A novel immunoassay strategy to measure total serum lymphotoxin-α levels in cynomolgus monkeys dosed with an anti-LTα antibody (62.5)

Abstract

Abstract Lymphotoxin α (LTα) is a TNF superfamily cytokine secreted from activated lymphocytes as a homotrimer (LTα3) or complexed on the cell-surface with LTβ as heterotrimers. Compelling evidence indicates that LTα is involved in the pathophysiology of autoimmune diseases. Given that blocking the activity of LTα holds promise as a treatment option, a neutralizing monoclonal antibody (mAb) is under clinical development as a potential therapeutic. In preclinical studies of mAb therapeutics, measuring the level of the target antigen is often of interest as a pharmacodynamic marker. We found that the presence of anti-LTα caused a dose-dependent decrease in ability to detect LTα by immunoassay due to the inability of detection antibodies to bind in the presence of bound therapeutic mAb. To circumvent this problem, an alternate assay strategy was devised to accurately measure total LTα3 (free LTα and LTα/anti-LTα complexes). Specifically, excess anti-LTα was added to fully saturate all binding sites on LTα3. Bound anti-LTα was detected using an anti-idiotype mAb. Using this approach, total LTα was measured in serum of cynomolgus monkeys dosed with anti-LTα. Circulating LTα concentrations showed dose related increases up to 8,000-fold that correlated with anti-LTα PK. This assay strategy may be useful in other situations where low molecular weight and/or multimeric structure of a target antigen makes accurate detection in the presence of therapeutic mAb problematic. KEY WORDS LTα mAb PD