Off-Target Toxicity of Antibody-Drug Conjugates (ADCs): How to Break Through the Efficacy-Safety Balance Challenge?

I. What are ADC Drugs and What Are Their Core Challenges?

Antibody-Drug Conjugates (ADCs) are a class of targeted therapeutics formed by covalently linking specific antibodies to highly potent cytotoxic drugs (payloads) via chemical linkers. The design concept of ADCs aims to utilize the targeting capability of antibodies to deliver cytotoxic drugs precisely to tumor cells, thereby improving the therapeutic index (TI) and reducing toxic side effects on normal tissues.

As of now, 16 ADC drugs have been approved globally, with over 900 ADC candidates in various stages of development, primarily focused on hematological malignancies and solid tumors. However, despite continuous optimization in target selectivity and drug design, dose-limiting toxicity (DLT) remains a major obstacle to their clinical application. Even when targeting antigens with high tumor-specific expression, ADCs often cause off-target toxicity at suboptimal doses, resulting in a narrow therapeutic window and limiting their full efficacy.

II. Is ADC Toxicity Solely Caused by the Payload?

Conventional views attribute ADC toxicity primarily to the cytotoxic payloads they carry. Clinical data show that adverse events such as myelosuppression, hepatotoxicity, and peripheral neuropathy reported for different ADCs are indeed highly correlated with the type of payload used. For example:

·       ADCs using tubulin inhibitors (e.g., MMAE, DM1) often lead to neutropenia and peripheral neuropathy;

·       ADCs using DNA-damaging agents (e.g., Calicheamicin) are more likely to cause thrombocytopenia and hepatotoxicity.

However, toxicity is not solely determined by the payload. ADCs are widely distributed in the body, with only about 0.1% of the administered dose ultimately accumulating in tumor tissue. The majority distributes to normal tissues and is taken up by normal cells through various mechanisms, leading to payload release and target-independent toxicity.

III. How Are ADCs Taken Up by Normal Cells? What Are the Mechanisms?

The pathways for normal cell uptake of ADCs are complex and diverse, primarily including the following three mechanisms:

1.     Premature Release Due to Linker-Payload Instability
ADCs may release payloads prematurely in the bloodstream due to chemical instability or enzymatic cleavage of the linker. These free payloads can enter normal cells via passive diffusion or transporter-mediated uptake, causing systemic toxicity. For example, cleavage of the valine-citrulline (VC) linker by extracellular serine proteases, releasing MMAE, is a significant cause of myelosuppression.

2.     Non-Specific Endocytosis
Various endocytic mechanisms contribute to the non-specific uptake of ADCs, including:

o   Macropinocytosis: Primarily occurs in corneal epithelial cells, megakaryocytes, etc., associated with ocular toxicity and thrombocytopenia.

o   Clathrin- and caveolin-mediated endocytosis: Widely present in endothelial and epithelial cells.

o   Phagocytosis: Active mainly in immune cells such as hepatic Kupffer cells and splenic macrophages.

The physicochemical properties of ADCs (e.g., surface charge, hydrophobicity) also influence their non-specific endocytosis rate. ADCs with positive charges or high hydrophobicity are more likely to interact with cell membranes, increasing the risk of non-targeted uptake.

3.     Receptor-Mediated Endocytosis
Various Fc receptors recognize the Fc region of the antibody portion of ADCs, mediating their internalization:

o   Fcγ receptors: Widely expressed on immune cells, endothelial cells, platelets, etc., involved in ADC clearance and toxicity.

o   FcRn receptor: Regulates the circulatory half-life of ADCs; its high expression in vascular endothelial cells and myeloid cells influences ADC tissue distribution.

o   C-type lectin receptors: Such as the macrophage mannose receptor (MR), highly expressed in liver sinusoidal endothelial cells, potentially associated with hepatotoxicity.

IV. How to Reduce Off-Target Toxicity by Optimizing ADC Design?

1.     Improve Linker Stability
Develop novel non-cleavable linkers or enzyme-specific responsive linkers to reduce premature payload release in circulation. For example, sulfonyl pyrimidine-based linkers can significantly enhance plasma stability.

2.     Modulate Antibody Physicochemical Properties
Use engineering approaches to adjust antibody isoelectric point, surface charge distribution, and hydrophobicity, reducing non-specific interactions with non-target cells.

3.     Optimize Payload Properties
Design membrane-impermeable payloads or prodrug forms of payloads to limit their diffusion capacity after release from target cells, thereby controlling the scope and intensity of the "bystander effect."

4.     Selectively Reduce Fc Receptor Binding
Introduce point mutations in the Fc region to lower its affinity for receptors like FcγR and FcRn, reducing receptor-mediated non-targeted internalization.

V. What Directions Should Future ADC Development Focus On?

Although ADC technology continues to advance, off-target toxicity remains a core challenge for clinical translation. Future research should focus on:

·       Establishing more accurate preclinical models to improve the prediction of human-specific toxicities.

·       Developing tissue-specific delivery systems to further enhance tumor targeting efficiency.

·       Exploring combination therapy strategies to reduce the required ADC dose through synergistic effects.

·       Leveraging artificial intelligence and multi-omics data for the rational design of safer ADC candidates.

VI. Which Manufacturers Provide DM-1/DM-4 Antibodies?

Hangzhou Starter Bio-tech Co., Ltd. independently developed the "Anti-DM-1/DM-4 Monoclonal Antibody" (Product Code: S0E0004), a core detection tool characterized by high specificity, high affinity, and excellent stability. This product can accurately recognize DM-1 and DM-4 class tubulin inhibitors and holds significant application value in the research, quality control, and pharmacological evaluation of Antibody-Drug Conjugates (ADCs).

Core Product Advantages:

·       High Specificity Recognition: Capable of simultaneously targeting DM-1 and DM-4 with no cross-reactivity, supporting various immunoassay platforms.

·       Exceptional Sensitivity and Stability: Performs reliably in applications such as ELISA, Western Blot, and Immunohistochemistry, with strong batch-to-batch consistency.

·       Applicable to Multiple Scenarios: Including quantitative analysis of ADC drug payloads, pharmacokinetic studies, and tissue distribution detection.

·       Professional Technical Support: Provides comprehensive product performance data and experimental protocol support to help customers accelerate R&D progress.

Hangzhou Starter Bio-tech is committed to providing high-quality antibodies and detection solutions for global innovative pharmaceutical companies and research institutions. For more product details or to request sample testing, please feel free to contact us.

Conclusion

The off-target toxicity of ADC drugs is a complex issue involving multiple factors and intertwined mechanisms, encompassing linker stability, payload properties, antibody physicochemical characteristics, and the host receptor expression profile. Only by systematicaling its molecular mechanisms and comprehensively considering various risk factors during the drug design phase can the ideal balance between efficacy and safety for ADC drugs be truly achieved, promoting the development and clinical application of next-generation ADCs.

Product Information