Biology subjects

Zingg, A.

Publications and source records attributed to Zingg, A..

10 recordsLinked to original sources

bioRxiv2026

alpha2,3-sialylation on human naive T cells restrains bispecific engager-mediated anti-tumor immunity

Aberrantly elevated cell-surface sialylation, or hypersialylation, is a common feature of human cancers and contributes to immune evasion. Sialidase-based therapies have therefore emerged as a strategy to disrupt this glyco-checkpoint. Although the immunosuppressive role of tumor-associated sialylation is well established, how sialylation on human T cells shapes anti-tumor responses remains poorly defined. Here, we identify surface sialoglycans on T cells, particularly 2,3-linked structures, as a cell-intrinsic restraint on human T cell activation, proliferation, and effector function. In vitro, enzymatic desialylation enhanced T cell activation, proliferation, cytokine production, and bispecific T cell engager (TCE)-mediated tumor-cell killing in healthy donor PBMC co-cultures. In ex vivo cultures of primary chronic lymphocytic leukemia (CLL) PBMCs, sialidase treatment combined with the CD20-directed TCE glofitamab enhanced cytotoxic effector transcriptional programming in autologous T cells. Single-cell RNA sequencing combined with lectin-based CITE-seq linked treatment-induced transcriptional states to lectin-defined cell-surface glycan signatures within the same single-cell dataset. This integrated analysis revealed that naive and, to a lesser extent, central memory T cells combined elevated baseline 2,3-sialylation signatures with the clearest transcriptional responses to glofitamab plus sialidase treatment. CD43 emerged as a major carrier of 2,3-linked sialoglycans, and its deletion attenuated sialidase-enhanced T cell activation. Together, these findings identify sialylation of the T cell surface as a subset-specific restraint on human TCE responses and provide a rationale for testing sialidase-TCE combinations designed to engage less-differentiated T cell populations. One-sentence summaryDesialylation enhances bispecific T cell engager responses by relieving a sialoglycan-dependent restraint in human T cells.

immunology↗

bioRxiv2026

Local glycan engineering induces systemic antitumor immune reactions via antigen cross-presentation

Immune checkpoint inhibitors (ICI) have revolutionized cancer therapy, yet response rates remain suboptimal across many solid tumors, and resistance mechanisms, particularly those involving glycans, are not fully understood. Recent studies have identified sialic acid-containing glycans and their interactions with Siglec receptors on tumor-associated macrophages as an important contributor to immune suppression within the tumor microenvironment (TME). Targeting this sialic acid-Siglec axis by glycan engineering with sialidases and other glycosidases has shown therapeutic potential in preclinical models. However, safe and effective delivery of sialidases to tumors remains a challenge. Here, we present a novel approach using adeno-associated virus (AAV)-mediated therapy to deliver sialidases (AAVSia) and other glycosidases, including fucosidase, directly to the TME. Intratumoral administration of AAVSia in mouse models resulted in significant tumor growth reduction, enhanced survival, and robust systemic antitumor immunity through improved cross-presentation and dendritic cell activation. Furthermore, combining local sialidase expression with fucosidase treatment and classical PD-1 blockade allowed a synergistic effect, amplifying antitumor response. Our findings highlight the therapeutic promise of glycoengineering the TME using local delivery systems and support the development of combination strategies to overcome glycan-mediated resistance in cancer immunotherapy. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=129 SRC="FIGDIR/small/720097v1_ufig1.gif" ALT="Figure 1"> View larger version (34K): org.highwire.dtl.DTLVardef@dc9d72org.highwire.dtl.DTLVardef@1e4e455org.highwire.dtl.DTLVardef@4a8f93org.highwire.dtl.DTLVardef@11813a3_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2025

Longitudinal profiling of tumor-reactive T cells during TIL therapy in metastatic melanoma

Adoptive cell therapy (ACT) with expanded autologous tumor-infiltrating lymphocytes (TILs) can induce durable responses in metastatic melanoma, yet many patients relapse. We profiled tumor-reactive T cell dynamics during TIL therapy using single-cell RNA and TCR sequencing from seven patients to elucidate underlying reasons. We found that tumor-reactive T cells preferentially expanded early during ex vivo TIL culture, transitioning from exhausted to reinvigorated effector states. Particularly, CD8+ exhausted T cells (Tex) and CD4+ follicular helper T cell (Tfh), but not CD4+ Tex, were efficiently reinvigorated. Further, we resolved the heterogeneity of tumor-reactive CD8+ and CD4+ subsets, defining unique signatures for their identification during TIL expansion. In addition, non-responders (NRs) exhibit increased levels of Type 17 T cells in TIL products, suggesting a potential association with resistance to therapy. After transfer, tumor-reactive clones rapidly extravasated and established a stem-like reservoir. However, in NRs, CD4+ regulatory T cells (Tregs) expanded de novo and tumor-reactive CD8+ T cells reacquired exhaustion markers, limiting their functionality. By contrast, responders (Rs) retained a pool of less differentiated, stem-like cells. Collectively, these data provide a comprehensive analysis of T cell fates during TIL-ACT providing the basis for new approaches to enhance therapeutic strategies.

immunology↗

bioRxiv2025

In vivo screening of TCR-based chimeric antigen receptors for improved anti-tumor functionality

Chimeric antigen receptor (CAR) T-cell therapy has demonstrated remarkable efficacy in hematological malignancies, but its success in solid tumors remains limited. Conventional CAR-T designs do not integrate into the T-cell receptor (TCR) and rely on non-natural CD3{zeta} signaling. They thus often suffer from tonic signaling, rapid T-cell exhaustion, and antigen escape due to reduced sensitivity. To address these limitations, we explored alternative CAR architectures that take into account the evolutionary optimized TCR signaling machinery. Specifically, we employed T-cell receptor fusion constructs (TRuCs), where a single-chain variable fragment (scFv) targeting the Sialyl-Thomson-Nouveau antigen (STn) is attached to the CD3{varepsilon} subunit. We then systematically screened a library of costimulatory domains and their combinations, fused to the TRuC, using a novel in vitro and in vivo screening approach. This screen identified a potent TRuC variant incorporating both CD28 and CD27 costimulatory domains. This optimized construct exhibited increased in vitro and in vivo proliferation and enhanced 4-1BB and IFN{gamma} expression upon repeated antigen stimulation. It further showed improved antitumor efficacy compared to conventional second-generation CAR-T cells in a mouse melanoma model. Moreover, we validated the adaptability of this approach by targeting B7H3 in a sarcoma model. The TRuC with costimulatory domains again outperformed other CAR-variants. Our findings highlight the potential of incorporating combined CD28 and CD27 costimulatory domains into TCR-based CAR architectures to overcome limitations associated with conventional CAR-T therapy and improve efficacy against solid tumors.

bioengineering↗

bioRxiv2025

CAR T cell engineering impacts antigen-independent activation and co-inhibition

Viral vectors have successfully modified T cells to express chimeric antigen receptors (CAR), leading to clinical approvals. However, their high cost and regulatory challenges hinder rapid and broad clinical translation. Here, we demonstrate that our lentivirally (LV) manufactured R110-CAR T cells, targeting a leukemia neoepitope, can also be engineered using non-viral Sleeping Beauty (SB) transposition with minimal-sized DNA vectors. Flow cytometry and single-cell sequencing was used to compare the two production modes using healthy donor and CLL patient-derived T cells and a CD19-CAR T cell control. SB products were shifted towards CD8+ subsets with expression of activation/co-inhibition markers (CD69, LAG-3, TIM-3) despite their naive-like phenotype and lack of antigenic challenge. The CAR binding moiety modulated these patterns with R110-CAR T cells showing more aberrant phenotypes. Moreover, SB engineering resulted in inflammatory signatures along with RIG-I-like and TOLL-like nucleotide sensing potentially resulting from the transfection procedure. Patient-derived products showed significantly fewer CAR-expressing cells, reduced proliferation clusters, and lower T cell diversity, particularly with SB manufacturing, pointing at potential challenges with this method when engineering CLL T cells. Together, our data suggest that the engineering mode may substantially influence T cell properties and that these are further modulated by the CAR binding moiety and the type of T cell donor.

immunology↗

bioRxiv2024

Targeting cancer-associated glycosylation for adoptive T cell therapy of gastro-intestinal and gynecological cancers

CAR-T cell therapy has provided a significant improvement for patients with chemotherapy-resistant B cell malignancies. However, CAR-T cell treatment of patients with solid cancers has been more difficult, in part because of the heterogeneous expression of tumor-specific cell surface antigens. Here, we describe the generation of a fully human CAR targeting altered glycosylation in secretory epithelial cancers. The expression of the target antigen - the truncated, sialylated O-glycan sialyl-Thomsen-Nouveau antigen (STn) - was studied with a highly STn-specific antibody across various different tumor tissues. Strong expression was found in a high proportion of gastro-intestinal cancers including pancreatic cancers and in gynecological cancers, in particular ovarian and endometrial tumors. T cells expressing anti-STn CAR were tested in vitro and in vivo. Anti-STn CAR-T cells showed activity in mouse models as well as in assays with primary ovarian cancer samples. No significant toxicity was observed in mouse models, although some intraluminal expression of STn was found in gastro-intestinal mouse tissue. Taken together, this fully human anti-STn CAR construct shows promising activity in preclinical tumor models supporting its further evaluation in early clinical trials.

cancer biology↗

bioRxiv2023

Engagement of sialylated glycans with Siglec receptors on myeloid suppressor cells inhibit anti-cancer immunity via CCL2

Overexpression of sialic acids on glycans, called hypersialylation is a common alteration found in cancer. Hypersialylation can, for example, enhance immune evasion via interaction with sialic acid-binding immunoglobulin-like lectin (Siglec) receptors on tumor-infiltrating immune cells. Here, we tested the role of sialic acid on myeloid-derived suppressor cells (MDSCs) and their interaction with Siglec receptors. We found that MDSCs derived from the blood of lung cancer patients and tumor-bearing mice strongly express inhibitory Siglec receptors. In murine cancer models of emergency myelopoiesis, Siglec-E knockout on myeloid cells resulted in prolonged survival and increased infiltration of activated T cells. Targeting suppressive myeloid cells by blocking Siglec receptors or desialylation led to strong reduction of their suppressive potential. We further identified CCL2 as mediator involved in T cell suppression upon interaction of sialoglycans and Siglec receptors on MDSCs. Our results provide mechanistic insights how sialylated glycans inhibit anti-cancer immunity by facilitating CCL2 expression. O_FIG O_LINKSMALLFIG WIDTH=142 HEIGHT=200 SRC="FIGDIR/small/547025v1_ufig1.gif" ALT="Figure 1"> View larger version (42K): org.highwire.dtl.DTLVardef@e7664forg.highwire.dtl.DTLVardef@1edd220org.highwire.dtl.DTLVardef@7e630dorg.highwire.dtl.DTLVardef@198ff40_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2020

MTX-COVAB, a human-derived antibody with potent neutralizing activity against SARS-CoV-2 infection in vitro and in a hamster model of COVID-19

Fast track microfluidic screening of the antibody repertoires of 12 convalescent COVID-19 donors comprising 2.8mio antibodies yielded MTX-COVAB, a human-derived monoclonal antibody with low picomolar neutralization IC50 of SARS-CoV-2. COVAB neutralization potency is on par with the Regeneron cocktail as demonstrated in a comparative neutralization assay. MTX-COVAB shows strong efficacy in vivo and binds to all currently identified clinically relevant variants of SARS-CoV-2. MTX-COVAB completes GMP manufacturing by the end of this year and will be tested in the clinic in March 2021.

immunology↗

bioRxiv2018

Synthetic standards combined with error and bias correction improves the accuracy and quantitative resolution of antibody repertoire sequencing in human and naive memory B cells

High-throughput sequencing of immunoglobulin repertoires (Ig-seq) is a powerful method for quantitatively interrogating B cell receptor sequence diversity. When applied to human repertoires, Ig-seq provides insight into fundamental immunological questions, and can be implemented in diagnostic and drug discovery projects. However, a major challenge in Ig-seq is ensuring accuracy, as library preparation protocols and sequencing platforms can introduce substantial errors and bias that compromise immunological interpretation. Here, we have established an approach for performing highly accurate human Ig-seq by combining synthetic standards with a comprehensive error and bias correction pipeline. First, we designed a set of 85 synthetic antibody heavy chain standards (in vitro transcribed RNA) to assess correction workflow fidelity. Next, we adapted a library preparation protocol that incorporates unique molecular identifiers (UIDs) for error and bias correction which, when applied to the synthetic standards, resulted in highly accurate data. Finally, we performed Ig-seq on purified human circulating B cell subsets (naive and memory), combined with a cellular replicate sampling strategy. This strategy enabled robust and reliable estimation of key repertoire features such as clonotype diversity, germline segment and isotype subclass usage, and somatic hypermutation (SHM). We anticipate that our standards and error and bias correction pipeline will become a valuable tool for researchers to validate and improve accuracy in human Ig-seq studies, thus leading to potentially new insights and applications in human antibody repertoire profiling.

immunology↗

bioRxiv2017

Demographic variability in spruce-fir-beech forest stands in Europe

1Many studies have predicted large changes in forest dynamics during the next century because of global warming. Although empirical approaches and studies based on species distribution models provide valuable information about future changes, they do not take into account biotic interactions and stand-level demographic variations. The objective of this study was to quantify the local and regional variability of the growth and regeneration of three important forest species growing often in mixed stands in Europe (Picea abies (L.) Karst., Abies alba Mill., Fagus sylvatica), and to assess the climatic drivers of this variability. For that purpose, we collected a large forestry data set compiling the long-term (up to 100 years) evolution of species and size distributions for 163 stands across Europe, in the mesic distribution area of these forests. We used an inverse modeling approach, Approximate Bayesian Computation, to calibrate an individual-based model of forest dynamics on these data. Our study revealed that the variability of the demographic processes was of the same order of magnitude between stands of a same forest as between different forests. Out of the three species and two demographic processes studied, only the fir growth strongly varied with temperature. Water availability did not explain any demographic variation over stands. For these forests experiencing mesic conditions, local unmeasured factors seem therefore to have an influence at least as important as macro-environmental factors on demographic variations. Efforts to include these important factors in projection scenarios should therefore be prioritized. Besides, our study demonstrates that inverse modelling methods make possible the analysis of long-term forestry data. Such data should therefore be more widely compiled and used for ecological and global change research.

ecology↗