Human organoids with an autologous tissue-resident immune compartment.

The intimate relationship between the epithelium and immune system is crucial for maintaining tissue homeostasis, with perturbations therein linked to autoimmune disease and cancer1-3. Whereas stem cell-derived organoids are powerful models of epithelial function4, they lack tissue-resident immune cells that are essential for capturing organ-level processes. We describe human intestinal immuno-organoids (IIOs), formed through self-organization of epithelial organoids and autologous tissue-resident memory T (TRM) cells, a portion of which integrate within the epithelium and continuously survey the barrier. TRM cell migration and interaction with epithelial cells was orchestrated by TRM cell-enriched transcriptomic programs governing cell motility and adhesion. We combined IIOs and single-cell transcriptomics to investigate intestinal inflammation triggered by cancer-targeting biologics in patients. Inflammation was associated with the emergence of an activated population of CD8+ T cells that progressively acquired intraepithelial and cytotoxic features. The appearance of this effector population was preceded and potentiated by a T helper-1-like CD4+ population, which initially produced cytokines and subsequently became cytotoxic itself. As a system amenable to direct perturbation, IIOs allowed us to identify the Rho pathway as a new target for mitigation of immunotherapy-associated intestinal inflammation. Given that they recapitulate both the phenotypic outcomes and underlying interlineage immune interactions, IIOs can be used to study tissue-resident immune responses in the context of tumorigenesis and infectious and autoimmune diseases.

Analysis of off-tumour toxicities of T-cell-engaging bispecific antibodies via donor-matched intestinal organoids and tumouroids.

Predicting the toxicity of cancer immunotherapies preclinically is challenging because models of tumours and healthy organs do not typically fully recapitulate the expression of relevant human antigens. Here we show that patient-derived intestinal organoids and tumouroids supplemented with immune cells can be used to study the on-target off-tumour toxicities of T-cell-engaging bispecific antibodies (TCBs), and to capture clinical toxicities not predicted by conventional tissue-based models as well as inter-patient variabilities in TCB responses. We analysed the mechanisms of T-cell-mediated damage of neoplastic and donor-matched healthy epithelia at a single-cell resolution using multiplexed immunofluorescence. We found that TCBs that target the epithelial cell-adhesion molecule led to apoptosis in healthy organoids in accordance with clinical observations, and that apoptosis is associated with T-cell activation, cytokine release and intra-epithelial T-cell infiltration. Conversely, tumour organoids were more resistant to damage, probably owing to a reduced efficiency of T-cell infiltration within the epithelium. Patient-derived intestinal organoids can aid the study of immune-epithelial interactions as well as the preclinical and clinical development of cancer immunotherapies.

Conserved γδ T cell selection by BTNL proteins limits progression of human inflammatory bowel disease.

Murine intraepithelial γδ T cells include distinct tissue-protective cells selected by epithelial butyrophilin-like (BTNL) heteromers. To determine whether this biology is conserved in humans, we characterized the colonic γδ T cell compartment, identifying a diverse repertoire that includes a phenotypically distinct subset coexpressing T cell receptor Vγ4 and the epithelium-binding integrin CD103. This subset was disproportionately diminished and dysregulated in inflammatory bowel disease, whereas on-treatment CD103+γδ T cell restoration was associated with sustained inflammatory bowel disease remission. Moreover, CD103+Vγ4+cell dysregulation and loss were also displayed by humans with germline BTNL3/BTNL8 hypomorphism, which we identified as a risk factor for penetrating Crohn's disease (CD). Thus, BTNL-dependent selection and/or maintenance of distinct tissue-intrinsic γδ T cells appears to be an evolutionarily conserved axis limiting the progression of a complex, multifactorial, tissue-damaging disease of increasing global incidence.

miR-29b directly targets activation-induced cytidine deaminase in human B cells and can limit its inappropriate expression in naïve B cells.

Class-switch recombination (CSR) is an essential B cell process that alters the isotype of antibody produced by the B cell, tailoring the immune response to the nature of the invading pathogen. CSR requires the activity of the mutagenic enzyme AID (encoded by AICDA) to generate chromosomal lesions within the immunoglobulin genes that initiate the class switching recombination event. These AID-mediated mutations also participate in somatic-hypermutation of the immunoglobulin variable region, driving affinity maturation. As such, AID poses a significant oncogenic threat if it functions outside of the immunoglobulin locus. We found that expression of the microRNA, miR-29b, was repressed in B cells isolated from tonsil tissue, relative to circulating naïve B cells. Further investigation revealed that miR-29b was able to directly initiate the degradation of AID mRNA. Enforced overexpression of miR-29b in human B cells precipitated a reduction in overall AID protein and a corresponding diminution in CSR to IgE. Given miR-29b's ability to potently target AID, a mutagenic molecule that can initiate chromosomal translocations and "off-target" mutations, we propose that miR-29b acts to silence premature AID expression in naïve B cells, thus reducing the likelihood of inappropriate and potentially dangerous deamination activity.