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Fall 2021 - Innovation

Hemophilia Gene Therapy: Cures May Finally Be at Hand

The concept of gene therapy can be traced back at least 80 years, but its real genesis began with Professor William Szybalski, who in 1962 performed the first virus-mediated gene transfer to mammalian cells to correct a genetic defect. Two decades later, scientists fully characterized the normal amino acid sequences of factor VIII (FVIII) and factor IX (FIX), and successfully induced immortalized hamster ovary or kidney cells to express them in large-scale cell culture for purification into concentrates for the treatment of hemophilia A and B. If mammalian cells could be coaxed to produce large quantities of functional clotting factor, it was not a major conceptual leap to imagine targeted cells in the hemophilia patient himself could do so as well. And thus was born the dream of exploiting gene therapy to essentially cure hemophilia. 

Where Clotting Factor Therapy Falls Short

Administered prophylactically, either recombinant or plasma-derived factor concentrates can sharply reduce the risk of spontaneous hemorrhage, limit the severity of traumatic bleeds and protect joints from chronic bleeding-related arthropathy. For people living with severe hemophilia, in whom less than 1 percent FVIII or FIX activity results in frequent spontaneous bleeding episodes, sustained maintenance of just 5 percent of the normal endogenous level of FVIII or FIX can improve the disease to a mild phenotype, sharply reducing the risk of frequent or serious bleeding events.

But as transformational as factor concentrates have been for persons with hemophilia, they are still well short of a functional cure. 

Most obviously, there is the requirement for frequent self-injections and the ever-present potential for breakthrough bleeds, particularly if a dose is missed. Additionally, because hemophilia patients don’t naturally produce normal clotting factor, their immune system can “see” the exogenous FVIII or FIX protein as a foreign antigen and generate inhibitor antibodies that block its critical function in the coagulation cascade. Inhibitors occur in an estimated 25 percent to 30 percent of persons with hemophilia A and about one in 20 persons with hemophilia B, but generally with higher severity. 

Thus, the decades-old dream is to correct the factor deficiency through a single infusion of a hemophilia gene therapy that produces steady high protective levels of FVIII or FIX. Ideally, a single injection of viral vectors carrying the normal gene would result in a permanent functional cure, freeing patients with severe or moderately severe disease from the lifelong need to regularly self-administer factor concentrates* and the residual risk of serious or potentially life-threatening bleeding events. 

* or emicizumab-kxwh (Hemlibra)

In addition, a bonus for the healthcare system is the potential for hemophilia gene therapy to dramatically reduce the lifetime cost of managing severe hemophilia, now estimated to well exceed $20 million with both prophylaxis and on-demand treatment strategies.1

Hurdles That Must Be Crossed

At least six companies currently have prospective hemophilia A and B gene therapies in various stages of clinical testing (Table). The prospect of eventual regulatory approvals of these treatments got a significant boost with the U.S. Food and Drug Administration’s (FDA) 2017 marketing clearance of the first gene therapy: Kymriah (tisagenlecleucel), a genetically modified autologous T-cell immunotherapy for the treatment of a form of acute lymphoblastic leukemia in children and young adults.2 But unlike in vivo hemophilia gene therapies, nearly all of which are given intravenously, Kymriah is the first of a class of chimeric antigen receptor T-cell (CAR-T) gene therapies wherein large numbers of patient T lymphocytes are collected by apheresis and modified ex vivo to incorporate a gene coding for a specific protein that directs the T cell to target and kill leukemia cells. 

More akin to hemophilia are a number of investigational gene therapies that similarly target serious or life-threatening genetic disorders, notably including sickle cell disease (SCD), beta thalassemia and severe combined immunodeficiency. These treatments similarly involve the infusion of many copies of a gene-carrying adeno-associated virus (AAV) or lentiviral vector designed to target and transduce liver or other cells to express the critical missing enzyme or other functional protein. 

But there are two fundamental hurdles that any hemophilia or other investigational gene therapy must address:

• Clinical trial results must demonstrate there are no serious safety concerns associated with infusing the viral vector. Specifically, patient studies need to show the gene therapy does not induce a serious immune response or cause hematological malignancies through insertional mutagenesis. While leukemias that plagued early human gene therapy experiments have not been identified in subjects receiving newer AAV or lentiviral vectors, in August 2021, a case of myelodysplastic syndrome prompted FDA to place a clinical hold on bluebird bio’s cerebral adrenoleukodystrophy (CALD) gene therapy program.3

• Transduced cells must express therapeutic levels of the fully functional clotting protein on a sustained basis. Simply put, the agency wants to see robust documentation of gene therapy durability, which can only come from extended follow-up of numbers of treated study participants.

These issues came into sharp focus with FDA’s recent response to BioMarin Pharmaceutical’s biologics license application (BLA) for its hemophilia A gene therapy, valoctocogene roxaparvovec in December 2019 — the first-ever U.S. filing for approval of any hemophilia gene therapy. The BLA was based on three years of clinical data from Phase I/II trials, as well as interim Phase III trial results. An approval announcement by the following August was widely anticipated in the hemophilia community. Instead, in August 2020, FDA issued a complete response letter, indicating the submission did not provide adequate assurance of safety or efficacy. The agency cited inconsistencies between Phase I/II and pivotal studies, and asked BioMarin to provide an additional two years of data from its ongoing Phase III trial, whose last-enrolled participant will complete two years of follow-up in November 2021.

This unexpected rejection of BioMarin’s approval submission sent shock waves through the hemophilia research and patient communities. But perhaps it shouldn’t have. Available treatments for genetic disorders such as SCD, beta thalassemia or CALD are accompanied by serious adverse effects, or are risky or challenging to perform. Patients with severe SCD, for example, require frequent transfusions that result in iron overload that in turn can damage vital organs; treatment of the iron overload requires iron chelating agents that have their own toxicities. Even with regular transfusions to keep circulating red blood cells with the HbS mutation below the target level, most patients with the severe phenotype experience hospitalizations for sickle cell crises and much-shortened lifespans.4 Assuming a well-matched donor can be found, the only available treatment option for CALD is hematopoietic stem cell transplant (HSCT), which is associated with a risk of infection, graft-versus-host disease, engraftment failure and death; five-year survival following HSCT for treatment of CALD is less than 80 percent.5 

Severe hemophilia presents a much different scenario. The available treatments for hemophilia — highly concentrated factor concentrates or emicizumab — are very safe and highly effective in reducing bleeding risk when self-administered prophylactically under the guidance of a trained specialist. Treatment-compliant individuals can live relatively normal lives, with a low risk of premature death. Even instances of severe inhibitors can be managed or eradicated with early, aggressive immune tolerance induction supported with bypassing agents.6

Thus, from a regulatory perspective, investigational gene therapies for hemophilia A and B might need to meet an incrementally higher standard for approval than other gene therapies that target severe genetically based diseases for which available treatment options are problematic or offer only limited benefits.

Hemophilia A Gene Therapies in the Pipeline

The simpler structure and much smaller number of amino acids that comprise the FIX molecule helped to give investigational hemophilia B gene therapies a substantial head start in their early development. But after years of lagging behind, hemophilia A gene therapies have caught up, with three prospective treatments now in Phase II or Phase III clinical testing.

BioMarin Pharmaceuticals (valoctocogene roxaparvovec). In early 2020, a United Kingdom research team reported that all 13 adult subjects with severe hemophilia A who received the two highest doses of valoctocogene roxaparvovec (also called AAV5-hFVIII-SQ) had complete resolution of bleeding in all previously affected target joints, no bleeding events and complete cessation of prophylactic FVIII use over two to three years of follow-up. Study participants who received the two highest doses — 4 x 1013 or 6 x 1013 vector genomes per kilogram of body weight (4e13 or 6e13 vg/kg) — had a median FVIII expression of 13 percent and 20 percent of normal — well above the protective FVIII threshold level.7

Subsequently, BioMarin presented detailed findings from its Phase III GENEr8-1 study, whose 134 enrolled participants make it the largest global pivotal trial to date to evaluate any hemophilia gene therapy. At the end of the first post-infusion year, the mean endogenous FVIII expression level climbed from a baseline of 1 IU/dL to 42.9 IU/dL (median 23.9), and remained steady in a subset of 17 patients who were out at least two years from their single dose of valoctocogene roxaparvovec. In a prespecified subset of 112 patients with a mean follow-up of 72 weeks, the annualized FVIII utilization rate was reduced by 99 percent, from a mean of 3,961 IU/kg/year (median 3,754) to just 57 IU/kg/year (median 0). The mean annualized infusion rate (AIR) was correspondingly reduced by 99 percent from 136 (median 129) to 2.0 (median 0) infusions per year. The annualized bleeding rate (ABR) fell by 84 percent from 2.8 (median 2.8) bleeding episodes at baseline to 0.8 (median 0.0) episodes per year following valoctocogene roxaparvovec gene therapy (p < 0.001).

Elevated alanine aminotransferase (ALT), an enzyme whose increased blood level is generally indicative of liver cell damage, was the most common adverse event identified in study participants, about three-quarters of whom were treated with corticosteroids. The average duration on corticosteroids was 33 weeks; predictably, its use was associated with side effects that included insomnia, cushingoid changes and increased weight. But importantly, no Grade 4 ALT elevations occurred, and no participants met Hy’s law criteria for drug-induced liver injury. “The demonstrated bleed control at 52 weeks and beyond in this pivotal study supports our thesis that gene therapy can play an important role in the treatment of severe hemophilia,” said lead principal investigator Margareth C. Ozelo, MD, PhD.

BioMarin plans to resubmit a BLA for valoctocogene roxaparvovec in the second quarter of 2022, which will include FDA’s requested two-year follow-up data for all 134 subjects in the 4e13 vg/kg and 6e13 vg/kg dosage cohorts of the company’s Phase III GENEr8-1 study.

Spark Therapeutics/Roche (SPK-8011). Spark’s SPK-8011 comprises a bioengineered AAV vector containing a codon-optimized FVIII gene under the control of a liver-specific promoter. To date, Spark has enrolled fewer study participants than BioMarin, but has reported on patients who have been followed for as long as four years following vector administration. A total of 18 Phase I/II study participants have received SPK-8011 in four dose cohorts, ranging from 5 x 1011 vg/kg to 2 x 1012 vg/kg. In the 16 patients with sustained FVIII expression, the ABR and annualized FVIII infusion rates were reduced by 91 percent and 97 percent, respectively.

Interim data indicate SPK-8011 has “an acceptable safety profile,” according to the company, with no deaths and no FVIII inhibitor development with up to four years of follow-up. Two of 17 participants with more than one year of data lost FVIII expression as the result of a presumptive immune response to the AAV capsid that was unresponsive to immunosuppression. Seven participants experienced transient, asymptomatic liver function test elevations, all of which were mild or moderate and have resolved; one experienced a grade 2 transaminitis, which resolved after intravenous steroid treatment. 

Earlier this year, Spark also reported stable and durable FVIII activity at more than 52 weeks of follow-up in all four adult Phase I/II study subjects with severe hemophilia and no history of inhibitors treated with a different investigational hemophilia A gene therapy — SPK-8016 — at a comparatively low vector dose of 5 x 1011 vg/kg. 

One of the four subjects who did not require immunomodulatory agents had the highest level of FVIII activity (21.8 percent of normal), while the others received tapering doses of oral corticosteroids and steroid-sparing immunomodulatory co-therapies and maintained FVIII levels of 5.9 percent of normal or higher. Collectively, they experienced a 98 percent reduction in the AIR and an 85 percent reduction in the ABR after a follow-up of 15 months to 18 months.

Pfizer (giroctocogene fitelparvovec; SB-525). Initially developed by Sangamo Therapeutics, this recombinant AAV serotype 6 vector (AAV6) carries DNA-encoding B domain-deleted human FVIII, with an expression cassette designed for optimal liver-specific expression of the protein. 

The five participants in the high-dose 3 x 1013 vg/kg cohort of the Phase I/II Alta study had sustained very high, steady-state FVIII activity levels, with a group median FVIII activity of 56.9 percent of normal from week 9 to week 52. Beyond week 3 following infusion of SB-525, none experienced any bleeding events or required prophylactic factor within the first year. One patient had one target joint bleed requiring FVIII therapy after week 52. Four of the five participants in the high-dose cohort received oral corticosteroids for ALT elevations, all of which fully resolved.

Over the next year, a total of 63 participants with severe or moderately severe hemophilia A will be enrolled in the AFFINE Phase III open-label registrational trial to evaluate the efficacy and safety of a single infusion of SB-525; they will be followed over a five-year study period after the single infusion to further assess the ABR relative to previous FVIII prophylaxis, as well as the magnitude and durability of the FVIII activity level. 

“Given the Phase I/II study findings to date, we believe that [SB-525] has the potential to sustain factor levels and reduce annual bleed rates, suggesting this one-time gene therapy could potentially transform the standard of care for eligible patients worldwide,” said Pfizer’s rare disease chief development officer Brenda Cooperstone.

A fourth company, privately held Expression Therapeutics based in Atlanta, has developed a novel ex vivo hemophilia A gene therapy that involves harvesting autologous hematopoietic stem cells from the patient, selecting for CD34+ cells, genetically modifying them using its proprietary lentivirus-FVIII vector and, following a conditioning regimen, reinfusing the transduced CD34+ cells so they can engraft in the stem cell compartment within the bone marrow. A Phase I clinical trial in seven patients with severe hemophilia A is expected to start enrollment early in 2022.8

Hemophilia B Gene Therapies in the Pipeline

The competitive field working to introduce a hemophilia B gene therapy has recently been pared back with the discontinuation of programs sponsored by Sangamo Therapeutics (SB-FIX) and Takeda (TAK-748), leaving at least four companies with promising candidate products in the race: Pfizer, Netherlands/U.S.-based uniQure, U.K.-based Freeline and Shanghai-based Belief Biomed.

uniQure (etranacogene dezaparvovec; AMT-061). AMT-061, an enhanced construct of uniQure’s original AMT-060 gene therapy candidate, comprises an AAV5 viral vector carrying a gene cassette with the high-functioning Padua variant of FIX. uniQure and CSL Behring have entered into a commercialization and license agreement providing CSL Behring exclusive global commercialization rights to AMT-061.

Interim clinical data from all 54 hemophilia B patients enrolled in the company’s pivotal Phase III HOPE-B trial have demonstrated durable, sustained increases in FIX activity at 52 weeks following infusion of a single dose of AMT-061, with a mean of 41.5 percent of normal, compared to a mean of 39.0 percent of normal at 26 weeks of follow-up.

The ABR during the 52-week follow-up period was reduced by 80 percent to 0.68 bleeding episodes per year, from the pre-treatment baseline of 3.39 episodes per year (p < 0.0001). The annualized rate of spontaneous bleeding requiring treatment dropped by 85 percent from 1.16 at baseline to 0.18 bleeds per year during the 52-week follow-up period. FIX replacement therapy in all patients declined by 96 percent, with 52 of 54 patients successfully discontinuing their prophylactic infusions.** 

** One nonresponder received less than 10 percent of the AMT-061 dosage due to an infusion reaction, while the second nonresponder had an unusually high preexisting neutralizing antibody titer against AAV5 that is expected in less than 1 percent of the general population.

“The 52-week data show mean FIX activity in the normal range and increases our confidence in the potential durability and long-term benefits of [AMT-061], bringing us one step closer to our goal of delivering this groundbreaking therapy,” said uniQure R&D president Ricardo Dolmetsh, PhD.

Pfizer (fidanacogene elaparvovec). Fidanacogene elaparvovec is a bioengineered AAV vector utilizing a high-activity FIX transgene. In 2018, Spark Therapeutics transferred responsibility for all clinical development, regulatory and manufacturing activities to Pfizer. Now dubbed “PF-06838435,” this gene therapy is currently in a Phase III trial enrolling 55 adult hemophilia B patients with severe or moderately severe disease (residual FIX activity less than or equal to 2 percent). 

At 52 weeks following vector infusion, 15 adult hemophilia B patients in an earlier Phase I/II study experienced a mean steady-state FIX level of 22.9 percent (± 9.9 percent), with a drop in the ABR from 8.9 ± 14.0 bleeds prior to treatment to 0.4 ± 1.1 bleeds. Twelve of 15 patients reported zero bleeds over the 52 weeks following fidanacogene elaparvovec infusion. Five patients required a total of 20 factor infusions. There were no reported serious adverse events, and all hepatic transaminase elevations responded to treatment with corticosteroids.

Thus far, with up to five years of follow-up in 14 participants, fidanacogene elaparvovec appears to be generally well tolerated, according to a very recent report by the Phase I/II clinical study team. Mild sustained elevations of uncertain etiology in some study subjects continue to be monitored.

Freeline Therapeutics (FLT180a). Another AAV-based gene therapy, FLT180a has been administered to 10 patients with severe hemophilia B in four dosage cohorts. With follow-up periods ranging from 26 weeks to 104 weeks, durable, long-term elevation in FIX activity was observed for up to two years in the two patients enrolled in the lowest dose cohort (4.5e11 vg/kg); both had levels of 38 percent of normal, with no evidence of transaminitis. 

Remarkably, three patients who received a dose of 9.75e11 vg/kg had week 3 FIX activity levels of 136 percent, 82 percent and 105 percent of normal; a fourth patient had a FIX expression of just 3 percent of normal, preceded by an increase in ALT. Average FIX expression in two patients who received a dose of 1.5e12 vg/kg averaged 160 percent of normal, which was deemed to be higher than required for the potential treatment of hemophilia B.

“These data suggest that FLT180a has the potential, using relatively low doses, to create durable FIX activity levels in the normal range … and provide functional cures,” said Freeline CEO Theresa Heggie. Freeline is currently screening potential participants for enrollment in a planned Phase III clinical trial.

Belief Biomed (BBM-H901). Specific findings have not yet been released for Belief’s bioengineered AAV containing a codon-optimized human FIX gene under the control of a liver-specific promoter, which is currently in Phase I clinical development. But the company reports an investigator-initiated trial of BBM-H901 has already demonstrated “high efficacy and safety,” with stable FIX activity, a significant decline in ABR and no evidence of serious adverse events.

Not If But When

Available data suggest that several hemophilia A and B gene therapy candidates have acceptable safety profiles, consistently and durably induce protective clotting factor levels, essentially end the need for prophylactic factor replacement therapy, and dramatically reduce the risk of bleeds requiring treatment. 

While the excellent effectiveness and safety of current prophylaxis therapies make it incumbent for gene therapies to demonstrate long-term safety and efficacy, all indications are that the dream of a functional cure for severe hemophilia will soon become reality for thousands of affected individuals in the U.S. and throughout the world.


1. Li N, Sawyer EK, Maruszczyk K, et al. Adult lifetime cost of hemophilia B management in the US: payer and societal perspective from a decision analytic model. J Med Econ 2021;24(1):363-72.

2. U.S. Food and Drug Administration. FDA approval brings first gene therapy to the United States. Accessed 8/27/2021 at

3. Bluebird bio. Bluebird bio reports second quarter financial results and provides operations update, Aug. 9, 2021. Accessed 8/30/2021 at

4. Lanzkron S, Carroll CP, Haywood C. Mortality rates and age at death from sickle cell disease: 1979 – 2005. Public Health Rep 2013 Mar-Apr;128(2):110-16.

5. Raymond GV, Aubourg P, Paker A, et al. Survival and functional outcomes in boys with cerebral adrenoleukodystrophy with and without hematopoietic stem cell transplantation. Biol Blood Marrow Transplant 2019 Mar;25(3):538-48.

6. Meeks S and Batsuli G. Hemophilia and inhibitors: current treatment options and potential new therapeutic approaches. Hematology Am Soc Hematol Educ Program 2016 Dec 2;657-62.

7. Pasi KJ, Rangarajan S, Mitchell N, et al. Multiyear follow-up of AAV5-hFVIII-SQ gene therapy for hemophilia A. N Engl J Med 2020 Jan 2;382(1):29-40.

8. Expression Therapeutics. Hematopoietic Stem Cell Transplantation Gene Therapy for Treatment of Severe Hemophilia A. Accessed 8/29/2021 at

Keith Berman, MPH, MBA
Keith Berman, MPH, MBA, is the founder of Health Research Associates, providing reimbursement consulting, business development and market research services to biopharmaceutical, blood product and medical device manufacturers and suppliers. He also serves as editor of International Blood/Plasma News, a blood products industry newsletter.