Prostate cancer (PCa) is the second most common cancer in men and the fifth leading cause of cancer death. PCa is a complicated disease that develops and progresses across time, with prostate tissue inflammation playing an important role. Several pro-inflammatory mediators are associated with tissue inflammation in PCa including the cytokine TNF-α. The progression of prostate cancer (PCa) has been shown to depend on a variety of signalling pathways including the phosphatidylinositol 3-kinase PI3K/Akt pathway, phosphorylation of glycogen synthase kinase 3-beta (GSK-3β ) and Proto-oncogene tyrosine-protein kinase Src signalling (c-Src or Src). The PI3K-Akt pathway is one of the most frequently over-activated intracellular pathways in tumour signalling as it regulates cell survival processes including apoptosis and proliferation. Glycogen synthase kinase-3β (GSK-3β) is a serine/ threonine kinase that plays a significant role in the progression of PCa that promotes cell proliferation and survival. SFKs comprise nine structurally similar non-receptor protein tyrosine kinases including Src, Fyn, Lyn, Yes, Blk, Lck, Hck, Fgr, and Yrk and some SFKs are overexpressed during PCa tumorigenesis and progression. In the current study, we aimed to further investigate the potential therapeutic effects of morphine against prostate cancer by studying its potential cytotoxic and anti-inflammatory properties using the in vitro PCa (PC3).

PCa cells (PC3) were treated with various concentrations of morphine (0.5-2.0  μmol/L), and an MTT assay was used to explore the cytotoxic effect of various concentrations of morphine (0.5-2.5 μmol/L), naloxone, combined naloxone and morphine, and β-endorphin (10-60 pg/ml) on the PC3 cells at 24, 48, and 72-hour incubation periods. In addition, an ELISA assay was used to assess the release of the pro-inflammatory cytokines IL-6, IL-8 and IL-1 β by the PC3 cells upon their incubation with TNF-α (10 ng/ml) for 24 hours with or without pre-incubation with morphine. Finally, the associated intracellular signalling events were assessed using a specific Proteome Profiler Human Phospho-Kinase Array Kit.

Interestingly, various concentrations of morphine exerted a significant cytotoxic effect on PC3 cells, which was maximal at 1.0 μmol/L concentration (Figure 1(A)). Moreover, 24-hour incubation with TNF-α induced significant release of the cytokines IL-1β, IL-6 and IL-8 by PC3 cells. Such release was significantly inhibited by pre-incubation with 1.0 μmol/L of morphine (Figure 1(B1-B3)). In addition, with 2, 4 and 6 hours of incubation, TNF-α induced a significant increase in the phosphorylation levels of the protein kinases JNK1/2/3 (T183/Y185, T221/Y223), Lck (Y394), Akt1/2/3 (S473), p53 (S15), p70 S6 Kinase (T421/S424), RSK1/2 (S221/S227), GSK-3β (S9) and RSK1/2 (S221/S227). These signalling events were significantly inhibited upon pre-incubation with 1.0 μmol/L of morphine (Figure 2).

Interestingly, various concentrations of morphine exerted a significant cytotoxic effect on PC3 cells, which was maximal at 0.5 μmol/L concentration. Moreover, 24-hour incubation with TNF-α induced significant release of the cytokines IL-1β, IL-6 and IL-8 by PC3 cells. In addition, upon 2, 4 and 6 hours of incubation, TNF-α induced a significant increase in the phosphorylation levels of the protein kinases JNK1/2/3 (T183/Y185, T221/Y223), Lck (Y394), Akt1/2/3 (S473), p53 (S15), p70 S6 Kinase (T421/S424), RSK1/2 (S221/S227), GSK-3β (S9) and RSK1/2 (S221/S227).

The current study demonstrates preliminary evidence suggesting that morphine could be used in PCa patients without oncological detriment.