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Published on: November 19, 2019
C16 ceramide accumulates following androgen ablation in LNCaP prostate cancer cells
Masatoshi Eto1, Jaafar Bennouna, Oriana C Hunter
1Department of Surgery/Division of Biological Therapeutics, University of Pittsburgh School of Medicine, University of Pittsburgh Cancer Institute, Pittsburgh, Pennsylvania 15219, USA.
Background:
Adenocarcinoma of the prostate is the most frequently diagnosed non-cutaneous cancer and the second leading cause of cancer-related deaths among men in the United States. The most successful therapies to date for this tumor have involved some form of androgen ablation. However, these therapies become ineffective as the tumor evolves to an androgen-insensitive state. Ceramide is a lipid second messenger that has been shown to mediate growth arrest or cell death when added exogenously to prostate cancer cells. As a first step toward understanding the events that lead to the transition of prostate cancer cells to an androgen-independent state, we considered investigating the effect of androgen ablation on endogenous ceramide levels in androgen-sensitive and androgen-insensitive prostate cancer cells.
Methods:
To investigate the mechanisms of growth arrest/apoptosis in androgen-sensitive (LNCaP) and insensitive (DU-145, PC-3) cells, we used various methods including nonyl acridine orange (NAO) staining, propidium iodide (PI) staining/cell-cycle analysis, lipid analysis, and Western blotting assays.
Results:
In this study, we demonstrate that androgen ablation drives G(0)/G(1)-phase cell-cycle arrest followed by progressive apoptosis in vitro, in LNCaP cells. Lipid analysis indicated an increase in C16 ceramide, which was generated via the de novo pathway as revealed by blockade of ceramide synthase by fumonisin B1. The addition of 5alpha-dihydrotestosterone (DHT) or fumonisin B1 rescued LNCaP cells from apoptosis induced by androgen ablation, and decreased levels of intracellular C16 ceramide. Neither apoptosis nor an increase in C16 ceramide was observed in androgen-independent cell lines following androgen ablation.
Insights
Androgen ablation in prostate cancer cells triggers cell cycle arrest and apoptosis, linked to increased C16 ceramide levels. Blocking ceramide production or adding androgens prevents this effect in sensitive cells.
Area of Science:
- Oncology
- Molecular Biology
- Biochemistry
Background:
- Prostate adenocarcinoma is a leading cancer in men, often treated with androgen ablation.
- Androgen ablation therapies eventually fail as tumors become androgen-insensitive.
- Ceramide, a lipid second messenger, induces growth arrest and apoptosis in prostate cancer cells.
Purpose of the Study:
- To investigate the impact of androgen ablation on endogenous ceramide levels.
- To understand the transition of prostate cancer cells to an androgen-independent state.
Main Methods:
- Cell cycle analysis using propidium iodide (PI) staining.
- Lipid analysis to quantify ceramide levels.
- Western blotting and nonyl acridine orange (NAO) staining.
Main Results:
- Androgen ablation induced G(0)/G(1) cell-cycle arrest and apoptosis in androgen-sensitive LNCaP cells.
- Lipid analysis revealed increased C16 ceramide via the de novo pathway.
- 5alpha-dihydrotestosterone (DHT) or fumonisin B1 treatment rescued cells from apoptosis and reduced C16 ceramide.
Conclusions:
- Androgen ablation increases C16 ceramide, leading to apoptosis in androgen-sensitive prostate cancer cells.
- The de novo pathway is involved in C16 ceramide generation.
- Ceramide modulation may offer therapeutic strategies for prostate cancer.
