scholarly journals The Molecular Imaging of Natural Killer Cells

2018 ◽  
Vol 17 ◽  
pp. 153601211879481 ◽  
Author(s):  
Mariya Shapovalova ◽  
Sean R. Pyper ◽  
Branden S. Moriarity ◽  
Aaron M. LeBeau
Keyword(s):  
Molecular Imaging ◽  
Natural Killer Cells ◽  
Nk Cells ◽  
Natural Killer ◽  
Solid Tumors ◽  
Nk Cell ◽  
Human Leukocyte ◽  
Cytotoxic Lymphocytes ◽  
Killer Cells

The recent success of autologous T cell-based therapies in hematological malignancies has spurred interest in applying similar immunotherapy strategies to the treatment of solid tumors. Identified nearly 4 decades ago, natural killer (NK) cells represent an arguably better cell type for immunotherapy development. Natural killer cells are cytotoxic lymphocytes that mediate the direct killing of transformed cells with reduced or absent major histocompatibility complex (MHC) and are the effector cells in antibody-dependent cell-mediated cytotoxicity. Unlike T cells, they do not require human leukocyte antigen (HLA) matching allowing for the adoptive transfer of allogeneic NK cells in the clinic. The development of NK cell-based therapies for solid tumors is complicated by the presence of an immunosuppressive tumor microenvironment that can potentially disarm NK cells rendering them inactive. The molecular imaging of NK cells in vivo will be crucial for the development of new therapies allowing for the immediate assessment of therapeutic response and off-target effects. A number of groups have investigated methods for detecting NK cells by optical, nuclear, and magnetic resonance imaging. In this review, we will provide an overview of the advances made in imaging NK cells in both preclinical and clinical studies.

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.

123 Natural killer cells engineered with an inducible, responsive genetic construct targeting TIGIT and CD73 to relieve immunosuppression within the GBM microenvironment

2020 ◽  
Vol 8 (Suppl 3) ◽  
pp. A133-A134
Author(s):  
Kyle Lupo ◽  
Sandro Matosevic
Keyword(s):  
Natural Killer Cells ◽  
Natural Killer Cell ◽  
Nk Cells ◽  
Natural Killer ◽  
Solid Tumors ◽  
Nk Cell ◽  
Killer Cell ◽  
Functional Responses ◽  
Killer Cells ◽  
Genetic Construct

BackgroundSolid tumors such as GBM are particularly difficult to treat, being largely resistant to traditional treatments, fueling interest in alternative treatment approaches, including cell-based immunotherapy. Natural killer (NK) cells have emerged as promising effectors to target GBM through genetic modifications and ex vivo manipulation. However, immunosuppressive conditions within the tumor microenvironment (TME) further complicate NK cell-based treatments. Specifically, within the TME tumor cells release of high levels of ATP extracellularly. While intracellular ATP is necessary for cell metabolism, extracellular ATP is converted into adenosine (ADO) by ectonucleotidases CD39 and CD73, both overexpressed on GBM.1 Extracellular ADO induces immunometabolic suppression of NK cells through binding with A2A adenosine receptors (A2ARs) on NK cells, suppressing cytokine secretion, proliferation, and other functional activities. 2–4 Adding to the suppression of NK cells is the interaction between CD155, expressed highly on GBM and other solid tumors, and T cell immunoreceptor with Ig and ITIM domains (TIGIT) expressed on NK cells. This interaction signals inhibition of NK cell cytolytic function, allowing for cancer cell immune-evasion.5,6MethodsTo restore impaired NK cell anti-tumor activity, we have engineered NK cells to concomitantly target CD155 and CD73-induced immunosuppression on GBM using a tumor-responsive genetic construct. The construct is capable of blocking the immunosuppressive CD155/TIGIT interaction, and, upon binding, release a CD73-blocking scFv to inhibit the accumulation of extracellular ADO and mitigate immunosuppression of NK cells. Such localized response enhances specificity and reduces off-target effects of NK-based targeting.ResultsPrimary NK cells were successfully electroporated to express our synthetic TIGIT-synNotch construct, as evidenced by increased expression levels of TIGIT (% and MFI) (figure 1). To evaluate the functionality of engineered NK cells against GBM targets, we tested the cytotoxicity of our engineered NK cells against a primary, patient-derived GBM cell line, GBM43. Overall, cytolytic function of engineered NK cells against GBM was significantly higher than that of non-engineered NK cells, with or without CD73 (10 ug/mL) and TIGIT (50 ug/mL) antibodies, for E:T ratios of 5:1 and 10:1 (figure 2), demonstrating the functional efficacy of our genetic construct. Further, engineered NK cells (T-PNK) expressed significantly higher levels of CD107a in response to GBM43 stimulation than non-engineered PNK at E:T ratios 2.5:1 and 10:1 (figure 3).Abstract 123 Figure 1TIGIT-synNotch gene expressionGene expression (left: %, right, MFI) of electroporated NK cells engineered with anti-CD73 and TIGIT blocking mRNAAbstract 123 Figure 2Engineered NK cell cytotoxicityCytotoxicity of NK cells against GBM43 cells at E:T ratios of 2.5:1, 5:1, and 10:1. NK cells were either un-transfected (with and without CD73 and TIGIT mAbs), transfected with the TIGIT-synNotch construct, or transfected with the TIGIT-synNotch and CD73 genetic constructsAbstract 123 Figure 3Engineered NK cell degranulationCD107a expression measured on transfected and non-transfected NK cells stimulated with GBM43 at E:T ratios of 2.5:1, 5:1, and 10:1ConclusionsOverall, we have shown that co-targeting CD155 and CD73 in a localized, responsive manner can dampen immunosuppression and significantly enhance the killing potential of engineered NK cells against aggressive patient-derived GBM tumors.ReferencesChambers AM, et al. Adenosinergic Signaling Alters Natural Killer Cell Functional Responses. Front. Immunol 2018;9:2533.Chambers AM, Lupo KB & Matosevic S. Tumor microenvironment-induced immunometabolic reprogramming of natural killer cells. Front Immunol 2018;9:2517.Chambers AM, et al. Adenosinergic signaling alters natural killer cell functional responses. Front. Immunol 2018;9:2533.Wang, J., Lupo, K. B., Chambers, A. M. & Matosevic, S. Purinergic targeting enhances immunotherapy of CD73+ solid tumors with piggyBac-engineered chimeric antigen receptor natural killer cells. J. immunotherapy cancer 2018;6:136.Zhang B, et al. Immunoreceptor TIGIT inhibits the cytotoxicity of human cytokine-induced killer cells by interacting with CD155. Cancer Immunol Immunother 2016;65:305–314.Lupo KB & Matosevic S. CD155 immunoregulation as a target for natural killer cell immunotherapy in glioblastoma. J Hematol Oncol 2020;13:76.

scholarly journals 130 Engineered natural killer cells reactively block TIGIT and CD73 in the GBM microenvironment

2021 ◽  
Vol 9 (Suppl 3) ◽  
pp. A139-A139
Author(s):  
Kyle Lupo ◽  
Sandro Matosevic
Keyword(s):  
Natural Killer Cells ◽  
Natural Killer Cell ◽  
Nk Cells ◽  
Natural Killer ◽  
Solid Tumors ◽  
Nk Cell ◽  
Killer Cell ◽  
Functional Responses ◽  
Killer Cells ◽  
Genetic Construct

BackgroundNatural killer (NK) cells have emerged as promising effectors to target GBM and other solid tumors through genetic modifications and ex vivo manipulation. However, immunosuppressive conditions within the tumor microenvironment (TME) and interactions between NK cell activating and inhibitory receptors further complicate NK cell-based treatments. In particular, the T cell immunoreceptor with Ig and ITIM domains (TIGIT) is expressed on NK cells and interacts with CD155 to induce immunosuppression of NK cell cytolytic functions.1 2 Although CD155 also binds with activating receptors DNAM-1 and CD96 on NK cells, spurring NK cell activity, TIGIT has predominantly been reported as having an inhibitory effect on NK cells.3–5 Further, tumor cells release of high levels of ATP extracellularly. While intracellular ATP is necessary for cell metabolism, extracellular ATP is converted into adenosine (ADO) by ectonucleotidases CD39 and CD73, both overexpressed on GBM and other solid tumors.6 Extracellular ADO induces immunometabolic suppression of NK cells through binding with A2A adenosine receptors (A2ARs) on NK cells, suppressing cytokine secretion, proliferation, and other functional activities.7–9 We found that TIGIT and CD73 are effective combination targets in GBM for both primary and iPSC-derived NK cells.MethodsIn order to effectively target immunometabolic reprogramming induced by CD73-produced adenosine and the immunosuppressive TIGIT-CD155 axis, we have engineered NK cells to concomitantly target CD155 and CD73-induced immunosuppression on GBM using a tumor-responsive genetic construct based on the synNotch signaling system. The construct is capable of blocking the immunosuppressive CD155/TIGIT interaction, and, upon binding, release a CD73-blocking scFv to inhibit the accumulation of extracellular ADO and mitigate immunosuppression of NK cells. Such localized response enhances specificity and reduces off-target effects of NK-based targeting.ResultsPrimary NK cells and iPSC-derived NK cells were successfully engineered to express the synthetic TIGIT-synNotch construct, measured through expression of TIGIT. To evaluate the functionality of engineered NK cells against GBM targets, we tested the cytotoxicity of our engineered NK cells against a primary, patient-derived GBM cell line, GBM43. Overall, cytolytic function of engineered NK cells against GBM was significantly higher than that of non-engineered NK cells, with or without CD73 (10 ug/mL) and TIGIT (50 ug/mL) antibodies, for E:T ratios of 5:1 and 10:1, demonstrating the functional efficacy of our genetic construct.ConclusionsOverall, we have shown that co-targeting CD155 and CD73 in a localized, responsive manner can dampen immunosuppression and significantly enhance the killing potential of engineered NK cells against aggressive patient-derived GBM tumors.ReferencesZhang B, et al. Immunoreceptor TIGIT inhibits the cytotoxicity of human cytokine-induced killer cells by interacting with CD155. Cancer Immunol Immunother 2016;65:305–314.Lupo KB & Matosevic S. CD155 immunoregulation as a target for natural killer cell immunotherapy in glioblastoma. J Hematol Oncol 2020;13:76.Hung AL, et al. TIGIT and PD-1 dual checkpoint blockade enhances antitumor immunity and survival in GBM. OncoImmunology 2018; e1466769. doi:10.1080/2162402X.2018.1466769.Mahnke K & Enk, AH. TIGIT-CD155 Interactions in Melanoma: A Novel Co-Inhibitory Pathway with Potential for Clinical Intervention. Journal of Investigative Dermatology 2016; 136, 9–11.Stanietsky N, et al. Mouse TIGIT inhibits NK-cell cytotoxicity upon interaction with PVR: Innate immunity. Eur J Immunol 2013; 43:2138–2150.Chambers AM, et al. Adenosinergic Signaling Alters Natural Killer Cell Functional Responses. Front Immunol 2018;9:2533.Chambers AM, Lupo KB & Matosevic S. Tumor Microenvironment-Induced Immunometabolic Reprogramming of Natural Killer Cells. Front Immunol 2018;9:2517.Chambers AM. et al. Adenosinergic Signaling Alters Natural Killer Cell Functional Responses. Front Immunol 2018;9:2533.Wang J, Lupo KB, Chambers AM & Matosevic S. Purinergic targeting enhances immunotherapy of CD73+ solid tumors with piggyBac-engineered chimeric antigen receptor natural killer cells. J Immunotherapy Cancer 2018;6:136.Ethics ApprovalPrimary human NK cells were obtained from healthy adult donors approved under Purdue University’s Institutional Review Board (IRB) (IRB-approved protocol #1804020540). Donors gave written informed consent prior to taking part in the study.

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 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.

scholarly journals 799 Neoantigen-cytokine-chemokine multifunctional natural killer cell engager for immunotherapy of solid tumors

2021 ◽  
Vol 9 (Suppl 3) ◽  
pp. A834-A834
Author(s):  
Xue Yao ◽  
Sandro Matosevic
Keyword(s):  
Tumor Microenvironment ◽  
Cell Therapy ◽  
Nk Cells ◽  
Natural Killer ◽  
Solid Tumors ◽  
Tumor Cells ◽  
Nk Cell ◽  
Killer Cell ◽  
Nk Cell Therapy

BackgroundThe effectiveness of natural killer (NK) cell-based immunotherapy against solid tumors is limited by the lack of specific antigens and the immunosuppressive tumor microenvironment (TME). Glioblastoma multiforme (GBM) is one such heavily immunosuppressive tumor that has been particularly hard to target and remains without a viable treatment. The development of novel approaches to enhance the efficacy of NK cells against GBM is urgently needed. NK cell engagers (NKCE) have been developed to enhance the efficacy of NK cell therapy.MethodsTo improve the clinical efficacy of NK cell therapy, we are developing a new generation of multi-specific killer engagers, which consists of a neoantigen-targeting moiety, together with cytokine and chemokine-producing domains. Neoantigens are new antigens formed specifically in tumor cells due to genome mutations, making them highly specific tools to target tumor cells. Our engager has been designed to target Wilms' tumor-1 (WT-1), a highly specific antigen overexpressed in GBM among other solid tumors. This is done through the generation of an scFv specific targeting the complex of WT-1126-134/HLA-A*02:01 on the surface of GBM. On the NK cell side, the engager is designed to target the activating receptor NKp46. Incorporation of the cytokine IL-15 within the engager supports the maturation, persistence, and expansion of NK cells in vivo while favoring their proliferation and survival in the tumor microenvironment. Additionally, our data indicated that the chemokine CXCL10 plays an important role in the infiltration of NK cells into GBM, however, GBM tumors produce low levels of this chemokine. Incorporation of a CXCL10-producing function into our engager supports intratumoral NK cell trafficking by promoting, through their synthetic production, increased levels of CXCL10 locally in the tumor microenvironment.ResultsCollectively, this has resulted in a novel multifunctional NK cell engager, combining neoantigen-cytokine-chemokine elements fused to an activating domain-specific to NK cells, and we have investigated its ability to support and enhance NK cell-mediated cytotoxicity against solid tumors in vitro and in vivo against patient-derived GBM models. The multi-specific engager shows both high tumor specificity, as well as the ability to overcome NK cell dysfunction encountered in the GBM TME.ConclusionsWe hypothesize that taking advantage of our multi-functional engager, NK cells will exhibit superior ex vivo expansion, infiltration, and antitumor activity in the treatment of GBM and other solid tumors.

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.

scholarly journals Nonstochastic Coexpression of Activation Receptors on Murine Natural Killer Cells

2000 ◽  
Vol 191 (8) ◽  
pp. 1341-1354 ◽  
Author(s):  
Hamish R.C. Smith ◽  
Hubert H. Chuang ◽  
Lawrence L. Wang ◽  
Margarita Salcedo ◽  
Jonathan W. Heusel ◽  
...  
Keyword(s):  
Natural Killer Cells ◽  
Nk Cells ◽  
Natural Killer ◽  
Nk Cell ◽  
Inhibitory Receptors ◽  
Antigen Receptors ◽  
Killer Cells ◽  
Histocompatibility Complex ◽  
Nk Cell Cytotoxicity

Murine natural killer cells (NK) express lectin-like activation and inhibitory receptors, including the CD94/NKG2 family of receptors that bind Qa-1, and the Ly-49 family that recognizes major histocompatibility complex class I molecules. Here, we demonstrate that cross-linking of NK cells with a new specific anti–Ly-49H mAb induced NK cell cytotoxicity and cytokine production. Ly-49H is expressed on a subset of NK cells and can be coexpressed with Ly-49 inhibitory receptors. However, unlike Ly-49 inhibitory receptors, Ly-49H is not detectable on naive splenic CD3+ T cells, indicating that Ly-49H may be an NK cell–specific activation receptor. In further contrast to the stochastically expressed Ly-49 inhibitory receptors, Ly-49H is preferentially expressed with the Ly-49D activation receptor, and expression of both Ly-49H and Ly-49D is augmented on NK cells that lack receptors for Qa-1 tetramers. On developing splenic NK1.1+ cells, Ly-49D and Ly-49H are expressed later than the inhibitory receptors. These results directly demonstrate that Ly-49H activates primary NK cells, and suggest that expression of Ly-49 activation receptors by NK cells may be specifically regulated on NK cell subsets. The simultaneous expression of multiple activation receptors by individual NK cells contrasts with that of T cell antigen receptors and is relevant to the role of NK cells in innate immunity.

scholarly journals Fewer circulating natural killer cells 28 days after double cord blood transplantation predicts inferior survival and IL-15 response

2016 ◽  
Vol 1 (3) ◽  
pp. 208-218 ◽  
Author(s):  
Rachel J. Bergerson ◽  
Robin Williams ◽  
Hongbo Wang ◽  
Ryan Shanley ◽  
Gretchen Colbenson ◽  
...  
Keyword(s):  
Natural Killer Cells ◽  
Cord Blood ◽  
Nk Cells ◽  
Natural Killer ◽  
Nk Cell ◽  
Cord Blood Transplantation ◽  
Killer Cells ◽  
Cell Numbers ◽  
Blood Transplantation ◽  
Key Points

Key Points Low numbers of reconstituting NK cells at D+28 after dUCBT are associated with inferior DFS. Patients with low NK cell numbers at D+28 have reduced phosphorylation of STAT5 upon IL-15 stimulation and less Eomes expression.

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