Design Principles and Mechanisms of ADCs
Figure 1. Structure of ADC[1].
An ADC consists of three key components: a targeting
antibody that serves as the “navigation system”, a highly potent
cytotoxic small molecule, and a chemical
linker that connects the two. Additional critical parameters
include the drug-to-antibody ratio (DAR) and the conjugation method, which influence the
ADC's efficacy and safety profile.
Mechanism of action of ADCs: The action begins with the antibody guiding the ADC
to bind to the target antigen on the cell surface. This ADC-antigen complex is then
internalized through antigen-mediated endocytosis. Once inside the tumor cell, the
linker is cleaved either by acidic conditions or enzymatic activity, releasing the
cytotoxic drug. This drug subsequently induces cell death by targeting
microtubules or DNA. If the cytotoxic agent is
membrane-permeable, it can also diffuse to neighboring cells, facilitating a bystander
killing effect.
Figure 2. Mechanism of action of ADCs[2].
As powerful tumor-targeted therapeutics, ADCs require careful optimization of each
structural component—antibody, cytotoxic payload, and linker—to achieve maximal
therapeutic efficacy. Using high-affinity antibodies, stable linkers, and potent cytotoxins is an essential prerequisite for creating a safe and effective ADC.
Selection of Target Antigen
The selection of the target antigen plays a pivotal role in determining the success of
ADCs. An ideal antigen should possess the following characteristics: (i) high
specificity and robust expression in tumor tissues; (ii) efficient internalization with
the ability to recycle back to the cell membrane; (iii) homogeneous antigen expression
within the
tumor microenvironment, with minimal antigen
presence in circulation.
Figure 3. Target antigens in ADCs[3].
Selection of Antibody
Antibody selection is closely linked to the target antigen and involves key attributes
such as antigen affinity, target specificity, robust retention, minimal immunogenicity,
low cross-reactivity, and favorable
pharmacokinetics in
plasma. Typically, the binding affinity of the antibodies in ADCs ranges from 0.1 to 1
nM. Early ADCs relied on murine-derived antibodies, which induced human anti-mouse
antibody responses in patients, leading to severe immunogenicity. Currently, most ADCs
utilize humanized or fully human antibodies, significantly reducing immunogenicity.
Among antibody subtypes,
IgG1 antibodies are
predominantly used due to their ability to induce
antibody-dependent cellular cytotoxicity (ADCC) and
complement-dependent cytotoxicity (CDC), promoting antitumor
immune responses, whereas antibodies of the IgG2 and IgG4 subtypes lack these functions.
Figure 4. Characteristics of IgG1–IgG4 subtype antibodies[4].
Table 1. Recommended ADC antibody (biosimilar) products
Note: MCE can provide products for research use only. We do not sell to patients.
Selection of ADC Linker
Once internalized by the tumor cells, the linker undergoes cleavage triggered by
intracellular factors, rapidly releasing the cytotoxic drug. The linker is a pivotal
component of an ADC, as its properties directly influence both therapeutic efficacy and
pharmacokinetics. A stable linker ensures high systemic antibody concentrations and
prevents premature release of the cytotoxic payload during circulation, minimizing
off-target toxicity. Upon reaching the tumor cell and being internalized, the linker
must respond to specific intracellular triggers, cleaving to release the cytotoxic drug
and effectively killing tumor cells.
ADCs employ two main types of linkers: cleavable and non-cleavable.
Cleavable
linkers include hydrazones, disulfides, and peptide linkers, each sensitive to
distinct tumor-associated intracellular conditions, such as low pH, elevated
glutathione, or protease activity.
Non-cleavable linkers, such as thioethers or
maleimidocaproyl (MC) groups, rely on complete
lysosomal
proteolytic degradation of the antibody after cellular uptake to release the cytotoxic
payload.
Figure 5. Classification and release mechanism of ADC linkers[5].
Table 2. Recommended ADC linkers products
Note: MCE can provide products for research use only. We do not sell to patients.
Selection of ADC Payload
An ideal ADC payload should exhibit sub-nanomolar
IC50 values against tumor cell lines in vitro.
Additionally, it must possess a functional group that ensures sufficient aqueous
solubility, enabling efficient chemical conjugation to the antibody while simultaneously
improving the overall solubility of the resulting ADC. The cytotoxic agents currently
used—predominantly derived from natural products—can be classified into three main
categories:
(i) microtubule inhibitors, such as
DM1,
DM4,
MMAF, and
MMAE;
(ii) DNA
damaging agents, such as
calicheamicin,
doxorubicin, duocarmycin, and pyrrolobenzodiazepines
(PBD);
(iii) traditional cytotoxins, such as
paclitaxel, mitomycin C,
methotrexate, and
α-amanitin,
which exert similar anti-tumor mechanisms.
Figure 6. Classification of ADC cytotoxins[6].
Table 3. Recommended ADC payload (cytotoxin) products
Note: MCE can provide products for research use only. We do not sell to patients.
Selection of Conjugation Strategy
The choice of conjugation strategy is a critical determinant of the drug-to-antibody
ratio (DAR), the distribution of conjugation sites, and the overall stability of an ADC.
Broadly, conjugation methods can be classified into two major categories:
non-site-specific and site-specific approaches.
Non-site-specific conjugation includes
lysine-based and
cysteine-based
conjugation.
Lysine-based conjugation utilizes the abundant surface-exposed
ε-amino groups of lysine residues (~40 per IgG) to form stable amide bonds with the
linker. This approach results in highly heterogeneous products with a wide DAR
distribution.
Cysteine-based conjugation involves reducing interchain disulfide
bonds to expose free thiol groups, which then react with the linker to form stable
thioether bonds. By controlling the extent of reduction, defined DAR values (commonly 2,
4, 6, or 8) can be achieved with reduced heterogeneity compared to lysine conjugation.
There are many ways to perform
site-specific conjugation: (i)
Engineered
cysteines: Site-directed mutagenesis introduces cysteine residues at
predetermined locations (e.g., HC-A114C), allowing controlled conjugation with defined
DARs (typically 2 or 4). (ii)
Non-canonical amino acids (ncAAs): An orthogonal
tRNA/aaRS pair incorporates ncAAs bearing bio-orthogonal functional groups (e.g.,
ketones, azides), allowing specific conjugation with precise DAR control, typically at
2. (iii)
Enzymatic conjugation: Enzymes such as
Sortase A or
microbial
transglutaminase (MTGase) recognize specific peptide tags to catalyze
site-specific conjugation. (iv)
Glycan-mediated conjugation: The Fc glycan is
oxidized (e.g., fucose oxidation to an aldehyde) and subsequently functionalized,
enabling site-directed attachment through the glycan chain.
Figure 7. Conjugation strategies in ADC development[3].
Structural and Technological Innovations of ADCs
To enhance the safety and therapeutic efficacy of ADCs, ongoing innovations in
structural design are being explored. These include the development of multivalent and
multispecific antibodies, site-specific conjugation strategies, novel linker
technologies, and new cytotoxic payloads. Together, these advancements aim to improve
tumor targeting, enhance payload delivery, and minimize off-target toxicity.
Figure 8. Structural innovation of ADCs[7].
Innovations in Targeting Vectors
One of the key frontiers in ADC innovation lies in the evolution of targeting vectors.
The replacement of traditional monoclonal antibodies with bispecific antibodies enhances
internalization efficiency and tumor selectivity. Additionally, low-molecular-weight
targeting moieties, such as
peptides, antibody fragments,
and small molecules, offer superior tissue penetration—particularly valuable in the
treatment of solid tumors. Among these,
peptide-drug
conjugates (PDCs) combine the excellent tissue permeability and low
immunogenicity of peptides to achieve ideal drug delivery while minimizing systemic
toxicity. Several peptide-radionuclide conjugates have been approved by the FDA and are
playing a growing role in both cancer diagnosis and therapy. Meanwhile, small molecule
targeting ligands are also being actively explored. These molecules either block
oncogenic pathways or act as homing ligands that bind to specific tumor-associated
receptors. Notable examples of target receptors include folate receptor (FR),
prostate-specific membrane antigen (PSMA), and somatostatin receptor (SSTR).
Innovations in ADC Payloads
In terms of payload design, most approved ADCs to date utilize microtubule targeting
agents, followed by DNA-damaging agents and
topoisomerase
I (TOP I) inhibitors. However, a growing trend is the shift toward non-traditional,
non-cytotoxic payloads. These include TOP II inhibitors, RNA polymerase inhibitors,
Bcl-xL inhibitors, and immune-stimulating agents.
Beyond single-agent strategies, dual-payload ADCs—featuring two cytotoxic or synergistic
payloads with complementary mechanisms—are gaining attention. These dual-payload ADCs
allow for improved tumor cell killing and help overcome challenges such as
chemotherapy resistance and antigen expression heterogeneity.
Furthermore, novel biomolecule-based payloads, such as
antisense oligonucleotides (ASOs) and
small
interfering RNAs (siRNAs), are being explored to extend ADC applications to
genetic diseases, including Duchenne muscular dystrophy (DMD). Another cutting-edge
approach involves using
proteolysis-targeting chimeras
(PROTACs) as payloads, which enable targeted protein degradation through
antibody guidance. Altogether, these novel payload strategies address current
limitations and promise to open new frontiers in cancer treatment.
Figure 9. Novel classes of ADC payloads[6].
Innovations in Conjugation Technologies
The rapid advancement and adoption of site-specific conjugation strategies have largely
addressed the limitations of stochastic conjugation, which typically produces
heterogeneous mixtures with broad DAR distributions (e.g., 2/4/6/8) and high product
variability. In contrast, site-specific approaches produce highly uniform conjugates
with a defined DAR and a single attachment site, resulting in a consistent chemical
entity that simplifies analytical characterization and quality control.
Heterogeneous ADCs often display variable clearance and tissue distribution, making
pharmacokinetics behavior difficult to predict. In comparison, structurally uniform ADCs
provide more stable plasma concentration profiles and reduce the risks of unexpected
elimination or toxic accumulation. Random conjugation may also lead to high-DAR species
that aggregate or cause off-target toxicity. By fixing the DAR (e.g., 2 or 4) and
selecting sites distal to the antigen-binding region, site-specific strategies markedly
lower systemic toxicity while increasing the maximum tolerated dose and broadening the
therapeutic window. Moreover, random conjugation is associated with batch-to-batch
variability and limited process reproducibility. Site-specific technologies—whether
genetic (e.g., engineered cysteines, ncAAs) or enzymatic/chemical—enable consistent DARs
and potency across production lots, streamlining manufacturing and regulatory
submissions.
Clinical Progress and Future Prospects of ADCs
As of July 2025, over 15 ADCs have received regulatory approval and entered global
commercialization. Meanwhile, the clinical development pipeline is rapidly expanding,
with approximately 189 ADCs currently in clinical trials. The majority of these are in
Phase I and II, with a smaller subset entering Phase III. More than 80% of these
clinical trials focus on evaluating safety and efficacy in solid tumors, while
approximately 20% target hematologic malignancies.
Figure 10. Clinical development stage and tumor types targeted by ADCs under
investigation[7].
Table 4. Selected approved ADC products
Note: MCE can provide products for research use only. We do not sell to patients.
The therapeutic landscape of ADCs is continuously broadening, with targets and
indications expanding beyond traditional ones such as
HER2,
EGFR,
c-MET. Newer targets—such as CDH17, DLL3, CLDN18.2, B7-H3, and
CEACAM5—are being actively explored. In parallel, innovations across antibody design,
payload development, and conjugation technologies are converging to enhance the
therapeutic window, fueling the evolution of next-generation ADCs.
Figure 11. Emerging targets of ADCs currently in clinical evaluation[8].
In terms of payload selection, microtubule targeting agents remain the most widely
utilized payload class in ADCs. Notably, novel ADCs with TOP I inhibitors as payloads
represent a rapidly expanding proportion of clinical candidates, far exceeding the
number of ADCs using DNA-damaging agents. In addition, ADCs containing other novel
payloads, such as immunomodulators and RNA polymerase II inhibitors, are actively
undergoing clinical evaluation.
Figure 12. Diversity of clinical-stage ADC payloads[8].
Although ADCs have demonstrated remarkable success in oncology, multiple challenges
remain unresolved—particularly limited penetration into solid tumors, acquired
resistance mechanisms, off-target toxicities, and suboptimal payload release profiles.
To overcome these barriers, ongoing research focuses on innovations such as improved
delivery systems, non-internalizing antigen targets, novel cytotoxic payloads, and
site-specific conjugation strategies.
In addition to structural refinements of ADCs themselves, translational and clinical
strategies are also being explored to maximize therapeutic potential. One promising
direction is fractionated or metronomic dosing, which maintains cumulative anti-tumor
exposure while lowering peak serum concentrations. This approach, reminiscent of
conventional chemotherapy regimens, may help reduce peak-associated toxicities, extend
drug exposure duration, and improve the likelihood of targeting tumor cells as they
cycle into vulnerable states.
Figure 13. Optimization strategies for ADC therapy[8].
Summary
As a groundbreaking class of targeted therapeutics, antibody–drug conjugates (ADCs) hold
immense clinical and commercial promise in oncology. Over the past decade, continuous
innovations in ADC design—ranging from novel targeting vectors such as bispecific
antibodies, peptides, and small molecules, to next-generation payloads including immune
agonists, PROTACs, and oligonucleotides—have significantly expanded the therapeutic
landscape. These advances have been further propelled by the emergence of site-specific
conjugation technologies, enabling greater precision, stability, and efficacy.
ADCs have now evolved into a diverse and modular platform, giving rise to a new
generation of conjugates where, effectively, “everything can be conjugated”—heralding
the era of xenobiotic-drug conjugates (XDCs). While many of these innovations are still
undergoing clinical validation, the field has already produced highly encouraging
outcomes. XDCs stand poised to reshape the landscape of cancer therapy.
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Design Principles and Mechanisms
Structural and Technological Innovations of
ADCs
Clinical Progress and Future Prospects of ADCs
