Lentiviral vector transduced autologous CD34+ cells with FVIII transgene for gene therapy of Hemophilia A with history of inhibitors

Abstract

Abstract There has been no report demonstrating measurable plasma FVIII after gene therapy for hemophilia A with history of inhibitors. We have previously reported lentiviral vector (LV) transduced autologous CD34+ cell-based gene therapy for hemophilia A without inhibitors. (N Engl J Med 2025; 392: 450-7) This report describes the first application of the same technology in a patient with hemophilia A after immune tolerance induction for inhibitors. The patient, a 31-years old man, with severe hemophilia A diagnosed at the age of 5 years, had been on episodic replacement therapy with clotting factor concentrates (>150 exposures) with no inhibitors. His past history was significant for treatment for possible tuberculosis in 2016 when a lymph node biopsy had shown granulomatous inflammation without acid fast bacilli and a negative GeneXpert® MTB/RIF assay. He was first consented for this LV vector transduced autologous CD34+ cell-based gene therapy in Sept, 2022. Mobilized peripheral blood CD34+ cells were collected in November 2022 and the transduced therapeutic product was prepared without any transduction enhancer, as described earlier (N Engl J Med 2025; 392: 450-7). The vector copy number (VCN) in the therapeutic product was 1.3. On the first day of his conditioning therapy in November 2022 for hematopoietic stem cell transplantation (HSCT) for gene therapy, the FVIII recovery assay showed an unexpectedly low level of 11% after a correction aimed at 100%. Inhibitor testing revealed presence of 2BU of FVIII inhibitors. The conditioning protocol was aborted. Daily infusion of FVIII was continued for two weeks but the FVIII recovery remained extremely low at <5% after corrections aimed at >50% FVIII activity. He was then started on immune tolerance induction (ITI) with 25 IU/kg of extended half-life FVIII (Eloctate®) three times a week in December, 2022. For a trauma related oral bleeding in January, 2023, he was given one dose of emicizumab 3mg/kg with which his bleeding stopped within 24 hours. It took nearly 16 months of ITI for FVIII recovery to normalize and FVIII inhibitors to become negative. After scientific review and repeat ethical approvals for proceeding with the planned gene therapy as the best long-term option for him, the patient and family were counselled again and reconsented. Gene therapy was undertaken in June, 2024. The conditioning protocol for the HSCT consisted of myeloablative doses of treosulfan along with fludarabine followed by infusion of the transduced CD34+ gene therapy product cryo-preserved from November, 2022. The cell dose was 5.19x106 CD34+ cells/kg. There were no major complications during the peri-HSCT period with neutrophil engraftment on day +10 and platelet engraftment on day +12. The duration of severe neutropenia was 8 days and that of severe thrombocytopenia was 5 days. The peripheral blood counts were normal by day +17. He had an episode of hypotension without fever on day +4 of HSCT which was attributed to hypocortisolism after exclusion of possible sepsis as the cause including a sterile blood culture. Replacement doses of prednisolone corrected the hypotension. He remained afebrile throughout the HSCT period. There was no mucositis. Exogenous FVIII replacement was stopped on day +13. A week later on day +20, a FVIII activity of ~3% could be measured in plasma. Plasma FVIII activity gradually increased to ~8% at 6 weeks, ~11% at 3 months and ~17% at 6 months after gene therapy. During follow-up from the 7th to the 12th month, the plasma FVIII activity has been maintained between 20-25% with good concordance between the one-stage and chromogenic assays. Inhibitors assays have remained negative. He has had no bleeding since gene therapy or any treatment associated adverse events since discharge from the hospital more than 1 year ago. This case demonstrates the first successful sustained expression of measurable FVIII activity in plasma after gene therapy for hemophilia A with history of inhibitors which had resolved after immune tolerance induction. Gene therapy with LV transduced autologous CD34+ cells technology holds promise for hemophilia A patients with inhibitors and needs to be further explored.