Cancer Res Treat.  2017 Apr;49(2):322-329. 10.4143/crt.2016.091.

Blocking Interleukin-4 Receptor α Using Polyethylene Glycol Functionalized Superparamagnetic Iron Oxide Nanocarriers to Inhibit Breast Cancer Cell Proliferation

• Shaik AP
• Shaik AS
• Majwal AA
• Faraj AA

• KMID: 2378104
• DOI: http://doi.org/10.4143/crt.2016.091

Abstract

PURPOSE
The specific targeting of interleukin-4 receptor α (IL4Rα) receptor offers a promising therapeutic approach for inhibition of tumor cell progression in breast cancer patients. In the current study, the in vitro efficacy of superparamagnetic iron oxide nanoparticles conjugated with anti-IL4Rα blocking antibodies (SPION-IL4Rα) via polyethylene glycol polymers was evaluated in 4T1 breast cancer cells.
MATERIALS AND METHODS
Cell viability, reactive oxygen species generation, and apoptosis frequency were assessed in vitro in 4T1 cancer cell lines following exposure to SPION-IL4Rα alone or combined with doxorubicin. In addition, immunofluorescence assessments and fluorimetrywere performed to confirm the specific targeting and interaction of the developed nanocarriers with IL4Rα receptors in breast cancer cells.
RESULTS
Blocking of IL4Rα receptors caused a significant decrease in cell viability and induced apoptosis in 4T1 cells. In addition, combined treatment with SPION-IL4Rα+doxorubicin caused significant increases in cell death, apoptosis, and oxidative stress compared to either SPION-IL4Rα or doxorubicin alone, indicating the enhanced therapeutic efficacy of this combination. The decrease in fluorescence intensity upon immunofluorescence and fluorimetry assays combined with increased viability and decreased apoptosis following the blocking of IL4Rα receptors confirmed the successful binding of the synthesized nanocarriers to the target sites on murine 4T1 breast cancerous cells.
CONCLUSION
These results suggest that SPION-IL4Rα nanocarriers might be used for successfulreduction of tumor growth and inhibition of progression of metastasis in vivo.

Keyword

Breast neoplasms; Dextran coated superparamagnetic iron oxide; Biomarkers; Tumor; Receptors; Interleukin-4; Drug delivery systems

MeSH Terms

Antibodies, Blocking
Apoptosis
Biomarkers
Breast Neoplasms*
Breast*
Cell Death
Cell Line
Cell Proliferation*
Cell Survival
Doxorubicin
Drug Delivery Systems
Fluorescence
Fluorescent Antibody Technique
Fluorometry
Humans
In Vitro Techniques
Interleukin-4*
Iron*
Nanoparticles
Neoplasm Metastasis
Oxidative Stress
Polyethylene Glycols*
Polyethylene*
Polymers
Reactive Oxygen Species
Antibodies, Blocking
Biomarkers
Doxorubicin
Interleukin-4
Iron
Polyethylene
Polyethylene Glycols
Polymers
Reactive Oxygen Species

Figure

• 

  Fig. 1. Cell viability and oxidative stress generation assessments of the in vitro effects of SPION-IL4Rα, DOX, or combined SPION-IL4Rα+DOX. MTT assay (A) and TBARS assay (B) were performed to assess cell viability and oxidative stress generation, respectively, in 4T1 cancer cells. Analyses were performed after incubation for 24, 48, and 72 hours. Data are expressed as the mean±standard error, *p < 0.05, **p < 0.01, ***p < 0.001. NT, no treatment; SPION-IL4Rα, superparamagnetic iron oxide nanoparticles conjugated with anti–interleukin-4 receptor α [IL4Rα] blocking antibodies; DOX, doxorubicin; MTT, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrasodium bromide; TBARS, thiobarbituric acid reactive substances.

• 

  Fig. 2. Assessments of apoptosis caused by SPION-IL4Rα, DOX, and SPION-IL4Rα+DOX. Analyses were performed after incubation for 24, 48, and 72 hours. Data are expressed as the mean±standard error, *p < 0.05, **p < 0.01, ***p < 0.001. NT, no treatment; SPION-IL4Rα, superparamagnetic iron oxide nanoparticles conjugated with anti–interleukin-4 receptor α [IL4Rα] blocking antibodies; DOX, doxorubicin.

• 

  Fig. 3. In vitro interaction studies of SPION-IL4Rα with IL4Rα receptors on 4T1 cells. Fluorescence intensity decreased upon blocking of receptors with anti-IL4Rα antibodies (A) while higher fluorescence intensity was observed upon blocking using isotype IgG antibody (B). (C) Fluorimetry data showing the difference in fluorescence intensity after blocking with either the anti-IL4Rα antibody or isotype IgG antibody (i.e., without blocking). All immunofluorescence images were captured at the same fluorescence exposure time of 300 milliseconds. Blocking was performed with anti-IL4Rα antibodies. FITC-conjugated SPION-IL4Rα were used to show the expression and appear in green; blue staining indicates staining of nuclei with DAPI. Data are expressed as the mean±standard error, ***p < 0.001. SPION-IL4Rα, superparamagnetic iron oxide nanoparticles conjugated with anti–interleukin-4 receptor α [IL4Rα] blocking antibodies; FITC, fluorescein isothiocyanate.

• 

  Fig. 4. Assessments of cell viability (A) and apoptosis (B) following prior blocking of IL4Rα receptors on 4T1 cells. Blocking was performed for 1 hour with either anti-IL4Rα antibody or isotype IgG antibody (i.e., without blocking) and assessments were performed after 12 hours of incubation following treatment with SPION-IgG1, anti-IL4Rα antibodies alone, SPION-IL4Rα, DOX, and SPION-IL4Rα+DOX. Data are expressed as the mean±standard error, *p < 0.05, **p < 0.01, ***p < 0.001. NT, no treatment; SPION-IL4Rα, superparamagnetic iron oxide nanoparticles conjugated with anti–interleukin-4 receptor α [IL4Rα] blocking antibodies; DOX, doxorubicin.

Reference

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