Based on the unique design and mechanism, T-DXd opens up a sea of stars for HER2-positive advanced gastric cancer

The ADC consists of antibodies, cytotoxic drug loaders, and linkers; The main mechanism is to use the specific targeting effect of antibodies to deliver the drug to the target cells, so that the drug can be concentrated on the tumor cells to exert anti-tumor killing effect, and can reduce the drug concentration in other normal tissues and organs, so as to obtain a wider treatment window
.
Specifically, after the ADC enters the body, it selectively binds to the antigen of the target cell through the targeting effect of the antibody; The formed ADC-antigen complex enters tumor cells
by endocytosis.
Subsequently, under the action of lysosomes, cytotoxic drugs are released through cleavage of linkers, which play the role of killing tumor cells ^[12].

In the above process, the affinity of antigens and antibodies, the stability of linkers, the potency of cytotoxic drug loading, membrane permeability, and release timing will all affect
the efficacy of ADC drugs.

Figure 1.
ADC structure and mechanism of action ^[12].

Figure A.
General ADC structure
.
These include antibodies, linkers, and
payloads.
Figure B The mechanism
of action of the general ADC.
After distribution through the hematological system to tumor tissue (I), the ADC binds to its cell surface antigen, and the ADC-antigen complex is internalized by antigen-mediated endocytosis (II).

ADC processing (linker cleavage and/or antibody degradation) occurs during endosome-lysosome (IIIa, IIIb) transport, resulting in the release of the drug load into the cytoplasm (IV) in a biologically active form, ultimately leading to cell death (V).

If the drug load is permeable to the cell membrane, it can enter adjacent cells to achieve the bystander effect (VI).

As an emerging anti-tumor drug, the research and development exploration of ADC has never stopped, and the continuous optimization of various components has effectively promoted the development process of ADC, which has undergone three generations of technological changes
so far.
T-DXd targeting HER2 has improved off-target toxicity, hydrophobicity and homogeneity, and has become a "bright star"
in the third-generation ADC drugs.
T-DXd is formed by humanized HER2 antibody conjugated to topoisomerase I inhibitor (DXd) via cleavable linkers, which has many advantages
in structure and mechanism of action.

Figure 2.
T-DXd structure ^{diagram [13].}

Structure of T-DXd: the antibody moiety is conjugated to the linker-drug-loading moiety by cysteine residues, and the DAR reaches a maximum of 8
.

Antibody conjugation of DXd did not affect its HER2 recognition ability and ADCC effect

The antibody component of T-DXd is humanized trastuzumab
.
Preclinical studies have found ^[14] that T-DXd and naked antibodies bind to HER2 at different concentrations with similar affinities and have similar dissociation constants (Kd, 7.
3 ng/mL and 7.
8 ng/mL, respectively).

Therefore, antibody conjugation of DXd did not affect its HER2 recognition ability
.
In addition to the direct killing of drugs, antibody-dependent cell-mediated cytotoxicity (ADCC) effects also play an important role
in the process of ADC drugs killing tumors.
Preclinical studies have found that T-DXd has similar ADCC effects
to naked antibodies.

Figure 3.
T-DXd contrasts HER2 binding capacity and ADCC activity of naked antibodies^[14].

Figure A.
The affinity of T-DXd (DS-8201a) and naked antibody (anti-HER2 Ab, trastuzumab) for HER2 protein is determined by ELISA
.
Figure B The ADCC activity
of T-DXd and naked antibody was compared by measuring the degree of lysis of SK-BR-3 cells by human peripheral blood mononuclear cells.

The linker is the key structure of the ADC, which determines whether the ADC drug has a bystander effect and the size of the drug-load/antibody ratio (DAR), which will seriously affect the efficacy
of the drug.
First, the linker must have some stability to ensure the integrity of the ADC drug during blood circulation before reaching the target cell to avoid damage to normal tissues or cells
by early drug release.
T-DXd uses tetrapeptide-based digestion to cleave linkers, which are more
stable than traditional aminobenzyl linkers.
Preclinical studies have shown [^14] that the presence of free DXd
in plasma is detected only at the beginning after a single injection into mice, rats, monkeys, and humans.
After 21 days, the release rate of DXd in plasma is only 1.
2%-3.
9%, of which the release rate in human plasma is about 2%, while the release rate of T-DM1 is as high as 18.
4%
on day 4.
It shows that the tetrapeptide linker of T-DXd is highly stable in the blood, and the DXd shedding rate is extremely low, which ensures the safety
of the system.

Figure 4.
The T-DXd linker is stable in plasma and has a very low drop-off rate ^[14].

Left: Pharmacokinetics
of T-DXd in cynomolgus monkeys.
T-DXd is administered at a dose of 3 mg/kg
intravenously.
Right: DXd release rate
in plasma.

When the ADC enters the target cell, the linker needs to be effectively lysed to quickly release the drug load to function
.
After T-DXd enters the cell, it only needs to be recognized by lysosomal protease to recognize the linker cleavage site to release the drug load ^[14,15], which is simple and more
efficient.
This characteristic makes T-DXd a bystander effect, which is of great significance
for the treatment of gastric cancer, a highly heterogeneous tumor.
Preclinical studies have demonstrated that T-DXd significantly reduces the volume of mixed tumors (HER2-positive/HER2-negative) ^[16].

Figure 5.
Only T-DXd can effectively kill rodent mixed graft tumors, and significantly reduce the volume of mixed tumors ^[16].

The conjugation method used for the linker is also important and will determine the homogeneity and DAR
of the drug.
Natural conjugation methods include lysine coupling and cysteine coupling, and cysteine conjugation results in better
homogeneity of ADC drugs.
The homogeneity will directly affect the distribution and metabolism of ADC drugs in vivo, and high homogeneity will improve drug purity and benefit quality control ^[13,17].

On the basis of cysteine coupling, the coupling technology of T-DXd was further optimized to make it more homogeneous and reach the highest theoretical value of DAR ^8[13].

Figure 6.
Lysine is coupled to cysteine^[13,17].

The drug carrier DXd is a topoisomerase I inhibitor, which is different from the mechanism of chemotherapy drugs commonly used in gastric cancer such as microtubule inhibitors (taxane), DNA damage factors (platinum), and antimetabolites (fluorouracil) to avoid cross-resistance ^[18].

Secondly, DXd has 10 times the antitumor activity of its metabolite SN38 compared to irinotecan ^[13], while SN38 is 100-1000 times
more active than irinotecan.

The system half-life of free DXd is short, and T-DXd is conducive to control system toxicity after drug release ^[19].

Moreover, DXd's self-deamination design gives it strong cell membrane permeability ^[13], which is another condition for
T-DXd to produce a bystander effect.

Overall, thanks to the optimized linker technology, the homogeneity of T-DXd is greatly improved and the DAR reaches 8
.
Moreover, the efficient release of drug loading can be achieved by lysosomal protease cleavage linkers, and because the drug load has high cell membrane permeability, T-DXd has a strong bystander effect, showing great potential
in the treatment of gastric cancer with high heterogeneity.
The unique mechanism and excellent pharmacokinetic characteristics of drug-loaded DXd are also indispensable for
T-DXd to exert strong anti-tumor effects.
All in all, the systematic optimization of each key element of the ADC has played a huge role in improving the clinical efficacy of the ADC, and T-DXd has "emerged"
with technological advancements.