scholarly journals In Vivo Survival and Homeostatic Proliferation of Natural Killer Cells

2003 ◽  
Vol 197 (8) ◽  
pp. 967-976 ◽  
Author(s):  
Martin Prlic ◽  
Bruce R. Blazar ◽  
Michael A. Farrar ◽  
Stephen C. Jameson
Keyword(s):  
T Cells ◽  
Natural Killer Cells ◽  
Nk Cells ◽  
Natural Killer ◽  
Cell Sorting ◽  
Functional Activity ◽  
Homeostatic Proliferation ◽  
Killer Cells ◽  
Bystander Cells

While the specificity and development of natural killer (NK) cells have been intensely studied, little is known about homeostasis of the mature NK population. Here we show that mouse NK cells undergo homeostatic proliferation when transferred into NK-deficient Rag−/− γC−/− hosts. Normal NK functional activity is maintained during this process, although there are some changes in NK phenotype. Using cell sorting, we demonstrate that mature (Mac-1hi) NK cells undergo homeostatic proliferation in an NK-deficient environment, yet immature (Mac-1lo) NK cells also proliferate in such hosts. We find that mature NK cells survive but do not proliferate in hosts which possess an endogenous NK pool. However, we go on to show that mature NK survival is critically dependent on interleukin (IL)-15. Surprisingly, NK survival is also compromised after transfer of cells into IL-15Rα−/− mice, implying that IL-15 responsiveness by bystander cells is critical for NK maintenance. These data imply that, similar to T cells, homeostasis of the NK pool is much more dynamic than previously appreciated and this may be relevant to manipulation of NK cells for therapeutic purposes.

Autologous Expanded Natural Killer Cells As a New Therapeutic Option for High-Risk Myeloma

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 2918-2918
Author(s):  
Tarun K. Garg ◽  
Junaid Khan ◽  
Susann Szmania ◽  
Amy D Greenway ◽  
Joshuah D Lingo ◽  
...  
Keyword(s):  
High Risk ◽  
Natural Killer Cells ◽  
Nk Cells ◽  
Natural Killer ◽  
Cell Line ◽  
Nk Cell ◽  
Tumor Burden ◽  
Cytolytic Activity ◽  
Killer Cells

Abstract Abstract 2918 Natural killer cells (NK) have the unique ability to kill target cells without priming. While their therapeutic potential against various malignancies is becoming more apparent, it has been restricted to the allogeneic setting; NK cells are inhibited by autologous targets by engaging killer immunoglobulin-like receptors with their ligands. Another major challenge to the clinical utility of NK cells is obtaining a sufficient number of NK cells for infusion. Co-culture of blood mononuclear cells (PBMNC) with the leukemic cell line K562, genetically modified to express membrane-bound IL15 and the co-stimulatory molecule 41BBL (K562mbIL15-41BBL) in the presence of IL2 results in robust expansion and activation of NK cells. To determine if NK cells derived from myeloma (MM) patients can be used therapeutically in the autologous setting, we explored the expansion of NK cells from MM patients, their gene expression profiles (GEP), and their ability to kill autologous and allogeneic MM cells from high-risk patients in vitro and in vivo, and compared these to NK cells from healthy donors (HD). PBMNC from MM patients (N=30) co-cultured with irradiated K562mbIL15-41BBL cells expanded a median of 351 fold (range20–10, 430), comparable to the expansion of HD-derived NK cells (N=15, median 803, range 127–1, 727; p=0.5). GEP of MM non-exp-NK differed from HD non-exp-NK in the expression of only one gene (PRKCi), underexpessed in MM (false discovery rate (FDR) <0.05, p-value <3×10−10). GEP of exp-NK cells from both MM patients and HD was very different from non-exp-NK cells (8 pairs each, 10, 639 differentially overexpressed and 26, 057 underexpressed probe sets, FDR <0.05). Genes associated with proliferation, cytolytic activity, activation, adhesion, migration and cell cycle regulation were highly up-regulated in exp-NK cells. Standard chromium release assays demonstrated that MM exp-NK cells killed both allogeneic and autologous primary MM cells more efficiently compared to non-exp-NK cells, via a perforin mediated mechanism. Blocking studies revealed that the natural cytotoxicity receptors, activating receptors, and DNAX accessory molecule (DNAM-1) played a central role in target cell lysis. The killing ability of MM patient and HD derived exp-NK cells was very similar against allogeneic targets, while primary MM targets were more resistant to killing by autologous exp-NK. The anti-MM activity of allogeneic and autologous exp-NK cells was further examined in vivo. NOD/SCID/IL2R γ-null mice were implanted subcutaneously with a human fetal bone, and primary MM cells or luciferase-transfected OPM2 MM cell line were engrafted into the bone. The tumor burden was determined by ELISA for human Ig and/or bio-imaging. The mice were randomized to control and exp-NK treatment groups. A total of 160 ×106 exp-NK cells, in 4 doses 48 hrs apart, were injected in the exp-NK treatment group via tail vein injection. The mice were administered 1000U of IL2 subcu daily to support the NK cells. The mice were bled on days 7, 14, 21 & 28 for the assessment of human Ig by ELISA and enumerating circulating NK cells by flow cytometry. Exp-NK treated mice had a significantly reduced MM burden by ELISA (p<0.04) on day 21, and exp-NK could be detected in the murine blood up to day 28 post-administration in both primary MM and OPM2 tumor bearing mice. The mice were sacrificed and the tumors were harvested after 4 weeks. A noticeable reduction in tumor burden in the exp-NK cell treated mice was confirmed by histology. NK cells were detected by immunohistochemistry (CD57 or CD16) in the hu-bone implants harvested 28 days after infusion. In conclusion, MM patient-derived NK cells have a similar expansion potential, and MM exp-NK cells have cytolytic activity against allogeneic targets similar to those of HD exp-NK cells, and somewhat reduced activity against autologous targets. These exp-NK cells have significant activity against the aggressive cell line OPM2 and high-risk autologous primary MM cells in vivo. Exp-NK cells trafficked to MM tumors and persisted in the myelomatous hu bone microenvironment for 4 weeks. The anti-MM activity of autologous exp-NK cells is exciting and avails a new therapeutic avenue for patients with GEP-defined high-risk disease. A phase II clinical trial of allogeneic and autologous exp-NK cell therapy for relapsed/refractory high-risk MM is in progress at our institution. Disclosures: No relevant conflicts of interest to declare.

scholarly journals The aryl hydrocarbon receptor is required for the maintenance of liver-resident natural killer cells

2016 ◽  
Vol 213 (11) ◽  
pp. 2249-2257 ◽  
Author(s):  
Luhua H. Zhang ◽  
June Ho Shin ◽  
Mikel D. Haggadone ◽  
John B. Sunwoo
Keyword(s):  
Natural Killer Cells ◽  
Aryl Hydrocarbon Receptor ◽  
Nk Cells ◽  
Natural Killer ◽  
Nk Cell ◽  
Memory Function ◽  
Contact Hypersensitivity ◽  
Killer Cells ◽  
Aryl Hydrocarbon

A tissue-resident population of natural killer cells (NK cells) in the liver has recently been described to have the unique capacity to confer immunological memory in the form of hapten-specific contact hypersensitivity independent of T and B cells. Factors regulating the development and maintenance of these liver-resident NK cells are poorly understood. The aryl hydrocarbon receptor (AhR) is a transcription factor modulated by exogenous and endogenous ligands that is important in the homeostasis of immune cells at barrier sites, such as the skin and gut. In this study, we show that liver-resident NK (NK1.1+CD3−) cells, defined as CD49a+TRAIL+CXCR6+DX5− cells in the mouse liver, constitutively express AhR. In AhR−/− mice, there is a significant reduction in the proportion and absolute number of these cells, which results from a cell-intrinsic dependence on AhR. This deficiency in liver-resident NK cells appears to be the result of higher turnover and increased susceptibility to cytokine-induced cell death. Finally, we show that this deficiency has functional implications in vivo. Upon hapten exposure, AhR−/− mice are not able to mount an NK cell memory response to hapten rechallenge. Together, these data demonstrate the requirement of AhR for the maintenance of CD49a+TRAIL+CXCR6+DX5− liver-resident NK cells and their hapten memory function.

Effects of Anti-CCR4 Antibody (KW-0761) on Regulatory T Cells and Natural Killer Cells in Patients with Cutaneous T-Cell Lymphoma

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 444-444 ◽  
Author(s):  
Xiao Ni ◽  
Jeffrey L. Jorgensen ◽  
Meghali Goswami ◽  
Pramoda Challagundla ◽  
William K Decker ◽  
...  
Keyword(s):  
T Cells ◽  
Natural Killer Cells ◽  
Regulatory T Cells ◽  
Nk Cells ◽  
Natural Killer ◽  
Board Of Directors ◽  
Research Funding ◽  
Advisory Committees ◽  
Killer Cells ◽  
Malignant T Cells

Abstract Abstract 444 The CC chemokine receptor 4 (CCR4) is expressed on malignant T-cells in cutaneous T-cell lymphoma (CTCL) and also on the surface of regulatory T cells (T-regs). T-regs also express the transcription factor, foxp3 and suppress effector immune cells, including natural killer (NK) cells. Thus, increased T-regs in the tumor microenvironment is associated with impaired anti-tumor immunity. KW-0761, a defucosylated, humanized monoclonal antibody, binds to CCR4 and induces effective antibody-dependent cellular cytotoxicity (ADCC) against CCR4+ malignant T-cells. The phase I/II clinical trial to determine the safety and efficacy of anti-CCR4 antibody (KW-0761) included a translational component to evaluate its effects on T-regs and NK cells in CTCL patients. Peripheral blood mononuclear cells (PBMCs) were collected from 20 patients [10 with mycosis fungoides (MF) and 10 with Sézary syndrome(SS)] pre- and post-treatment at two centers for flow cytometry analysis of CD3+CD4+CD25+CD127- T-regs, CCR4+ T-regs, and CD3-CD56+CD16+ NK cell subsets. Total RNA was extracted from PBMCs, and foxp3 and CCR4 mRNA were quantified using real-time PCR. The standard 4-hour 51Cr release assay was used to assess the cytotoxicity of NK cells. Fifteen of 20 patients (75.0%) had detectable CD3+CD4+CD25+CD127- T-regs (1.26±1.09 % or 33.79±46.88 /μl) at baseline with 60 –100 % of T-regs positive for CCR4. After 4–6 weeks of treatment with anti-CCR4 antibody (KW-0761), all 15 patients had a decrease in T-reg numbers (0.39±0.49 % or 5.65±8.95 /μl, *p<0.05, Figure 1). CCR4+ T-regs were significantly reduced from an average of 67.2% to 24.6% (p<0.01). In parallel, foxp3 and CCR4 mRNA levels were also significantly decreased from baseline levels (foxp3: from 0.57±0.91 to 0.07±0.08, **p<0.01, Figure 1; CCR4: from 23.40±33.50 to 1.73±2.35, p<0.05). The CD3-CD56+CD16+ NK cells were found at baseline in all 15 patients tested. Of 14 paired samples, 10 (71.4%) had increased NK cells after 4–16 weeks of treatment (pre-treatment: 16.02±15.86 % vs. post-treatment: 22.64±13.93 %, p=0.05, Figure 2) with reduced numbers of T-regs. Five of 6 patients studied also showed a dose dependent increase in NK cell cytotoxicity to target cells by 51Cr release assay (Figure 2). Ten patients with SS had a lower NK cells (13.37±18.48 %) and higher foxp3 (0.81±1.19) and CCR4 mRNA (34.60±39.90) at baseline compared to ten patients with MF (19.04±12.98%, 0.31±0.31; 10.94±20.07) respectively. Seven paired samples from SS patients all had increased NK cells post-treatment with a reduction of T-regs and foxp3 and CCR4 mRNA. Six of 7 SS patients had blood improvement with 3 complete and 3 partial blood responses.Figure 1.Effect of KW-0761 on regulatory T cells in CTCL patients.Figure 1. Effect of KW-0761 on regulatory T cells in CTCL patients.Figure 2.Effect of KW-0761 on natural killer cells in CTCL patients.Figure 2. Effect of KW-0761 on natural killer cells in CTCL patients. Our results suggest that in addition to ADCC towards malignant T-cells, the anti-neoplastic activity of the anti-CCR4 antibody (KW-0761) may include a reduction in T-regs in most CTCL patients and a subsequent increase in NK numbers and function in some patients. Follow up studies need to be performed to confirm these findings. Disclosures: Ni: KYOWA HAKKO KIRIN CO., LTD: Research Funding. Kim:kyowa: Consultancy, Research Funding; Merck: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Allos: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Seattle Genetics: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Millenium: Consultancy. Duvic:KYOWA HAKKO KIRIN CO., LTD: Research Funding.

scholarly journals Memory-Like Natural Killer Cells

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. SCI-8-SCI-8 ◽  
Author(s):  
Melissa M Berrien-Elliott ◽  
Julia A Wagner ◽  
Amanda F Cashen ◽  
Todd A Fehniger
Keyword(s):  
Natural Killer Cells ◽  
Nk Cells ◽  
Natural Killer ◽  
Target Cell ◽  
Research Funding ◽  
Nk Cell ◽  
Cytokine Receptors ◽  
Inhibitory Receptor ◽  
Killer Cells

Abstract Natural killer (NK) cells, classically considered to be part of the innate immune system, are specialized for rapid responses that protect the host against pathogens and eliminate malignant cells. These functional responses are multi-faceted, and include not only direct target cell killing, but also production of cytokines and chemokines, proliferative expansion, and cross-talk with various immune cells to help orchestrate cellular immunity. NK cell recognition of a target cell is controlled by the integration of signals from germline-DNA encoded activating, inhibitory, and cytokine receptors. NK cell tolerance toward healthy host cells arises via education or licensing that requires self-inhibitory receptor engagement. Over the past decade, it has become clear that NK cells have the capacity to remember prior activation events, including hapten-exposure,1 viral infection,2 and combined cytokine stimulation.3 These studies have defined "memory", "adaptive", and "memory-like" responses by NK cells within both murine and human immune systems, which can result in long-lived NK cell populations with enhanced effector functionality.4-6 While certain types of NK cell "memory" can be specific to the original stimulation, other types of "memory-like" responses are non-specific, providing flexibility in the enhanced response to subsequent activating triggers.7 Cytokine-induced memory-like NK cells were originally discovered in mice, following a brief exposure to the potent activating combination of IL-12, IL-15 and IL-18.3 After this activation, murine memory-like NK cells differentiated in vivo, and demonstrated enhanced IFN-γ recall responses to IL-12 and IL-15 re-stimulation, even after extensive cell division. Subsequent studies identified human IL-12/15/18-induced memory-like NK cells that displayed enhanced function after re-stimulation via cytokine receptors, activating receptors, or tumor targets.8 Adoptive transfer of murine memory-like NK cells into syngeneic or immunodeficient mice resulted in an enhanced ability to control lymphomas and solid tumors in vivo,9 as well as the ability to persist in the recipient for months. In addition, memory-like NK cell differentiation restored the anti-tumor function of unlicensed NK cells, demonstrating that cytokine receptor signals can overcome a lack of NK cell education.10 Other studies showed that memory-like NK cells can ignore inhibitory receptor signals, and have enhanced anti-leukemia responses.11 Based on these pre-clinical findings, we translated allogeneic memory-like NK cells into the clinic in a first-in-human adoptive cell therapy trial for patients with relapsed/refractory (rel/ref) myeloid malignancies. This study demonstrated that rel/ref AML patients were able to safely receive IL-12/15/18-activated donor NK cells (up to 10x106/kg) without developing cytokine release syndrome, neurotoxicity, or graft-versus-host disease. Immune monitoring revealed that memory-like NK cells expanded, trafficked to the bone marrow, and exhibited enhanced anti-leukemia function ex vivo. Clinical responses (CR/CRi) were observed in >50% of patients with active rel/ref AML.11 Ongoing studies are exploring memory-like NK cell adoptive immunotherapy in a phase 2 trial for rel/ref AML, combined with same-donor allogeneic hematopoietic cell transplantation (HCT), and for relapse after allogeneic HCT. Active areas of investigation in memory-like NK cell biology and therapeutics include defining mechanisms that regulate memory-like differentiation and enhanced function, elucidating memory-like NK cell checkpoints, evaluating autologous memory-like NK cell responses against cancers, and developing strategies to enhance memory-like NK cell targeting of resistant malignancies. O'Leary JG, Goodarzi M, Drayton DL, Yu H, von Andrian UH. T cell- and B cell-independent adaptive immunity mediated by natural killer cells. Nat Immunol. 2006;7(5):507-16. Sun JC, Beilke JN, Lanier LL. Adaptive immune features of natural killer cells. Nature. 2009;457(7229):557-61. Cooper MA, Elliott JM, Keyel PA, et al. Cytokine-induced memory-like natural killer cells. Proc Natl Acad Sci USA. 2009;106(6):1915-9. Rölle A, Pollmann J, Cerwenka A. Memory of Infections: An Emerging Role for Natural Killer Cells. PLoS Pathog. 2013;9(9):1-3. Schlums H, Cichocki F, Tesi B, et al. Cytomegalovirus Infection Drives Adaptive Epigenetic Diversification of NK Cells with Altered Signaling and Effector Function. Immunity. 2015;42(3):443-456. Lee J, Zhang T, Hwang I, et al. Epigenetic Modification and Antibody-Dependent Expansion of Memory-like NK Cells in Human Cytomegalovirus-Infected Individuals. Immunity. 2015;42(3):431-442. Fehniger TA, Cooper MA. Harnessing NK Cell Memory for Cancer Immunotherapy. Trends Immunol. 2016;Epub Oct 2:10.1016/j.it.2016.09.005. Romee R, Schneider SE, Leong JW, et al. Cytokine activation induces human memory-like NK cells. Blood. 2012;120(24):4751-4760. Ni J, Miller M, Stojanovic A, Garbi N, Cerwenka A. Sustained effector function of IL-12/15/18-preactivated NK cells against established tumors. J Exp Med. 2012;209(13):2351-2365. Wagner JA, Berrien-Elliott MM, Rosario M, et al. Cytokine-Induced Memory-Like Differentiation Enhances Unlicensed NK Cell Anti-Leukemia and FcγRIIIa-Triggered Responses. Biol. Blood Marrow Transplant. 2016;dx.doi.org: Romee R, Rosario M, Berrien-Elliott MM, et al. Cytokine-induced memory-like natural killer cells exhibit enhanced responses against myeloid leukemia. Sci. Transl. Med. 2016;8(357):357:doi: 10.1126/scitranslmed.aaf2341. Figure. Figure. Disclosures Fehniger: Altor BioScience: Research Funding; Cyto-Sen Therapeutics: Consultancy; Celgene: Research Funding; NIH/NCI: Other: R01 CA205239, P50CA171963; Affimed: Research Funding.

The Opposite Effect of Tumor-Infiltrating Natural Killer Cells on In Vivo Priming of Tumor-Specific CD8+ T Cells and CD4+ T Cells

Immunobiology ◽  
1996 ◽  
Vol 195 (2) ◽  
pp. 172-186 ◽  
Author(s):  
Hiroshi Terao ◽  
Mamoru Harada ◽  
Shin Kurosawa ◽  
Yoshihiro Shinomiya ◽  
Osamu Ito ◽  
...  
Keyword(s):  
T Cells ◽  
Natural Killer Cells ◽  
Natural Killer ◽  
Cd8 T Cells ◽  
Cd4 T Cells ◽  
Opposite Effect ◽  
Killer Cells

scholarly journals 128 KIR haplotype can inform donor selection production of allogeneic memory-like CAR NK cells for clinical application

2021 ◽  
Vol 9 (Suppl 3) ◽  
pp. A137-A137
Author(s):  
Hadia Lemar ◽  
Anmol Vohra ◽  
Ming-Hong Xie ◽  
Ivan Chan ◽  
Sasha Lazetic ◽  
...  
Keyword(s):  
Natural Killer Cells ◽  
Nk Cells ◽  
Natural Killer ◽  
Nk Cell ◽  
Killer Cell ◽  
Donor Selection ◽  
Myelogenous Leukemia ◽  
Killer Cells ◽  
Group B

BackgroundNK cells expanded on membrane-bound (mb) IL-15 and 41BBL expressing K562 stimulatory cells (NKSTIM) for clinical use can be genetically modified to express activating chimeric receptors.1 2 3 NK cells activated in the presence of IL-12, IL-15 and IL-18 develop cytokine induced memory-like (CIML) phenotype and function; these cells have shown clinical promise.4 Additionally, HSCT AML transplants using NK KIR Haplotype Group B donors with better and best Group B profiles (≥2 activating genes) show better survival.5 6 Here we investigate whether KIR profiles impact healthy allogeneic donor NK cell function and phenotype when these cells are expanded on NKSTIM in the presence of IL-12 and IL-18 (12–18).MethodsHealthy donor PBMC NK were genotyped for HLA and KIR and expanded on K562-mbIL15-41BBL stimulatory cells with IL-2 alone or with IL-2 plus IL-12 and IL-18 (12–18). Expanded NK were transduced with CAR constructs including CD19, and then evaluated for NK cell expansion, cytokine secretion, RNA profiles, cytotoxicity against tumor lines, and cell surface phenotypes. Expanded CD19 NK donors with varying numbers of activating KIR vs inhibitory KIR were tested for effector function, and these donors were then tested for in vivo efficacy and pharmacokinetics. A KIR ranking score was developed by considering both the number of activating and inhibitory KIR genes expressed by each donor. This score was correlated with functional properties of CAR NK cells.ResultsAddition of 12–18 to the K562-mbIL15-41BBL stimulatory cells improves CD19-CAR NK potency 2-fold relative to the stimulatory cell line alone (P=.02) while NK cell expansion is unchanged. 12–18 also drove an increase in effector cytokine accumulation on exposure of CAR-NK to CD19 tumor. CIML CAR NK cells from donors with higher KIR scoring also had higher cytotoxicity (Pearson’s R=0.74, P=0.006); this correlation was not observed following expansion in the absence of 12–18. 12–18 also drove more potent in vivo activity against tumor with an increased presence of circulating NK cells over 4 weeks in the mice.ConclusionsCIML CAR NK cells derived from donors with favorable KIR scoring have greater cytotoxic activity, effector cytokine production, and in vivo pharmacokinetics and efficacy. These findings may provide an important criterion for donor selection in the development of more robust and potent engineered NK cells for clinical use.ReferencesLapteva N, Durett AG, Sun J, Rollins LA, Huye LL, Fang J, Dandekar V, Mei Z, Jackson K, Vera J, Ando J, Ngo MC, Coustan-Smith E, Campana D, Szmania S, Garg T, Moreno-Bost A, Vanrhee F, Gee AP, Rooney CM. Large-scale ex vivo expansion and characterization of natural killer cells for clinical applications. Cytotherapy 2012;14(9):1131–1143.Chihaya I, Iwamoto S, Campana D. Genetic modification of primary natural killer cells overcomes inhibitory signals and induces specific killing of leukemic cells. Blood 2005;106:376–383.Yang Y, Connolly J, Shimasaki N, Mimura K, Kono K, Campana D. A Chimeric Receptor with NKG2D Specificity Enhances Natural Killer Cell Activation and Killing of Tumor Cells. Cancer Res 2013;73(6):1777–1786.Romee R, Rosario M, Berrien-Elliott MM, Wagner JA, Jewell BA, Schappe T, Leong JW, Abdel-Latif S, Schneider SE, Willey S, Neal CC, Yu L, Oh ST, Lee YS, Mulder A, Claas F, Cooper MA, Fehniger TA. Cytokine-induced memory-like natural killer cells exhibit enhanced responses against myeloid leukemia. Sci Trans Med 2016;8(357): 357ra123.Cooley S, Weisdorf DJ, Guethlein LA, Klein JP, Wang T, Le CT, Marsh SGE, Geraghty D, Spellman S, Haagenson MD, Ladner M, Trachtenberg E, Parham P, and Miller JS. Donor selection for natural killer cell receptor genes leads to superior survival after unrelated transplantation for acute myelogenous leukemia. Blood 2010;116(14):2414–2419.Cooley S, Weisdorf DJ, Guethlein LA, Klein JP, Wang T, Marsh SGE, Spellman S, Haagenson MD, Saeturn K, Ladner M, Trachtenberg E, Parham P, and Miller JS. Donor Killer Cell Ig-like Receptor B Haplotypes, Recipient HLA-C1, and HLA-C Mismatch Enhance the Clinical Benefit of Unrelated Transplantation for Acute Myelogenous Leukemia. JI, 2014;192(10):4592–600.Ethics ApprovalAnimal studies were conducted with IACUC approval.

scholarly journals Natural Killer Cells Activated By Oncolytic Reovirus Enhance Cetuximab Mediated Antibody Dependent Cellular Cytotoxicity in an in Vitro and In Vivo Model of Colorectal Cancer

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 3439-3439
Author(s):  
Xing Zhao ◽  
Narendiran Rajasekaran ◽  
Cariad Chester ◽  
Atsushi Yonezawa ◽  
Suparna Dutt ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Natural Killer Cells ◽  
Nk Cells ◽  
Natural Killer ◽  
Colorectal Cancer Cell Line ◽  
In Vivo Model ◽  
Cellular Cytotoxicity ◽  
Antibody Dependent Cellular Cytotoxicity ◽  
Killer Cells

Abstract The naturally occurring oncolytic virus, reovirus, exhibits cytotoxic effects on cancer cells. Reovirus is currently being tested in multiple clinical trials for the treatment of different cancers. In addition, they also activate the innate and adaptive immune responses targeting immune cells like dendritic cells and macrophages. In this study we investigated the direct effect of reovirus on Natural Killer cells (NK cells) and its effect on NK cell mediated antibody dependent cellular cytotoxicity (ADCC) against the EGFR (Epidermal Growth Factor) positive colorectal cancer cell line: DLD-1 (KRAS mutant). NK cells isolated from human PBMCs were cultured with 1pfu of reovirus for 12 hrs and subsequently co-cultured with DLD-1 cells coated with increasing concentrations anti-EGFR antibody cetuximab. ADCC was measured after 4 hrs using a lactate dehydrogenase (LDH) based cytotoxicity assay. We observed that the reovirus treated NK cells (Reo-NK cells) exhibited a ~16-fold increase in cytotoxicity against DLD-1 (16.3% ±1.5, n=3) compared to untreated NK cells (NK), even in the absence of any cetuximab. In the presence of cetuximab, NK cells showed a dose dependent increase in ADCC, with maximum ADCC, observed at 0.1 µg/ml of cetuximab (DLD-1+NK: 33.4%± 7.1, n=3). Interestingly, Reo-NK cells showed maximum ADCC even at 0.01 µg /ml of cetuximab (DLD-1+Reo-NK: 39.1±7.4, DLD-1+NK: 26.7±2.4%, n=3). Reo-NK cells also exhibited an increased expression of activation marker CD69 (Reo-NK: 70.4%, NK: 35.2%) and degranulation maker CD107a (Reo-NK: 14.6%; NK: 4.45%) compared to the untreated NK cells. We further characterized the Reo-NK cells by using the HIMChip microarray platform; a custom Agilent SurePrint HD 8x15k format array containing over 7,000 unique probes for over 4,274 human immune-related genes. In ingenuity pathway analysis, we observed that the Interferon pathway (2.13E-20) and pathway controlling activation of IRF by cytosolic pattern recognition receptors (1.27E-11) were the predominant pathways observed in the Reo-NK cells. These results suggest an interferon-mediated response could be contributing to the increased cytoxicity of the NK cells. In an in vivo study, DLD-1 cells were grown subcutaneously in athymic nude mice and injected intravenously with reovirus (5x 108 pfu), followed by intraperitoneal injection of Cetuximab (200 ug/mice) every week. We observed a significant regression of tumors in the Reovirus+Cetuximab combination group compared to the Reovirus treated (Reovirus+Cet: 349.9 mm3, Reovirus: 623.8 mm3; n=9; P=0.0028) or Cetuximab treated (Reovirus+Cet: 349.9 mm3, Cet: 730.5 mm3;n=9; P= 0.030) groups on day 28 post treatment. Thus, in this study our results demonstrated that human NK cells when treated with reovirus show increases in activation, degranulation and cytotoxicity when compared to untreated NK cells. Further, in the in vivo model we observed increased tumor regression in mice treated with reovirus in combination with cetuximab. We propose that reovirus activated NK cells are a potential candidate for cell based immunotherapy in combination with FDA approved tumor targeting antibodies to treat malignancies, including lymphomas. Further studies are ongoing to investigate the underlying mechanisms that contribute to the increase in cytotoxicity by NK cells treated with reovirus. Disclosures No relevant conflicts of interest to declare.

In vitro expanded human invariant natural killer T-cells promote functional activity of natural killer cells

2008 ◽  
Vol 129 (1) ◽  
pp. 145-154 ◽  
Author(s):  
María Moreno ◽  
Johan W. Molling ◽  
Silvia von Mensdorff-Pouilly ◽  
René H.M. Verheijen ◽  
B. Mary E. von Blomberg ◽  
...  
Keyword(s):  
T Cells ◽  
Natural Killer Cells ◽  
Natural Killer ◽  
Functional Activity ◽  
Natural Killer T Cells ◽  
Killer Cells ◽  
Killer T Cells ◽  
Invariant Natural Killer ◽  
Natural Killer T
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