Research Abstract Title
MDM2 enhances taxane sensitivity by inducing STMN1 degradation
Presenter Status
Student
Abstract Type
Research
Primary Mentor
Tomoo Iwakuma
Start Date
5-5-2022 11:30 AM
End Date
5-5-2022 1:30 PM
Presentation Type
Poster Presentation
Description
Background: MDM2, a major E3 ubiquitin ligase of p53, is overexpressed in ~30% of human cancers. Evidence showed that overexpressed MDM2 has p53-independent oncogenic functions and is correlated with poor prognosis. Given that MDM2 overexpression is cancer-specific, therapies targeting vulnerabilities imposed by MDM2 overexpression may cause cell death specifically in cancer cells with minimum side effects.
Objectives/Goal: Our goal is to identify drugs that specifically kill MDM2-overexpressing cancer cells in a p53- independent manner.
Methods/Design: We generated p53-knockout (p53KO) and p53/MDM2-double knockout (DKO) SJSA-1 osteosarcoma cells in which MDM2 gene is amplified by using the CRISPR-Cas9 system. We used MTT assay to measure cell viability, flow cytometry to measure cell cycle, western blot for measuring microtubule polymerization markers, microtubule destabilizing proteins levels. To validate our findings in SJSA1 cells in other cell lines, we use RNAi and exogenous protein expression techniques to generate MDM2 knock down or overexpression cell lines.
Results: We treated p53KO and DKO SJSA-1 cells with 18 available FDA-approved chemotherapeutic agents and found that paclitaxel and docetaxel, which are a group of drugs called “taxanes”, specifically killed MDM2-overexpressing p53KO cells at significantly lower concentrations than that in MDM2-lacking DKO cells. Similar observations were obtained using other cell lines with or without MDM2 knockout or overexpression. Also, MDM2 knockout inhibited taxane’s mechanism of action - microtubule polymerization, mitotic arrest, and cell death, which was rescued by re-introduction of wild-type MDM2, but not an E3 ubiquitin ligase dead mutant MDM2C464A. We furthermore identified that protein levels and stability of a microtubule destabilizing protein, STMN1, were significantly increased by MDM2 knockout, and concomitant depletion of STMN1 re-sensitized MDM2-knockout cells to taxanes.
Conclusions: Our study discovers a yet unidentified p53-independent function of MDM2, in which MDM2 induces degradation of STMN1, leading to increased sensitivity to taxanes. Our findings enhance our knowledge about the taxane’s mechanism of action, which can be exploited clinically by using taxanes for MDM2-overexpressing cancers.
MeSH Keywords
neoplasms drug therapy
Additional Files
MDM2 enhances taxane sensitivity by inducing STMN1 degradation.pdf (234 kB)Abstract
Included in
MDM2 enhances taxane sensitivity by inducing STMN1 degradation
Background: MDM2, a major E3 ubiquitin ligase of p53, is overexpressed in ~30% of human cancers. Evidence showed that overexpressed MDM2 has p53-independent oncogenic functions and is correlated with poor prognosis. Given that MDM2 overexpression is cancer-specific, therapies targeting vulnerabilities imposed by MDM2 overexpression may cause cell death specifically in cancer cells with minimum side effects.
Objectives/Goal: Our goal is to identify drugs that specifically kill MDM2-overexpressing cancer cells in a p53- independent manner.
Methods/Design: We generated p53-knockout (p53KO) and p53/MDM2-double knockout (DKO) SJSA-1 osteosarcoma cells in which MDM2 gene is amplified by using the CRISPR-Cas9 system. We used MTT assay to measure cell viability, flow cytometry to measure cell cycle, western blot for measuring microtubule polymerization markers, microtubule destabilizing proteins levels. To validate our findings in SJSA1 cells in other cell lines, we use RNAi and exogenous protein expression techniques to generate MDM2 knock down or overexpression cell lines.
Results: We treated p53KO and DKO SJSA-1 cells with 18 available FDA-approved chemotherapeutic agents and found that paclitaxel and docetaxel, which are a group of drugs called “taxanes”, specifically killed MDM2-overexpressing p53KO cells at significantly lower concentrations than that in MDM2-lacking DKO cells. Similar observations were obtained using other cell lines with or without MDM2 knockout or overexpression. Also, MDM2 knockout inhibited taxane’s mechanism of action - microtubule polymerization, mitotic arrest, and cell death, which was rescued by re-introduction of wild-type MDM2, but not an E3 ubiquitin ligase dead mutant MDM2C464A. We furthermore identified that protein levels and stability of a microtubule destabilizing protein, STMN1, were significantly increased by MDM2 knockout, and concomitant depletion of STMN1 re-sensitized MDM2-knockout cells to taxanes.
Conclusions: Our study discovers a yet unidentified p53-independent function of MDM2, in which MDM2 induces degradation of STMN1, leading to increased sensitivity to taxanes. Our findings enhance our knowledge about the taxane’s mechanism of action, which can be exploited clinically by using taxanes for MDM2-overexpressing cancers.
