Activity-based sensors for reactive aldehydes and their corresponding enzymatic machinery

Bearrood, Thomas Edward

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https://hdl.handle.net/2142/109493 Copy

Description

Title

Activity-based sensors for reactive aldehydes and their corresponding enzymatic machinery

Author(s)

Bearrood, Thomas Edward

Issue Date

2020-11-25

Director of Research (if dissertation) or Advisor (if thesis)

Chan, Jefferson

Doctoral Committee Chair(s)

Chan, Jefferson

Committee Member(s)

• Burke, Martin
• Sarlah, David
• Zimmerman, Steven

Department of Study

Chemistry

Discipline

Chemistry

Degree Granting Institution

University of Illinois at Urbana-Champaign

Degree Name

Ph.D.

Degree Level

Dissertation

Keyword(s)

• molecular imaging
• activity-based sensors
• aldehydes
• aldehyde dehydrogenase
• 4-hydroxynonenal

Abstract

Molecular imaging agents are chemical tools that have become essential to the study of life. The field of molecular imaging encompasses a wide range of modalities and mechanism, but an exciting and growing subfield is that of activity-based sensing. Activity-based sensors provides new insights into biological processes since they report on the activity of biological species rather than just the presence. Naturally these are ideal probes for enzymes, affording multiple turnovers and higher signal from a single active enzyme. Activity-based sensors are also excellent sensors for reactive small molecules with unique chemical reactivity. Here, we report the development and/or progress on activity-based sensors for reactive biological aldehydes and the enzymatic machinery tasked with aldehyde metabolism. We first developed AlDeSense, the first turn on fluorescent probe selective for aldehyde dehydrogenase (ALDH) 1A1. We pursued a probe for this enzyme since it’s frequently cited as a cancer stem cell marker. AlDeSense had remarkable selectivity for ALDH1A1, producing a 20-fold increase in fluorescence signal upon oxidation by the enzyme. We confirmed ALDH1A1 activity, as measured by AlDeSense, was a reliable cancer stem cell marker in leukemia, melanoma, and breast cancer models. AlDeSense was then applied to image ALDH1A1 activity in aggressive and non-aggressive melanoma ex vivo and in vivo. We demonstrated that not only do the cancer stem cells have elevated ALDH1A1 activity in cell culture, but they maintain elevated activity through establishment of a tumor microenvironment. We next sought to improve on certain limitations of AlDeSense, namely the green fluorescence and cell impermeability. Towards this goal, eleven possible Si-xanthene probes were synthesized. Each iteration provided new insights into the electronic and steric requirements for isoform selectivity and fluorescence turn on. In the end, the careful incorporation of two fluorine atoms on the reactive benzaldehyde moiety produced a selective molecule named red-AlDeSense. Red-AlDeSense was used to study ALDH1A1 activity in lung cancer models. We also aimed to develop a selective probe for 4-hydroxynonenal (4HNE). It is one of the most reactive and abundant aldehydes produced during oxidative stresses (i.e. lipid peroxidation) and has been connected to many age-associated conditions including cancer and neurodegeneration. We hypothesized selectivity could be achieved against the other 10000-plus aldehydes reported in biological systems using an activity-based sensor. We are currently investigating synthetic strategies to achieve a fluorophore capped by our hydrazinoacetate trigger. We anticipate this probe will be extremely useful to study the effects of 4HNE in living systems of many disease models. We are particularly interested in its connection to cancer stem cells.

Graduation Semester

2020-12

Type of Resource

Thesis

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http://hdl.handle.net/2142/109493

Copyright and License Information

Copyright 2020 Thomas Bearrood

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