Application data and FAQs for LymphoONE medium
The development of therapies utilizing adoptive T-cell transfer has provided powerful new approaches for combating previously intractable diseases such as cancer. However, the broader application of these approaches will require additional technological innovation and refinement of existing protocols for culturing and engineering T cells. For example, in order to obtain sufficient quantities of patient-derived T cells for therapeutic applications, conventional culture systems typically require supplementation of culture medium with serum or plasma, which can be problematic for a variety of reasons:
- Supplies of autologous serum that can be obtained from a patient are limited and vary depending on the patient's condition
- Supplementation with donor serum of bovine or human origin is costly, introduces a source of unwanted variability in protocol performance, and raises the risk of pathogen exposure
- Culturing in the absence of serum yields T cells that exhibit improved downstream performance (Janetzki et al. 2010; Medvec et al. 2018)
Efficient T-cell expansion without serum
To better facilitate research involving T cell-based therapies, Takara Bio has developed LymphoONE T-Cell Expansion Xeno-Free Medium, a chemically defined medium that enables efficient T-cell expansion without the addition of serum or plasma. As demonstrated by the data below, LymphoONE medium enables greater expansion relative to Takara Bio's previous offering (GT-T551) and commonly used serum-free media from other suppliers (X-VIVO 15, AIM V).
T-cell expansion comparison using various serum-free culture media. Peripheral blood mononuclear cells (PBMCs) were isolated from blood obtained from two healthy donors and cultured for four days under stimulation by anti-CD3 monoclonal antibody and IL-2 in CultiLife 215 Culture Bags using LymphoONE T-Cell Expansion Xeno-Free Medium, GT-T551 (Takara Bio), X-VIVO 15 (Lonza), or AIM V (Thermo Fisher Scientific). The cells were then cultured at appropriate dilutions for an additional 10 days in each of the respective media supplemented with IL-2. By Day 14, cells cultured in LymphoONE medium exhibited higher fold expansion than cells cultured in either of the other three media.
Higher proportions of naïve T cells
In addition to expansion efficiency, another critical factor in the development of T cell-based therapies involves the phenotypic characteristics of expanded populations. Research over the past decade has established that T cells with a naïve phenotype provide greater therapeutic efficacy as compared to more differentiated populations (Klebanoff et al. 2015), an understanding that has informed the development of LymphoONE medium, as demonstrated below.
Phenotypic analysis of T cells expanded using various culture media in the presence or absence of serum. Peripheral blood mononuclear cells (PBMCs) were cultured for four days under stimulation by anti-CD3 monoclonal antibody, IL-2, and RetroNectin reagent in CultiLife 215 Culture Bags with or without addition of human serum using LymphoONE T-Cell Expansion Xeno-Free Medium, X-VIVO 15 (Lonza), or AIM V (Thermo Fisher Scientific). The cells were then cultured at appropriate dilutions with or without serum for an additional six days in each of the respective media supplemented with IL-2. Human serum was added at a concentration of 1% from Day 0 to Day 7, and then at a concentration of 0.5% from Day 7 to Day 10. Expansion in LymphoONE medium resulted in higher proportions of cells expressing markers associated with a naïve T-cell phenotype (CD45RA+/CCR7+), with or without addition of serum.
References
Janetzki, S. et al. Performance of serum-supplemented and serum-free media in IFNgamma Elispot Assays for human T cells. Cancer Immunol. Immunother. 59, 609–18 (2010).
Klebanoff et al. Memory T cell-driven differentiation of naive cells impairs adoptive immunotherapy. J. Clin. Invest. 126, 318–334 (2016).
Medvec, A. R. et al. Improved expansion and in vivo function of patient T cells by a serum-free medium. Mol. Ther. Methods Clin. Dev. 8, 65–74 (2018).
FAQs about LymphoONE T-Cell Expansion Xeno-Free Medium
How does LymphoONE medium perform with the addition of serum?
While LymphoONE medium was developed to enable efficient T-cell expansion in the absence of serum, researchers always have the option of supplementing the medium with serum or plasma in order to achieve greater proliferation. As demonstrated below by the data involving stimulation by IL-2, anti-CD3 mAb, and RetroNectin reagent, supplementation of LymphoONE medium with human serum results in greater T-cell expansion and yields higher proportions of naïve T cells (CD45RA+/CCR7+ phenotype) compared to serum-free conditions. The data also demonstrate that LymphoONE medium outperforms competitor media under both scenarios (with or without addition of serum).
T-cell expansion via stimulation by anti-CD3 antibody, IL-2, and RetroNectin in the presence or absence of human serum. Peripheral blood mononuclear cells (PBMCs) were cultured for four days under stimulation by anti-CD3 monoclonal antibody, IL-2, and RetroNectin reagent in CultiLife 215 Culture Bags with or without addition of human serum using LymphoONE T-Cell Expansion Xeno-Free Medium, X-VIVO 15 (Lonza), or AIM V (Thermo Fisher Scientific). The cells were then cultured at appropriate dilutions with or without serum for an additional six days in each of the respective media supplemented with IL-2. Human serum was added at a concentration of 1% from Day 0 to Day 7, and then at a concentration of 0.5% from Day 7 to Day 10. Top panel. By Day 10, cells cultured in LymphoONE medium exhibited higher fold expansion than cells cultured in either of the other two media, regardless of whether serum was added. Bottom panel. Expansion in LymphoONE medium also resulted in higher proportions of cells expressing markers associated with naïve T-cell phenotype (CD45RA+/CCR7+).
Does LymphoONE medium contain antibiotics?
Yes, the version of LymphoONE medium that is currently available (Cat. #s WK552S & WK552) contains streptomycin. In the future, we will launch a GMP-grade version of the medium without antibiotics.
Does LymphoONE medium contain any animal-derived components?
No. LymphoONE medium is xeno-free, meaning that it does not include any animal-derived (nonhuman) components.
Does LymphoONE medium contain any human-derived components?
Yes. LymphoONE medium contains highly purified pharmaceutical-grade human serum albumin.
What other components does LymphoONE medium contain?
In addition to streptomycin and human serum albumin, LymphoONE medium also includes L-glutamine and recombinant human insulin.
Is LymphoONE medium chemically defined?
Yes. The identities and quantities of all medium components are known.
How does LymphoONE medium compare to Takara Bio's previous T-cell medium, GT-T551?
The composition of LymphoONE T-Cell Expansion Xeno-Free Medium is similar to that of its predecessor, GT-T551. However, the formulation of LymphoONE medium has been optimized to enable greater T-cell proliferation, as demonstrated by the data presented above.
Will there be a GMP-grade version of LymphoONE medium?
We plan to launch a GMP-grade version of LymphoONE medium in 2021.
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