AAV-based neonatal gene therapy for hemophilia A: long-term correction and avoidance of immune responses in mice

Abstract One major obstacle in gene therapy is the generation of immune responses directed against transgene product. Five consecutive anti-CD3 treatments concomitant with factor VIII (FVIII) plasmid injection prevented the formation of inhibitory antibodies against FVIII and achieved persistent, therapeutic levels of FVIII gene expression in treated hemophilia A mice. Repeated plasmid gene transfer is applicable in tolerized mice without eliciting immune responses. Anti-CD3 treatment significantly depleted both CD4+ and CD8+ T cells, whereas increased transforming growth factor-β levels in plasma and the frequency of both CD4+CD25+FoxP3+ and CD4+CD25−Foxp3+ regulatory T cells in the initial few weeks after treatment. Although prior depletion of CD4+CD25+ cells did not abrogate tolerance induction, adoptive transfer of CD4+ cells from tolerized mice at 6 weeks after treatment protected recipient mice from anti-FVIII immune responses. Anti-CD3–treated mice mounted immune responses against both T-dependent and T-independent neo-antigens, indicating that anti-CD3 did not hamper the immune systems in the long term. Concomitant FVIII plasmid + anti-CD3 treatment induced long-term tolerance specific to FVIII via a mechanism involving the increase in transforming growth factor-β levels and the generation of adaptive FVIII-specific CD4+Foxp3+ regulatory T cells at the periphery. Furthermore, anti-CD3 can reduce the titers of preexisting anti-FVIII inhibitory antibodies in hemophilia A mice.

Download Full-text

Assessing Patient Experiences with Prophylactic Treatments for Hemophilia a: Concept Elicitation for Gene Therapy

Blood ◽

10.1182/blood-2020-140307 ◽

2020 ◽

Vol 136 (Supplement 1) ◽

pp. 33-34

Author(s):

Shawn X. Sun ◽

Oyebimpe Olayinka-Amao ◽

Dana DiBenedetti

Keyword(s):

Gene Therapy ◽

Physical Health ◽

Pharmaceutical Company ◽

Prophylactic Treatment ◽

Hemophilia A ◽

Current Therapy ◽

Specific Patient ◽

Hypothetical Scenario ◽

The Impact

Background: Gene therapy for hemophilia A is designed to be a one-time infusion to deliver functional copies of the defective factor VIII (FVIII) gene, to facilitate the endogenous production of therapeutic FVIII levels. The aim is to achieve long-term protection from bleeds without the burden of regular infusions. Aims: To better understand patients' experiences of living with hemophilia A, the impact of traditional hemophilia A treatments, and patients' perceptions of the potential value of gene therapy versus traditional prophylactic treatments. Methods: Patients were identified from the database of a US rare diseases patient organization, who also recruited and screened patients for the study using materials developed by the outcomes research organization (RTI-HS) and Takeda. Adult males aged ≥18 years with a self-reported diagnosis of moderate or severe hemophilia A, who reported using factor or nonfactor prophylactic treatment and were not currently receiving treatment for inhibitors, were eligible for the study. Eligible patients provided verbal informed consent to participate in a semi-structured, 60-minute telephone interview conducted in English by 2 members of RTI-HS who did not have access to any patient-identifying information at any time during the study. Targeted questions probed perceptions of treatment burden, impact of hemophilia A on daily life, and time spent on treatment. Additionally, questions were posed to assess patients' perceptions of the impact of traditional treatments and the potential benefits they anticipate from gene therapy. Results: Nineteen patients aged 19-55 years with moderate (n = 1) or severe (n = 18) hemophilia A were interviewed. Most (16/19, 89.5%) received prophylactic FVIII therapy, (3/19, 15.8%) were receiving nonfactor prophylactic treatment, of which 1 patient also used FVIII treatment. The aspects of current or past treatments most frequently disliked by patients were lack of efficacy, frequency of infusions, intravenous administration, vein health/scar tissue, and dosing volume. Most patients expressed satisfaction with their current treatment (18/19; 94.7%), though all listed ≥1 negative treatment impact, most frequently related to difficulties with travel (13/19; 68.4%), mood/emotions (12/19; 63.2%), day-to-day activities (10/19; 52.6%), and physical health/activities (7/19; 36.8%), including having to give up or reduce particular activities because of their treatment and needing to be more cautious, especially on nontreatment days. When presented with a hypothetical scenario for gene therapy - a one-time long-acting intravenous infusion to provide a constant level of FVIII that could reduce future bleeds - all patients stated they would choose gene therapy over their current therapy, although several said they would have initial questions regarding safety, efficacy, and duration of protection. Commonly expressed reasons for preferring gene therapy (Figure 1) included fewer infusions and less worry about the need to infuse. All 19 patients said they expected to be highly satisfied with this treatment, largely because of the long-term protection from bleeds, fewer infusions, and less concern about inhibitors. The most commonly anticipated improvements were in mood/emotions (15/19; 78.9%), specifically related to reduced concern about infusions and bleed protection. Other commonly anticipated improvements included gain in time usually spent infusing (13/19; 68.4%), easier travel (12/19; 63.2%), and improved physical health and ability to perform activities (10/19; 52.6%). Conclusions: This study identified specific patient priorities, including treatment convenience, long-lasting bleed protection, frequency of intravenous infusions, and infusion volumes. The results suggest that gene therapy clinical trials should consider evaluating patient concerns in relation to the level of patient confidence in bleed protection. A study limitation is that, at the time of the survey, data on the efficacy and safety of gene therapy were limited. In the future, the study will be expanded to include a larger population of patients with hemophilia. Disclosures Sun: Takeda Pharmaceutical Company Ltd.: Current Employment. Olayinka-Amao:Takeda Pharmaceutical Company Ltd: Other: RTI-HS was contracted by Takeda Pharmaceutical Company Ltd to conduct this work; RTI Health Solutions: Current Employment. DiBenedetti:RTI Health Solutions: Current Employment; Takeda Pharmaceutical Company Ltd: Other: RTI-HS was contracted by Takeda Pharmaceutical Company Ltd to conduct this work.

Download Full-text

34. Retroviral Vector-Mediated Neonatal Gene Therapy Results in Long-Term and Therapeutic Levels of Canine Factor VIII Expression in Hemophilia A Mice and Dogs

Molecular Therapy ◽

10.1016/j.ymthe.2004.05.057 ◽

2004 ◽

Vol 9 ◽

pp. S14-S15

Keyword(s):

Gene Therapy ◽

Factor Viii ◽

Hemophilia A ◽

Retroviral Vector

Download Full-text

Animal Testing of Retroviral-Mediated Gene Therapy for Factor VIII Deficiency

Thrombosis and Haemostasis ◽

10.1055/s-0037-1615880 ◽

1999 ◽

Vol 82 (08) ◽

pp. 555-561 ◽

Cited By ~ 31

Author(s):

Douglas Jolly ◽

Judith Greengard

Keyword(s):

Gene Therapy ◽

Factor Viii ◽

Hemophilia A ◽

Joint Damage ◽

Coagulation Factor ◽

The United States ◽

Severe Hemophilia ◽

Factor Viii Gene ◽

Bleeding Episodes

IntroductionHemophilia A results from the plasma deficiency of factor VIII, a gene carried on the X chromosome. Bleeding results from a lack of coagulation factor VIII, a large and complex protein that circulates in complex with its carrier, von Willebrand factor (vWF).1 Severe hemophilia A (<1% of normal circulating levels) is associated with a high degree of mortality, due to spontaneous and trauma-induced, life-threatening and crippling bleeding episodes.2 Current treatment in the United States consists of infusion of plasma-derived or recombinant factor VIII in response to bleeding episodes.3 Such treatment fails to prevent cumulative joint damage, a major cause of hemophilia-associated morbidity.4 Availability of prophylactic treatment, which would reduce the number and severity of bleeding episodes and, consequently, would limit such joint damage, is limited by cost and the problems associated with repeated venous access. Other problems are associated with frequent replacement treatment, including the dangers of transmission of blood-borne infections derived from plasma used as a source of factor VIII or tissue culture or formulation components. These dangers are reduced, but not eliminated, by current manufacturing techniques. Furthermore, approximately 1 in 5 patients with severe hemophilia treated with recombinant or plasma-derived factor VIII develop inhibitory humoral immune responses. In some cases, new inhibitors have developed, apparently in response to unnatural modifications introduced during manufacture or purification.5 Gene therapy could circumvent most of these difficulties. In theory, a single injection of a vector encoding the factor VIII gene could provide constant plasma levels of factor in the long term. However, long-term expression after gene transfer of a systemically expressed protein in higher mammals has seldom been described. In some cases, a vector that appeared promising in a rodent model has not worked well in larger animals, for example, due to a massive immune response not seen in the rodent.6 An excellent review of early efforts at factor VIII gene therapy appeared in an earlier volume of this series.7 A summary of results from various in vivo experiments is shown in Table 1. This chapter will focus on results pertaining to studies using vectors based on murine retroviruses, including our own work.

Download Full-text