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The Synergistic Use of IL-15 and IL-21 for the Generation of NK Cells From CD3/CD19-Depleted Grafts Improves Their ex vivo Expansion and Cytotoxic Potential Against Neuroblastoma: Perspective for Optimized Immunotherapy Post Haploidentical Stem Cell Transplantation.

ABSTRACT: Neuroblastoma (NB) is the most common solid extracranial tumor in childhood. Despite therapeutic progress, prognosis in high-risk NB is poor and innovative therapies are urgently needed. Therefore, we addressed the potential cytotoxic capacity of interleukin (IL)-activated natural killer (NK) cells compared to cytokine-induced killer (CIK) cells for the treatment of NB. NK cells were isolated from peripheral blood mononuclear cells (PBMCs) by indirect CD56-enrichment or CD3/CD19-depletion and expanded with different cytokine combinations, such as IL-2, IL-15, and/or IL-21 under feeder-cell free conditions. CIK cells were generated from PBMCs by ex vivo stimulation with interferon-?, IL-2, OKT-3, and IL-15. Comparative analysis of expansion rate, purity, phenotype and cytotoxicity was performed. CD56-enriched NK cells showed a median expansion rate of 4.3-fold with up to 99% NK cell content. The cell product after CD3/CD19-depletion consisted of a median 43.5% NK cells that expanded significantly faster reaching also 99% of NK cell purity. After 10-12 days of expansion, both NK cell preparations showed a significantly higher median cytotoxic capacity against NB cells relative to CIK cells. Remarkably, these NK cells were also capable of efficiently killing NB spheroidal 3D culture in long-term cytotoxicity assays. Further optimization using a novel NK cell culture medium and a prolonged culturing procedure after CD3/CD19-depletion for up to 15 days enhanced the expansion rate up to 24.4-fold by maintaining the cytotoxic potential. Addition of an IL-21 boost prior to harvesting significantly increased the cytotoxicity. The final cell product consisted for the major part of CD16^-, NCR-expressing, poly-functional NK cells with regard to cytokine production, CD107a degranulation and antitumor capacity. In summary, our study revealed that NK cells have a significantly higher cytotoxic potential to combat NB than CIK cell products, especially following the synergistic use of IL-15 and IL-21 for NK cell activation. Therefore, the use of IL-15+IL-21 expanded NK cells generated from CD3/CD19-depleted apheresis products seems to be highly promising as an immunotherapy in combination with haploidentical stem cell transplantation (SCT) for high-risk NB patients.

SUBMITTER: Heinze A

PROVIDER: S-EPMC6901699 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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The Synergistic Use of IL-15 and IL-21 for the Generation of NK Cells From CD3/CD19-Depleted Grafts Improves Their <i>ex vivo</i> Expansion and Cytotoxic Potential Against Neuroblastoma: Perspective for Optimized Immunotherapy Post Haploidentical Stem Cell Transplantation.

Heinze Annekathrin A Grebe Beatrice B Bremm Melanie M Huenecke Sabine S Munir Tasleem Ah TA Graafen Lea L Frueh Jochen T JT Merker Michael M Rettinger Eva E Soerensen Jan J Klingebiel Thomas T Bader Peter P Ullrich Evelyn E Cappel Claudia C

Frontiers in immunology 20191203

Neuroblastoma (NB) is the most common solid extracranial tumor in childhood. Despite therapeutic progress, prognosis in high-risk NB is poor and innovative therapies are urgently needed. Therefore, we addressed the potential cytotoxic capacity of interleukin (IL)-activated natural killer (NK) cells compared to cytokine-induced killer (CIK) cells for the treatment of NB. NK cells were isolated from peripheral blood mononuclear cells (PBMCs) by indirect CD56-enrichment or CD3/CD19-depletion and ex ...[more]

PMID: 31849984

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Project description:BackgroundT-cell depleted (TCD) haploidentical transplantation using CD3+/CD19+ and TCRαβ+/CD19+ depletion techniques has been increasingly used in children with hematological malignancies. We present a retrospective study aimed to compare transplant outcomes in children with leukemia receiving a TCD haploidentical transplant using either CD3+/CD19+ or TCRαβ+/CD19+ platforms.MethodsA total of 159 children with leukemia (ALL=80) (AML=79) that received a TCD haploidentical transplantation using either CD3+/CD19+ (n=79) or TCRαβ+/CD19+ (n=80) platforms between 2005 and 2020 were included. Median age was 9 years in both groups. There were no differences in patient, donor, and transplant characteristics between groups except for donor KIR B genotype more frequent in the TCRαβ+/CD19+ group (91%) than in the CD3+/CD19+ group (76%) (p=0.009) and a high number of NK+ cells and lower CD19+ cells infused in the TCRαβ+/CD19+ group (35.32x106/kg and 0.06 x106/Kg) than in the CD3+/CD19 group (24.6x106/Kg and 0.25 x106/Kg) (p=0.04 and p=0.0001), respectively. Conditioning was based on TBF. Median follow-up for survivors was 11 years (range; 8-16 y) in CD3+/CD19+ group and 5 years (range; 2-9 y) in the TCRαβ+/CD19+ group.ResultsEngraftment kinetics were similar in both groups (13 days for neutrophils and 10 days for platelets). There was no difference in the incidence of acute GvHD II-IV (29 ± 5% in the CD3+/CD19+ group vs 38 ± 5% in the TCRαβ+/CD19+ group) and chronic GvHD (32 ± 5% vs 23 ± 4%, respectively). NRM was 23 ± 5% in the CD3+/CD19+group vs 21 ± 4% in the TCRαβ+/CD19+group. Relapse incidence was also similar, 32 ± 5% vs 34 ± 6%, respectively. DFS and OS were not different (45 ± 5% vs 45 ± 6% and 53 ± 6% vs 58 ± 6% respectively). As there were no differences on transplant outcomes between groups, we further analyzed all patients together for risk factors associated with transplant outcomes. On multivariate analysis, we identified that early disease status at transplant (HR: 0.16; 95%CI (0.07-0.35) (p=0.0001), presence of cGvHD (HR: 0.38; 95%CI (0.20-0.70) (p= 0.002), and donor KIR-B genotype (HR: 0.50; 95%CI (0.32-0.90) (p=0.04) were associated with better DFS.ConclusionsOur data suggest that there are no advantages in transplant outcomes between TCD platforms. Risk factors for survival are dependent on disease characteristic, donor KIR genotype, and chronic GvHD rather than the TCD platform used.

Dataset Information

The Synergistic Use of IL-15 and IL-21 for the Generation of NK Cells From CD3/CD19-Depleted Grafts Improves Their ex vivo Expansion and Cytotoxic Potential Against Neuroblastoma: Perspective for Optimized Immunotherapy Post Haploidentical Stem Cell Transplantation.

Publications

The Synergistic Use of IL-15 and IL-21 for the Generation of NK Cells From CD3/CD19-Depleted Grafts Improves Their <i>ex vivo</i> Expansion and Cytotoxic Potential Against Neuroblastoma: Perspective for Optimized Immunotherapy Post Haploidentical Stem Cell Transplantation.

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