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) 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 severa...
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MDPI AG
2020-04-01
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| Series: | International Journal of Molecular Sciences |
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| Online Access: | https://www.mdpi.com/1422-0067/21/8/2717 |
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| author | Dzjemma Sarkisjan Joris R. Julsing Btissame El Hassouni Richard J. Honeywell Ietje Kathmann Larry H. Matherly Young B. Lee Deog J. Kim Godefridus J. Peters |
| author_facet | Dzjemma Sarkisjan Joris R. Julsing Btissame El Hassouni Richard J. Honeywell Ietje Kathmann Larry H. Matherly Young B. Lee Deog J. Kim Godefridus J. Peters |
| author_sort | Dzjemma Sarkisjan |
| collection | DOAJ |
| description | (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: O<sup>6</sup>-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 <i>PCFT</i> 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. |
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| issn | 1661-6596 1422-0067 |
| language | English |
| last_indexed | 2024-03-10T20:29:04Z |
| publishDate | 2020-04-01 |
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| spelling | doaj.art-68b4de910bbf4c838cb3c06a99ffccf72023-11-19T21:34:55ZengMDPI AGInternational Journal of Molecular Sciences1661-65961422-00672020-04-01218271710.3390/ijms21082717RX-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 CarrierDzjemma Sarkisjan0Joris R. Julsing1Btissame El Hassouni2Richard J. Honeywell3Ietje Kathmann4Larry H. Matherly5Young B. Lee6Deog J. Kim7Godefridus J. Peters8Laboratory Medical Oncology, Amsterdam UMC, location VU University Medical Center, 1081 HV Amsterdam, The NetherlandsLaboratory Medical Oncology, Amsterdam UMC, location VU University Medical Center, 1081 HV Amsterdam, The NetherlandsLaboratory Medical Oncology, Amsterdam UMC, location VU University Medical Center, 1081 HV Amsterdam, The NetherlandsLaboratory Medical Oncology, Amsterdam UMC, location VU University Medical Center, 1081 HV Amsterdam, The NetherlandsLaboratory Medical Oncology, Amsterdam UMC, location VU University Medical Center, 1081 HV Amsterdam, The NetherlandsDepartment of Oncology, Wayne State University School of Medicine, Detroit, and Molecular Therapeutics Program, Barbara Ann Karmanos Cancer Institute, Detroit, MI 48201-1976, USARexahn Pharmaceuticals, Inc., Rockville, MD 20850, USARexahn Pharmaceuticals, Inc., Rockville, MD 20850, USALaboratory Medical Oncology, Amsterdam UMC, location VU University Medical Center, 1081 HV Amsterdam, The Netherlands(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: O<sup>6</sup>-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 <i>PCFT</i> 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.https://www.mdpi.com/1422-0067/21/8/2717RX-31175-aza-2′-deoxycytidinehypomethylationproton-coupled folate receptormethotrexate |
| spellingShingle | Dzjemma Sarkisjan Joris R. Julsing Btissame El Hassouni Richard J. Honeywell Ietje Kathmann Larry H. Matherly Young B. Lee Deog J. Kim Godefridus J. Peters 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 RX-3117 5-aza-2′-deoxycytidine hypomethylation proton-coupled folate receptor methotrexate |
| title | 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 |
| title_full | 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 |
| title_fullStr | 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 |
| title_full_unstemmed | 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 |
| title_short | 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 |
| title_sort | 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 |
| topic | RX-3117 5-aza-2′-deoxycytidine hypomethylation proton-coupled folate receptor methotrexate |
| url | https://www.mdpi.com/1422-0067/21/8/2717 |
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