In the last 20 years, dendritic cells (DCs) have been largely used as a platform for therapeutic vaccination in cancer patients. However, despite its proven safety and ability to induce cancer specific immune responses, the clinical benefits of DC-based immunotherapy are currently very limited. Thus, novel approaches are still needed to boost its efficacy. Our group recently showed that squaric acid treatment of antigens is an important adjuvant that can increase vaccine-induced downstream immune responses and therapeutic outcomes. Here we further improved this dendritic cell vaccine formulation by developing a new method for differentiating and maturing DCs from their bone marrow precursors. Our data demonstrate that bone marrow-derived DCs differentiated with GM-CSF and IL-15 and matured with a maturation cocktail in two steps present a more mature and immunogenic phenotype, compared to standard DC preparations. Further suppression of the prostaglandin E2 pathway achieved even more immunogenic DC phenotypes. This vaccine was more potent at delaying tumor growth, improved animal survival and induced a more immunogenic and Th1-skewed T cell response in an ovarian cancer mouse model. These promising results support future efforts for the clinical translation of this approach.