Antibody-drug conjugates, or ADCs, are a class of therapeutic modalities where a target-specific antibody is conjugated to cytotoxic molecules via a linker moiety. The concept of a ‘magic bullet’ discriminately targeting diseased, unhealthy cells was envisioned in the early 20th century by 1908 Nobel Prize laureate Paul Ehrlich. Nevertheless, it took almost a century for the first ADC, gemtuzumab ozogamicin, to be approved by the FDA in 2000.
The first ADCs entered clinical trials in the 1980s, and the subsequent evolution of multiple ADC generations witnessed the transition to fully humanized antibodies, site-specific conjugation technologies, a broader array of cleavable and non-cleavable linker technologies, and a more diverse cytotoxic payload toolbox (1, 2). With two recent approvals of telisotuzumab vedotin and datopotamab deruxtecan in 2025, currently there are 15 ADCs approved by the FDA with more than 300 modalities in clinical trials (3; Figure 1).
The IgG1 subclass of antibody is typically selected to target surface-exposed antigens possessing endocytic and lysosomal-trafficking properties (2).
The FDA-approved cytotoxic payloads can be divided into three major categories: tubulin inhibitors, DNA-damaging agents, and topoisomerase I inhibitors (4). Tubulin inhibitors can be further subdivided into auristatins (e.g. MMAE and MMAF) and maytansinoids (e.g.DM1, DM4). DNA-damaging agents can be subclassed into calicheamicins, pyrrolobenzodiazepine (PBD) dimers, and duocormycins. Exemplary topoisomerase I inhibitors are campothecins (e.g. SN-38, exatecan derivative/deruxtecan/Dxd).
Linkers can be sub-grouped into non-cleavable and cleavable, with the latter currently dominating the ADC landscape (5). The cleaving mechanism is typically triggered chemically (e.g. pH, glutathione) or enzymatically (e.g. by tumor proteases).
The conjugation of linker-payload moiety to an antibody can be either non-specific or site-specific. Earlier ADCs utilized stochastic lysine and cysteine-based conjugation methods with constructs displaying high degrees of heterogeneity. Recent conjugation trends are predominantly based on site-specific strategies such as various enzymatic methods, incorporation of non-canonical amino acids, engineered cysteines, selective reduction of disulfide bonds, Fc affinity reagents, and glycoengineering (6).
The choice of conjugation strategy impacts the position of linker-payload attachment as well as the drug-to-antibody ratio (DAR). The position of conjugation can impact the biological functions of the antibody (e.g. Fc receptor binding) whereas DAR affects potency, tendency to aggregate, immunogenicity, and pharmacokinetics of the conjugate.
The clinical success of conventional ADCs as well as the remaining unsolved critical challenges, namely adverse side effects, drug resistance, and tumor heterogeneity, fueled the expansion of the field from ADCs to a broader range of conjugates (4). Collectively, they are known as XDCs (X-any targeting moiety, D-broader range of drugs, C-conjugate), with innovations happening for each component of the XDC (Figure 2).
To improve the targeting component of the ADC, non IgG-1 scaffolds are being tested as targeting moieties for payload delivery. Reduced size of peptides, as discussed previously in the BioPharmaSpec blog on peptide conjugates, scFv’s, nanobodies, aptamers and other antibody mimetics hold potential in reducing immunogenicity and off-target interactions whilst enhancing tumor penetration of the conjugate (4). Bispecific ADCs are being developed to address tumor heterogeneity and drug resistance (7). Non-internalizing ADCs targeting stromal, vascular, and non-cellular antigens are emerging as an alternative approach to address tumor resistance (8). Conditionally active ADCs (probody-drug conjugates/ PDCs) are designed to bind antigen selectively in the tumor microenvironment and hold promise to minimize on-target, off-tumor toxicities (4).
Extending beyond the traditional cytotoxic payloads is another area of extensive research. Immune-stimulating antibody conjugates (ISACs) with TLR-7/8 and STING agonists are currently being tested in clinical trials (9). Proteolysis targeting chimera (PROTAC) degrader- antibody conjugates (DACs) and molecular glue-antibody conjugates (MACs) are designed to degrade specific protein targets and are actively explored at pre-clinical settings (10, 11). Antibody-oligonucleotide conjugates (AOCs) bearing gene-modulating siRNA and ASO payloads open additional avenues for cancer treatment (12). Finally, ADCs with two different payloads hold promise to tackle tumor resistance, with the first dual-payload ADC having entered clinical trials recently (13).
Linker plays a critical and delicate balance between payload stability and safety as well as efficient payload release and efficacy of the ADC. 13 out of 15 FDA-approved ADCs and more than 50% of those developed clinically employ cleavable linkers (3), with a lot of research dedicated towards novel ways to efficiently and selectively release the payload from the ADC/XDC in the tumor microenvironment. A notable example is the next-generation linker technology developed by TagWorks Therapeutics, where trans-cyclooctane (TCO) linker-bound payload is released from an ADC by administration of tetrazine reagent (14).
Recent clinical success of ADCs, as well as decades-long accumulated clinical evidence of their shortcomings, namely toxicity, drug resistance, and tumor heterogeneity, spurred the development of a broader range of conjugates called XDCs. Their molecular architecture entails a unique combination of targeting moiety, linker, and payload, thus requiring careful consideration of structural characterization studies, which can be performed across BioPharmaSpec’s global laboratories.
As ADCs evolve into increasingly complex XDC modalities, robust analytical characterization becomes essential for clinical and regulatory success. BioPharmaSpec partners with developers to deliver the structural insight and confidence needed across ADC and XDC development.
