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Iron-free and iron-saturated bovine lactoferrin inhibit survivin expression and differentially modulate apoptosis in breast cancer.

Gibbons JA, Kanwar JR, Kanwar RK - BMC Cancer (2015)

Bottom Line: Iron binding, naturally occurring protein bovine lactoferrin (bLf) has attracted attention as a safe anti-cancer agent capable of inducing apoptosis.Apo-bLf induced significantly greater cytotoxicity and reduction in cell proliferation in both cancer cells showing a time and dose dependent effect.Key apoptotic molecules including p53, Bcl-2 family proteins, IAP members and their inhibitors were significantly modulated by both forms of bLf, though differentially in each cell line.

View Article: PubMed Central - PubMed

Affiliation: Nanomedicine - Laboratory for Immunology and Molecular Biomedical Research, Molecular and Medical Research Facility, School of Medicine, Faculty of Health, Deakin University, Geelong, Victoria, Australia. jgib@deakin.edu.au.

ABSTRACT

Background: Iron binding, naturally occurring protein bovine lactoferrin (bLf) has attracted attention as a safe anti-cancer agent capable of inducing apoptosis. Naturally, bLf exists partially saturated (15-20%) with Fe(3+) however, it has been demonstrated that manipulating the saturation state can enhance bLf's anti-cancer activities.

Methods: Apo-bLf (Fe(3+) free) and Fe-bLf (>90% Fe(3+) Saturated) were therefore, tested in MDA-MB-231 and MCF-7 human breast cancer cells in terms of cytotoxicity, proliferation, migration and invasion. Annexin-V Fluos staining was also employed in addition to apoptotic protein arrays and Western blotting to determine the specific mechanism of bLf-induced apoptosis with a key focus on p53 and inhibitor of apoptosis proteins (IAP), specifically survivin.

Results: Apo-bLf induced significantly greater cytotoxicity and reduction in cell proliferation in both cancer cells showing a time and dose dependent effect. Importantly, no cytotoxicity was detected in normal MCF-10-2A cells. Both forms of bLf significantly reduced cell invasion in cancer cells. Key apoptotic molecules including p53, Bcl-2 family proteins, IAP members and their inhibitors were significantly modulated by both forms of bLf, though differentially in each cell line. Most interestingly, both Apo-bLf and Fe-bLf completely inhibited the expression of survivin protein (key IAP), after 48 h at 30 and 40 nM in cancer cells.

Conclusions: The capacity of these forms of bLf to target survivin expression and modulation of apoptosis demonstrates an exciting potential for bLf as an anti-cancer therapeutic in the existing void of survivin inhibitors, with a lack of successful inhibitors in the clinical management of cancer.

No MeSH data available.


Related in: MedlinePlus

Cytotoxicity and proliferation in MDA-MB-231, MCF-7 and MCF-10-2A cells following Apo-bLf and Fe-bLf treatments. Lactate dehydrogenase assay (LDH) results demonstrating cytotoxicity in cells after 24 and 48 h Apo and Fe-bLf treatments in MDA-MB-231 (a), MCF-7 (b) and MCF-10-2A cells (e). CyQUANT® assay results represent cell proliferation levels after 24 and 48 h Apo and Fe-bLf treatments in MDA-MB-231 (c) and MCF-7 cells (d) including high (20% FBS) and low (untreated) controls. Data represented as mean with + SEM (n = 6). * = p < 0.05 compared with untreated (0 nM) group, ** = p < 0.01 compared with untreated (0 nM). Statistical analysis performed using Student’s t-test
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Fig1: Cytotoxicity and proliferation in MDA-MB-231, MCF-7 and MCF-10-2A cells following Apo-bLf and Fe-bLf treatments. Lactate dehydrogenase assay (LDH) results demonstrating cytotoxicity in cells after 24 and 48 h Apo and Fe-bLf treatments in MDA-MB-231 (a), MCF-7 (b) and MCF-10-2A cells (e). CyQUANT® assay results represent cell proliferation levels after 24 and 48 h Apo and Fe-bLf treatments in MDA-MB-231 (c) and MCF-7 cells (d) including high (20% FBS) and low (untreated) controls. Data represented as mean with + SEM (n = 6). * = p < 0.05 compared with untreated (0 nM) group, ** = p < 0.01 compared with untreated (0 nM). Statistical analysis performed using Student’s t-test

Mentions: Apo-bLf was the most effective at inducing cell cytotoxicity with significant increases in cytotoxicity at concentrations of 20, 30 and 40 nM in MDA-MB-231 (133.05%, p = 0.003, 112.53%, p = 0.003 and 110.17%, p = 0.007, respectively) and MCF-7 cells (68.58%, p = 0.001, 90.68%, p = 0.00001 and 99.31%, p = 0.000006, respectively) after 48 h (Fig. 1a). Furthermore, Apo-bLf demonstrated significant toxicity after 24 h in MCF-7 cells at concentrations of 20 nM (25.93%, p = 0.02), 30 nM (40.09%, p = 0.001) and 40 nM (61.97%, p = 0.001) (Fig. 1b). Fe-bLf demonstrated significant increases in cytotoxicity in MDA-MB-231 cells at concentrations of 10, 20 and 30 nM (51.81%, p = 0.04, 61.76%, p = 0.01 and 42.01%, p = 0.048) after 48 h (Fig. 1a) yet no significant cytotoxicity in MCF-7 cells (Fig. 1b).Fig. 1


Iron-free and iron-saturated bovine lactoferrin inhibit survivin expression and differentially modulate apoptosis in breast cancer.

Gibbons JA, Kanwar JR, Kanwar RK - BMC Cancer (2015)

Cytotoxicity and proliferation in MDA-MB-231, MCF-7 and MCF-10-2A cells following Apo-bLf and Fe-bLf treatments. Lactate dehydrogenase assay (LDH) results demonstrating cytotoxicity in cells after 24 and 48 h Apo and Fe-bLf treatments in MDA-MB-231 (a), MCF-7 (b) and MCF-10-2A cells (e). CyQUANT® assay results represent cell proliferation levels after 24 and 48 h Apo and Fe-bLf treatments in MDA-MB-231 (c) and MCF-7 cells (d) including high (20% FBS) and low (untreated) controls. Data represented as mean with + SEM (n = 6). * = p < 0.05 compared with untreated (0 nM) group, ** = p < 0.01 compared with untreated (0 nM). Statistical analysis performed using Student’s t-test
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4440599&req=5

Fig1: Cytotoxicity and proliferation in MDA-MB-231, MCF-7 and MCF-10-2A cells following Apo-bLf and Fe-bLf treatments. Lactate dehydrogenase assay (LDH) results demonstrating cytotoxicity in cells after 24 and 48 h Apo and Fe-bLf treatments in MDA-MB-231 (a), MCF-7 (b) and MCF-10-2A cells (e). CyQUANT® assay results represent cell proliferation levels after 24 and 48 h Apo and Fe-bLf treatments in MDA-MB-231 (c) and MCF-7 cells (d) including high (20% FBS) and low (untreated) controls. Data represented as mean with + SEM (n = 6). * = p < 0.05 compared with untreated (0 nM) group, ** = p < 0.01 compared with untreated (0 nM). Statistical analysis performed using Student’s t-test
Mentions: Apo-bLf was the most effective at inducing cell cytotoxicity with significant increases in cytotoxicity at concentrations of 20, 30 and 40 nM in MDA-MB-231 (133.05%, p = 0.003, 112.53%, p = 0.003 and 110.17%, p = 0.007, respectively) and MCF-7 cells (68.58%, p = 0.001, 90.68%, p = 0.00001 and 99.31%, p = 0.000006, respectively) after 48 h (Fig. 1a). Furthermore, Apo-bLf demonstrated significant toxicity after 24 h in MCF-7 cells at concentrations of 20 nM (25.93%, p = 0.02), 30 nM (40.09%, p = 0.001) and 40 nM (61.97%, p = 0.001) (Fig. 1b). Fe-bLf demonstrated significant increases in cytotoxicity in MDA-MB-231 cells at concentrations of 10, 20 and 30 nM (51.81%, p = 0.04, 61.76%, p = 0.01 and 42.01%, p = 0.048) after 48 h (Fig. 1a) yet no significant cytotoxicity in MCF-7 cells (Fig. 1b).Fig. 1

Bottom Line: Iron binding, naturally occurring protein bovine lactoferrin (bLf) has attracted attention as a safe anti-cancer agent capable of inducing apoptosis.Apo-bLf induced significantly greater cytotoxicity and reduction in cell proliferation in both cancer cells showing a time and dose dependent effect.Key apoptotic molecules including p53, Bcl-2 family proteins, IAP members and their inhibitors were significantly modulated by both forms of bLf, though differentially in each cell line.

View Article: PubMed Central - PubMed

Affiliation: Nanomedicine - Laboratory for Immunology and Molecular Biomedical Research, Molecular and Medical Research Facility, School of Medicine, Faculty of Health, Deakin University, Geelong, Victoria, Australia. jgib@deakin.edu.au.

ABSTRACT

Background: Iron binding, naturally occurring protein bovine lactoferrin (bLf) has attracted attention as a safe anti-cancer agent capable of inducing apoptosis. Naturally, bLf exists partially saturated (15-20%) with Fe(3+) however, it has been demonstrated that manipulating the saturation state can enhance bLf's anti-cancer activities.

Methods: Apo-bLf (Fe(3+) free) and Fe-bLf (>90% Fe(3+) Saturated) were therefore, tested in MDA-MB-231 and MCF-7 human breast cancer cells in terms of cytotoxicity, proliferation, migration and invasion. Annexin-V Fluos staining was also employed in addition to apoptotic protein arrays and Western blotting to determine the specific mechanism of bLf-induced apoptosis with a key focus on p53 and inhibitor of apoptosis proteins (IAP), specifically survivin.

Results: Apo-bLf induced significantly greater cytotoxicity and reduction in cell proliferation in both cancer cells showing a time and dose dependent effect. Importantly, no cytotoxicity was detected in normal MCF-10-2A cells. Both forms of bLf significantly reduced cell invasion in cancer cells. Key apoptotic molecules including p53, Bcl-2 family proteins, IAP members and their inhibitors were significantly modulated by both forms of bLf, though differentially in each cell line. Most interestingly, both Apo-bLf and Fe-bLf completely inhibited the expression of survivin protein (key IAP), after 48 h at 30 and 40 nM in cancer cells.

Conclusions: The capacity of these forms of bLf to target survivin expression and modulation of apoptosis demonstrates an exciting potential for bLf as an anti-cancer therapeutic in the existing void of survivin inhibitors, with a lack of successful inhibitors in the clinical management of cancer.

No MeSH data available.


Related in: MedlinePlus