dc.contributor.advisor |
Loomes, Kerry |
|
dc.contributor.advisor |
Squire, Chris |
|
dc.contributor.author |
Krinkel, Benjamin Andrew |
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dc.date.accessioned |
2022-06-29T01:45:36Z |
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dc.date.available |
2022-06-29T01:45:36Z |
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dc.date.issued |
2022 |
en |
dc.identifier.uri |
https://hdl.handle.net/2292/60214 |
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dc.description |
Full Text is available to authenticated members of The University of Auckland only. |
en |
dc.description.abstract |
Cancer is a leading cause of death. Consequently, efficacious anticancer treatments are still crucial to improve treatment outcomes and quality of life. Many cancers adapt their cellular metabolism to facilitate rapid cell growth. One feature is reliance on glutamine as a source for mitochondrial metabolism and increased expression of malic enzyme (ME) isoform 2, which catalyses the conversion of the substrate, L-malate, to pyruvate with the reduction of NAD(P)+.
There are three human malic enzyme (ME) isoforms: ME1 (cytosolic) and ME2/ME3 (mitochondrial). Overexpression of ME1 and ME2 is associated with various cancers, with ME2 knockdowns generating ROS, diminishing NADPH generation, and inducing cellular senescence. Thus, targeting malic enzyme isoforms in cancer may be an attractive therapeutic approach. Currently, two small molecule inhibitors of ME2 are known, serving as possible drug leads to target cancer cell metabolism. These comprise of NPD-389 and embonic acid that inhibit ME2 activity. ME3 remains fundamentally uncharacterised, and the effect of these inhibitors has not been investigated.
The aim of this project was to determine the inhibitory profile of NPD-389 and embonic acid across the three malic enzyme isoforms. Specific objectives: (i) express and purify human recombinant ME 1-3 isoforms; (ii) characterise their kinetic parameters; and (iii) profile the inhibitory potencies of NPD-389 and embonic acid. Kinetic parameters derived for ME1 and ME2 relative to substrate (L-malate), cofactor (NAD(P)+) and allosteric activator, fumarate closely matched the literature. The ME3 data is novel, with a KM of 0.89 mM for L-malate and a KM of 0.26 mM for NADP+. Fumarate also inhibited ME3 activity, indicating a putative fumarate binding site.
Derived inhibitory constants (Ki) showed NPD-389 was a potent inhibitor of all three ME isoforms, putatively targeting the malate substrate binding site. Embonic acid potently inhibited ME2 and ME3 but was a poor inhibitor of ME1. Findings suggest a distinct binding site for embonic acid in the vicinity of the fumarate binding site of ME2 and possibly ME3. This study suggests that NPD-389 and embonic acid are promising leads toward the development of new anticancer treatments targeting malic enzymes in tumour metabolism. |
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dc.publisher |
ResearchSpace@Auckland |
en |
dc.relation.ispartof |
Masters Thesis - University of Auckland |
en |
dc.relation.isreferencedby |
UoA |
en |
dc.rights |
Restricted Item. Full Text is available to authenticated members of The University of Auckland only. |
en |
dc.rights |
Items in ResearchSpace are protected by copyright, with all rights reserved, unless otherwise indicated. |
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dc.rights.uri |
https://researchspace.auckland.ac.nz/docs/uoa-docs/rights.htm |
en |
dc.rights.uri |
http://creativecommons.org/licenses/by-nc-sa/3.0/nz/ |
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dc.title |
Kinetic characterisation of recombinant human malic enzymes with small molecule inhibitors |
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dc.type |
Thesis |
en |
thesis.degree.discipline |
Biological Sciences |
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thesis.degree.grantor |
The University of Auckland |
en |
thesis.degree.level |
Masters |
en |
dc.date.updated |
2022-06-03T03:51:29Z |
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dc.rights.holder |
Copyright: the author |
en |