TROP-2 and the Promise and Peril of High-Affinity in ADCs

Over the last decades, Antibody–drug conjugates (ADCs) have emerged as a transformative class of cancer therapeutics, offering the promise of targeted cytotoxicity with the precision of monoclonal antibodies.* To fulfill this promise, the ideal ADC includes an antibody that is specific for binding to the target antigen, has high binding affinity for that target, and, at the same time, has low immunogenicity.  In addition, the antibody should have a long plasma half-life and rapid internalization. [1][2][3]

Targeting TROP-2
Trophoblastic Cell Surface Antigen 2 (TROP-2) stands out as one of the most compelling ADC targets because it is a transmembrane glycoprotein with both extracellular and intracellular domains that is involved in calcium signal transduction. Its extracellular domain is overexpressed in a wide range of epithelial cancers—including breast, lung, and gastrointestinal tumors—and its heightened expression often correlates with poor clinical prognosis. TROP-2 also has relatively low or no baseline expression in most normal or healthy tissues. [4][5][6]

These features make TROP-2 an appealing therapeutic target for next-generation ADCs. To date, the safety and clinical benefit of Trop 2-based ADCs have been demonstrated in several clinical trials across multiple tumor types, including those with limited treatment options, such as triple-negative breast cancer (TNBC), platinum-resistant urothelial cancer, and heavily pretreated non-small cell lung cancer. [5]

The clinical success of TROP-2-targeting ADCs, including sacituzumab govitecan (Trodelvy®; Gilead Sciences) and datopotamab deruxtecan (Datroway®; Daiichi Sankyo and AstraZeneca), has catalyzed a wave of innovation in the field. [7]

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Sacituzumab govitecan, a TROP-2-directed antibody and topoisomerase I inhibitor payload (SN-38) drug conjugated via a hydrolyzable linker, is designed to treat metastatic triple-negative breast cancer (mTNBC) and HR+/HER2- breast cancer. Datopotamab deruxtecan*** uses a tetrapeptide-based cleavable linker to covalently attach the humanized anti-TROP-2 antibody to the topoisomerase I inhibitor payload (DXd) and is primarily developed for the treatment of advanced hormone receptor-positive/HER2-negative breast cancer and EGFR-mutated non-small cell lung cancer (NSCLC). [7]

A remaining challenge
Despite successes, a critical challenge remains toxicity. TROP-2 is not exclusive to cancer cells; it is also present, albeit at lower levels, in healthy epithelial tissues such as the skin and gastrointestinal tract. This shared expression between tumor and normal tissues creates a classic ‘on-target, off-tumor’ toxicity dilemma. The result? ADCs often reach dose-limiting toxicity thresholds before achieving their maximum tumor-killing potential, constraining their clinical utility.

For example, sacituzumab govitecan is associated with significant, manageable toxicities, primarily hematologic and gastrointestinal. Common adverse events (AEs) include high-grade neutropenia, diarrhea, fatigue/asthenia, nausea/vomiting, and alopecia. Severe (Grade 3/4) neutropenia, which can be life-threatening, is the most common serious adverse event.[8]

And while generally considered to have a more manageable safety profile than other ADCs, common AEs of datopotamab deruxtecan include stomatitis/mucositis (often >50%; Grade 3-4), nausea, fatigue, alopecia, dry eye/keratitis, conjunctivitis, and increased lacrimation. The most serious, potentially fatal adverse event is interstitial lung disease (ILD) or pneumonitis, occurring in ~7% of cases (5% Grade), requiring close monitoring, prophylactic steroid mouth rinses and good oral hygiene, and dose interruptions.[9]

Solving a problem
A groundbreaking study by Aiko Yamaguchi, Junping Hong, Leike Li, et al, published in the March 13, 2026, edition of Antibody Therapeutics [10], now proposes a radical yet elegant solution to solve this problem: affinity optimization. In a counterintuitive twist, the authors of this study demonstrate that reducing antibody affinity—rather than maximizing it—can significantly expand the therapeutic window of TROP-2-targeted ADCs. In their study, they explored the science, significance, and future implications of this novel, affinity-optimized approach. [10]

Traditional Approach: High Affinity at Any Cost
ADCs exploit the high affinity and exquisite selectivity of an antibody for a specific epitope on the target antigen to deliver a payload to the tumor site. [11][12] Their efficacy is deeply rooted in the high-affinity binding between the antibody’s paratope (binding site) and the target antigen’s epitope. High affinity ensures the ADC binds preferentially to cells expressing the target antigen, typically overexpressed on tumor cells, while sparing healthy tissue. Once bound, the antibody-antigen complex is often internalized through endocytosis. High binding affinity (characterized by a low dissociation constant, K_d)^# ensures the complex remains stable long enough to be pulled into the cell. Proper binding affinity (generally a K_d in the low nanomolar range  (10^-9 to 10^-10 M)), along with the antibody’s Fc region properties, is essential for rapid internalization and for maintaining the ADC in systemic circulation, thereby maximizing the chance of reaching the tumor.

Hence, in the world of antibody engineering, the dogma has long been ‘the tighter, the better.’ High-affinity antibodies are prized for their presumed ability to bind targets with maximal specificity and strength, theoretically improving tumor selectivity and payload delivery. This pursuit of picomolar affinities has been a key driver behind the remarkable efficacy of many modern biologics.

There is, nevertheless, an upper limit to the benefit of high affinity. If the affinity is too high, the ADC may bind so tightly to the first cells it encounters (the ‘perivascular’ cells near blood vessels) that it fails to penetrate deeper into the tumor mass—a phenomenon known as the Binding Site Barrier (BSB)****.  To improve drug distribution and enhance ADC penetration throughout solid tumors, strategies to intentionally moderate affinity include adjusting antibody dose, reducing affinity, or using smaller antibodies (e.g., single-domain antibodies or scFv).[13][14]

However, when the target antigen is also expressed in normal tissues, the strategy designed to exploit high affinity can backfire.^## The obvious reason is that high-affinity antibodies are indiscriminate—they bind to the intended target (TROP-2) wherever it is found, regardless of whether it is on a tumor cell or a healthy epithelial cell.  The result is collateral damage: skin rashes, gastrointestinal toxicity, and other adverse effects that curtail dosing and limit patient benefit.[15]

The Achilles’ heel of high-affinity ADCs, therefore, is their inability to distinguish between ‘good’ and ‘bad’ TROP-2. The field has recognized this challenge, but until recently, a practical solution has remained elusive.

The Counterintuitive, Goldilocks Binding Strategy
The newly published research upends conventional wisdom by demonstrating the power of affinity tuning—deliberately engineering antibodies with reduced (but not weak) affinity for TROP-2.

This strategy is guided by a simple yet profound insight: tumor cells typically express TROP-2 at much higher densities than normal cells. By designing antibodies with ‘just right’ or ‘Goldilocks’ affinity, it is possible to exploit this difference.

How does this work in practice?

• Low-Density Tissues (Healthy Cells): The antibody’s lower affinity means that, in tissues where TROP-2 is expressed at low levels, binding is weak or negligible. The antibody essentially ‘ignores’ healthy cells, reducing off-tumor toxicity.
• High-Density Tissues (Tumor Cells): In tumors, where TROP-2 is overexpressed, the high local concentration of antigens enables multivalent (avidity-driven) binding. The cumulative effect of multiple weak interactions results in strong, stable attachment to the tumor cell surface, ensuring effective delivery of the cytotoxic payload.

This approach leverages a fundamental principle of molecular biology: the context-dependent nature of binding interactions. Rather than seeking the highest possible affinity, the goal is to achieve a level that is ‘good enough’ to capture tumor cells but ‘not good enough’ to stick to healthy tissues.

Discovery and Engineering
To realize this strategy, the research team employed a sophisticated antibody discovery approach using an in-house phage display library. They identified and characterized a fully human monoclonal antibody that binds a unique conformational epitope on TROP-2. Notably, this antibody exhibited reduced binding affinity compared to traditional anti-TROP-2 antibodies.

The importance of targeting a unique conformational epitope cannot be overstated. By focusing on a three-dimensional structure specific to TROP-2 in its tumor-associated conformation, the antibody achieves heightened selectivity. This spatial targeting ensures that the antibody is more likely to bind the intended target in the tumor microenvironment, further mitigating the risk of off-tumor effects.

The researchers then engineered this antibody into homogeneous ADCs, conjugating it with highly potent cytotoxic payloads: auristatin and duocarmycin derivatives. For benchmarking, they also developed a surrogate ADC modeled after sacituzumab govitecan, the current standard-bearer in TROP-2-targeted therapy.

Potent Efficacy with Lower Toxicity
The preclinical data are striking. In their study, Aiko Yamaguchi, Junping Hong, Leike Li, et al, confirmed that in both mouse xenograft and syngeneic tumor models, the affinity-optimized ADCs demonstrated robust antitumor activity—comparable to, and in some cases exceeding, that of traditional high-affinity ADCs. Most importantly, these effects were achieved without the significant toxicities typically observed with existing TROP-2-targeting agents.

The study results demonstrated:

• Effective Tumor Killing: Affinity-optimized ADCs induced rapid and durable tumor regression in models of TROP-2-expressing cancers.
• Reduced Off-Tumor Toxicity: Unlike their high-affinity counterparts, the novel ADCs showed minimal adverse effects in healthy tissues, as evidenced by improved tolerability in animal studies.
• Efficient Payload Delivery: The antibody’s unique conformational epitope and rapid internalization properties ensured that the cytotoxic drug was delivered effectively into tumor cells, maximizing selectivity and potency.
• Comparison with Existing ADCs: When compared head-to-head with a sacituzumab govitecan surrogate, the novel ADCs maintained or exceeded efficacy while displaying a more favorable safety profile.

These results underscore the potential of affinity optimization to widen the therapeutic window—the range between effective and toxic doses—of TROP-2-targeted ADCs. By dialing in the ‘just right‘ affinity, the therapeutic index can be substantially improved.

Rethinking Antibody Design
This breakthrough challenges a core assumption of antibody engineering: that higher affinity is always better. In the context of ADCs targeting antigens expressed on both tumors and healthy tissues, extreme affinity becomes a liability rather than an asset. The implications extend beyond TROP-2. Many clinically relevant tumor-associated antigens, such as Human Epidermal Growth Factor Receptor 2 (HER2/ErbB2) and Epidermal Growth Factor Receptor (EGFR/HER1), are also expressed at lower levels on normal or healthy tissues. The affinity optimization strategy could therefore serve as a generalizable blueprint for designing safer and more effective ADCs across a wide range of targets.

Moreover, the study published in Antibody Therapeutics highlights the importance of epitope selection. By mapping antibodies to unique conformational epitopes that are preferentially exposed or internalized on tumor cells, it is possible to further enhance selectivity and efficacy.[10] This approach also dovetails with advances in ADC payload chemistry, linker stability, and site-specific conjugation, offering a multi-pronged toolkit for next-generation therapeutics.

Towards an Era of ‘Smart’ Antibodies
The ADC field has been characterized by relentless innovation, but also by persistent challenges related to safety and efficacy. The findings from this study suggest that ‘dumbing down’ antibody affinity—far from a compromise—may represent a sophisticated strategy to improve clinical outcomes.

By intentionally tuning antibody properties, researchers can create ‘smart’ antibodies that respond to the nuanced biology of tumors versus normal tissues. This paradigm shift is likely to influence drug development pipelines, regulatory considerations, and clinical trial designs in the coming years.

It also invites a re-examination of past failures in the ADC space. Some agents that showed preclinical promise but failed due to toxicity may find new life when paired with affinity-optimized antibodies. Conversely, projects that stalled due to insufficient efficacy may benefit from combining optimized binding with more potent or tumor-selective payloads.

Future Directions and Challenges
While the preclinical data are compelling, several important questions remain:

• Translational Relevance: Will the affinity-optimized strategy hold up in human clinical trials, where the complexity of tumor heterogeneity and patient variation comes into play?
• Biomarker Development: How can we best identify patients who will benefit most from affinity-optimized ADCs? Quantitative assays for TROP-2 density and epitope exposure may be required.
• Combination Therapies: Can affinity-optimized ADCs be effectively combined with other modalities, such as immune checkpoint inhibitors or targeted therapies, to further boost efficacy?
• Manufacturing and Scalability: Are there technical or economic hurdles to producing homogeneous, affinity-tuned ADCs at commercial scale?

Addressing these questions will be crucial for translating this promising preclinical science into real-world patient benefit.

A New Era for TROP-2-Targeted Therapies
The advent of affinity-optimized TROP-2 antibodies marks a turning point in the design of ADCs and, more broadly, in the philosophy of antibody engineering. By embracing the nuance of ‘just right’ binding—rather than maximal binding—researchers have opened the door to safer, more effective therapies for patients with solid tumors.

This approach promises to expand the therapeutic window, reduce dose-limiting toxicities (DLT), and unlock new potential for ADCs targeting not just TROP-2 but also a host of other clinically relevant target antigens.

Following a long history of setbacks and a high number of discontinued ADCs in preclinical and early clinical development, the complexity of ADCs has become clearer, leading to more approved drugs. And while the design of ADC required understanding the interactions among several critical design functions, a firm grasp of the structure-function relationship of an investigational ADC may translate into improved clinical success and, ultimately, more approved ADCs. [3]

As the field moves forward, affinity optimization may not just be another aspect of ADC development; it may represent a transformative advancement. As such, it may be poised to become a cornerstone of rational drug design—heralding an era where smarter, safer, and more selective biologics redefine the landscape of ADCs in cancer therapy and beyond. In turn, with ongoing clinical evaluation and continued innovation, affinity-optimized ADCs may fulfill their promise as the next wave of transformative cancer treatments—offering renewed hope to patients, clinicians, and researchers alike.

__

Note:* Antibody-drug Conjugates or ADCs function by precisely delivering a cytotoxic payload directly to tumor cells, which enhances selectivity and reduces systemic toxicity, thereby improving the therapeutic index. When an ADC binds to its target receptor on a cancer cell, the resulting complex is internalized via mechanisms such as clathrin- or caveolae-mediated endocytosis or pinocytosis. This process leads to the formation of endosomes, which subsequently fuse with lysosomes. Within these acidic, proteolytic, or reductive intracellular compartments, the cytotoxic drug is released from the antibody. The released payload then diffuses into the cytoplasm, where it targets its intended targets and induces cell death.[16][17]

**Trophoblastic Cell Surface Antigen 2 (TROP-2) is a surface transmembrane glycoprotein member of the epithelial cell adhesion molecule (EpCAM) family.

*** Datopotamab deruxtecan differs from sacituzumab govitecan in that it has a cleavable tetrapeptide linker and a more potent topoisomerase inhibitor payload. This construct is highly stable in circulation, with a longer half-life than sacituzumab govitecan, and undergoes cleavage by intracellular lysosomal proteases.

**** Tumor penetration of ADCs is distinctly affected by their ability to breach the binding site barrier (BSB) in perivascular regions of tumor vasculature. Limited distribution within solid tumors may be due to the BSB.

^# K_d is defined as the concentration of ligand at which half of a partner’s binding sites are occupied when at equilibrium. A low K_d constant, typically in the picomolar and nanomolar range, describes a tight binding interaction, while a high K_d constant describes a weak interaction (typically in the micromolar range or higher).

^## Lowering the target binding affinity of the antibody-component of an ADC may mitigate the on-target, off-tumor toxicity due to decreased target-mediated drug disposition (TMDD) within normal, or healthy tissues, while maintaining robust pharmacological efficacy through sufficient TMDD to tumor tissue, which often over-expresses the antigen for adequate delivery of the potent cytotoxic agent directly to the tumor. [18]

Highlights of prescription information
Sacituzumab govitecan (Trodelvy®; Gilead Sciences)[Prescribing Information]
Datopotamab deruxtecan (Datroway®; Daiichi Sankyo and AstraZeneca)[Prescribing Information]

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DOI: 10.14229/jadc.2026.31.03.001