Discovery and interrogation of new mechanism to control MYC oncogene by Mitogen‐Activated Protein Kinase Kinase 3
Authors: Yang Xuan , Du Yuhong , Fu Haian , Ivanov Andrey A 
Published: June 2021

Transcription factor MYC is a master regulator that modulates the expression of more than 15% of all genes in the human genome. MYC is frequently amplified across the vast majority of cancer types, and its amplification is correlated with a poor survival and prognosis of cancer patients. Moreover, the elevated expression of MYC has been demonstrated to play an essential role in cardiac myocyte hypertrophy, chronic liver diseases, neurodegenerative disorders, including Alzheimer and Parkinson diseases. Extensive clinical and biological studies have established MYC as a highly attractive therapeutic target. However, MYC lacks an enzymatic activity, and its targeting for drug discovery is highly challenging. Currently, there are no FDA‐approved MYC inhibitors, and novel therapeutic strategies to control MYC‐dependency are urgently needed.

Through the integration of the high‐throughput protein‐protein interaction (PPI) screening, bioinformatics, and biochemical studies we have discovered a new mechanism to regulate MYC activity through its interaction with Mitogen‐Activated Protein Kinase Kinase 3 (MKK3), a known activator of p38‐signaling. We found that in contrast to other MYC‐regulating kinases, such as ERK, MKK3 enhances MYC protein stability and transcriptional activity in a phosphorylation‐independent manner. Through biochemical studies guided by computational molecular modeling, we identified specific domains of MKK3 and MYC that are responsible for their interaction. Furthermore, we found that MKK3‐MYC PPI can be disrupted by a short 27 amino acid residue fragment derived from MYC helix‐loop‐helix domain. Overexpression of this peptide was correlated with the cell growth inhibition and induction of apoptosis in multiple cancer cell lines.

This data suggests that specific small molecule inhibitors of MKK3‐MYC PPI would provide new tools to investigate and control MKK3‐dependent MYC activity in different biological models. Toward this goal, we have developed a time‐resolved fluorescence resonance energy transfer (TR‐FRET) assay to monitor the interaction of MKK3 and MYC in mammalian cells. We have further optimized the assay for ultra‐high‐throughput screening (uHTS) in 1536‐well plate format. A pilot screening with a library of 3,280 compounds has identified several positive hits. The hits were further validated in a secondary dose‐dependent TR‐FRET assay as well as in orthogonal affinity pull‐down PPI inhibition assays.

Together, our data has revealed a novel biological role for MKK3 as a direct regulator of MYC‐driven cell growth and proliferation, and provides a new strategy for therapeutic discovery through interrogation of MKK3‐MYC PPI in MYC‐dependent disease.

Support or Funding Information

This research was supported in part by the National Cancer Institute of the NIH (Cancer Target Discovery and Development Network grants U01CA168449 and U01CA217875, H.F.), Emory University Research Committee grant (A.A.I.), and by the Fadlo R. Khuri Translational Research Award of the Winship Cancer Institute of Emory University (A.A.I).

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.