Increased Accumulation and Retention of rhFVIIa (eptacog beta) in Knee Joints of Hemophilia A Mice Compared to Wild-Type Mice

Jhansi Magisetty; Usha R. Pendurthi; SubbaRao V. Madhunapantula; Jerry Grandoni; L. Vijaya Mohan Rao

doi:10.1055/s-0039-1688907

Thrombosis and Haemostasis, Table of Contents

Thromb Haemost 2019; 119(08): 1283-1294
DOI: 10.1055/s-0039-1688907

Coagulation and Fibrinolysis

Georg Thieme Verlag KG Stuttgart · New York

Increased Accumulation and Retention of rhFVIIa (eptacog beta) in Knee Joints of Hemophilia A Mice Compared to Wild-Type Mice

Jhansi Magisetty

¹Department of Cellular and Molecular Biology, The University of Texas Health Science Center at Tyler, Tyler, Texas, United States

,

Usha R. Pendurthi

¹Department of Cellular and Molecular Biology, The University of Texas Health Science Center at Tyler, Tyler, Texas, United States

,

SubbaRao V. Madhunapantula

²Center of Excellence in Molecular Biology and Regenerative Medicine Laboratory, Department of Biochemistry, JSS Medical College, JSS Academy of Higher Education & Research, Mysuru, Karnataka, India

,

Jerry Grandoni

³LFB USA, Framingham, Massachusetts, United States

,

L. Vijaya Mohan Rao

¹Department of Cellular and Molecular Biology, The University of Texas Health Science Center at Tyler, Tyler, Texas, United States

› Author Affiliations

Abstract

Our earlier studies showed that recombinant human factor VIIa (rhFVIIa) administered intravascularly in mice disappeared rapidly from the circulation. However, a small fraction of rhFVIIa that entered extravascular remained functionally active for an extended period. The present study aims to investigate the dose-dependency of rhFVIIa accumulation and retention in mouse knee joints and test whether the hemophilic condition affects rhFVIIa sequestration in joints. Wild-type and FVIII^−/− mice were injected with three doses of rhFVIIa (eptacog beta, 90, 250, and 500 μg/kg) via the tail vein. At varying times following rhFVIIa administration, blood and knee joints were collected to measure FVIIa activity and antigen levels in plasma and joint tissues. Joint tissue sections were analyzed by immunohistochemistry for the presence of rhFVIIa. Vascular permeability was assessed by either Evans Blue dye or fluorescein dextran extravasation. The study showed that rhFVIIa accumulated in knee joints of wild-type and FVIII^−/− mice in a dose-dependent manner. rhFVIIa antigen and FVIIa activity could be detectable in joints for at least 7 days. Significantly higher levels of rhFVIIa accumulation were observed in knee joints of FVIII^−/− mice compared with that of wild-type mice. Immunohistochemical analyses confirmed higher levels of rhFVIIa retention in FVIII^−/− mice compared with wild-type mice. Additional studies showed that FVIII^−/− mice were more permissible to vascular leakage. In conclusion, the present data demonstrate a dose-dependent accumulation of rhFVIIa in knee joints, and the hemophilic condition enhances the entry of rhFVIIa from circulation to the extravascular. The present data will be useful in improving rhFVIIa prophylaxis.

Keywords

factor VIIa - eptacog beta - hemophilia - permeability - prophylaxis

Full Text

References

References
1 Hedner U. Factor VIIa and its potential therapeutic use in bleeding-associated pathologies. Thromb Haemost 2008; 100 (04) 557-562
2 Abshire T, Kenet G. Recombinant factor VIIa: review of efficacy, dosing regimens and safety in patients with congenital and acquired factor VIII or IX inhibitors. J Thromb Haemost 2004; 2 (06) 899-909
3 Logan AC, Yank V, Stafford RS. Off-label use of recombinant factor VIIa in U.S. hospitals: analysis of hospital records. Ann Intern Med 2011; 154 (08) 516-522
4 Mathew P, Young G. Recombinant factor VIIa in paediatric bleeding disorders--a 2006 review. Haemophilia 2006; 12 (05) 457-472
5 Witmer CM, Huang YS, Lynch K, Raffini LJ, Shah SS. Off-label recombinant factor VIIa use and thrombosis in children: a multi-center cohort study. J Pediatr 2011; 158 (05) 820-825
6 Saxon BR, Shanks D, Jory CB, Williams V. Effective prophylaxis with daily recombinant factor VIIa (rFVIIa-Novoseven) in a child with high titre inhibitors and a target joint. Thromb Haemost 2001; 86 (04) 1126-1127
7 Young G, McDaniel M, Nugent DJ. Prophylactic recombinant factor VIIa in haemophilia patients with inhibitors. Haemophilia 2005; 11 (03) 203-207
8 Blatny J, Kohlerova S, Zapletal O, Fiamoli V, Penka M, Smith O. Prophylaxis with recombinant factor VIIa for the management of bleeding episodes during immune tolerance treatment in a boy with severe haemophilia A and high-response inhibitors. Haemophilia 2008; 14 (05) 1140-1142
9 Morfini M, Auerswald G, Kobelt RA. , et al. Prophylactic treatment of haemophilia patients with inhibitors: clinical experience with recombinant factor VIIa in European Haemophilia Centres. Haemophilia 2007; 13 (05) 502-507
10 Tcheng WY, Donkin J, Konzal S, Wong WY. Recombinant factor VIIa prophylaxis in a patient with severe congenital factor VII deficiency. Haemophilia 2004; 10 (03) 295-298
11 Mathijssen NC, Masereeuw R, Verbeek K. , et al. Prophylactic effect of recombinant factor VIIa in factor VII deficient patients. Br J Haematol 2004; 125 (04) 494-499
12 Konkle BA, Ebbesen LS, Erhardtsen E. , et al. Randomized, prospective clinical trial of recombinant factor VIIa for secondary prophylaxis in hemophilia patients with inhibitors. J Thromb Haemost 2007; 5 (09) 1904-1913
13 Young G, Auerswald G, Jimenez-Yuste V. , et al. PRO-PACT: retrospective observational study on the prophylactic use of recombinant factor VIIa in hemophilia patients with inhibitors. Thromb Res 2012; 130 (06) 864-870
14 Gopalakrishnan R, Hedner U, Ghosh S. , et al. Bio-distribution of pharmacologically administered recombinant factor VIIa (rFVIIa). J Thromb Haemost 2010; 8 (02) 301-310
15 Gopalakrishnan R, Hedner U, Clark C, Pendurthi UR, Rao LV. rFVIIa transported from the blood stream into tissues is functionally active. J Thromb Haemost 2010; 8 (10) 2318-2321
16 Sen P, Clark CA, Gopalakrishnan R. , et al. Factor VIIa binding to endothelial cell protein C receptor: differences between mouse and human systems. Thromb Haemost 2012; 107 (05) 951-961
17 Clark CA, Vatsyayan R, Hedner U, Esmon CT, Pendurthi UR, Rao LV. Endothelial cell protein C receptor-mediated redistribution and tissue-level accumulation of factor VIIa. J Thromb Haemost 2012; 10 (11) 2383-2391
18 Vatsyayan R, Kothari H, Mackman N, Pendurthi UR, Rao LV. Inactivation of factor VIIa by antithrombin in vitro, ex vivo and in vivo: role of tissue factor and endothelial cell protein C receptor. PLoS One 2014; 9 (08) e103505
19 Chevreux G, Faid V, Scohyers JM, Bihoreau N. N-/O-glycosylation analysis of human FVIIa produced in the milk of transgenic rabbits. Glycobiology 2013; 23 (12) 1531-1546
20 Chevreux G, Tilly N, Leblanc Y. , et al. Biochemical characterization of LR769, a new recombinant factor VIIa bypassing agent produced in the milk of transgenic rabbits. Haemophilia 2017; 23 (04) e324-e334
21 Grandoni J, Perret G, Forier C. Kinetic analysis and binding studies of a new recombinant human factor VIIa for treatment of haemophilia. Haemophilia 2017; 23 (02) 300-308
22 Morrissey JH, Macik BG, Neuenschwander PF, Comp PC. Quantitation of activated factor VII levels in plasma using a tissue factor mutant selectively deficient in promoting factor VII activation. Blood 1993; 81 (03) 734-744
23 Nguyen DH, Zhou T, Shu J, Mai J-H. Quantifying chromogen intensity in immunohistochemistry via reciprocal intensity. Cancer Incytes 2013; 2 (01)
24 Gopalakrishnan R, Pendurthi UR, Hedner U, Agersø H, Esmon CT, Rao LV. Influence of endothelial cell protein C receptor on plasma clearance of factor VIIa. J Thromb Haemost 2012; 10 (05) 971-973
25 Lopez-Fernandez M, Altisent C, Jimenez-Yuste V. , et al. Effective prophylaxis with rFVIIa in young haemophiliacs with inhibitors using a schedule similar to FVIII prophylaxis in non-inhibitor patients. Health 2013; 5: 1151-1157
26 Leissinger CA, Singleton T, Kruse-Jarres R. How I use bypassing therapy for prophylaxis in patients with hemophilia A and inhibitors. Blood 2015; 126 (02) 153-159
27 Wang M, Lawrence JB, Quon DV. , et al. PERSEPT 1: a phase 3 trial of activated eptacog beta for on-demand treatment of haemophilia inhibitor-related bleeding. Haemophilia 2017; 23 (06) 832-843
28 Neri Serneri GG, Gensini GF, Gensini RA. Increased capillary permeability in haemophilia and afibrinogenaemia. Acta Haematol 1974; 52 (06) 336-344
29 O'Doherty MJ, Page CJ, Harrington C, Nunan T, Savidge G. Haemophilia, AIDS and lung epithelial permeability. Eur J Haematol 1990; 44 (04) 252-256
30 Sundaram J, Keshava S, Gopalakrishnan R, Esmon CT, Pendurthi UR, Rao LVM. Factor VIIa binding to endothelial cell protein C receptor protects vascular barrier integrity in vivo. J Thromb Haemost 2014; 12 (05) 690-700
31 Cooke EJ, Zhou JY, Wyseure T. , et al. Vascular permeability and remodelling coincide with inflammatory and reparative processes after joint bleeding in factor VIII-deficient mice. Thromb Haemost 2018; 118 (06) 1036-1047
32 Valentino LA, Hakobyan N. Histological changes in murine haemophilic synovitis: a quantitative grading system to assess blood-induced synovitis. Haemophilia 2006; 12 (06) 654-662
33 Ducore J, Alexander WA, Mitchell IS, Lawrence J, Grandoni J, Bonzo D. Potential clinical trial validation of differential EPCR-dependent activity by eptacog beta, a rhFVIIa, in the treatment of bleeding in hemophilia patients with inhibitors. Research and Practice in Thrombosis and Haemostasis 2018; 2 (S1) PB197

Supplementary Material

Supplementary Material