Identification of methotrexate as a heterochromatin-promoting drug

Andre C. Loyola,Lin Zhang,Pranabananda Dutta,Robin Shang,Jinghong Li,Willis X. Li

doi:10.1038/s41598-019-48137-w

Andre C. Loyola, Lin Zhang + Show 4 more

Open Access

https://doi.org/10.1038/s41598-019-48137-w

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Journal: Scientific Reports	Publication Date: Aug 12, 2019
Citations: 13	License type: open-access

Affiliation: University of California, San Diego

Abstract

Heterochromatin is a tightly packed form of DNA involved in gene silencing, chromosome segregation, and protection of genome stability. Heterochromatin is becoming more recognized in tumor suppression and may thus serve as a potential target for cancer therapy. However, to date there are no drugs that are well established to specifically promote heterochromatin formation. Here, we describe a screening method using Drosophila to identify small molecule compounds that promote heterochromatin formation, with the purpose of developing epigenetic cancer therapeutics. We took advantage of a Drosophila strain with a variegated eye color phenotype that is sensitive to heterochromatin levels, and screened a library of 97 FDA approved oncology drugs. This screen identified methotrexate as the most potent small molecule drug, among the 97 oncology drugs screened, in promoting heterochromatin formation. Interestingly, methotrexate has been identified as a JAK/STAT inhibitor in a functional screen, causing reduced phosphorylation of STAT proteins. These findings are in line with our previous observation that unphosphorylated STAT (uSTAT) promotes heterochromatin formation in both Drosophila and human cells and suppresses tumor growth in mouse xenografts. Thus, Drosophila with variegated eye color phenotypes could be an effective tool for screening heterochromatin-promoting compounds that could be candidates as cancer therapeutics.

Highlights

In eukaryotic cells, DNA is folded with histone and non-histone proteins in order to form chromatin
We obtained from the National Cancer Institute (NCI) Developmental Therapeutics Program (DTP) a small-molecule drug library, Oncology Set III, consisting of 97 Food and Drug Administration (FDA) approved oncology drugs (Table S1)
Due to the metastable nature of heterochromatin induced at the P[mini-white+] tandem repeats, transcription of P[mini-white+] and the degree of variegation in the eye color of DX1 flies is sensitive to heterochromatin levels

Summary

Introduction

DNA is folded with histone and non-histone proteins in order to form chromatin. Nucleosomes, the basic units of chromatin consist of 146 bp of DNA wrapped around a histone octomer core and linker histone protein H1, the core being made up of two copies of H2A, H2B, H3 and H4 histone proteins This chromosomal template is the foundation for many important cellular processes, such as gene transcription, DNA repair and recombination[1,2]. Constitutive heterochromatin domains occur in regions that are highly dense in repetitive DNA elements and possess structural importance These regions remain condensed throughout the cell cycle. Tri-methylation of histone 3 lysine 9 (H3K9me3) and recruitment of HP1 chromo domain proteins is well established to be associated with heterochromatin and epigenetic silencing[1,2] These proteins are indicators and markers for heterochromatin formation. Retinoblastoma (Rb) tumor surpressor, a protein that suppresses transcription of E2F genes upon DNA damage in order to prevent the division of damaged cells, mediates its effects through stabilizing heterochromatin[8,9,10,11,12]

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