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Targeted therapy of glioblastoma stem-like cells and tumor non-stem cells using cetuximab-conjugated iron-oxide nanoparticles.

Kaluzova M, Bouras A, Machaidze R, Hadjipanayis CG - Oncotarget (2015)

Bottom Line: Transmission electron microscopy revealed cetuximab-IONP GBM cell binding and internalization.Treatment with cetuximab-IONPs resulted in a significant antitumor effect that was greater than with cetuximab alone due to more efficient, CD133-independent cellular targeting and uptake, EGFR signaling alterations, EGFR internalization, and apoptosis induction in EGFR-expressing GSCs and neurospheres.A significant increase in survival was found after cetuximab-IONP convection-enhanced delivery treatment of 3 intracranial rodent GBM models employing human EGFR-expressing GBM xenografts.

View Article: PubMed Central - PubMed

Affiliation: Brain Tumor Nanotechnology Laboratory, Department of Neurosurgery, Emory University School of Medicine, Winship Cancer Institute of Emory University, Atlanta, GA, USA.

ABSTRACT
Malignant gliomas remain aggressive and lethal primary brain tumors in adults. The epidermal growth factor receptor (EGFR) is frequently overexpressed in the most common malignant glioma, glioblastoma (GBM), and represents an important therapeutic target. GBM stem-like cells (GSCs) present in tumors are felt to be highly tumorigenic and responsible for tumor recurrence. Multifunctional magnetic iron-oxide nanoparticles (IONPs) can be directly imaged by magnetic resonance imaging (MRI) and designed to therapeutically target cancer cells. The targeting effects of IONPs conjugated to the EGFR inhibitor, cetuximab (cetuximab-IONPs), were determined with EGFR- and EGFRvIII-expressing human GBM neurospheres and GSCs. Transmission electron microscopy revealed cetuximab-IONP GBM cell binding and internalization. Fluorescence microscopy and Prussian blue staining showed increased uptake of cetuximab-IONPs by EGFR- as well as EGFRvIII-expressing GSCs and neurospheres in comparison to cetuximab or free IONPs. Treatment with cetuximab-IONPs resulted in a significant antitumor effect that was greater than with cetuximab alone due to more efficient, CD133-independent cellular targeting and uptake, EGFR signaling alterations, EGFR internalization, and apoptosis induction in EGFR-expressing GSCs and neurospheres. A significant increase in survival was found after cetuximab-IONP convection-enhanced delivery treatment of 3 intracranial rodent GBM models employing human EGFR-expressing GBM xenografts.

No MeSH data available.


Related in: MedlinePlus

Apoptosis studies and cell signaling in human GSCs after treatment with cetuximab-IONPsGSCs and GBM CD133-negative neurospheres (5×105 cells) from N08-74 (A) and N08-30 (B) were treated with free IONPs, cetuximab-IONPs, and cetuximab for 3 hs and expression of cleaved caspase 3, caspase 3, cleaved PARP, and PARP was determined by Western blot analysis. (C) Expression of cleaved caspase 9, caspase 9 after 3 h treatment in GSCs and GBM CD133-negative neurospheres (5×105 cells) from N08-30. GSCs and GBM CD133-negative cells from neurospheres N08-74 were treated with free IONPs (0.2 mg/ml), cetuximab-IONPs (0.2 mg/ml), or cetuximab (50 μg/ml) for 3 hs and analyzed by Western blotting with phospho-ERK44/42 and total ERK44/42 antibodies (D) or GSCs from N08-74 for 72 hs (E) and analyzed by Western blotting with phospho-ERK44/42 and β-actin antibodies.
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Figure 6: Apoptosis studies and cell signaling in human GSCs after treatment with cetuximab-IONPsGSCs and GBM CD133-negative neurospheres (5×105 cells) from N08-74 (A) and N08-30 (B) were treated with free IONPs, cetuximab-IONPs, and cetuximab for 3 hs and expression of cleaved caspase 3, caspase 3, cleaved PARP, and PARP was determined by Western blot analysis. (C) Expression of cleaved caspase 9, caspase 9 after 3 h treatment in GSCs and GBM CD133-negative neurospheres (5×105 cells) from N08-30. GSCs and GBM CD133-negative cells from neurospheres N08-74 were treated with free IONPs (0.2 mg/ml), cetuximab-IONPs (0.2 mg/ml), or cetuximab (50 μg/ml) for 3 hs and analyzed by Western blotting with phospho-ERK44/42 and total ERK44/42 antibodies (D) or GSCs from N08-74 for 72 hs (E) and analyzed by Western blotting with phospho-ERK44/42 and β-actin antibodies.

Mentions: Apoptosis and cleavage of caspase 3 was investigated as a mechanism of cell death in GSCs and GBM CD133-negative cells. Both populations from the neurospheres N08-74 and N08-30 were incubated with free IONPs, cetuximab-IONPs, and cetuximab alone. Cetuximab-IONPs induced apoptosis in N08-74 GSCs and GBM CD133-negative cells, as indicated by the presence of cleaved PARP and caspase 3 after treatment for 3 hs (Figure 6A) and cleaved PARP after 6, 8, and 24 hs (data not shown). We did not find any increase in cleaved caspase 3 or PARP after treatment with free IONPs and cetuximab alone. Only cetuximab-IONP-treated N08-30 GSCs underwent apoptosis, as shown by cleaved PARP and caspase 3, whereas cetuximab alone had less of an effect. In CD133-negative GBM cells from N08-30, the opposite was observed: cetuximab alone was more potent than cetuximab-IONPs in inducing cleavage of PARP (Figure 6B).


Targeted therapy of glioblastoma stem-like cells and tumor non-stem cells using cetuximab-conjugated iron-oxide nanoparticles.

Kaluzova M, Bouras A, Machaidze R, Hadjipanayis CG - Oncotarget (2015)

Apoptosis studies and cell signaling in human GSCs after treatment with cetuximab-IONPsGSCs and GBM CD133-negative neurospheres (5×105 cells) from N08-74 (A) and N08-30 (B) were treated with free IONPs, cetuximab-IONPs, and cetuximab for 3 hs and expression of cleaved caspase 3, caspase 3, cleaved PARP, and PARP was determined by Western blot analysis. (C) Expression of cleaved caspase 9, caspase 9 after 3 h treatment in GSCs and GBM CD133-negative neurospheres (5×105 cells) from N08-30. GSCs and GBM CD133-negative cells from neurospheres N08-74 were treated with free IONPs (0.2 mg/ml), cetuximab-IONPs (0.2 mg/ml), or cetuximab (50 μg/ml) for 3 hs and analyzed by Western blotting with phospho-ERK44/42 and total ERK44/42 antibodies (D) or GSCs from N08-74 for 72 hs (E) and analyzed by Western blotting with phospho-ERK44/42 and β-actin antibodies.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4496184&req=5

Figure 6: Apoptosis studies and cell signaling in human GSCs after treatment with cetuximab-IONPsGSCs and GBM CD133-negative neurospheres (5×105 cells) from N08-74 (A) and N08-30 (B) were treated with free IONPs, cetuximab-IONPs, and cetuximab for 3 hs and expression of cleaved caspase 3, caspase 3, cleaved PARP, and PARP was determined by Western blot analysis. (C) Expression of cleaved caspase 9, caspase 9 after 3 h treatment in GSCs and GBM CD133-negative neurospheres (5×105 cells) from N08-30. GSCs and GBM CD133-negative cells from neurospheres N08-74 were treated with free IONPs (0.2 mg/ml), cetuximab-IONPs (0.2 mg/ml), or cetuximab (50 μg/ml) for 3 hs and analyzed by Western blotting with phospho-ERK44/42 and total ERK44/42 antibodies (D) or GSCs from N08-74 for 72 hs (E) and analyzed by Western blotting with phospho-ERK44/42 and β-actin antibodies.
Mentions: Apoptosis and cleavage of caspase 3 was investigated as a mechanism of cell death in GSCs and GBM CD133-negative cells. Both populations from the neurospheres N08-74 and N08-30 were incubated with free IONPs, cetuximab-IONPs, and cetuximab alone. Cetuximab-IONPs induced apoptosis in N08-74 GSCs and GBM CD133-negative cells, as indicated by the presence of cleaved PARP and caspase 3 after treatment for 3 hs (Figure 6A) and cleaved PARP after 6, 8, and 24 hs (data not shown). We did not find any increase in cleaved caspase 3 or PARP after treatment with free IONPs and cetuximab alone. Only cetuximab-IONP-treated N08-30 GSCs underwent apoptosis, as shown by cleaved PARP and caspase 3, whereas cetuximab alone had less of an effect. In CD133-negative GBM cells from N08-30, the opposite was observed: cetuximab alone was more potent than cetuximab-IONPs in inducing cleavage of PARP (Figure 6B).

Bottom Line: Transmission electron microscopy revealed cetuximab-IONP GBM cell binding and internalization.Treatment with cetuximab-IONPs resulted in a significant antitumor effect that was greater than with cetuximab alone due to more efficient, CD133-independent cellular targeting and uptake, EGFR signaling alterations, EGFR internalization, and apoptosis induction in EGFR-expressing GSCs and neurospheres.A significant increase in survival was found after cetuximab-IONP convection-enhanced delivery treatment of 3 intracranial rodent GBM models employing human EGFR-expressing GBM xenografts.

View Article: PubMed Central - PubMed

Affiliation: Brain Tumor Nanotechnology Laboratory, Department of Neurosurgery, Emory University School of Medicine, Winship Cancer Institute of Emory University, Atlanta, GA, USA.

ABSTRACT
Malignant gliomas remain aggressive and lethal primary brain tumors in adults. The epidermal growth factor receptor (EGFR) is frequently overexpressed in the most common malignant glioma, glioblastoma (GBM), and represents an important therapeutic target. GBM stem-like cells (GSCs) present in tumors are felt to be highly tumorigenic and responsible for tumor recurrence. Multifunctional magnetic iron-oxide nanoparticles (IONPs) can be directly imaged by magnetic resonance imaging (MRI) and designed to therapeutically target cancer cells. The targeting effects of IONPs conjugated to the EGFR inhibitor, cetuximab (cetuximab-IONPs), were determined with EGFR- and EGFRvIII-expressing human GBM neurospheres and GSCs. Transmission electron microscopy revealed cetuximab-IONP GBM cell binding and internalization. Fluorescence microscopy and Prussian blue staining showed increased uptake of cetuximab-IONPs by EGFR- as well as EGFRvIII-expressing GSCs and neurospheres in comparison to cetuximab or free IONPs. Treatment with cetuximab-IONPs resulted in a significant antitumor effect that was greater than with cetuximab alone due to more efficient, CD133-independent cellular targeting and uptake, EGFR signaling alterations, EGFR internalization, and apoptosis induction in EGFR-expressing GSCs and neurospheres. A significant increase in survival was found after cetuximab-IONP convection-enhanced delivery treatment of 3 intracranial rodent GBM models employing human EGFR-expressing GBM xenografts.

No MeSH data available.


Related in: MedlinePlus