Antibody-drug conjugates (ADCs) are emerging anti-cancer treatment agents that combine the strong cytotoxicity of small molecule drugs with the specificity of monoclonal antibodies (mAb). Studies have shown that human epidermal growth factor receptor-2 (HER2) is overexpressed in many tumors and is one of the most common target antigens for ADCs.
Introduction of HER2
Human epidermal growth factor receptor-2 (HER2) is a transmembrane receptor for tyrosine kinases and a member of the epidermal growth factor receptor (EGFR) family, which also includes HER1, HER3 and HER4, each of which includes an extracellular domain, lipophilic transmembrane region, intracellular domain containing tyrosine kinase, and a carboxy-terminal region. All of these four members are essential for regulating cell proliferation and differentiation through ligand-dependent active forms or independent homodimers or heterodimers.
However, unlike the other three members, the extracellular domain of HER2 is the only one that dimerizes with any of the other members in the activated state without binding ligands and then triggers signaling pathways that regulate cell proliferation and survival, including the PI3K/AKT, MAPK and JAK/STAT pathways, etc. (Figure 1).
Figure 1. Overview of the HER2 signaling pathway. Source: Reference 
Currently, there is increasing evidence that HER2 is overexpressed in various cancers, such as breast, gastric, lung, and ovarian cancers. In particular, up to 15-30% of breast cancer patients carry HER2 gene amplification and overexpression. Therefore, HER2 is considered a valuable prognostic factor for breast cancer and an effective therapeutic target for the treatment of various HER2-positive cancers.
Approved HER2 ADCs
Currently, three HER2 ADC drugs are approved worldwide, T-DM1 (Kadcyla, Trastuzumab emtansine), T-DXd (DS-8201, Enhertu, Trastuzumab deruxtecan）and RC48 (Disitamab vedotin).
|Fam-trastuzumab deruxtecan-nxki||Trastuzumab (IgG1)||DM1 (Maytansine)||3.5||Lysine-SMCC||Metastatic breast cancer||Approved|
|Ado-trastuzumab emtansine||Trastuzumab (IgG1)||DXd (Topoisomerase I inhibitor)||8.0||Cysteine-maleimide||Metastatic breast cancer||Approved|
|Disitamab vedotin||Disitamab (IgG1)||MMAE (Auristatin)||4.0||Cysteine-maleimide||Metastatic urothelial cancer||Phase III/Approved in China|
Table 1. List of Approved ADCs, source: references 
T-DM1 (Kadcyla, Trastuzumab emtansine)
T-DM1, developed by Roche, was the first anti-HER2 ADC to receive FDA approval. It consists of trastuzumab conjugated to the maytansine derivative DM1 via a thioether linker. DM1 effectively inhibits microtubulin polymerization and blocks mitosis, which leads to apoptosis.
Figure 2. Structure of Kadcyla. Source: https://doi.org/10.3390/ph13090245
In 2013, T-DM1 was approved by the FDA for the treatment of advanced HER2+ breast cancer.
In 2019, T-DM1 was also approved for adjuvant (after surgery) treatment of people with HER2-positive early breast cancer (EBC) who have residual invasive disease after neoadjuvant (before surgery) taxane and Herceptin® (trastuzumab)-based treatment.
T-DXd (DS-8201, Enhertu, Trastuzumab deruxtecan）
T-DXd was the second anti-HER2 ADC to receive FDA approval. DS-8201a has a drug-to-antibody-ratio (DAR) of 8, which is higher than that of T-DM1 (3.5). Meanwhile, T-DXd uses DXd, a potent topoisomerase I inhibitor, as a carrier drug, which has approximately 1000 times more antitumor activity than conventional chemotherapy drugs and can effectively avoid cross-resistance. In addition, DXd has better cell membrane permeability and can penetrate cell membranes into adjacent tumor cells to exert potent bystander effects.
Figure 3. DS-8201 Structure, source: doi:10.1248/cpb.c18-00744
In 2019, T-DXd was approved by FDA for unresectable or metastatic HER2-positive breast cancer following two or more prior anti-HER2 based regimens.
After that, T-DXd was successively approved for the treatment of unresectable or metastatic non-small cell lung cancer (NSCLC) and HER2-positive gastric or gastroesophageal junction adenocarcinoma.
At the 2022 ASCO Annual Meeting, the Phase 3 clinical results of T-DXd for HER2-low metastatic breast cancer were announced, which received a standing ovation. According to the study, T-DXd reduced the risk of disease progression or death by a whopping 50 percent, compared with conventional chemotherapy, in HER2-low metastatic breast cancer, which opened a new era of breast cancer treatment. In August 2022, FDA approved T-DXd for the treatment of adult patients with unresectable or metastatic HER2-low breast cancer.
In addition, DS-8201 is also in clinical trials for pan-cancer types, including colorectal cancer, uterine cancer, biliary tract tumors, and pancreatic cancer, etc.
RC48 (Disitamab vedotin)
RC48 (Disitamab vedotin), developed by RemeGen, is an innovative HER2 ADC comprising a novel HER2-directed monoclonal antibody, disitamab, conjugated to monomethyl auristatin E (MMAE) via a protease-cleavable mc-vc linker. The antibody disitamab targets different epitopes of the HER2 receptor and has a better molecular affinity for HER2 targeting compared to trastuzumab. The mc-vc linker is stable and can only be cleaved by cathepsin when RC48 is cytosolicized into the lysosome, thus releasing a payload to kill tumor cells. The toxin MMAE is a synthetic derivative with anti mitogenic effects.
Figure 4. Structure of Disitamab Vedotin, source: doi: 10.1080/10717544.2022.2069883
In 2021, RC48 received conditional marketing approval in China for the treatment of patients with HER2-overexpressing locally advanced or metastatic gastric cancer (including gastroesophageal junction adenocarcinoma) who have received at least two systemic chemotherapy regimens.
HER2 ADCs Under Investigation
There are many HER2 ADCs in clinical trials for cancer treatment.
|SYD985||Trastuzumab (IgG1)||DUBA (Duocarmycin)||2.8||Cysteine-maleimide||Metastatic breast cancer||FDA rejected/Phase III|
|ZRC-3256||Trastuzumab (IgG1)||DM1 (Maytansine)||-||Lysine-SMCC||Metastatic breast cancer||Phase III|
|MRG002||IgG1||MMAE (Auristatin)||-||Unknown||Advanced solid tumors||Phase II|
|ARX788||Transtuzumab (IgG1)||MMAF(Auristatin)||1.9||pAF-hydroxylamine-PEG4||Metastatic breast cancer/gastric cancer||Phase II/III|
|BDC-1001||Trastuzumab (IgG1)||TLR7/8 agonist||-||Unknown||Metastatic breast cancer/gastric cancer||Phase I/II|
|A166||Trastuzumab (IgG1)||MMAF (Auristatin)||2||Lysine site-specific||Metastatic breast cancer||Phase I/II|
|FS-1502||Trastuzumab (IgG1)||MMAF (Auristatin)||2.0||Cysteine-maleimide||Breast cancer||Phase I|
|SHR-A1201||Trastuzumab (IgG1)||DM1 (Maytansine)||-||Lysine SMCC||Metastatic breast cancer||Phase I/II|
|DP303c||Trastuzumab (IgG1)||DP104n||2.0||Unknown||Gastric cancer||Phase II|
|BI-CON-02||Trastuzumab (IgG1)||Unknown||-||Unknown||Metastatic breast cancer||Phase I|
|ALT-P7||Trastuzumab biobetter HM2 (IgG1)||MMAE (Auristatin)||2.0||Cysteine-maleimide||Metastatic breast cancer||Phase I|
|DX126-262||DX-CHO9 (IgG1)||Tubulysin||-||Cysteine-maleimide||Breast/gastric cancer||Phase I|
|ZW49||IgG1||Auristatin||-||Cysteine-MC||Metastatic cancers||Phase I|
|HS630||Trastuzumab (IgG1)||DM1 (Maytansine)||-||Unknown||Breast cancer||Phase I|
|B003||Trastuzumab (IgG1)||DM1 (Maytansine)||-||Lysine-SMCC||Metastatic breast cancer||Phase I|
|SBT6050||IgG1||TLR8 agonist||-||Unknown||Advanced solid tumors||Phase I/II|
|SHR-A1811||Unknown||Unknown||-||Unknown||Advanced solid tumors||Phase I/II|
|MT-5111||Unknown||SLTA||-||Unknown||Advanced solid tumors||Phase I|
|GQ1001||Unknown||DM1 (Maytansine)||2.0||Unknown||Advanced solid tumors||Phase I
Table 2. List of HER2 ADCs in Clinical Trails, source: reference 
SYD985 (Trastuzumab duocarmazine)
SYD985 is an ADC that targets HER2 and consists of the monoclonal antibody trastuzumab and a cleavable linker known as valine-citrulline-seco-DUocarmycin-hydroxyBenzamide-Azaindole (vc-seco-DUBA). The antibody of SYD985 is able to bind to HER2 targets on the surface of cancer cells, resulting in ADC internalization by the cells. Following hydrolytic cleavage of the linker, the inactivated cytotoxin is activated, which induces DNA damage and ultimately tumor cell death. In addition, uptake of the activated payload by neighboring tumor cells with lower HER2 expression may improve the efficacy potential, the so-called bystander effect.
In July 2022, The FDA accepted a Biologics License Application (BLA) for SYD985 in patients with HER2-positive unresectable locally advanced or metastatic breast cancer (MBC).
Meanwhile, on July 18, 2022, the EMA validated the marketing authorization application (MAA) for SYD985 for the treatment of patients with HER2-positive unresectable, locally advanced or metastatic breast cancer. In May 2023, the FDA has suspended its decision on approving SYD985 following the complete response letter, and requested additional information requiring time and resources beyond the current review period for the agent.
ARX788 is a homogeneous and highly stable ADC with a similar mechanism of action to T-DM1. It is comprised of an anti-HER2 mAb Herceptin® (trastuzumab) site-specifically conjugated a potent tubulin inhibitor payload AS269 is a non-natural amino acid incorporated into the antibody.
Results from a phase I trial of the safety, tolerability, and pharmacokinetics of ARX788 monotherapy in Chinese patients with advanced HER2 breast cancer showed a favorable safety profile and a tumor treatment response in the target dose group following efficacy evaluation.
On March 2023, positive clinical data were obtained from a randomized Phase 3 clinical trial (ACE-Breast-02) of ARX788,. The trial enrolled 441 patients with HER2-positive breast cancer who were refractory to prior treatment with paclitaxel and trastuzumab. Eligible patients were randomized 1:1 to receive either the new experimental drug ARX788 or the control drug lapatinib in combination with capecitabine. Results showed that ARX788 significantly improved progression-free survival (PFS) in patients with HER2-positive locally advanced or metastatic breast cancer.
Last December, Ambrx announced the data of ACE-Breast-03, a phase II clinical trial of ARX788, which showed that among seven patients with advanced breast cancer with evaluable efficacy, five achieved PR and tumor shrinkage of more than 60% in all cases. It is worth mentioning that all seven patients had previously received T-DM1 treatment.
In this published Phase II/III clinical analysis, preliminary clinical data showed that ARX788 was approximately 25% effective in patients who were resistant to DS-8201 for 12 months. In this regard, Ambrx's CEO believes that ARX788 may become the first choice for DS-8201-resistant patients.
MRG-002 is under clinical development by Shanghai Miracogen and currently in Phase II for HER2 positive breast cancer. It is composed of a humanized anti-HER2 mAb trastuzumab conjugated to MMAE via a cleavable vc linker.
The high-affinity of trastuzumab specifically binds to HER2 antigen on the surface of tumor cells, internalizes and releases the cytotoxic payload MMAE, which effectively inhibits microtubule protein polymerization, thereby interfering with mitosis and exerts tumor cell killing effects. The innovatively modified trastuzumab has a selectively high level of fucosylation in the Fc region, resulting in a reduced Fc binding to CD16a expressed on effector immune cells, minimizing potential killing of CD16a expressing immune cells by MRG002 and therefore reducing potential adverse effects on patients.
In the multicenter phase II study of MRG002 for advanced HER2 low-expressing breast cancer, results showed that of 18 patients with HER2 low-expressing breast cancer who had previously failed standard first-line therapy, 5 had achieved PR on first efficacy assessment. In a clinical study of patients with HER2-positive breast cancer after multiple lines of therapy, an ORR of 55% was achieved in 55 patients with HER2-positive breast cancer who had received a median of 5 lines of prior therapy, with a common adverse effect of decreased neutrophils.
HER2 ADCs Failed in Clinic Settings
Though many HER2 ADCs approved, there are also many HER2 ADCs failed in clinical settings.
|TAA013||Trastuzumab (IgG1)||DM1 (Maytansine)||3.5||Lysine-SMCC||Metastatic breast cancer|
|XMT-1522||HT-19||Auristatin F-HPA (Auristatin)||12||Cysteine-fleximer polymer||Breast/gallbladder/gastric cancer|
|ADCT-502||Trastuzumab||SG3199 (PBD)||1.7||Cysteine site-specific||Breast/bladder/gastric cancer|
|MEDI4276 (Bispecific)||Trastuzumab& 39S||AZ13599185 (Tubulysin)||4||Cysteine site-specific||Breast/gastric cancer|
|NJH395||Unknown||TLR7 agonist||Unknown||Unknown||Solid tumors|
|MM-302||F5||Doxorubicin||Unknown||Unknown||Advanced solid tumors|
|BAT8001||BAT0606||Maytansinoid||3.5||Cysteine-3AA||Advanced solid tumors|
Table 3. List of anti-Her2 ADCs failed in clinic settings, source: reference 
The approved HER2-targeted ADCs have significantly benefited cancer patients, and the ones being currently investigated may provide even further benefit.
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 Najjar MK, Manore SG, Regua AT, Lo HW. Antibody-Drug Conjugates for the Treatment of HER2-Positive Breast Cancer. Genes (Basel). 2022 Nov 8;13(11):2065. doi: 10.3390/genes13112065. PMID: 36360302; PMCID: PMC9691220.
 Iqbal N, Iqbal N. Human Epidermal Growth Factor Receptor 2 (HER2) in Cancers: Overexpression and Therapeutic Implications. Mol Biol Int. 2014;2014:852748. doi: 10.1155/2014/852748. Epub 2014 Sep 7. PMID: 25276427; PMCID: PMC4170925.
 Xinling Zhang, Andrew C Huang, Fahai Chen, Hu Chen, Lele Li, Nana Kong, Wenting Luo, Jianmin Fang, Novel development strategies and challenges for anti-HER2 antibody-drug conjugates, Antibody Therapeutics, Volume 5, Issue 1, January 2022, Pages 18–29, https://doi.org/10.1093/abt/tbac001
 Ferraro, E., Drago, J.Z. & Modi, S. Implementing antibody-drug conjugates (ADCs) in HER2-positive breast cancer: state of the art and future directions. Breast Cancer Res 23, 84 (2021). https://doi.org/10.1186/s13058-021-01459-y
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