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6 results on '"Blomberg, Anne Louise"'

1. Cell-based glycoengineering of extracellular vesicles through precise genome editing

Author

Tian, Weihua, Zagami, Chiara, Chen, Jiasi, Blomberg, Anne Louise, Guiu, Laura Salse, Skovbakke, Sarah Line, Goletz, Steffen, Tian, Weihua, Zagami, Chiara, Chen, Jiasi, Blomberg, Anne Louise, Guiu, Laura Salse, Skovbakke, Sarah Line, and Goletz, Steffen

Abstract

Engineering of extracellular vesicles (EVs) towards more efficient targeting and uptake to specific cells has large potentials for their application as therapeutics. Carbohydrates play key roles in various biological interactions and are essential for EV biology. The extent to which glycan modification of EVs can be achieved through genetic glycoengineering of their parental cells has not been explored yet. Here we introduce targeted glycan modification of EVs through cell-based glycoengineering via modification of various enzymes in the glycosylation machinery. In a “simple cell” strategy, we modified major glycosylation pathways by knocking-out (KO) essential genes for N-glycosylation (MGAT1), O-GalNAc glycosylation (C1GALT1C1), glycosphingolipids (B4GALT5/6), glycosaminoglycans (B4GALT7) and sialylation (GNE) involved in the elongation or biosynthesis of the glycans in HEK293F cells. The gene editing led to corresponding glycan changes on the cells as demonstrated by differential lectin staining. Small EVs (sEVs) isolated from the cells showed overall corresponding glycan changes, but also some unexpected differences to their parental cell including enrichment preference for certain glycan structures and absence of other glycan types. The genetic glycoengineering did not significantly impact sEVs production, size distribution, or syntenin-1 biomarker expression, while a clonal influence on sEVs production yields was observed. Our findings demonstrate the successful implementation of sEVs glycoengineering via genetic modification of the parental cell and a stable source for generation of glycoengineered sEVs. The utilization of glycoengineered sEVs offers a promising opportunity to study the role of glycosylation in EV biology, as well as to facilitate the optimization of sEVs for therapeutic purposes.

Published
2024

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Author

Romanò, Cecilia, Jiang, Hao, Tahvili, Sahar, Wei, Peng, Keiding, Ulrik B., Clergeaud, Gael, Skovbakke, Sarah Line, Blomberg, Anne Louise, Hafkenscheid, Lise, Henriksen, Jonas R., Andresen, Thomas L., Goletz, Steffen, Hansen, Anders E., Christensen, Dennis, Clausen, Mads H., Romanò, Cecilia, Jiang, Hao, Tahvili, Sahar, Wei, Peng, Keiding, Ulrik B., Clergeaud, Gael, Skovbakke, Sarah Line, Blomberg, Anne Louise, Hafkenscheid, Lise, Henriksen, Jonas R., Andresen, Thomas L., Goletz, Steffen, Hansen, Anders E., Christensen, Dennis, and Clausen, Mads H.

Published
2024

4. Novel genetically glycoengineered human dendritic cell model reveals regulatory roles of α2,6-linked sialic acids in DC activation of CD4+ T cells and response to TNFα.

Author

Tian, Weihua, Blomberg, Anne Louise, Steinberg, Kaylin Elisabeth, Henriksen, Betina Lyngfeldt, Jørgensen, Josefine Søborg, Skovgaard, Kerstin, Skovbakke, Sarah Line, and Goletz, Steffen

Subjects
*T cells, *DENDRITIC cells, *SIALIC acids, *HUMAN biology, *GLYCOCALYX, *IMMUNOREGULATION, *T cell receptors, *GLYCAN structure
Abstract

Dendritic cells (DCs) are central for the initiation and regulation of appropriate immune responses. While several studies suggest important regulatory roles of sialoglycans in DC biology, our understanding is still inadequate primarily due to a lack of appropriate models. Previous approaches based on enzymatic- or metabolic-glycoengineering and primary cell isolation from genetically modified mice have limitations related to specificity, stability, and species differences. This study addresses these challenges by introducing a workflow to genetically glycoengineer the human DC precursor cell line MUTZ-3, described to differentiate and maturate into fully functional dendritic cells, using CRISPR-Cas9, thereby providing and validating the first isogenic cell model for investigating glycan alteration on human DC differentiation, maturation, and activity. By knocking out (KO) the ST6GAL1 gene, we generated isogenic cells devoid of ST6GAL1-mediated α(2,6)-linked sialylation, allowing for a comprehensive investigation into its impact on DC function. Glycan profiling using lectin binding assay and functional studies revealed that ST6GAL1 KO increased the expression of important antigen presenting and co-stimulatory surface receptors and a specifically increased activation of allogenic human CD4 + T cells. Additionally, ST6GAL1 KO induces significant changes in surface marker expression and cytokine response to TNFα-induced maturation, and it affects migration and the endocytic capacity. These results indicate that genetic glycoengineering of the isogenic MUTZ-3 cellular model offers a valuable tool to study how specific glycan structures influence human DC biology, contributing to our understanding of glycoimmunology. [ABSTRACT FROM AUTHOR]

Published
2024
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5. Chemical synthesis and immunological evaluation of cancer vaccines based on ganglioside antigens and α-galactosylceramideElectronic supplementary information (ESI) available. See DOI: https://doi.org/10.1039/d4md00387j

Author

Romanò, Cecilia, Jiang, Hao, Tahvili, Sahar, Wei, Peng, Keiding, Ulrik B., Clergeaud, Gael, Skovbakke, Sarah Line, Blomberg, Anne Louise, Hafkenscheid, Lise, Henriksen, Jonas R., Andresen, Thomas L., Goletz, Steffen, Hansen, Anders E., Christensen, Dennis, and Clausen, Mads H.

Abstract

iNKT cells – often referred as the “Swiss Army knife” of the immune system – have emerged as central players in cancer vaccine therapies. Glycolipids activating iNKT cells, such as α-galactosylceramide (αGalCer), can enhance the immune response against co-delivered cancer antigens and have been applied in the design of self-adjuvanting anti-tumor vaccines. In this context, this work focuses on the chemical synthesis of ganglioside tumor-associated carbohydrate antigens (TACAs), namely GM3 and (Neu5Gc)GM3 antigens, their conjugation to αGalCer, and their formulation into liposomes as an efficient platform for their in vivodelivery. Liposomes containing GM3–αGalCer, (Neu5Gc)GM3–αGalCer, and equimolar amounts of the two conjugates have been fully characterized and their ability to activate iNKT cell has been confirmed ex vivoin mouse and human cell assays. The candidates were tested in in vivoimmunization studies, demonstrating an ability to induce both TH1 and TH2 cytokines leading to the production of all subclasses of IgG antibodies. Notably, the study also demonstrated that serum antibodies raised against the two TACAs, alone and in combination, were cross-reactive. This finding has consequences for future vaccine designs – even if a highly tumor-selective antigen is chosen, the resulting antibody response may be broader than anticipated.

Published
2024
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