Use of a standardized MxA protein measurement-based assay for validation of assays for the assessment of neutralizing antibodies against interferon-ß.

Effective monitoring of the development of neutralizing antibodies (NAbs) against IFN-ß in multiple sclerosis (MS) patients on IFN-ß therapy is important for clinical decision making and disease management. To date, antiviral assays have been the favored approach for NAb determination, but variations in assay conditions between laboratories and the increasing use of novel assays have contributed to the reporting of inconsistent antibody data between laboratories and between products. This study, undertaken at the request of the Committee for Medicinal Products for Human Use (CHMP) of the European Medicines Agency (EMA), is a joint effort by manufacturers of IFN-ß products (approved in Europe) towards harmonization of a NAb assay that facilitates generation of comparable NAb data, which, in conjunction with clinical outcomes, should prove useful for clinicians treating MS patients with IFN-ß products. This article describes the standardized cellular myxovirus resistance protein A (MxA) protein measurement-based assay for detection of IFN-ß NAbs and its use for the validation of assays used for the quantitative determination of such antibodies. Although titers varied between laboratories and the products used, utilization of IFN-ß1a rather than IFN-ß1b as the challenge antigen produced more consistent results in the NAb assay. Adoption of the standardized assay improves comparability between laboratories circumventing problems that arise when different, nonstandardized assays are employed for immunogenicity assessment. Based on the data, the EMA recommended for standardization purposes, the use of IFN-ß1a in NAb assays, independent of the therapeutic product used for therapy and validation of new NAb procedures against the standardized assay described.

Recommendations for the design, optimization, and qualification of cell-based assays used for the detection of neutralizing antibody responses elicited to biological therapeutics.

The administration of biological therapeutics can evoke some level of immune response to the drug product in the receiving subjects. An immune response comprised of neutralizing antibodies can lead to loss of efficacy or potentially more serious clinical sequelae. Therefore, it is important to monitor the immunogenicity of biological therapeutics throughout the drug product development cycle. Immunoassays are typically used to screen for the presence and development of anti-drug product antibodies. However, in-vitro cell-based assays prove extremely useful for the characterization of immunoassay-positive samples to determine if the detected antibodies have neutralizing properties. This document provides scientific recommendations based on the experience of the authors for the development of cell-based assays for the detection of neutralizing antibodies in non-clinical and clinical studies.

A multicenter, open-label, phase II study of the immunogenicity and safety of a new prefilled syringe (liquid) formulation of Avonex in patients with multiple sclerosis.

BACKGROUND: A new liquid formulation of Avonex (interferon beta-1a [IFNbeta-1a]) in a prefilled syringe has been developed to make administration of the drug easier for patients with multiple sclerosis (MS). This formulation does not contain human serum albumin (HSA), often added to interferon (IFN) products for stabilization. However, formulation changes may alter the secondary, tertiary, and quaternary structures of IFNbeta products. These kinds of structural changes could lead to the formation of antibodies directed against IFNbeta. Some of these anti-IFN antibodies may neutralize the biologic activity of IFNbeta. OBJECTIVE: This study was designed to determine the immunogenicity and safety of the new prefilled syringe (liquid) HSA-free formulation of Avonex in patients with relapsing MS. METHODS: This was a multicenter, single-arm, open-label study. Patients with relapsing MS received liquid, HSA-free Avonex 30 microg by IM injection from a prefilled syringe once weekly for up to 24 months. Immunogenicity and safety were assessed every 3 months. Serum levels of neutralizing antibodies (NAbs) were measured at baseline and every 3 months using a 2-step enzyme-linked immunosorbent assay and antiviral cytopathic effect assay. RESULTS: A total of 153 patients (121 women, 32 men; mean [SD] age, 39.6 [9.9] years; age range, 19.0-59.0 years) were enrolled in the study. Sera were available for analysis from 125 and 119 patients after 18 and 24 months of treatment, respectively. By 18 months, 1 patient (1%) had > or =2 consecutive titers of > or =20, a level at which the persistent presence of NAbs has been shown in some studies to have clinical consequences. By 24 months, 1 additional patient (total 2%) had > or =2 consecutive titers of > or =20. At 18 months, 5 patients (4%) had > or =1 NAb titer of > or =5; at 24 months, 6 patients (5%) had > or =1 NAb titer of > or =5. The safety profile of liquid Avonex was comparable to the lyophilized form containing HSA. CONCLUSIONS: The prefilled syringe (liquid) HSA-free formulation of Avonex was well tolerated and showed a low level of immunogenicity. Over 24 months, 2% of patients developed persistent NAbs (> or =2 consecutive titers of > or =20).