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Table 11 Protocol features for Prodigy systems

From: Lymphocyte expansion in bioreactors: upgrading adoptive cell therapy

Authors Protocol features Starting material Culturing Expansion Purity Functionality
IL-2 [IU/mL] Stimmul. strategy Medium Serum Source Seed [cell/ml] Vol. [mL] Tubing Set Shaking Fold Days Compared system Comp. Fold
T cells
Mock [187] - 2016 Used CD3/CD28; Transd. TexMACS GMP 3% HS T cells from PBMC 7.45–10 × 108 100 TS520 Shaker from day 4/5. More vigorous shaking depending on cell density 16.2 ± 7.9 8–10 G-rex 10 22.3 ± 12.2 No difference with G-rex;98.0% max T cell purity Exhaustion profile did not differ. Prodigy cells produced less IFN-γ. Secretion of TNF-α and IL-2 was lower but not significant specific lysis to target
Priesner [199] - 2016 Not used CD3/CD28; Transd.; IL7; IL15 TexMACS GMP 3% HS CD62L+ cells from PBMC 3 × 109 total NP TS520 Static culture until day 3; clusters were dispersed 13–23 (cells); 28–42 (Tcells) 10–13 Not used 83.4–98.9% CD3 + CD45+ T cells Transduction efficiency of 83%
Lock [200] - 2017 Not used CD3/CD28(transact); Transd.; IL7; IL15 TexMACS GMP 3% HS CD4 + CD8 + CD45+ from PBMC 2–10 × 107 total 250 TS520 Automated media exchange every day 65 ± 36 12 Not used 91.3–5.0% for Healthy donor and 88.3–7.1% for Melanoma Patient sourced Transduct.: 34.5–11.7% for HD and 36.4–17.7% for PM. Secretion of GM-CSF, IFN-γ, IL-2, and TNF-α, no IL-4, IL-5, or IL-10 detected.
Blaeschke [201] - 2018 Not used CD3/CD28(transact); Transd.; IL7; IL15 TexMACS GMP 3% HS CD4+/CD8+ from PBMC 2.86 × 107 total NP TS520 NP 14.7–102.4 12 Not used 50.3% CD4+ and 38.7% CD8+ cells; 10.2% of the cells were NKT cells. Transduction: 26.95%; No increased expression of exhaustion markers. CD19 CAR T cells killed 80% of the target (5:1); cells were able to secrete GM-CSF, IFN-γ, IL-2, and TNF-α
Zhang [202] - 2018 200 CD3/CD28(transact); Transd. TexMACS GMP   CD4+/CD8+ from PBMC 108 total 250 TS520 Media exchanges performed without disrupting the cells 16 (cells);
20 (T cells)
  Not used CD4+ and CD8+ T cells changed from 45 and 34% to 22 and 74%.The frequency of CD4+ Tregs decreased Transduction 60% in CD4+ and 50% in CD8+ T cells. Cells produced significant amounts of Th1 cytokine, IFN-γ and IL-2 after stimulation. Cytotoxicity ~ 60% at a 3:1 effector-to-target ratio
Zhu [197] - 2018 Used CD3/CD28(transact); Transduct. TexMACS GMP 3% HS CD4+/CD8+ from PBMC 108 total NP TS520 As medium is added or exchanged, it is programmed to shake 24.5–41.0 13 G-rex100 Signific. better CD4+ decreased; CD8+ increased. Treg decreased. Little difference between Prodigy and G-rex in regard to phenotype Low levels of IFN-γ. IFN-γ; all CAR-T-cell products lysed Raji cells when tested; transduction: 21 to 56.6%. better transduction in Prodigy.
 Aleksandrova [203] - 2019 Not used CD3/CD28(transact); Transd.; IL7; IL15 TexMACS GMP HS CD4+/CD8+ from PBMC NR NP TS520 NP 46–81 12 Not used 97% T cells (range 96–99); some impurities of NKT and NK cells: median 2.9 and 0.07%, respectively. CD4/CD8 ratio of 2.4 decreased to 1.7 (range 1.1–2.3) in the final product. Transduction: 22% on day 5 (range 17–41%) and 23% in the final product (range 21–45%).; CAR T cells were also cytotoxic against target cells
Fernández [204] - 2019 100 CD3/CD28(transact); Transd. TexMACS GMP   CD45RA− from PBMC 1 × 108 total NP TS520 NP 13.4–38.6 10–13 Not used Enrichment in CD4+ vs. CD8+ T cells. Effector memory (TEM) phenotype Cytotoxicity ≥20%. The expression of NKG2D increased during the expansion of cells. NKG2D CAR expression 55–60.6%
Marín [205] - 2019 1000 T reg beads (aCD3/aCD8); Rapamycin TexMACS GMP 5%AB CD4 + CD25− from PBMC Mean: 56.9 × 106 250 TS510 Shaked from day 5; media exchange was conducted. Agitation paused to facilitate bead to cell contact Similar kinetics G-rex G-rex 95.6% (range 93.7–97.0; n = 4). Prodigy R 92.8% (range 89.8–94.5%, n = 4; p = 0.08). Suppressive capacity varied between donors but appeared to be independent of the manufacturing regimen
NK cells
Granzin [198] - 2015 500 Irr. EBV-LCL TexMACS GMP 5% AB CD56 + CD3- from PBMC 2 × 106 210 TS730 Shaking after day 7 850 ± 509 14 T75 1344 ± 1135 > 99% CD3e/CD56þ,T or B cells could not be detected. No differences in the production of IFN-γ and TNF-α and similar levels of degranulation. No reduction in telomere length. T75 and Prodigy had a comparable marker profile. NKG2C, CX3CR1 and KIR2DL2/DL3 higher in Prodigy
 Oberschmidt [188] - 2019 500 IL-21, IL-15; NK MACS supplement NK MACS 5%AB CD56 + CD3- from PBMC 2–4 × 106 250 TS310 0.17–0.3 g shaking mode was activated on day 7. 11.31–17.94 3–14 Not used 98.8–99.2% Total NK cells expressed initially low to moderate levels of NKG2D, NKp30, NKp44, and NKp46. The surface markers increased during the 14-day expansion. Death receptors and activation markers increased. NK cell cytotoxicity increased