Combining cancer cell-seeking antibodies with tumor-killing drugs or radionuclides is an old idea whose current embodiment, antibody-drug conjugates (ADCs), results from several generations of improvements leading from murine or chimeric antibodies unstably linked to poorly characterized payloads, to fully humanized antibodies, site-specific conjugation, and precisely calculated drug-antibody combinations. Yet ADC products still face existential issues like off-target toxicity, tumor heterogeneity, and unpredictable pharmacokinetics (PK).

Managing off-target effects

“ADCs can damage healthy tissues through non-specific binding or premature release of the cytotoxic drug,” says Namrata Jayanth, Ph.D., Research Leader at Charles River. “These ‘bystander effects’ limit ADCs’ therapeutic window.” Improving target specificity and site-specific conjugation, while optimizing linkers to release the drug only at its intended destination, mitigates some of these toxicities.

An example is Sacituzumab Govitecan (Trodelvy), which targets the Trop-2 tumor marker with a SN-38 payload and a pH-sensitive linker. SN-38 is an active metabolite of the potent cytotoxic agent irinotecan, while the pH-sensitive linker ensures drug release within the tumor’s acidic environment.

“Tumor heterogeneity and drug resistance are significant challenges that significantly limit ADCs’ therapeutic effectiveness,” Jayanth says. “Intra- and inter-tumoral heterogeneity, characterized by variability in antigen expression, genetic mutations, and cellular characteristics within a single tumor or across tumors, can reduce ADC uptake and efficacy, causing incomplete tumor eradication. Simultaneously, resistance mechanisms stemming from antigen downregulation, drug efflux pumps expelling cytotoxic agents, and altered intracellular processing, all reduce ADC delivery and therapeutic success.”

“Tumor heterogeneity leading to treatment resistance also leads to different responses to treatment, which gives rise to ADC-resistant tumor types,” adds Petra Dieterich, Ph.D., Scientific Leader at Abzena.

ADC toxicities tend to be serious. For example, Mytotarg, the first U.S.-approved ADC (for leukemia), was withdrawn in 2010 as a result of more deaths among treated vs. untreated patients but was re-approved in 2017 at a lower dose. And 11.4% of patients receiving Enhertu, an ADC targeting Her2-positive solid tumors, develop life-threatening lung inflammation.

“Drug developers are investigating several alternative formats to address these shortcomings,” Dieterich says, “including dual payload ADCs, bispecific ADCs and proteolytically activated antibody prodrugs (probodies).” Probodies mitigate “on-target, off-tumor” side effects from ADCs targeting receptors on both cancer and healthy cells by only releasing the drug inside the tumor.

Cytomx Therapeutics has developed several PDCs currently in preclinical and clinical development. Primate studies report a tenfold increase in therapeutic index, suggesting these drug candidates are effective at much lower doses than conventional ADCs.

Another potential fix for ADC toxicity are bispecific ADCs, which, as their name suggests, carry a cytotoxic payload but target two receptors simultaneously. Bispecific ADCs promise enhanced binding affinity, rapid internalization, and therefore higher potency.

“One thing to watch out for with bispecific ADCs is that target expression ratios on tumor cells can vary between tumors and between patients,” Dieterich tells Biocompare. “This can complicate the selection of targets, and requires the careful selection of epitopes and binding modes, and a greater understanding the drugs’ underlying biology.”

Rounding out other drug-antibody combinatorial possibilities are dual-drug ADCs, which seek to overcome tumor resistance to individual cytotoxic payloads by providing a one-two punch through a single dosing regime.

“Dual response may also be accompanied by additive or synergistic toxicities,” Dieterich explains, “so payloads need to be carefully selected based on their mechanism of action to achieve optimal therapeutic outcomes.” These efforts are still in the early stages of exploration.

Making connections

The success of an ADC depends not only on the effectiveness of its main actors (antibody and drug) but on how well they work together. Linker selection is therefore critical to ensure ADC safety and effectiveness, according to Yuerong Wang, Marketing Manager at Sino Biological.

“Early-generation ADCs used non-cleavable linkers requiring lysosomal degradation within cancer cells to release cytotoxic drugs, but this approach led to uncontrolled payload release and limited efficacy, especially in tumors with heterogeneous antigen expression or antigen down-regulation.”

In response, researchers turned to cleavable linkers that release the drug under specific conditions such as low pH or biochemical triggers inside the tumor. Targeted release minimizes off-target toxicity while enhancing effectiveness.

Some cleavable linkers can trigger “bystander killing,” where the drug diffuses to and kills neighboring tumor cells expressing reduced or even no antigen. “This feature is critical for overcoming resistance within tumors but must be balanced to avoid off-target effects,” Yuerong says.

Today, linker research focuses on circulation stability to prevent premature drug release and biocompatibility to minimize adverse immune reactions. “Smart” linkers responding to multiple stimuli within tumors are particularly noteworthy, Yuerong adds, because they improve the precision of payload delivery.

PK considerations

Due to their structures and biological interactions, ADCs exhibit unusual pharmacokinetics. Once administered, ADCs circulate in the bloodstream either as intact ADC, naked antibody, or free cytotoxic payload, and all three forms undergo dynamic changes through target antigen recognition, internalization, and dissociation within cells.

ADC PKs are further complicated by cytotoxic payload metabolism in the liver, which is influenced by drug interactions and liver or kidney impairment.

“These factors, plus patient variability, challenge the development of robust PK models that accurately describe ADC behavior in patients and assist in the design of new ADCs,” Yuerong says. Useful PK models are emerging, however, through the integration of standard analytics (e.g., liquid chromatography-mass spectrometry, enzyme-linked immunosorbent assays, and capillary electrophoresis).

Often less than 1% of the administered ADC dose reaches target cells, which Jayanth attributes to slow tissue penetration and its consequence—limited drug delivery to tumor cells. Both issues are related to ADCs’ large size compared with small molecule drugs and conventional antibodies.

“PK issues result in suboptimal drug delivery to tumors, especially those located farther from blood vessels. Utilizing smaller antibody fragments such as single-chain variable fragments (scFv), Fab fragments, or nanobodies reduces the molecules’ size, improves their diffusivity, and may improve tissue penetration,” Jayanth adds.

Another work-around involves increasing the drug-to-antibody ratio to enhance payload delivery. For example, PBD-based ADCs (pyrrolobenzodiazepine) have a higher DAR, providing increased cytotoxicity to target cells compared to traditional ADCs.

“ADCs represent a transformative approach to targeted cancer therapy,” Jayanth says, “offering the potential for more effective, less toxic treatments. As the field continues to evolve, advances in ADC design, targeting strategies, and combination therapies are expected to enhance ADC effectiveness and broaden their clinical application.”