Antibody drugs have emerged in a variety of innovative and diverse forms, including Antibody Drug Conjugates (ADCs), bispecific antibodies, nanobodies, and other forms.

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Bispecific Antibody Drug Conjugates (ADCs):
Emerging Trends
Since the FDA approved the first monoclonal antibody in 1986, antibody drugs have
emerged in a variety of innovative and diverse forms, including Antibody Drug Conjugates
(ADCs), bispecific antibodies, nanobodies, and other forms.
ADCs are known as biological missiles, and there are currently 14 drugs approved
worldwide. The primary mechanism of action is internalization of the ADC, with the
release of the payload into the cytoplasm after cleavage of the linker. This makes the ADC
drugs very demanding in terms of the targets to which the antibodies bind.
Bispecific antibodies have long been highly anticipated. Unlike monoclonal antibodies,
bispecific antibodies can recognize two epitopes of a target or two different targets at the
same time, thus producing better efficacy through multiple different molecular
mechanisms. However, the design principles require a high level of immunological
understanding, and the challenges of light and heavy chain mismatches, lack of purity,
and lack of thermal stability at the manufacturing level have resulted in few approved
bispecific antibodies. There are currently only five bispecific antibodies approved
worldwide.
The rapid development of bispecific antibody (BsAb) technology contributes to more
choices of antibodies as a partner of ADCs. Conjugating payload into BsAb to improve
specificity and internalization, called bispecific ADCs (BsADCs), has been studied in
recent years, which may be a solution to the existing problems of endocytosis, toxicity and
drug resistance of ADC drugs.
What Are The Features Of Bispecific ADCs?

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The dual anti-ADCs currently under investigation can be divided into two main
categories: bispecific ADCs targeting dual tumor associated antigens (TAA) (e.g.,
MUC1/EGFR-ADC), and bispecific ADCs targeting dual epitopes (e.g.,
HER2/HER2-ADC).
Tumor-associated antigen (TAA) is an important class of druggable targets that are highly
expressed specifically on tumor cells. With the rapid development of linker and payload
technologies, significant progress has been made in the development of TAA-based
antibody-coupled drugs (ADCs), with 14 ADCs targeting 11 TAA targets approved to date.
Bispecific ADC drugs targeting dual TAA are a brand new form of escalating on top of the
advantages of ADC.
(i) Two targets can enhance tumor targeting and reduce the toxicity to normal tissues.
(ii) Certain dual-epitope bispecific ADCs or dual-target bispecific ADCs can produce better
internalization and enhance the killing effect.
(iii) Overcome the problem of drug resistance caused by decreased single-target
expression. However, while dual-targeting brings better efficacy, it may also have the
problem of a too narrow therapeutic window. Especially in TAA, the expression of this
target also exists in non-tumor tissues, and the toxicity of the two combinations together
may exceed the simple sum of the two.

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Fig. 1. Schematic representation of fragment expression, the split intein trans-splicing,
and drug conjugation process used to generate reactivated BsADC. Source: reference 1
In terms of global drug development, the research and development of the bispecific ADC
pipeline are still in its early stage and there are few preclinical products in development.
There are three major challenges ahead.
 • First, there are few antibody backbone molecules that can be used for bispecific
antibody construction, and the targets are very concentrated, with only a few targets such
as HER2, HER3, EGFR, MUC1, etc. There is a lack of sufficient "backbone" for the rapid
construction of stably expressed bispecific ADC molecules.
 • Secondly, most of the current antibodies against these molecules are screened
by blocking activity, and it is not yet known whether their endocytosis activity meets the
demand and whether they can be the best candidate antibody molecules for ADC drugs.
 • From the perspective of the production process, bispecific antibody production
suffers from low efficiency due to mismatch, on top of which the coupling reaction is
performed, further increasing the difficulty and challenge of the production process.
Bispecific ADCs Under Investigation
M1231
M1231 is an EGFR/MUC1 bispecific ADC developed by SUTRO that uses the company's
proprietary fixed-point coupling technology to attach Hemiasterlin derivatives to specific
amino acid sites of the antibody via a cleavable linker with a DAR value of about 4.

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Figure 2. Structure of M1231, source: reference 1
EMD Serono presented preclinical data of M1231 at 2020 AACR. The results showed that
a single dose of M1231 exhibited some tumor curative effect in NSCLC and sqCC
Esophageal PDX models, and there was a correlation between tumor remission effect and
target expression.
Figure 3. Strong anti-tumor activity in patient-derived xenograft models of M1231, source:
reference [2]
In January 2021, an open-label, non-randomized, parallel, dose-escalating,
dose-expanding Phase I clinical trial (NCT04695847) was initiated in the United States
and Canada to evaluate the safety, pharmacokinetics, tolerated dose and clinical efficacy
of M-1231 as a single agent in the treatment of patients with advanced solid tumors
(expected n=84).

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ZW-49
ZW-49 is a bispecific ADC drug developed by Zymeworks that specifically binds both
non-overlapping epitopes of the HER2 receptor (ECD4/trastuzumab and
ECD2/pertuzumab). The bispecific ADC is based on ZW25 coupled to Auristatin toxin A
via a protease cleavage linker.
Figure 4. Structure of ZW49, and its advantages, source: reference [3]
In preclinical studies, ZW-49 has shown excellent druggability. According to data from the
company's preclinical animal model study published in 2020, ZW-49 showed superior
anti-cancer effects in both low and high HER2 expressing breast cancer models and
showed some efficacy in a HER2 high expressing breast cancer brain metastasis model.

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Figure 5. Preclinical efficacy of ZW49, source: Zymeworks website
Compared with the marketed Trastuzumab emtansine (Roche anti-HER2-ADC, launched
in 2013) and Trastuzumab deruxtecan (AstraZeneca and Daiichi Sankyo anti-HER2-ADC,
launched in 2019), ZW-49 has a more superior performance in shrinking tumor volume
and improving survival rates.
In January 2021, Zymeworks published selected data from the Phase I clinical trial of
ZW-49. The results showed that ZW-49 demonstrated antitumor activity across all
treatment regimens and dose levels. Partial remission (PR) and stable disease (SD) were
observed in both Q2W and Q3W dosing regimens. And in all 35 patients treated with
ZW-49, no treatment-related hematotoxicity, pulmonary toxicity and hepatotoxicity were
observed. More than 90% of the adverse reactions were mild or moderate.
REGN5093- M114

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REGN5093- M114 is a bispecific ADC developed by Regenron targeting two different
epitopes of MET. The antibody is a 1+1 asymmetric bispecific antibody that links the
antibody to the toxin M24 ( maytansine derivative) through the M114 linker with a DAR
value of about 3.2.
Figure 6. Structure of regn5093-m114
REGN5093-M114 has been initiated in Phase I/II clinical trial (NCT04982224) in the U.S.
in July 2021, with an expected completion date of December 2026.
REGN5093-M114 naked antibody itself also has some tumor suppressive effect and has
demonstrated good efficacy in a variety of preclinical models. In various animal tumor
models, REGN5093-M114 at 10 mg/kg could completely inhibit tumor cells or even clear
them, while METxMET bispecific antibody without toxin coupling could only inhibit tumor
growth at doses up to 25 mg/kg, and could not clear tumor cells. And compared with the
MET ADC drugs already in the clinic, the bispecific ADC REGN5093-M114 could not only
inhibit tumor growth but even remove tumor cells at the same dose (Figure 7 below), so its
efficacy is much better than the ADCs in the clinic.

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Figure 7. METxMET-M114 compares favorably to a clinical-stage MET ADC. Source:
reference [4]
BL-B01D1
BL-B01D1 is an EGFR/HER3 bispecific ADC developed by Baili Pharmaceuticals, which
adopts its unique AC linker to link the camptothecin derivative ED04 to the cysteine site of
the antibody with a DAR value of about 7.5. The results of the preclinical study showed
that BL-B01D1 was able to exert good anti-tumor effects at an administered dose of 10
mg/kg. On October 19, 2021, Baili Pharma's clinical trial application for BL-B01D1
received implied permission from the NMPA for the treatment of advanced or metastatic
epithelial tumors.
Figure 8. Structure of BL-B01D1
ADC drugs have been developed over several decades before they have begun to show
results. The emergence of bispecific ADCs is an outgrowth of the trend toward antibody
engineering and coupling technology. Although there are still a few projects carried out

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and failures have occurred, this is just the general rule of drug development. It is believed
that the concept of bispecific antibodies can provide more ideas in the current context of
multiple factors that emphasize differentiation and temporary target "depletion".
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References:
1. Zong, Hui-Fang & Zhang, Bao-Hong & Zhu, Jianwei. (2022). Generating a Bispecific Antibody Drug
Conjugate Targeting PRLR and HER2 with Improving the Internalization. Pharmaceutical Fronts. 04.
e113-e120. 10.1055/s-0042-1749334.
2. R&D update call 2021, Merck KgaA
https://www.merckgroup.com/investors/events-and-presentations/conferences-and-roadshows/2021/en/
211122_RD_UpdateCall_Final_EN.pdf
3. https://www.zymeworks.com/_publications/12-8-2018-PRESENTATION.pdf
4. DaSilva JO, Yang K, Surriga O, et al. A Biparatopic Antibody-Drug Conjugate to Treat
MET-Expressing Cancers, Including Those that Are Unresponsive to MET Pathway Blockade. Mol
Cancer Ther. 2021;20(10):1966-1976. doi:10.1158/1535-7163.MCT-21-0009
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