very often leads to cell death. Because of the role of mitochondria in several physiological processes necessary for survival and the various disease states that exist because of dysfunctional mitochondria, mitochondria are an important target for probes and medications for therapeutic intervention.
Triphenylphosphonium (TPP+) is a diffused lipophilic cation known for its tendency to accumulate in the mitochondria. Because of this, it is commonly used in drug conjugates and probes to direct cargo to the mitochondria. TPP+ effectively targets mitochondria because of its lipophilic and diffused cationic character. These qualities allow TPP+ to readily cross lipid bilayer membranes and accumulate across the negative inner mitochondrial membrane against their concentration gradient, respectively.
While TPP+ is effective in mitochondrial accumulation, it has been shown to cause a dose- mediated uncoupling of OXPHOS, which depolarizes the mitochondrial membrane and leads to cell death. Previous research has identified phenyl-substituted conjugates with modulated uncoupling potential as a function of the electron-density on the central phosphorus atom of TPP+. Electron-donating substituents such as methoxy increase the electron density on the central phosphorus atom and cause a relative increase in uncoupling potential as compared to unsubstituted TPP+. Electron-withdrawing substituents such as trifloromethyl (CF3) decrease the electron density of the central phosphorus cation, and effectively decrease dose-mediated uncoupling of OXPHOS.
The first goal of this work was to develop novel TPP+ conjugates with electron-withdrawing phenyl substitutions for further analysis for mitochondrial uncoupling potential and cytotoxicity. This was attempted by three different approaches, first to change the identity of the phenyl substituents on TPP+, second to change the number of substituted phenyls on phosphonium conjugates, and third to change the number of substitutions on triphenylphosphonium. Though modulating the identity and quantity of electron-withdrawing groups, it was hypothesized that uncoupling potential could be modulated in a step-wise fashion.
The second goal of this work was to analyze the stability of the previously reported CF3-para- substutited conjugate with decyl linker (CF3-TPP+-DC) after potential stability concerns with this conjugate had been identified. This section aimed to monitor the stability of CF3-TPP+-DC in a variety of experimentally relevant buffers and solvents, as well as biologically relevant conditions that mimic environments found in the mitochondria.
Results of the stability experiments suggested a potential interaction between CF3-TPP+-DC and the metal chelator, DTPA, which led to the final goal of this dissertation, to probe potential interactions between metal chelators of divalent cations and TPP+ conjugates. This section aimed to elucidate interactions between EDTA, DTPA, and Chelex100 with TPP+-based drugs and probes.
Results of these studies identify specific conditions that cause degradation of CF3-TPP+-DC, and several conditions of stability for previously reported TPP+ conjugates and commercially available probes. These studies identify and provide the first report of interactions between metal chelators and TPP+ conjugates, including commercially available probes.
Details
- Title: Subtitle
- Synthesis, stability, and metal chelator interactions of triphenylphosphonium conjugates
- Creators
- Hannah Gruenwald
- Contributors
- Robert J. Kerns (Advisor)Ethan Anderson (Committee Member)Jonathan Doorn (Committee Member)David Roman (Committee Member)
- Resource Type
- Dissertation
- Degree Awarded
- Doctor of Philosophy (PhD), University of Iowa
- Degree in
- Pharmacy
- Date degree season
- Autumn 2022
- Publisher
- University of Iowa
- DOI
- 10.25820/etd.006767
- Number of pages
- xvi, 192 pages
- Copyright
- Copyright 2022 Hannah Gruenwald
- Language
- English
- Description illustrations
- Illustrations, charts, graphs
- Description bibliographic
- Includes bibliographical references (pages 180-192).
- Public Abstract (ETD)
Mitochondria are organelles of human cells that play a role in several processes that support cellular function, life, and death and mitochondrial dysfunction can lead to several diseases. Because of the essential role of the mitochondria in maintaining life, it is a common site for the delivery of drugs and diagnostic agents. Triphenylphosphonium (TPP+) is a chemical moiety that enables this targeting as it readily accumulates in mitochondria. Through linkage to TPP+, drug cargo can effectively reach the mitochondria for therapeutic intervention. While effective in mitochondrial targeting, TPP+ is not an inert carrier, and increased dose leads to increased cytotoxicity. This cytotoxicity can be modulated, however, through modifications to the TPP+ moiety, such as through electron-withdrawing substitutions.
The first goal of this study is to develop novel, modified conjugates for further analysis of their cytotoxicity. The second goal of this study is to assess the stability of the least cytotoxic of these modified derivatives with CF3 substitutions to the para-position of the TPP+ phenyl groups and decyl linker (CF3-TPP+-DC) in experimentally and biologically relevant conditions. The final goal of this study is to analyze potential interactions between metal chelators and TPP+ conjugates, after results from the stability studies suggested a potential interaction between the metal chelator, DTPA, and CF3-TPP+-DC.
There have been no reported stability studies on modified TPP+ conjugates, and little to no reported data on stability of commercially available TPP+ based molecules. These results provide the first comprehensive report of TPP+ conjugate stability, and the first report of any interactions between metal chelators and TPP+ conjugates.
- Academic Unit
- Pharmacy
- Record Identifier
- 9984362857902771
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