Optimization of a GMP-compliant method in G-Rex devices to expand large numbers of CIK cells rapidly and reproducibly from healthy donors’ peripheral and cord blood and BETs from leukemia patients, with minimal manipulation. CIK cells expanded in G-Rex are cytotoxic against CD19+ leukemia cell and lacked GvHD activity supporting their use for clinical immunotherapy studies.
Optimization of therapeutic T cell expansion in G-Rex device and applicability to large-scale production for clinical use
Publication: Cytotherapy
Published Date : 1/19/2022
Notable Quotes:
“CIK cells expanded in G-Rex (CIK-Gs) are phenotypically very similar to those expanded in T-flasks for expression of a quite extensive panel of activation, differentiation and inhibitory molecules and showed a similar T cell subset composition (CD4, CD8, TH1, TH2, TH17, Treg, αβ and γδ as well as naïve/memory subsets). CIK-Gs are functional in vitro (strong cytotoxic activity against K562 and REH leukemia targets) and in vivo (therapeutic activity in the CD19+ orthotopic pre-B acute leukemia model ALL-2) and do not induce GvHD."
“In the context of clinical trials involving treatments with cell products dedicated to single, severely ill patients, speed of production and release of ATMPs are crucial elements for the success of therapy. A more efficient and rapid expansion was demonstrated for CIKs (either from PB or CB) cultured in G-Rex in comparison with T-flasks (9.7 days and 21 days, respectively). Similarly, Blinatumomab-Expanded T cells cultured in G-Rex (BET-Gs) were obtained in a mean of 10.8 days in G-Rex with less variability of final yields of CD3+ cells compared to BETs expanded in T-flasks and a similar composition of T cell subsets was demonstrated."
“Conditions set up to expand CIKs or BETs in the small vessels (G-Rex10M single vessels or G-Rex6M multi-well vessels) can be scaled up to the G-Rex100M vessels, just applying the same plating density and using medium and additives in proportion to total volume. The constant proportion between gas-permeable surface area and maximal volume of culture medium means that the scale-up is straightforward and reproducible."
“The G-Rex vessels are different from the rocking motion, hollow fiber, stirred vessels or Prodigy systems in that they are disposable, single-use culture vessels that can easily be implemented at low cost. Several vessels can be inserted in a single standard incubator, potentially allowing parallel expansions of multiple products with just one instrument. In contrast, other platforms require specific and quite expensive bioreactors in addition to single-use culture bags or vessels, and the machinery generally needs to be multiplied to be able to perform more than one expansion run in parallel. This of course increases the costs significantly."
“CIKs and BETs can be conveniently and safely expanded in G-Rex devices for clinical purposes. The method to expand CIK-Gs from healthy donors and BET-Gs from B-NHL patient PB was validated in GMP; all product attributes (including cytotoxic potential, endotoxin contamination, sterility, and mycoplasma) were compliant with the specifications. The lower risk of microbial contamination during culture using G-Rex, thanks to the reduced manipulation, facilitate early release of products in case of urgent clinical need before full quality control data have been obtained."
“CIK cells expanded in G-Rex (CIK-Gs) are phenotypically very similar to those expanded in T-flasks for expression of a quite extensive panel of activation, differentiation and inhibitory molecules and showed a similar T cell subset composition (CD4, CD8, TH1, TH2, TH17, Treg, αβ and γδ as well as naïve/memory subsets). CIK-Gs are functional in vitro (strong cytotoxic activity against K562 and REH leukemia targets) and in vivo (therapeutic activity in the CD19+ orthotopic pre-B acute leukemia model ALL-2) and do not induce GvHD."
“In the context of clinical trials involving treatments with cell products dedicated to single, severely ill patients, speed of production and release of ATMPs are crucial elements for the success of therapy. A more efficient and rapid expansion was demonstrated for CIKs (either from PB or CB) cultured in G-Rex in comparison with T-flasks (9.7 days and 21 days, respectively). Similarly, Blinatumomab-Expanded T cells cultured in G-Rex (BET-Gs) were obtained in a mean of 10.8 days in G-Rex with less variability of final yields of CD3+ cells compared to BETs expanded in T-flasks and a similar composition of T cell subsets was demonstrated."
“Conditions set up to expand CIKs or BETs in the small vessels (G-Rex10M single vessels or G-Rex6M multi-well vessels) can be scaled up to the G-Rex100M vessels, just applying the same plating density and using medium and additives in proportion to total volume. The constant proportion between gas-permeable surface area and maximal volume of culture medium means that the scale-up is straightforward and reproducible."
“The G-Rex vessels are different from the rocking motion, hollow fiber, stirred vessels or Prodigy systems in that they are disposable, single-use culture vessels that can easily be implemented at low cost. Several vessels can be inserted in a single standard incubator, potentially allowing parallel expansions of multiple products with just one instrument. In contrast, other platforms require specific and quite expensive bioreactors in addition to single-use culture bags or vessels, and the machinery generally needs to be multiplied to be able to perform more than one expansion run in parallel. This of course increases the costs significantly."
“CIKs and BETs can be conveniently and safely expanded in G-Rex devices for clinical purposes. The method to expand CIK-Gs from healthy donors and BET-Gs from B-NHL patient PB was validated in GMP; all product attributes (including cytotoxic potential, endotoxin contamination, sterility, and mycoplasma) were compliant with the specifications. The lower risk of microbial contamination during culture using G-Rex, thanks to the reduced manipulation, facilitate early release of products in case of urgent clinical need before full quality control data have been obtained."
Review of a serum-free method of ex vivo CIK cell expansion based on the use of G-Rex static bioreactors, which drastically reduces culture manipulations and can be easily reproduced under GMP conditions as well as scaled for larger cell numbers.
A serum-free protocol for the ex vivo expansion of Cytokine-Induced Killer cells using gas-permeable static culture flasks
Publication: Cytotherapy
Published Date : 9/1/2020
Notable Quotes:
“G-Rex led to large numbers of CIK cells, with a minimal need for technical interventions, thus reducing the time and costs of culture manipulation....The described procedure can be easily translated to large-scale production under Good Manufacturing Practice.”
“Thus, the higher proportion of naïve and central memory CD3+CD56-T cells in the final CIK cell product, which we demonstrate to occur in the G-Rex cultures, could ensure a source of new post-infusion CIK effector cells that may possibly contribute to higher survival rates and long-term clinical responses.”
“G-Rex led to large numbers of CIK cells, with a minimal need for technical interventions, thus reducing the time and costs of culture manipulation....The described procedure can be easily translated to large-scale production under Good Manufacturing Practice.”
“Thus, the higher proportion of naïve and central memory CD3+CD56-T cells in the final CIK cell product, which we demonstrate to occur in the G-Rex cultures, could ensure a source of new post-infusion CIK effector cells that may possibly contribute to higher survival rates and long-term clinical responses.”
Provides details on using G-Rex for an efficient approach to generate highly functional Vγ9Vδ2 T cells in massive numbers suitable for clinical application in an allogeneic setting.
Large-scale expansion ofVγ9Vδ2T cells with engineered K562 feeder cells in G-Rex vessels and their use as chimeric antigen receptor– modified effector cells
Publication: Cytotherapy
Published Date : 12/17/2017
Notable Quotes:
“optimized the co-culture ratio of K562 aAPCs to immune cells, and migrated this method to a G-Rex cell growth platform to derive clinically relevant cell numbers in a Good Manufacturing Practice (GMP)-compliant manner”
“optimized the co-culture ratio of K562 aAPCs to immune cells, and migrated this method to a G-Rex cell growth platform to derive clinically relevant cell numbers in a Good Manufacturing Practice (GMP)-compliant manner”
Clinically relevant numbers of NK cells from unseparated apheresis or PBMCs in 10 days. Methods Chapter describing Baylor College of Medicine producing clinically relevant numbers of NK cells from unseparated apheresis or PBMCs in 10 days using G-Rex. Describing the methodologies as GMP compliant and suitable for use in clinical-grade product manufacturing.
Large-Scale Culture and Genetic Modification of Human Natural Killer Cells for Cellular Therapy
Publication: Methods in Microbiology
Published Date : 1/1/2016
Notable Quotes:
“Cultures in G-Rex (optimal 10 mL of medium per cm2 of surface area of a gas-permeable membrane) typically require no cell manipulation or feeding during the 10 days of culture period.”
“Cultures in G-Rex (optimal 10 mL of medium per cm2 of surface area of a gas-permeable membrane) typically require no cell manipulation or feeding during the 10 days of culture period.”
Expansion of Human and Murine Hematopoietic Stem and Progenitor Cells Ex Vivo without Genetic Modification Using MYC and Bcl-2 Fusion Proteins
Publication: PLOS One
Published Date : 8/1/2014
Clinical Grade Purification and Expansion of Natural Killer Cells
Publication: Critical Reviews in Oncogenesis
Published Date : 1/1/2014
Robust and cost effective expansion of human regulatory T cells highly functional in a xenograft model of graft-versus-host disease
Publication: Haematologica
Published Date : 1/1/2013
Cellular therapy of cancer with natural killer cells—where do we stand?
Publication: Cytotherapy
Published Date : 1/1/2013
Will T-cell therapy for cancer ever be a standard of care?
Publication: Cancer Gene Therapy
Published Date : 9/1/2012