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Published on: November 15, 2013
A simple yeast-based system for analyzing inhibitor resistance in the human cancer drug targets Hsp90alpha/beta
Stefan H Millson1, Chrisostomos Prodromou, Peter W Piper
1Department of Molecular Biology and Biotechnology, The University of Sheffield, Western Bank, UK.
Abstract:
Heat shock protein 90 (Hsp90), a highly conserved molecular chaperone, is one of the most promising targets for cancer drug development. Whether any resistance to these Hsp90 inhibitor drugs could arise by Hsp90 mutation is still unknown. Yeast is readily engineered so that its essential Hsp90 function is provided by either isoform of the human cytosolic Hsp90, Hsp90alpha or Hsp90beta. However, its high intrinsic resistance to most drugs poses a major obstacle to the use of such Hsp90alpha- or Hsp90beta-expressing yeast cells as a model system to analyse whether drug resistance might arise by Hsp90 mutation. In order to overcome this problem, we have generated a strain that is both hypersensitive to Hsp90 inhibitors as it lacks multiple drug resistance genes, and in which different heterologous and mutant Hsp90s can be expressed by plasmid exchange. It is not rendered appreciably stress sensitive when made to express Hsp90alpha or Hsp90beta as its sole form of Hsp90. Should there be any development of resistance to the Hsp90 drugs now in cancer clinic trials, this system can provide a rapid initial test of whether any single nucleotide polymorphism appearing within the coding regions of Hsp90alpha or Hsp90beta could be a contributory factor in this resistance. We have used this strain to demonstrate that significant levels of resistance to the Hsp90 inhibitors radicicol and 17-allylamino-demethoxygeldanamycin (17-AAG) are generated as a result of the same single point mutation within the native Hsp90 of yeast (A107N), the human Hsp90alpha (A121N) and the human Hsp90beta (A116N).
Insights
A new yeast strain sensitive to Hsp90 inhibitors allows rapid testing for drug resistance mutations. This system can identify if Hsp90 mutations contribute to resistance against cancer drugs targeting heat shock protein 90 (Hsp90).
Area of Science:
- Oncology
- Molecular Biology
- Biochemistry
Background:
- Heat shock protein 90 (Hsp90) is a key target for cancer drug development.
- Understanding potential drug resistance mechanisms, such as Hsp90 mutations, is crucial for effective cancer therapy.
Purpose of the Study:
- To develop a yeast model system for analyzing Hsp90 mutations as a cause of drug resistance.
- To create a yeast strain hypersensitive to Hsp90 inhibitors, enabling rapid screening of resistance mechanisms.
Main Methods:
- Engineered a yeast strain lacking multiple drug resistance genes for increased sensitivity to Hsp90 inhibitors.
- Utilized plasmid exchange to express heterologous and mutant human Hsp90alpha and Hsp90beta.
- Tested the strain's response to Hsp90 inhibitors radicicol and 17-allylamino-demethoxygeldanamycin (17-AAG).
Main Results:
- Demonstrated that specific single point mutations in yeast, human Hsp90alpha, and human Hsp90beta confer significant resistance to Hsp90 inhibitors.
- Identified conserved mutations (A107N in yeast, A121N in Hsp90alpha, A116N in Hsp90beta) as drivers of resistance.
Conclusions:
- The developed yeast system is effective for rapidly assessing the impact of Hsp90 mutations on drug resistance.
- This model can predict whether specific Hsp90 polymorphisms may contribute to resistance against Hsp90-targeting cancer drugs currently in clinical trials.
