Limits...
Insulin-like-growth-factor-binding-protein-3 (IGFBP-3) contrasts melanoma progression in vitro and in vivo.

Naspi A, Panasiti V, Abbate F, Roberti V, Devirgiliis V, Curzio M, Borghi M, Lozupone F, Carotti S, Morini S, Gaudio E, Calvieri S, Londei P - PLoS ONE (2014)

Bottom Line: Insulin-like-factor-binding-protein 3 (IGFBP-3) is known to modulate the activity of insulin-like growth factors (IGFs) besides having a number of IGF-independent effects on cell growth and survival.In the present work, we have evaluated the levels of IGFBP-3 in the blood serum and tissues of patients affected by cutaneous melanoma, showing that loss of IGFBP-3 from both is strongly correlated with disease progression and reduced survival.In summary, IGFBP-3 appears to exert a specific inhibitory effect on melanoma growth and dissemination, suggesting that it may qualify as a useful therapeutic agent in melanomas and perhaps other cancers, at the least as a valid adjuvant therapy during treatment with conventional anti-tumoral drugs.

View Article: PubMed Central - PubMed

Affiliation: Istituto Pasteur-Fondazione Cenci-Bolognetti, Dpt. Biotecnologie Cellulari ed Ematologia, University of Rome Sapienza, Rome, Italy.

ABSTRACT
Insulin-like-factor-binding-protein 3 (IGFBP-3) is known to modulate the activity of insulin-like growth factors (IGFs) besides having a number of IGF-independent effects on cell growth and survival. IGFBP-3 has been reported to decrease significantly in the blood serum of patients affected by certain cancers. In the present work, we have evaluated the levels of IGFBP-3 in the blood serum and tissues of patients affected by cutaneous melanoma, showing that loss of IGFBP-3 from both is strongly correlated with disease progression and reduced survival. In vitro treatment with IGFBP-3 of human and murine metastatic melanoma cell lines specifically inhibited the cells' migratory and invasive behaviour, inducing up-regulation of melanocytic differentiation markers such as tyrosinase activity and melanin content. A molecular analysis of the cellular pathways transducing the effect of IGFBP-3 implicated the Akt-GSK3β axis. Moreover, administration of IGFBP-3 in vivo to SCID mice inoculated with human metastatic melanoma cells strongly reduced or completely inhibited tumor growth. In summary, IGFBP-3 appears to exert a specific inhibitory effect on melanoma growth and dissemination, suggesting that it may qualify as a useful therapeutic agent in melanomas and perhaps other cancers, at the least as a valid adjuvant therapy during treatment with conventional anti-tumoral drugs.

Show MeSH

Related in: MedlinePlus

Different expression of MMP-9 in primary and metastatic melanoma.(A) Detection of MMP-2 and MMP-9 activity in the culture media of WM793 and Me501 cells by zymographic assay. The arrows point to the areas of degradation of the gelatine matrix produced by the indicated proteases (B) Western blot analysis showing degradation over time of IGFBP-3 by melanoma-produced proteases. Human recombinant IGFBP-3 was incubated for the indicated times with the culture media of the primary melanoma line WM793 or of the metastatic line Me501, in the absence (96 hrs) or in the presence (96i hrs) of protease inhibitors. A Red Ponceau staining of the original gel is shown on the bottom as the loading control. (C,D) MMP-9 immunostaining of primary (C) and metastatic (D) melanoma. Asterisks indicate melanocytes nests and the arrows indicate stromal cells. Original magnification, X100. (E) Comparison of the MMP-9 reactivity score of primary and metastatic melanoma. Central boxes represent values from lower to upper quartile (25th–75th percentile). Middle lines represent median. Vertical lines extend from minimum to maximum value. P value was calculated by Mann-Whitney U test. The reactive cells were counted, and the scores determined, as described in Fig. 1 and in the Methods section.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4048209&req=5

pone-0098641-g002: Different expression of MMP-9 in primary and metastatic melanoma.(A) Detection of MMP-2 and MMP-9 activity in the culture media of WM793 and Me501 cells by zymographic assay. The arrows point to the areas of degradation of the gelatine matrix produced by the indicated proteases (B) Western blot analysis showing degradation over time of IGFBP-3 by melanoma-produced proteases. Human recombinant IGFBP-3 was incubated for the indicated times with the culture media of the primary melanoma line WM793 or of the metastatic line Me501, in the absence (96 hrs) or in the presence (96i hrs) of protease inhibitors. A Red Ponceau staining of the original gel is shown on the bottom as the loading control. (C,D) MMP-9 immunostaining of primary (C) and metastatic (D) melanoma. Asterisks indicate melanocytes nests and the arrows indicate stromal cells. Original magnification, X100. (E) Comparison of the MMP-9 reactivity score of primary and metastatic melanoma. Central boxes represent values from lower to upper quartile (25th–75th percentile). Middle lines represent median. Vertical lines extend from minimum to maximum value. P value was calculated by Mann-Whitney U test. The reactive cells were counted, and the scores determined, as described in Fig. 1 and in the Methods section.

Mentions: The progression of primary melanoma to metastatic disease may be influenced by the IGFBP-3 in the tissutal microenvironment. Most of the circulating IGFBP-3 is produced by the liver; however, the protein is also secreted by several other inflammatory and mesenchymal cell types and by melanocytes themselves. Notably, an inverse correlation has been observed between the amount of IGFBP-3 produced by melanoma tissue and the metastatic capacity of the cells [24]. To evaluate the presence of IGFBP-3 in the immediate environs of the tumour, immuno-histochemical analyses of primary and metastatic tumour samples taken from patients were performed. As shown in Fig. 1 (D,G), a diffuse intracellular IGFBP-3 staining was observed in primary melanomas as well as in peri-tumoral stromal cells including monocytes/macrophages, lymphocytes, granulocytes and fibroblasts. In metastatic tumours IGFBP-3 staining was much weaker or absent (Fig. 1 E,F,H,I). Previously, we hypothesized that IGFBP-3 loss in both blood and tissues could be accounted for by degradation by tumour-produced proteases. This was further investigated. Firstly, secretion of proteases by cultured primary and metastatic melanoma cells was evaluated by zymography. As shown in Fig. 2A, the primary melanoma line WM793 secreted only small amounts of a gelatinase (presumably metalloprotease-2) (MMP-2). By contrast, the metastatic cells Me501 produced large amounts of proteases, especially MMP-2 and metalloprotease-9 (MMP-9). Accordingly, the culture media of metastatic, but not primary, cells caused extensive degradation of recombinant IGFBP-3 in vitro. (Fig. 2B). This result agrees with our previous finding that recombinant IGFBP-3 was degraded upon incubation with blood serum from stage-IV melanoma patients [15].


Insulin-like-growth-factor-binding-protein-3 (IGFBP-3) contrasts melanoma progression in vitro and in vivo.

Naspi A, Panasiti V, Abbate F, Roberti V, Devirgiliis V, Curzio M, Borghi M, Lozupone F, Carotti S, Morini S, Gaudio E, Calvieri S, Londei P - PLoS ONE (2014)

Different expression of MMP-9 in primary and metastatic melanoma.(A) Detection of MMP-2 and MMP-9 activity in the culture media of WM793 and Me501 cells by zymographic assay. The arrows point to the areas of degradation of the gelatine matrix produced by the indicated proteases (B) Western blot analysis showing degradation over time of IGFBP-3 by melanoma-produced proteases. Human recombinant IGFBP-3 was incubated for the indicated times with the culture media of the primary melanoma line WM793 or of the metastatic line Me501, in the absence (96 hrs) or in the presence (96i hrs) of protease inhibitors. A Red Ponceau staining of the original gel is shown on the bottom as the loading control. (C,D) MMP-9 immunostaining of primary (C) and metastatic (D) melanoma. Asterisks indicate melanocytes nests and the arrows indicate stromal cells. Original magnification, X100. (E) Comparison of the MMP-9 reactivity score of primary and metastatic melanoma. Central boxes represent values from lower to upper quartile (25th–75th percentile). Middle lines represent median. Vertical lines extend from minimum to maximum value. P value was calculated by Mann-Whitney U test. The reactive cells were counted, and the scores determined, as described in Fig. 1 and in the Methods section.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4048209&req=5

pone-0098641-g002: Different expression of MMP-9 in primary and metastatic melanoma.(A) Detection of MMP-2 and MMP-9 activity in the culture media of WM793 and Me501 cells by zymographic assay. The arrows point to the areas of degradation of the gelatine matrix produced by the indicated proteases (B) Western blot analysis showing degradation over time of IGFBP-3 by melanoma-produced proteases. Human recombinant IGFBP-3 was incubated for the indicated times with the culture media of the primary melanoma line WM793 or of the metastatic line Me501, in the absence (96 hrs) or in the presence (96i hrs) of protease inhibitors. A Red Ponceau staining of the original gel is shown on the bottom as the loading control. (C,D) MMP-9 immunostaining of primary (C) and metastatic (D) melanoma. Asterisks indicate melanocytes nests and the arrows indicate stromal cells. Original magnification, X100. (E) Comparison of the MMP-9 reactivity score of primary and metastatic melanoma. Central boxes represent values from lower to upper quartile (25th–75th percentile). Middle lines represent median. Vertical lines extend from minimum to maximum value. P value was calculated by Mann-Whitney U test. The reactive cells were counted, and the scores determined, as described in Fig. 1 and in the Methods section.
Mentions: The progression of primary melanoma to metastatic disease may be influenced by the IGFBP-3 in the tissutal microenvironment. Most of the circulating IGFBP-3 is produced by the liver; however, the protein is also secreted by several other inflammatory and mesenchymal cell types and by melanocytes themselves. Notably, an inverse correlation has been observed between the amount of IGFBP-3 produced by melanoma tissue and the metastatic capacity of the cells [24]. To evaluate the presence of IGFBP-3 in the immediate environs of the tumour, immuno-histochemical analyses of primary and metastatic tumour samples taken from patients were performed. As shown in Fig. 1 (D,G), a diffuse intracellular IGFBP-3 staining was observed in primary melanomas as well as in peri-tumoral stromal cells including monocytes/macrophages, lymphocytes, granulocytes and fibroblasts. In metastatic tumours IGFBP-3 staining was much weaker or absent (Fig. 1 E,F,H,I). Previously, we hypothesized that IGFBP-3 loss in both blood and tissues could be accounted for by degradation by tumour-produced proteases. This was further investigated. Firstly, secretion of proteases by cultured primary and metastatic melanoma cells was evaluated by zymography. As shown in Fig. 2A, the primary melanoma line WM793 secreted only small amounts of a gelatinase (presumably metalloprotease-2) (MMP-2). By contrast, the metastatic cells Me501 produced large amounts of proteases, especially MMP-2 and metalloprotease-9 (MMP-9). Accordingly, the culture media of metastatic, but not primary, cells caused extensive degradation of recombinant IGFBP-3 in vitro. (Fig. 2B). This result agrees with our previous finding that recombinant IGFBP-3 was degraded upon incubation with blood serum from stage-IV melanoma patients [15].

Bottom Line: Insulin-like-factor-binding-protein 3 (IGFBP-3) is known to modulate the activity of insulin-like growth factors (IGFs) besides having a number of IGF-independent effects on cell growth and survival.In the present work, we have evaluated the levels of IGFBP-3 in the blood serum and tissues of patients affected by cutaneous melanoma, showing that loss of IGFBP-3 from both is strongly correlated with disease progression and reduced survival.In summary, IGFBP-3 appears to exert a specific inhibitory effect on melanoma growth and dissemination, suggesting that it may qualify as a useful therapeutic agent in melanomas and perhaps other cancers, at the least as a valid adjuvant therapy during treatment with conventional anti-tumoral drugs.

View Article: PubMed Central - PubMed

Affiliation: Istituto Pasteur-Fondazione Cenci-Bolognetti, Dpt. Biotecnologie Cellulari ed Ematologia, University of Rome Sapienza, Rome, Italy.

ABSTRACT
Insulin-like-factor-binding-protein 3 (IGFBP-3) is known to modulate the activity of insulin-like growth factors (IGFs) besides having a number of IGF-independent effects on cell growth and survival. IGFBP-3 has been reported to decrease significantly in the blood serum of patients affected by certain cancers. In the present work, we have evaluated the levels of IGFBP-3 in the blood serum and tissues of patients affected by cutaneous melanoma, showing that loss of IGFBP-3 from both is strongly correlated with disease progression and reduced survival. In vitro treatment with IGFBP-3 of human and murine metastatic melanoma cell lines specifically inhibited the cells' migratory and invasive behaviour, inducing up-regulation of melanocytic differentiation markers such as tyrosinase activity and melanin content. A molecular analysis of the cellular pathways transducing the effect of IGFBP-3 implicated the Akt-GSK3β axis. Moreover, administration of IGFBP-3 in vivo to SCID mice inoculated with human metastatic melanoma cells strongly reduced or completely inhibited tumor growth. In summary, IGFBP-3 appears to exert a specific inhibitory effect on melanoma growth and dissemination, suggesting that it may qualify as a useful therapeutic agent in melanomas and perhaps other cancers, at the least as a valid adjuvant therapy during treatment with conventional anti-tumoral drugs.

Show MeSH
Related in: MedlinePlus