Dowdy Jackson, Ph.D.
May 10th 2024
Abstract
Antibody drug conjugates (ADCs) have become one of the most sought after modalities to treat cancer patients. Several high-profile deals have been announced recently between smaller biotech companies developing ADCs and larger companies looking to enhance their oncology pipelines. There is significant competition in the ADC space and one of the key questions for companies developing ADCs is how can they differentiate from their competition? Since the majority of ADCs in development are monospecific, targeting only one antigen/target, one way to differentiate from the competition is by developing ADCs that bind to two or more antigens or two epitopes on the same target.
There are least two important questions that need to be addressed when developing bispecific ADCs. The first question is whether or not the bispecific ADCs has superior efficacy and safety compared to the monospecific ADCs individually or when the monospecific ADCs are combined. The second question is whether or not the development of bispecific ADCs will be cost effective given the complexities of developing bispecific ADCs.
Introduction
ADCs can be placed into essentially four broad categories. The first is the traditional monospecific ADC, where the ADC binds with high affinity to only one epitope on a cell surface antigen/target. The second is the biparatopic ADC, where the ADC binds to two different epitopes on the same antigen/target. The third is the bispecific ADCs, where the ADC binds to two distinct cell surface antigens/targets on a tumor. The fourth category would be bispecific antibody fragments where smaller antibody scaffolds are used to improve antibody penetration into solid tumors. Bispecific ADCs appear to be the next wave of ADCs entering clinical development.
Bispecific ADC targets
The number of bispecific ADCs in clinical development is small. There are approximately ten bispecific or biparatopic ADCs in clinical development (Table 1). How do you select which antigen to target with your bispecific ADC? If you were to evaluate the possible number of combinations of ADC targets, starting with the ADCs that have gained FDA approval, (BCMA, Nectin-4, CD19, CD22, CD33, CD79b, CD30, TROP2, Her2, Tissue Factor, and Folate receptor-a), you’d have an 11 x 11 matrix or 56 possible combinations. If you remove targets that have limited tumor expression, such as Folate receptor (Ovarian, Fallopian tube, Peritoneal cancer) and Nectin-4 (Bladder and Renal cancer) or heme targets, this leaves you with a 3 x 3 matrix (HER2, TROP2 and Tissue Factor) or six possible combinations.
To expand the number of possible bispecific targets, you could consider only focusing on ADC targets in phase 3 of clinical development because these targets are clinically validated ADC targets. There are approximately eight ADC targets in phase 3. This provides 36 possible bispecific and biparatopic ADC target combinations (Table 2).
As you begin to include ADC targets in earlier phases of clinical development, the number of possible target combinations increase. There are approximately thirteen ADC targets in phase 2 for solid tumor indications. This would have the potential for over 100 possible combinations. The majority of bispecific ADCs, which includes biparatopic ADCS, are in early clinical development Figure 1.
Selecting targets in early clinical development may increase the risk of failure because the targets lack clinical validation. It is important to note that the majority of oncology drugs, including ADCs, don’t progress past phase 2 [1, 2].
With numerous possible bispecific target combinations, how would you identify which targets to combine? One approach is to identify targets that are co-expressed in the tumor indications of interest as described in the literature.
Zalontamab brengitecan (BL-B01D1) is a bispecific ADC that targets both EGFR and HER3. BL-B01D1 is in phase 3 in China and phase 1 in the US. EGFR was expressed in 52.3% and HER3 was expressed in 82.7% of non-small cell lung cancer (NSCLC) patients’ primary tumors. EGFR was expressed in 62.7% and HER3 was expressed in 91.2% of brain metastasis [3].
Another approach would be to use single-cell transcriptomic profiling and multiomic cross-validation methods. This method was used to identify eleven novel cell surface targets for multiple myeloma [4].
Lastly bioinformatics techniques could be used to identify potential bispecific targets. Co-expression analysis could be used to identify potential bispecific antibody partners [5].
Cell surface antigen binding, internalization and intracellular trafficking
Biparatopic ADCs can induce cell surface clustering, which results in enhanced antibody internalization and delivery to the lysosome. Data from the Her2 biparatopic ADCs and Met biparatopic ADCs have reported enhanced receptor clustering and lysosomal trafficking [6-8].
MEDI4276, for example, is a Her2 biparatopic ADC, which was reported to have enhanced lysosomal trafficking and enhanced in vitro cytotoxicity compared to monospecific Her2 ADCs [6]. The preclinical data package for MEDI4276 showed great promise but the clinical development of MEDI4276 was discontinued in phase 2.
Different ADC targets are internalized at different rates [9]. This could impact the ADC’s response and, in some cases, favor one target over another. Another consideration is the affinity of each arm to the antigen. In some cases, you may need to lower the affinity for one antigen while raising the affinity for another to minimize on-target toxicity in normal tissues. EGFR is an example where an EGFR ADC, RN765C, which has lower affinity for EGFR, had reduced in vitro cytotoxicity against normal keratinocytes compared to EGFR ADCs with higher affinities and maintained potent in vivo anti-tumor efficacy against EGFR expressing tumor xenograft models [10].
Bispecific antibody formats and manufacturing challenges
The major issue with bispecific antibody production is ensuring you have proper formation of the bispecific antibody with high efficiency and yields. There are several bispecific antibody formats currently used for ADCs (Figure 2).
Some bispecific antibody formats require two cell lines to stably produce each arm of the antibody. Each arm of the antibody will be purified and after purification, each arm is assembled in-vitro to produce the bispecific antibody. This approach would require each arm of the antibody to be engineered, using the knob into hole format for example, to ensure proper assembly of the bispecific antibody by favoring the thermodynamics of the formation of the heterodimeric bispecific antibody [11, 12]. Some bispecific antibody formats require substantial antibody engineering to ensure the proper formation of the heterodimeric antibody. IMGN151, which is a biparatopic ADC targeting folate receptor-a, contains the VH44-VL100 cysteine mutation to prevent aggregation of the scFv and a C220S mutation in the Fc domain to reduce the formation of free cysteines [13]. Identifying the correct expression constructs and the expression construct ratios for stable single cell expression can be a challenge.
Some of the things to consider when developing bispecific ADCs are in Figure 3. It is important to show that the bispecific ADC provides superior efficacy and safety compared to the monospecific ADCs. The addition of the monospecific ADCs individually and in combination will be important control groups to include in your preclinical studies. It will also be important to show the bispecific ADC is superior to the monospecific ADCs and the combination of the monospecific ADCs in tumor models with heterogeneous expression of each antigen. Lastly having antibodies with similar binding affinities to the human and non-human primate antigens will be important for the non-GLP and GLP toxicology studies.
Discussion
Bispecific ADCs offer another way of addressing the potential issue of drug resistance due to the heterogeneous expression of ADC targets in tumors. Bispecific ADCs may also improve tumor targeting since both antigens may have higher expression in tumors compared to normal tissues. Bispecific ADCs, including biparatopic ADCs, can increase the internalization rates and lysosomal trafficking of ADCs and may result in a more effective ADC compared to monospecific ADCs. For monospecific ADCs, when the expression of one ADC target is low, the ADC may be ineffective therefore targeting two tumor antigens with one ADC will minimize the chance the bispecific ADC loses its effectiveness, if one tumor antigen has low expression.
Currently there are no approved bispecific ADCs and the majority of bispecific ADCs are in early clinical development. The most clinically advanced bispecific ADCs, JSKN-003 (Her2 biparatopic) and BL-B01D1 (EGFR x Her3) are in phase 3 in China and in early clinical development in the US. It will be interesting to see if these bispecific ADCs provide significant advantages over the monospecific ADCs.
The manufacturing challenges for bispecific ADCs need to be considered. The manufacturing challenges could be significant depending on the antibody format, linker/payload selected and the conjugation method. Stringent QC criteria and robust lot release assays need to be implemented early during development to minimize the probability of having a manufacturing batch failure in late development, which could be costly and result in significant delays.
Bispecific ADCs could offer significant advantages over the monospecific ADCs in terms of efficacy and safety by improving the delivery of the ADC to the tumor and less to normal tissues. The ADC space is very competitive and the avenues available to significantly differentiate from competitors may be limited. Bispecific ADCs may provide a point of significant differentiation in the ADC space and more importantly provide significant benefit to cancer patients.
References
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