A new method for in vitro expansion of cytotoxic human CD3−CD56+ natural killer cells

Carlens, S; Gilljam, Mari; Chambers, Benedict J.; Aschan, J; Guvén, Hayrettin; Ljunggren, Hans‐Gustaf; Christensson, Birger; Dilber, M. Siraç

doi:10.1016/s0198-8859(01)00313-5

Cited by 118 publications

(91 citation statements)

References 23 publications

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“…27 In short, peripheral blood was obtained from B-CLL patients and cultures were initiated on the same day (day 0). PBMCs were isolated by gradient centrifugation, using Lymphoprep (Nyegaard, Oslo, Norway).…”

Section: Cell Cultures and Quantificationmentioning

confidence: 99%

Expansion of natural killer (NK) and natural killer-like T (NKT)-cell populations derived from patients with B-chronic lymphocytic leukemia (B-CLL): a potential source for cellular immunotherapy

et al. 2003

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B-cell chronic lymphocytic leukemia (B-CLL) is the most common leukemia in the Western world. It is currently an incurable disease, making new treatment options such as immunotherapy desirable. Monoclonal antibodies (Mabs) to surface antigens of the tumor cell is one option. Administration of cytotoxic cells such as natural killer (NK) and natural killerlike T (NKT) cells expanded in vitro might be a useful treatment modality alone or in combination with MAbs. A limiting step in the development of successful cellular immunotherapy has been the availability of appropriate cytotoxic cells. Here, we report the feasibility of expanding populations of the human killer cells, CD3ÀCD56 þ NK and CD3 þ CD56 þ NKT cells, from peripheral blood mononuclear cells (PBMCs) of B-CLL patients. The influence of tumor B cells on the in vitro expansion of killer cells was assessed by depleting B cells from PBMCs by microbead separation before culture. The 21-day cultures from both B-cell-and non-B-cell-depleted PBMC showed a marked expansion of NK cells, and also of T cells, among which almost half had the NKT phenotype. Depletion of B cells before culture did not change the expansion rates of NK and NKT cells significantly. In patients with progressive B-CLL, NK cell expansion capacity was improved after fludarabine treatment when compared to samples obtained before treatment. Repeated samples of PBMCs from individual untreated patients with both indolent and progressive disease cultured under identical conditions gave similar NK cell expansion rates. Expanded killer cell populations had cytotoxic function against the NK-sensitive target K562 cell line and expressed high levels of Granzyme B. From our studies, we conclude that NK cells as well as NKT cells from the peripheral blood of B-CLL patients can be expanded, and that these cells have cytotoxic capacity.

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Section: Cell Cultures and Quantificationmentioning

confidence: 99%

Expansion of natural killer (NK) and natural killer-like T (NKT)-cell populations derived from patients with B-chronic lymphocytic leukemia (B-CLL): a potential source for cellular immunotherapy

et al. 2003

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“…14,15 Nevertheless, because NK cells represent a small fraction of peripheral blood mononuclear cells (PBMCs), generating them in a sufficient number to meet clinical requirements is challenging. 16 The proliferation, survival and function of NK cells are improved by engagement of NKG2D with solid-phase immobilized chimeric molecules that resemble NKG2DLs, such as MICA-Fc and ULBP1-Fc. 17 In this study, we investigated whether immobilized MICA (iMICA) synergizes with soluble 4-1BBL (s4-1BBL) and IL-15 to expand NK cells efficiently.…”

Section: Introductionmentioning

confidence: 99%

Immobilized MHC class I chain-related protein A synergizes with IL-15 and soluble 4-1BB ligand to expand NK cells with high cytotoxicity ex vivo

Gong

Xiao

et al. 2010

Cell Mol Immunol

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Major histocompatibility complex (MHC) class I chain-related protein A (MICA), which is a ligand for human NKG2D, is expressed by a variety of epithelial tumor cells and promotes the activation of natural killer (NK), CD8 1 and cd-T cells. Although ectopic expression of MICA on tumor cells elicits anti-tumor responses, soluble MICA downregulates the activities of lymphocytes. In this study, we showed that recombinant, immobilized MICA (iMICA) molecules coated on plastic wells weakly promote peripheral NK cell activation, secretion of interferon (IFN)-c and degranulation without inducing apoptosis. In addition, iMICA synergized with IL-15 and soluble 4-1BB ligand (s4-1BBL) to expand NK cells 25-to 42-fold in a 13-day culture, whereas NK cells stimulated only with IL-15 and s4-1BBL expanded 10-to 16-fold. In contrast to NK cells expanded by IL-15 and s4-1BBL stimulation, NK cells expanded long term in the presence of iMICA exhibited increased cytotoxicity against leukemia cells. These results suggest that large numbers of NK cells with high cytotoxicity can be generated by stimulation with IL-15 and s4-1BBL in the presence of iMICA and that these cells can be used for adoptive cancer immunotherapy. Studies on the 4-1BB signaling pathway have shown that ligation of 4-1BB promotes the survival and multiplication of CD8 1 T and NK cells. 6 IL-15 is essential for the development and function of NK cells. 7 Coculture of NK cells with K562 cells ectopically coexpressing 4-1BB ligand (4-1BBL) and membrane-bound IL-15 effectively promotes the expansion of NK cells in vitro and mediates their cytotoxicity against some leukemia cells. 8,9 In addition, IL-15 upregulates the expression of NKG2D on NK and T cells, even in the presence of soluble NKG2DL. 10 IL-15 and NKG2D signals cross-regulate each other and

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“…46,47 Ex vivo expansion of NK and NKT cells from healthy donors and CLL patients have shown exciting results, where Guven et al reported even better expansion rates for functionally active NK cells from CLL patients despite initial low numbers. 3,48 However, possible side effects of immunoligand-associated NK cell activation in patients such as autoimmune reactions toward healthy B cells should be considered. Limitations of autologous and allogeneic NK cell infusions have been discussed elsewhere 49 and must be pointed out in the context of adoptive transfer of NK cells with immunoligand.…”

Section: Discussionmentioning

confidence: 99%

Mono- and dual-targeting triplebodies activate natural killer cells and have anti-tumor activityin vitroandin vivoagainst chronic lymphocytic leukemia

et al. 2016

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Chronic lymphocytic leukemia (CLL) is the most common form of leukemia that affects B lymphocytes in adults. Natural killer (NK) cells in CLL patients are intrinsically potent but display poor in situ effector functions. NKG2D is an activating receptor found on NK and CD8 C T cells and plays a role in immunosurveillance of CLL. In this study, we developed mono-and dual-targeting triplebodies utilizing a natural ligand for human NKG2D receptor (ULBP2) to retarget NK cells against tumor cells. Triplebodies in both formats showed better ability to induce NK-cell-dependent killing of target cells compared to bispecific counterparts. A mono-targeting triplebody ULBP2-aCD19-aCD19 successfully triggered NK cell effector functions against CLL cell line MEC1 and primary tumor cells in allogenic and autologous settings. Additionally, a dual-targeting triplebody ULBP2-aCD19-aCD33 specific for two distinct tumor-associated antigens was developed to target antigen loss variants, such as mixed lineage leukemia (MLL). Of note, this triplebody exhibited cytotoxic activity against CD19/CD33 double positive cells and retained its binding features even in the absence of one of the tumor antigens. Further, ULBP2-aCD19-aCD19 showed significant in vivo activity in immune-deficient (NSG) mouse model transplanted with CLL cell line as target cells and human immune cells as an effector population providing a proof-of-principle for this therapeutic concept.

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A new method for in vitro expansion of cytotoxic human CD3−CD56+ natural killer cells

Cited by 118 publications

References 23 publications

Expansion of natural killer (NK) and natural killer-like T (NKT)-cell populations derived from patients with B-chronic lymphocytic leukemia (B-CLL): a potential source for cellular immunotherapy

Expansion of natural killer (NK) and natural killer-like T (NKT)-cell populations derived from patients with B-chronic lymphocytic leukemia (B-CLL): a potential source for cellular immunotherapy

Immobilized MHC class I chain-related protein A synergizes with IL-15 and soluble 4-1BB ligand to expand NK cells with high cytotoxicity ex vivo

Mono- and dual-targeting triplebodies activate natural killer cells and have anti-tumor activityin vitroandin vivoagainst chronic lymphocytic leukemia

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