Nonneutralizing Factor VIII Antibodies as a Marker for Inhibitor Development in a Longitudinal Cohort of Hemophilia A Patients

The development of neutralizing antibodies to FVIII (Nabs) is still the most severe complication in modern hemophilia A treatment. In clinical routine, Nabs are determined by Bethesda assay, which detects Nabs, but not nonneutralizing antibodies to FVIII (NNAs). The primary objective of this study was the characterization anti-FVIII drugs antibodies (ADA) development, ADA function, and signature in correlation to treatment outcome in longitudinal samples of previously untreated patients.

Validated FVIII-ADA assays for the detection of ADA, Nabs, and IgG subclasses (IgG1, IgG2, IgG3, and IgG4) were used in combination with assays to identify the binding domains on FVIII (human porcine FVIII constructs, FVIII heavy and light chain, and individually expressed FVIII domains). A total of 21 previously untreated patients were analyzed for >50 exposure days.

In this retrospective longitudinal analysis, 10 patients had developed Nabs. For 6 with Nabs, NNAs were detected up to 150 days before Nabs. Two out of 11 Nab-negative patients showed ADAs in screening, but were not confirmed in competition tests. IgG subclass characterization showed a broad response in the beginning including IgG3, with a change to IgG1 and IgG4 only. Epitopes on FVIII were localized on FVIII heavy and light chain including domains A2, C1, and C2 with a change to A2 with Nab development.

For most patients, the development of NNAs preceded the detection of Nabs. None of the patients who did not develop Nabs showed confirmed ADAs. The relevance of ADA detection before Nabs development and possible clinical interventions needs to be assessed in clinical trials.

Publication History

Article published online:
13 November 2020