Pharmacologic inhibition of lysine-specific demethylase 1 as a therapeutic and immune-sensitization strategy in pediatric high-grade glioma

Abstract Background Diffuse midline gliomas (DMG), including brainstem diffuse intrinsic pontine glioma (DIPG), are incurable pediatric high-grade gliomas (pHGG). Mutations in the H3 histone tail (H3.1/3.3-K27M) are a feature of DIPG, rendering them therapeutically sensitive to small-molecule inhibition of chromatin modifiers. Pharmacological inhibition of lysine-specific demethylase 1 (LSD1) is clinically relevant but has not been carefully investigated in pHGG or DIPG. Methods Patient-derived DIPG cell lines, orthotopic mouse models, and pHGG datasets were used to evaluate effects of LSD1 inhibitors on cytotoxicity and immune gene expression. Immune cell cytotoxicity was assessed in DIPG cells pretreated with LSD1 inhibitors, and informatics platforms were used to determine immune infiltration of pHGG. Results Selective cytotoxicity and an immunogenic gene signature were established in DIPG cell lines using clinically relevant LSD1 inhibitors. Pediatric HGG patient sequencing data demonstrated survival benefit of this LSD1-dependent gene signature. Pretreatment of DIPG with these inhibitors increased lysis by natural killer (NK) cells. Catalytic LSD1 inhibitors induced tumor regression and augmented NK cell infusion in vivo to reduce tumor burden. CIBERSORT analysis of patient data confirmed NK infiltration is beneficial to patient survival, while CD8 T cells are negatively prognostic. Catalytic LSD1 inhibitors are nonperturbing to NK cells, while scaffolding LSD1 inhibitors are toxic to NK cells and do not induce the gene signature in DIPG cells. Conclusions LSD1 inhibition using catalytic inhibitors is selectively cytotoxic and promotes an immune gene signature that increases NK cell killing in vitro and in vivo, representing a therapeutic opportunity for pHGG. Key Points 1. LSD1 inhibition using several clinically relevant compounds is selectively cytotoxic in DIPG and shows in vivo efficacy as a single agent. 2. An LSD1-controlled gene signature predicts survival in pHGG patients and is seen in neural tissue from LSD1 inhibitor–treated mice. 3. LSD1 inhibition enhances NK cell cytotoxicity against DIPG in vivo and in vitro with correlative genetic biomarkers.

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Pharmacologic inhibition of lysine specific demethylase-1 (LSD1) as a therapeutic and immune-sensitization strategy in diffuse intrinsic pontine glioma (DIPG)

10.1101/690966 ◽

2019 ◽

Author(s):

Cavan P. Bailey ◽

Megan M. Romero ◽

Oren J. Becher ◽

Michelle Monje ◽

Dean A. Lee ◽

...

Keyword(s):

Nk Cells ◽

Nk Cell ◽

Pediatric Brain Tumor ◽

High Grade Glioma ◽

Gene Signature ◽

Immune Gene ◽

High Grade ◽

Cell Cytotoxicity ◽

Selective Cytotoxicity ◽

Pediatric High Grade Glioma

AbstractBackgroundDiffuse intrinsic pontine glioma (DIPG) is an incurable pediatric brain tumor. Mutations in the H3 histone tail (H3.1/3.3-K27M) are a feature of DIPG, potentially rendering them therapeutically sensitive to small-molecule inhibition of chromatin modifiers. Pharmacological inhibition of lysine specific demethylase-1 (LSD1) shows promise in pediatric cancers such as Ewing’s sarcoma, but has not been investigated in DIPG, which was the aim of our study.MethodsPatient-derived DIPG cell lines and pediatric high-grade glioma (pHGG) datasets were used to evaluate effects of several LSD1 inhibitors on selective cytotoxicity and immune gene expression. Immune cell cytotoxicity was assessed in DIPG cells treated with LSD1 inhibitors and informatics platforms were used to determine immune infiltration of pHGG and impact on survival.ResultsSelective cytotoxicity and an immunogenic gene signature was established in DIPG lines using several clinically-relevant LSD1 inhibitors. Pediatric high-grade glioma patient sequencing data demonstrated survival benefit using this LSD1-dependent gene signature. On-target binding of catalytic LSD1 inhibitors was confirmed in DIPG and pre-treatment of DIPG with these inhibitors increased lysis by natural killer (NK) cells. CIBERSORT analysis of patient data confirmed NK infiltration is beneficial to patient survival while CD8 T-cells are negatively prognostic. Catalytic LSD1 inhibitors are non-perturbing to NK cells while scaffolding LSD1 inhibitors are toxic to NK cells and do not induce the gene signature in DIPG cells.ConclusionsLSD1 inhibition using catalytic inhibitors are both selectively cytotoxic and promote an immune gene signature that is associated with NK cell killing, representing a therapeutic opportunity for pHGG.Key pointsLSD1 inhibition using several clinically relevant compounds is selectively cytotoxic in DIPG.An LSD1-controlled gene signature predicts survival in pediatric high-grade glioma patients.LSD1 inhibition enhances NK cell cytotoxicity against DIPG with correlative genetic biomarkers.Importance of the studyThis is the first study to evaluate inhibition of LSD1 in a uniformly lethal type of pediatric brain tumor: DIPG. We demonstrate selective cytotoxicity of several clinically relevant compounds against patient derived DIPG cells, and identify an immune gene signature that is upregulated in DIPG cells by catalytic inhibitors of LSD1. This immune gene signature is predictive of prognosis in pHGG, consistent with the rationale of promoting this signature through LSD1 inhibition. NK cell killing of DIPG is enhanced by LSD1 inhibition, providing functional confirmation of this gene signature, and represents the first report of LSD1 inhibition promoting NK cell cytotoxicity of cancer cells. Given the poor prognosis of pHGGs and lack of effective treatments, our results suggest use of LSD1 inhibition as a single agent or in combination with NK cell therapy may be a safe and efficacious strategy.

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In Vitro and In Vivo Sensitization of Malignant Cells to Autologous Natural Killer Cell Cytotoxicity Following Exposure to Bortezomib.

Blood ◽

10.1182/blood.v108.11.925.925 ◽

2006 ◽

Vol 108 (11) ◽

pp. 925-925 ◽

Cited By ~ 1

Author(s):

Andreas Lundqvist ◽

Kristy Greeneltch ◽

Maria Berg ◽

Shivani Srivastava ◽

Nanae Harashima ◽

...

Keyword(s):

Tumor Growth ◽

Nk Cells ◽

Tumor Cells ◽

Nk Cell ◽

Cell Cytotoxicity ◽

Malignant Cells ◽

Nk Cell Cytotoxicity ◽

Tumor Death

Abstract Killer IgG like receptor (KIR) inactivation of NK cells by self HLA molecules has been proposed as a mechanism through which malignant cells evade host NK cell-mediated immunity. To overcome this limitation, we sought to develop a method to sensitize the patient’s tumor to autologous NK cell cytotoxicity. The proteasome inhibitor bortezomib has recently been shown to enhance the activity of tumor death receptors. We found that exposure of a variety of different leukemia, lymphoma and solid tumor cancer cell lines to sub-apoptotic doses of bortezomib sensitized tumor cells in vitro to lysis by allogeneic NK cells. Importantly, this sensitizing effect also occurs with autologous NK cells normally rendered inactive via tumor KIR ligands; NK cells expanded from patients with metastatic renal cell carcinoma were significantly more cytotoxic against the patient’s own autologous tumor cells when pretreated with bortezomib compared to untreated tumors. This sensitization to autologous NK cell killing was also observed in vivo in two different murine tumor models. A significant delay in tumor growth in C57BL/6 mice bearing LLC1 tumors (figure) and a delay in tumor growth and a significant prolongation (p<0.01) in survival were observed in RENCA tumor bearing Balb/c mice treated with bortezomib and syngeneic NK cell infusions compared to untreated mice or animals treated with bortezomib alone or NK cells alone. An investigation into the mechanism through which NK cell cytotoxicity was potentiated revealed bortezomib enhanced the activity of tumor death receptor-dependent and -independent apoptotic pathways. More specifically, bortezomib sensitized human and murine tumor cells to TRAIL and perforin/granzyme mediated NK cell cytotoxicity respectively. These observations suggest that pretreatment of malignant cells with bortezomib could be used as a strategy to override NK cell inhibition via tumor KIR ligands, thus potentiating the activity of adoptively infused autologous NK cells. A clinical trial evaluating the safety and anti-tumor efficacy of adoptively infused autologous NK cells in patients with advanced malignancies with and without tumor sensitization using bortezomib is currently being explored. Figure: Tumor growth in LLC1 bearing C57BL/6 mice. Fourteen days following s.c. injection of 3x105 LLC1 tumor cells, mice received 15μg (i.p) bortezomib and/or an adoptive infusion of 1x106 NK cells from C57BL/6 mice (i.v) given on day 15. Each dot represents the tumor volume of individual mice measured on day 28 post tumor injection. Tumors were significantly smaller in mice treated with bortezomib followed by NK cells compared to controls or mice that received either NK cells alone or bortezomib alone (p<0.04 for all groups). Figure:. Tumor growth in LLC1 bearing C57BL/6 mice. . / Fourteen days following s.c. injection of 3x105 LLC1 tumor cells, mice received 15μg (i.p) bortezomib and/or an adoptive infusion of 1x106 NK cells from C57BL/6 mice (i.v) given on day 15. Each dot represents the tumor volume of individual mice measured on day 28 post tumor injection. Tumors were significantly smaller in mice treated with bortezomib followed by NK cells compared to controls or mice that received either NK cells alone or bortezomib alone (p<0.04 for all groups).

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Impaired NK cell cytotoxicity by high level of interferon-γ in concanavalin A-induced hepatitis

Canadian Journal of Physiology and Pharmacology ◽

10.1139/y05-093 ◽

2005 ◽

Vol 83 (11) ◽

pp. 1045-1053 ◽

Cited By ~ 11

Author(s):

Zhongjun Dong ◽

Cai Zhang ◽

Haiming Wei ◽

Rui Sun ◽

Zhigang Tian

Keyword(s):

Nk Cells ◽

Cell Lines ◽

Nk Cell ◽

Interferon Γ ◽

Cell Cytotoxicity ◽

Wild Mice ◽

Ifn Γ ◽

Nk Cell Cytotoxicity

Unlike T cells, the role of natural killer (NK) cells is not well documented in the concanavalin (ConA)- induced hepatitis model. This study aimed to investigate the regulatory effect of high levels of interferon-γ (IFN-γ) on NK cells in ConA-induced hepatitis. The cytotoxicities of NK cells from ConA-injected mice or NK cell lines (NK92 and NKL) were detected by the 4-h 51Cr release assay. Depletion of NK cells with AsGM1 antibody was used to assess the NK cell role in ConA-induced hepatitis. Expression of NK cell receptors and cytotoxic molecules was measured by reverse transcription – polymerase chain reaction. Twelve hours after ConA injection, serum IFN-γ was significantly increased in wild mice, but not in severe combined immunodeficiency mice, and hepatic NK cells exerted impaired cytotoxicity against YAC-l cells in wild mice. Eight hours after NK cells were incubated in serum from ConA-treated mice, NK cell cytotoxicity was down-modulated and the effect was abolished by pretreatment with neutralizing serum IFN-γ with specific antibody in vitro. A high concentration of IFN-γ (> 1000 U/mL) inhibited the cytotoxicities of 2 NK cell lines in vitro, accompanied with down-regulation of NKG2D transcripts and up-regulation of NKG2A/B and KIR2DL transcripts. The inhibitive role of IFN-γ was not seen in NKG2D ligand negative cells. These results suggest that NK cell cytotoxicity was inhibited by high levels of IFN-γ in ConA-induced hepatitis, which may relate to the dispensable role of NK cells.Key words: cytotoxicity, hepatoimmunology, interferon-γ, liver injury.

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MicroRNA-20a mediates the cytotoxicity of natural killer cells in endometriosis via ERG/HLX/STAT4/perforin axis

10.21203/rs.2.17459/v1 ◽

2019 ◽

Author(s):

Li-Juan Chen ◽

Bin Hu ◽

Zhi-Qiang Han ◽

Jian Ni ◽

Yong-Ming Zhou ◽

...

Keyword(s):

Nk Cells ◽

Natural Killer ◽

Nude Mice ◽

Nk Cell ◽

Protective Role ◽

Cell Cytotoxicity ◽

Nk Cell Cytotoxicity

Abstract Background: Intriguingly, microRNA-20a (miR-20a) has been recently witnessed to be involved in the pathogenesis of endometriosis (EMS) but the molecular mechanism controlled by miR-20a is to be undefined. The present study is designed to probe into how miR-20a acts to regulate the cytotoxicity of natural killer (NK) cells. Methods: Most of all, consistent with the clinical determination in EMS patients, miR-20a was determined to be down-regulated in NK cells isolated from nude mice. miR-20a could specifically bind to ERG and negatively regulates its expression in NK cells. Additionally, shRNA-mediated silencing of ERG decreased the expression of HLX. HLX up-regulated STAT4 by inducing proteasome degradation and inhibited NK cell cytotoxicity. Results: Of great importance, forced expression of miR-20a consequently induced NK cell cytotoxicity in vitro by increasing perforin expression via enhancement of STAT4 that was caused by impairing the binding of ERG to HLX enhancer. The in vivo experiments further confirmed the promoting role of miR-20a in the cytotoxicity of NK cells isolated from EMS nude mice and subsequent protective role of miR-20a against EMS-induced endometrial injury. Conclusion: The aforementioned data suggest that miR-20a potentiates the cytotoxicity of NK via up-regulating perforin mediated by ERG/HLX/STAT4, highlighting potential novel mechanisms associated with EMS progression.

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Primary B-CLL Resistance to NK Cell Cytotoxicity can be Overcome In Vitro and In Vivo by Priming NK Cells and Monoclonal Antibody Therapy

Journal of Clinical Immunology ◽

10.1007/s10875-011-9624-5 ◽

2012 ◽

Vol 32 (3) ◽

pp. 632-646 ◽

Cited By ~ 27

Author(s):

Caroline Veuillen ◽

Thérèse Aurran-Schleinitz ◽

Rémy Castellano ◽

Jérôme Rey ◽

Françoise Mallet ◽

...

Keyword(s):

Monoclonal Antibody ◽

Nk Cells ◽

Nk Cell ◽

Antibody Therapy ◽

Monoclonal Antibody Therapy ◽

Cell Cytotoxicity ◽

Nk Cell Cytotoxicity

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NK cells promote cancer immunoediting through tumor-intrinsic loss of interferon-stimulated gene expression

10.1101/2020.11.23.394353 ◽

2020 ◽

Author(s):

Yung Yu Wong ◽

Luke Riggan ◽

Edgar Perez-Reyes ◽

Christopher Huerta ◽

Matt Moldenhauer ◽

...

Keyword(s):

Tumor Cell ◽

Nk Cells ◽

Tumor Cells ◽

Cell Lines ◽

Nk Cell ◽

Metastatic Cancer ◽

Cancer Immunoediting ◽

Ifn Γ

AbstractNatural killer (NK) cells are innate lymphocytes that constantly patrol host tissues against transformed cells in a process known as cancer immunosurveillance. Previous evidence in mice has demonstrated that NK cell-derived IFN-γ can promote immunoevasion by sculpting the immunogenicity of developing tumors in a process known as cancer immunoediting. This process involves the elimination of highly immunogenic “unedited” tumor cells followed by the eventual escape of less immunogenic “edited” tumor cell variants that are able to escape recognition or elimination by the immune system. Here, we show that NK cell-edited fibrosarcomas decrease the expression of 17 conserved IFN-γ-inducible genes compared to unedited tumor cells. High expression of 3 of these identified genes (Psmb8, Trim21, Parp12) in human tumor samples correlates with enhanced survival in breast cancer, melanoma, and sarcoma patients. While NK cell-edited fibrosarcomas displayed resistance to IFN-γ growth suppression in vitro, functional knockouts of individual interferon stimulated genes (ISGs) were not required for outgrowth of unedited tumor cell lines in vitro and in vivo compared to complete loss of IFN signaling. Furthermore, knockout of IFN-γ-intrinsic signaling via deletion of Ifngr in edited B16 F10 and E0771 LMB metastatic cancer cell lines did not impact host survival following lung metastasis. Together, these results suggest that unedited tumors can be selected for decreased IFN-γ signaling to evade immune responses in vivo, and as a consequence, tumor-extrinsic IFN signaling may be more important for potentiating durable anti-tumor responses to advanced solid tumors.

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NK cell activation in visceral leishmaniasis requires TLR9, myeloid DCs, and IL-12, but is independent of plasmacytoid DCs

Journal of Experimental Medicine ◽

10.1084/jem.20061293 ◽

2007 ◽

Vol 204 (4) ◽

pp. 893-906 ◽

Cited By ~ 124

Author(s):

Ulrike Schleicher ◽

Jan Liese ◽

Ilka Knippertz ◽

Claudia Kurzmann ◽

Andrea Hesse ◽

...

Keyword(s):

Visceral Leishmaniasis ◽

Nk Cells ◽

Cell Activation ◽

Nk Cell ◽

Dependent Manner ◽

Cell Cytotoxicity ◽

Ifn Γ ◽

Nk Cell Cytotoxicity

Natural killer (NK) cells are sentinel components of the innate response to pathogens, but the cell types, pathogen recognition receptors, and cytokines required for their activation in vivo are poorly defined. Here, we investigated the role of plasmacytoid dendritic cells (pDCs), myeloid DCs (mDCs), Toll-like receptors (TLRs), and of NK cell stimulatory cytokines for the induction of an NK cell response to the protozoan parasite Leishmania infantum. In vitro, pDCs did not endocytose Leishmania promastigotes but nevertheless released interferon (IFN)-α/β and interleukin (IL)-12 in a TLR9-dependent manner. mDCs rapidly internalized Leishmania and, in the presence of TLR9, produced IL-12, but not IFN-α/β. Depletion of pDCs did not impair the activation of NK cells in L. infantum–infected mice. In contrast, L. infantum–induced NK cell cytotoxicity and IFN-γ production were abolished in mDC-depleted mice. The same phenotype was observed in TLR9−/− mice, which lacked IL-12 expression by mDCs, and in IL-12−/− mice, whereas IFN-α/β receptor−/− mice showed only a minor reduction of NK cell IFN-γ expression. This study provides the first direct evidence that mDCs are essential for eliciting NK cell cytotoxicity and IFN-γ release in vivo and demonstrates that TLR9, mDCs, and IL-12 are functionally linked to the activation of NK cells in visceral leishmaniasis.

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Mechanism of rejection of virus persistently infected tumor cells by athymic nude mice.

Journal of Experimental Medicine ◽

10.1084/jem.149.5.1117 ◽

1979 ◽

Vol 149 (5) ◽

pp. 1117-1133 ◽

Cited By ~ 96

Author(s):

N Minato ◽

B R Bloom ◽

C Jones ◽

J Holland ◽

L M Reid

Keyword(s):

Cytotoxic Activity ◽

Nk Cells ◽

Cell Line ◽

Tumor Cells ◽

Cell Lines ◽

Nude Mice ◽

Nk Cell ◽

Persistently Infected

Cell lines known to be tumorigenic in the nude mouse were modified by rendering them persistently infected (P.I.) with a variety of RNA viruses, including measles, mumps, vesicular stomatitis virus, and influenza. Although as few as 100 HeLa or BHK cells produced tumors in 100% of nude mice, as many as 2 x 10(7) of the same cells P.I. with viruses failed to produce tumors. An active host response responsible for restricting the growth of the P.I. cells was suggested by the findings of marked mononuclear cell infiltrates at the inoculation sites and the inability of irradiated nude mice to reject them. An analysis of the in vitro cytotoxic activity of spleen cells from normal nude mice indicated that: (a) P.I. cell lines, but not uninfected cell lines, were susceptible to spontaneous cytotoxicity; (b) in vivo inoculation of P.I. lines induced an enhanced cytotoxic activity for P.I. targets in vitro, and this induction was not specific either for inducing virus or cell line; and (c) the effector cell had the characteristics for natural killer (NK) cells. Although the specificity of recognition of the various P.I. cell lines remains unclear, cold competition experiments indicated that blocking the killing of one P.I. cell line, e.g. HeLa-measles, could be achieved only by unlabeled homologous cells, i.e. HeLa-measles, and not by uninfected cells or other P.I. lines. A variant subline of BHK cells P.I. with VSV was selected for its ability to withstand the rejection process in nude mice. These cells formed metastatic and invasive tumors in nude mice. Although they were the most potent inducers in vivo of NK cell activity against various P.I. targets, they were the most resistant of the P.I. lines to NK cell cytotoxicity in vitro. In this system there was a good correlation between tumor rejection in vivo and susceptibility to NK cells in vitro. The present results suggest that NK cells may play a significant role in both rejection of tumor cells, and in resistance to viruses, particularly persistent infections.

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562 SO-C101 induces NK cell cytotoxicity and potentiates antibody-dependent cell cytotoxicity and anti-tumor activity

Journal for ImmunoTherapy of Cancer ◽

10.1136/jitc-2020-sitc2020.0562 ◽

2020 ◽

Vol 8 (Suppl 3) ◽

pp. A596-A596

Author(s):

Zuzana Antosova ◽

Nada Podzimkova ◽

Marketa Jiratova ◽

Eva Nedvedova ◽

Guy de Martynoff ◽

...

Keyword(s):

Monoclonal Antibodies ◽

Nk Cells ◽

Nk Cell ◽

Therapeutic Antibodies ◽

Cell Cytotoxicity ◽

Sequential Administration ◽

Dependent Cell ◽

Anti Tumor Activity

BackgroundSO-C101 is a superagonist fusion protein of interleukin (IL)-15 and the IL-15 receptor α (IL-15Rα) sushi+ domain, representing a promising clinical candidate for the treatment of cancer. SO-C101 specifically stimulates natural killer (NK) cells and memory CD8+ T cells with no significant expansion and activation of regulatory T cell compartment.MethodsHuman NK cell proliferation, the expression of NK cell receptors and ADCC activity of human PBMC after stimulation with SO-C101 in vitro in combination with monoclonal antibodies were detected by flow cytometry. The anti-tumor efficacy of SO-C101 in combination with Daratumumab was assessed in a multiple myeloma SCID xenograft mouse model in vivo.ResultsIn this study, we show that SO-C101 induced proliferation and expansion of both major subsets of human NK cells, CD56brightCD16- and CD56dimCD16+. Furthermore, SO-C101 induced expression of the cytotoxic receptors NKp30 and NKG2D whereas no upregulation of the inhibitory receptors CD158a, CD158b and NKG2A was detected. Both NK cell subsets activated by SO-C101 exhibited cytotoxicity towards cancer cells in vitro. Using human PBMCs, we show that SO-C101 potentiated killing of tumor cells induced by several clinically approved therapeutic monoclonal antibodies such as Cetuximab, Daratumumab and Obinutuzumab in vitro. SO-C101 and Daratumumab monotherapy treatment inhibited tumor growth in vivo, however, their combination showed the strongest anti-tumor efficacy. Specifically, sequential administration of Daratumumab, followed by SO-C101 promoted complete tumor regression, compared to partial anti-tumor responses induced by the respective monotherapies.ConclusionsSO-C101 augments the anti-tumor activity of therapeutic antibodies by increasing NK cells mediated antibody-dependent cell cytotoxicity. These results support the evaluation of SO-C101 in combination with monoclonal therapeutic antibodies in clinical studies.Ethics ApprovalThe anti-tumor efficacy studies in mice were approved by the internal ethics board of the respective contract research organization (CRO).

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Glucose Oligosaccharide and Long-Chain Glucomannan Feed Additives Induce Enhanced Activation of Intraepithelial NK Cells and Relative Abundance of Commensal Lactic Acid Bacteria in Broiler Chickens

Veterinary Sciences ◽

10.3390/vetsci8060110 ◽

2021 ◽

Vol 8 (6) ◽

pp. 110

Author(s):

Nathalie Meijerink ◽

Jean E. de Oliveira ◽

Daphne A. van Haarlem ◽

Guilherme Hosotani ◽

David M. Lamot ◽

...

Keyword(s):

Immune System ◽

Nk Cells ◽

Relative Abundance ◽

Nk Cell ◽

Feed Additives ◽

Microbiota Composition ◽

In Ovo ◽

Long Chain

Restrictions on the use of antibiotics in the poultry industry stimulate the development of alternative nutritional solutions to maintain or improve poultry health. This requires more insight in the modulatory effects of feed additives on the immune system and microbiota composition. Compounds known to influence the innate immune system and microbiota composition were selected and screened in vitro, in ovo, and in vivo. Among all compounds, 57 enhanced NK cell activation, 56 increased phagocytosis, and 22 increased NO production of the macrophage cell line HD11 in vitro. Based on these results, availability and regulatory status, six compounds were selected for further analysis. None of these compounds showed negative effects on growth, hatchability, and feed conversion in in ovo and in vivo studies. Based on the most interesting numerical results and highest future potential feasibility, two compounds were analyzed further. Administration of glucose oligosaccharide and long-chain glucomannan in vivo both enhanced activation of intraepithelial NK cells and led to increased relative abundance of lactic acid bacteria (LAB) amongst ileum and ceca microbiota after seven days of supplementation. Positive correlations between NK cell subsets and activation, and relative abundance of LAB suggest the involvement of microbiota in the modulation of the function of intraepithelial NK cells. This study identifies glucose oligosaccharide and long-chain glucomannan supplementation as effective nutritional strategies to modulate the intestinal microbiota composition and strengthen the intraepithelial innate immune system.

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