Background/Purpose: Aberrant activation and persistence of adaptive immune responses play a central role in autoimmune pathogenesis. Where traditional immunosuppressive therapies offer a broad targeting approach, reprogramming the adaptive immune system at a metabolic level represents a novel therapeutic strategy to selectively modulate disease-driving populations. Methylenetetrahydrofolate dehydrogenase 2 (MTHFD2), a mitochondrial enzyme critical to the folate cycle is robustly expressed in inflammatory disease tissue and upregulated in activated lymphocytes, providing a unique opportunity to target disease-relevant effector populations. We sought to develop MTHFD2-targeting inhibitors and to evaluate the effects of pharmacological MTHFD2 inhibition on key adaptive immune cell subsets and their therapeutic potential in models of autoimmunity.

Methods: The in vitro pharmacology of MTHFD2 inhibitors in specific lymphocyte subsets was tested in human primary immune cell cultures- key readouts included cell proliferation and cytokine production which were assessed using flow cytometry and HTRF. Human primary CD4⁺ T cells were stimulated with anti-CD3 and anti-CD28 to assess impact on T helper function. Th17 polarization was performed using purified human memory CD4⁺ T cells cultured under Th17-skewing conditions to assess effects on lineage commitment and effector cytokine production. Th1 antigen recall responses were explored in human PBMCs stimulated ex vivo using REVAXIS® (Diptheria, Tetanus and Poliomyelitis) antigens. B cell responses were analysed following BCR stimulation. In vivo efficacy was tested in a keyhole limpet hemocyanin (KLH)–induced delayed-type hypersensitivity (DTH) model and an adjuvant- induced arthritis (AIA) model in rat to assess impact of inhibition in autoimmune disease progression.

Results: MTHFD2 inhibition led to a marked reduction in the proliferation of CD4⁺ T cells, accompanied by a decrease in Th1 and Th17 signatures, without affecting overall cell viability. During Th17 polarisation, IL-17 production was significantly diminished. Antigen recall assays showed reduced proliferation and IFNγ production upon MTHFD2 inhibition, indicating impaired memory Th1 cell function. In B cells, treatment suppressed proliferation and IgG secretion upon stimulation. In vivo, MTHFD2 inhibition reduced KLH-specific IgG levels and ear swelling in the DTH model. In the AIA model, treatment significantly decreased paw swelling, correlating with reduced inflammation and periosteal bone formation.

Conclusion: MTHFD2 inhibition modulates key adaptive immune pathways by targeting memory Th1/Th17 cells and B cell effector functions- suppressing pathogenic cytokine production and antibody responses central to autoimmune pathology. These findings highlight MTHFD2 as a promising metabolic checkpoint for therapeutic intervention in autoimmune disease, with dual action on both T and B cell arms of adaptive immunity.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/mthfd2-is-a-novel-metabolic-target-in-autoimmune-disease/