Overexpression of the anti-apoptotic protein BCL-2 is frequently observed in multiple malignancies, including about 85% of patients with estrogen receptor positive (ER⁺) breast cancer. Besides being studied as a prognostic marker, BCL-2 is investigated as a therapeutic target in ER⁺ breast cancer. Here, we introduce a new exosome-based strategy to target BCL-2 using genetically modified natural killer (NK) cells. The NK cell line NK92MI was lentivirally transduced to express and load BCL-2 siRNAs (siBCL-2) into exosomes (NKExos) and then evaluated for its potential to treat ER⁺ breast cancer. Transfected NK92MI cells produced substantial levels of BCL-2 siRNAs, without substantially affecting NK cell viability or effector function and led to loading of siBCL-2 in NKExos. Remarkably, targeting BCL-2 via siBCL-2 NKExos led to enhanced intrinsic apoptosis in breast cancer cells, without affecting non-malignant cells. Together, our prototypical results for BCL-2 in breast cancer provide proof of concept for a novel strategy to utilize NKExos as a natural delivery vector for siRNA targeting of oncogenes.

Vascular endothelial growth factor (VEGF) is the important angiogenic factor associated with tumor growth and metastasis in a wide variety of solid tumors. The aim of this study is to investigate the tumor suppressive effect of chitosan/small interfering RNA (siRNA)-VEGF nanoplexes in the rat breast cancer model. Chitosan/siRNA nanoplexes (siVEGF-A, siVEGFR-1, siVEGFR-2) and NRP-1 were prepared in a 15 to1 ratio and injected (intratumorally) into the breast-tumor-bearing Sprague-Dawley rats. Tumor volumes were measured during 21 days. To investigate the effect of chitosan/siRNA nanoplexes on VEGF expression in tumors, VEGF was analyzed with immunohistochemistry and western blotting. The mRNA levels of VEGF in tumor samples were determined with real-time PCR (RT-PCR). After siRNA treatment, a marked reduction in tumor volumes was measured in complex-injected rats (97%). Free siRNA injection showed lower tumor inhibition. Reduction of VEGF protein was also shown with western blotting and immunohistochemistry. Similar results were obtained with RT-PCR also. These results indicate that the chitosan/siRNA targeting to VEGF nanoplexes have a remarkably suppressive effect on VEGF expression and tumor volume in breast cancer model of rats.

The 3D polymeric network structure of hydrogels imitates the extracellular matrix, thereby facilitating cell growth and differentiation. In the current study, chitosan/hyaluronic acid/honey coacervate hydrogels were produced without any chemicals or crosslinking agents and investigated for their wound-healing abilities. Chitosan/hyaluronic acid/honey hydrogels were characterized by FTIR, SEM, and rheology analysis. Moreover, their water content, water uptake capacities, and porosity were investigated. In FT-IR spectra, it was discovered that the characteristic band placement of chitosan with hyaluronic acid changed upon interacting with honey. The porosity of the honey-containing hydrogels (12%) decreased compared to those without honey (17%). Additionally, the water-uptake capacity of honey-containing hydrogels slightly decreased. Also, it was observed that hydrogels’ viscosity increased with the increased hyaluronic acid amount and decreased with the amount of honey. The adhesion and proliferation of fibroblast cells on the surface of hydrogel formulations were highest in honey-containing hydrogels (144%). In in vivo studies, wound healing was accelerated by honey addition. It has been demonstrated for the first time that honey-loaded chitosan-hyaluronic acid hydrogels, prepared without the use of toxic covalent crosslinkers, have potential for use in wound healing applications.

Vascular endothelial growth factor (VEGF) plays a critical role in angiogenesis. The aim of this study was to use chitosan/short hairpin VEGF (shVEGF) [short hairpin RNA (shRNA)-expressing pDNA targeting VEGF-A] complexes in the treatment of rat breast cancer model. Therefore, chitosan/shVEGF complexes were prepared in (2/1) ratio and injected to the breast-tumor bearing Sprague-Dawley rats. Intratumoral and intraperitoneal injections were applied and compared. Tumor volumes were measured during the 36 days. To investigate the effect of complexes on the VEGF expression, VEGF were analyzed by immunohistochemistry and western blotting. The mRNA levels of VEGF were determined by real-time polymerase chain reaction. Tumor volume decreased at the end of experiments after shRNA treatment. The highest suppression in the tumor volume was observed after intratumoral complex injection to rats (96%). Compared with intratumoral and intraperitoneal injection, higher tumor inhibition was obtained with intratumoral injection. Free shRNA injection indicated lower tumor suppression. The immunohistochemistry and western blotting results correlated with the real-time polymerase chain reaction and tumor volume measurements. The data suggest that chitosan/shVEGF complexes can be used to inhibit tumor growth in breast carcinoma model of rats.