Human RANKL Antibody - Denosumab Biosimilar

Anti-human RANKL (TNFSF11) - Denosumab biosimilar - CAS #615258-40-7

Anti-hRANKL-hIgG2 is a biosimilar antibody of Denosumab, a therapeutic antibody that targets the human RANKL (receptor activator of nuclear factor-κB ligand, or TNFSF11). This monoclonal antibody (mAb) blocks the interaction of RANKL with its receptor. The FDA has approved Denosumab for the treatment of osteoporosis and bone metastasis.

 More details

Anti-hRANKL-hIgG2 comprises the variable region of Denosumab and the IgG2 constant region of Denosumab with high effector functions. 

This antibody can be used with HEK-Blue™ RANKL cells for screening and neutralization assays to block recombinant human RANKL-induced signaling (see figure).

Key features

• Each lot is functionally tested and validated.
• The complete sequence of the antibody construct was verified.
• The absence of endotoxins is determined using the EndotoxDetect™ assay.

All InvivoGen products are for internal research use only, and not for human or veterinary use.

Figures

Binding of Anti-hRANKL-hIgG2 mAb to coated human RANKL protein. Human RANKL (1 µg/ml) was coated on ELISA plates overnight. A 3-fold serial dilution of the  Anti-hRANKL-hIgG2 mAb (red curve) or the Anti-β-Gal-hIgG2 control antibody (grey curve) was performed for the capture step. An HRP-labelled anti-hIgG antibody (1/1000 dilution) and the HRP substrate OPD (o-phenylenediamine dihydrochloride) were used for the detection step. Absorbance was read at 490 nm.

Dose-dependent inhibition of HEK-Blue™ RANKL cells response using denosumab biosimilar. A serial dilution of denosumab, a biosimilar Anti-hRANKL-hIgG2 monoclonal antibody (mAb) was incubated with 10 ng/ml of recombinant human RANKL for 2 hours prior to the addition of the HEK-Blue™ RANKL cells. After overnight incubation, the NF-kB response was determined using QUANTI-Blue™ Solution, a SEAP detection reagent. Data are presented as a percentage of activity.

Specifications

Application: Neutralization assay, ELISA

Isotype:  Human IgG2, kappa

Recommended isotype control:Human IgG2

Target: Human receptor activator of nuclear factor-κB ligand (RANKL, TNFSF11, CD254)

Species reactivity: Human

Clone: Denosumab, AMG-162

Cas number: 615258-40-7

Source: CHO cells 

Production: Animal-free

Purification: Protein A

Molecular weight: ~ 150 kDa

Physical form: Lyophilized

Formulation buffer: Sodium phosphate buffer with glycine, saccharose, and stabilizing agents

Preservative: Azide-free

Reconstitution buffer: Sterile water (not provided)

Purity: ≥ 95 %

Quality control: Each lot is functionally tested and validated.

Contents

Anti-hRANKL-hIgG2 purified monoclonal antibody is provided azide-free and lyophilized. It is available in two quantities:

• hrankl-mab2: 100 µg
• hrankl-mab2-03: 3 x 100 µg

 The product is shipped at room temperature.

Upon receipt, store lyophilized antibody at -20°C.

 Lyophilized product is stable for at least 1 year.

 Avoid repeated freeze-thaw cycles.

Details

Denosumab background

Denosumab is a therapeutic, fully human IgG2 mAb that blocks the interaction of human (h)RANKL with its receptor RANK and thus the downstream signaling. It targets the transmembrane and soluble forms of the human RANKL (receptor activator of nuclear factor-κB ligand, also known as member 11 of the tumor necrosis factor ligand superfamily (TNFSF11)) [1].  RANKL binding to its receptor RANK induces signaling in osteoclasts and is responsible for bone resorption. RANKL-RANK also plays a pivotal role in dendritic cell maturation and T-cell differentiation [2].

Human IgG2 is the second most common antibody present in serum. It is resistant to cleavage by proteolytic enzymes, due to a short hinge region. This isotype displays low complement-dependent cytotoxicity (CDC), very low antibody-dependent cellular phagocytosis (ADCP), and very low antibody-dependent cellular cytotoxicity (ADCC).

Denosumab has been approved by the FDA for the treatment of osteoporosis and bone metastasis [3].

References:

1. Ahern E. et al., 2018. Roles of the RANKL-RANK axis in antitumour immunity -implications for therapy. Nat Rev Clin Oncol. 15(11):676-693.
2. Cheng ML. & Fong L., 2014. Effects of RANKL-Targeted Therapy in Immunity and Cancer. Front Oncol. 3:329.
3. Zaheer S, et al., 2015. Denosumab for the treatment of osteoporosis. Expert Opin Drug Metab Toxicol. 2015 Mar;11(3):461-70. doi: 10.1517/17425255.2015.1000860.