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Identification and multidimensional optimization of an asymmetric bispecific IgG antibody mimicking the function of factor VIII cofactor activity.

Sampei Z, Igawa T, Soeda T, Okuyama-Nishida Y, Moriyama C, Wakabayashi T, Tanaka E, Muto A, Kojima T, Kitazawa T, Yoshihashi K, Harada A, Funaki M, Haraya K, Tachibana T, Suzuki S, Esaki K, Nabuchi Y, Hattori K - PLoS ONE (2013)

Bottom Line: Since the therapeutic potential of the lead bispecific antibody was marginal, FVIII-mimetic activity was improved by modifying its binding properties to FIXa and FX, and the pharmacokinetics was improved by engineering the charge properties of the variable region.Importantly, the activity of hBS910 was not affected by FVIII inhibitors, while anti-hBS910 antibodies did not inhibit FVIII activity, allowing the use of hBS910 without considering the development or presence of FVIII inhibitors.We expect that hBS910 (investigational drug name: ACE910) will provide significant benefit for severe hemophilia A patients.

View Article: PubMed Central - PubMed

Affiliation: Research Division, Chugai Pharmaceutical Co., Ltd., Gotemba, Shizuoka, Japan.

ABSTRACT
In hemophilia A, routine prophylaxis with exogenous factor VIII (FVIII) requires frequent intravenous injections and can lead to the development of anti-FVIII alloantibodies (FVIII inhibitors). To overcome these drawbacks, we screened asymmetric bispecific IgG antibodies to factor IXa (FIXa) and factor X (FX), mimicking the FVIII cofactor function. Since the therapeutic potential of the lead bispecific antibody was marginal, FVIII-mimetic activity was improved by modifying its binding properties to FIXa and FX, and the pharmacokinetics was improved by engineering the charge properties of the variable region. Difficulties in manufacturing the bispecific antibody were overcome by identifying a common light chain for the anti-FIXa and anti-FX heavy chains through framework/complementarity determining region shuffling, and by pI engineering of the two heavy chains to facilitate ion exchange chromatographic purification of the bispecific antibody from the mixture of byproducts. Engineering to overcome low solubility and deamidation was also performed. The multidimensionally optimized bispecific antibody hBS910 exhibited potent FVIII-mimetic activity in human FVIII-deficient plasma, and had a half-life of 3 weeks and high subcutaneous bioavailability in cynomolgus monkeys. Importantly, the activity of hBS910 was not affected by FVIII inhibitors, while anti-hBS910 antibodies did not inhibit FVIII activity, allowing the use of hBS910 without considering the development or presence of FVIII inhibitors. Furthermore, hBS910 could be purified on a large manufacturing scale and formulated into a subcutaneously injectable liquid formulation for clinical use. These features of hBS910 enable routine prophylaxis by subcutaneous delivery at a long dosing interval without considering the development or presence of FVIII inhibitors. We expect that hBS910 (investigational drug name: ACE910) will provide significant benefit for severe hemophilia A patients.

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Therapeutic potential of multidimensionally optimized bispecific antibody, hBS910.(A) FVIII-mimetic activity of hBS910 in thrombin generation assay (TGA). Effect of hBS910 (circles) or recombinant human FVIII (squares) on thrombin generation in FVIII-deficient plasma is shown. The reaction was triggered by FXIa, synthetic phospholipid, and Ca2+. The Y-axis indicates the peak height, a thrombin generation parameter (in many cases, the bars depicting s.d. are shorter than the height of the symbols). Data were collected in triplicate for each plasma lot and are expressed as mean ± s.d. (B) Pharmacokinetics of hBS910 in cynomolgus monkeys. Time profiles of plasma concentration of hBS910 after intravenous (circles) or subcutaneous (squares) injection are shown.
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pone-0057479-g007: Therapeutic potential of multidimensionally optimized bispecific antibody, hBS910.(A) FVIII-mimetic activity of hBS910 in thrombin generation assay (TGA). Effect of hBS910 (circles) or recombinant human FVIII (squares) on thrombin generation in FVIII-deficient plasma is shown. The reaction was triggered by FXIa, synthetic phospholipid, and Ca2+. The Y-axis indicates the peak height, a thrombin generation parameter (in many cases, the bars depicting s.d. are shorter than the height of the symbols). Data were collected in triplicate for each plasma lot and are expressed as mean ± s.d. (B) Pharmacokinetics of hBS910 in cynomolgus monkeys. Time profiles of plasma concentration of hBS910 after intravenous (circles) or subcutaneous (squares) injection are shown.

Mentions: To examine the therapeutic potential of hBS910, its FVIII-mimetic activity was compared with human FVIII by using thrombin generation assay (TGA) [23], [35], [36] in commercially available human FVIII-deficient plasma which was derived from a single donor with severe hemophilia A without FVIII inhibitors (Fig. 7A). hBS910 dose-dependently increased peak height (defined as the peak concentration of free thrombin) in the same manner as recombinant human FVIII (rhFVIII). The thrombin generation activity of hBS910 was also observed even in plasma of a hemophilia A donor who has FVIII inhibitors, whereas 1 IU/mL of rhFVIII did not exhibit any effects (data not shown). On the other hand, while polyclonal anti-idiotype antibodies to the anti-FIXa Fab or anti-FX Fab of hBS910 completely inhibited the activity of hBS910, they did not interfere with rhFVIII activity at all (data not shown).


Identification and multidimensional optimization of an asymmetric bispecific IgG antibody mimicking the function of factor VIII cofactor activity.

Sampei Z, Igawa T, Soeda T, Okuyama-Nishida Y, Moriyama C, Wakabayashi T, Tanaka E, Muto A, Kojima T, Kitazawa T, Yoshihashi K, Harada A, Funaki M, Haraya K, Tachibana T, Suzuki S, Esaki K, Nabuchi Y, Hattori K - PLoS ONE (2013)

Therapeutic potential of multidimensionally optimized bispecific antibody, hBS910.(A) FVIII-mimetic activity of hBS910 in thrombin generation assay (TGA). Effect of hBS910 (circles) or recombinant human FVIII (squares) on thrombin generation in FVIII-deficient plasma is shown. The reaction was triggered by FXIa, synthetic phospholipid, and Ca2+. The Y-axis indicates the peak height, a thrombin generation parameter (in many cases, the bars depicting s.d. are shorter than the height of the symbols). Data were collected in triplicate for each plasma lot and are expressed as mean ± s.d. (B) Pharmacokinetics of hBS910 in cynomolgus monkeys. Time profiles of plasma concentration of hBS910 after intravenous (circles) or subcutaneous (squares) injection are shown.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3585358&req=5

pone-0057479-g007: Therapeutic potential of multidimensionally optimized bispecific antibody, hBS910.(A) FVIII-mimetic activity of hBS910 in thrombin generation assay (TGA). Effect of hBS910 (circles) or recombinant human FVIII (squares) on thrombin generation in FVIII-deficient plasma is shown. The reaction was triggered by FXIa, synthetic phospholipid, and Ca2+. The Y-axis indicates the peak height, a thrombin generation parameter (in many cases, the bars depicting s.d. are shorter than the height of the symbols). Data were collected in triplicate for each plasma lot and are expressed as mean ± s.d. (B) Pharmacokinetics of hBS910 in cynomolgus monkeys. Time profiles of plasma concentration of hBS910 after intravenous (circles) or subcutaneous (squares) injection are shown.
Mentions: To examine the therapeutic potential of hBS910, its FVIII-mimetic activity was compared with human FVIII by using thrombin generation assay (TGA) [23], [35], [36] in commercially available human FVIII-deficient plasma which was derived from a single donor with severe hemophilia A without FVIII inhibitors (Fig. 7A). hBS910 dose-dependently increased peak height (defined as the peak concentration of free thrombin) in the same manner as recombinant human FVIII (rhFVIII). The thrombin generation activity of hBS910 was also observed even in plasma of a hemophilia A donor who has FVIII inhibitors, whereas 1 IU/mL of rhFVIII did not exhibit any effects (data not shown). On the other hand, while polyclonal anti-idiotype antibodies to the anti-FIXa Fab or anti-FX Fab of hBS910 completely inhibited the activity of hBS910, they did not interfere with rhFVIII activity at all (data not shown).

Bottom Line: Since the therapeutic potential of the lead bispecific antibody was marginal, FVIII-mimetic activity was improved by modifying its binding properties to FIXa and FX, and the pharmacokinetics was improved by engineering the charge properties of the variable region.Importantly, the activity of hBS910 was not affected by FVIII inhibitors, while anti-hBS910 antibodies did not inhibit FVIII activity, allowing the use of hBS910 without considering the development or presence of FVIII inhibitors.We expect that hBS910 (investigational drug name: ACE910) will provide significant benefit for severe hemophilia A patients.

View Article: PubMed Central - PubMed

Affiliation: Research Division, Chugai Pharmaceutical Co., Ltd., Gotemba, Shizuoka, Japan.

ABSTRACT
In hemophilia A, routine prophylaxis with exogenous factor VIII (FVIII) requires frequent intravenous injections and can lead to the development of anti-FVIII alloantibodies (FVIII inhibitors). To overcome these drawbacks, we screened asymmetric bispecific IgG antibodies to factor IXa (FIXa) and factor X (FX), mimicking the FVIII cofactor function. Since the therapeutic potential of the lead bispecific antibody was marginal, FVIII-mimetic activity was improved by modifying its binding properties to FIXa and FX, and the pharmacokinetics was improved by engineering the charge properties of the variable region. Difficulties in manufacturing the bispecific antibody were overcome by identifying a common light chain for the anti-FIXa and anti-FX heavy chains through framework/complementarity determining region shuffling, and by pI engineering of the two heavy chains to facilitate ion exchange chromatographic purification of the bispecific antibody from the mixture of byproducts. Engineering to overcome low solubility and deamidation was also performed. The multidimensionally optimized bispecific antibody hBS910 exhibited potent FVIII-mimetic activity in human FVIII-deficient plasma, and had a half-life of 3 weeks and high subcutaneous bioavailability in cynomolgus monkeys. Importantly, the activity of hBS910 was not affected by FVIII inhibitors, while anti-hBS910 antibodies did not inhibit FVIII activity, allowing the use of hBS910 without considering the development or presence of FVIII inhibitors. Furthermore, hBS910 could be purified on a large manufacturing scale and formulated into a subcutaneously injectable liquid formulation for clinical use. These features of hBS910 enable routine prophylaxis by subcutaneous delivery at a long dosing interval without considering the development or presence of FVIII inhibitors. We expect that hBS910 (investigational drug name: ACE910) will provide significant benefit for severe hemophilia A patients.

Show MeSH
Related in: MedlinePlus