antibody-drug conjugate (ADC)

Antibody-drug conjugate

Antibody-drug conjugates (ADCs) are drug delivery systems composed of three elements: an antibody (targeting), a payload (cytotoxin), and a linker.Combining the target specificity of antibodies with the potent cell-killing ability of low-molecular-weight anticancer drugsand delivers the drug selectively to cancer cells. After binding to the antigen, it is internalized into the cell, undergoes degradation and linker cleavage in the lysosome, and releases the payload to induce apoptosis. Furthermore, a bystander effect by the released drug is also expected, and it is attracting attention as a cutting-edge drug discovery approach aiming for treatment including the cancer microenvironment.

Representative examples of antibody-drug conjugates

Enhertu (trastuzumab deruxtecan: T-DXd)

Enhertu (trastuzumab deruxtecan: T-DXd) is an HER2-targeting ADC.Potent bystander effect driven by a high DAR (approx. 8) and DXd with high membrane permeability, and induces cell death through topoisomerase I inhibitory activity. On the other hand, it is a representative ADC that requires careful attention to respiratory toxicities such as interstitial lung disease (ILD).

Kadcyla (Trastuzumab emtansine: T-DM1)

Kadcyla (trastuzumab emtansine: T-DM1) is a first-generation HER2-targeted ADC.High blood stability due to the non-cleavable MCC linker and DM1, with a limited bystander effectThis is characterized by being degraded along with the antibody within the cell, inhibiting microtubules as an amino acid-conjugated DM1, and inducing cell death.

Adcetris (brentuximab vedotin)

Adcetris (brentuximab vedotin) is an ADC targeting CD30-positive lymphoma.Intracellular payload release by MMAE and Val-Cit linkercharacterized by, and induces cell death by inhibiting microtubules. The DAR is approximately 4, and toxicity evaluations, such as for thrombocytopenia and peripheral neuropathy, are also important.

Key challenges in antibody-drug conjugate development

Linker cleavage (premature release) in the systemic circulation, off-target toxicity, and non-specific cellular uptake

In ADCs, premature cleavage of the linker in the bloodstream before reaching the tumor leads to early release of the potent payload, resulting in systemic toxicities such as myelosuppression, hepatotoxicity, and interstitial lung disease. Also,Non-specific uptake via FcγR and recycling by FcRn complicate toxicity prediction.Points are also an important issue.

The analytical difficulty of DAR (Drug-to-Antibody Ratio) heterogeneity and the "complex PK/PD kinetics by molecular species"

Because ADCs produced by conventional random conjugation methods are a mixture of heterogeneous molecular species with a DAR of 0 to 8, high-DAR species have a rapid hepatic clearance and low-DAR species tend to lack therapeutic efficacy.It is necessary to individually analyze the pharmacokinetics of Total Antibody, Conjugated Antibody, and Free Payload.The point is a crucial issue.

Quantitative evaluation of intra-tumoral antigen expression heterogeneity and the bystander effect

In actual solid tumors, the expression of target antigens is heterogeneous, with a mixture of positive and negative cells. Therefore,Quantitative evaluation of the therapeutic window and effective range of the bystander effectis important. Since excessive payload diffusion leads to toxicity in normal tissues, evaluation in co-culture systems and disease models remains a challenge.

Nonclinical evaluation required for the development of antibody-drug conjugates

Evaluation 1: "Multi-analyte (Total/ADC/Free) LC-MS/MS & DAR distribution tracking PK analysis" to elucidate molecular species-specific behavior

Measuring Total Antibody alone cannot determine the effective concentration of the ADC or the leakage of the free payload. Therefore,Simultaneously quantify Total Antibody, Conjugated Antibody, and Free Payload, and track the DAR distribution and linker cleavage rate.It is important to evaluate the in vivo behavior of ADCs in detail by combining LC-MS/MS, HRMS, immunoprecipitation, and ELISA.

Rating 2: "Antigen-positive/negative cell 3D co-culture organoid & patient-derived tumor tissue (PDX) model drug efficacy evaluation" proving the bystander effect and tumor permeability

A single antigen-overexpressing cell line cannot sufficiently reproduce the heterogeneity within actual tumors. Therefore,Evaluating ADC penetration deep into tumors and bystander effects using 3D co-cultures combining antigen-positive/negative cells and PDX modelsIt is important to analyze the correlation between the intra-tissue distribution of the payload and its efficacy by using techniques such as MSI in combination.

Rating 3: "Fc/Fab-mediated uptake assay & human iPSC-derived tissue (cardiac, liver, lung, etc.) toxicity screen" for predicting human-specific off-target toxicity

In ADCs, in addition to the toxicity of the payload itself, toxicity caused by non-specific uptake via FcγR and antigen expression in normal tissues poses problems.Combining Fc/Fab-mediated uptake assays and human iPS-derived tissue models to predict human-specific toxicity risks at an early stageis important.

Rating 4: "Live-Cell Imaging & Microenvironment (pH/Enzyme) Responsive Linker Cleavage Assay" for Visualizing ADC Internalization and Lysosomal Trafficking Dynamics

For an ADC, it is important not only to bind to the antigen, but also to be efficiently internalized and reach the lysosome.Visualize internalization and lysosomal translocation using live-cell imaging, and quantitatively evaluate the cleavage rate of the linker by pH or enzymes.This allows us to analyze the payload release efficiency within cells. We select the optimal combination of antibody and linker.

Key points when choosing a contract research organization (CRO) for non-clinical studies

In ADC development, it is important to conduct a multifaceted evaluation of molecule-specific PK/PD, the bystander effect within tumors, off-target toxicity, and intracellular kinetics. Strategic CRO selection capable of accurately assessing the complex challenges unique to ADCs is the key to development success. Moving forward, safer and more effective ADC drug discovery is anticipated through high-precision non-clinical evaluation.