Where do the NK cells used for immunotherapy come from?

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Where do the NK cells used for immunotherapy come from?

Where do the NK cells used for immunotherapy come from? Current immunotherapy uses NK cells from various sources: primary peripheral blood NK cells (PB-NK Figure 3a, e), primary cord blood NK cells (CB-NK Figure 3b), immortalized NK cell lines ( NK-92, Figure 3c), and more recently iPSC-derived NK cells (Figure 3d), each cell has its own advantages and disadvantages.

Where do the NK cells used for immunotherapy come from?

fig. 3. Sources of NK cells.

Current NK therapies use NK cells from the following sources: primary PB-NK cells, primary CB-NK cells, NKcell lines, and recently, iPSC-NK cells, each with its own benefits and limitations.

Research from the late 1990s to the early 2000s focused on the use of cytokine-activated autologous primary NK cells to improve the anti-tumor activity of NK cells. However, it has proven difficult to isolate a large number of primary PB-NK cells because only 5-15% of circulating blood lymphocytes are NK cells, and the number of leukocyte removal products is highly dependent on the donor.

In addition, separation from peripheral blood lymphocytes usually results in a heterogeneous population of lymphocytes, and the yield of NK cells is only 10-20%. Since primary NK cells have reduced cytotoxicity after cryopreservation, their use is more complicated. Although the low yield and cytotoxic loss of cryopreservation can be partially overcome by cytokine support and feeder cell expansion, this approach makes PB-NK cells not the best choice for “off the shelf” treatment.

In contrast, CB-NK cells recover well after cryopreservation and may be a potential candidate for “off the shelf” therapy using primary NK cells. Although the percentage of NK cells in CB (23%) is higher than that in PB (11%), one disadvantage is that the number of NK cells per cord blood unit is limited, which may require multiple units per dose. Compared with PB-NK cells, CB-NK cells express lower levels of KIR and Granzyme B, and higher levels of NKG2A inhibitory receptors, indicating that CB-NK cells have an immature phenotype. Although this special phenotype leads to lower cytotoxicity, CB-NK cell effector function can be enhanced to PB-NK cell function through cytokine stimulation.

Recently, because iPSC-NK therapy can solve the supply chain bottleneck associated with primary and cell line NK therapy, people are increasingly interested in iPSC-NK therapy.

Advantages include iPSCs can be produced from readily available sources (such as fibroblasts or peripheral blood), maintain pluripotency during expansion, and can be stored for long periods of time. NK cells from iPSC have been proven to be as effective as primary NK cells and NK-92 cells.

A study showed that in MA148 and A1847 ovarian tumor xenograft models, the cytotoxicity of iPSC-NK is comparable to activated and expanded PB-NK. Compared with donor PB-NK cells, donor peripheral blood iPSC-NK cells are also more cytotoxic to SKOV3, SW480, HCT-8, MCF7 and SCC-25 cancer cell lines, and have greater efficacy and donor efficacy against K562s. Body PB is similar to NK cells. iPSC-NK cells also proved to be superior to the established NK-92 cell line because iPSC-NK cells do not require irradiation [80-82].

Another benefit of iPSC-NK cells is that they express the CD16 receptor and are therefore capable of ADCC [81, 82]. Zeng et al. proved that PB-iPSC-NK cells can perform ADCC by showing successful killing of Raji cells opsonized with anti-CD20-hIgG1 antibody [81]. The typical degranulation of NK cells is achieved by phosphorylation of the Syk-PLCγ-DAG/inositol triphosphate or Syk-Vav-RAC-PAK-MEK-ERK pathway, while the secretion of inflammatory cytokines is achieved by the NF-κB pathway Caused by phosphorylation [83, 84].

However, whether iPSC-NK cells need the same activation pathway to induce granule and cytokine secretion is not fully understood. Nevertheless, these data indicate that the iPSC-NK-based strategy combines the most attractive features of primary NK cells and NK-92 cell lines, namely the high potential for cytotoxicity, including ADCC function, as well as in vivo expansion and persistence The potential even in cryopreservation.

Compared with primary NK cells, the research team has successfully differentiated pluripotent stem cells (PSC) into NK cells with similar phenotypes and effector functions. Differentiated cells express multiple NK-related receptors, such as CD56, KIR, CD16, NKp44, NKp46, NKG2D and TRAIL. However, it is unclear whether the derived cells are similar to the bright or dim subsets of the CD56 population, which suggests that further studies of the expression profile of differentiated NK cells may be needed.

A recent study by Dege et al. The ontogeny of hPSC-derived NK cells was further explored, and two different CD34+ populations were discovered, which determine the final effector function. HOXAneg/low/CD34+ progenitor cells produced a population that showed more effective degranulation, while HOXA+/CD34+ progenitor cells produced a NK population with strong inflammatory cytokine secretion [97].

These findings indicate that the origin of differentiated NK cells should be considered when adapting to immunotherapy applications, and it may be beneficial to enrich the progenitor cell population during the differentiation process to obtain more effective markers of degranulation characteristics.

In view of the therapeutic potential of iPSC-NK products, the FDA has approved a phase I clinical trial to study Fate Therapeutics’ ready-made iPSC-NK product FT500. FT500 is the first clinical study of iPSC-derived cell products approved by the US FDA. NK cells are developed from the cloned master iPSC line cell bank, so iPSC-NK cells that are relatively homogeneous, controllable in quality, and capable of freezing for long-term storage can be produced in large quantities.

This trial aims to evaluate the safety and tolerability of multi-dose FT500 combined with checkpoint blocking therapy in the treatment of adults with advanced solid tumors (ClinicalTrials.gov, NCT03841110). A separate observational study has been initiated to evaluate the long-term safety and effectiveness of patients treated with FT500 (ClinicalTrials.gov: NCT04106167).

Where do the NK cells used for immunotherapy come from?