NLRP3 inflammasome and progression of Oral Squamous Cell Carcinoma (OSCC): differential influence in neoplastic and immune cells

• Grant number: 22/05120-3
• Support Opportunities: Scholarships in Brazil - Doctorate
• Start date: August 01, 2023
• End date: January 31, 2026
• Field of knowledge: Health Sciences - Dentistry
• Principal Investigator: Carlos Rossa Junior
• Grantee: Álvaro Formoso Pelegrin
• Host Institution: Faculdade de Odontologia (FOAr). Universidade Estadual Paulista (UNESP). Campus de Araraquara. Araraquara , SP, Brazil
• Associated research grant: 20/00394-2 - Head and neck squamous cell carcinoma (HNSCC) invasion and distant metastasis: relevance and crosstalk between GALR2 and efferocytosis in the tumor microenvironment and hematogenic dissemination, AP.TEM

Abstract

NLRP3 inflammasome has a relevant role on innate immunity in both infectious and non-infectious conditions and the activation status of NLRP3 also influences adaptive immunity. Genetic variations and gain of function of this inflammasome are linked to multiple conditions and diseases presenting an important inflammatory component, indicating both the relevance and influence of NLRP3 on the immune response. Reports indicate ambiguous roles for NLRP3 in cancer, ranging from a pro-tumoral/oncogenic to as anti-tumoral/tumor suppressor influence. These controversies may stem from the type of cancer considered, the stage of development of the tumor, and from specificities in experimental approaches and models used. Nevertheless, most studies indicate the presence of genetic variations and gain of function/increased expression of NLRP3 in various types of cancer, including oral squamous cell carcinoma (OSCC). Activation mechanisms of NLRP3 and include both exogenous (PAMPs) and endogenous (DAMPs) stimuli, triggering activation via a canonical pathway (which includes a NF-kB-dependent first/priming phase followed by a secondary/activation phase) or via a non-canonical pathway (through direct activation of Caspase-4/-5 in humans). Most importantly for this proposal, it was recently shown that the small GTPase Rac1 (a downstream mediator of GALR2 signaling and the master switch in efferocytosis - two biological processes of interest investigated in the process 2020/00394-2) physically interacts with NLRP3 and influences the activation of the inflammasome. Other recent evidence supports a direct role of NLRP3 in the transcription/expression of IL-4 in immune cells, suggesting that NLRP3 may directly promote a suppressive/pro-tumoral immune phenotype. The main biological outcomes subsequent to NLRP3 activation derive from the activation of Caspase-1 and the enzymatic activation (cleavage) of inflammatory cytokines IL-1b and IL-18, as well as of Gasdermin D, inducing inflammatory cell death by pyroptosis. The specific biological consequences of NLRP3 activation in neoplastic or stromal/immune cells for OSCC progression is not known, but its improved comprehension may provide insights into the ambiguous role of NLRP3 in tumor progression, as a biomarker of tumor aggressiveness/response to treatment and into its potential as a therapeutic target. The main hypothesis in this proposal is that the NLRP3 inflammasome has relevant and distinct influences on progression of OSCC, depending on the cell type considered: neoplastic cells or immune cells. To test this hypothesis, we propose in vitro experiments investigating the influence of NLRP3 activity on indirect and direct interactions between neoplastic cells and immune cells, assessing phenotype of immune cells, tumor cell invasion and efferocytosis activity as outcomes. In vivo, we will use a murine syngeneic immunocompetent orthotopic model of OSCC using both WT and NLRP3 KO mice to dissect the influences of NLRP3 in neoplastic cells (with and without shRNA-mediated silencing of NLRP3) and stromal/immune cells on tumor progression, immune infiltration and phenotype and the prevalence of apoptosis and pyroptosis in the tumor microenvironment. The association between NLRP3 expression/activation and OSCC aggressiveness, immune infiltration and efferocytosis in the tumor microenvironment will also be studied in patient-derived samples. (AU)