Downregulation of thymidylate synthase and E2F1 by arsenic trioxide in mesothelioma

Our plain-English summary of Downregulation of thymidylate synthase and E2F1 by arsenic trioxide in mesothelioma, by S.K. Lam, Y.Y. Li, C.Y. Zheng, et al (International Journal of Oncology, 2015).

The study

• Authors: S.K. Lam, Y.Y. Li, C.Y. Zheng, J.C. Ho
• Journal: International Journal of Oncology
• Published: 2015;46(1):113-22
• Cited by: 14 articles, as counted by NIH iCite
• Read it: PubMed · Publisher

Abstract

Malignant pleural mesothelioma is a global health issue. Arsenic trioxide (ATO) has been shown to suppress thymidylate synthase (TYMS) in lung adenocarcinoma and colorectal cancer, and induce apoptosis in acute promyelocytic leukemia. With TYMS as a putative therapeutic target, the effect of ATO in mesothelioma was therefore studied. A panel of 5 mesothelioma cell lines was used to study the effect of ATO on cell viability, protein expression, mRNA expression and TYMS activity by MTT assay, western blot, qPCR and tritium-release assay, respectively. The knockdown of TYMS and E2F1 was performed with a specific siRNA. Phosphatidylserine externalization and mitochondrial membrane depolarization were measured by Annexin V and JC-1 staining respectively. The in vivo effect of ATO was studied using a nude mouse xenograft model. Application of ATO demonstrated anticancer effects in the cell line model with clinically achievable concentrations. Downregulation of TYMS protein (except H226 cells and 1.25 µM ATO in H2052 cells) and mRNA expression (H28 cells), pRB1 (H28 cells) and E2F1 and TYMS activity (except H226 cells) were also evident. E2F1 knockdown decreased cell viability more significantly than TYMS knockdown. In general, thymidine kinase 1, ribonucleotide reductase M1, c-myc and skp2 were downregulated by ATO. p-c-Jun was downregulated in H28 cells while upregulated in 211H cells. Phosphatidylserine externalization, mitochondrial membrane depolarization, downregulation of Bcl-2 and Bcl-xL, and upregulation of Bak and cleaved caspase-3 were observed. In the H226 xenograft model, the relative tumor growth was aborted, and E2F1 was downregulated while cleaved caspase-3 was elevated and localized to the nucleus in the ATO treatment group. ATO has potent antiproliferative and cytotoxic effects in mesothelioma in vitro and in vivo, partially mediated through E2F1 targeting (less effect through TYMS targeting). There is sound scientific evidence to support the clinical application of ATO in treatment of mesothelioma.

In plain English

This laboratory study tested arsenic trioxide, a drug approved for a type of leukemia, against malignant pleural mesothelioma. It did not study asbestos exposure or patients. The authors treated five purchased mesothelioma cell lines with the drug. They also gave it daily by abdominal injection to mice carrying tumors grown from one line, H226. The drug reduced growth in all five cell lines. The concentration that halved growth was 4.7 to 7.8 micromolar at 48 hours and 1.7 to 7 micromolar at 72 hours. It lowered the protein E2F1 in all cell lines. It lowered the enzyme thymidylate synthase in some lines but raised its level and activity in H226 cells. Silencing E2F1 cut viability by 37 to 72% in four lines. Silencing thymidylate synthase, tested in three lines, cut viability by 25% in two and not at all in the third. Cleaved caspase-3, a marker of programmed cell death, rose in all lines. After 23 days, relative tumor volume in the two dose groups of mice was 42.9% and 46.5% that of controls. The authors concluded the effect was at least partly through E2F1 and that their findings support exploring clinical use.

We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

What the study did and found

• Design: Laboratory study using mesothelioma cell lines (in vitro) and a mouse tumor xenograft experiment (in vivo)
• Who was studied: Five commercially purchased human mesothelioma cell lines: NCI-H28 (sarcomatoid), MSTO-211H (biphasic), NCI-H226 (epithelioid), NCI-H2052 (sarcomatoid) and NCI-H2452 (epithelioid). Also 18 female nude mice injected with H226 cells. The work was done at the University of Hong Kong.
• Exposure: Arsenic trioxide (ATO) as a drug treatment, given to cells at various concentrations. Mice received PBS (control) or ATO at 3.75 or 7.5 mg/kg daily by intraperitoneal injection. Asbestos exposure was not studied.
• Main results:
  • ATO reduced growth in all five cell lines. The concentrations that halved growth (IC50) were 4.7 to 7.8 micromolar after 48 hours and 1.7 to 7 micromolar after 72 hours.
  • E2F1 protein decreased significantly in all cell lines after ATO treatment.
  • Thymidylate synthase (TYMS) protein decreased in H28 and H2452 cells and increased in H226 cells after 48 hours of ATO.
  • ATO inhibited TYMS activity in H28, H2052 and H2452 cells but increased TYMS activity in H226 cells.
  • Only H28, H226 and H2052 cells were used in the TYMS silencing experiment, because 211H cells did not express TYMS and H2452 cells could not survive the low-serum conditions.
  • Silencing TYMS was associated with a 25% reduction in cell viability in H28 and H226 cells and no change in viability in H2052 cells.
  • Silencing E2F1 was associated with reductions in cell viability of 57%, 72%, 37% and 45% in H28, 211H, H226 and H2052 cells, respectively.
  • Cleaved caspase-3, a marker of programmed cell death, increased with ATO in all cell lines.
  • In mice with H226 tumors, relative tumor volume after 23 days of treatment was 42.9% (3.75 mg/kg group) and 46.5% (7.5 mg/kg group) that of the control group.
  • Body weight did not differ between the mouse groups, and no pathological changes were seen in the livers of ATO-treated mice.
• The authors' conclusion: The authors concluded that ATO had an antiproliferative effect at least partially mediated through downregulation of E2F1, and a cytotoxic effect through apoptosis, in both the cell line and mouse models. They stated that their findings provide a scientific basis for future exploration of clinical use of ATO in malignant pleural mesothelioma.
• Limitations the authors state:
  • Mouse tumor models could be made only from H226 cells. Xenografts from the other four cell lines could not be established.
  • H2452 cells could not survive the low-serum conditions used for the gene-silencing experiments, so they were not included.

Topics

The National Library of Medicine indexes this study under subjects our site covers: Lung cancer, Mesothelioma.

Medical terms in this study, explained in our Lexicon: Mesothelioma, Pleural Mesothelioma, Lung Adenocarcinoma, Colorectal Cancer, Apoptosis.

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