As a new type of anti-tumor drug, Antibody-Durg Conjugate (ADC) consists of antibody, linker and cytotoxic drug. One end of the ADC is an antibody that specifically recognizes cancer cells, and the other end is connected to cytotoxic chemotherapy drugs. Antibodies can accurately deliver chemotherapy drugs to cancer cells, so ADC has both the powerful lethality of small molecule drugs and the high targeting of monoclonal antibodies, which is a hot field of anticancer drug research and development these years. In recent years, driven by the development of antibody drugs and the progress and iteration of ADC technology, ADC drug research and development has developed rapidly.
The more popular targets in ADC drugs include HER2, TROP2, EGFR, CLDN18.2, c-Met, CD19, PSMA, Muc1, BCMA and PDL1, and most of them are mature targets that have been verified by the market. According to statistics, there are more than 140 ADC-targeted drugs in clinical trials worldwide.
Here we introduce a new ADC target TPBG. TPBG, the full name is trophoblast glycoprotein. Recently, TPBG has attracted extensive interest from many pharmaceutical companies. According to clinical data from ClinicalTrials, more than 10 drugs targeting TPBG are currently under development. Among them, the carcinoembryonic antigen tumor vaccine (Trovax) targeting TPBG has made the fastest progress, and has completed phase II and phase III clinical trials of various tumors. A large number of studies have revealed that TPBG is a dual-function carcinoembryonic antigen, which is not only indispensable for embryonic growth and development, but also promotes the development of tumors, making it an opportunity target for tumor immunotherapy.
The Structure of TPBG
Trophoblast glycoprotein (TPBG) is a glycoprotein present in embryonic trophoblast cells, also known as carcinoembryonic protein 5T4 and WAIF1.
TPBG is an extracellular highly glycosylated type I single transmembrane protein, containing 420 amino acids, including an extracellular domain of 310 amino acids, a transmembrane region of 20 amino acids and a cytoplasmic tail of 44 amino acids. TPBG can influence cytoskeletal reconstruction and intercellular integrity through its PDZ domain. There are 7 N-glycosylation sites in the extracellular amino acid part, which has a variety of structures and can prevent protein hydrolysis.
The TPBG glycoprotein also has seven leucine-rich repeats (LRRs), which are involved in the interaction between proteins (Figure 1). Compared with the human and mouse TPBG sequences, the two species are highly conserved, and the amino acid identity level of the transmembrane region and the cytoplasmic part reaches 81%. A large amount of evidence shows that TPBG is closely related to different physiological and pathological processes, such as intercellular connections, cell morphology and movement, cell adhesion, cell membrane integrity, etc.
Figure 1 The structure of TPBG
(Source: doi: 10.1007/s00262-019-02419-4)
Expression And Function of TPBG
The expression of TPBG in adult normal tissues is limited to certain special epithelial cells, such as basal stratified squamous epithelium, glandular and ductal epithelium, as well as retinal secondary neurons and olfactory bulb. TPBG is relatively widely expressed during embryonic development, and TPBG is strongly expressed in placental trophoblast cells throughout pregnancy. TPBG not only plays an important role in embryonic cell development, imbedding and implantation, but also can guide cell differentiation and embryonic organogenesis during the embryonic period.
Successive studies have shown that TPBG is highly expressed in a variety of solid tumor malignancies, including mesothelioma, non-small cell lung cancer, breast cancer, head and neck cancer, cervical cancer, kidney cancer, gastric cancer and colorectal cancer, the expression level of TPBG is positively correlated with tumor metastasis, poor prognosis and clinical stage. TPBG, however, is rarely found in normal mature tissues and is considered to be a tumor-associated antigen, which is an important design target for anticancer drugs. In other words, TPBG is not only essential for embryonic growth and development, but also may promote tumor cells to have similar local infiltration and ability to evade immune surveillance as trophoblast cells.
Figure 2 Protein expression of TPBG in cancer cells of different organs
Mechanism of Action of TPBG
At present, the signaling pathway and molecular mechanism of TPBG in the process of tumorigenesis and development have not yet been clarified. Existing studies have shown that TPBG may play a role through these three mechanisms: Wnt signal transduction, Epithelial-mesenchymal transition (EMT) and Regulation of the CXCL12/CXCR4 biological axis.
1.Wnt signal transduction
At present, the mechanism of TPBG's effect on tumor is not very clear. Studies have shown that TPBG can block both classical Wnt/β-catenin and non-classical Wnt signaling pathways. The Wnt signaling pathway is closed in normal mature cells, but its activity is enhanced in tumor cells. On the one hand, TPBG inhibits the activation of Wnt/β-catenin classical pathway through its interaction with LRP6. On the other hand, TPBG affects the Wnt pathway by activating non-classical Wnt signaling pathways.
Figure 3 TPBG affects Wnt signaling pathway
(Source: DOI 10.1016/j.devcel.2011.10.015)
In addition to affecting the transduction of the Wnt signaling pathway, some studies believe that TPBG can promote cell morphology changes as well as metastasis by affecting epithelial mesenchymal transition, and some studies have suggested that TPBG affects the metastasis and proliferation of tumor cells through the CXCRL12-related pathway. Regardless of the specific mechanism of influence, the high expression of TPBG in tumor cells and its correlation with metastasis and poor prognosis make it one of the ideal targets for tumor drugs.
It has been found that in breast cancer, gastric cancer and melanoma, TPBG can allosteric Wnt auxiliary receptor LRP5/6 through DKK1 protein, thus inhibiting Frizzled-LRP5/6 generation, preventing LRP5/6 signal amplification, blocking classical Wnt signal, and thus preventing normal cell development and differentiation. On the other hand, TPBG activates non-classical Wnt signaling pathway through DKKl protein and JNK gene to promote cytoskeletal recombination and enhance cell motility, resulting in the spread of cancer cells and the decline of clinical survival of patients.
2. Epithelial-mesenchymal transition (EMT)
Epithelial-mesenchymal transition (EMT) plays an important role in the process of tumor invasion and metastasis. TPBG induces down-regulation of E-cadherin expression, enhances cytoskeletal actin reorganization, weakens cell adhesion, and promotes cell morphology changes and metastasis.
3. Regulation of the CXCL12/CXCR4 biological axis.
TPBG promotes chemotaxis and metastasis of tumor cells through the CXCR4/CXCL12 pathway; or further selectively expresses CXCR7 to promote tumor cell proliferation through the CXCR7/CXCL12 pathway.
Clinical Drugs Under Development for TPBG
Many pharmaceutical companies around the world are carrying out clinical trials of drugs targeting TPBG, including Aptevo Therapeutics, Inc., Inhibrx, Inc., Byondis BV, Imbioray (Hangzhou) Biomedicine Co., Ltd., Xadcera Biopharmaceutical (Suzhou) Co., Ltd. etc.
At present, there are more than 10 clinical drugs in research for TPBG in the world, none of which are listed, and the fastest one is in clinical phase III. These drugs are of various types (tumor vaccine, fusion protein, double antibody, monoclonal ADC, CAR-NK), and their indications include solid tumors such as non-small cell lung cancer, renal cell carcinoma, bladder cancer, colon cancer, breast cancer, prostate cancer, and ovarian cancer. Multiple immunotherapies are involved, including monoclonal antibodies, bispecific antibodies, trispecific antibodies, ADCs, and tumor vaccines (Table 1).
Table 1 Clinical research progress of TPBG
As mentioned above, data from several pharmaceutical companies have revealed that TPBG drugs are closely related to clinical benefits, showing that TPBG antibodies or vaccines have important value in cancer treatment. Therefore, TPBG is expected to become a potential target for the next anti-tumor immunotherapy.
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 Zhao, Yuguang, et al. "Structural insights into the inhibition of Wnt signaling by cancer antigen 5T4/Wnt-activated inhibitory factor 1." Structure 22.4 (2014): 612-620.
 Stern, Peter L., and Richard Harrop. "5T4 oncofoetal antigen: an attractive target for immune intervention in cancer. "Cancer Immunology, Immunotherapy 66.4 (2017): 415-426.
 Spencer, Helen L., et al. "Role of TPBG (trophoblast glycoprotein) antigen in human pericyte migratory and angiogenic activity. "Arteriosclerosis, Thrombosis, and Vascular Biology 39.6 (2019): 1113-1124.
 Harrop, Richard, Eric O'Neill, and Peter L. Stern. "Cancer stem cell mobilization and therapeutic targeting of the 5T4 oncofetal antigen. " Therapeutic advances in vaccines and immunotherapy 7 (2019): 2515135518821623.
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 Tsuboi, Akio. "LRR-containing oncofetal trophoblast glycoprotein 5T4 shapes neural circuits in olfactory and visual systems. "Frontiers in Molecular Neuroscience 13 (2020): 581018.
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