Introduction
Despite advances in biologic therapies, up to 70% of Crohn's disease (CD) patients require ileocolic resection (IR) within 10 years of diagnosis, and over 60% develop postoperative recurrence (POR) within one year(1). Biologic efficacy in preventing POR varies substantially, underscoring the need to identify immunological mechanisms that drive recurrence versus sustained remission (2).
Aims & Methods
This study aimed to identify transcriptomic and immune cell signatures in peripheral blood mononuclear cells (PBMCs) associated with endoscopic recurrence following IR in CD.
Bulk RNA sequencing was performed on PBMCs from 34 CD patients (44% male) following IR: 17 in endoscopic remission (Rutgeerts i0–i1) and 17 with endoscopic recurrence (i2–i4). Differential gene expression (DGE) analysis was conducted using the limma-voom pipeline, adjusting for biological therapy, sex, as well as four surrogate variables (SVA)(3,4). Immune cell composition was estimated using EPIC deconvolution. Pathway enrichment analysis was performed using Gene Set Enrichment Analysis (GSEA) with gene sets from MSigDB Hallmark, Reactome, and C7 immunologic.
Results
Cell-type deconvolution revealed a striking shift in circulating immune cell composition. NK cells were enriched (median 0.127 vs. 0.088; p=0.0175) while CD8 T cells were markedly depleted (median 0.001 vs. 0.050; p=0.008) in endoscopic recurrence patients. These findings were consistent across biological therapy subgroups. At the gene level, DGE analysis identified 661 nominally significant genes (p<0.05), with none surviving FDR correction. Among the most differentially expressed genes, CXCR2 was notably upregulated in patients with recurrence (logFC = +0.613; p = 0.0008), reflecting neutrophil recruitment and innate immune activation. In a targeted analysis of canonical Crohn's disease marker genes, ITGA4 (logFC = −0.159; p = 0.004) and STAT1 (logFC = −0.390; p = 0.019) were significantly downregulated in recurrence compared to remission, suggesting reduced gut-homing lymphocyte trafficking via the α4β7 integrin pathway and attenuation of JAK/STAT interferon signalling in recurrence. Pathway analysis revealed a broad and coordinated suppression programme in patients with recurrence. Immune pathways, including IFN-γ response (NES=−2.89), IFN-α response (NES=−2.83), TNF-α/NF-κB signalling (NES=−2.71), were suppressed, alongside metabolic programmes such as oxidative phosphorylation, fatty acid metabolism, and myogenesis. Reactome analysis confirmed suppression of TCR/BCR signalling and MHC class I/II antigen presentation, with activation of neutrophil degranulation. C7 signatures showed suppression of CD4 memory T cell and effector T cell programmes (all FDR<0.05). Mechanistically, CD8 T cell abundance correlated positively with both IFN-γ (ρ=0.399; p=0.019) and TNF-α/NF-κB transcriptional scores (ρ=0.345; p=0.046), identifying CD8 T cell depletion as a likely driver of the observed interferon suppression.
Conclusion
Post-operative CD recurrence is associated with a systemic immune signature in PBMCs. This signature is characterised by coordinated suppression of interferon signalling, lymphocyte activation, and cellular metabolic programmes including oxidative phosphorylation, fatty acid metabolism, and myogenesis. This is accompanied by NK cell enrichment and CD8 T cell depletion. These findings reveal a dissociation between systemic and mucosal immune responses and support the use of PBMC transcriptomic and cellular features as candidate biomarkers for post-operative monitoring in CD.
References
1. Sensi B, Siragusa L, Efrati C, Petagna L, Franceschilli M, Bellato V, et al. The Role of Inflammation in Crohn’s Disease Recurrence after Surgical Treatment. J Immunol Res. 2020 Dec 26;2020:8846982. doi:10.1155/2020/8846982 PubMed PMID: 33426097; PubMed Central PMCID: PMC7781709.
2. Cruz PD, Kamm MA, Hamilton AL, Ritchie KJ, Krejany EO, Gorelik A, et al. Crohn’s disease management after intestinal resection: a randomised trial. The Lancet. 2015 Apr 11;385(9976):1406–17. doi:10.1016/S0140-6736(14)61908-5 PubMed PMID: 25542620.
3. voom: precision weights unlock linear model analysis tools for RNA-seq read counts | Genome Biology | Springer Nature Link. Available from: https://link.springer.com/article/10.1186/gb-2014-15-2-r29
4. Leek JT, Johnson WE, Parker HS, Jaffe AE, Storey JD. The sva package for removing batch effects and other unwanted variation in high-throughput experiments. Bioinformatics. 2012 Mar 15;28(6):882–3. doi:10.1093/bioinformatics/bts034
Disclosure
Robert Battat: Speaker/consulting/moderator: Bristol Myers Squibb, Johnson and Johnson Innovative Medicine, AbbVie, Takeda, Ferring, Celltrion, Pfizer, Merck, Eli Lilly
Advisory boards: Celltrion, AbbVie, Pfizer, Janssen, Bristol Myers Squibb, Takeda, Merck, Eli Lilly
Educational grants and educational sponsorships: Abbvie, Johnson and Johnson Innovative Medicine, Pfizer, Eli Lilly, Amgen, Pendopharm, Merck, Celltrion, Organon
Travel support: Pfizer, Celltrion, Johnson and Johnson Innovative Medicine, Abbvie.