Automated isolation of primary antigen‐specific T cells from donor lymphocyte concentrates: results of a feasibility exercise

Bunos, Milica; Hümmer, Christiane; Wingenfeld, Eva; Sorg, Nadine; Pfirrmann, Verena; Bader, Peter; Seifried, Erhard; Bönig, Halvard

doi:10.1111/vox.12291

Cited by 22 publications

(23 citation statements)

References 20 publications

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“…Other investigators have also validated the feasibility of clinical-scale CMVspecific T-cell selection using the CliniMACS Prodigy [18,20,21]. However, full characterization of the final product has been insufficient due to the limited numbers of isolated cells.…”

Section: Discussionmentioning

confidence: 99%

“…The principle of the CCS has been described previously [19], and the CCS process using the CliniMACS Prodigy has also been described recently by Bunos and colleagues [18,20,21]. Briefly, PBS/EDTA buffer and elution buffer were first prepared by adding 0.5% and 2.5% HSA (w/v), respectively.…”

Section: Isolation Of CMV Pp65-specific T Cells Using the Clinimacs Pmentioning

confidence: 99%

“…The Clini-MACS Prodigy (Miltenyi Biotec) was initially described for its automated density gradient separation, including red blood cell depletion, magnetic cell separation, and cell culture [17]. Recently, the integration of IFN-γ CCS software into the CliniMACS Prodigy has proven to be feasible for the automated production of CMV-specific T cells [18].…”

Section: Introductionmentioning

confidence: 99%

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Robust Production of Cytomegalovirus pp65-Specific T Cells Using a Fully Automated IFN-γ Cytokine Capture System

Kim

Nam

et al. 2017

Transfus Med Hemother

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Background: Cytomegalovirus(CMV)-related diseases are a serious cause of morbidity and mortality following hematopoietic stem cell transplantation (HSCT). CMV-specific cytotoxic T lymphocytes (CMV-CTLs) have been reported as an alternative to antiviral drugs that provide long-term CMV-specific immunity without major side effects. However, their application has been limited by the prolonged manufacturing process required. Methods: In this study, we applied the IFN-γ cytokine capture system (CCS) using the fully automated CliniMACS Prodigy device for rapid production of CMV-CTLs, which may be applicable in clinically urgent CMV-related diseases. Five validation runs were performed using apheresis samples from randomly selected CMV-seropositive healthy blood donors. Successive processes, including antigen stimulation, anti-IFN-γ labeling, magnetic enrichment and elution, were then performed automatically using the CliniMACS Prodigy, which took approximately 13 h. Results: The original apheresis samples consisted mainly of CD45RA+ CD62L+ naïve T cells as well as 0.3% IFN-γ-secreting CD3+ T cells that showed a response to the CMV pp65 antigen (CD3+ IFN-γ+ cells). Following IFN-γ enrichment, the target fraction contained 51.3% CD3+ IFN-γ+ cells with a reduction in naïve T cells and selection of CD45RA- CD62L- and CD45RA+ CD62L- memory T cells. Furthermore, extended culture of these isolated cells revealed functional activity, including efficient proliferation, sustained antigen-specific IFN-γ secretion, and cytotoxicity against pp65-pulsed target cells. Conclusion: The findings reported here suggest that the IFN-γ CCS by the CliniMACS Prodigy is a simple and robust approach to produce CMV-CTLs, which may be applicable for the treatment of clinically urgent CMV-related diseases.

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Section: Discussionmentioning

confidence: 99%

Section: Isolation Of CMV Pp65-specific T Cells Using the Clinimacs Pmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Robust Production of Cytomegalovirus pp65-Specific T Cells Using a Fully Automated IFN-γ Cytokine Capture System

Kim

Nam

et al. 2017

Transfus Med Hemother

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“…Such techniques are time-consuming and costly and are very difficult to perform under true Good Manufacturing Practice conditions [29]. In contrast, multimer-based [30] or interferon-g catch [7,31] direct selection of CMV-specific T cells from seropositive donors allows a more rapid production of a cellular product. We demonstrated that a single stimulation with a CMVpp65 peptide pool at day 0 of the expansion period suffices to increase the frequency of CMV-specific cytotoxic cells within CIK cells from CMV-seropositive donors.…”

Section: Discussionmentioning

confidence: 99%

Cytomegalovirus-specific cytokine-induced killer cells: concurrent targeting of leukemia and cytomegalovirus

et al. 2015

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“…Various culture vessels, including G-Rex flasks (Wilson Wolf Manufacturing), cell differentiation bags and bioreactors have proven suitability for T-cell manufacturing [5][6][7]. Current procedures involve a variety of hands-on steps and transfer of cells between cultivation vessels, each bearing the risk of contamination, human errors and cell loss.To simplify the protocol and minimize potential risks during the process, the CliniMACS Prodigy [8] has been adapted to automate clinical-scale manufacturing of cell products [9][10][11]. This platform allows for magnetic beadbased cell separation followed by activation, transduction, multiple media-exchanges and washing steps with a single-use, closed tubing system ( Figure 1A, Figure 1B).…”

Section: Introductionmentioning

confidence: 99%

Automated manufacturing of chimeric antigen receptor T cells for adoptive immunotherapy using CliniMACS Prodigy

et al. 2016

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Novel cell therapies derived from human T lymphocytes are exhibiting enormous potential in early-phase clinical trials in patients with hematologic malignancies. Ex vivo modification of T cells is currently limited to a small number of centers with the required infrastructure and expertise. The process requires isolation, activation, transduction, expansion and cryopreservation steps. To simplify procedures and widen applicability for clinical therapies, automation of these procedures is being developed. The CliniMACS Prodigy (Miltenyi Biotec) has recently been adapted for lentiviral transduction of T cells and here we analyse the feasibility of a clinically compliant T-cell engineering process for the manufacture of T cells encoding chimeric antigen receptors (CAR) for CD19 (CAR19), a widely targeted antigen in B-cell malignancies. Using a closed, single-use tubing set we processed mononuclear cells from fresh or frozen leukapheresis harvests collected from healthy volunteer donors. Cells were phenotyped and subjected to automated processing and activation using TransAct, a polymeric nanomatrix activation reagent incorporating CD3/CD28-specific antibodies. Cells were then transduced and expanded in the CentriCult-Unit of the tubing set, under stabilized culture conditions with automated feeding and media exchange. The process was continuously monitored to determine kinetics of expansion, transduction efficiency and phenotype of the engineered cells in comparison with small-scale transductions run in parallel. We found that transduction efficiencies, phenotype and function of CAR19 T cells were comparable with existing procedures and overall T-cell yields sufficient for anticipated therapeutic dosing. The automation of closed-system T-cell engineering should improve dissemination of emerging immunotherapies and greatly widen applicability.

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Automated isolation of primary antigen‐specific T cells from donor lymphocyte concentrates: results of a feasibility exercise

Cited by 22 publications

References 20 publications

Robust Production of Cytomegalovirus pp65-Specific T Cells Using a Fully Automated IFN-γ Cytokine Capture System

Robust Production of Cytomegalovirus pp65-Specific T Cells Using a Fully Automated IFN-γ Cytokine Capture System

Cytomegalovirus-specific cytokine-induced killer cells: concurrent targeting of leukemia and cytomegalovirus

Automated manufacturing of chimeric antigen receptor T cells for adoptive immunotherapy using CliniMACS Prodigy

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Automated isolation of primary antigen‐specific T cells from donor lymphocyte concentrates: results of a feasibility exercise

Cited by 22 publications

References 20 publications

Robust Production of Cytomegalovirus pp65-Specific T Cells Using a Fully Automated IFN-&#x03B3; Cytokine Capture System

Robust Production of Cytomegalovirus pp65-Specific T Cells Using a Fully Automated IFN-&#x03B3; Cytokine Capture System

Cytomegalovirus-specific cytokine-induced killer cells: concurrent targeting of leukemia and cytomegalovirus

Automated manufacturing of chimeric antigen receptor T cells for adoptive immunotherapy using CliniMACS Prodigy

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Product

Resources

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Robust Production of Cytomegalovirus pp65-Specific T Cells Using a Fully Automated IFN-γ Cytokine Capture System

Robust Production of Cytomegalovirus pp65-Specific T Cells Using a Fully Automated IFN-γ Cytokine Capture System