Targeted erythrophagocyte reprogramming of Kupffer cells halts cancer immunotherapy associated liver toxicity [Multiplexed_CD40]

Agonistic CD40 monoclonal antibodies have emerged as promising immunotherapeutic compounds with impressive anti-tumoral effects. However, one of the main limitations of their use in patients is the severe liver necro-hepatitis they can induce. Up to this point, no effective treatment for anti-CD40 liver toxicity that does not hinder anti-tumor efficacy has been found. In the present study , we show that anti-CD40 liver toxicity is dependent on liver macrophages and recruitment of monocytes and neutrophils. We specifically reprogrammed the phenotype of liver macrophages to anti-inflammatory erythrophagocytes by using repeated injection of anti-erythrocyte murine Ter119 (mTer119) antibody. mTer119 administration induced phagocytosis of erythrocytes by liver macrophages and their transformation to Hmoxhigh/Marcohigh/MHC-IIlow erythrophagocytes. mTer119 treatment prevented anti-CD40 induced liver toxic side-effect, with significantly reduced elevation of plasma transaminases level and area of liver necrosis on histology, while preserving the anti-tumoral efficiency of anti-CD40. Our study offers a novel targeted therapeutic approach to treat immune-related liver adverse side effects of immunotherapies. Overall design: C57BL/6J mice were injected with 20 mg/kg anti-CD40 antibody (InVivoPlus BP0016-2) intraveinously or saline (Control mouse). At 7, 14 or 22 hours post injection, mice were euthanized and their liver digested into a non-parenchymal single-cell suspension. Cells were enriched for leucocytes using CD45+ coated magnetic beads (MagniSort™ Mouse CD45 Positive Selection Kit, 8802-6865-74), stained with Totalseq B hashtag antibodies (Biolegend) and pooled together at equal cell count before processing for single-cell RNA sequencing (Chromium Single Cell 3' Reagent Kits User Guide (v3.1 Chemistry Dual Index) with Feature Barcoding technology for Cell Surface Protein).