Anti-PD-L1 (Atezolizumab biosimilar - IgG1 isotype)

Human IgG1 monoclonal antibody (mAb) against PD-L1

Principle of PD-1/PD-L1 cellular assay
(click to enlarge)

Anti-hPD-L1-hIgG1 is a recombinant monoclonal antibody (mAb) featuring a variable region equivalent to atezolizumab, that recognizes human (h)PD-L1, and the constant region of the human (h)IgG1 isotype. In contrast to the N298A mutant, the 'wild-type' hIgG1 isotype is known to bind with high affinity to the Fc receptor on phagocytic cells. Therefore, Anti-hPD-L1-hIgG1 displays high effector function, including antibody‑dependent cell-mediated cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC). Anti-hPD-L1-hIgG1 was generated by recombinant DNA technology, has been produced in Chinese hamster ovary (CHO) cells, and purified by affinity chromatography with protein G.

PD-L1 background

Programmed cell death ligand 1 (PD-L1), also known as cluster of differentiation 274 (CD274) or B7 homolog 1 (B7-H1) is a transmembrane protein that can be constitutively expressed or induced in myeloid, lymphoid, and normal epithelial cells, as well as in cancer [1, 2]. PD-L1 is the principal ligand for programmed cell death protein 1 (PD-1) and under physiological conditions, this interaction is essential in the development of immune tolerance preventing excessive immune cell activity. However, PD-L1 expression is an immune evasion mechanism exploited by various malignancies and is generally associated with poorer prognosis [3]. Specifically, over-expressed PD-L1 on tumor cells and tumor-infiltrating immune cells, such as macrophages, can bind to PD-1 on cytotoxic T cells, and ultimately inhibit the anti-tumor T cell response [2, 4]. Thus, due to PD-L1’s instrumental role in immune evasion by cancer cells, there are numerous inhibitors in development as promising immuno-oncology therapies. Notably, Atezolizumab (also known as MPDL3280A), a fully humanized IgG1 (N298A) mAb that blocks the interaction of PD-L1 with PD-1 and induces anti-tumor immune reactivation, has been approved by the FDA for combinational use in the treatment of lung and breast cancer [2, 5].

Key features of Anti-hPD-L1-hIgG1:

• Clinically relevant variable region targeting PD-L1
• Features the 'wild-type' hIgG1 constant region for high effector function
• Functionally validated by flow cytometry

The terms “Atezolizumab” and “MPDL3280A” are only used as references. Anti-hPD-L1-hIgG1 is not a pharmaceutical biosimilar of Atezolizumab. It has not been developed nor approved by the Atezolizumab owner(s) and is not intended for any therapeutic or diagnostic use in humans or animals.

References

1. Juneja, V.R. et al. 2017. PD-L1 on tumor cells is sufficient for immune evasion in immunogenic tumors and inhibits CD8 T cell cytotoxicity. J Exp Med 214, 895-904.
2. Kythreotou, A. et al. 2018. PD-L1. J Clin Pathol 71, 189-194.
3. Sun, C. et al. 2018. Regulation and Function of the PD-L1 Checkpoint. Immunity 48, 434-452.
4. Lau, J. et al. 2017. Tumour and host cell PD-L1 is required to mediate suppression of anti-tumour immunity in mice. Nat Commun 8, 14572.
5. Heimes, A.S. & Schmidt, M. 2019. Atezolizumab for the treatment of triple-negative breast cancer. Expert Opin Investig Drugs 28, 1-5.

Figures

Comparison of ADCC potency for native and engineered anti-human PD-L1 antibody isotypes. Raji-hPD-L1 cells were incubated with gradient concentrations of Anti-hPD-L1 or Anti-β-galactosidase (β-gal) mAbs for 1 hour. Jurkat-Lucia™ NFAT-CD16 effector cells were then co-incubated with target cells for 6 hours. NFAT activation, reflecting the induced ADCC response, was assessed by determining Lucia luciferase activity in the supernatant using QUANTI-Luc™. Percentages of the maximal response normalized to the IgG1 isotype are shown.

Activation of Jurkat-Lucia™ TCR-hPD-1 cells.
Raji-APC-hPD-L1 and Jurkat-Lucia™ TCR-hPD-1 cells were incubated with gradient concentrations of AntihPD-1 hIgG1 (Ni: Nivolumab variable region; Pem: Pembrolizumab variable region) or Anti-hPD-L1 mAbs for 6 hours. NFAT activation, reflecting the disruption of PD-1/PD-L1 inhibitory interaction, was assessed by determining Lucia luciferase activity in the supernatant using QUANTI-Luc™.
The fold increase over non induced cells (no mAbs) is shown.

Specifications

Specificity: Targets cells expressing human or murine PD-L1

Clonality: Monoclonal antibody

Clone: Atezolizumab (anti-hPD-L1-hIgG1 (N298A), kappa)

Isotype: Human IgG1, kappa

Control: Human IgG1 control

Source: CHO cells

Formulation: 0.2 µm filtered solution in 68 mM sodium phosphate buffer (pH 7.4) with 91 mM glycine, 5% w/v saccharose and stabilizing agents

Purification: Purified by affinity chromatography with protein G

Tested applications: Flow cytometry, ADCC, Disruption of PD-1/PD-L1 inhibitory interaction

Quality Control:

• Binding of Anti-hPD-L1-hIgG1 to human PD-L1 has been validated using flow cytometry.
• The complete sequence of this antibody has been verified.
• The absence of bacterial contamination (e.g. lipoproteins and endotoxins) has been confirmed using HEK-Blue™ TLR2 and HEK-Blue™ TLR4 cells.

Contents

Anti-hPD-L1-hIgG1 purified monoclonal antibody is provided azide-free and lyophilized. It is available in two quantities:

• hpdl1-mab1: 100 µg
• hpdl1-mab1-03: 3 x 100 µg

Product is shipped at room temperature.

 Upon receipt, store at -20°C.