Effect of α2- Macroglobulin on GPCR-induced Prostate Cancer Cell Proliferation

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Background: α2- Macroglobulin (α2-M) is classified as an acute phase protein since its blood level in animals can be elevated nearly 10-fold by inducing sepsis and it has the ability to complex with and inhibit proteases. Blood levels of α2-M is decreased in prostate cancer (PC), perhaps due to the increased secretion of the protease prostate specific antigen. Stromal cells of the normal human prostate can also synthesize α2-M and this activity is upregulated in patients with benign prostate hyperplasia. During proteinase entrapment, α2-M can become activated as it undergoes a conformational change. Activated α2-M can bind to certain receptors related to the LDL receptor and some studies indicate an ability to induce cellular signaling and the activation of growth pathways. Thus the present study was aimed at examining the effect of α2-M on G protein-coupled receptor (GPCR) agonist-induced prostate cancer cell growth.
Methods: PC cell proliferation was measured by DNA synthesis assay. Release of EGF-like material in the conditioned medium was measured by using a cell-based radioreceptor assay. Western blot was done to see the ERK, AKT and EGFR phosphorylation. Human plasma was separated by chromatography on Biogel A 0.5M, and α2-M protein fractions were identified by SDS gel electrophoresis and western blot analysis.
Results: Human α2-M dose-responsively inhibited basal DNA synthesis in PC3 cells, although it had no effect in LNCaP and MLL cells. In contrast, DNA synthesis induced by neurotensin (NT) or by bombesin (BOM) was inhibited by α2-M in each of these cell lines. Since the EGF receptor is thought to mediate the effects of NT and BOM on DNA synthesis, we also tested the effect of α2-M on the response to EGF. The fact that α2-M did not alter the response to EGF indicated that its inhibitory actions were exerted proximal to the EGF receptor. NT and BOM are known to transactivate the EGF receptor in PC3 cells by stimulating cell-surface shedding of EGF-like substances. α2-M inhibited the ability of NT and BOM to enhance the release of EGF-like material from each of the PC cell-lines. Thus, α2-M exhibited a general ability to disrupt growth factor shedding, a mechanism that is considered important for PC growth in vivo. Since the concentration of α2-M in human plasma (@2 mg/ml) is 100-1000 times that shown here to inhibit DNA synthesis in PC3 cells, we hypothesized that blood levels of α2-M might exert important effects on PC growth. In plasma chromatogram fractions we could demonstrate the growth inhibitory properties expected for α2-M and dose-dependently inhibited NT-induced DNA synthesis in PC3 and LNCaP cells without altering the response to EGF. Furthermore, human α2-M and chromatogram fractions for α2-M inhibited NT/BOM-induced but not EGF-induced ERK, AKT and EGFR phosphorylation in prostate cancer cells.
Conclusions: Altogether we have identified that α2-M can inhibit GPCR agonist-induced prostate cancer cell proliferation and considering the unique features of its protein binding and targeting properties, α2-M may play an important role in regulating benign and malignant prostatic growth.

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