Single-Cell RNA Sequencing of Tocilizumab-Treated Peripheral Blood Mononuclear Cells as an in vitro Model of Inflammation

Abstract

COVID-19 has posed a significant threat to global health. Early data has revealed that IL-6, a key regulatory cytokine, plays an important role in the cytokine storm of COVID-19. Multiple trials are therefore looking at the effects of Tocilizumab, an IL-6 receptor antibody that inhibits IL-6 activity, on treatment of COVID-19, with promising findings. As part of a clinical trial looking at the effects of Tocilizumab treatment on kidney transplant recipients with subclinical rejection, we performed single-cell RNA sequencing of comparing stimulated PBMCs before and after Tocilizumab treatment. We leveraged this data to create an in vitro cytokine storm model, to better understand the effects of Tocilizumab in the presence of inflammation. Tocilizumab-treated cells had reduced expression of inflammatory-mediated genes and biologic pathways, particularly amongst monocytes. These results support the hypothesis that Tocilizumab may hinder the cytokine storm of COVID-19, through a demonstration of biologic impact at the single-cell level.

Role of Tocilizumab in Down Regulating sCD163 Plasmatic Levels in a Cohort of COVID-19 Patients

“…Similarly, Guo et al., performing a scRNA-seq of two patients with severe COVID-19 pre- and post-treatment with tocilizumab, observed a reduced enrichment of inflammatory pathways as well as a reduced expression of IL-6 receptor related pathways genes in tocilizumab-treated cells. Moreover, the authors showed an enrichment in CD14 expression associated with the presence of non-inflammatory classical monocytes, in tocilizumab-treated cells ( 30 , 31 ). All these findings, together with the available clinical data, support the belief that tocilizumab may be effective in reducing the monocytes-related inflammatory burden that results in the adverse outcomes of COVID-19.…”

“…The idea that in COVID-19 patients tocilizumab may suppress the cytokine storm by decreasing the activity of IL-6, is corroborated by the findings of Zarinsefat et al., who speculated on the mechanistic/biologic effects of this drug on immune system cells using an in vitro cytokine storm model of peripheral blood mononuclear cells (PBMC) ( 30 ). Specifically, the authors comparing single-cell RNA sequencing (scRNA-seq) of stimulated PBMC from kidney transplant recipients with subclinical rejection with and without tocilizumab treatment, showed that tocilizumab-treated PBMC had reduced expression of inflammatory-mediated genes and biologic pathways, particularly amongst monocytes ( 30 , 31 ). Similarly, Guo et al., performing a scRNA-seq of two patients with severe COVID-19 pre- and post-treatment with tocilizumab, observed a reduced enrichment of inflammatory pathways as well as a reduced expression of IL-6 receptor related pathways genes in tocilizumab-treated cells.…”

Identifying COVID‐19 subtypes by single‐sample gene set enrichemnt analysis and providing guidance for sensitive drug selection

“…Immune responses against SARS‐Cov‐2 infection differ between individuals. Many in vitro experiments show drugs targeting inflammatory cytokines seem to be effective for experiment subjects, 21–24 but in vivo experimental results are not ideal or even have no effect 25–31 . We notice that COVID‐19 patients participating in drug trials are selected randomly and whether expression levels of genes that drugs target have changed related to baseline levels is not considered.…”

Case Report: Tocilizumab for Acute Kidney Graft Dysfunction in Patient Affected by COVID-19

“…Several reports have described the use of tocilizumab administration promoting a favorable outcome in some transplant patients focused on reduced requirement of oxygen (O 2 ) therapy and on improvement of lung lesions and other studies reported higher mortality in transplant patients treated with tocilizumab ( 18 – 20 ); however, none of these reported the effects on graft function. Alternatively, there is a paucity of data on the mechanistic and biological impact of tocilizumab on the immune system; recently, Zarinsefat et al ( 21 ) explored the impact of tocilizumab on immune system activation through stimulated cells derived by kidney-transplanted patients with subclinical rejection enrolled in an investigator-initiated clinical trial ( NIAID U01 AI113362-01 ; https://grantome.com/grant/NIH/U01-AI113362-06 ). Interestingly, the authors showed that many inflammation-related pathways linked to acute rejection were suppressed in cells exposed to tocilizumab.…”