Epirubicin, Identified Using a Novel Luciferase Reporter Assay for Foxp3 Inhibitors, Inhibits Regulatory T Cell Activity.

Abstract

Forkhead box protein p3 (Foxp3) is crucial to the development and suppressor function of regulatory T cells (Tregs) that have a significant role in tumor-associated immune suppression. Development of small molecule inhibitors of Foxp3 function is therefore considered a promising strategy to enhance anti-tumor immunity. In this study, we developed a novel cell-based assay system in which the NF-κB luciferase reporter signal is suppressed by the co-expressed Foxp3 protein. Using this system, we screened our chemical library consisting of approximately 2,100 compounds and discovered that a cancer chemotherapeutic drug epirubicin restored the Foxp3-inhibited NF-κB activity in a concentration-dependent manner without influencing cell viability. Using immunoprecipitation assay in a Treg-like cell line Karpas-299, we found that epirubicin inhibited the interaction between Foxp3 and p65. In addition, epirubicin inhibited the suppressor function of murine Tregs and thereby improved effector T cell stimulation in vitro. Administration of low dose epirubicin into tumor-bearing mice modulated the function of immune cells at the tumor site and promoted their IFN-γ production without direct cytotoxicity. In summary, we identified the novel action of epirubicin as a Foxp3 inhibitor using a newly established luciferase-based cellular screen. Our work also demonstrated our screen system is useful in accelerating discovery of Foxp3 inhibitors.

• Authors: Hajime Kashima, Fumiyasu Momose, Hiroshi Umehara, Nao Miyoshi, Naohisa Ogo, Daisuke Muraoka, Hiroshi Shiku, Naozumi Harada, Akira Asai
• Journal: PloS one (PLoS One) Vol. 11 Issue 6 Pg. e0156643 ( 2016) ISSN: 1932-6203 [Electronic] United States
• PMID: 27284967 (Publication Type: Journal Article)

Chemical References

• Antineoplastic Agents
• FOXP3 protein, human
• Forkhead Transcription Factors
• Epirubicin
• Luciferases

Topics

• Animals
• Antineoplastic Agents (isolation & purification, pharmacology)
• Cells, Cultured
• Down-Regulation (drug effects)
• Drug Screening Assays, Antitumor (methods)
• Epirubicin (isolation & purification, pharmacology)
• Female
• Forkhead Transcription Factors (antagonists & inhibitors)
• Genes, Reporter
• HEK293 Cells
• Humans
• Luciferases (genetics, metabolism)
• Lymphocyte Activation (drug effects)
• Mice
• Mice, Inbred BALB C
• T-Lymphocytes, Regulatory (drug effects, immunology)