Pre-clinical Development of a Lentiviral Vector Expressing the Anti-sickling βAS3 Globin for Gene Therapy for Sickle Cell Disease

Introduction: In the United States, most children with sickle cell disease (SCD) survive into adulthood and transfer from pediatric to adult-centered care. Cognitive deficits begin during childhood and are highly prevalent among individuals with SCD, potentially affecting their functional ability to establish adult care and navigate the new adult care environment. Lack of engagement in adult care can place youth with SCD at higher risk for care discontinuity and higher disease morbidity and mortality. The relationship between cognition and transition to adult care has not been examined. We hypothesized that better performance on measures of neurocognition were associated with decreased latency in initiating adult care, greater retention in adult care, and increased utilization of adult ambulatory services. As a secondary objective, we examined the relationship of environmental outcomes to transition outcomes. Methods: We included participants enrolled in the Sickle Cell Research and Intervention Program (SCCRIP; Hankins J. et al, Pediatric Blood and Cancer 2018), a longitudinal lifetime cohort study of individuals with SCD that monitors neurocognition. Participants were included if they underwent neurocognitive screening assessment in adolescence, prior to their transfer to adult care and if they satisfied their first appointment in adult care. The neurocognitive screening battery included measures of estimated global intelligence (Wechsler Abbreviated Scales of Intelligence, 2nd Ed; WASI-2) and sustained attention (Continuous Performance Test, 2nd Ed; CPT-2). Environmental factors included the Economic Hardship Index (EHI), guardian employment status while in pediatric care, and the number of persons living in the household. Use of adult ambulatory services was measured by the number of outpatient visits per patient-year. The association between cognitive performance and the latency from pediatric to adult care, adult care retention and environmental variables was examined using the 2-sample t test if the data were normally distributed or the Wilcoxon rank-sum test otherwise. Categorical variables were analyzed with the Chi-square test or Fisher's exact test. Transition outcomes were also analyzed as continuous variables using univariate linear regression. All reported p-values are two-sided. Results: Eighty adolescents with SCD ages 15-18 years at the time of their cognitive assessment (58% male, 63% HbSS/HbSβ0-thalassemia) were included; most transferred <6 months from the last pediatric visit Table 1). Of these 80 patients, 61 and 43 had sufficient follow-up time to examine their retention in adult care 12 and 24 months after transfer, respectively. Fifty out of the 61 patients (82%) remained in adult care > 12 months, and 31 of the 43 (72%) remained in adult care >24 months after their first adult visit. Higher Full-Scale IQ was associated with establishing adult care ≤2 months from last pediatric visit (Table 1; Figure 1A, 1B). Belonging to families with fewer children, smaller households and a higher WASI-2 Verbal Comprehension Index were associated with establishing adult care ≤6 months from last pediatric visit. Better CPT-2 Commissions performance (less attention deficit) was associated with increased adult care retention at 12 and 24 months (Table 2; Figure 1C,1D). Having a working guardian was associated with less retention at 12 months (p=0.01), whereas having an unemployed primary guardian was associated with greater retention at 24 months (p=0.02). Further, an employed guardian was associated with greater utilization of adult ambulatory services (p=0.01). EHI was not significantly related to transition outcomes. No relationship was found between adult ambulatory services and neurocognitive assessment. Conclusion: Neurocognitive deficit (lower IQ and attention deficits) may decrease short and long-term engagement in adult care among youth with SCD as demonstrated by longer latency periods between pediatric and adult care and shorter adult care retention. Socio-economic factors may also play a role in transition outcomes but require further investigation. Investigation of disease modifying therapies that preserve cognitive function should be prioritized. Interventions that account for patients' cognitive level and their environment should be considered in the individualization of transition plans. Disclosures King: Magenta Therapeutics: Membership on an entity's Board of Directors or advisory committees; Novimmune: Research Funding; Amphivena Therapeutics: Research Funding; Incyte: Consultancy; Tioma Therapeutics (formerly Vasculox, Inc.):: Consultancy; Cell Works: Consultancy; Bioline: Consultancy; Celgene: Consultancy; RiverVest: Consultancy; WUGEN: Equity Ownership. Wang:Agios Pharmaceuticals: Consultancy; Novartis: Consultancy. Zhao:MBIO: Other: St. Jude Children's Research Hospital has an existing exclusive license and ongoing partnership with Mustang Bio for the further clinical development and commercialization of this XSCID gene therapy. Kang:MBIO: Other: St. Jude Children's Research Hospital has an existing exclusive license and ongoing partnership with Mustang Bio for the further clinical development and commercialization of this XSCID gene therapy. Hankins:National Committee for Quality Assurance: Consultancy; NHLBI: Research Funding; Global Blood Therapeutics: Research Funding; Novartis: Research Funding; LYNKS Foundation: Research Funding; NHLBI: Honoraria; ASPHO: Honoraria; Bluebird Bio: Consultancy.

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Myelodysplastic syndrome unrelated to lentiviral vector in a patient treated with gene therapy for sickle cell disease

Blood Advances ◽

10.1182/bloodadvances.2019001330 ◽

2020 ◽

Vol 4 (9) ◽

pp. 2058-2063

Author(s):

Matthew M. Hsieh ◽

Melissa Bonner ◽

Francis John Pierciey ◽

Naoya Uchida ◽

James Rottman ◽

...

Keyword(s):

Gene Therapy ◽

Sickle Cell Disease ◽

Adverse Events ◽

Myelodysplastic Syndrome ◽

Sickle Cell ◽

Lentiviral Vector ◽

Cell Disease ◽

Novel Treatments ◽

Key Points

Key Points Ability to accurately attribute adverse events post–gene therapy is required to describe the benefit-risk of these novel treatments. A SCD patient developed myelodysplastic syndrome post-LentiGlobin treatment; we show how insertional oncogenesis was excluded as the cause.

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Safety and Efficacy of Gene Therapy of the Sickle Cell Disease by Transplantation of an Autologous CD34+ Enriched Cell Fraction That Contains CD34+ Cells Transduced ex Vivo With the GLOBE1 Lentiviral Vector Expressing the βAS3 Globin Gene in Patients With Sickle Cell Disease (DREPAGLOBE)

Case Medical Research ◽

10.31525/ct1-nct03964792 ◽

2019 ◽

Author(s):

Keyword(s):

Gene Therapy ◽

Sickle Cell Disease ◽

Sickle Cell ◽

Lentiviral Vector ◽

Ex Vivo ◽

Globin Gene ◽

Cell Fraction ◽

Cell Disease ◽

Safety And Efficacy ◽

Cd34 Cells

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A Study Evaluating Gene Therapy With BB305 Lentiviral Vector in Sickle Cell Disease

Case Medical Research ◽

10.31525/ct1-nct04293185 ◽

2020 ◽

Author(s):

Keyword(s):

Gene Therapy ◽

Sickle Cell Disease ◽

Sickle Cell ◽

Lentiviral Vector ◽

Cell Disease

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Increasing Transduction Efficiency in Human Hematopoietic Stem Cells for Gene Therapy

Blood ◽

10.1182/blood.v120.21.2052.2052 ◽

2012 ◽

Vol 120 (21) ◽

pp. 2052-2052

Author(s):

Kismet M Baldwin ◽

Fabrizia Urbinati ◽

Zulema Romero-Garcia ◽

Donald B. Kohn

Keyword(s):

Stem Cells ◽

Gene Therapy ◽

Clinical Trials ◽

Sickle Cell Disease ◽

Sickle Cell ◽

Lentiviral Vector ◽

Transduction Efficiency ◽

Cell Disease ◽

Hematopoietic Stem ◽

Cd34 Cells

Abstract Abstract 2052 Background: Sickle cell disease (SCD) is a multisystem disease, associated with severe episodes of acute illness and progressive organ damage. Currently, the only curative treatment is allogeneic hematopoietic stem cell transplant (HSCT); however, this is limited by availability of HLA compatible donors and by immunological complications of graft rejection or graft-versus-host disease. Autologous stem cell gene therapy for SCD has the potential to treat this illness without the immune suppression needed for current allogeneic HSCT approaches. Previous studies have demonstrated that addition of a β-globin gene, modified to have the anti-sickling properties of fetal (γ-) globin (βAS3), to bone marrow (BM) stem cells in murine models of SCD normalizes RBC physiology and prevents the manifestations of sickle cell disease (Levasseuer Blood 102:4312–9, 2003). Initial evidence for the efficacy of the modification of human SCD BM CD34+ cells with the βAS3lentiviral (LV) vector for gene therapy of sickle cell disease has been demonstrated in our lab. However, this complex lentiviral vector is produced at a sub-optimal titer and large production batches would be needed to supply clinical trials. Hypothesis: Although, it has been proven that the βAS3 gene can be transduced into CD34+ hematopoietic stem/progenitor cells (HSPC), the transduction efficiency is still not optimal. The CD34+ cell population includes rare long-lived stem cells but also more abundant progenitors, which would be short-lived after transplant. We hypothesize that isolating the more primitive HSPC population (CD34+/CD38− cells approximately 1% of all CD34+ cells) and transducing them with the βAS3 lentiviral vector will increase transduction efficiency and greatly reduce vector needs. Methods: CD34+/CD38− cells were isolated from cord blood (CB) CD34+ cells obtained from healthy donors by fluorescence activated cell sorting (FACS) and transduced with the CCL.βAS3.FB LV vector. After 14 days in culture, vector copy number (VCN) was determined by qPCR. Isolation of a more primitive cell was confirmed via long term culture (LTC) assay for 90 days. At 2–3 weeks intervals, non-adherent cell number was obtained, VCN was analyzed and CFU assays were performed to assess their capability to fully maintain their hematopoietic potential after transduction. Results: CD34+/CD38− cells were effectively isolated using FACS (n=7; 6,329–33,742 cells; 34–99% theoretical yield). The isolated CD34+/CD38- cells were able to generate progeny over an extended period of LTC compared to the CD34+ cells whose cell expansion declined ∼60 days in culture. CFU assays demonstrated that βAS3 gene-modified CB CD34+/CD38- cells were fully capable of maintaining their hematopoietic potential. The isolated CD34+/CD38- cells required 3–40 fold less vector for transduction compared to an equivalent number of these cells contained within the larger, non-fractionated CD34+ preparations. Transduction of CD34+/CD38- cells measured at day 14, by qPCR, was improved relative to CD34+ cells, mean VCN 2.5, +/− SEM 0.33 (range 2–3.5) vs. VCN 1.3, +/− 0.40 (range 0.5–2), respectively (p=0.03). In LTC, VCN remained higher over time in the CD34+/CD38- cells compared to the CD34+ cells, mean VCN 2.0, +/− SEM 0.13 (range 1.6–2.7) vs. VCN 0.5, +/− 0.09 (range 0.2–0.9) respectively. In vivo studies are ongoing to investigate the transduction efficiency of stem/progenitor cells engrafting from CD34+ and CD34+/CD38- cells transplanted in the NSG mouse model. Immunomagnetic isolation of CD34+/CD38- cells using columns is underway in anticipation of potential use in future clinical trials. Further investigations into the mechanisms for increased transduction in the CD34+/CD38- cells are ongoing. Conclusions: This work provides initial evidence for the beneficial effects from isolating human CB CD34+/CD38− cells to improve transduction with the βAS3LV vector for gene therapy of sickle cell disease. Disclosures: No relevant conflicts of interest to declare.

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