Comparison of neutralizing antibody assays for receptor binding and enzyme activity of the enzyme replacement therapeutic Naglazyme (galsulfase).

Most patients receiving Naglazyme (galsulfase, rhASB) enzyme replacement therapy for mucopolysaccharidosis type VI develop an antibody response. To evaluate the impact of this response, two in vitro neutralizing antibody (NAb) assays were developed based on the two steps of the mechanism of action. Neutralization of enzyme activity was detected by inhibition of rhASB cleavage of a fluorogenic substrate. Neutralization of receptor binding was detected by decreased binding of labeled rhASB to immobilized soluble receptor. For the enzyme activity NAb assay, serum pretreatment was required to isolate antibodies from interfering phosphate ions, with sensitivity of < or =5 microg/mL. The receptor binding NAb assay used a five-fold dilution, with sensitivity of < or =40 microg/mL. Cutpoints for percent inhibition were based on 95% confidence intervals from naïve sera. Clinical samples were similarly likely to be positive in both assays than positive for neutralization of only one step in the mechanism of action. The two NAb assays yielded complementary information about potential neutralization of rhASB. Relative estimated sensitivity between neutralization assays did not correlate with the number of positive clinical samples or patients. In vitro NAb assays based on a well-understood mechanism of action provide specific information about the NAb mechanism.