Arthritis & Rheumatism, Volume 60,
October 2009 Abstract Supplement
The 2009 ACR/ARHP Annual Scientific Meeting
Philadelphia October 16-21, 2009.
The PEG Component of Certolizumab Pegol Inhibits Degranulation by Stimulated Mast Cells
Lamour, Sabrina, Bracher, Marguerite, Nesbitt, Andrew
Purpose:
The administration of some conventional injectable TNF inhibitors is associated with severe injection-site pain (ISP), including stinging and burning. ISP may be linked to the inflammatory mediators released upon degranulation of mast cells, which are highly responsive skin cells that can rapidly secrete an array of inflammatory mediators. In contrast to conventional TNF inhibitors, the anti-TNF certolizumab pegol (CZP) lacks an Fc region and instead consists of a Fab' molecule site-specifically linked to a 40 kDa PEG moiety. A low incidence of ISP has been observed in patients receiving CZP in clinical trials in patients with rheumatoid arthritis.(1–3) The objective of our investigations was to determine if the PEG moiety of CZP inhibits the non-immune–stimulated degranulation of mast cells and may be responsible for the low incidence of ISP associated with CZP.
Methods:
Mast cells were cultured in vitro from stem cells over an 8–12 week period using the method of Saito et al.(4) The development of stem cells into mast cells was confirmed by identification of mast cell markers, including CD117, CD203c and CD32, using flow cytometry. Mast cell degranulation, as measured by b hexosaminidase release, was stimulated by the addition of compound 48/80, a known non-immune activator of mast cells, which causes degranulation. Titrations of CZP, PEG, and a mixture of PEG and naked Fab' at a PEG concentration of 45mg/mL were incubated with the mast cells and a fixed amount of compound 48/80 to determine the effect on mast cell degranulation. Mast cell viability at the end of the experiment was assessed using the Promega Cell titer 96 Aqueous One Solution cell proliferation assay.
Results:
Compound 48/80 stimulated mast cell degranulation as measured by b hexosaminidase release, although the absolute level varied between cell preparations. PEG (45 mg/mL) inhibited mast cell degranulation stimulated by 20mM and 200mM compound 48/80 by 66% (n=3) and 57.5% (n=4) respectively (P< 0.001). CZP (100 mg/mL), PEG alone (45mg/mL) and the mixture of PEG (19.8mg/mL) and naked Fab' (23.9 mg/mL) all inhibited the majority of mast cell degranulation. None of the reagents affected overall cell viability.
Conclusion:
PEG inhibited compound 48/80–stimulated degranulation of mast cells. The concentrations at which an effect is observed are what might be expected at the injection site but not systemically. This beneficial effect of PEG on mast cells may explain the low level of ISP observed with CZP in clinical trials. However, the exact mechanism behind this activity is unclear and warrants further investigation.
1.Keystone, EC, et al. Arthritis Rheum 2008;58:3319-3329.
2.Smolen, J, et al. Ann Rheum Dis 2009;69:797-804.
3.Fleischman, R, et al. Ann Rheum Dis 2009;69: 805-811.
4.Saito, H, et al. Nature Protocols 2006;1:2178-2183.
To cite this abstract, please use the following information:
Lamour, Sabrina, Bracher, Marguerite, Nesbitt, Andrew; The PEG Component of Certolizumab Pegol Inhibits Degranulation by Stimulated Mast Cells [abstract]. Arthritis Rheum 2009;60 Suppl 10 :44
DOI: 10.1002/art.25127
