The epigenetic impact of suberohydroxamic acid and 5‑Aza‑2'‑deoxycytidine on DNMT3B expression in myeloma cell lines
Katerina Smesny Trtkova1, Petra Luzna1, Denisa Weiser Drozdkova1
1Department of Clinical and Molecular Pathology, Faculty of Medicine and Dentistry, Palacky University Olomouc, 777 15 Olomouc, Czech Republic.
Abstract:
Gene inactivation of the cyclin‑dependent kinase inhibitors p16INK4a, p15INK4b and p21WAF is frequently mediated by promoter gene methylation, whereas histone deacetylases (HDACs) control gene expression through their ability to deacetylate proteins. The effect of suberohydroxamic acid (SBHA) and 5‑Aza‑2'‑deoxycytidine (Decitabine) (DAC) treatments on the transcription of CDKN2A, CDKN2B and CDKN1A genes, and their effects on molecular biological behavior were examined in two myeloma cell lines, RPMI8226 and U266, which differ in p53‑functionality and IL‑6 expression. In both tested myeloma cell lines, a non‑methylated state of the CDKN2B gene promoter region was detected with normal gene expression, and the same level of p15INK4b protein was detected by immunocytochemical staining. Furthermore, in myeloma cells treated with SBHA and DAC alone, the expression of both p15INK4b and p21WAF was significantly upregulated in RPMI8226 cells (p53‑functional, without IL‑6 expression), whereas in the U266 cell line (p53 deleted, expressing IL‑6) only p21WAF expression was significantly increased. Moreover, the analysis revealed that treatment with DAC induced DNMT3B enhancement in U266 cells. In conclusion, in myeloma cells with IL‑6 expression, significantly increased DNMT3B expression indicated the tumorigenic consequences of 5‑Aza‑2'deoxycytidine treatment, which requires careful use in diseases involving epigenetic dysregulation, such as multiple myeloma (MM).
Insights
Suberoylanilide hydroxamic acid (SBHA) and Decitabine (DAC) differentially affect gene expression in multiple myeloma cells. DAC treatment increased DNMT3B in IL-6 expressing cells, suggesting potential tumorigenic effects requiring cautious use in epigenetic dysregulation.
Area of Science:
- Molecular Biology
- Epigenetics
- Cancer Research
Background:
- Gene inactivation of cyclin-dependent kinase inhibitors (e.g., p16INK4a, p15INK4b, p21WAF) is often driven by promoter methylation.
- Histone deacetylases (HDACs) regulate gene expression via protein deacetylation.
- Myeloma cell lines RPMI8226 and U266 exhibit distinct p53 functionality and IL-6 expression, providing a model to study epigenetic drug effects.
Purpose of the Study:
- To investigate the effects of suberohydroxamic acid (SBHA) and Decitabine (DAC) on CDKN2A, CDKN2B, and CDKN1A gene transcription in two myeloma cell lines.
- To analyze the impact of these treatments on the molecular biological behavior of myeloma cells with differing p53 and IL-6 statuses.
- To assess the potential tumorigenic consequences of epigenetic drug treatments in multiple myeloma.
Main Methods:
- Treatment of RPMI8226 (p53-functional, no IL-6) and U266 (p53-deleted, IL-6 expressing) myeloma cell lines with SBHA and DAC.
- Analysis of gene promoter methylation status, specifically for the CDKN2B gene.
- Assessment of p15INK4b and p21WAF protein expression via immunocytochemical staining.
- Evaluation of DNMT3B expression levels following DAC treatment.
Main Results:
- The CDKN2B gene promoter was non-methylated with normal gene expression in both cell lines, and p15INK4b protein levels were consistent.
- SBHA and DAC treatments significantly upregulated p15INK4b and p21WAF in RPMI8226 cells.
- In U266 cells, only p21WAF expression was significantly increased, while DAC treatment induced DNMT3B enhancement.
- The p15INK4b upregulation was not observed in IL-6 expressing U266 cells.
Conclusions:
- Decitabine (DAC) treatment in IL-6 expressing myeloma cells led to increased DNMT3B expression, indicating potential tumorigenic effects.
- The differential response of p15INK4b and p21WAF expression in the two cell lines highlights the role of p53 functionality and IL-6 signaling in epigenetic drug response.
- Caution is advised when using DAC in epigenetic dysregulation diseases like multiple myeloma due to potential adverse outcomes.
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