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Caspase 3 promotes surviving melanoma tumor cell growth after cytotoxic therapy.

Donato AL, Huang Q, Liu X, Li F, Zimmerman MA, Li CY - J. Invest. Dermatol. (2014)

Bottom Line: In this study, we aim to define the role of caspase 3 in melanoma tumor growth after cytotoxic therapy.Furthermore, we observed that caspase 3 gene knockdown attenuated the growth-stimulating effect of irradiated, dying cells on living melanoma cell growth.Finally, we showed that caspase 3-mediated dying melanoma cell stimulation of living cell growth involves secreted prostaglandin E2 (PGE2).

View Article: PubMed Central - PubMed

Affiliation: 1] Department of Dermatology, Duke University School of Medicine, Durham, North Carolina, USA [2] Howard Hughes Medical Institute, Chevy Chase, Maryland, USA.

ABSTRACT
Metastatic melanoma often relapses despite cytotoxic treatment, and hence the understanding of melanoma tumor repopulation is crucial for improving our current therapies. In this study, we aim to define the role of caspase 3 in melanoma tumor growth after cytotoxic therapy. We examined a paradigm-changing hypothesis that dying melanoma cells undergoing apoptosis during cytotoxic treatment activate paracrine signaling events that promote the growth of surviving tumor cells. We propose that caspase 3 has a key role in the initiation of the release of signals from dying cells to stimulate melanoma tumor growth. We created a model for tumor cell repopulation in which a small number of luciferase-labeled, untreated melanoma cells are seeded onto a layer of a larger number of unlabeled, lethally treated melanoma cells. We found that dying melanoma cells significantly stimulate the growth of living melanoma cells in vitro and in vivo. Furthermore, we observed that caspase 3 gene knockdown attenuated the growth-stimulating effect of irradiated, dying cells on living melanoma cell growth. Finally, we showed that caspase 3-mediated dying melanoma cell stimulation of living cell growth involves secreted prostaglandin E2 (PGE2). Our study therefore suggests a counterintuitive strategy to inhibit caspase 3 for therapeutic gain in melanoma treatment.

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Caspase 3 regulates radiation-induced PGE2 secretion and indomethacin decreases dying melanoma cell stimulated living cell growthMelanoma cells were irradiated and PGE2 concentrations were measured in the supernatant using an ELISA assay for PGE2 72 hours after radiation. A) Graph showing PGE2 concentration per cell number in irradiated or untreated A375 and A375 C3shRNA cells. B) Graph showing PGE2 concentration per cell in irradiated or untreated A375 cells treated with indomethacin 25 or 50 μM. C) Graph showing reporter luciferase activity vs. time when seeded onto irradiated A375 cells treated with indomethacin 25, 50 or 100 μM with luminescence images on day 10, Error bars are mean +/− SEM, n= 3. *p < 0.05, one-way ANOVA test.
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Figure 5: Caspase 3 regulates radiation-induced PGE2 secretion and indomethacin decreases dying melanoma cell stimulated living cell growthMelanoma cells were irradiated and PGE2 concentrations were measured in the supernatant using an ELISA assay for PGE2 72 hours after radiation. A) Graph showing PGE2 concentration per cell number in irradiated or untreated A375 and A375 C3shRNA cells. B) Graph showing PGE2 concentration per cell in irradiated or untreated A375 cells treated with indomethacin 25 or 50 μM. C) Graph showing reporter luciferase activity vs. time when seeded onto irradiated A375 cells treated with indomethacin 25, 50 or 100 μM with luminescence images on day 10, Error bars are mean +/− SEM, n= 3. *p < 0.05, one-way ANOVA test.

Mentions: Given that we previously identified PGE2 as the downstream secreted factor from dying cell stimulated living cell growth in breast cancer cells (Huang et al., 2011), we sought to determine if there is a role for PGE2 in a caspase 3-mediated mechanism of melanoma tumor repopulation. We measured PGE2 production induced by irradiation of melanoma cells in the supernatant of cells and found that PGE2 levels are increased after radiation of melanoma cells (Figure 5a & 5b). In comparison with the supernatant of irradiated wild-type A375 cells, PGE2 levels were significantly lower in irradiated A375 caspase 3 shRNA cells (Figure 5a) or after treatment with indomethacin (a cyclooxygenase I and II inhibitor that should significantly reduce PGE2 production) (Figure 5b). Further, treatment of irradiated A375 cells with 100 μM indomethacin in a transwell plate decreased the growth-stimulating effect of dying melanoma cells on living melanoma cells by greater than 32-fold (Figure 5c). Finally, we also obtained evidence that PGE2 was induced by radiation in A2508 cells and its production was decreased with indomethacin (supplementary Figure S5a). The importance of PGE2 was again demonstrated by that fact that indomethacin significantly reduced cell death stimulated A2508Fluc repopulation (supplementary Figure S5b). Therefore, we conclude that PGE2 secretion is crucial to caspase 3-mediated dying melanoma cell stimulation of living cell growth.


Caspase 3 promotes surviving melanoma tumor cell growth after cytotoxic therapy.

Donato AL, Huang Q, Liu X, Li F, Zimmerman MA, Li CY - J. Invest. Dermatol. (2014)

Caspase 3 regulates radiation-induced PGE2 secretion and indomethacin decreases dying melanoma cell stimulated living cell growthMelanoma cells were irradiated and PGE2 concentrations were measured in the supernatant using an ELISA assay for PGE2 72 hours after radiation. A) Graph showing PGE2 concentration per cell number in irradiated or untreated A375 and A375 C3shRNA cells. B) Graph showing PGE2 concentration per cell in irradiated or untreated A375 cells treated with indomethacin 25 or 50 μM. C) Graph showing reporter luciferase activity vs. time when seeded onto irradiated A375 cells treated with indomethacin 25, 50 or 100 μM with luminescence images on day 10, Error bars are mean +/− SEM, n= 3. *p < 0.05, one-way ANOVA test.
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Figure 5: Caspase 3 regulates radiation-induced PGE2 secretion and indomethacin decreases dying melanoma cell stimulated living cell growthMelanoma cells were irradiated and PGE2 concentrations were measured in the supernatant using an ELISA assay for PGE2 72 hours after radiation. A) Graph showing PGE2 concentration per cell number in irradiated or untreated A375 and A375 C3shRNA cells. B) Graph showing PGE2 concentration per cell in irradiated or untreated A375 cells treated with indomethacin 25 or 50 μM. C) Graph showing reporter luciferase activity vs. time when seeded onto irradiated A375 cells treated with indomethacin 25, 50 or 100 μM with luminescence images on day 10, Error bars are mean +/− SEM, n= 3. *p < 0.05, one-way ANOVA test.
Mentions: Given that we previously identified PGE2 as the downstream secreted factor from dying cell stimulated living cell growth in breast cancer cells (Huang et al., 2011), we sought to determine if there is a role for PGE2 in a caspase 3-mediated mechanism of melanoma tumor repopulation. We measured PGE2 production induced by irradiation of melanoma cells in the supernatant of cells and found that PGE2 levels are increased after radiation of melanoma cells (Figure 5a & 5b). In comparison with the supernatant of irradiated wild-type A375 cells, PGE2 levels were significantly lower in irradiated A375 caspase 3 shRNA cells (Figure 5a) or after treatment with indomethacin (a cyclooxygenase I and II inhibitor that should significantly reduce PGE2 production) (Figure 5b). Further, treatment of irradiated A375 cells with 100 μM indomethacin in a transwell plate decreased the growth-stimulating effect of dying melanoma cells on living melanoma cells by greater than 32-fold (Figure 5c). Finally, we also obtained evidence that PGE2 was induced by radiation in A2508 cells and its production was decreased with indomethacin (supplementary Figure S5a). The importance of PGE2 was again demonstrated by that fact that indomethacin significantly reduced cell death stimulated A2508Fluc repopulation (supplementary Figure S5b). Therefore, we conclude that PGE2 secretion is crucial to caspase 3-mediated dying melanoma cell stimulation of living cell growth.

Bottom Line: In this study, we aim to define the role of caspase 3 in melanoma tumor growth after cytotoxic therapy.Furthermore, we observed that caspase 3 gene knockdown attenuated the growth-stimulating effect of irradiated, dying cells on living melanoma cell growth.Finally, we showed that caspase 3-mediated dying melanoma cell stimulation of living cell growth involves secreted prostaglandin E2 (PGE2).

View Article: PubMed Central - PubMed

Affiliation: 1] Department of Dermatology, Duke University School of Medicine, Durham, North Carolina, USA [2] Howard Hughes Medical Institute, Chevy Chase, Maryland, USA.

ABSTRACT
Metastatic melanoma often relapses despite cytotoxic treatment, and hence the understanding of melanoma tumor repopulation is crucial for improving our current therapies. In this study, we aim to define the role of caspase 3 in melanoma tumor growth after cytotoxic therapy. We examined a paradigm-changing hypothesis that dying melanoma cells undergoing apoptosis during cytotoxic treatment activate paracrine signaling events that promote the growth of surviving tumor cells. We propose that caspase 3 has a key role in the initiation of the release of signals from dying cells to stimulate melanoma tumor growth. We created a model for tumor cell repopulation in which a small number of luciferase-labeled, untreated melanoma cells are seeded onto a layer of a larger number of unlabeled, lethally treated melanoma cells. We found that dying melanoma cells significantly stimulate the growth of living melanoma cells in vitro and in vivo. Furthermore, we observed that caspase 3 gene knockdown attenuated the growth-stimulating effect of irradiated, dying cells on living melanoma cell growth. Finally, we showed that caspase 3-mediated dying melanoma cell stimulation of living cell growth involves secreted prostaglandin E2 (PGE2). Our study therefore suggests a counterintuitive strategy to inhibit caspase 3 for therapeutic gain in melanoma treatment.

Show MeSH
Related in: MedlinePlus