T Cell, NK Cell Differentiation
iPSC differentiation to Immune Cells, Blood Cells
iPSC-derived T cells, NK cells, HPCs CD34+, dendritic cells and others provide a next generation toolset for understanding cancer and disease pathology as well as developing successful immunotherapies (adoptive therapies like CAR-T, CAR-NK, TCR-T and more). Applied StemCell with its world-class comprehensive stem cell platform which includes iPSC generation, genome editing and differentiation, can differentiate your iPSCs into CD34+ hematopoietic lineage progenitor cells and further into high-quality lineage committed immune cells: CD8+, CD4+ T cells, CD34+ HPCs, NK cell, dendritic cells, monocytes, and more. These cells are ideal for immunotherapy research for cancer and immune disorders as well as to develop complex co-culture models to screen drugs and understand immune cell activation and function.
Figure 1: iPSC Differentiation Process.
T lymphocytes (T cells): With the FDA approval of two novel T cell-based therapies for treating cancer, Kymriah™ and Yescarta™, the race for immunotherapies for different types of cancer and immune system-based disorders has intensified. These are a type of adoptive cell transfer (ACT) therapies called chimeric antigen receptor-T cell (CAR-T) where a patient’s (autologous) T cells are genetically engineered ex vivo, to recognize and kill tumor-specific antigens expressed on the surface of tumor cells. They leverage the central role played by T lymphocytes in the immune response to treat cancer. Another type of cytotoxic T lymphocyte cell therapy includes directly delivering ex vivo engineered autologous T cell receptors (TCR-T) that recognize tumor-specific intracellular proteins presented as protein fragments by the major histocompatibility complex (MHC) class I proteins.
Although a great success initially in cancer remission and treatment, there are limitations in the technology such as: limited to autologous (patient) T cells, loss of naïve T cells and persistence of long-term immunologic memory as T cells acquire effector functions, and T cell exhaustion due to repeated activation of T cells in cancer patients which severely reduces the amount of cells that can be harvested from the patient. Such limitations may be overcome by using T cells differentiated ex vivo from iPSCs reprogrammed from patient cells. These T cells have rejuvenated T cell characteristics and can be produced in high numbers with consistent and reliable quality. Additionally, iPSCs can be genetically engineering more efficiently than primary T cells, and can be used for generating CAR-T and TCR-T cells for immunotherapy applications.
Natural Killer (NK) cells are a type of lymphoid cells that originate from the same progenitor as T cells and B cells. They are an important part of the innate immune response. NK cells are activated in response to interferons or macrophage-derived cytokines. They recognize “non-self” cells without the need for antigen presentation or recognition, executing a rapid immune reaction. The broad cytotoxicity and rapid apoptosis induced by the NK cells helps contain virus-infected cells and controlling early signs of cancer while the adaptive immune response is activated to produce cytotoxic T cells to clear the antigens. With the improving success rate of CAR-T cell therapy, scientists are exploring the NK cells to engineer them for CAR-NK cell therapy to target cancer cells. CAR-NK cells have certain advantages over CAR-T cells in that they retain their natural tumor recognition and killing ability; they do not require strict HLA matching and lack the potential to cause graft-versus-host disease (GvHD), they do not have the same safety concerns as seen with CAR-T cells such as cytokine release syndrome, and therefore they can be generated from allogenic donors and possibly developed as off-shelf therapeutic products.
However, purification of NK cells from allogenic hosts is critical since residual T and B cells may cause GvHD and other complications. Primary NK cells are difficult to harvest, purify, and transfect/transduce, and result in a heterogenous population. They are also difficult to standardize due to the heterogeneity in starting material from different donors. As well, the generation of large quantities of highly pure NK cells requires an extended manufacturing process which can compromise recovery of NK cells, their viability, and potency. Therefore, iPSC-derived NK cells offer a consistent and renewable starting material to provide standardizable NK cells without the dependency on donors and complex harvesting and purification processes. They offer a method to generate homogenous populations of CD56+ CD45+ NK cells and subsequently to generate CAR-NK cells for targeted allogenic cancer immunotherapy, even solid tumors.
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Kawamoto, H., Masuda, K., Nagano, S., & Maeda, T. (2018). Cloning and expansion of antigen-specific T cells using iPS cell technology: development of “off-the-shelf” T cells for the use in allogeneic transfusion settings. International journal of hematology, 107(3), 271-277.
Minagawa, A., Yoshikawa, T., Yasukawa, M., Hotta, A., Kunitomo, M., Iriguchi, S., ... & Kawai, Y. (2018). Enhancing T cell receptor stability in rejuvenated iPSC-derived T cells improves their use in cancer immunotherapy. Cell stem cell, 23(6), 850-858.
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Development of cellular immunotherapy (such as CAR-T, CAR-NK cell therapies and other adoptive cell transfers) is gaining momentum for targeted treatment of cancer and immune disorders, and in understanding role of immune response in pathology of diseases. Therefore, an in-depth knowledge of the hematopoiesis of immune cells, their activation, and function is critical to understanding disease pathology and developing safe and effective therapies. Current processes for developing such cell-based therapies are complex and involve: harvesting of patient (autologous) cells, handling and processing patient-derived primary cells, including genome engineering the cells to target specific antigens, and re. And, result in only a single-dose of the product being manufactured that is heterogenous in its characteristics and very expensive to develop and engineer.
The revolutionary iPSC technology may provide a reliable and standardizable starting material to generate various immune cells for therapeutic purposes, to screen drugs and for disease modeling; and thus, overcome the difficulties of sourcing patient’s primary cells, genome engineering these notoriously difficult to engineer cells, and avoid the variance in cellular characteristics associated with current methods:
- A more physiologically relevant cell line model combining the advantages of cell culture and primary cells
- Unlimited self-renewal potential of iPSCs would assure a steady and consistent supply of cells
- Single cell clonability for a homogeneous population of cells
- CRISPR and genetic engineering amenability to engineer antigen-specific cells for targeted therapy
- Potential to differentiate into many different cell types with same genetic background, including iPSC-derived CD34+ progenitor cells, CD4+T helper cells, CD8+ cytotoxic T cells, natural killer (NK) cells, dendritic cells, macrophages and more
- Full characterization possible
- Scalability for mass production
- Potential for developing into cryopreserved, off-the-shelf therapeutic products
The iPSC-based cellular immunotherapy has great potential to make successful bench-to-bedside translation a reality.
Applied StemCell can generate lineage-committed immune cells (CD34+ progenitor cells, CD8+T cells, NK cells, monocytes, dendritic cells) differentiated from clonal master iPSC line(s) using highly optimized and efficient differentiation protocols, for preclinical applications:
1. Customer provided patient iPSC (healthy/ disease)
2. ASC’s fully characterized control “master” iPSC lines
3. CRISPR/Cas9 engineered “master” iPSC lines
Have patient samples but don’t have an iPSC starting material? We can reprogram patient PBMCs/ fibroblasts/ other patient samples into functional iPSCs and proceed with differentiation.
Applications for iPSC-derived immune cells include:
- Disease modeling for cancer and autoimmune disorders
- iPSC-derived antigen-specific “CAR-T” research
- Adoptive transfer and other cell-based therapies
- Antibody discovery
- Drug target discovery and drug screening
- Cell line models for drug toxicity screening
- Cell-based therapies
- Co-culture cells of different lineages to model in vivo heterogeneity of human immune system and cancer pathology
- Multiplexing target attributes