Abstract
Abstract BACKGROUND Vedolizumab (VDZ) is an anti-a4b7 integrin monoclonal antibody effective for treating ulcerative colitis (UC). The cellular and genetic factors that mediate VDZ response and non-response remain unclear. Our previous data employing different multi-omics revealed an impact of VDZ on myeloid dendritic cells (mDCs) and mononuclear phagocytes (MNPs), in addition to lymphocytes, influencing stromal and epithelial compartments. METHODS We performed a mass cytometry (CyTOF) experiment on peripheral blood and colon biopsies in healthy controls (HC, n=6), patients with UC on anti-TNF or 5-ASA (UC, n=5/6) and patients with UC on VDZ (UC-VDZ, n=5). Subsequently, we carried out a retrospective longitudinal spatial analysis using 1000-plex RNA and 62-plex protein CosMx on archived formalin-fixed paraffin-embedded (FFPE) biopsies before and after treatment in VDZ responders (VDZ-R) and non-responders (VDZ-NR) (Fig.1). Lastly, gene set enrichment analysis (GSEA) using VDZ response and non-response signatures obtained from our longitudinal spatial transcriptomic data were validated in an external publicly available dataset. RESULTS CyTOF results confirmed that VDZ is associated with a shift of a4b7+ cells from tissue to blood across multiple subsets. Spatial data showed elevated activated MNPs in UC compared to HC and UC-VDZ. Following treatment, these levels decreased in VDZ-R but increased in VDZ-NR. Neighborhood enrichment analysis revealed a trend toward increased proximity of activated fibroblasts and activated MNPs in UC, which was reduced following treatment. Furthermore, to explore differences pre-treatment, we performed pseudobulk differentially expressed (DE) gene analysis comparing VDZ-NR versus VDZ-R. Stromal and MNP-expressed genes including MMP1, MMP2, and THBS1 were among the top DE genes in VDZ-NR, while genes associated with the IEC crypt base including REG1A, OLFM4, AGR2, SPINK1, and LYZ were related to VDZ-R. GSEA confirmed significant enrichment of the identified VDZ-NR and VDZ-R gene signatures in patients pre-VDZ, aligning with their outcome. The VDZ-R signature was specific to VDZ and was not associated with anti-TNF response. The VDZ-NR signature was also associated with anti-TNF non-response. CONCLUSION Longitudinal spatial transcriptomics of archived FFPE specimens highlighted potential pathways associated with VDZ responsiveness, identifying epithelial-, MNP-, and fibroblast-enriched genes. Validation in an external dataset emphasized the specificity of the VDZ-associated response signature. Fig1. a, Schematic of retrospective, longitudinal analysis of archived FFPE specimens using 1000-plex CosMx spatial transcriptomics of 126,368 cells. b, Dot plot representation of a subset of genes from pseudobulk DE gene analysis for the indicated subsets.
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