CD3 proteins: Targets for bispecific antibodies

A bispecific antibody (BsAb) is a type of artificial protein with the ability to bind to two distinct antigens or two epitopes of one antigen at the same time. Bispecific antibodies can assume a synergistic role of the two monoclonal antibodies and have benefits over the simple combination.

Due to its special biological mechanism, the bispecific antibody has turned out to be an essential branch of therapeutic antibody research and development and has been drawing investments. As per statistics, there are over 180 bispecific antibodies in clinical trials and nearly 260 in pre-clinical development around the world. Among them, CD3 is considered to be the primary target.

CD3 proteins: Targets for bispecific antibodies

Image Credit: ACROBiosystems

CD3 molecules tend to bind to TCR non-covalently to develop a TCR/CD3 receptor complex on the T-cell’s surface. There are four subtypes of CD3 — CD3ε, CD3δ, CD3γ and CD3ζ. CD3δ/CD3ε and CD3γ/CD3ε have the ability to develop the TCR-CD3 complexes through the α/β chain of TCR as heterodimers.

Generally, CD3-targeted therapeutic antibodies recognize the CD3ε in the heterodimer complex to initiate the anti-tumor activity of T cells. But during recombinant expression, CD3E and CD3D have the ability to randomly form faulty heterogeneous complexes, which might result in the loss of bioactivity and huge batch-to-batch inconsistency.

ACROBiosystems has designed a range of homogeneous CD3δ/CD3ε and CD3γ/CD3ε proteins with good bioactivity. They are confirmed as 1:1 heterodimers by non-reductive electrophoresis and MALS as well.

In addition, they exhibit superior performance in therapeutic antibody screening, identification, characterization as well as clinical pharmacokinetic assays, etc. Furthermore, the CD3 products have been successfully utilized for the clinical development of a bispecific antibody. This greatly expedited the process of therapeutic antibody development.

Product features

  • MALS verified heterodimer
  • Bioactivity confirmed by a clinical bispecific antibody
  • Various tags: his, his&Avi, Fc tags, etc.
  • High binding ability and low batch-to-batch variations

Product list

CD3E and CD3D

Table 1. Source: ACROBiosystems

Molecule Cat. No. Species Product Description Structure
CD3E & CD3D CDD-H5255 HEK293 Human CD3 epsilon&CD3 delta Heterodimer Protein, Fc Tag
&Fc Tag (MALS verified)New
CDD-H52Wa HEK293 Human CD3 epsilon&CD3 delta Heterodimer Protein, Fc, His Tag&Fc, Flag Tag (MALS verified)Hot
CDD-H82W6 HEK293 Biotinylated Human CD3 epsilon&CD3 delta Heterodimer Protein, His, Avitag™&Tag Free (MALS verified)Hot
CDD-H52W1 HEK293 Human CD3 epsilon & CD3 delta Heterodimer Protein, His Tag&Tag Free (MALS verified)Hot

 

Click here to learn about many more CD3E and CD3Ds.

CD3E and CD3G

Table 2. Source: ACROBiosystems

Molecule Cat. No. Host Product Description Structure
CD3E & CD3G CDG-H5253 HEK293 Human CD3 epsilon&CD3 gamma Heterodimer Protein, Fc Tag
&Fc Tag (MALS verified)New
CDG-H52W9 HEK293 Human CD3 epsilon&CD3 gamma Heterodimer Protein, Fc, His Tag&Fc, Flag Tag (MALS verified)Hot
CDG-H52W5 HEK293 Human CD3 epsilon&CD3 gamma Heterodimer Protein, His Tag&Tag Free (MALS verified)
CDG-C5257 HEK293 Cynomolgus CD3E&CD3G Heterodimer Protein, Fc Tag&Fc Tag (MALS verified)

 

Click here to learn about many more CD3E and CD3Gs.

CD3 epsilon

Table 3. Source: ACROBiosystems

Molecule Cat. No. Host Product Description Structure
CD3
epsilon
CDE-M525a HEK293 Biotinylated Monoclonal Anti-Human CD3 Antibody, Mouse IgG2a, Avitag™ (Clone: OKT3), Ultra-low endotoxin  
CDE-H5223 HEK293 Human CD3 epsilon Protein, His Tag (MALS verified)Hot
CDE-H5256 HEK293 Human CD3 epsilon Protein, Fc, His TagHot
CDE-C5226 HEK293 Cynomolgus CD3 epsilon Protein, His Tag (MALS verified)Hot

 

Click here to learn about many more CD3 epsilons.

CD3 delta

Table 4. Source: ACROBiosystems

Molecule Cat. No. Host Product Description Structure
CD3 delta CDD-H5224 HEK293 Human CD3 delta Protein, His Tag
CDD-C52H8 HEK293 Cynomolgus CD3 delta Protein, His Tag

 

CD3 gamma

Table 5. Source: ACROBiosystems

Molecule Cat. No. Host Product Description Structure
CD3
gamma
CDA-H5254 HEK293 Human CD3 gamma / CD3G Protein, Fc Tag (MALS verified)

 

More BsAb targets

Table 6. Source: ACROBiosystems

.
CD19 CD20 CD38 CD40 CD47
IL-3 R alpha Her2 ErbB3 LAG-3 4-1BB
PD-1 PD-L1 CTLA-4 TIM-3 EpCAM
EGF R BCMA PSMA HGF R ROR1
VEGF R2 FAP IL-1 beta IL-4 IL-6 R alpha
IL-13 IL-17A CD16 TNF-alpha ANGTP2
OX40 ICOS HGF B7-H3 TGF-beta 1
LRP-5 LRP-6 CEACAM-5 Mucin-1 DLL3

 

Data

1:1 heterodimer verified by nonreductive electrophoresis and MALS

ACROBiosystems has come up with a range of homogeneous CD3 proteins with ideal bioactivity, which is confirmed as 1:1 heterodimer by nonreductive electrophoresis and MALS.

Human CD3E&CD3D Heterodimer Protein (Cat. No. CDD-H52Wa) on SDS-PAGE under reducing (R) and non-reducing (NR) condition and the purity of the protein is greater than 95%. The purity of the protein is more than 85% and around 80–90 kDa verified by SEC-MALS.

Figure 1. Human CD3E&CD3D Heterodimer Protein (Cat. No. CDD-H52Wa) on SDS-PAGE under reducing (R) and non-reducing (NR) condition and the purity of the protein is greater than 95%. The purity of the protein is more than 85% and around 80–90 kDa verified by SEC-MALS. Image Credit: ACROBiosystems

High binding ability and low batch-to-batch variations

Such validated authentic 1:1 heterodimer proteins exhibit high bioactivity in various applications. The products are homogenous with rigorous quality control to ensure the least batch-to-batch variations.

Immobilized Biotinylated Human CD3E&CD3D Heterodimer Protein, His, Avitag&Tag Free (Cat. No. CDD-H82W6) at 1 μg/mL (100 μL/well) on Streptavidin (Cat. No. STN-N5116) precoated (0.5 μg/well) plate, can bind Monoclonal Anti-Human CD3 Antibody, Mouse IgG2a (Cat. No. CDE-M120a) with a linear range of 0.2–6 ng/mL (QC tested), and batch differences EC50 < 0.0001 μg/mL.

Figure 2. Immobilized Biotinylated Human CD3E&CD3D Heterodimer Protein, His, Avitag&Tag Free (Cat. No. CDD-H82W6) at 1 μg/mL (100 μL/well) on Streptavidin (Cat. No. STN-N5116) precoated (0.5 μg/well) plate, can bind Monoclonal Anti-Human CD3 Antibody, Mouse IgG2a (Cat. No. CDE-M120a) with a linear range of 0.2–6 ng/mL (QC tested), and batch differences EC50 < 0.0001 μg/mL. Image Credit: ACROBiosystems

Designed for different applications of drug development

Such products are appropriate for several applications at various stages of drug development, like antibody immunotiter detection, antibody screening, biopanning, affinity characterization and clinical pharmacokinetic assays.

Application 1: Development and screening of antibody drugs

Immobilized Biotinylated Human CD3E&CD3D Heterodimer Protein, His, Avitag&Tag Free (Cat. No. CDD-H82W6) at 1 μg/mL (100 μL/well) on streptavidin precoated (0.5 μg/well) plate, can bind Monoclonal Anti-Human CD3 Antibody, Mouse IgG2a (Cat. No. CDE-M120a) with a linear range of 0.2–6 ng/mL.

Figure 3. Immobilized Biotinylated Human CD3E&CD3D Heterodimer Protein, His, Avitag&Tag Free (Cat. No. CDD-H82W6) at 1 μg/mL (100 μL/well) on streptavidin precoated (0.5 μg/well) plate, can bind Monoclonal Anti-Human CD3 Antibody, Mouse IgG2a (Cat. No. CDE-M120a) with a linear range of 0.2–6 ng/mL. Image Credit: ACROBiosystems

Immobilized Human CD3E&CD3D Heterodimer Protein, His Tag&Tag Free (Cat. No. CDD-H52W1) at 1 μg/mL (100 μL/well) can bind Anti-CD3 antibody with a linear range of 0.2–6 ng/mL.

Figure 4. Immobilized Human CD3E&CD3D Heterodimer Protein, His Tag&Tag Free (Cat. No. CDD-H52W1) at 1 μg/mL (100 μL/well) can bind Anti-CD3 antibody with a linear range of 0.2–6 ng/mL. Image Credit: ACROBiosystems

Application 2: Identification and quality control of antibody drugs

Immobilized Human CD3E&CD3D Heterodimer Protein, His Tag&Tag Free (Cat. No. CDD-H52W1) at 1 μg/mL (100 μL/well) can bind Monoclonal Anti-Human CD3 Antibody, Mouse IgG2a (Cat. No. CDE-M120a) with a linear range of 0.2–6 ng/mL.

Figure 5. Immobilized Human CD3E&CD3D Heterodimer Protein, His Tag&Tag Free (Cat. No. CDD-H52W1) at 1 μg/mL (100 μL/well) can bind Monoclonal Anti-Human CD3 Antibody, Mouse IgG2a (Cat. No. CDE-M120a) with a linear range of 0.2–6 ng/mL. Image Credit: ACROBiosystems

Immobilized Human CD3E&CD3D Heterodimer Protein, Fc Tag&Fc Tag (Cat. No. CDD-H5255) at 1 μg/mL (100 μL/well) can bind Bispecific BCMA×CD3 T cell-engaging Antibody with a linear range of 0.8–6 ng/mL.

Figure 6. Immobilized Human CD3E&CD3D Heterodimer Protein, Fc Tag&Fc Tag (Cat. No. CDD-H5255) at 1 μg/mL (100 μL/well) can bind Bispecific BCMA×CD3 T cell-engaging Antibody with a linear range of 0.8–6 ng/mL. Image Credit: ACROBiosystems

Biotinylated Human CD3E&CD3D Heterodimer Protein, His, Avitag&Tag Free (Cat. No. CDD-H82W6) captured on Biotin CAP - Series S sensor Chip can bind Anti-Human CD3 Antibody, Mouse IgG2a with an affinity constant of 22.5 nM as determined in an SPR assay (Biacore T200).

Figure 7. Biotinylated Human CD3E&CD3D Heterodimer Protein, His, Avitag&Tag Free (Cat. No. CDD-H82W6) captured on Biotin CAP - Series S sensor Chip can bind Anti-Human CD3 Antibody, Mouse IgG2a with an affinity constant of 22.5 nM as determined in an SPR assay (Biacore T200). Image Credit: ACROBiosystems

Bispecific T-cell Engager (CD3 X BCMA) immobilized on CM5 Chip can bind Human CD3E&CD3D Heterodimer Protein, His Tag&Tag Free (Cat. No. CDD-H52W1) with an affinity constant of 31.8 nM as determined in an SPR assay (Biacore T200).

Figure 8. Bispecific T-cell Engager (CD3 X BCMA) immobilized on CM5 Chip can bind Human CD3E&CD3D Heterodimer Protein, His Tag&Tag Free (Cat. No. CDD-H52W1) with an affinity constant of 31.8 nM as determined in an SPR assay (Biacore T200). Image Credit: ACROBiosystems

Application 3: Clinical PK assay

Immobilized Human BCMA, His Tag (Cat. No. BCA-H522y) at 2 μg/mL, add increasing concentrations of Bispecific T cell Engager (CD3 X BCMA) in 10% human serum and then add Biotinylated Human CD3E&CD3D Heterodimer Protein, His, Avitag&Tag Free (Cat. No. CDD-H82W6) at 0.2 μg/mL. Detection was performed using HRP-conjugated streptavidin with a sensitivity of 15 ng/mL.

Figure 9. Immobilized Human BCMA, His Tag (Cat. No. BCA-H522y) at 2 μg/mL, add increasing concentrations of Bispecific T cell Engager (CD3 X BCMA) in 10% human serum and then add Biotinylated Human CD3E&CD3D Heterodimer Protein, His, Avitag&Tag Free (Cat. No. CDD-H82W6) at 0.2 μg/mL. Detection was performed using HRP-conjugated streptavidin with a sensitivity of 15 ng/mL. Image Credit: ACROBiosystems