Overcoming delayed in-vitro response to rFVIIa: effects of rFVIIa and rFVIIa analogue (vatreptacog alfa) concentration escalation in whole blood assays.

In a previous pharmacokinetic/pharmacodynamic study in nonbleeding hemophilia patients, variability in laboratory response to recombinant factor VIIa (rFVIIa) 90 µg/kg was noted, and the patients were described as delayed or rapid laboratory responders based on time to clot formation. The current study determined whether in-vitro experiments could reproduce previous in-vivo findings; whether the delayed laboratory response to rFVIIa 90 µg/kg is improved by spiking with high-dose rFVIIa or rFVIIa analogue (vatreptacog alfa); whether a dose-response is observed with our method. In-vitro experiments were conducted in our previous patient cohort using rFVIIa 1.28 and 3.84 µg/ml and vatreptacog alfa 0.28 and 0.56 µg/ml. Whole blood studies were conducted using the Hemodyne Hemostasis Analysis System (platelet contractile force, clot elastic modulus, force onset time) and rotational thromboelastometry (clotting time, maximum clot firmness). Spiking with rFVIIa 1.28 µg/ml showed the same distribution of delayed and rapid laboratory response as observed previously. Increasing in-vitro rFVIIa concentrations improved the coagulation parameters; however, there remained delayed and rapid responders. Vatreptacog alfa improved the coagulation parameters at all concentrations tested, and the 0.56 µg/ml concentration normalized the force onset time, platelet contractile force, clot elastic modulus and clotting time parameters. A dose-response was observed with both assays. There was good agreement between the laboratory responses obtained after intravenous administration of rFVIIa 90 µg/kg and in-vitro spiking studies. Escalating rFVIIa and vatreptacog alfa concentrations improved coagulation parameters in all patients compared to rFVIIa 1.28 µg/ml. Vatreptacog alfa produced more pronounced coagulation effects at lower concentrations than rFVIIa; and the 0.56 µg/ml concentration completely normalized responses in all patients.

Factor VIIa analog has marked effects on platelet function and clot kinetics in blood from patients with hemophilia A.

To evaluate the hemostatic effects of NN1731 and rFVIIa, an ex-vivo study in hemophilia patients used the Hemodyne Hemostasis Analysis System (HAS) to measure platelet contractile force (PCF), clot elastic modulus (CEM), and force onset time (FOT), and the Haemoscope Thrombelastograph (TEG) to measure reaction time (R), kinetics time (K), and maximum amplitude (MA). Blood samples from 10 healthy volunteers and 10 Factor VIII-deficient patients of varying severity (mild, moderate, severe), were spiked with rFVIIa and NN1731 (both 0.64 and 1.28 microg/ml, respectively) and analyzed to characterize platelet function and clot kinetics. There was wide variability in the rFVIIa response. NN1731 had greater and more consistent effects on PCF, CEM, FOT, R, and K relative to rFVIIa, in all hemophilia groups. The lowest NN1731 concentration (0.64 microg/ml) shortened R and FOT, and increased CEM and PCF more than rFVIIa 1.28 microg/ml. NN1731 normalized clotting parameters equivalent to values obtained in healthy volunteers. FOT and R were highly correlated (r = 0.96). No correlation was observed between CEM and MA. NN1731 produced less variable, more pronounced and predictable ex-vivo hemostatic effects on PCF, CEM, FOT, R and K than rFVIIa in all hemophilia groups. HAS and TEG assays provided similar estimates of FOT and R, however CEM appeared to be more sensitive than MA to changes in clot firmness.

Post-transplantation Lymphoproliferative Disease.

Post-transplant lymphoproliferative disorders are a varied group of disease processes that follow both solid organ and bone marrow transplantation. The disorders are typically associated with the Epstein-Barr virus and, as a consequence, have contributed greatly to our understanding of the role of human viruses in the pathogenesis of malignant disease. They have also expanded our understanding of the role of immunosurveillance in the prevention and control of malignancies which has, in turn, led to exciting developments in the use of transferred or adoptive immunotherapy for the treatment of cancer. This technology holds great potential for the treatment of these and other diseases associated with immunodeficiency states. When recognized early in their natural history, the post-transplant lymphoproliferative disorders appear to be curable; later detection frequently leads to a fatal outcome. Prompt diagnosis of these disorders by the clinician caring for the transplant patient is therefore critical.