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AZD1480 delays tumor growth in a melanoma model while enhancing the suppressive activity of myeloid-derived suppressor cells.

Maenhout SK, Du Four S, Corthals J, Neyns B, Thielemans K, Aerts JL - Oncotarget (2014)

Bottom Line: Here we investigated the effects of AZD1480 on the function of different immune cell populations in a melanoma model.Moreover, a significant decrease in the percentage of myeloid-derived suppressor cells (MDSCs) was observed after treatment with AZD1480.The addition of AZD1480 to co-cultures of human MDSCs and T cells does not affect the suppressive activity of MDSCs but it does reduce the IFN-γ secretion and the proliferative capacity of T cells.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Molecular and Cellular Therapy, Department of Immunology-Physiology, Vrije Universiteit Brussel, Brussels, Belgium.

ABSTRACT
AZD1480 is a potent, competitive small-molecule inhibitor of JAK1/2 kinase which inhibits STAT3 phosphorylation and tumor growth. Here we investigated the effects of AZD1480 on the function of different immune cell populations in a melanoma model. When MO4 tumor-bearing mice were treated with AZD1480 we observed a strong inhibition of tumor growth as well as a prolonged survival. Moreover, a significant decrease in the percentage of myeloid-derived suppressor cells (MDSCs) was observed after treatment with AZD1480. However, AZD1480 enhanced the suppressive capacity of murine MDSCs while at the same time impairing the proliferative as well as the IFN-γ secretion capacity of murine T cells. The addition of AZD1480 to co-cultures of human MDSCs and T cells does not affect the suppressive activity of MDSCs but it does reduce the IFN-γ secretion and the proliferative capacity of T cells. We showed that although AZD1480 has the ability to delay the tumor growth of MO4 tumor-bearing mice, this drug has detrimental effects on several aspects of the immune system. These data indicate that systemic targeting of the JAK/STAT pathway by JAK1/2 inhibition can have divergent effects on tumor growth and anti-tumor immune responses.

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AZD1480 inhibits P-STAT3 expression but does not induce apoptosis in murine melanoma cell lines in vitroMO4 cells, MO4 cells pretreated with IL-6 (50 ng/ml) and K1735-C4 melanoma cells were treated with the indicated doses of AZD1480 and after 48 hours the percentage of apoptotic cells was determined by flow cytometry using Annexin-V/7-AAD staining. A. Represenative FACS profile of Annexin-V/7-AAD staining of the different cell lines treated with AZD1480. B. Overview of the percentage of viable cells (defined as Annexin-V−/7-AAD− cells) in different mouse melanoma cell lines 48 hours after the addition of different concentrations of AZD1480. Results of 3 independent experiments are shown as mean ± SEM. C. MO4 cells pretreated with IL-6 (50 ng/ml) were treated with the indicated doses of AZD1480 and after 6 hours the level of P-STAT3 expression was determined by western blot. One representative blot of 2 indepenent experiments is shown.
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Figure 1: AZD1480 inhibits P-STAT3 expression but does not induce apoptosis in murine melanoma cell lines in vitroMO4 cells, MO4 cells pretreated with IL-6 (50 ng/ml) and K1735-C4 melanoma cells were treated with the indicated doses of AZD1480 and after 48 hours the percentage of apoptotic cells was determined by flow cytometry using Annexin-V/7-AAD staining. A. Represenative FACS profile of Annexin-V/7-AAD staining of the different cell lines treated with AZD1480. B. Overview of the percentage of viable cells (defined as Annexin-V−/7-AAD− cells) in different mouse melanoma cell lines 48 hours after the addition of different concentrations of AZD1480. Results of 3 independent experiments are shown as mean ± SEM. C. MO4 cells pretreated with IL-6 (50 ng/ml) were treated with the indicated doses of AZD1480 and after 6 hours the level of P-STAT3 expression was determined by western blot. One representative blot of 2 indepenent experiments is shown.

Mentions: Since it is well known that inhibition of STAT3 signalling can induce apoptosis in tumor cells[25] we determined the apoptotic effects of AZD1480 in different murine melanoma cell lines in vitro. The MO4 cell line, which lacks constitutive expression of P-STAT3, was treated with IL-6, in order to induce P-STAT3 expression and the K1735-C4 cell line, characerized by constitutive expression of P-STAT3, was treated with different doses of AZD1480 for 48 hours. Even at the highest concentration we tested (5 μM), AZD1480 failed to induce apoptosis in all tested cell lines as evaluated by Annexin-V/7-AAD staining (Figure 1A,B). However, a dose-dependent downregulation of P-STAT3 expression in MO4 melanoma cells was apparant 6 hours after the treatment with AZD1480. At a concentration of 5 μM, AZD1480 completely abrogated the P-STAT3 expression in IL-6 treated MO4 cells (Figure 1C). Comparable effects were seen in a human melanoma cell line (Supplementary Figure 1) These data demonstrate that AZD1480 is able to inhibit induced activation of P-STAT3 in the MO4 cell line, without affecting the viability of these cells.


AZD1480 delays tumor growth in a melanoma model while enhancing the suppressive activity of myeloid-derived suppressor cells.

Maenhout SK, Du Four S, Corthals J, Neyns B, Thielemans K, Aerts JL - Oncotarget (2014)

AZD1480 inhibits P-STAT3 expression but does not induce apoptosis in murine melanoma cell lines in vitroMO4 cells, MO4 cells pretreated with IL-6 (50 ng/ml) and K1735-C4 melanoma cells were treated with the indicated doses of AZD1480 and after 48 hours the percentage of apoptotic cells was determined by flow cytometry using Annexin-V/7-AAD staining. A. Represenative FACS profile of Annexin-V/7-AAD staining of the different cell lines treated with AZD1480. B. Overview of the percentage of viable cells (defined as Annexin-V−/7-AAD− cells) in different mouse melanoma cell lines 48 hours after the addition of different concentrations of AZD1480. Results of 3 independent experiments are shown as mean ± SEM. C. MO4 cells pretreated with IL-6 (50 ng/ml) were treated with the indicated doses of AZD1480 and after 6 hours the level of P-STAT3 expression was determined by western blot. One representative blot of 2 indepenent experiments is shown.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4196164&req=5

Figure 1: AZD1480 inhibits P-STAT3 expression but does not induce apoptosis in murine melanoma cell lines in vitroMO4 cells, MO4 cells pretreated with IL-6 (50 ng/ml) and K1735-C4 melanoma cells were treated with the indicated doses of AZD1480 and after 48 hours the percentage of apoptotic cells was determined by flow cytometry using Annexin-V/7-AAD staining. A. Represenative FACS profile of Annexin-V/7-AAD staining of the different cell lines treated with AZD1480. B. Overview of the percentage of viable cells (defined as Annexin-V−/7-AAD− cells) in different mouse melanoma cell lines 48 hours after the addition of different concentrations of AZD1480. Results of 3 independent experiments are shown as mean ± SEM. C. MO4 cells pretreated with IL-6 (50 ng/ml) were treated with the indicated doses of AZD1480 and after 6 hours the level of P-STAT3 expression was determined by western blot. One representative blot of 2 indepenent experiments is shown.
Mentions: Since it is well known that inhibition of STAT3 signalling can induce apoptosis in tumor cells[25] we determined the apoptotic effects of AZD1480 in different murine melanoma cell lines in vitro. The MO4 cell line, which lacks constitutive expression of P-STAT3, was treated with IL-6, in order to induce P-STAT3 expression and the K1735-C4 cell line, characerized by constitutive expression of P-STAT3, was treated with different doses of AZD1480 for 48 hours. Even at the highest concentration we tested (5 μM), AZD1480 failed to induce apoptosis in all tested cell lines as evaluated by Annexin-V/7-AAD staining (Figure 1A,B). However, a dose-dependent downregulation of P-STAT3 expression in MO4 melanoma cells was apparant 6 hours after the treatment with AZD1480. At a concentration of 5 μM, AZD1480 completely abrogated the P-STAT3 expression in IL-6 treated MO4 cells (Figure 1C). Comparable effects were seen in a human melanoma cell line (Supplementary Figure 1) These data demonstrate that AZD1480 is able to inhibit induced activation of P-STAT3 in the MO4 cell line, without affecting the viability of these cells.

Bottom Line: Here we investigated the effects of AZD1480 on the function of different immune cell populations in a melanoma model.Moreover, a significant decrease in the percentage of myeloid-derived suppressor cells (MDSCs) was observed after treatment with AZD1480.The addition of AZD1480 to co-cultures of human MDSCs and T cells does not affect the suppressive activity of MDSCs but it does reduce the IFN-γ secretion and the proliferative capacity of T cells.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Molecular and Cellular Therapy, Department of Immunology-Physiology, Vrije Universiteit Brussel, Brussels, Belgium.

ABSTRACT
AZD1480 is a potent, competitive small-molecule inhibitor of JAK1/2 kinase which inhibits STAT3 phosphorylation and tumor growth. Here we investigated the effects of AZD1480 on the function of different immune cell populations in a melanoma model. When MO4 tumor-bearing mice were treated with AZD1480 we observed a strong inhibition of tumor growth as well as a prolonged survival. Moreover, a significant decrease in the percentage of myeloid-derived suppressor cells (MDSCs) was observed after treatment with AZD1480. However, AZD1480 enhanced the suppressive capacity of murine MDSCs while at the same time impairing the proliferative as well as the IFN-γ secretion capacity of murine T cells. The addition of AZD1480 to co-cultures of human MDSCs and T cells does not affect the suppressive activity of MDSCs but it does reduce the IFN-γ secretion and the proliferative capacity of T cells. We showed that although AZD1480 has the ability to delay the tumor growth of MO4 tumor-bearing mice, this drug has detrimental effects on several aspects of the immune system. These data indicate that systemic targeting of the JAK/STAT pathway by JAK1/2 inhibition can have divergent effects on tumor growth and anti-tumor immune responses.

Show MeSH
Related in: MedlinePlus