Dissertação de Mestrado Investigação Biomédica 2024 Faculdade de Ciências Médicas, Universidade NOVA de Lisboa

Abstract Breast cancer (BC) is the leading cause of cancer-related deaths in women worldwide, largely due to metastasis. Immunotherapy is an emerging treatment for metastatic BC (mBC), although, its success is currently limited. The resistance to these therapies is mainly imposed by the tumor microenvironment (TME). Neutrophils play a significant role within the TME, where they can exhibit either antitumor (N1) or protumor (N2) functions. Interestingly, circulating neutrophils, divided into high density neutrophils (HDN) and low density neutrophils (LDN), functionally mirror N1 and N2 cells, respectively. Recently, there has been growing interest in targeting LDN for the development of novel immunotherapies, as they can promote cancer progression, potentiate metastasis, and accumulate in the blood of critically ill patients. However, a great challenge of neutrophil-targeted approaches is the discrimination of the pathological neutrophils from the remaining neutrophil populations. Thus, one promising avenue of research is the identification of particular subsets of LDN, which have been shown to modulate cancer progression and the efficacy of immunotherapies, in order to unveil possible biomarkers and/or therapeutic targets. Interestingly, we identified a novel subset of neutrophils that express CCR4, a chemokine receptor never described in these cells before. This new subset was found mainly within the population of LDN, exhibits increased immunosuppressive features, and is prevalent in the blood of mBC patients. Moreover, we demonstrated that CCR4+LDN can efficiently impair the activation of T lymphocytes and present enhanced migratory ability towards the chemokines CCL17 and CCL22, which may promote their migration to the tumor/metastasis sites. Also, we observed that these chemokines are increased in the blood of mBC patients, therefore, suggesting the relevance of the CCR4-expressing LDN - CCL17/22 axis in cancer progression. Concerning the clinical impact of these neutrophils, we verified that they are correlated with a decreased life expectancy of mBC patients. Altogether, the obtained results suggest that this recently identified subset of LDN may play a critical role in the progression of BC, contributing to a worse prognosis and potentially impairing patients’ response to therapies. Therefore, we anticipate the potential of CCR4+LDN either as clinically meaningful biomarkers of BC prognosis and/or as novel therapeutic targets for advanced BC. Ultimately, the findings resulting from this thesis’ work will contribute to improving mBC patients’ care and the survival rate of this disease