SAFETY & IMMUNOGENICITY
Cytokine release assays for preclinical CRS risk assessment
Could your biologic trigger excessive immune activation as you move toward IND and first-in-human studies?
Explicyte uses primary human immune-cell models to characterize cytokine-release liability across donors, concentrations and time points, with mechanistic follow-up when deeper investigation is needed.
FROM PRECLINICAL SAFETY TO FIRST-IN-HUMAN
Characterize cytokine-release liability before clinical exposure
For immune-activating biologics, the question is not simply whether cytokines are released, but how strongly, at what concentrations, with what kinetics, and how consistently across donors. When needed, complementary cellular readouts can help identify the immune-cell populations and biological programs associated with the response.
A fit-for-purpose human-cell assay helps characterize cytokine-release liability in the context of the molecule’s mechanism of action before first-in-human studies.”
Imane Nafia, PhD
Chief Scientific Officer, Explicyte
Cytokine release assay: format and readouts
- Relevant modalities
- T-cell engagers · multispecific antibodies · immune agonists · immunomodulatory antibodies · other immune-stimulating biologics
- Model
- Primary human PBMCs from multiple healthy donors.
- Format
- Concentration-response testing in soluble and/or immobilized formats.
- Donors & kinetics
- Multi-donor assessment across early, intermediate and late time points.
- Readouts
- Multiplex cytokine profiling, including IL-6, TNF-α, IFN-γ, IL-2 and IL-10.
- Controls
- Fit-for-purpose positive and negative controls, including TGN1412 where relevant.
- On request
- Flow cytometry, effector profiling and Xenium STAMP mechanistic analysis.
ASSAY PRINCIPLE
How the cytokine release assay works
1
Expose primary human PBMCs to the drug candidate
Healthy-donor PBMCs are treated across a defined concentration range using the assay format best suited to the molecule.
2
Capture cytokine release over time
3
Interpret magnitude, kinetics & donor variability
ANALYTICAL DEPTH
From cytokine-release screening to mechanistic characterization
Cytokine-release hazard assessment
Characterize cytokine release across healthy donors, concentrations and time points using fit-for-purpose controls.
- Primary human PBMCs
- Multi-donor testing
- Dose-response & kinetics
- Multiplex cytokine profiling
Resolve the immune-cell response
Add cellular readouts to identify responding immune populations and characterize their activation and functional state.
- Multiplex flow cytometry
- Immune-cell activation markers
- Intracellular effector markers
- Cytotoxic effector profiling
Identify cells & programs driving the response
Use Xenium STAMP to connect secreted cytokines with responding immune-cell populations and transcriptional programs.
- Cell-state resolution
- Cytokine-source identification
- Differential-expression analysis
- Pathway analysis by cell population
REPRESENTATIVE DATA
Representative cytokine-release studies
Regulatory context
Human-cell cytokine release assays in first-in-human risk assessment
For immune-stimulating biologics, cytokine-release assessment should be tailored to the molecule’s mechanism of action and the biological context of immune activation. FDA guidance recognizes soluble and immobilized PBMC or whole-blood assays, as well as more complex co-culture formats, and emphasizes fit-for-purpose methods with appropriate controls and measurable outcomes.
Interpretation should consider response magnitude and duration, the cytokines affected, donor variability and the underlying pharmacology. Together, these data can inform the broader first-in-human risk assessment, including starting-dose rationale, clinical monitoring and dose-escalation strategy.
Making a difference as a preclinical CRO in oncology
Why work with Explicyte for cytokine-release studies
Human primary-cell expertise
Multi-donor PBMC studies designed around immune responsiveness, assay sensitivity and inter-donor variability.
Fit-for-purpose study design
A PhD-level study director adapts donor number, exposure format, concentrations, controls, time points and readouts to the molecule’s mechanism.
One program, from efficacy to safety
For immune-redirecting biologics, connect tumor-cell killing and target-dependent immune activation with cytokine-release liability assessment.
Paul Marteau, PharmD (study director), Imane Nafia, PhD (CSO), Loïc Cerf, MSc (COO), Alban Bessede, PhD (founder, CEO), Jean-Philippe Guégan, PhD (CTO)
Contact our team
Discuss your cytokine-release study
Tell us your modality, target, development stage and the cytokine-release question you need to address. We'll help define the donor panel, assay format, concentration range, controls, kinetics and mechanistic readouts for a fit-for-purpose study.
CYTOKINE RELEASE ASSAY FAQ
Frequently asked questions about cytokine-release risk assessment
What does a cytokine release assay measure?
A cytokine release assay evaluates whether a drug candidate induces cytokine secretion from primary human immune cells. Responses are assessed across concentrations, donors and time points to characterize the magnitude, kinetics and inter-donor variability of cytokine release.
Can an in-vitro cytokine release assay predict clinical CRS?
No single in-vitro assay can predict whether a patient will develop CRS or its clinical severity. Cytokine release assays are used to identify and characterize immune-activation hazards under defined experimental conditions and contribute to the broader nonclinical risk assessment before first-in-human studies.
How many PBMC donors do you use?
We use multi-donor panels of primary human PBMCs to capture inter-individual variability. The number of donors is defined according to the molecule, study objective and development stage to provide a fit-for-purpose assessment of cytokine-release liability.
Which cytokines do you measure?
We use multiplex cytokine profiling covering key pro-inflammatory, T-cell activation and regulatory signals. Panels can include CRS-associated cytokines such as IL-6, TNF-α, IFN-γ, IL-2 and IL-10 and can be adapted to the biology of the drug candidate.
Do you use soluble and immobilized assay formats?
Yes. Drug candidates can be evaluated in soluble and/or immobilized formats depending on their mechanism of action and on whether receptor clustering or cross-linking may influence immune activation. The exposure format, concentration range, controls, and kinetics are defined during study design.
Can you assess target-dependent cytokine release?
Yes. When relevant to the mechanism of action, healthy-donor PBMC assays can be complemented with tumor–immune co-cultures incorporating target-positive and target-negative tumor cells. This can help distinguish cytokine release associated with target-dependent pharmacology from broader immune activation.
How can you investigate the mechanism behind a cytokine-release signal?
The study can be extended beyond secreted cytokine measurements using flow-cytometry immunophenotyping, activation and effector markers, additional kinetic measurements or Xenium STAMP. These complementary approaches can help identify responding immune-cell populations, characterize changes in cell state over time and resolve transcriptional programs associated with the cytokine response.
What controls are included in a cytokine release assay?
Positive and negative controls are selected according to the molecule’s mechanism of action and the assay format. Reference immune-activating biologics, including TGN1412 where relevant, can be incorporated to confirm assay responsiveness and support interpretation of the test article response.