Biology subjects

Paulo, P.

Publications and source records attributed to Paulo, P..

4 recordsLinked to original sources

bioRxiv2025

CD276 Immature Glycosylation Drives Colorectal Cancer Aggressiveness and T-cell Mediated Immune Escape

Colorectal cancer (CRC) progression is fueled by immune evasion, yet the underlying molecular mechanisms remain to be fully characterized. CD276 (B7-H3), an immune checkpoint glycoprotein frequently overexpressed in aggressive tumours, is extensively modified by glycosylation, a process known to regulate protein stability, localization, and immune interactions. However, its glycosylation-dependent functions in CRC remain unclear. This study shows that poor-prognosis CRC tumours exhibit dysregulated glycogene expression, leading to an immature O-glycosylation phenotype that enhances malignancy. Using C1GALT1 knockout CRC cells, which recapitulate the cancer glycocalyx, it was demonstrated that this glycosylation shift promotes proliferation and invasion. We further identify CD276 as a metastasis-associated glycoprotein whose function is shaped by O-glycan modifications. Mechanistically, immature O-glycosylation of CD276 enhances invasion, suppresses T-cell activation, and induces an immunosuppressive cytokine milieu, reinforcing its role in tumour immune escape. These findings establish CD276 as a glycosylation-dependent immune checkpoint and a promising therapeutic target to overcome immune evasion in CRC. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=184 SRC="FIGDIR/small/668091v1_ufig1.gif" ALT="Figure 1"> View larger version (101K): org.highwire.dtl.DTLVardef@2af030org.highwire.dtl.DTLVardef@fee9a8org.highwire.dtl.DTLVardef@19e1c49org.highwire.dtl.DTLVardef@152fd73_HPS_FORMAT_FIGEXP M_FIG C_FIG Table of ContentsColorectal cancer (CRC) progression is closely linked to immune evasion, yet the molecular mechanisms underlying this process remain poorly understood. This study identifies CD276 (B7-H3) as a glycosylation-driven regulator of CRC aggressiveness and demonstrates how O-glycosylation remodeling promotes tumour immune escape. These findings establish CD276 as a potential therapeutic target and highlight the role of glycoproteoform-specific immune modulation in cancer progression.

cancer biology↗

bioRxiv2023

Multilevel Plasticity and Altered Glycosylation Drive Aggressiveness in Hypoxic and Glucose-Deprived Bladder Cancer Cells

Bladder tumours with aggressive characteristics often present with microenvironmental niches marked by low oxygen levels (hypoxia) and limited glucose supply due to inadequate vascularization. The molecular mechanisms facilitating cellular adaptation to these stimuli remain largely elusive. Employing a multi-omics approach, we discovered that hypoxic and glucose- deprived cancer cells enter a quiescent state supported by mitophagy, fatty acid {beta}-oxidation, and amino acid catabolism, concurrently enhancing their invasive capabilities. Reoxygenation and glucose restoration efficiently reversed cell quiescence without affecting cellular viability, highlighting significant molecular plasticity in adapting to microenvironmental challenges. Furthermore, cancer cells exhibited substantial perturbation of protein O-glycosylation, leading to simplified glycophenotypes with shorter glycosidic chains. Exploiting glycoengineered cell models, we established that immature glycosylation contributes to reduced cell proliferation and increased invasion. Our findings collectively indicate that hypoxia and glucose deprivation trigger cancer aggressiveness, reflecting an adaptive escape mechanism underpinned by altered metabolism and protein glycosylation, providing grounds for clinical intervention.

biochemistry↗

bioRxiv2021

Glycoproteogenomics characterizes the CD44 splicing code driving bladder cancer invasion

Bladder cancer (BC) management demands the introduction of novel molecular targets for precision medicine. Cell surface glycoprotein CD44 has been widely studied as a potential biomarker of BC aggressiveness and cancer stem cells. However, significant alternative splicing and multiple glycosylation generate a myriad of glycoproteoforms with potentially distinct functional roles. The lack of tools for precise molecular characterization has led to conflicting results, delaying clinical applications. Addressing these limitations, we have interrogated the transcriptome of a large BC patient cohort for splicing signatures. Remarkable CD44 heterogeneity was observed, as well as associations between short CD44 standard splicing isoform (CD44s), invasion and poor prognosis. In parallel, immunoassays showed that targeting short O-glycoforms could hold the key to improve CD44 cancer specificity. This prompted the development of a glycoproteogenomics approach, building on the integration of transcriptomics-customized datasets and glycomics for protein annotation from nanoLC-ESI-MS/MS experiments. The concept was applied to invasive human BC cell lines, glycoengineered cells, and tumor tissues, enabling unequivocal CD44s identification. Finally, we confirmed the link between CD44s and invasion in vitro by siRNA knockdown, supporting findings from BC tissues. The key role played by short-chain O-glycans in CD44-mediated invasion was also demonstrated through glycoengineered cell models. Overall, CD44s emerged as biomarker of poor prognosis and CD44-Tn/STn as promising molecular signatures for targeted interventions. This study materializes the concept of glycoproteogenomics and provides a key vision to address the cancer splicing code at the protein level, which may now be expanded to better understand CD44 functional role in health and disease. Significance StatementThe biological role of CD44, a cell membrane glycoprotein involved in most cancer hallmarks and widely explored in BC, is intimately linked to its protein isoforms. mRNA alternative splicing generates several closely related polypeptide sequences, which have so far been inferred from transcripts analysis, in the absence of workflows for unequivocal protein annotation. Dense O-glycosylation is also key for protein function and may exponentiate the number of proteoforms, rendering CD44 molecular characterization a daunting enterprise. Here, we integrated multiple molecular information (RNA, proteins, glycans) for definitive CD44 characterization by mass spectrometry, materializing the concept of glycoproteogenomics. BC specific glycoproteoforms linked to invasion have been identified, holding potential for precise cancer targeting. The approach may be transferable to other tumors, paving the way for precision oncology.

cancer biology↗

bioRxiv2021

Metabolomics, Transcriptomics and Functional Glycomics Reveals Bladder Cancer Cells Plasticity and Enhanced Aggressiveness Facing Hypoxia and Glucose Deprivation

Bladder cancer constitutes one of the deadliest genitourinary diseases, especially when diagnosed at late stages. These tumours harbour microenvironmental niches characterized by low levels of oxygen (hypoxia) and limited glucose supply due to poor vascularization. However, the synergic contribution of these features to disease development is poorly understood. Here, we demonstrated that cells with distinct histopathological and molecular backgrounds responded similarly to such stimuli. Cancer cells arrested proliferation, significantly increased invasive capacity in vitro and enhanced tolerance to cisplatin-based chemotherapy. Reoxygenation and access to glucose restored basal proliferation and invasion levels without triggering stress-induced apoptosis, denoting significant cellular plasticity in adapting to microenvironmental cues. Whole transcriptomics showed major molecular reprogramming, supporting main functional alterations. Metabolomics evidenced fatty acids {beta}-oxidation as main bioenergetic pathway rather than anaerobic glycolysis generally adopted by hypoxic cells. Joint pathway analysis also suggested relevant alterations in mucin-type O-glycan biosynthesis. Glycomics confirmed a major antagonization of O-glycosylation pathways, leading to simple cell glycophenotypes characterized by the accumulation of immature short-chain O-glycans such as Tn and STn antigens at the cell surface. Glycoengineered models reflecting simple cell glycophenotypes were developed and functional studies in vitro and in vivo showed that Tn and STn overexpression decreased proliferation and promoted chemoresistance, reinforcing their close link with tumour aggressiveness. Collectively, we have demonstrated that hypoxia and glucose deprivation trigger more aggressive cell behaviours, in what appears to be an escape mechanism from microenvironmental stress. We propose that, altered glycosylation may be used to target these subpopulations, paving the way for precision oncology.

cancer biology↗