The close relation between Severe Acute Respiratory Syndrome (SARS), unpaired adaptive immune responses and overexpression of inflammatory cytokines has been described more than a decade ago (1, 2).
More recently, the notion of “cytokine storm” or “cytokine release syndrome” has been used by Life Scientists when they have monitored the expression level dynamics of cytokines in the sera of COVID-19 patients with acute lung damages and respiratory distress. (3)
The numerous publications focused on the recent COVID-19 outbreak have confirmed this intimate link between the “cytokine storm” and acute SARS-CoV-2 infections. For example, severe respiratory syndromes have been associated with an enhanced production of pro-inflammatory factors (e.g.
IL-6, GM-CSF, TNFα, IL-10 and soluble IL-2R) and type-I IFN modulation. A decreased IFNγ-expression in CD4+ T cells has been published in COVID-19 patients with acute lung damages. In parallel, high sera levels of IP-10, MCP-3 and IL-1r have been also associated with SARS-CoV disease severity. (3-5)
Why monitor cytokine expression in early antiviral drug discovery?
The monitoring of the dynamic expression of INFs and/or pro-inflammatory cytokines in the early stages of antiviral drug development is of interest for multiple reasons mainly.
First, it allows a clear understanding of the mechanisms of action (MoA) of viral-induced immune responses and the Cytokine Release Syndrome (CRS) related lung damages. (5)
Secondly, it helps in the identification of new potentially interesting inflammatory-related druggable targets to cure SARS-CoV infected patients. For example, the E3 UB ligase inhibitor Thalidomide, the Sphingosine-1-phosphate receptor agonist Fingolimod (FTY-720), and the MEK inhibitor Trametinib have been described to limit the “Cytokine Storm” induced by SARS-CoV-2 & MERS-CoV. They are now potential “Cytokine storm” drugs.
More recently, the humanized monoclonal antibody against the interleukin-6 receptor (IL-6R) (Tocilizumab, used in rheumatoid arthritis) have been described to improve significantly clinical outcomes of patients with moderate or severe COVID-19 pneumonia.
To perform such monitoring, various experimental approaches can be considered.
Inflammatory cytokines monitoring tool box
Researchers and drug discoverers can decide to analyze a range of cytokines of interest in one experiment either semi-quantitatively (profiling) or quantitatively (multiplexing). These Antibody Planar Arrays are ideal for biomarker identification.
Popular Inflammation Antibody Array Kits & Services
| Array | Type | #Targets | Targets Detected |
|---|---|---|---|
| Human Inflammation Array GS1 | Semi-quantitative | 10 | CCL2, CXCL8, IFNG, IL10, IL13, IL1A, IL1B, IL4, IL6, TNF |
| Human Inflammation Array G3 | Semi-Quantitative | 40 | CCL1, CCL11, CCL15, CCL2, CCL24, CCL3, CCL4, CCL5, CCL8, CSF1, CSF2, CSF3, CXCL10, CXCL8, CXCL9, ICAM1, IFNG, IL10, IL11, IL12A, IL12B, IL13, IL15, IL16, IL17A, IL1A, IL1B, IL2, IL3, IL4, IL6, IL6R, IL7, LTA, PDGFB, TGFB1, TIMP2, TNF, TNFRSF1A, TNFRSF1B |
| Human Inflammation Array Q1 | Quantitative | 10 | IL-1 alpha, IL-1 beta, IL-4, IL-6, IL-8, IL-10, IL-13, MCP-1, IFN-gamma, TNF alpha |
| Human Inflammation Array Q3 | Quantitative | 40 | BLC, Eotaxin-1, Eotaxin-2, G-CSF, GM-CSF, I-309, ICAM-1, IFN-gamma, IL-1 alpha, IL-1 beta, IL-1 ra, IL-2, IL-4, IL-5, IL-6, IL-6 sR, IL-7, IL-8, IL-10, IL-11, IL-12 p40, IL-12 p70, IL-13, IL-15, IL-16, IL-17, MCP-1, M-CSF, MIG, MIP-1 alpha, MIP-1 beta, MIP-1 delta, PDGF-BB, RANTES, TIMP-1, TIMP-2, TNF alpha, TNF beta, sTNFRI, sTNFRII |
| Human Th1/Th2 Array Q1 | Quantitative | 10 | GM-CSF, IFN-gamma, IL-10, IL-13, IL-2, IL-4, IL-5, IL-6, IL-8 (CXCL8), TNF alpha |
ELISA Kits & Services for “cytokine storm” monitoring
In parallel to broad cytokine analysis, researchers might prefer to focus on a small number of cytokine(s) of interest. For this purpose, conventional ELISA tests are ideal mainly for biomarker validation or later in the clinical stages to evaluate the expression dynamics of human cytokines.
Selection of SARS-CoV related ELISAs
- IL-6, IL-10, TNFα, IFNγ, IP-10, MCP-3, IL-1ra, IL-1, GM-CSF,
- Hyaluronan, CXCL10/IFN-γ-inducible protein 10,
- CCL2/monocyte chemotactic protein 1
- D-dimer, Procalcitonin…
Interestingly, new multiplex technologies now allow reproducible and simultaneous quantification of a small onset of cytokines of interest in one experiment (pg/ml range).
Such Multiplex Assays are performed in tebu-bio’s laboratories as Services. They are not only a cost-effective alternative for measuring inflammatory cytokine(s) from less than 50µl of serum/plasma or cell culture supernatants, but also when high assay reproducibility is required when quantifying one cytokine in hundreds of samples.
| MultiPlex ELISA | #targets | Targets |
| Human Cytokine | 4-Plex | IL-6, TNFα, IFNγ, IL-10 |
| Human Cytokine Inflammation | 9-Plex | IL-1α, IL-1β, IL-2, IL-4, IL-6, IL-8, IL-10, IFNγ, TNFα |
| Human IFNs & pro-inflammatory cytokines | 9-Plex | IFN-alpha, IFN-beta, IFN-gamma, IFN-lambda, IFN-omega, IL-1alpha, IL-6, IP-10 and TNF-alpha |
| NEW! Human Cytokine Release Syndrome | 16-Plex | IL-1b, IL-1Ra, IL-2, IL-2Ra, IL-6, IL-6R, IL-8, IL-10, IL-12p70, IL-13, IFNy, INFa, TNFa, GM-CSF, MCP-1, MIP-1a |
Monitoring inflammatory cytokines?
What about your project? Which inflammatory cytokines are you monitoring in your antiviral drug discovery or repositiong programs?
Leave a message below or do not hesitate to contact us for further information concerning your projects and our cytokine profiling / quantification services!
Sources
1- Chung Y. et al. “Cytokine Responses in Severe Acute Respiratory Syndrome Coronavirus-Infected Macrophages In Vitro: Possible Relevance to Pathogenesis” (2005) Journal of Virology, 79(12): 7819–7826. doi: 10.1128/JVI.79.12.7819-7826.2005
2- He L. “Expression of elevated levels of pro-inflammatory cytokines in SARS-CoV-infected ACE2+ cells in SARS patients: relation to the acute lung injury and pathogenesis of SARS” (2006) Jpurnal of Pathology, 210(3):288-97. doi: doi.org/10.1002/path.2067
3- Pedersen S.F. et al. “SARS-CoV-2: A Storm is Raging” (2020) The Journal of Clinical Investigation
4- Yang Y. et al. “Exuberant elevation of IP-10, MCP-3 and IL-1ra during SARS-CoV-2 infection is associated with disease severity and fatal outcome” (2020) medRxiv; doi: doi.org/10.1101/2020.03.02.20029975
5- Shi et. al. “COVID-19 infection: the perspectives on immune responses” (2020) Cell Death Differ (2020). doi: doi.org/10.1111/bph.13947
