Investigation of the immunostimulatory effect of human dendritic cells induced by formulations containing human breast cancer cell lysate and different adjuvants on T lymphocytes

Breast cancer is the most common cancer among women in both developing and developed countries. According to the World Health Organization, in 2018, breast cancer, which is considered to constitute 24.2% of the cancers in women worldwide, ranks first among all cancer cases. Treatment of breast cancer mainly consists of surgery, chemotherapy and radiotherapy. However, these treatments also harm the healthy cells of the patient, can not prevent the recurrence of the tumor and can not extend the survival period sufficiently. In recent years, the giving importance to cancer vaccines has increased with the understanding of therapeutic potential of immunotherapy in the cancer cases. There are different cancer vaccine approaches such as peptide-based vaccines all tumor cell vaccines and genetically based vaccines, but the expected results could not be obtained because they could not activate the patients' weak immune systems. For this reason, the interest in dendritic cell-based vaccine approaches, which are the professional antigen presenting cells, has increased in recent years. The first immunotherapeutic vaccine approved by the FDA, the Sipuleucel-T (Provenge) vaccine, has proven the success of dendritic cell-based cancer vaccines in immunotherapy. This xiv success, which has been achieved by using dendritic cell based vaccines against prostate cancer, has increased the interest in breast cancer in recent years. The basis of dendritic cell-based vaccines consist of dendritic cell-based vaccine studies which are loaded with various antigens, such as Her-2, Muc-1, loaded with killed or freeze-thawed tumor lysates or fused with tumor cells. Although the Lapuleucel-T vaccine, which is prepared by loading Her2 peptide on dendritic cells, shows that studies in this area are promising, but its efficacy is limited only on the types of cancer that express Her2. However, until now, the reasons of not gaining expected results of dendritic cell based vaccine studies against breast cancer are that breast cancer's highly heterogeneous antigenic structure, the fusion technique is immunosuppressive and toxic, and no addition of an effective adjuvant. Therefore, it is very important to develop new vaccine formulations that do not have the disadvantages of current vaccines, but using dendritic cell based which are the most promising among the available approaches. The aim of this thesis is the development of whole tumor cell lysates and various adjuvants-induced dendritic cell-based vaccine formulations against breast cancer and investigating the efficancy of them on T lymphocytes. In order to achieve this goal, autologous dendritic cells were differentiated by using GM-CSF and IL-4 cytokines from peripheral blood mononuclear cells isolated from donors' blood. Human breast cancer cells (MCF-7) were cultured and all antigens were isolated by various methods. Dendritic cells were loaded with antigen and each of the Saponin, Polioxidonium, CFA and CpG-ODN adjuvants. Obtained dendritic cells were incubated with T lymphocytes and in vitro immunostimulatory activities were compared. As a result of the analysis, the differentiation of peripheral blood mononuclear cells into dendritic cells has been shown morphologically. Immunophenotypic analyzes of dendritic cells loaded with various formulations have shown that adjuvants increase the expression of CD83 maturation and CD80 costimulatory molecules, but have less effect on CD40 and CD86 helper molecules. However, it has been determined that when the induced dendritic cells are cocultured with T lymphocytes, a high amount of cytokines is secreted in the culture supernatant. The highest IL-12p70 cytokine release was obtained with the vaccine formulation using 100 μg/ml POX adjuvant, while the highest IFN-gamma cytokine release was achieved with the vaccine formulation using 100 μg/ml Saponin adjuvant. In addition, 100 μg/ml POX adjuvant was found to significantly reduce IL-10 and IL-4 immunosuppression cytokines. Furthermore, it has been found that dendritic cells induced by the vaccine formulation using lysate and 100 μg/ml POX adjuvant stimulate CTL transformation in T lymphocytes and these T lymphocytes kill MCF-7 cells by 80%. As a result, within the scope of FYL-2017-3202 project supported by YTU BAP Unit, a new generation of dendritic cell-based vaccines induced by various formulations have been developed for the first time in the world and in our country against breast cancer. These results indicate that breast cancer antigens and adjuvant-induced dendritic cells induce T lymphocytes at a high level and therefore combination of adjuvants use may be appropriate for immunotherapy in breast cancer.