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| Author(s): |
Amanda Pires Bonfanti
Total Authors: 1
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| Document type: | Doctoral Thesis |
| Press: | Campinas, SP. |
| Institution: | Universidade Estadual de Campinas (UNICAMP). Instituto de Biologia |
| Defense date: | 2024-05-15 |
| Examining board members: |
Catarina Raposo Dias Carneiro;
Vinícius de Oliveira Boldrini;
Isabel Kinney Ferreira de Miranda Santos;
Priscila Pini Zenatti;
Ana Lucia Tasca Gois Ruiz
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| Advisor: | Fernando Guimarães; Catarina Raposo Dias Carneiro |
| Abstract | |
Gliomas are the most common primary malignant tumors of the central nervous system in adults. They are highly invasive and heterogeneous, making treatment difficult. Glioblastomas (GBMs) are the most common and aggressive type of glioma. Despite currently available treatments, GBMs continue to be associated with high morbidity and mortality. Cancer immunotherapy involves interfering with the immune system to enhance or modify anti-tumor immune responses. Studies conducted by our group have shown that the venom of the spider Phoneutria nigriventer has cytotoxic effects on the human GBM cell line NG97 and activates dendritic cells, both in vitro. However, we had not yet investigated the effects of the venom on innate immune cells. This led us to the objectives of the present study: to clarify the effects of the venom and its purified components on macrophages and NK cells, including in vitro analyzes (to evaluate the effects on the phenotype and function of these cells), as well as in vivo, in a xenogenic GBM model (to evaluate the clinical efficacy of the treatments). Initially, a proof of concept was performed, through the treatment of animals with GBM induced on the back, by inoculation of cells from the NG97 lineage. The results showed that the administration of the venom (100 ?g/Kg) every 48 hours, for 14 days, reduced the tumor size and promoted large necrotic areas (histopathology), which justifies the low uptake of 18F-FDG observed in the PET-CT images. It increased the infiltration of macrophages in the peri and intratumoral regions (immunohistochemistry) and increased the number of circulating monocytes and NK cells (blood count and flow cytometry). Analysis of the effects of the venom on the phenotype and function of isolated macrophages and NK cells showed that PnV (14 ?g/mL, for 24 h) and its isolated component (called LW-9; 1 ?g/mL) modulated macrophages with LPS-induced M1 profile, increasing their phagocytic, cytotoxic and migratory capacity. In the M1 profile polarized with LPS and IFN-?, LW-9 increased IL-6 (1 ?g/mL, but there was no change with 0.1 ?g/mL) and reduced IL-10 (in both concentrations), indicating both phenotypic and functional activation of this profile. In NK cells, LW-9 (1 ?g/mL, for 5 h) increased the release of IFN-?, suggesting activation of these cells. It was possible to confirm that GBM NG97 cells are targeted by NK, even if not stimulated and activation with IL-2 (control) and with LW-9 did not increase the effector action of NK. However, LW-9 increased NK cytotoxicity against their control target cells (YAC-1). Finally, adoptive cell therapy was performed (previously activated NK cells and M1 macrophages, with LW-9 at 1 ?g/mL for 5 hours for NK cells and LW-9 at 0.1 ?g/mL for 2 hours for M1 macrophages) in animals with GBM (intracerebral inoculation of NG97 cells), with the following experimental groups: 1) NK, 2) NK + M1, 3) M1, 4) Systemic LW-9; three control groups were conducted: Tumor-free control, Sham control, and Tumor-bearing control without treatment. All three experimental treatments (NK, NK+M1, LW-9) reduced tumor size compared to the untreated group (significant only for the NK-treated group). All treatments reduced the quantity of infiltrated macrophages in the TME, except the group that received M1. Systemic LW-9 treatment induced an increase in IFN-? and a reduction in TNF-?. Histopathological analysis showed tumors with lower cellularity and vascularization in animals treated with NK cells and systemic LW-9. Therefore, this study contributed to clarifying the effects of LW-9 on macrophages and NK cells and to initiating the establishment of an adoptive cell therapy protocol. The study also provided insights into the crosstalk between macrophages and NK cells in the context of the TME. Other protocols will be tested by increasing the treatment duration and reducing the administration interval, as well as combining therapy with temozolomide (the gold standard chemotherapy for GBM). This study presents an innovative and promising approach, aiming to contribute to GBM treatment and with the potential to be tested in other solid tumors (AU) | |
| FAPESP's process: | 18/23559-7 - Cell therapy with macrophages and NK cells modulated ex vivo by peptide isolated from animal venom: a new approach in immunotherapy for cancer |
| Grantee: | Amanda Pires Bonfanti |
| Support Opportunities: | Scholarships in Brazil - Doctorate |