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Updated: Nov 1, 2025
In Vitro Transcription Assays and Their Application in Drug Discovery
Published on: September 20, 2016
Norstictic Acid Is a Selective Allosteric Transcriptional Regulator
Julie M Garlick1,2, Steven M Sturlis1,2, Paul A Bruno1,2
1Life Sciences Institute, University of Michigan, Ann Arbor, Michigan 48109, United States.
Abstract:
Inhibitors of transcriptional protein-protein interactions (PPIs) have high value both as tools and for therapeutic applications. The PPI network mediated by the transcriptional coactivator Med25, for example, regulates stress-response and motility pathways, and dysregulation of the PPI networks contributes to oncogenesis and metastasis. The canonical transcription factor binding sites within Med25 are large (∼900 Å2) and have little topology, and thus, they do not present an array of attractive small-molecule binding sites for inhibitor discovery. Here we demonstrate that the depsidone natural product norstictic acid functions through an alternative binding site to block Med25-transcriptional activator PPIs in vitro and in cell culture. Norstictic acid targets a binding site comprising a highly dynamic loop flanking one canonical binding surface, and in doing so, it both orthosterically and allosterically alters Med25-driven transcription in a patient-derived model of triple-negative breast cancer. These results highlight the potential of Med25 as a therapeutic target as well as the inhibitor discovery opportunities presented by structurally dynamic loops within otherwise challenging proteins.
Insights
Norstictic acid blocks Med25-mediated transcription by targeting a dynamic loop, offering new therapeutic strategies. This discovery opens avenues for inhibiting protein-protein interactions (PPIs) in cancer, particularly triple-negative breast cancer.
Area of Science:
- Biochemistry
- Molecular Biology
- Oncology
Background:
- Transcriptional protein-protein interactions (PPIs) are crucial for cellular processes, including stress response and motility.
- Dysregulation of PPI networks, like those involving Med25, is linked to cancer development and metastasis.
- Traditional binding sites on proteins like Med25 are large and lack topology, posing challenges for small-molecule inhibitor discovery.
Purpose of the Study:
- To identify novel therapeutic strategies targeting Med25-mediated transcriptional regulation.
- To explore alternative binding sites on Med25 for inhibitor development.
- To investigate the potential of norstictic acid as an inhibitor of Med25-transcriptional activator PPIs.
Main Methods:
- In vitro assays to assess the inhibitory activity of norstictic acid on Med25-transcriptional activator PPIs.
- Cell culture experiments to evaluate the effects of norstictic acid on Med25-driven transcription.
- Utilized a patient-derived model of triple-negative breast cancer for in vivo validation.
Main Results:
- Norstictic acid effectively blocks Med25-transcriptional activator PPIs.
- The natural product norstictic acid targets a unique, highly dynamic loop on Med25, distinct from canonical binding sites.
- Norstictic acid modulates Med25-driven transcription both orthosterically and allosterically, impacting cancer cell behavior.
- Demonstrated efficacy in a triple-negative breast cancer model.
Conclusions:
- Med25 represents a promising therapeutic target for cancer treatment.
- Structurally dynamic loops on proteins offer new opportunities for inhibitor discovery, even for challenging targets.
- Norstictic acid serves as a valuable lead compound for developing novel anti-cancer therapeutics targeting PPIs.
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