MODALITY-SPECIFIC PRECLINICAL ASSAYS
ADC preclinical assays & CRO services: target qualification, efficacy and translational profiling
Understand what drives ADC activity — from target expression and internalization to payload sensitivity, tumor heterogeneity and patient-derived response.
Explicyte designs fit-for-purpose in vitro and ex vivo studies for antibody-drug conjugates, combining human tissue target profiling, functional ADC characterization, kinetic tumor cell killing in 2D cultures and 3D models, including patient-derived models.
Target qualification · Binding & internalization · Kinetic cytotoxicity · Payload sensitivity · Bystander activity · Patient-derived models · Mechanistic profiling
ADC BIOLOGY
Target expression is only the beginning
A tumor can express the ADC target and still respond poorly. Conversely, ADC activity may persist in tumors with relatively low or heterogeneous target expression. ADC response reflects multiple interconnected biological determinants:
Target biology
Is the target expressed on the tumor cell surface, at what level, and how heterogeneous is its distribution across tumors and models?
ADC engagement and uptake
Does the ADC bind selectively to target-expressing cells and undergo efficient target-dependent internalization?
Payload sensitivity
How intrinsically sensitive is the tumor cell to the cytotoxic mechanism delivered by the ADC?
Tumor heterogeneity and bystander activity
Can cytotoxic activity extend from target-positive cells to neighboring target-low or target-negative populations?
Cellular context and resistance
Which tumor cell states or molecular programs are associated with sensitivity or resistance?
Explicyte integrates all these dimensions into one study strategy, built around the biology of the ADC and your development question, to provide an integrated understanding of the response.
The value of ADC biology is not just in showing where the target is expressed. It is in understanding how tumor biology shapes response — and generating the evidence to support translational and development decisions.
Imane Nafia, PhD
Chief Scientific Officer, Explicyte
CASE STUDY · ANTI-TROP2 ADC
Anti-TROP2 ADC: target expression guides model selection, functional assays explain response
Model selection based on TROP2 expression
Step 1 — Qualify TROP2 expression across a cancer cell line panel
Cell surface TROP2 expression was characterized across a panel of human cancer cell lines including SKOV3, T24, MDA-MB-231, PC3 and A549.
This first step identified relevant TROP2-positive models for functional testing and provided the biological context needed to interpret differences in ADC response.
Key outputs
- Cell surface TROP2 expression
- Inter-model comparison
- Model selection
Real-time ADC cytotoxicity
Step 2 — Measure functional ADC response over time
TROP2-positive SKOV3 and MDA-MB-231 cells were treated with sacituzumab govitecan and monitored by live-cell imaging.
Although both models expressed the therapeutic target, they displayed distinct sensitivity profiles to the ADC, highlighting the importance of pairing target-expression data with functional response measurements.
Key outputs
- Proliferation & apoptosis kinetics
- Concentration-dependent activity
- Differential response between tumor models
ADC vs payload
Step 3 — Put ADC response in the context of payload sensitivity
In SKOV3 cells, sacituzumab govitecan was compared with irinotecan, a prodrug of SN-38, to contextualize ADC activity relative to sensitivity to topoisomerase-I inhibition.
This comparison provides additional information on the intrinsic sensitivity of the tumor model to the payload mechanism, and can help guide subsequent mechanistic investigations.
Key outputs
- Comparative growth inhibition
- Apoptosis kinetics
- AUC-based treatment response
Build the study around the biological question
Core ADC characterization
Four questions structure the core ADC framework — from target biology to functional response in heterogeneous tumors. Each question is mapped to models, readouts and platforms selected according to the molecule and development objective.
| Step | Stage & development question | Models | Key readouts | Platforms & technologies |
|---|---|---|---|---|
| 01 | ADC target qualification & model selection Is the target expressed in the relevant tumors and models? Characterize tumor vs normal tissue expression, membrane localization, prevalence and heterogeneity, to inform model selection and translational context, and potential on-target/off-tumor liabilities. | FFPE tumor cohorts & relevant normal tissue panels; cancer cell line panels stratified into target-high, target-low and target-negative tiers | Target prevalence · membrane localization · intra-tumor heterogeneity · cell surface expression (MFI) · target expression in relevant normal tissue | Multiplex IHC/IF, flow cytometry |
| 02 | ADC binding & internalization Does the ADC bind specifically and internalize efficiently? Target expression alone does not establish productive ADC uptake; binding and internalization kinetics provide a functional assessment of target engagement. | Target-high, target-low and target-negative cell lines with isotype-matched controls | Binding intensity & specificity · dose response · internalization kinetics · intracellular uptake | Flow cytometry, live-cell imaging, pH-sensitive fluorescent probes |
| 03 | ADC efficacy & payload contribution Does the ADC induce target-dependent tumor cell killing, and how does this relate to payload sensitivity? Kinetic live-cell imaging captures how tumor cell killing develops over time, with control arms helping contextualize the contribution of target engagement and payload sensitivity. | 2D tumor cell lines spanning differential target-expression levels; comparator arms with backbone antibody, ADC isotype, and free payload. | Live-cell imaging, dose-response modelling | |
| 04 | Heterogeneity & bystander activity How does target heterogeneity influence response? Mixed cell models can test how ADC activity changes as target-positive and target-negative populations are combined at defined proportions. | Mixed cell co-cultures at defined target-positive / target-negative ratios; patient-derived organoids and fragments where relevant | Bystander killing · target-density dependence · population-specific tumor cell elimination kinetics · response across mix ratios | Live-cell imaging; flow cytometry; 3D models |
PRECLINICAL ADC CHARACTERIZATION
ADC assays aligned with your development questions
Build a fit-for-purpose ADC study from target qualification to functional efficacy, mechanism and translational biomarkers. Each program can combine the assays and models most relevant to the molecule, indication and stage of development.
Is the target expressed in the right tumors and models?
Characterize target prevalence, membrane localization and heterogeneity in tumor and normal tissues, and quantify cell-surface expression across cancer cell lines to select relevant experimental models.
Does the ADC bind specifically and internalize efficiently?
Confirm target-dependent binding and quantify ADC uptake over time in models with defined antigen expression.
Does the ADC induce target-dependent tumor-cell killing?
Quantify dose- and time-dependent effects on proliferation, viability and apoptosis, with appropriate comparators such as unconjugated antibody, non-targeting ADC and free payload or relevant payload controls.
How does target heterogeneity influence response?
Use target-high, target-low and target-negative mixed cultures and, where appropriate, patient-derived 3D tumor models to investigate differential sensitivity and bystander killing. Fc-dependent immune mechanisms can also be evaluated where relevant.
What drives response or resistance?
Integrate functional data with flow cytometry, transcriptomics, spatial profiling or Xenium STAMP to identify the cellular and molecular programs associated with ADC response or resistance.
EXTEND THE STUDY WHEN THE BIOLOGY REQUIRES IT
Add complexity when it answers a development question
Modules can be added to core framework when a specific biological or translational question requires deeper resolution.
Fc-dependent immune activity
For ADCs with an Fc capable of engaging immune effector mechanisms, tumor cell killing can be evaluated in the presence of human PBMCs or isolated effector cell populations. This allows Fc-mediated activity, including ADCC, to be evaluated alongside payload-driven cytotoxicity.
Patient-derived 3D tumor models
Move beyond established cell lines into models that better represent patient-specific tumor biology, target heterogeneity, and tissue context. Early patient-derived organoids provide flexible 3D systems for functional treatment testing, while intact tumor fragments additionally retain native tissue architecture and spatial organization.
Mechanism of response and resistance
When models with similar target expression show different sensitivity profiles to the ADC, deeper profiling can generate hypotheses around cellular states, pathways, and molecular features associated with sensitivity or resistance. Functional data can be integrated with flow cytometry, single-cell RNA-seq and spatial biology.
Why work with Explicyte as your ADC preclinical CRO?
Connect human target biology with functional testing
Every ADC program is different, but a few principles run through every Explicyte study — from model selection to functional characterization and deeper mechanistic investigations.
Dissect ADC response
Target-stratified models and comparator arms help distinguish the contributions of target expression, ADC uptake and intrinsic payload sensitivity.
Model tumor heterogeneity directly
Mixed tumor cell systems and patient-derived models investigate ADC activity in heterogeneous settings where target expression alone may not explain response.
Escalate profiling when it adds value
Scientific continuity end-to-end
A dedicated Study Director oversees study performance and data interpretation, with pathology, molecular and bioinformatics expertise available as needed
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 ADC preclinical program
Tell us your target, ADC format, payload, indications and current development stage. We can help define the models, assays, and complementary profiling needed to characterize target biology, internalization, functional activity, heterogeneity, and translational response.
ADC assays FAQ
Frequently asked questions about ADC preclinical testing
What controls are needed to demonstrate target-dependent ADC activity?
Control selection depends on the ADC and the development question. Where available, studies can combine target-negative or target-low tumor models with the unconjugated antibody, a non-targeting ADC and the free payload. These controls address complementary questions and help distinguish target-dependent ADC delivery from antibody-intrinsic activity, nonspecific conjugate effects and intrinsic payload sensitivity.
How important is target expression for ADC efficacy?
Target expression is an important determinant of ADC activity, but expression alone does not predict response. Cell-surface abundance and heterogeneity, ADC binding and internalization, intracellular processing, intrinsic payload sensitivity, linker–payload properties and bystander activity can all influence efficacy.
Explicyte therefore connects target profiling with functional testing to place target expression in the broader biological context of ADC response.
How do you assess ADC internalization?
ADC uptake can be characterized using flow-cytometry and imaging-based methods in tumor models with defined target-expression profiles. Time- and concentration-dependent internalization can then be interpreted alongside surface expression and functional cytotoxicity to determine whether differences in uptake may contribute to differential ADC response.
How do you distinguish ADC activity from payload sensitivity?
ADC response can be compared with the unconjugated antibody, a non-targeting ADC and the free payload when these controls are available and appropriate.
The free payload provides information on intrinsic cellular sensitivity to the payload mechanism, while target-negative models and non-targeting conjugates help determine how strongly cytotoxicity depends on specific ADC delivery.
How is ADC bystander activity evaluated?
Bystander activity can be assessed using mixed tumor-cell cultures containing distinguishable target-positive and target-low or target-negative populations.
Population-specific killing is monitored after ADC treatment to determine whether cytotoxic activity extends beyond cells directly expressing the target.
Can ADC activity be tested in patient-derived models?
Yes. ADC response can be evaluated in patient-derived organoids and intact tumor fragments.
These models capture patient-specific tumor characteristics and heterogeneous target expression, while intact tumor fragments additionally preserve native tissue architecture and spatial organization.
Can Fc-mediated activity be evaluated?
Yes, when relevant to the ADC. Tumor cells can be combined with human PBMCs or selected immune-effector populations to determine whether Fc-mediated mechanisms contribute additional antitumor activity alongside payload-mediated cytotoxicity.
Can molecular biomarkers be integrated with functional ADC studies?
Yes. Responsive and less-responsive models can be compared using flow cytometry, transcriptomics, spatial biology or other molecular profiling approaches.
