Anti-EGFR (Cetuximab) h(41BB-CD3ζ) CAR, pCDCAR1 vector

All products and services are For Research Use Only and CANNOT be used in the treatment or diagnosis of disease.

Details

Target
EGFR

Targeting Cell Type
T cell

Targeting Diseases
Human primary glioblastoma

Generation
Second

Vector Name
pCDCAR1

Vector Length
~8kb

Vector Type
Lentiviral

Receptor Construction
scFv-41BB-CD3ζ

Discription of Signaling Cassetes
41BB
CD137 (also known as 4-1BB) is a surface co-stimulatory glycoprotein originally described as present on activated T lymphocytes, which belongs to the tumor necrosis factor (TNF) receptor superfamily. It is expressed mainly on activated CD4+ and CD8+ T cells, and binds to a high-affinity ligand (4-1BBL) expressed on several antigen-presenting cells such as macrophages and activated B cells. On the basis of preclinical observation, this molecule can promote the persistence of antigen-specific and antigen-nonspecific chimeric antigen receptor T-cells to significantly increases antitumor activity.
CD3ζ
CD3ζ, also known as T-cell receptor zeta, which together with T-cell receptor and CD3γ, δ , ε chain, forms the TCR-CD3 complex. ζ was expressed independently from the complex. The zeta chain plays an important role in coupling antigen recognition to several intracellular signal-transduction pathways. CD3-zeta, which contains 3 ITAMs, is the most commonly used endodomain component of CARs. It transmits an activation signal to the T cell after antigen is bound. CD3-zeta may not provide a fully competent activation signal and additional co-stimulatory signaling is needed. For example, chimeric CD28 and OX40 can be used with CD3-zeta to transmit a proliferative/survival signal, or all three can be used together.

Target

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Published Data

CAR scFv data FCM

Fig.1 Surface expression of EGFR on osteosarcoma and Ewing's sarcoma cell lines was measured by flow cytometry using the anti-EGFR mAb cetuximab followed by the Alexa Fluor-647 goat anti-human IgG secondary antibody.

CAR Construction :

EGFR is expressed on the surface of sarcoma cell lines

Pahl, J. H., Ruslan, S. E. N., Buddingh, E. P., Santos, S. J., Szuhai, K., Serra, M., ... & Lankester, A. C. (2012). Anti-EGFR Antibody Cetuximab Enhances the Cytolytic Activity of Natural Killer Cells toward OsteosarcomaCetuximab Enhances NK Cell Killing of Sarcoma Cells. Clinical cancer research, 18(2), 432-441.

Complete CAR data FCM

Fig.2 HEK293T cells transfected with a CTL vector orthe two EGFR CAR expression vectors.

CAR Construction : Cetuximab scFv-28ζ

Fig.2 HEK293T cells transfected with a CTL vector orthe two EGFR CAR expression vectors.

Two EGFR CAR expression vectors as described inawere stained with an APC-conjugated anti-human IgG-Fc antibody or a FITC-conjugatedanti-human IgG (Fab)2 antibody followed by flow cytometry analysis.

Xia, L., Zheng, Z. Z., Liu, J. Y., Chen, Y. J., Ding, J. C., Xia, N. S., ... & Liu, W. (2020). EGFR‐targeted CAR‐T cells are potent and specific in suppressing triple‐negative breast cancer both in vitro and in vivo. Clinical & translational immunology, 9(5), e1135.

Complete CAR data FCM

Fig.3 CTL T or EGFR CAR-T cells were incubated with MDA-MB-231 or MDA-MB-468 cells and stained with CD8-APC and CD69-PE or CD25-FITC followed by flow cytometry analysis.

CAR Construction : Cetuximab scFv-28ζ

Fig.3 CTL T or EGFR CAR-T cells were incubated with MDA-MB-231 or MDA-MB-468 cells and stained with CD8-APC and CD69-PE or CD25-FITC followed by flow cytometry analysis.

Experiments were repeated three times, and representative histograms are shown.

Complete CAR data Cyt

Fig.4 EGFR CAR-T cells were incubated with MCF10A, MCF7, BT474, MDA-MB-468 or MDA-MB-231 cells at a ratio of 1:1 for the indicated durations followed by the cytotoxicity assay.

CAR Construction : Cetuximab scFv-28ζ

Fig.4 EGFR CAR-T cells were incubated with MCF10A, MCF7, BT474, MDA-MB-468 or MDA-MB-231 cells at a ratio of 1:1 for the indicated durations followed by the cytotoxicity assay.

Data were obtained from three replicates and are presented as means.e.m..

Complete CAR data FuncS

Fig.5 Tumor growth was monitored by using bioluminescence imaging for 20 days.

CAR Construction : Cetuximab scFv-28ζ

Fig.5 Tumor growth was monitored by using bioluminescence imaging for 20 days.

Representative data of two independentexperiments are shown.

Complete CAR data TCA

Fig.6 Primary T lymphocytes infected with EGFR CAR lentivirus (EGFR CAR-T) labelled with CFSE were incubated with or without MDA-MB-231 or MDA-MB-468 cells.

CAR Construction : Cetuximab scFv-28ζ

Fig.6 Primary T lymphocytes infected with EGFR CAR lentivirus (EGFR CAR-T) labelled with CFSE were incubated with or without MDA-MB-231 or MDA-MB-468 cells.

Experiments were repeated three times, and representative histograms are shown.

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