D J Gottlieb

Westmead Hospital, Sydney, New South Wales, Australia

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Publications (112)483.5 Total impact

  • [Show abstract] [Hide abstract]
    ABSTRACT: Protocols for the production of CD19-specific chimeric antigen receptor (CAR19) T cells are often complex and expensive because of the use of retroviral and lentiviral vectors or the need for CAR19 T-cell enrichment. We aimed to simplify the generation of CAR19 T cells from the peripheral blood of normal donors and patients using the piggyBac transposon system of gene modification. We varied electroporation voltage, cytokines and stimulation conditions for the generation and expansion of CAR19 T cells over a 3-week culture period. Using optimized electroporation voltage, interleukin-15 alone and co-culturing CAR T cells with peripheral blood mononuclear cells, we were able to expand CAR19 T-cell cultures by up to 765-fold over 3 weeks in normal donors and 180-fold in patients with B-cell malignancies. Final median CAR19 expression of 72% was seen in normal donors, and 81% was seen in patients with acute lymphoblastic leukaemia, chronic lymphocytic leukemia or non-Hodgkin lymphoma. CAR19 T cells produced interferon gamma on stimulation with CD19(+) cell lines and efficiently lysed both CD19(+) cell lines and primary leukemia cells. In addition, combining CAR expression with an inducible caspase safety switch allowed elimination of CAR19 T cells by the application of a small molecule dimerizer. We have produced a simple, inexpensive and easily adoptable protocol for the generation of CAR19 T cells suitable for use in clinical trials using the piggyBac transposon system. This provides a robust platform for further enhancing the T-cell product and testing new CAR technologies. Copyright © 2015 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.
    Cytotherapy 07/2015; DOI:10.1016/j.jcyt.2015.05.013 · 3.10 Impact Factor
  • Cytotherapy 06/2015; 17(6). DOI:10.1016/j.jcyt.2015.03.312 · 3.10 Impact Factor
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    ABSTRACT: We compared outcomes for 2 retrospective cohorts of patients undergoing reduced-intensity conditioning (RIC) therapy transplants using haploidentical related donors and post-transplant prophylaxis against graft-versus-host disease (GVHD) with high-dose cyclophosphamide, tacrolimus, and mycophenolate. The first cohort of 13 was transplanted with bone marrow (BM) as the stem cell source, whereas the second cohort of 23 used peripheral blood stem cells (PBSCs) mobilized with granulocyte colony-stimulating factor. The BM cohort received a single 60-mg/kg dose of cyclophosphamide on day +3, whereas the PBSC cohort received 2 doses on days +3 and +4. Patients in the first cohort were slightly older and had a higher proportion of acute myeloid leukemia, but there were no differences in the distribution of Disease Risk Index scores between the 2 groups. Patients in the PBSC group received double the number of CD34+ cells in the stem cell graft. Times to neutrophil and platelet recovery were not different between the 2 groups. Three patients, all in the PBSC group, failed to engraft but recovered with autologous hemopoiesis and survived. The 6-month cumulative incidences of acute GVHD were 55.1% for BM and 48.5% for PBSCs (P = .651), whereas 24-month cumulative rates for chronic GHVD were 28.6% for BM and 32.3% for PBSCs (P = .685). Only 2 patients, both in the BM group, died of nonrelapse causes, both of second cancers. The 2-year cumulative incidences of relapse were 43.9% for BM and 23.5% for PBSCs (P = .286). Overall survival at 2 years was significantly better for PBSC patients (P = .028), at 83.4% versus 52.7% for BM. Relapse-free and event-free survival did not differ significantly between BM and PBSC groups. In this retrospective analysis, we conclude that the use of PBSCs for haploidentical RIC transplants is a feasible strategy, with equivalent rates of acute and chronic GVHD and risk of relapse and low nonrelapse mortality compared with BM.
    Biology of blood and marrow transplantation: journal of the American Society for Blood and Marrow Transplantation 06/2015; DOI:10.1016/j.bbmt.2015.06.006 · 3.35 Impact Factor
  • E. Blyth · L. Clancy · D. Gottlieb
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    ABSTRACT: Infection remains a large contributor to mortality post-allogeneic haemopoietic stem cell transplantation (HSCT). Non-bacterial pathogens cause disease post-HSCT due to impaired cellular immunity that persists for months to years. Treatment with antiviral or antifungal pharmacotherapy is problematic due to drug toxicity, treatment failure and high cost. Pathogen-specific T cells (PSTs) from transplant donors provide a potential solution to the problem of impaired recipient immunity by directly and rapidly reconstituting immunity, thereby preventing or controlling infections. A number of phases I and II clinical trials of PSTs that have focused primarily on cytomegalovirus, Epstein–Barr virus and adenovirus have now been performed in the prophylactic, pre-emptive and treatment settings. These trials show that transferred cells are safe and that they expand in vivo, respond to viral antigens and can prevent or control disease. At the current time, manufacture remains the domain of large centres with the necessary expertise and infrastructure, but automation and standardization will allow techniques to be more widely adopted. It is clear that adoptive T cell transfer will form part of the future of transplantation, with the current challenge being to perform large randomized clinical trials demonstrating safety and efficacy to justify the investment in infrastructure required for widespread incorporation of this therapy into routine clinical practice.
    ISBT Science Series 04/2015; 10(S1). DOI:10.1111/voxs.12130
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    ABSTRACT: Cytomegalovirus (CMV) is a significant cause of morbidity, mortality and graft loss in solid organ transplantation (SOT). Treatment options for ganciclovir-resistant CMV are limited. We describe a case of ganciclovir-resistant CMV disease in a renal transplant recipient manifested by thrombotic microangiopathy-associated glomerulopathy. Adoptive T cell immunotherapy using CMV-specific T cells from a donor bank was used as salvage therapy. This report is a proof-of-concept of the clinical and logistical feasibility of this therapy in SOT recipients. © Copyright 2015 The American Society of Transplantation and the American Society of Transplant Surgeons.
    American Journal of Transplantation 02/2015; 15(3). DOI:10.1111/ajt.13023 · 6.19 Impact Factor
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    ABSTRACT: High dose cyclophosphamide given early after allogeneic hemopoietic cell transplantation has been shown to be effective prophylaxis against graft-versus-host disease (GVHD) in the setting of HLA-matched myeloablative bone marrow grafts, allowing avoidance of long-term immunosuppression with calcineurin inhibitors in some patients. Whether this approach is feasible using G-CSF-mobilized peripheral blood stem cell grafts is unknown. We conducted an exploratory Phase 2 trial of cyclophosphamide given at 50 mg/kg IV on days 3 and 4 post-transplant as sole GVHD prophylaxis in recipients of G-CSF-mobilized peripheral blood stem cell grafts from HLA-matched related or unrelated donors after reduced intensity conditioning therapy with fludarabine, carmustine and melphalan. Five patients aged 52 to 67 years with high risk hematologic malignancies were enrolled. Four of the 5 developed severe acute GVHD of grades 3 to 4, requiring treatment with methylprednisolone and cyclosporine; 3 were steroid refractory and were given salvage therapy. One of these 4 patients died of hepatic GVHD, one died of sepsis, and 2 survive. We conclude that post-transplant cyclophosphamide is inadequate as sole GVHD prophylaxis in the context of peripheral blood reduced intensity conditioning transplants from HLA-matched donors. This trial is registered at ACTRN12613001154796. Copyright © 2015 American Society for Blood and Marrow Transplantation. Published by Elsevier Inc. All rights reserved.
    Biology of blood and marrow transplantation: journal of the American Society for Blood and Marrow Transplantation 01/2015; 21(5). DOI:10.1016/j.bbmt.2015.01.020 · 3.35 Impact Factor
  • D C Bishop · A J Johnston · J M W Kwan · V Antonenas · D J Gottlieb
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    ABSTRACT: Elderly patients with acute myeloid leukaemia (AML) have a poor prognosis with standard chemotherapy. Two elderly AML patients treated with infusion of family-derived partially human leukocyte antigen (HLA)-matched peripheral blood stem cells following each cycle of chemotherapy entered morphological complete remission without graft versus host disease or major toxicity. Our results support this as a non-toxic approach for inducing a graft versus leukaemia effect in patients not suitable for allogeneic transplantation. Additional resources required for donor assessment and harvest may be reduced by using banked partially HLA-matched mononuclear cells from unrelated donors.
    Internal Medicine Journal 10/2014; 44(10):1038-40. DOI:10.1111/imj.12551 · 1.70 Impact Factor
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    ABSTRACT: Regulation of the lectin galectin 9 (Gal-9) was investigated for the first time during human cytomegalovirus (HCMV) infection. Gal-9 transcription was significantly upregulated in transplant recipients with reactivated HCMV in vivo. In vitro, Gal-9 was potently upregulated by HCMV independently of viral gene expression, with interferon beta (IFN-β) identified as the mediator of this effect. This study defines an immunoregulatory protein potently increased by HCMV infection and a novel mechanism to control Gal-9 through IFN-β induction.
    Journal of Virology 07/2014; 88(18). DOI:10.1128/JVI.01259-14 · 4.65 Impact Factor
  • Seminars in Thrombosis and Hemostasis 02/2014; 40(1):1-4. DOI:10.1055/s-0033-1364205 · 3.69 Impact Factor
  • TRANSPLANTATION; 01/2014
  • Peter Mollee · Patricia Renaut · David Gottlieb · Hugh Goodman
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    ABSTRACT: Amyloidosis is a rare but devastating condition caused by deposition of misfolded proteins as aggregates in the extracellular tissues of the body, leading to impairment of organ function. High clinical suspicion is required to facilitate early diagnosis. Correct identification of the causal amyloid protein is absolutely crucial for clinical management in order to avoid misdiagnosis and inappropriate, potentially harmful treatment, to assess prognosis and to offer genetic counselling if relevant. This review summarises the current evidence on which the diagnosis and subtyping of amyloidosis is based, outlines the limitations of various diagnostic techniques particularly in an Australian and New Zealand context, and discusses optimal strategies for the diagnostic approach to these patients. Recommendations are provided for when to particularly suspect amyloidosis, what investigations are required, as well as an approach to accurate subtyping of amyloidosis.
    Internal Medicine Journal 09/2013; 44(1). DOI:10.1111/imj.12288 · 1.70 Impact Factor
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    ABSTRACT: Several human cytomegalovirus (HCMV) genes encode products that modulate cellular functions in a manner likely to enhance viral pathogenesis. This includes UL111A, which encodes homologs of human interleukin-10 (hIL-10). Depending upon signals received, monocytes and macrophages become polarized to either classically activated (M1 pro-inflammatory) or alternatively activated (M2 anti-inflammatory) subsets. Skewing of polarization towards an M2 subset may benefit the virus by limiting the pro-inflammatory responses to infection and so we determined whether HCMV encoded viral IL-10 influenced monocyte polarization. Recombinant viral IL-10 protein polarized CD14(+) monocytes towards an anti-inflammatory M2 subset with an M2c phenotype, as demonstrated by high expression of CD163 and CD14, and suppression of MHC class II. Significantly, in the context of productive HCMV infection, viral IL-10 produced by infected cells polarized uninfected monocytes towards an M2c phenotype. We also assessed the impact of viral IL-10 on heme oxygenase 1 (HO-1), which is an enzyme linked with suppression of inflammatory responses. Polarization of monocytes by viral IL-10 resulted in upregulation of HO-1 and inhibition of HO-1 function resulted in a loss of capacity of viral IL-10 to suppress TNF-α and IL-1β, implicating HO-1 in viral IL-10 induced suppression of pro-inflammatory cytokines by M2c monocytes. In addition, a functional consequence of monocytes polarized with viral IL-10 was a decreased capacity to activate CD4(+) T cells. This study identifies a novel role for viral IL-10 in driving M2c polarization which may limit virus clearance by restricting pro-inflammatory and CD4(+) T cell responses at sites of infection.
    Journal of Virology 07/2013; 87. DOI:10.1128/JVI.00912-13 · 4.65 Impact Factor
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    ABSTRACT: Uncontrolled cytomegalovirus (CMV) reactivation after allogeneic hematopoietic stem cell transplantation causes significant morbidity and mortality. Adoptive transfer of CMV-specific cytotoxic T lymphocytes (CTLs) is a promising therapy to treat reactivation and prevent viral disease. In this article, we describe the generation of clinical-grade CMV-specific CTLs directly from granulocyte colony-stimulating factor–mobilized hemopoietic progenitor cell (G-HPC) products collected for transplantation. This method requires less than 2.5% of a typical G-HPC product to reproducibly expand CMV-specific CTLs ex vivo. Comparison of 11 CMV CTL lines generated from G-HPC products with 52 CMV CTL lines generated from nonmobilized peripheral blood revealed similar expansion kinetics and phenotype. G-HPC–derived CTLs produced IFN-γ after reexposure to CMVpp65 antigen and exhibited CMV-directed cytotoxicity but no alloreactivity against transplantation recipient–derived cells. Seven patients received CMV-specific CTL lines expanded from G-HPC products in a prophylactic adoptive immunotherapy phase I/II clinical trial. Use of G-HPC products will facilitate integration of CTL generation into established quality systems of transplantation centers and more rapid inclusion of T cell therapies into routine clinical care.
    Biology of blood and marrow transplantation: journal of the American Society for Blood and Marrow Transplantation 05/2013; 19(5):725–734. DOI:10.1016/j.bbmt.2013.01.021 · 3.35 Impact Factor
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    ABSTRACT: BACKGROUND: Hematopoietic stem cell (HSC) transplantation using bone marrow and peripheral blood stem cells is a lifesaving treatment for patients with leukemia or other blood disorders. However, donors face the risk of physical and psychosocial complications. We aimed to synthesize qualitative studies on the experiences and perspectives of HSC donors. METHODS: We searched MEDLINE, Embase, PsycINFO, CINAHL, Google Scholar, and reference lists of relevant articles to 13th November 2012. Thematic synthesis was used to analyze the findings. RESULTS: Thirty studies involving 1552 donors were included. The decision to donate included themes of: saving life, family loyalty, building a positive identity, religious conviction, fear of invasive procedures, and social pressure and obligation. Five themes about the donation experience were identified: mental preparedness (pervasive pain, intense disappointment over recipient death, exceeding expectations and valuing positive recipient gains); burden of responsibility (striving to be a quality donor, unresolved guilt, and exacerbated grief); feeling neglected (medical dismissiveness and family inattention); strengthened relationships (stronger family ties, establishing blood bonds); and personal sense of achievement (satisfaction and pride, personal development, hero status and social recognition). CONCLUSIONS: Although HSC donation was appreciated as an opportunity to save life, some donors felt anxious and unduly compelled to donate. HSC donors became emotionally invested and felt responsible for their recipient's outcomes, and were profoundly grieved and disappointed if the transplant was unsuccessful. To maximize donor satisfaction and mitigate the psychosocial risks for HSC donors, strategies to address the emotional challenges of anxiety, sense of coercion, guilt, and grief in donors are warranted.
    Biology of blood and marrow transplantation: journal of the American Society for Blood and Marrow Transplantation 04/2013; 19(7). DOI:10.1016/j.bbmt.2013.04.012 · 3.35 Impact Factor
  • Cytotherapy 04/2013; 15(4):S53. DOI:10.1016/j.jcyt.2013.01.207 · 3.10 Impact Factor
  • Cytotherapy 04/2013; 15(4):S24–S25. DOI:10.1016/j.jcyt.2013.01.092 · 3.10 Impact Factor
  • Cytotherapy 04/2013; 15(4):S38. DOI:10.1016/j.jcyt.2013.01.142 · 3.10 Impact Factor
  • C.K. Ma · L. Clancy · D. Gottlieb
    Cytotherapy 04/2013; 15(4):S38. DOI:10.1016/j.jcyt.2013.01.144 · 3.10 Impact Factor
  • Cytotherapy 04/2013; 15(4):S10. DOI:10.1016/j.jcyt.2013.01.034 · 3.10 Impact Factor
  • M M Sartor · D J Gottlieb
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    ABSTRACT: Levels of residual disease (RD) are an independent predictor of progression-free survival (PFS) and overall survival (OS) in patients treated for chronic lymphocytic leukemia (CLL). We modified the international standardized approach (ISA) to RD detection using flow cytometry by developing a single tube 10 color antibody assay. METHOD: A single tube incorporated the following monoclonal antibodies: CD81FITC, CD22PE, CD3ECD, CD5PercP5.5, CD20PECY7, CD79bAPC, CD38A700, CD43APC Alexa750, CD19eFluor 450, and CD45KO. A modified ISA gating strategy was developed that removed contaminating events. Sensitivity assays were performed using dilution with normal peripheral blood and bone marrow. Clinical samples were compared using the ISA and the single tube assay. RESULTS: Dilution studies showed that sensitivity of 0.001% was achievable when a minimum of 1.8 × 10(6) total events were acquired. One hundred twenty-nine samples were analyzed and showed RD levels from 0.003 to 22%. In 80 samples analyzed with both assays, there was an excellent correlation between the two methods (slope = 1.0, intercept = 0.07 and R2 = 0.992) and results from Bland-Altman analysis showed a bias of 0.04 ± 0.38 with 95% confidence interval of -0.71 to 0.79. Removal of contaminating events in the single tube assay led to a significant reduction in RD values (P = 0.0014). CONCLUSION: The single tube 10-color assay for the detection of RD in CLL provides equivalent results to the ISA but requires fewer cells, uses fewer reagents, and allows for simpler analysis. By directly removing contaminating events, it improves the accuracy of CLL RD detection and may reclassify the status of some patients following chemotherapy. © 2012 International Clinical Cytometry Society.
    Cytometry Part B Clinical Cytometry 03/2013; 84B(2). DOI:10.1002/cyto.b.21067 · 2.28 Impact Factor

Publication Stats

2k Citations
483.50 Total Impact Points

Institutions

  • 1993–2015
    • Westmead Hospital
      • • Department of Haematology
      • • Department of Medicine
      Sydney, New South Wales, Australia
  • 2000–2014
    • Westmead Millennium Institute
      Sydney, New South Wales, Australia
  • 1997–2013
    • University of Sydney
      Sydney, New South Wales, Australia
  • 2011
    • University of North Carolina at Chapel Hill
      North Carolina, United States
  • 2009
    • University of Queensland
      Brisbane, Queensland, Australia
  • 2008
    • University of New South Wales
      Kensington, New South Wales, Australia