DOP132 An integrated single-cell atlas enables the discovery and validation of a novel candidate therapeutic target for Inflammatory Bowel Disease
Authors: De Baets G , D'Rozario J , Mattson J , Castiblanco D , Kaymak I , Moldoveanu A L , Camilli G , Prandi F , Chouri E , Popov N , Jawale C , Escudero-Ibarz L , Oliphant C , Nayar S , Kline-Schoder A , Leonardi I , Joseph A , Geller E , Vollmer E , Pizzarella V , Honan A , Mediratta D , Freinkman E , Vijay P , Wroblewska A , Platt A , Ort T , Corridoni D 
Published: January 2025

Background

Single-cell technologies enable the fine mapping of disease and treatment mechanisms in inflammatory bowel disease (IBD). We sought to leverage these insights to discover novel drug targets with precise therapeutic hypotheses. To this end, we previously constructed one of the largest integrated single-cell atlas of IBD patient-derived tissue samples1. We then used Immunai’s ImmunoDynamics EngineTM, a proprietary machine learning (ML) framework, to identify transcriptional signatures of inflammation and non-response to anti-TNF treatment in specific cell populations, and extracted a ranked gene list that was highly enriched in targets of currently marketed IBD therapies. We selected top-ranked genes for in vitro functional genomic validation and leveraged the IBD single-cell atlas to support clinical relevance of in vitro observations. Here, we present a target case study demonstrating this approach.

Methods

The top 400 genes were triaged to choose 20 candidate targets based on novelty, druggability, genetic association with IBD, and experimental feasibility. Each of these 20 targets was individually deleted under optimized culture conditions in the primary human cell type(s) in which it showed a significant disease association in the IBD single-cell atlas, including monocyte-derived macrophages (MDM), CD8, CD4 helper and/or regulatory T cells, and intestinal fibroblasts. Knockout (KO) cells were characterized by flow cytometry, secreted proteome profiling, and bulk transcriptomics, with suitable positive and negative controls per cell type. KO-induced transcriptomic changes were interpreted via gene-set enrichment of canonical pathways and of IBD-associated transcriptional signatures derived from orthogonal clinical cohorts. KO-induced transcriptomic signatures were also computed in each sample in the IBD single-cell atlas.

Results

Deletion of one selected target in MDMs led to a reduction in TNFα and IFNγ signaling and in the expression of IBD inflammation-associated signatures from orthogonal patient cohorts. Genes downregulated by the deletion of this target in MDMs were enriched in macrophages from inflamed tissue in the IBD single-cell atlas.

Conclusion

The functional genomic data indicate that deletion of the selected target shifts MDMs away from a disease-associated inflammatory state, suggesting that its pharmacologic inhibition in macrophages may have a therapeutic benefit. These findings highlight the relevance of cell type-specific methods to understand disease mechanisms and characterize putative novel targets. Applying these experimental and clinical contextualization methods to additional candidate genes will further expand the basis for novel therapeutic strategies in IBD.

References

1. De Baets G, d’Rozario J, Gehrmann U, et al. Inflammatory Bowel Disease single cell atlas construction to enable cell type-specific target identification. J Crohns Colitis 2024;18(Supplement_1), i143. doi: 10.1093/ecco-jcc/jjad212.0080