Antibody-Drug Conjugates (ADCs): a new therapeutic class​

Antibody–Drug conjugates (ADCs) are a unique class of targeted immuno-oncology therapeutics that combines the tumor-targeting selectivity, stability and favorable pharmacokinetic profile of monoclonal antibodies with the potent antitumor activity of cytotoxic drugs. The ultimate goal of ADCs is to increase the therapeutic index of potent chemical agents, by selective delivery of the chemotherapy to the tumor instead of the healthy tissues.

This new class of medicines ultimately offers new treatment modalities for clinicians in oncology and hematology, and can be effective in patients who no longer respond to other treatments. As of early 2020, there are five approved ADCs on the market, with more than 140 different ADCs undergoing clinical trials for both hematological malignancies and solid tumors. The vast majority of these clinical ADCs are based on random payload attachment (Generation 1 ADCs, see below) and a clear trend towards site-specifically homogeneously conjugated ADCs is observed since the past few years (Generation 2 ADCs, see below).

ADCs: mechanism of action

The general mechanism of action of ADCs is well described by Tsuchikama et al. : once ADCs are injected into the bloodstream, the antibody component recognizes and binds to extracellular antigens that are expressed in target cancer cells. Upon internalization of the ADC-antigen complex through endocytosis, the complex is processed within lysosomes, which releases the cytotoxic payload in its bioactive form inside the cell. The released payload binds to its target (microtubules, DNA/RNA strands, enzymes…), leading to cell death.

Some payloads attached via a “cleavable linker” are able to diffuse outside the cell by passively crossing the external cell membrane (“bystander” effect) in order to exert cell-killing activities to surrounding tumor cells, which themselves may or may not express the target antigen. These types of ADCs are generally preferred for the treatment of heterogeneous solid tumors.

Mechanism of action of an ADC. The conjugate binds to its target cell-surface antigen receptor (Step 1) to form an ADC-antigen complex, leading to endocytosis of the complex (Step 2). The internalized complex undergoes lysosomal processing (Step 3) and the cytotoxic payload is released inside the cell (Step 4). The released payload binds to its target (Step 5), leading to cell death (Step 6). Image taken from Tsuchikama, K. & An, Z. Protein Cell (2018) 9: 33. https://doi.org/10.1007/s13238-016-0323-0.

Current challenges in the ADC field

To date, ADCs generated with conventional (Generation 1) or site-specific conjugation by re-engineering and/or enzymes (Generation 2) face several challenges:

Excessive hydrophobicity and limited drug-antibody ratio (DAR)

Overall hydrophobicity of ADC is now considered to be a key physicochemical parameter, directly influencing ADC pharmacokinetics, efficacy and tolerability (therapeutic index). Hydrophobic ADCs are more easily recognized and sequestered by the clearance organs and the immune system, thus leading to dose-limiting toxicities such as liver toxicity or neutropenia. Cytotoxic payloads attached to the antibody inherently confer increased hydrophobic character and aggregation potential to the final conjugate.

It has long been considered that a 2-to-4 drug-antibody ratio (DAR2-4) achieves the optimal balance between physicochemical properties, pharmacokinetics, in vivo potency and tolerability. This assertion is not true anymore, with the use of

  • New hydrophilic drug-linker chemistries,
  • New drug-linker design and architecture (linear versus orthogonal; length of molecular spacers; presence and chemical nature of enzyme-cleavable entities or lack thereof),
  • hydrophobic dibenzocyclooctyne (DBCO) or bicyclononyne (BCN) scaffolds used in copper-free click chemistries,
  • less hydrophobic payloads,
  • “hydrophobicity masking” entities (such as our PSARlink technology)

For example, we routinely observe in our labs homogeneous DAR8 ADCs that are less hydrophobic than DAR4 ADCs while bearing the same MMAE warhead. High DAR ADCs allow the use of moderately potent drugs with innovative mechanism of action as ADC payloads (e.g. topoisomerase I inhibitors such as SN-38 or Exatecan). These ADCs can also be more potent towards tumors expressing low antigen levels (therefore reaching patient populations hitherto ineligible for treatment), lower-expressing tumor antigens and resistant tumors.

Heterogeneity, time and cost-consuming bioconjugation procedures

Heterogeneous ADCs (Generation 1) resulting from simple stochastic coupling procedures contain fractions that have sub-optimal drug-antibody ratios (DAR) and that are known to possess unfavorable pharmacological properties. The broad distribution of DARs negatively impacts ADC efficacy and therapeutic window, which ultimately leads to high attrition rates in clinical trials. In light of this findings, a great emphasis on site-specific bioconjugation technologies is observed in the ADC field and began to translate into the clinic (Generation 2 ADCs). These techniques require antibody re-engineering and/or the use of coupling enzymes and are time-consuming, expensive and difficult to transpose to large-scale production.

Mablink’s technological approach yields clearly defined homogeneous ADCs having a drug-antibody ratio of 8 (DAR8) or 16 (DAR16). The procedure is directly applicable to “off-the-shelf” native monoclonal antibodies (no re-engineering required) and rely on well-known easy-to-implement stable maleimide chemistry procedures. The time for ADC development is drastically reduced (it takes approximately a few days for payload grafting) and the approach is thus amenable to high-throughput screening.

Premature payload deconjugation (plasma instability)

Conventional bioconjugation chemistries such as classic maleimides (including SMCC) or disulfide-based linkers suffers from premature loss of payload cargo in the bloodstream. This directly translates to loss of efficacy and off-site toxicity. At Mablink, we use new bioconjugation chemistries in our drug-linker designs to ensure absolute plasma stability and avoid premature deconjugations.

Mablink, pioneering 3rd generation ADC

1st generation ADC

2nd generation ADC

Logo Mablink

3rd generation ADC

 

Description

 

Stochastic random conjugation of the payload to lysine or cysteine residues of the mAb.

Heterogeneous DAR leading to suboptimal therapeutic index and limited outcomes in the clinic.

Bioconjugation procedure lacks reproducibility.

High DAR is possible but leads to detrimental hydrophobicity of the ADC.

The five ADCs approved as of today are 1st generation ADCs.

Requires re-engineering of the antibody to add specific conjugations sites and/or the use of one coupling enzymes for bioconjugation.

Homogeneous ADCs with DAR2 or DAR4.

Often paired with plasma-stable chemistries between the antibody and the linker-drug.

Long and expensive process, which can ultimately alter the native antibody properties.

Limited to low DAR values because of the hydrophobicity of the payload and the required inclusion of conjugations sites on the protein sequence.

Straightforward and fast chemical coupling to the native unmodified monoclonal antibody.

Homogeneous DAR.

Hydrophobicity masking moieties embedded into the drug-linker structure, allowing high DAR values while preserving or improving the overall hydrophilicity of the ADC.

Global improvement of physicochemical and pharmacological properties of the conjugate, and therefore of the therapeutic index.

 

Features

 

Average DAR 3-4

DAR 2 or DAR 4

DAR 8 or more

Antibody re-engineering not required

Antibody re-engineering required

Antibody re-engineering not required

Few days of development

Several months of development

Few days of development

Heterogenous coupling

Homogenous coupling

Homogenous coupling

Moderate plasma stability

Improved plasma stability

High plasma stability

Ease of bioconjugation but lack of reproducibility

Tedious bioconjugation procedures but good reproducibility

Ease of bioconjugation and excellent reproducibility, even at an industrial scale

Narrow therapeutic index

Improved therapeutic index

Further improved therapeutic index