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A Neoglycoconjugate Containing the Human Milk Sugar LNFPIII Drives Anti-Inflammatory Activation of Antigen Presenting Cells in a CD14 Dependent Pathway.

ABSTRACT: The milk pentasaccharide LNFPIII has therapeutic action for metabolic and autoimmune diseases and prolongs transplant survival in mice when presented as a neoglycoconjugate. Within LNFPIII is the Lewisx trisaccharide, expressed by many helminth parasites. In humans, LNFPIII is found in human milk and also known as stage-specific embryonic antigen-1. LNFPIII-NGC drives alternative activation of macrophages and dendritic cells via NF?B activation in a TLR4 dependent mechanism. However, the connection between LNFPIII-NGC activation of APCs, TLR4 signaling and subsequent MAP kinase signaling leading to anti-inflammatory activation of APCs remains unknown. In this study we determined that the innate receptor CD14 was essential for LNFPIII-NGC induction of both ERK and NFkB activation in APCs. Induction of ERK activation by LNFPIII-NGC was completely dependent on CD14/TLR4-Ras-Raf1/TPL2-MEK axis in bone marrow derived dendritic cells (BMDCs). In addition, LNFPIII-NGC preferentially induced the production of Th2 "favoring" chemokines CCL22 and matrix metalloprotease protein-9 in a CD14 dependent manner in BMDCs. In contrast, LNFPIII-NGC induces significantly lower levels of Th1 "favoring" chemokines, MIP1?, MIP1? and MIP-2 compared to levels in LPS stimulated cells. Interestingly, NGC of the identical human milk sugar LNnT, minus the alpha 1-3 linked fucose, failed to activate APCs via TLR4/MD2/CD14 receptor complex, suggesting that the alpha 1-3 linked fucose in LNFPIII and not on LNnT, is required for this process. Using specific chemical inhibitors of the MAPK pathway, we found that LNFPIII-NGC induction of CCL22, MMP9 and IL-10 production was dependent on ERK activation. Over all, this study suggests that LNFPIII-NGC utilizes CD14/TLR4-MAPK (ERK) axis in modulating APC activation to produce anti-inflammatory chemokines and cytokines in a manner distinct from that seen for the pro-inflammatory PAMP LPS. These pathways may explain the in vivo therapeutic effect of LNFPIII-NGC treatment for inflammation based diseases.

SUBMITTER: Tundup S

PROVIDER: S-EPMC4560409 | biostudies-literature | 2015

REPOSITORIES: biostudies-literature

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A Neoglycoconjugate Containing the Human Milk Sugar LNFPIII Drives Anti-Inflammatory Activation of Antigen Presenting Cells in a CD14 Dependent Pathway.

Tundup Smanla S Srivastava Leena L Norberg Thomas T Watford Wendy W Harn Donald D

PloS one 20150904 9

The milk pentasaccharide LNFPIII has therapeutic action for metabolic and autoimmune diseases and prolongs transplant survival in mice when presented as a neoglycoconjugate. Within LNFPIII is the Lewisx trisaccharide, expressed by many helminth parasites. In humans, LNFPIII is found in human milk and also known as stage-specific embryonic antigen-1. LNFPIII-NGC drives alternative activation of macrophages and dendritic cells via NFκB activation in a TLR4 dependent mechanism. However, the connect ...[more]

PMID: 26340260

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Project description:BackgroundTNF receptor family agonists and checkpoint blockade combination therapies lead to minimal tumor clearance of poorly immunogenic tumors. Therefore, a need to enhance the efficacy of this combination therapy arises. Antigen-presenting cells (APCs) present antigen to T cells and steer the immune response through chemokine and cytokine secretion. DRibbles (DR) are tumor-derived autophagosomes containing tumor antigens and innate inflammatory adjuvants.MethodsUsing preclinical murine lung and pancreatic cancer models, we assessed the triple combination therapy of GITR agonist and PD-1 blocking antibodies with peritumoral injections of DRibbles-pulsed-bone marrow cells (BMCs), which consisted mainly of APCs, or CD103+ cross-presenting dendritic cells (DCs). Immune responses were assessed by flow cytometry. FTY720 was used to prevent T-cell egress from lymph nodes to assess lymph node involvement, and MHC-mismatched-BMCs were used to assess the necessity of antigen presentation by the peritumorally-injected DR-APCs.ResultsTritherapy increased survival and cures in tumor-bearing mice compared to combined antibody therapy or peritumoral DR-BMCs alone. Peritumorally-injected BMCs remained within the tumor for at least 14 days and tritherapy efficacy was dependent on both CD4+ and CD8+ T cells. Although the overall percent of tumor-infiltrating T cells remained similar, tritherapy increased the ratio of effector CD4+ T cells-to-regulatory T cells, CD4+ T-cell cytokine production and proliferation, and CD8+ T-cell cytolytic activity in the tumor. Despite tritherapy-induced T-cell activation and cytolytic activity in lymph nodes, this T-cell activation was not required for tumor regression and enhanced survival. Replacement of DR-BMCs with DR-pulsed-DCs in the tritherapy led to similar antitumor effects, whereas replacement with DRibbles was less effective but delayed tumor growth. Interestingly, peritumoral administration of DR-pulsed MHC-mismatched-APCs in the tritherapy led to similar antitumor effects as MHC-matched-APCs, indicating that the observed enhanced antitumor effect was mediated independently of antigen presentation by the administered APCs.ConclusionsOverall, these results demonstrate that peritumoral DR-pulsed-BMC/DC administration synergizes with GITR agonist and PD-1 blockade to locally modulate and sustain tumor effector T-cell responses independently of T cell priming and perhaps through innate inflammatory modulations mediated by the DRibbles adjuvant. We offer a unique approach to modify the tumor microenvironment to benefit T-cell-targeted immunotherapies.

Dataset Information

A Neoglycoconjugate Containing the Human Milk Sugar LNFPIII Drives Anti-Inflammatory Activation of Antigen Presenting Cells in a CD14 Dependent Pathway.

Publications

A Neoglycoconjugate Containing the Human Milk Sugar LNFPIII Drives Anti-Inflammatory Activation of Antigen Presenting Cells in a CD14 Dependent Pathway.

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