Introduction:: In 2020, cancer is still accountable for around 10 millions of deaths worldwide despite years of research. The rapid proliferation of abnormal cells and invasion of other body parts and organs contributes to the cancer lethality. While surgery and chemotherapy can remove tumor and cancer cells from patients, the therapeutic side effects such as pain and hair loss hinders patients life routine. Therefore, effective methods to treat cancers are in great need.
Dendritic cells (DCs) play an important role in the human immune system to fight cancer. The dendritic cells present antigens which can activate T cells with three distinctive signals for eliminating antigen specific cancer cells. Thus, it is possible to fight against cancer cells by engineering dendritic cells with polymer-based biomaterials. Here, we use alginate hydrogel which the pore surfaces are coated with chemokines such as granulocyte-macrophage colony-stimulating factor (GM-CSF) which recruit Dendritic cells. When the dendritic cells are migrated to the materials, the presence of antigens and adjuvants can further activate DCs via the uptake process.
Materials and Methods:: The DC-recruiting alginate hydrogel is fabricated by mixing the GM-CSF, porogen beads, and alginate hydrogel derived from medium viscous alginate salt. The porogen beads are then bleached under oven heating and leaves pores inside Alginate hydrogel. The surface morphology of the fabricated porous hydrogel was analyzed using Scanning Electron Microscopy.
For in vitro studies, bone marrow derived DCs (BMDCs) were isolated and differentiated from C57BL/6 mice. The DCs are then transfected with Ovalbumin (OVA) mRNA by Lipofectamine 2000 in the porous alginate hydrogel for 2 days. The DCs antigen presentation and activation are detected by OVA257-264 (SIINFEKL) with APC, and the DC population is quantified through FACS assay. In addition, the OT-1 cells are prepared from spleen of mice with mouse Tcra-V2 and Tcrb-V5 genes. The OVA mRNA transfected DCs are then cocultured with CFSE-stained OT-1 cells, and the OT-1 cell proliferation index was determined by FACS Assay.
For in vivo studies, EG.7-OVA cells are subcutaneously injected to C57BL/6 as a tumor model. The GM-CSF coated porous alginate hydrogel was then subcutaneously implanted on the lymph node for DC recruiting. The tumor efficacy was determined by comparing the tumor volume and survival rate between alginate hydrogel treated groups and untreated group for 90 days.
Results, Conclusions, and Discussions:: First, the porosity of the Alginate hydrogel is verified by SEM image. The SEM image shows the pore size of alginate hydrogel is around XXX which fits DCs recuiting mechanism. Then, we confirmed the OVA mRNA transfection with different Lipofectamine 2000 concentrations. The SIINFEKL positive population of the best performing group is 20% higher than the PBS group and the mRNA only group. Moreover, the OVA-mRNA transfected DCs leads to higher proliferation index than the non-treated DCs group. More data will be collected for in vivo study and upon the presentation at BMES conference.
Despite the in vitro transfection of OVA mRNA with lipofectamine 2000 showed comparable transfection efficiency than the non-treated group. The in-vitro transfection efficiency. More conclusions and discussions will be included after examining the in vivo tumor treatment efficacy for the actual presentation at BMES conference.
