Browsing by Author "Peixoto, Andreia"
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- Deciphering the influence of CD276 glycocode in gastric cancer immunoregulationPublication . Magalhães, Mariana; Dourado-Gonçalves, Martina; Santos, Lúcio Lara; Peixoto, Andreia; Ferreira, José AlexandreGastric cancer (GC) ranks 5th globally in incidence and cancer-related mortality, with advanced-stage 5-year survival rates below 20%1,2, unveiling the need for novel immunotherapies. CD276, an immune checkpoint molecule, is related to immune evasion and tumour progression in GC3-5. In turn, aberrant glycosylation, especially truncated O-glycans, Tn and Sialyl-Tn, correlates with GC aggressiveness and poor prognosis6. Its clinical impact relies on the modulation of protein function and immune escape5,7. This study hypothesizes that CD276 glycosylation may offer new targets for immunotherapy in GC. Methods: CD276 expression was assessed in AGS and MKN-45 cell lines using qPCR and immunoblotting. Glycoengineered AGS and MKN-45 cell lines with truncated glycosylation (C1GALT1 KO) and wild-type (WT) controls were analysed similarly. In vitro proliferation, migration and invasion assays were conducted. CD276 expression was assessed in AGS and MKN-45 cell lines using qPCR and immunoblotting. Glycoengineered AGS and MKN-45 cell lines with truncated glycosylation (C1GALT1 KO) and wild-type (WT) controls were analysed similarly. In vitro proliferation, migration and invasion assays were conducted. Both cell lines have CD276 expression in WT cells. CRISPR/Cas9-mediated knockout of C1GALT1 did not significantly alter CD276 protein or mRNA expression levels. The proliferation assay did not show any changes in either cell line. However, migratory and invasive assays promoted a cell line-dependent response. The migratory capacity of AGS decreased with truncated glycosylation while increasing for MKN-45. C1GALT1 KO AGS cells showed enhanced invasiveness. Aberrant glycosylation in AGS and MKN-45 cells did not affect CD276 expression in vitro, suggesting a CD276 expression independent of O-glycan elongation in GC. Tn/STn glycophenotypes have a cell line-dependent functional impact in GC, suggesting glycosylation-related migration and invasion capacities. Future proteomic studies will clarify the results. Moreover, further research will identify CD276 glycoproteoforms and assess CD276 abrogation effects on T-cell immunomodulation and cytokine profiles. This research may contribute to the development of novel targeted therapies and improved patient outcomes in GC.
- P53 and Cancer-Associated Sialylated Glycans Are Surrogate Markers of Cancerization of the Bladder Associated with Schistosoma haematobium InfectionPublication . Santos, Júlio; Fernandes, Elisabete; Ferreira, José Alexandre; Lima, Luís; Tavares, Ana; Peixoto, Andreia; Parreira, Beatriz; Costa, José Manuel Correira da; Brindley, Paul J; Lopes, Carlos; Santos, LúcioBACKGROUND: Bladder cancer is a significant health problem in rural areas of Africa and the Middle East where Schistosoma haematobium is prevalent, supporting an association between malignant transformation and infection by this blood fluke. Nevertheless, the molecular mechanisms linking these events are poorly understood. Bladder cancers in infected populations are generally diagnosed at a late stage since there is a lack of non-invasive diagnostic tools, hence enforcing the need for early carcinogenesis markers. METHODOLOGY/PRINCIPAL FINDINGS: Forty-three formalin-fixed paraffin-embedded bladder biopsies of S. haematobium-infected patients, consisting of bladder tumours, tumour adjacent mucosa and pre-malignant/malignant urothelial lesions, were screened for bladder cancer biomarkers. These included the oncoprotein p53, the tumour proliferation rate (Ki-67>17%), cell-surface cancer-associated glycan sialyl-Tn (sTn) and sialyl-Lewisa/x (sLea/sLex), involved in immune escape and metastasis. Bladder tumours of non-S. haematobium etiology and normal urothelium were used as controls. S. haematobium-associated benign/pre-malignant lesions present alterations in p53 and sLex that were also found in bladder tumors. Similar results were observed in non-S. haematobium associated tumours, irrespectively of their histological nature, denoting some common molecular pathways. In addition, most benign/pre-malignant lesions also expressed sLea. However, proliferative phenotypes were more prevalent in lesions adjacent to bladder tumors while sLea was characteristic of sole benign/pre-malignant lesions, suggesting it may be a biomarker of early carcionogenesis associated with the parasite. A correlation was observed between the frequency of the biomarkers in the tumor and adjacent mucosa, with the exception of Ki-67. Most S. haematobium eggs embedded in the urothelium were also positive for sLea and sLex. Reinforcing the pathologic nature of the studied biomarkers, none was observed in the healthy urothelium. CONCLUSION/SIGNIFICANCE: This preliminary study suggests that p53 and sialylated glycans are surrogate biomarkers of bladder cancerization associated with S. haematobium, highlighting a missing link between infection and cancer development. Eggs of S. haematobium express sLea and sLex antigens in mimicry of human leukocytes glycosylation, which may play a role in the colonization and disease dissemination. These observations may help the early identification of infected patients at a higher risk of developing bladder cancer and guide the future development of non-invasive diagnostic tests.