Biology subjects

Wisnovsky, S.

Publications and source records attributed to Wisnovsky, S..

5 recordsLinked to original sources

bioRxiv2026

A genome-wide CRISPR screen reveals cancer-specific regulators of hyaluronan binding and cellular invasion

Metastatic spread of cancer cells is driven by binding between the cell-surface receptor CD44 and hyaluronan (HA) in the extracellular matrix. The specific oncogenes that drive increased CD44-HA binding in cancer remain poorly defined. Using a fluorescently labeled hyaluronan probe, we performed a genome-wide CRISPR screen to identify genes whose knockdown disrupts HA binding in breast cancer cells. We subsequently developed a bioinformatic analysis pipeline that enabled stratification and prioritization of cancer-specific regulators. This work provides a first-in-class resource for the identification of druggable targets to inhibit HA binding. We further validate RAB4A, a top hit from our screen, as a novel regulator of this process. Mechanistically, RAB4A KO dramatically reduces CD44 expression and inhibits the invasion of breast cancer cells through HA-rich matrices. This study validates a novel strategy for identifying regulators of cancer cell invasion and identifies immediate actionable targets for anti-metastatic therapy.

cancer biology↗

bioRxiv2025

A lectin-based magnetic CRISPR screening protocol permits rapid discovery of genes regulating cell-surface glycosylation

FACS-based CRISPR screening has emerged as a potent tool for dissecting the genetic networks that regulate cell-surface glycosylation. However, existing protocols are tedious and poorly suited to many cell models. We developed a lectin-based magnetic-activated cell sorting platform (Lec-MACS) that enables rapid identification of genes controlling expression of specific cell-surface glycans. Lec-MACS is faster and easier to perform than FACS-based screening while producing data of similar quality. We subsequently applied Lec-MACS to produce a genomic atlas of genes regulating breast cancer hypersialylation. This method will dramatically expand the scope and throughput of genetic screens targeted at cell-surface glycans.

genomics↗

bioRxiv2025

CRISPR activation screens map the genomic landscape of cancer glycome remodeling

Many cancer types upregulate expression of sialic acid-containing glycans. These oligosaccharides subsequently engage inhibitory Siglec receptors on immune cells, allowing cancer cells to evade immune surveillance. The genetic mechanisms by which this glycome remodeling occurs remain poorly defined. Understanding the ways that cancer cells change their cell surface glycosylation is critical for identification of biomarkers and targets for glycan-directed immunotherapy. In this study, we performed multiple gain-of-function CRISPR activation (CRISPRa) screens to broadly define genetic pathways that regulate expression of Siglec-binding glycans. We show that Siglec ligand expression is largely controlled through genetic competition between genes that catalyze 2-3 sialylation and GlcNAcylation of galactose residues. Perturbation of enzyme expression at this key biosynthetic node provides multiple "paths" by which cancers can acquire elevated expression of Siglec ligands. We further show that cancer glycome remodeling is aided by overexpression of novel "professional ligands" that facilitate Siglec-glycan binding. Notably, we also find that expression of the CD24 gene is genetically dispensable for cell-surface binding of the inhibitory receptor Siglec-10. Finally, by integrating our functional genetic model with clinical tumor genomic data, we identify the sulfotransferase enzyme GAL3ST4 as a potential novel driver of immune evasion in glioma cells. Taken together, this study provides a first-in-class genomic atlas to aid understanding of cancer-associated glycosylation and identifies immediately actionable targets for cancer immunotherapy.

genomics↗

bioRxiv2024

A Genome-Wide CRISPR Screen Identifies Sortilin as the Receptor Responsible for Galectin-1 Lysosomal Trafficking

Galectins are a family of mammalian glycan-binding proteins that have been implicated as regulators of myriad cellular processes including cell migration, apoptosis, and immune modulation. Several members of this family, such as galectin-1, exhibit both cell-surface and intracellular functions. Interestingly, galectin-1 can be found in the endomembrane system, nucleus, or cytosol, as well as on the cell surface. The mechanisms by which galectin-1 traffics between cellular compartments, including its unconventional secretion and internalization processes, are poorly understood. Here, we determined the pathways by which exogenous galectin-1 enters cells and explored its capacity as a delivery vehicle for protein and siRNA therapeutics. We used a galectin-1-toxin conjugate, modelled on antibody-drug conjugates, as a selection tool in a genome-wide CRISPR screen. We discovered that galectin-1 interacts with the endosome-lysosome trafficking receptor sortilin in a glycan-dependent manner, which regulates galectin-1 trafficking to the lysosome. Further, we show that this pathway can be exploited for delivery of a functional siRNA. This study sheds light on the mechanisms by which galectin-1 is internalized by cells and suggests a new strategy for intracellular drug delivery via galectin-1 conjugation.

molecular biology↗

bioRxiv2022

Antibody-lectin chimeras for glyco-immune checkpoint blockade

Despite the curative potential of checkpoint blockade immunotherapy, most patients remain unresponsive to existing treatments. Glyco-immune checkpoints - interactions of cell-surface glycans with lectin, or glycan-binding, immunoreceptors - have emerged as prominent mechanisms of immune evasion and therapeutic resistance in cancer. Here, we describe antibody-lectin chimeras (AbLecs), a modular platform for glyco-immune checkpoint blockade. AbLecs are bispecific antibody-like molecules comprising a cell-targeting antibody domain and a lectin "decoy receptor" domain that directly binds glycans and blocks their ability to engage inhibitory lectin receptors. AbLecs potentiate anticancer immune responses including phagocytosis and cytotoxicity, outperforming most existing therapies and combinations tested. By targeting a distinct axis of immunological regulation, AbLecs synergize with blockade of established immune checkpoints. AbLecs can be readily designed to target numerous tumor and immune cell subsets as well as glyco-immune checkpoints, and therefore represent a new modality for cancer immunotherapy.

biochemistry↗