Observations on Marketed Alpha-1-Proteinase Inhibitor Products Ewa Marszal, PhD FDA/CBER BPAC, November 2005

Presentation Contents Differences between a 1 -PI products Consistency of manufacturing

Differences between a 1 -PI products Contaminating plasma proteins Inactive a 1 -PI (e.g., polymer/latent form) Modifications of the primary structure

Protein impurities (SDS-PAGE, reducing) Prolastin Aralast Zemaira albumin → a 1 -PI → Specific activity: ≥ 0.35 ≥ 0.55 ≥ 0.7

mg functional a 1 -PI/mg total protein

Presence of high molecular weight species (SE-HPLC, absorbance@215 nm) Zemaira Aralast Prolastin

M M%

(a 1 -PI

+

albumin) 95.4

89.8

88.1

Isoelectric focusing (IEF) cathode (-) higher pH zero net charge pH gradient anode (+) lower pH

Heterogeneity of a 1 -PI in plasma by IEF (charge heterogeneity) 3 polysaccharide chains (sialic acid) N-terminal pentapeptide ED PNG Polymorphism, e.g., E 342 K

Dominant a 1 -PI isoforms in plasma (charge heterogeneity) M6 ED PNG N N N M4 ED PNG N N N M2 ED PNG N N N

M6 → M4 → M2 → Heterogeneity of a 1 -PI products by IEF desialylated

Consistency of manufacturing by IEF Analysis of patients’ plasma from Aralast pivotal clinical trial Analysis of current and historical lots of a 1 -PI products

Conclusions a 1 -PI products contain: different levels of contaminating plasma proteins different amounts of inactive a 1 -PI species different levels of primary structure modifications a 1 -PI in each product differs somewhat from a 1 -PI in plasma (IEF, polymer) Manufacturing of each a 1 -PI product appears to be consistent over time by IEF

Consistency of manufacturing Bayer Baxter Behring Desialylated Non-desialylated