Novel Approaches for Hemophilia Treatment
Novel Approaches for Hemophilia Treatment
The fusion proteins rFVIII-Fc and rFIXFc are composed of a single molecule of recombinant factor covalently fused to the dimeric Fc domain of IgG1. This approach combines two molecular structures, each with a long history of safety and efficacy in clinical use. The immunoglobulin G constant region (Fc) has been molecularly engineered to create fusion proteins that prolong the circulating half-life (T1/2) of Fc fusion–based drugs used clinically (eg, etanercept, romiplostim). After nearly a decade of experience in clinical use, these protein fusions support the safety of the Fc fusion approach.
With Fc fusion proteins, the neonatal Fc receptor interacting with the endogenous IgG recycling pathway delays lysosomal degradation of IgG (half-life of 3 weeks) and the fusion proteins, recycling them back into circulation and thus prolonging the plasma half-life. Although Fc fusions are typically expressed as homodimers formed through a disulfide bond, this dimeric structure was not effective for large clotting factors. Fusion of the monomeric form of the IgG1 Fc to human FIX, FVIIa, and B domain-deleted FVIII was effective and demonstrated increases in plasma half-life. In addition, the larger-sized molecules are expected to experience slower renal clearance. There are no intervening linker sequences. A human cell line, HEK-293H cells, is used for expression and co-transfected with the expression cassette for PC5, a processing enzyme, to ensure full cleavage of the FIX propeptide. The glycosylation of recombinant proteins expressed in HEK-293H cells sometimes varies from the cognate human natural protein, but any clinical implications remain unclear.
Preclinical studies of Fc fusion proteins with both rFIXFc and rFVIII-Fc demonstrated safety and prolonged half-life of efficacy in bleeding models when compared with current recombinant clotting factors in mice, rats, hemophilic dogs and cynomolgus monkeys. Biochemical characterization confirmed proper propeptide processing and appropriate extensive posttranslational modification after expression in HEK-293H cells. Peters and colleagues showed that rFIXFc has a three- to fourfold longer terminal T1/2 in mice expressing human FcRn compared with rFIX, whereas both proteins have similar short terminal half-lives in mice lacking FcRn, confirming that FcRn mediates the longer half-life of rFIXFc through use of the natural pathway responsible for protecting IgG antibodies from degradation. The half-life extension of FVIII-Fc was due to similar mechanisms but was additionally limited by the close interaction of FVIII with the chaperone protein von Willebrand factor (vWF).
Shapiro and colleagues reported the safety and pharmacokinetics of the first-in-human phase 1/2a clinical trial of rFIXFc (Clinicaltrials.gov identifier: NCT00716716) in previously treated adults with hemophilia B. A dose-proportional increase in FIX activity was observed immediately after infusion and the mean FIX activity terminal half-life for rFIXFc was 56.7 hours, approximately threefold longer than reported for current rFIX products. A phase 3 nonrandomized, open-label, multicenter study assessed the safety and efficacy of repeated administration of rFIXFc in 123 adolescents and adults with severe hemophilia B (ClinicalTrials.gov identifier: NCT01027364). The participants were representative of the general adult population with severe hemophilia B.
Reflecting different clinical regimens, the study included four treatment groups: group 1 received weekly dose-adjusted prophylaxis (50 IU of rFIXFc per kilogram of body weight to start), with the dose adjusted as needed clinically; group 2 received interval-adjusted prophylaxis (100 IU/kg every 10 days to start), with the interval adjusted as needed; group 3 received treatment as needed (episodic or on-demand treatment) for bleeding episodes (20 to 100 IU/kg), with the dose adjusted according to bleeding severity; and group 4 received treatment as needed for surgical procedures.
A total of 5243 rFIXFc administrations occurred during the study, and overall, 96.6% of participants in the prophylaxis groups were adherent to their treatment regimen; most self-administered rFIXFc at home. The terminal half-life of rFIXFc was significantly longer than that of rFIX (geometric mean, 82.1 vs 33.8 hours; P < .001). The incremental recoveries were similar. The time to reach a factor IX level of 1 IU/dL (1%) was 11.2 days with rFIXFc and 5.1 days with rFIX. In the group that received prophylaxis dosing based on individualized intervals to maintain trough FIX levels above 3% (Group 2), over half of the participants had dosing intervals of 14 or more days during the last 3 months on the study. Participants in groups 1 and 2, with the highest bleeding frequency before study entry, had median annualized bleeding rates of 2.05 and 2.76, respectively, while enrolled in the study. Among the participants receiving prophylaxis, 23.0% in group 1 and 42.3% in group 2 had no bleeding episodes during the study. Adverse events observed in this study were consistent with those expected in the population of persons with hemophilia. In the surgery arm of this phase 3 trial, rFIXFc also demonstrated safe and effective perioperative hemostasis during the perioperative period in persons requiring major (mostly orthopedic) surgery. No inhibitors were detected for subjects switching to the fusion protein. There was no evidence of allergic reactions or thrombogenicity as reported previously for rFIX.
Similar results were demonstrated in the two clinical trials for rFVIII-Fc, with safety confirmed and no inhibitor development in previously treated patients. However, probably due to the interaction of FVIII with its chaperone protein vWF, there was less impressive improvement in pharmacokinetic parameters, with the half-life essentially limited to somewhat less than that of vWF. In the phase 3 study, the terminal half-life of rFVIII-Fc was 1.53-fold longer compared with recombinant full-length FVIII (19.0 vs 12.4 hours; P < .001). Extended-interval dosing, with some patients able to infuse every 5 or 6 days, supported low rates of spontaneous bleeding, and the individual increases in half-life correlated with endogenous levels of vWF.
Fc Fusion Proteins
The fusion proteins rFVIII-Fc and rFIXFc are composed of a single molecule of recombinant factor covalently fused to the dimeric Fc domain of IgG1. This approach combines two molecular structures, each with a long history of safety and efficacy in clinical use. The immunoglobulin G constant region (Fc) has been molecularly engineered to create fusion proteins that prolong the circulating half-life (T1/2) of Fc fusion–based drugs used clinically (eg, etanercept, romiplostim). After nearly a decade of experience in clinical use, these protein fusions support the safety of the Fc fusion approach.
With Fc fusion proteins, the neonatal Fc receptor interacting with the endogenous IgG recycling pathway delays lysosomal degradation of IgG (half-life of 3 weeks) and the fusion proteins, recycling them back into circulation and thus prolonging the plasma half-life. Although Fc fusions are typically expressed as homodimers formed through a disulfide bond, this dimeric structure was not effective for large clotting factors. Fusion of the monomeric form of the IgG1 Fc to human FIX, FVIIa, and B domain-deleted FVIII was effective and demonstrated increases in plasma half-life. In addition, the larger-sized molecules are expected to experience slower renal clearance. There are no intervening linker sequences. A human cell line, HEK-293H cells, is used for expression and co-transfected with the expression cassette for PC5, a processing enzyme, to ensure full cleavage of the FIX propeptide. The glycosylation of recombinant proteins expressed in HEK-293H cells sometimes varies from the cognate human natural protein, but any clinical implications remain unclear.
Preclinical studies of Fc fusion proteins with both rFIXFc and rFVIII-Fc demonstrated safety and prolonged half-life of efficacy in bleeding models when compared with current recombinant clotting factors in mice, rats, hemophilic dogs and cynomolgus monkeys. Biochemical characterization confirmed proper propeptide processing and appropriate extensive posttranslational modification after expression in HEK-293H cells. Peters and colleagues showed that rFIXFc has a three- to fourfold longer terminal T1/2 in mice expressing human FcRn compared with rFIX, whereas both proteins have similar short terminal half-lives in mice lacking FcRn, confirming that FcRn mediates the longer half-life of rFIXFc through use of the natural pathway responsible for protecting IgG antibodies from degradation. The half-life extension of FVIII-Fc was due to similar mechanisms but was additionally limited by the close interaction of FVIII with the chaperone protein von Willebrand factor (vWF).
Shapiro and colleagues reported the safety and pharmacokinetics of the first-in-human phase 1/2a clinical trial of rFIXFc (Clinicaltrials.gov identifier: NCT00716716) in previously treated adults with hemophilia B. A dose-proportional increase in FIX activity was observed immediately after infusion and the mean FIX activity terminal half-life for rFIXFc was 56.7 hours, approximately threefold longer than reported for current rFIX products. A phase 3 nonrandomized, open-label, multicenter study assessed the safety and efficacy of repeated administration of rFIXFc in 123 adolescents and adults with severe hemophilia B (ClinicalTrials.gov identifier: NCT01027364). The participants were representative of the general adult population with severe hemophilia B.
Reflecting different clinical regimens, the study included four treatment groups: group 1 received weekly dose-adjusted prophylaxis (50 IU of rFIXFc per kilogram of body weight to start), with the dose adjusted as needed clinically; group 2 received interval-adjusted prophylaxis (100 IU/kg every 10 days to start), with the interval adjusted as needed; group 3 received treatment as needed (episodic or on-demand treatment) for bleeding episodes (20 to 100 IU/kg), with the dose adjusted according to bleeding severity; and group 4 received treatment as needed for surgical procedures.
A total of 5243 rFIXFc administrations occurred during the study, and overall, 96.6% of participants in the prophylaxis groups were adherent to their treatment regimen; most self-administered rFIXFc at home. The terminal half-life of rFIXFc was significantly longer than that of rFIX (geometric mean, 82.1 vs 33.8 hours; P < .001). The incremental recoveries were similar. The time to reach a factor IX level of 1 IU/dL (1%) was 11.2 days with rFIXFc and 5.1 days with rFIX. In the group that received prophylaxis dosing based on individualized intervals to maintain trough FIX levels above 3% (Group 2), over half of the participants had dosing intervals of 14 or more days during the last 3 months on the study. Participants in groups 1 and 2, with the highest bleeding frequency before study entry, had median annualized bleeding rates of 2.05 and 2.76, respectively, while enrolled in the study. Among the participants receiving prophylaxis, 23.0% in group 1 and 42.3% in group 2 had no bleeding episodes during the study. Adverse events observed in this study were consistent with those expected in the population of persons with hemophilia. In the surgery arm of this phase 3 trial, rFIXFc also demonstrated safe and effective perioperative hemostasis during the perioperative period in persons requiring major (mostly orthopedic) surgery. No inhibitors were detected for subjects switching to the fusion protein. There was no evidence of allergic reactions or thrombogenicity as reported previously for rFIX.
Similar results were demonstrated in the two clinical trials for rFVIII-Fc, with safety confirmed and no inhibitor development in previously treated patients. However, probably due to the interaction of FVIII with its chaperone protein vWF, there was less impressive improvement in pharmacokinetic parameters, with the half-life essentially limited to somewhat less than that of vWF. In the phase 3 study, the terminal half-life of rFVIII-Fc was 1.53-fold longer compared with recombinant full-length FVIII (19.0 vs 12.4 hours; P < .001). Extended-interval dosing, with some patients able to infuse every 5 or 6 days, supported low rates of spontaneous bleeding, and the individual increases in half-life correlated with endogenous levels of vWF.