Abstract

The clinical picture of hereditary angio-edema (HAE) is characterized by acute attacks of circumscribed swellings of the skin and sub-mucosa which recur after symptomless intervals of days to years. Episodes of swelling may involve the upper respiratory tract, including the tongue, pharynx, and larynx. This contributes to the mortality of up to 30% (or more in some families) due to suffocation as complication of laryngeal edema. This increased vascular permeability and massive local uncontrolled edema is most probably caused by uncontrolled activation of the complement system and by increased formation of bradykinin and C2 kinin due to deficiency or complete absence of C1-esterase inhibitor (C1-INH). The first line therapy for treatment of an acute attack and for short-term prophylaxis is intravenous administration of a plasma-derived C1-INH concentrate (where licensed). Such a C1-INH product is Berinert® P, licensed in e.g., Germany, Switzerland, France, and Japan. As, in general, plasma-derived products may potentially transmit viruses, special care is taken to minimize this risk by careful selection of plasma collection centers and donors. Furthermore, each donation is tested thoroughly for the absence of hepatitis A virus (HAV), hepatitis B virus (HBV), hepatitis C virus (HCV), human immunodeficiency viruses (HIV), and high titers of parvovirus B19 (B19V) by serological and/or NAT/PCR methods. In addition, the plasma pool for fractionation is tested and only released for further processing if the pool is non-reactive (negative) for HBsAg and non-reactive for antibodies against HCV and HIV-1/-2 as well as non-reactive for genomic sequences of HAV, HBV, HCV, HIV-1, and high titers of B19V (not exceeding 105 IU/ml). Selecting and testing the starting material for absence of blood borne viruses is a very important prerequisite to the production of the C1-INH concentrate; however, the manufacturing process of this product with its inherent capacity to inactivate and/or remove potentially present viruses in the plasma pool for fractionation complements the effort for a final product with a high margin of safety with regard to viruses. Two manufacturing steps - pasteurization (heat treatment of stabilized aqueous solution at 60°C for 10 hours) and hydrophobic interaction chromatography (HIC) - were validated for their capacity to inactivate and/or remove viruses. In these laboratory studies, the relevant blood borne viruses or specific and non-specific model viruses were employed: HIV and HAV as viruses of risk, bovine viral diarrhea virus (BVDV) as model virus for HCV and related viruses as West Nile virus (WNV), and canine parvovirus (CPV) as model virus for parvovirus B19 (B19V) demonstrating a very effective reduction of the infectivity by both manufacturing steps independently. Based on these virus validation studies it is evident that two complementary virus reduction steps are inherent in the manufacturing process of Berinert® P.

Mean overall virus reduction factors for Berinert® P

VirusVirus Reduction by Two Manufacturing Steps [log10]
HI ChromatographyPasteurizationOverall Virus Reduction
HIV ≥ 4.5 ≥ 6.6 11.1 
BVDV ≥ 5.1 ≥ 9.2 14.3 
PRV ≥ 6.7 6.6 13.3 
HAV ≥ 3.3 ≥6.4 9.7 
CPV 6.7 1.4 8.1 
VirusVirus Reduction by Two Manufacturing Steps [log10]
HI ChromatographyPasteurizationOverall Virus Reduction
HIV ≥ 4.5 ≥ 6.6 11.1 
BVDV ≥ 5.1 ≥ 9.2 14.3 
PRV ≥ 6.7 6.6 13.3 
HAV ≥ 3.3 ≥6.4 9.7 
CPV 6.7 1.4 8.1 

Furthermore, investigational studies demonstrate (i) an effective inactivation of B19V by pasteurization resulting in a virus reduction factor of ≥ 4.3 log10 and (ii) the removal of two different prion preparations by a single manufacturing step (microsomes and purified PrPSc by 3.1 log10 and 3.2 log10, respectively). Therefore, a high safety margin for pathogens can be attributed to Berinert® P.

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