Supplementary Figure S12. Individual tumor growth curves for each treatment group. OCI-Ly18 (DLBCL)-bearing humanized NSG mice were pre-treated with 30 mg/kg Gazyva (Gpt, grey arrow) and treated weekly with escalating doses of CD20-TCB (0.5 mg/kg, 1 mg/kg, 2 mg/kg, red arrows). Adalimumab (25 mg/kg, orange arrow, TNF-α blockade) and tocilizumab (10 mg/kg, green arrow, IL-6R blockade) were given 1 day and 1 hour before the first CD20-TCB infusion. Dexamethasone (1 mg/kg, blue arrow) was given 1 hour prior each therapy. Anakinra (10 mg/kg, purple arrow, IL-1R blockade) and the NLRP3 inhibitor (20 mg/kg, light blue arrow) were administered 1 hour before, 4 hrs and 24 hrs after the first CD20-TCB infusion as well as 1 hr before and 4 hrs after the second CD20-TCB treatment. Individual tumor growth curves for each treatment group.
ARTICLE ABSTRACT
Purpose: Target-dependent TCB activity can result in strong and systemic release of cytokines that may develop into Cytokine Release Syndrome (CRS), highlighting the need to understand and prevent this complex clinical syndrome. Experimental design: We explored the cellular and molecular players involved in TCB-mediated cytokine release by single cell RNA sequencing of whole blood treated with CD20-TCB together with bulk RNA sequencing of endothelial cells exposed to TCB-induced cytokine release. We used the in vitro whole blood assay and an in vivo DLBCL model in immunocompetent humanized mice to assess the effects of dexamethasone, anti-TNF-a, anti-IL-6R, anti-IL-1R and inflammasome inhibition, on TCB-mediated cytokine release and anti-tumor activity. Results: Activated T cells release TNF-α, IFN-γ, IL-2, IL-8 and MIP-1β, which rapidly activate monocytes, neutrophils, DCs and NKs along with surrounding T cells to amplify the cascade further, leading to TNF-α, IL-8, IL-6, IL-1β, MCP-1, MIP-1α, MIP-1β and IP-10 release. Endothelial cells contribute to IL-6 and IL-1β release and at the same time release several chemokines (MCP-1, IP-10, MIP-1α and MIP-1β). Dexamethasone and TNF-α blockade efficiently reduced CD20-TCB-mediated cytokine release while IL-6R blockade, inflammasome inhibition and IL1-R blockade induced a less pronounced effect. Dexamethasone, IL-6R blockade, IL-1R blockade and the inflammasome inhibitor did not interfere with CD20-TCB activity, in contrast to TNF-α blockade, which partially inhibited anti-tumor activity. Conclusion: Our work sheds a new light on the cellular and molecular players involved in cytokine release driven by TCBs and provides rationale for the prevention of CRS in patients treated with TCBs.
