Anti-EGFRvIII (MAb 139) 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
EGFRvIII

Targeting Cell Type
T Cell

Targeting Diseases
Glioma

Generation
Second

Vector Name
pCDCAR1

Vector Length
~8kb

Vector Type
Lentiviral vector

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

Complete CAR data FCM

Fig.1 EGFRvIII mCAR retroviral gene delivery to murine T cells confers antigen-specific activity.

CAR Construction : MAb 139 scFv-28ζ

Fig.1 EGFRvIII mCAR retroviral gene delivery to murine T cells confers antigen-specific activity.

VM/Dk-derived glioma SMA560vIII shows EGFRvIII expression compared with SMA560 by surface antibody staining (L8A4) and flow cytometry.

Sampson, J. H., Choi, B. D., Sanchez-Perez, L., Suryadevara, C. M., Snyder, D. J., Flores, C. T., ... & Johnson, L. A. (2014). EGFRvIII mCAR-Modified T-Cell Therapy Cures Mice with Established Intracerebral Glioma and Generates Host Immunity against Tumor-Antigen LossEGFRvIII mCARs for Malignant Glioma. Clinical Cancer Research, 20(4), 972-984.

Complete CAR data FuncS

Fig.2 EGFRvIII mCAR T-cell therapy against intracerebral tumors requires lymphodepletive host-conditioning for efficacy, and can be abrogated by soluble peptide administration.

CAR Construction : MAb 139 scFv-28ζ

Fig.2 EGFRvIII mCAR T-cell therapy against intracerebral tumors requires lymphodepletive host-conditioning for efficacy, and can be abrogated by soluble peptide administration.

To evaluate the impact of lymphopenia on the antitumor efficacy of EGFRvIII mCAR T cells, VM/Dk mice (8 per group) with 3-day established intracerebral SMA560vIII tumors.

Complete CAR data FCM

Fig.3 Treatment of mice bearing EGFRvIIIPOS tumors with mCARs provides long-term protection against EGFRvIIINEG tumor

CAR Construction : MAb 139 scFv-28ζ

Fig.3 Treatment of mice bearing EGFRvIIIPOS tumors with mCARs provides long-term protection against EGFRvIIINEG tumor

SMA560 (EGFRvIIINEG) gliomas were maintained in vitro or passaged in a mouse subcutaneously for 14 days, then harvested and both were stained for EGFRvIII expression using L8A4 mAb and analyzed by flow cytometry.

Complete CAR data FCM

Fig.4 Development of CARs targeting EGFRvIII. NIH-3T3 cells and BHK cells were transduced with a retroviral vector expressing EGFRvIII and a neomycin resistance gene.

CAR Construction : MAb 139 scFv-28ζ

Fig.4 Development of CARs targeting EGFRvIII. NIH-3T3 cells and BHK cells were transduced with a retroviral vector expressing EGFRvIII and a neomycin resistance gene.

Based on the amino acid sequence for anti-EGFRvIII mAbs, scFv genes were synthesized and fused to T-cell signaling domains from CD28 and CD3f followed by insertion into c-retroviral vector MSGV1 (diagram).

Morgan, R. A., Johnson, L. A., Davis, J. L., Zheng, Z., Woolard, K. D., Reap, E. A., ... & Rosenberg, S. A. (2012). Recognition of glioma stem cells by genetically modified T cells targeting EGFRvIII and development of adoptive cell therapy for glioma. Human gene therapy, 23(10), 1043-1053.

Complete CAR data ELISA

Fig.5 Equivalent function of second- and third-generation anti-EGFRvIII CAR vectors.

CAR Construction : MAb 139 scFv-28ζ

Fig.5 Equivalent function of second- and third-generation anti-EGFRvIII CAR vectors.

Using anti-EGFRvIII human mAb 139, a third-generation CAR vector was assembled using CD28, 4-1BB, and CD3f T-cell signaling domains.

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For research use only. Not intended for any clinical use. No products from Creative Biolabs may be resold, modified for resale or used to manufacture commercial products without prior written approval from Creative Biolabs.