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Article

RX-3117 (Fluorocyclopentenyl-Cytosine)-Mediated Down-Regulation of DNA Methyltransferase 1 Leads to Protein Expression of Tumor-Suppressor Genes and Increased Functionality of the Proton-Coupled Folate Carrier

1
Laboratory Medical Oncology, Amsterdam UMC, location VU University Medical Center, 1081 HV Amsterdam, The Netherlands
2
Department of Oncology, Wayne State University School of Medicine, Detroit, and Molecular Therapeutics Program, Barbara Ann Karmanos Cancer Institute, Detroit, MI 48201-1976, USA
3
Rexahn Pharmaceuticals, Inc., Rockville, MD 20850, USA
4
Department of Biochemistry, Medical University of Gdansk, 80-210 Gdansk, Poland
*
Author to whom correspondence should be addressed.
Int. J. Mol. Sci. 2020, 21(8), 2717; https://doi.org/10.3390/ijms21082717
Received: 14 January 2020 / Revised: 31 March 2020 / Accepted: 10 April 2020 / Published: 14 April 2020
(This article belongs to the Special Issue Immunoepigenetics: A Future Sentinel for Tumor Directed Therapies)
(1) Background: RX-3117 (fluorocyclopentenyl-cytosine) is a cytidine analog that inhibits DNA methyltransferase 1 (DNMT1). We investigated the mechanism and potential of RX-3117 as a demethylating agent in several in vitro models. (2) Methods: we used western blotting to measure expression of several proteins known to be down-regulated by DNA methylation: O6-methylguanine-DNA methyltransferase (MGMT) and the tumor-suppressor genes, p16 and E-cadherin. Transport of methotrexate (MTX) mediated by the proton-coupled folate transporter (PCFT) was used as a functional assay. (3) Results: RX-3117 treatment decreased total DNA-cytosine-methylation in A549 non-small cell lung cancer (NSCLC) cells, and induced protein expression of MGMT, p16 and E-cadherin in A549 and SW1573 NSCLC cells. Leukemic CCRF-CEM cells and the MTX-resistant variant (CEM/MTX, with a deficient reduced folate carrier) have a very low expression of PCFT due to promoter hypermethylation. In CEM/MTX cells, pre-treatment with RX-3117 increased PCFT-mediated MTX uptake 8-fold, and in CEM cells 4-fold. With the reference hypomethylating agent 5-aza-2′-deoxycytidine similar values were obtained. RX-3117 also increased PCFT gene expression and PCFT protein. (4) Conclusion: RX-3117 down-regulates DNMT1, leading to hypomethylation of DNA. From the increased protein expression of tumor-suppressor genes and functional activation of PCFT, we concluded that RX-3117 might have induced hypomethylation of the promotor. View Full-Text
Keywords: RX-3117; 5-aza-2′-deoxycytidine; hypomethylation; proton-coupled folate receptor; methotrexate RX-3117; 5-aza-2′-deoxycytidine; hypomethylation; proton-coupled folate receptor; methotrexate
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MDPI and ACS Style

Sarkisjan, D.; Julsing, J.R.; El Hassouni, B.; Honeywell, R.J.; Kathmann, I.; Matherly, L.H.; Lee, Y.B.; Kim, D.J.; Peters, G.J. RX-3117 (Fluorocyclopentenyl-Cytosine)-Mediated Down-Regulation of DNA Methyltransferase 1 Leads to Protein Expression of Tumor-Suppressor Genes and Increased Functionality of the Proton-Coupled Folate Carrier. Int. J. Mol. Sci. 2020, 21, 2717. https://doi.org/10.3390/ijms21082717

AMA Style

Sarkisjan D, Julsing JR, El Hassouni B, Honeywell RJ, Kathmann I, Matherly LH, Lee YB, Kim DJ, Peters GJ. RX-3117 (Fluorocyclopentenyl-Cytosine)-Mediated Down-Regulation of DNA Methyltransferase 1 Leads to Protein Expression of Tumor-Suppressor Genes and Increased Functionality of the Proton-Coupled Folate Carrier. International Journal of Molecular Sciences. 2020; 21(8):2717. https://doi.org/10.3390/ijms21082717

Chicago/Turabian Style

Sarkisjan, Dzjemma, Joris R. Julsing, Btissame El Hassouni, Richard J. Honeywell, Ietje Kathmann, Larry H. Matherly, Young B. Lee, Deog J. Kim, and Godefridus J. Peters 2020. "RX-3117 (Fluorocyclopentenyl-Cytosine)-Mediated Down-Regulation of DNA Methyltransferase 1 Leads to Protein Expression of Tumor-Suppressor Genes and Increased Functionality of the Proton-Coupled Folate Carrier" International Journal of Molecular Sciences 21, no. 8: 2717. https://doi.org/10.3390/ijms21082717

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