Relação entre a expressão de CD44-ICD e a inibição de metaloproteinases na progressão de melanomas: estudo in vitro e in vivo

• Ano de defesa: 2025
• Autor(a) principal: Bruna Mendes Lima
• Tipo de documento: Tese
• Tipo de acesso: Acesso aberto
• Idioma: por
• Instituição de defesa: Universidade Federal de Minas Gerais
• Palavras-chave em Português: Patologia; Melanoma; Metástase Neoplásica; Doxiciclina; Progressão neoplásica; Metástase pulmonar
• Link de acesso: https://hdl.handle.net/1843/80476

Melanoma is a highly metastatic neoplasm, with aggressive, invasive behavior and high resistance to treatment. During its progression, the importance of extracellular matrix remodeling via metalloproteinases has been demonstrated, facilitating processes such as migration, invasion and metastasis formation. Matrix metalloproteinases MMPs interact with adhesion molecules, such as CD44, promoting their cleavage, generating the intracytoplasmic domain (ICD) that is translocated to the nucleus, activating the transcription of genes related to migration, invasion and tumor implantation. Hence, the CD44 cleavage inhibition through metalloproteinase blockage, could function as a possible target for inhibition of tumor progression and metastasis. Thus, this work aimed to investigate the effects of metalloproteinase inhibition from the use of doxycycline on melanoma progression in the murine model. B16-F10 cells grown under normal conditions and treated with 1, 5 and 10 μM of doxycycline for 24 hours were used. Viability was evaluated by the MTT assay, followed by migration, invasion, cell cycle and zymography assays. For the in vivo assay, cells were treated and inoculated into 10-week-old female C57BL/6 mice via the tail vein. After 21 days, the animals were anesthetized and lung function was evaluated by spirometry, later they were euthanized and the lung collected for histopathological and immunohistochemical evaluation of metastases. The results to date showed that doxycycline induced an increase in the viability of B16-F10 cells, with significant results at 24 hours and 48 hours in the 10μM treated group, compared to the control and 1μM treated groups (p<0.05 and p<0.01 respectively). The analysis of gelatinolytic capacity indicated a reduction in MMP-9 activity at a concentration of 1 μM in 24 hours and inhibition at a concentration of 10 μM in 24 and 48 hours (p<0.01; p<0.001 respectively).The analysis of lung mechanics during the formation of metastases revealed a reduction in the levels of FVC, Cdyn and FEV100 in the groups treated with 10 μM when compared to the control (p<0.05). The lung areas affected by metastases in the animals treated with 1 μM and 10 μM of doxycycline were larger than those of the control animals (p<0,05). Immunohistochemistry revealed a lower expression of MMP-14 and CD44 in cases treated with doxycycline when compared to the control (p<0.0001), with no relevant changes in the expression of CD44-ICD (p<0.0001) and a decrease in the expression of MMP-9. Furthermore, a relationship was observed between the CDC47 proliferative index and the area affected by metastases (r=0.9505; p<0,05) and expression of MMP-9 and 14 (r=0.7959; r=0.814 respectively). Our data suggest that in vitro treatment of B16-F10 cells with doxycycline, considered a selective inhibitor of metalloproteinases, increases cell viability, inhibits the gelatinolytic activity of MMP-9, and promotes, in the in vivo model, a reduction in MMP-14 and CD44, without considerable changes in CD44-ICD and MMP-9, increases the development of lung metastases and reduces lung function in an experimental model of B16-F10 cells.