Tumor-associated myeloid cells, including dendritic cells and macrophages, are immune suppressive. The current study demonstrates a novel mechanism involving FOXO3 and NF-κB RelA that controls myeloid cell signaling and impacts their immune suppressive nature. We find that FOXO3 binds NF-κB RelA in the cytosol, impacting both proteins by preventing FOXO3 degradation and preventing NF-κB RelA nuclear translocation. The location of protein-protein interaction was determined to be near the FOXO3 transactivation domain. In turn, NF-κB RelA activation was restored upon deletion of the same sequence in FOXO3 containing the DNA binding domain. We have identified for the first time, a direct protein-protein interaction between FOXO3 and NF-κB RelA in tumor-associated dendritic cells. These detailed biochemical interactions provide the foundation for future studies to utilize the FOXO3-NF-κB RelA interaction as a target to enhance tumor-associated dendritic cell function to support or enhance anti-tumor immunity.

T regulatory cells (Treg) avert autoimmunity but their increased levels in melanoma confer a poor prognosis. To explore the basis for Treg accumulation in melanoma, we evaluated chemokine expression in patients. A 5-fold increase was documented in the Treg chemoattractants CCL22 and CCL1 in melanoma-affected skin versus unaffected skin, as accompanied by infiltrating FoxP3+ T cells. In parallel, there was a ~2-fold enhancement in expression of CCR4 in circulating Treg but not T effector cells. We hypothesized that redirecting Treg away from tumors might suppress autoimmune side-effects caused by immune checkpoint therapeutics now used widely in the clinic. In assessing this hypothesis, we observed a marked increase in skin Treg in mice vaccinated with CCL22, with repetitive vaccination sufficient to limit Treg accumulation and melanoma growth in the lungs of animals challenged by tumor cell injection, whether using a prevention or treatment protocol design. The observed change in Treg accumulation in this setting could not be explained by Treg conversion. Overall, our findings offered a preclinical proof of concept for the potential use of CCL22 delivered by local injection as a strategy to enhance the efficacious response to immune checkpoint therapy while suppressing its autoimmune side-effects.

Tumors evade immune recognition and destruction in many ways including the creation of an immune-suppressive tumor microenvironment (TME). Dendritic cells (DC) that infiltrate the TME are tolerogenic, and suppress effector T cells and anti-tumor activity. Previous reports demonstrated that a key regulator of tolerance in DC is the transcription factor FOXO3. Gender disparity has been studied in cancer in relation to incidence, aggressiveness, and prognosis. Few studies have touched on the importance in relation to impact on the immune system. In the current study, we show that there are significant differences in tumor growth between males and females. Additionally, frequencies and the function of FOXO3 expressed by DC subsets that infiltrate tumors vary between genders. Our results show for the first time that DC FOXO3 expression and function is altered in females. In vitro results indicate that these differences may be the result of exposure to estrogen. These differences may be critical considerations for the enhancement of immunotherapy for cancer.

Sex-specific differences exist in innate and adaptive immune responses and are mediated by hormone signaling. Estrogen is able to differentially modulate the development and differentiation of immune cells, including T cells. However, the effect of estrogen on T cell function, especially at concentrations other than physiological, remains controversial and incompletely understood. Immunotherapy is one of the most promising cancer treatments to date with a high probability of future enhancements. The adoptive transfer of genetically modified T cells can mediate tumor regression but there are still many hurdles to enhancing the proficiency of this treatment. This study demonstrates for the first time that one major aspect to consider for designing potent immunotherapies for cancer is the impact of the patient's sex. Herein, using two different Ag-specific T cell groups, we investigated the effect of sex and estrogen in antitumor effector responses, T helper cytokine secretion, and, importantly, on T cell whole polyfunctionality important for memory T cell development and survival. Major differences were observed in T cell function and polyfunctionality between sexes and on E2 treatment. The findings of this study may be critical to understand the results of immunotherapy on different patients and for the enhancement of immunotherapy for cancer.

Background/Aim. Irreversible electroporation (IRE) showed promising results for small-size tumors and very early cancers. However, further development is needed to evolve this procedure into a more efficient ablation technique for long-term control of tumor growth. In this work, we show that it is possible to increase the antitumor efficiency of IRE by simmultaneously injecting c-di-GMP, a STING agonist, intratumorally. Materials and Methods. Intratumoral administration of c-di-GMP simultaneously to IRE was evaluated in murine models of melanona (B16.OVA) and hepatocellular carcinoma (PM299L). Results. The combined therapy increased the number of tumor-infiltrating IFN-γ/TNF-α-producing CD4 and CD8 T cells and delayed tumor growth, as compared to the effect observed in groups treated with c-di-GMP or IRE alone. Conclusion. These results can lead to the development of a new therapeutic strategy for the treatment of cancer patients refractory to other therapies.