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Incorporation and Repair of Epigenetic Intermediates as Potential Chemotherapy Agents
Jason L Herring1, Mark L Sowers1,2, James W Conrad1
1Department of Pharmacology and Toxicology, University of Texas Medical Branch, 301 University Boulevard, Galveston, TX 77555, USA.
Abstract:
The incorporation of nucleoside analogs into DNA by polymerases, followed by their removal through base excision repair (BER), represents a promising strategy for cancer chemotherapy. In this study, we investigated the incorporation and cytotoxic effects of several nucleoside analogs-some of which are epigenetic reprogramming intermediates-in the U87 glioblastoma cell line. We found that two analogs, 5-hydroxymethyl-2'-deoxyuridine (5HmdU) and trifluorothymidine (TFT), are both cytotoxic and are efficiently incorporated into genomic DNA. In contrast, the 5-carboxy analogs-5-carboxy-2'-deoxyuridine (5CadU) and 5-carboxycytidine (5CadC)-showed no cytotoxicity and were not incorporated into DNA. Interestingly, 5-hydroxymethyl-2'-deoxycytidine (5HmdC) was cytotoxic but was not directly incorporated into DNA. Instead, it was deaminated into 5HmdU, which was then incorporated and likely responsible for the observed toxicity. 5HmdU is actively removed from DNA through the BER pathways. In contrast, TFT remains stably incorporated and is neither excised by BER nor does it hydrolyze into 5CadU-a known substrate for the DNA glycosylase SMUG1. We also found that N6-benzyladenosine (BzAdo), an inhibitor of the enzyme 2'-deoxynucleoside 5'-phosphate N-hydrolase (DNPH1), enhances the cytotoxicity of 5HmdU. However, the thymidine phosphorylase inhibitor tipiracil hydrochloride (TPI) does not increase the cytotoxic effect of TFT in U87 cells. Together, these findings highlight 5HmdU and TFT as promising chemotherapeutic agents for glioblastoma, each with distinct mechanisms of action and cellular processing.
Insights
Two nucleoside analogs, 5-hydroxymethyl-2'-deoxyuridine (5HmdU) and trifluorothymidine (TFT), show promise as cancer chemotherapy agents for glioblastoma. They are cytotoxic and incorporated into DNA, with distinct mechanisms of action and cellular processing.
Area of Science:
- Biochemistry
- Molecular Biology
- Cancer Research
Background:
- Nucleoside analogs incorporated into DNA and subsequently removed by base excision repair (BER) offer a potential cancer chemotherapy strategy.
- Investigating epigenetic reprogramming intermediates as nucleoside analogs is crucial for developing novel cancer treatments.
Purpose of the Study:
- To evaluate the incorporation and cytotoxic effects of various nucleoside analogs in the U87 glioblastoma cell line.
- To understand the mechanisms of action and cellular processing of promising nucleoside analogs for glioblastoma treatment.
Main Methods:
- Assessing cytotoxicity of nucleoside analogs in U87 glioblastoma cells.
- Quantifying the incorporation of nucleoside analogs into genomic DNA.
- Investigating the role of base excision repair (BER) in the removal of incorporated nucleoside analogs.
- Evaluating the effects of enzyme inhibitors (DNPH1, TPI) on nucleoside analog cytotoxicity.
Main Results:
- 5-hydroxymethyl-2 '-deoxyuridine (5HmdU) and trifluorothymidine (TFT) exhibited cytotoxicity and were incorporated into DNA.
- 5-carboxy analogs (5CadU, 5CadC) were not cytotoxic and not incorporated.
- 5-hydroxymethyl-2 '-deoxycytidine (5HmdC) was cytotoxic, deaminated to 5HmdU, and then incorporated.
- 5HmdU was actively removed by BER, while TFT remained stably incorporated.
- N 6-benzyladenosine (BzAdo) enhanced 5HmdU cytotoxicity, but tipiracil hydrochloride (TPI) did not affect TFT cytotoxicity.
Conclusions:
- 5HmdU and TFT are effective chemotherapeutic agents for glioblastoma with distinct mechanisms.
- The cellular processing and DNA repair pathways influence the efficacy of these nucleoside analogs.
- Further research into 5HmdU and TFT could lead to improved glioblastoma treatment strategies.
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