BAX 855 is a PEGylated recombinant factor VIII preparation that showed prolonged circulatory half-life in nonclinical and clinical studies. This paper describes the development, validation, and application of a novel ligand-binding assay (LBA) to selectively measure BAX 855 in plasma. The LBA is based on PEG-specific capture of BAX 855, followed by immunological factor VIII (FVIII)-specific detection of the antibody-bound BAX 855. This assay principle enabled sensitive measurement of BAX 855 down to the low nanomolar range without interference from non-PEGylated FVIII as demonstrated by validation data for plasma from animals typically used for nonclinical characterization of FVIII. The selectivity of an in-house-developed anti-PEG and a commercially available preparation, shown by competition studies to primarily target the terminating methoxy group of PEG, also allowed assessment of the intactness of the attached PEG chains. Altogether, this new LBA adds to the group of methods to selectively, accurately, and precisely measure a PEGylated drug in complex biological matrices. The feasibility and convenience of using this method was demonstrated during extensive nonclinical characterization of BAX 855.

Screening with T cell receptor incision circles (TREC) assay identifies newborns with severe T cell deficiency, including Severe Combined Immune Deficiency syndromes (SCID) and other causes. METHODS: A 3 week old Caucasian female born at 37 weeks gestation by vaginal delivery to a 28 year old mother with a history of myasthenia gravis post thyroidectomy requiring azathioprine 100 mg daily, prednisone 10 mg daily, and omeprazole during the pregnancy and with breastfeeding. Birth weight 2669 grams. She had a hip click, but did not have cardiac or any other malformation. Newborn SCID screen was positive. Breastfeeding was stopped to prevent further immunosuppression. RESULTS: At 3 weeks old she had very low total white blood cell count (0.8 TH/cmm), severe neutropenia (absolute 0.03 TH/cmm), severe lymphopenia (absolute 0.35 TH/cmm), anemia (hemoglobin 7.9 g/dL), and borderline low platelets (277 TH/cmm). Lymphocyte subsets showed profoundly low T cells (absolute CD3 206 cells/uL, absolute CD4 177 cells/uL, absolute CD8 83 cells/uL), no B cells (CD19 <20 cells/uL), and normal NK cells (171 cells/ uL). CBC and lymphocyte subsets were repeated weekly for one month and then monthly. At 3 months old, her total white blood cell count was 5.90 TH/ cmm, neutrophils 1.06 TH/cmm, and lymphocytes 3.95 TH/cmm. CD4 cells were normal (1540 cells/uL), and CD8 cells were moderately low (289 cells/ uL). CD19 B cells were normal (975 cells/uL). She remained healthy. She tolerated killed vaccines but has not had live viral vaccines. CONCLUSIONS: Taking azathioprine with prednisone during pregnancy may cause secondary, reversible immune suppression in the fetus.

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