Lightning Talk & Poster Presentation CD1-MR1 Workshop 2025

Characterization of bile acids as novel MR1 ligands (#21)

Alejandro Briones-Contreras 1 2 , Lucía Mosquera-Ferreiro 1 2 , Sofía Carreira-Santos 1 2 , Ana Ramírez-Iglesias 1 2 , Martin Kotrulev 1 2 , Beatriz García-Pinel 1 2 , Iria Gomez-Tourino 1 2
  1. Centre for Research in Molecular Medicine and Chronic Diseases (CiMUS), University of Santiago de Compostela, Santiago de Compostela, Spain
  2. Foundation Health Research Institute of Santiago de Compostela (IDIS), Santiago de Compostela, Spain

Background

Mucosal-associated invariant T (MAIT) cells are characterized by a TCR repertoire restricted by MR1, through which they can recognize bacterial and, interestingly, also self-metabolites. MAIT cells are the predominant T cell population in the liver, and recently some studies reported that bile acids (BAs) affect them. Cholic acid 3-sulfate (CA7S) is a BA and a MAIT antigen1, and there are conjugated BA that can modulate MAIT function2, but it is not known whether they modulate as MR1 ligands.

Aims

To identify bile acids capable of binding MR1 and modulating MAIT cell function.

Methods

We selected low-molecular weight BAs (<500 Da), as well as MAIT-modulatory BAs reported in the literature. We performed in silico virtual docking of BAs via Autodock-Vina into published MR1 crystal structures, followed by biophysical MR1 binding assays through surface plasmon resonance (SPR), and activation, blocking and competition assays employing MAIT TCR-transduced Jurkat cell lines.

Results

Virtual docking analyses identified six novel BAs that constitute in silico MR1 ligands. SPR assays showed that most of these BAs can indeed bind the binding groove of MR1. Coculture activation assays of MAIT-TCR-transduced Jurkat cells and BA-primed C1R.MR1 cells showed downregulation of MR1 expression in C1R.MR1 cells with no CD69 activation in Jurkat cells.

Competition assays show a significant reduction of 5-OP-RU-dependent Jurkat.MAIT cell activation, with this reduction being of different magnitude depending on the transduced TCR, indicating TCR specificity. We also performed competition assays inhibiting BA receptor (TGR5) signaling with 2',3'-dideoxyadenosine (DDA), demonstrating that the reduced CD69 levels in competition assays are not due to signaling through TGR5, but instead to an antagonistic effect of our novel BAs with regards to 5-OP-RU-mediated Jurkat.MAIT cell activation. Ongoing and future work includes validation in primary MAIT cells.

Conclusion

We identify four BAs that can bind MR1 and interfere with 5-OP-RU mediated MAIT cell activation, having great potential in inflammatory liver diseases.

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