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Published on: March 6, 2018
Characterization of calcium-independent purinergic receptor-mediated apoptosis in hormone-refractory prostate cancer
Majid Shabbir1, Mina Ryten, Cecil Thompson
1Autonomic Neuroscience Centre, Royal Free and University College Medical School, London, UK.
Objective:
To investigate the nature of purinergic signalling in hormone-refractory prostate cancer (HRPC) cells in vitro, as extracellular ATP inhibits the growth of HRPC in vitro via the activation of P2 purinergic receptors, and to characterize which P2 receptors subtypes and secondary mechanisms are involved.
Materials And Methods:
The effect of extracellular ATP on HRPC cell lines PC-3 and DU-145, and the normal prostate cell line PNT-2, were investigated. Reverse-transcription polymerase chain reaction was used to assess P2 purinergic receptors, which were pharmacologically characterized using various receptor agonists and antagonists. The effect of ATP on intracellular Ca(2+) concentration ([Ca(2+)](i)) was examined to asses its role in growth inhibition. The effect of combining ATP with the chemotherapeutic drug mitoxantrone was also assessed.
Results:
PC-3 cells expressed mRNA for P2X(4,5,7), P2Y(1,2,4,6); DU-145 cells expressed mRNA for P2X(4,5), P2Y(1,2,4,6,11); PNT-2 cells expressed mRNA for P2X(4,5,7) and P2Y(1,2,4,6,11). ATP (10(-4)m) inhibited HRPC PC-3 cell growth by approximately 90%, an effect partially inhibited by the nonselective P2 receptor antagonists pyridoxal-5'-phosphate-6-azophenyl-2',4' disulphonic acid (PPADS) and suramin. The order of potency of agonists was: adenosine 5'-O-(3 thiotriphosphate) > ATP > benzoyl benzoyl ATP >> 2-methylthio ATP. DU-145 cells responded similarly. Pharmacological profiling implicated P2X(5) and/or P2Y(11) receptors in the antineoplastic response in HRPC. ATP induced apoptosis in a [Ca(2+)](i)-independent mechanism. ATP was significantly less effective on PNT-2 cells, which also had a different order of agonist potency. ATP combined with mitoxantrone in an additive manner in HRPC.
Conclusions:
ATP effectively reduces growth of HRPC cells via calcium-independent apoptosis. Pharmacological profiling indicates P2X(5) and/or P2Y(11) receptors in this process, with a different functional purinergic receptor profile and sensitivity in normal vs cancer cells.
Insights
Extracellular adenosine triphosphate (ATP) inhibits hormone-refractory prostate cancer (HRPC) cell growth by inducing apoptosis. Specific P2 purinergic receptors, P2X5 and P2Y11, are implicated in this anti-cancer effect.
Area of Science:
- Cell biology
- Molecular pharmacology
- Cancer research
Background:
- Prostate cancer (PC) progression to hormone-refractory (HRPC) status presents significant therapeutic challenges.
- Purinergic signaling, mediated by extracellular nucleotides like adenosine triphosphate (ATP), plays a role in cellular processes, including cancer cell growth.
Purpose of the Study:
- To investigate the role of purinergic signaling in hormone-refractory prostate cancer (HRPC) cells.
- To identify specific P2 purinergic receptor subtypes and downstream mechanisms involved in ATP-mediated inhibition of HRPC growth.
Main Methods:
- Investigated the effects of extracellular ATP on HRPC cell lines (PC-3, DU-145) and a normal prostate cell line (PNT-2).
- Utilized reverse-transcription polymerase chain reaction (RT-PCR) to detect P2 receptor expression.
- Pharmacologically characterized receptor subtypes using agonists and antagonists.
- Assessed intracellular calcium concentration ([Ca(2+)](i)) and apoptosis induction.
- Evaluated the combined effect of ATP and mitoxantrone.
Main Results:
- HRPC cell lines (PC-3, DU-145) expressed various P2X and P2Y receptor subtypes.
- Extracellular ATP significantly inhibited HRPC cell growth (approx. 90% in PC-3 cells) via a calcium-independent apoptosis mechanism.
- Pharmacological profiling suggested involvement of P2X5 and/or P2Y11 receptors in the anti-cancer effect.
- ATP demonstrated reduced efficacy on normal prostate cells (PNT-2) with a distinct receptor profile.
- ATP and mitoxantrone exhibited an additive effect in HRPC cells.
Conclusions:
- Extracellular ATP effectively reduces HRPC cell growth through calcium-independent apoptosis.
- P2X5 and/or P2Y11 receptors are key mediators of this anti-neoplastic response in HRPC.
- Normal prostate cells exhibit differential purinergic receptor sensitivity compared to cancer cells.
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