Background/Purpose: In healthy individuals, CD8 Treg activation leads to selective elimination of aberrantly activated self-reactive CD4 T cells to maintain immune balance. The CD8 Treg network appears dysfunctional in autoimmune diseases and insufficient to kill self-reactive CD4 T cells, in part due to expression of inhibitory KIR2DL1/2/3 that serves as an autoimmune checkpoint. We have developed MTX-101, a bispecific CD8 Treg modulator targeting CD8 and KIR2DL1/2/3, which are co-expressed on the surface of CD8 Treg cells. MTX-101 selectively binds CD8 Treg and enhances their killing of self-reactive CD4 T cells by blocking KIR2DL1/2/3 binding to its ligand. Enhanced CD8 Treg function prevents self-reactive CD4 T cell expansion and inflammation, without increasing unwanted immune cell activation or pro-inflammatory cytokines.

Methods: To determine the applicability of MTX-101 to the treatment of rheumatologic autoimmune disorders, we evaluated PBMCs derived from patients with systemic lupus erythematosus (SLE), Sjögren’s, ankylosing spondylitis, and psoriatic arthritis for KIR2DL1/2/3 expressing CD8 Treg. We observed that KIR2DL1/2/3 expressing CD8 Treg are present in all patients including in patients without active disease. Immunophenotyping and functional analysis suggest that CD8 Treg are impaired in these patient populations, showing reduced responsiveness to stimulation and expression of proteins critical to CD8 Treg functions relative to healthy donor CD8 Treg.

Results: We used human PBMCs and tissue resident immune cells to demonstrate selective binding to and activation of CD8 Treg of MTX-101. In vitro and in mouse models engrafted with human immune cells, MTX-101 increased cytolytic capacity, activation, and prevalence in healthy donor and autoimmune patient-derived CD8 Treg. In a model of acute inflammation, we observed CD8 Treg binding in peripheral blood and tissues, a reduction in inflammatory cytokines, reduced tissue pathology, and delayed onset of disease in mice treated with MTX-101. Pharmacokinetic and tolerability studies performed using IL-15 transgenic humanized mice, in which human lymphocytes engraft at physiologic ratios, demonstrated a half-life of 11.5 days following a single dose, and selective CD8 Treg binding and expansion, without unwanted immune cell activation or pro-inflammatory serum cytokines in animals treated with doses up to 10 mg/kg. Treatment with MTX-101 reduced the prevalence of activated CD4 T cells, and selectively increased the Granzyme B content and prevalence of the CD8 Treg population, but not of non-Treg CD8 T cells or NK cells, which may serve as clinical biomarkers.

Conclusion: Collectively, our data suggest that MTX-101-mediated disruption of the inhibitory autoimmune checkpoint KIR2DL1/2/3 can improve CD8 Treg dysfunctions observed in patients with rheumatologic autoimmune diseases to support durable re-balancing of the immune system.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/pre-clinical-characterization-of-mtx-101-a-novel-bispecific-cd8-treg-modulator-with-the-potential-to-restore-cd8-treg-functions-in-patients-with-rheumatological-autoimmune-diseases/