ALBERT

Description: We report preliminary results in 3 children with cerebral X-linked adrenoleukodystrophy (ALD) who received in September 2006, January 2007 and June 2008 lentiviral vector transduced autologous hematopoietic stem cell (HSC). We have previously demonstrated that cerebral demyelination associated with cerebral ALD can be stopped or reversed within 12–18 months by allogeneic HSC transplantation. The long term beneficial effects of HCT transplantation in ALD are due to the progressive turn-over of brain macrophages (microglia) derived from bone-marrow cells. For the current HSC gene therapy procedure, we used mobilized peripheral blood CD34+ cells that were transduced ex vivo for 18 hours with a non-replicative HIV1-derived lentiviral vector (CG1711 hALD) at MOI25 and expressing the ALD cDNA under the control of the MND (myeloproliferative sarcoma virus enhancer, negative control region deleted, dl587rev primer binding site substituted) promoter, and in the presence of 4 human recombinant cytokines (Il- 3, Stem Cell Factor [SCF], Flt3-ligand and Megakaryocyte Growth and Differentiation Factor [MGDF]) and CH-296 retronectine. Transduced cells were frozen to perform the required (RCL) safety tests. After thawing and prior to reinjection, 50%, 30% and 40% of transduced CD34+ cells expressed the ALD protein with a mean of 0.7, 0.6 and 0.65 copies of integrated provirus per cell. Transduced CD34+ cells were infused to ALD patients after a conditioning regimen including full doses of cyclophosphamide and busulfan. Hematopoietic recovery occured at day 13–15 post-transplant and the procedure was uneventful. In patient P1 and P2, the percentage of lymphocytes and monocytes expressing the ALD protein declined from day 60 to 6 months after gene therapy (GT) and remained stable up to 16 months post-GT. In P1, 9 to 13% of CD14+, CD3+, CD19+ and CD15+ cells expressed ALD protein 16 months post-transplant. In P2 and at the same time-point after transplant, 10 to 18% of CD14+, CD3+, CD19+ and CD15+ cells expressed ALD protein. ALD protein was expressed in 18–20% of bone marrow CD34+ cells from patients P1 and P2, 12 months post-transplant. In patient P3, 20 to 23% of CD3+, CD14+ and CD15+ cells expressed ALD protein 2 months after transplant. Tests assessing vector-derived RCL and vector mobilization were negative up to the last followups in the 3 patients. Integration of the vector was polyclonal and studies of integration sites arein progress. At 16 months post-transplant, HSC gene therapy resulted in neurological effects comparable with allogeneic HSC transplantation in patient P1 and P2. These results support that: ex-vivo HSC gene therapy using HIV1-derived lentiviral vector is not associated with the emergence of RCL and vector mobilization; a high percentage of hematopoietic progenitors were transduced expressing ALD protein in long term; no early evidence of selective advantage of the transduced ALD cells nor clonal expansion were observed. (This clinical trial is sponsored by Institut National de la Santé et de la Recherche Médicale and was conducted in part under a R&D collaboration with Cell Genesys, Inc., South San Francisco, CA)

Description: Background: Hemophagocytic lymphohistiocytosis (HLH) syndrome is due to an abnormal and sustained activation of T lymphocytes and macrophages releasing pro-inflammatory cytokines in the microenvironment. HLH is characterized by clinical and biological features that includes fever, hepatomegaly, splenomegaly, high ferritin level, pancytopenia and liver biology abnormalities. HLH is commonly divided into genetic/hereditary cases (HLHg) and non genetic/ sporadic (HLHs) with no hints for an inherited causes. However, similarity of both clinical symptoms and biological abnormalities between HLHg and HLHs raised, however, the hypothesis that HLHs could have a genetic component that may be related to HLHg. However contribution of genetic factors in both, pathophysiology and prognosis of sporadic hemophagocytic lymphohistiocytosis (HLHs) remains unknown. To explore the hypothesis of a genetic susceptibility in HLHs, from 2010 to 2017, we constituted a cohort of 205 HLHs patients (adults and children) and analysed genes involved in genetic HLH (HLHg). Methods: Patients fulfillied modified HLH-2004 diagnostic criteria. Data collected included (i) clinical and biological features, (ii) associated diseases (AID), (iii) infectious events, (iv) treatment, and (v) clinical outcome during one year of follow up. HLHg related genes LYST, PRF1, UNC13-D, STX11, STXBP2, RAB27A, SH2D1A, XIAP were NGS sequenced in 172 patients. Selected variants were (i) unknown but met at least two criteria among MAF10, and/or Polyphen HDIV (〉0.485) (n=50), (ii) previously related to HLHg (n=2), PRF1 A755G/N252S (MAF 80%), or no etiology (idiopathic) (n=64) with a greater infectious diversity. Fifty-two variants met the selection criteria, they were distributed overall in 115 alleles in 84/172 patients. Variants were globally heterozygous. They were recurrent with occurrence in 2 patients for STX11, RAB27A and LYST genes to 10 patients for the N252S and 18 patients for the p.Ala91Val in PRF1. Overall the frequency of individuals exhibiting at least one variant was significantly in excess (48.8%, n=84) in the HLHs versus in in-house (35.9%, n=1826) (p=0.001, CAST test) or versus in 1000G population patients (38.6%, n=194) (p=0.02, CAST test). Importantly, the distribution of patients carrying either 0, 1, 2, 3 or 4 mutated alleles in HLHs cohort was statistically different from the two control populations (Trend test: HLH vs in-house p