Lipid Autacoids Research Articles

Abstract 7458: Inhibition of brain cancer progression via resolution of inflammation

Abstract While inflammation is a hallmark of cancer, the mechanisms that disrupt its resolution in cancer are not well understood. A paradigm shift is emerging in our understanding that the resolution of inflammation is an active process controlled by specialized pro-resolving mediators (SPMs), including protectins and resolvins, which are endogenous lipid autacoids present in multiple tissues including the brain and cerebrospinal fluid. SPMs counter-regulate pro-inflammatory cytokine production and stimulate phagocytosis of cellular debris (“pro-resolution”) without being immunosuppressive. We hypothesized treatment with pro-resolving agonists such as SPMs presents a novel debris-clearing and resolution-enhancing therapeutic approach to complement cytotoxic brain cancer therapies. Here, we demonstrate the failure of resolution of inflammation via dysregulated SPMs may be a critical risk factor for aggressive brain cancer growth. Moreover, chemotherapy and targeted therapy further disrupt inflammation resolution via a tumor cell death (“debris”)-induced cytokine storm by macrophages. The SPM receptors GPR37 (protectin D1), GPR18/DRV2 (resolvin (Rv) D2), GPR32 (RvD1), and ChemR23/ERV (RvE1) are expressed in tumor-infiltrating macrophages and endothelial cells in the brain tumor microenvironment. Plasma levels of SPMs were significantly reduced in the medulloblastoma-bearing mice, suggesting that brain cancer progression is associated with a failure of endogenous resolution of inflammation. To determine whether failed resolution in brain cancer can be rescued by lipid mediator agonists, we systemically administered SPMs to subcutaneous and orthotopic glioblastoma and medulloblastoma murine brain tumor models. Significantly, RvD1 and RvD2 each inhibited the growth of human glioblastoma tumors by 15-fold at nanogram concentrations without toxicity. Likewise, RvD2 inhibited the growth of human medulloblastoma tumors by 3-fold. Protectins, PDX and PD1n-3, each inhibited growth of human medulloblastoma in mice by approximately 5-fold. Systemic administration of PDX or RvD2 prolonged survival of mice injected with orthotopic human D556 medulloblastoma and also glioblastoma models. Flow cytometry analysis of orthotopic glioblastoma tumors revealed increased immune infiltration into the brain tumor microenvironment in the PCTR1-treatment group when compared to the tumor-bearing non-treated group. At nanogram per day doses, protectins and resolvins suppressed orthotopic debris-stimulated brain tumor growth by activating macrophage and microglial phagocytosis of tumor cell debris and counter-regulating pro-inflammatory cytokines, including TNF-α, CCL2, CXCL1, and CCL4. Thus, stimulating the resolution of inflammation via the clearance of cellular debris with pro-resolving agonists represents a novel therapeutic approach to brain cancer, including glioblastoma and medulloblastoma. Citation Format: Jacqueline Capuano, Kimberly Vasquez, Katherine Quinlivan, Rachel Bayer, Sarina Virani, Michael Gillespie, Mark Kieran, Charles Serhan, Diane Bielenberg, Dipak Panigrahy. Inhibition of brain cancer progression via resolution of inflammation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 7458.

Abstract 6817: Control of breast cancer via pro-resolving mediators

Abstract Tumor recurrence is the principal cause of mortality in breast cancer, yet the underlying mechanisms are not well understood. Anti-estrogen therapy, including tamoxifen, reduces tumor burden by inducing apoptosis and necrosis of cancerous tissue. The resulting cellular debris may stimulate tumor growth by disrupting inflammation resolution and creating pro-inflammatory and pro-tumorigenic microenvironments. Compared to estrogen receptor (ER)-positive breast cancer, ER-negative behaves aggressively, and patients have a poor prognosis due to the lack of treatment options. Here, we demonstrate that breast tumor cells killed by cytotoxic anti-estrogen therapy or chemotherapy (“tumor cell debris”) stimulate primary tumor growth when co-injected with a subthreshold (nontumorigenic) inoculum of tumor cells by triggering a macrophage-derived pro-inflammatory and pro-angiogenic “cytokine storm.” Thus, tumor cell debris is a critical risk factor for aggressive breast cancer growth. To assess whether stimulating the clearance of debris would impact breast cancer progression in vivo, we utilized maresin 1 (MaR1), maresin conjugates in tissue regeneration (MCTR1, MCTR2) and protectin conjugates in tissue regeneration (PCTRs), which are specialized pro-resolving lipid autacoid mediators (SPMs). We treated established models of ER-positive (EO771) and negative (4T1) breast cancer. Each maresin sharply reduced tumor growth in debris-stimulated and spontaneous (e.g. MMTV-PyMT) breast cancer models at nanogram concentrations without toxicity. Notably, PCTR1 inhibited orthotopic triple-negative (4T1) growth and proliferation compared to chemotherapy (paclitaxel) or immunotherapy (anti-cytotoxic T-lymphocyte-associated protein 4, CTLA4). Similarly, PCTR1 alone or in combination with chemotherapy (paclitaxel) reduced gene expression and protein levels of pro-angiogenic factor CXCL12/SDF1 in the ER-negative (4T1) tumor microenvironment. In ER-positive (EO771), a triple combination of PCTR1, chemotherapy (paclitaxel), and immunotherapy (anti-CTLA4) treatment potentiated the immune checkpoint blockade. Maresins also stimulated macrophage phagocytosis of therapy-generated breast tumor cell debris, inhibited tumor angiogenesis, and dampened a therapy-induced cytokine storm, including TNF-α, MIP-2/CXCL2, CCL2/MCP-1, IL- 1ra/IL-1F3, and G-CSF. Expression of SPM receptors (e.g. resolvin D1 (RvD1) receptor ALX/FPR2, resolvin D2 (RvD2) receptor GPR18, maresin 1 (MaR1) receptor LGR6, and resolvin E1 (RvE1) receptor ChemR23) were specifically identified in breast cancer tissue. Taken together, the maresin and protectin pathway mediators may represent a new therapeutic approach to stimulate the resolution of inflammation in breast cancer. Citation Format: Sarina A. Virani, Michael Gillespie, Haixia Yang, Diane R. Bielenberg, Mark W. Kieran, Sui Huang, Charles N. Serhan, Dipak Panigrahy. Control of breast cancer via pro-resolving mediators [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6817.

The 5-lipoxygenase/cyclooxygenase-2 cross-over metabolite, hemiketal E2, enhances VEGFR2 activation and promotes angiogenesis

Consecutive oxygenation of arachidonic acid by 5-lipoxygenase and cyclooxygenase-2 yields the hemiketal eicosanoids, HKE2 and HKD2. Hemiketals stimulate angiogenesis by inducing endothelial cell tubulogenesis in culture; however, how this process is regulated has not been determined. Here, we identify vascular endothelial growth factor receptor 2 (VEGFR2) as a mediator of HKE2-induced angiogenesis invitro and invivo. We found that HKE2 treatment of human umbilical vein endothelial cells dose-dependently increased the phosphorylation of VEGFR2 and the downstream kinases ERK and Akt that mediated endothelial cell tubulogenesis. Invivo, HKE2 induced the growth of blood vessels into polyacetal sponges implanted in mice. HKE2-mediated effects invitro and invivo were blocked by the VEGFR2 inhibitor vatalanib, indicating that the pro-angiogenic effect of HKE2 was mediated by VEGFR2. HKE2 covalently bound and inhibited PTP1B, a protein tyrosine phosphatase that dephosphorylates VEGFR2, thereby providing a possible molecular mechanism for how HKE2 induced pro-angiogenic signaling. In summary, our studies indicate that biosynthetic cross-over of the 5-lipoxygenase and cyclooxygenase-2 pathways gives rise to a potent lipid autacoid that regulates endothelial cell function invitro and invivo. These findings suggest that common drugs targeting the arachidonic acid pathway could prove useful in antiangiogenic therapy.

Essential lipid autacoids rewire mitochondrial energy efficiency in metabolic dysfunction-associated fatty liver disease.

Injury to hepatocyte mitochondria is common in metabolic dysfunction-associated fatty liver disease. Here, we investigated whether changes in the content of essential fatty acid-derived lipid autacoids affect hepatocyte mitochondrial bioenergetics and metabolic efficiency. The study was performed in transgenic mice for the fat-1 gene, which allows the endogenous replacement of the membrane omega-6-polyunsaturated fatty acid (PUFA) composition by omega-3-PUFA. Transmission electron microscopy revealed that hepatocyte mitochondria of fat-1 mice had more abundant intact cristae and higher mitochondrial aspect ratio. Fat-1 mice had increased expression of oxidative phosphorylation complexes I and II and translocases of both inner (translocase of inner mitochondrial membrane 44) and outer (translocase of the outer membrane 20) mitochondrial membranes. Fat-1 mice also showed increased mitofusin-2 and reduced dynamin-like protein 1 phosphorylation, which mediate mitochondrial fusion and fission, respectively. Mitochondria of fat-1 mice exhibited enhanced oxygen consumption rate, fatty acid β-oxidation, and energy substrate utilization as determined by high-resolution respirometry, [1- 14 C]-oleate oxidation and nicotinamide adenine dinucleotide hydride/dihydroflavine-adenine dinucleotide production, respectively. Untargeted lipidomics identified a rich hepatic omega-3-PUFA composition and a specific docosahexaenoic acid (DHA)-enriched lipid fingerprint in fat-1 mice. Targeted lipidomics uncovered a higher content of DHA-derived lipid autacoids, namely resolvin D1 and maresin 1, which rescued hepatocytes from TNFα-induced mitochondrial dysfunction, and unblocked the tricarboxylic acid cycle flux and metabolic utilization of long-chain acyl-carnitines, amino acids, and carbohydrates. Importantly, fat-1 mice were protected against mitochondrial injury induced by obesogenic and fibrogenic insults. Our data uncover the importance of a lipid membrane composition rich in DHA and its lipid autacoid derivatives to have optimal hepatic mitochondrial and metabolic efficiency.

COVID-19 and cancer: start the resolution!

Coronavirus disease 2019 (COVID-19) due to infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been an ongoing pandemic causing significant morbidity and mortality worldwide. The “cytokine storm” is a critical driving force in severe COVID-19 cases, leading to hyperinflammation, multi-system organ failure, and death. A paradigm shift is emerging in our understanding of the resolution of inflammation from a passive course to an active biochemical process driven by endogenous specialized pro-resolving mediators (SPMs), such as resolvins, protectins, lipoxins, and maresins. SPMs stimulate macrophage-mediated debris clearance and counter pro-inflammatory cytokine production, a process collectively termed as the “resolution of inflammation.” Hyperinflammation is not unique to COVID-19 and also occurs in neoplastic conditions, putting individuals with underlying health conditions such as cancer at elevated risk of severe SARS-CoV-2 infection. Despite approaches to block systemic inflammation, there are no current therapies designed to stimulate the resolution of inflammation in patients with COVID-19 or cancer. A non-immunosuppressive therapeutic approach that reduces the cytokine storm in patients with COVID-19 and cancer is urgently needed. SPMs are potent immunoresolvent and organ-protective lipid autacoids that stimulate the resolution of inflammation, facilitate clearance of infections, reduce thrombus burden, and promote a return to tissue homeostasis. Targeting endogenous lipid mediators, such as SPMs, offers an entirely novel approach to control SARS-CoV-2 infection and cancer by increasing the body’s natural reserve of pro-resolving mediators without overt toxicity or immunosuppression.

Resolvin D1 reduces cancer growth stimulating a protective neutrophil-dependent recruitment of anti-tumor monocytes

BackgroundInnovative therapies to target tumor-associated neutrophils (PMN) are of clinical interest, since these cells are centrally involved in cancer inflammation and tumor progression. Resolvin D1 (RvD1) is a lipid autacoid that promotes resolution of inflammation by regulating the activity of distinct immune and non-immune cells. Here, using human papilloma virus (HPV) tumorigenesis as a model, we investigated whether RvD1 modulates PMN to reduce tumor progression.MethodsGrowth-curve assays with multiple cell lines and in vivo grafting of two distinct HPV-positive cells in syngeneic mice were used to determine if RvD1 reduced cancer growth. To investigate if and how RvD1 modulates PMN activities, RNA sequencing and multiplex cytokine ELISA of human PMN in co-culture with HPV-positive cells, coupled with pharmacological depletion of PMN in vivo, were performed. The mouse intratumoral immune cell composition was evaluated through FACS analysis. Growth-curve assays and in vivo pharmacological depletion were used to evaluate anti-tumor activities of human and mouse monocytes, respectively. Bioinformatic analysis of The Cancer Genome Atlas (TCGA) database was exploited to validate experimental findings in patients.ResultsRvD1 decreased in vitro and in vivo proliferation of human and mouse HPV-positive cancer cells through stimulation of PMN anti-tumor activities. In addition, RvD1 stimulated a PMN-dependent recruitment of classical monocytes as key determinant to reduce tumor growth in vivo. In human in vitro systems, exposure of PMN to RvD1 increased the production of the monocyte chemoattractant protein-1 (MCP-1), and enhanced transmigration of classical monocytes, with potent anti-tumor actions, toward HPV-positive cancer cells. Consistently, mining of immune cells infiltration levels in cervical cancer patients from the TCGA database evidenced an enhanced immune reaction and better clinical outcomes in patients with higher intratumoral monocytes as compared to patients with higher PMN infiltration.ConclusionsRvD1 reduces cancer growth by activating PMN anti-cancer activities and encouraging a protective PMN-dependent recruitment of anti-tumor monocytes. These findings demonstrate efficacy of RvD1 as an innovative therapeutic able to stimulate PMN reprogramming to an anti-cancer phenotype that restrains tumor growth.

Roles of Specialized Pro-Resolving Lipid Mediators in Autophagy and Inflammation.

Autophagy is a catabolic pathway that accounts for degradation and recycling of cellular components to extend cell survival under stress conditions. In addition to this prominent role, recent evidence indicates that autophagy is crucially involved in the regulation of the inflammatory response, a tightly controlled process aimed at clearing the inflammatory stimulus and restoring tissue homeostasis. To be efficient and beneficial to the host, inflammation should be controlled by a resolution program, since uncontrolled inflammation is the underlying cause of many pathologies. Resolution of inflammation is an active process mediated by a variety of mediators, including the so-called specialized pro-resolving lipid mediators (SPMs), a family of endogenous lipid autacoids known to regulate leukocyte infiltration and activities, and counterbalance cytokine production. Recently, regulation of autophagic mechanisms by these mediators has emerged, uncovering unappreciated connections between inflammation resolution and autophagy. Here, we summarize mechanisms of autophagy and resolution, focusing on the contribution of autophagy in sustaining paradigmatic examples of chronic inflammatory disorders. Then, we discuss the evidence that SPMs can restore dysregulated autophagy, hypothesizing that resolution of inflammation could represent an innovative approach to modulate autophagy and its impact on the inflammatory response.

Inflammation resolution: a dual-pronged approach to averting cytokine storms in COVID-19?

Severe coronavirus disease (COVID-19) is characterized by pulmonary hyper-inflammation and potentially life-threatening “cytokine storms”. Controlling the local and systemic inflammatory response in COVID-19 may be as important as anti-viral therapies. Endogenous lipid autacoid mediators, referred to as eicosanoids, play a critical role in the induction of inflammation and pro-inflammatory cytokine production. SARS-CoV-2 may trigger a cell death (“debris”)-induced “eicosanoid storm”, including prostaglandins and leukotrienes, which in turn initiates a robust inflammatory response. A paradigm shift is emerging in our understanding of the resolution of inflammation as an active biochemical process with the discovery of novel endogenous specialized pro-resolving lipid autacoid mediators (SPMs), such as resolvins. Resolvins and other SPMs stimulate macrophage-mediated clearance of debris and counter pro-inflammatory cytokine production, a process called inflammation resolution. SPMs and their lipid precursors exhibit anti-viral activity at nanogram doses in the setting of influenza without being immunosuppressive. SPMs also promote anti-viral B cell antibodies and lymphocyte activity, highlighting their potential use in the treatment of COVID-19. Soluble epoxide hydrolase (sEH) inhibitors stabilize arachidonic acid-derived epoxyeicosatrienoic acids (EETs), which also stimulate inflammation resolution by promoting the production of pro-resolution mediators, activating anti-inflammatory processes, and preventing the cytokine storm. Both resolvins and EETs also attenuate pathological thrombosis and promote clot removal, which is emerging as a key pathology of COVID-19 infection. Thus, both SPMs and sEH inhibitors may promote the resolution of inflammation in COVID-19, thereby reducing acute respiratory distress syndrome (ARDS) and other life-threatening complications associated with robust viral-induced inflammation. While most COVID-19 clinical trials focus on “anti-viral” and “anti-inflammatory” strategies, stimulating inflammation resolution is a novel host-centric therapeutic avenue. Importantly, SPMs and sEH inhibitors are currently in clinical trials for other inflammatory diseases and could be rapidly translated for the management of COVID-19 via debris clearance and inflammatory cytokine suppression. Here, we discuss using pro-resolution mediators as a potential complement to current anti-viral strategies for COVID-19.

Pro‐Resolving Lipid Mediators and Anti‐Angiogenic Therapy Exhibit Synergistic Anti‐Tumor Activity via Resolvin Receptor Activation

Inflammation and angiogenesis are interdependent hallmarks of cancer. Tumor growth is angiogenesis‐dependent and inflammation is a risk factor for many cancers. Inflammation is regulated by endogenous specialized pro‐resolving lipid‐autacoid mediators (SPMs), including resolvins, protectins, and maresins, which are present in multiple tissues. Unlike the majority of anti‐inflammatory agents, SPMs are non‐immunosuppressive, and non‐toxic. These lipid autacoids inhibit angiogenesis and clear cellular debris by macrophages resulting in reduced localized inflammatory cytokines. SPMs mediate their pro‐resolution and anti‐inflammatory activity through at least five known human G‐coupled protein receptors (GPR32, GPR18, ChemR23, GPR37, and LGR6) for resolvin (Rv) D1, RvD2, RvE1, protectin D1, and maresin 1, respectively, as well as a murine RvD1 receptor ALX/FPR2. We show that stimulating resolution of inflammation (RvD4 or RvD5) and anti‐angiogenic therapy (e.g. the thrombospondin (TSP)‐1 peptide 3TSR or anti‐VEGF via DC101) induced synergistic antitumor activity in human xenograft models including ovarian cancer (e.g. 36M2) compared with either treatment alone. The triple therapy 3TSR, anti‐VEGF, and resolvins displayed additive anti‐tumor activity. As the role of SPM receptors in cancer is unknown, we screened various stromal cells in the tumor microenvironment (e.g. macrophages, pericytes, fibroblasts, and neutrophils) for expression of SPM receptors. SPM receptors were expressed in various clinical specimens including breast cancer, head and neck cancer (e.g. oral squamous cell carcinoma), and brain cancer. Flow cytometry and double immunohistochemistry staining confirmed that SPM receptors were expressed in non‐tumor cells in the tumor microenvironment (e.g. macrophages, fibroblasts, pericytes and blood vessels) in vivo. Accordingly, SPMs inhibited tumor cell proliferation in vivo. Thus, SPMs exhibit anti‐tumor activity via the tumor stroma. Loss of SPM receptor expression may be associated with cancer progression in clinical cancer specimens. Notably, resolvins (RvD4 or RvD5) inhibited tumor growth at doses 10,000 times lower than anti‐inflammatory agents such as aspirin and NSAIDs. Thus, stimulating the expression or activity of SPM receptors may enhance current anti‐angiogenic therapy for various cancers.

Abstract 503: Prostaglandin E2 produced by tumor cells or by the host tumor microenvironment is not completely abolished by aspirin or celecoxib and limits the ability of the host immune system to control tumor growth

Abstract Prostaglandins are lipid autacoids derived from arachidonic acid (AA). AA released from the cell membrane is metabolized by the sequential actions of prostaglandin G/H synthase or cyclooxygenase (COX) and respective synthases, such as mPGES-1, which forms the most abundant prostaglandin, PGE2. Prostaglandin biosynthesis is blocked by nonsteroidal anti-inflammatory drugs (NSAIDs), which inhibit the activity of either one or both cyclooxygenases. Besides their role in homeostasis, prostanoids can modulate immune responses and contribute to chronic inflammation associated with tumor initiation, vascularization, and growth. The prostanoid pathway is activated in many solid tumors, including breast cancer, and high levels of PGE2 have been associated with aggressive tumors and metastatic spread. The objective of this work was to define the pathways and cellular compartments that contribute to tumor PGE2 levels and their role in tumor growth and immune evasion. The anti-tumor impact of classical NSAIDs combined with immune checkpoint antibodies PD1 or CTLA4 (ICPAb) was also evaluated. 4T1 or CT26 murine tumors were grown in immune competent mice to confirm the presence of PGE2 in tumors. Mice were dosed with NSAIDs Aspirin or Celecoxib monotherapy, given at maximal tolerated dose, or combined with ICPAb. While ICPAb reduced tumor growth, nor Aspirin neither celecoxib was effective in monotherapy or combination with ICPAb. Interestingly, PGE2 concentration in tumors was reduced by NSAIDs but not abolished. To examine the production of PGE2 by tumors or host, we have used CRISPR/cas9 mutagenesis to generate tumor lines lacking various enzymes required for PGE2 production. Those lines were combined with mice carrying null mPGES1 alleles. We show that, while the host tumor microenvironment contributes to overall PGE2 levels in the tumor, large amounts of PGE2 are produced by the tumor epithelial cells. Despite high expression of Ptgs1 in these tumor lines, PGE2 production was dependent on COX-2 expression. Similarly, while 4T1 tumors express all three PGE2 synthases at relatively high levels, we found that mPGES1 was solely responsible for all PGE2 released from epithelial cells. Examining several independent Ptgs1/2 null 4T1 tumor lines, we show that loss of Ptgs1/2 results in decreased tumor growth in immune competent mice but not in an allogenic Rag1/2-/- line. Knowledge of tumor PGE2 metabolic and functional pathways is a critical first step in designing the most appropriate therapeutic interventions. Classical NSAIDs given as monotherapy or combined in mice with ICPAb are ineffective against tumors, possibly because they do not abolish PGE2 production, suggesting that better drugs against PGE2 pathway would be required to counter the immune suppression induced by PGE2. Citation Format: John N. Snouwaert, Leigh Jania, MyTrang Nguyen, Pragnya Dontu, Jérôme Besse, Barbara Akla, Pierre J. Ferré, Beverly H. Koller. Prostaglandin E2 produced by tumor cells or by the host tumor microenvironment is not completely abolished by aspirin or celecoxib and limits the ability of the host immune system to control tumor growth [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 503.

Brain Cancer: Failure of Resolution of Inflammation?

Medulloblastoma therapy, including radiation and chemotherapy, induces cell death resulting in tumor cell debris assumed to be inert or inhibitory to tumor growth. However, tumor cell death (debris) produced by cytotoxic therapy can accelerate tumor progression through the inflammatory response, suggesting that every attempt to kill tumor cells is a double‐edged sword. Inflammation is regulated by endogenous specialized pro‐resolving lipid mediators (SPMs), such as resolvins and protectins, which are present in the brain and CSF. These lipid autacoids stimulate the clearance of cellular debris by macrophages, resulting in reduced localized pro‐inflammatory cytokines. A loss of SPMs is associated with several neurodegenerative diseases but has not been studied in brain cancer. We hypothesize that reduced levels of SPMs correlate with brain cancer progression and that control of tumor cell debris‐mediated inflammation through resolvins and protectins represents a novel approach to brain cancer treatment. Here, we show that SPM receptors are expressed by human brain tumors including glioblastoma and medulloblastoma. Medulloblastoma tumor cell debris generated by front‐line chemotherapy or targeted therapy stimulates primary tumor growth when co‐injected with a subthreshold (non‐tumorigenic) inoculum of tumor cells by triggering the release of pro‐inflammatory cytokines. Stimulation of the resolution of inflammation via protectins and resolvins suppressed orthotopic medulloblastoma. Protectins and resolvins down‐regulated proinflammatory cytokine release in debris‐activated microglia and stimulate the clearance of tumor debris by inducing macrophage phagocytosis of apoptotic tumor cells. Histological analysis of SPM‐treated medulloblastoma and glioblastoma tumors revealed suppressed Ki‐67‐stained tumor cell proliferation. Thus, brain tumor progression results from a loss of resolution of tumor debris‐associated inflammation. Enhancing endogenous clearance of tumor cell debris via protectins and resolvins is a novel biological target to complement current cytotoxic brain cancer therapy and addresses the intrinsic limitation of current cancer therapy involving cell death. Our studies provide a new strategy for the prevention and treatment of therapy‐induced resistance in medulloblastoma and other brain cancers.Support or Funding InformationThis work was supported by the National Cancer Institute grant RO1 01CA170549‐02 (D.P.); ROCA148633‐01A4 (D.P.); and the Credit Unions Kids at Heart Team (D.P.)This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Synergy between Resolvins and Immune Checkpoint Blockade in a Novel Transplantable FANCC−/− Murine Head and Neck Tumor Model

Fanconi anemia (FA) patients have increased incidence of malignancies including aggressive head and neck cancer. Traditional cancer therapies, including chemotherapy and radiation, in head and neck cancer patients with mutations in the FA pathway (e.g. FANCC−/−) have failed and the development of novel therapeutics has been hindered by a lack of relevant pre‐clinical models. While FA knockout mice recapitulate tumorigenic properties and the stromal microenvironment, these genetic models exhibit extreme variability in time to develop invasive disease (e.g. 80–400 days), presenting a tremendous challenge to evaluate novel FA cancer therapies. Transplantable tumor systems are optimal to assess the anti‐tumor activity of new therapeutic approaches, however, few to no such models for FA have been characterized to date. Controlling inflammation is now recognized as a critical component of successful cancer treatment. While FA patients exhibit excessive inflammatory cytokines, a new direction has emerged with the discovery of endogenous omega‐3 fatty acid‐derived lipid mediators, such as resolvins, which have potent pro‐resolving activity without being immunosuppressive. These lipid autacoids act through promoting the clearance of cellular debris by macrophage phagocytosis, resulting in reduced localized pro‐inflammatory cytokines. As immunotherapy response can be predicted by DNA repair deficiencies that occur in FA, we hypothesize that stimulation of resolution of inflammation and checkpoint blockade would inhibit FANCC−/− tumor growth. Utilizing FANCC−/− and wild type (WT) murine head and neck squamous cell carcinoma cells, we have developed a robust transplantable tumor model in immunocompetent mice. FANCC−/− tumors exhibited accelerated growth and histologic findings demonstrated numerous FA tumor cells with dysplastic nuclei and Ki‐67‐positive proliferating cells compared to WT tumors. FANCC−/− tumors also expressed the immune checkpoint proteins PD1, PD‐L1, and CTLA4. Immune checkpoint blockade (e.g. anti‐PD1) and resolvins alone or in combination suppressed FANCC−/− subcutaneous and orthotopic (e.g. tongue) tumor growth and prolonged survival. Therefore, we have established a transplantable FANCC−/− murine head and neck tumor model whereby various novel therapies can be efficiently evaluated prior to clinical translation. The synergy between resolvins and immune checkpoint inhibitors to suppress FANCC−/− head and neck tumor growth provides a rationale for targeting immunotherapy and the resolution of inflammation for the treatment of FA cancers.Support or Funding InformationNational Cancer Institute grant RO1 01CA170549‐02; ROCA148633‐01A4; Credit Unions Kids at Heart Team.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Resolvins suppress tumor growth and enhance cancer therapy.

Cancer therapy reduces tumor burden by killing tumor cells, yet it simultaneously creates tumor cell debris that may stimulate inflammation and tumor growth. Thus, conventional cancer therapy is inherently a double-edged sword. In this study, we show that tumor cells killed by chemotherapy or targeted therapy ("tumor cell debris") stimulate primary tumor growth when coinjected with a subthreshold (nontumorigenic) inoculum of tumor cells by triggering macrophage proinflammatory cytokine release after phosphatidylserine exposure. Debris-stimulated tumors were inhibited by antiinflammatory and proresolving lipid autacoids, namely resolvin D1 (RvD1), RvD2, or RvE1. These mediators specifically inhibit debris-stimulated cancer progression by enhancing clearance of debris via macrophage phagocytosis in multiple tumor types. Resolvins counterregulate the release of cytokines/chemokines, including TNFα, IL-6, IL-8, CCL4, and CCL5, by human macrophages stimulated with cell debris. These results demonstrate that enhancing endogenous clearance of tumor cell debris is a new therapeutic target that may complement cytotoxic cancer therapies.

Fundamentals of and Critical Issues in Lipid Autacoid Medicine: A Review.

The identification of a number of families of lipid signal molecules since the 1990s created new therapeutic possibilities for a great number of disorders characterized by chronic inflammation and pain. These lipid autacoids have been explored in a great variety of animal models related to inflammation, pain, (neuro-)protection, and repair. Based on the data from these models, as well as on a number of proof of principle studies in the clinic in indications such as neuropathic pain, a new chapter in medicine is about to begin. We would like to introduce the term “Autacoid Pain Medicine” for this chapter. There are, however, a number of methodological and strategic issues to overcome in this field. One of the roadblocks is related to patent strategies around families of these molecules. As this is not always recognized we will present a number of examples.

Resolvins and aliamides: lipid autacoids in ophthalmology - what promise do they hold?

Resolvins are a novel class of lipid-derived endogenous molecules (autacoids) with potent immunomodulating properties, which regulate the resolution phase of an active immune response. These modulating factors are locally produced, influencing the function of cells and/or tissues, which are produced on demand and subsequently metabolized in the same cells and/or tissues. This review is focused on certain lipid autacoids with putative relevance for ophthalmology in general and for dry eye more specifically. We also briefly investigate the concept of aliamides and the role of palmitoylethanolamide in ophthalmology, and analyze in more detail the putative role and the preclinical and clinical development of resolvins as emerging treatments for dry eye and related disorders, with a focus on one of the lead resolvin derivatives – RX-10045.

Abstract 5133: Targeting the eicosanoid 20-HETE suppresses pancreatic cancer growth and metastasis through regulation of inflammation

Abstract Pancreatic cancer incidence and death rates are on the rise. Most available therapies fail partly due to the uniquely inflammatory pancreatic tumor stromal compartment. Prolonged and unresolved inflammation is implicated in the pathogenesis of pancreatic cancer, yet the critical inflammatory mediators remain largely unknown. The role of lipid autacoids in pancreatic cancer, which are metabolites of arachidonic acid and other related fatty acids, has not been thoroughly studied. Among these, metabolites of the cytochrome P450 (CYP) epoxygenases and ω-hydroxylases, including EETs and 20-HETE, respectively, have been mostly neglected in cancer. We previously showed that epoxyeicosatrienoic acids (EETs) stimulate multi-organ metastasis and tumor dormancy escape. Here we show that elevated levels of 20-hydroxyeicosatetraenoic acid (20-HETE), a product of ω-hydroxylases of the CYP4A/F subfamilies, stimulated pancreatic primary tumor growth and metastasis. We measured 20-HETE and other lipid autacoids via LC/MS/MS in multiple cell lines and found that 20-HETE was produced primarily by pancreatic cancer cells, which express the 20-HETE generating enzyme, CYP4F2. Overexpression of CYP4F2 and exogenous 20-HETE induced Ras activity in HEK293 cells or pancreatic cancer and macrophage cell lines, respectively. 20-HETE attracted human macrophages in vitro, while macrophage migration was reduced towards pancreatic cancer cells treated with an inhibitor of 20-HETE synthesis, HET0016. Inhibition of 20-HETE synthesis in human macrophages and pancreatic cancer cells decreased HB-EGF and M-CSF production. Systemic administration of 20-HETE promoted primary human and murine pancreatic tumor growth in subcutaneous and orthotopic murine models. Transgenic mice overexpressing 20-HETE (Tie2-CYP4F2 Tr) exhibited spontaneous liver and mediastinal lymph node metastasis following resection of subcutaneous primary pancreatic tumors, in contrast to wild type control mice. Targeting 20-HETE by HET0016 or siRNA towards CYP4F2 suppressed primary pancreatic tumor growth. Systemic treatment of orthotopic pancreatic tumors with HET0016, 14,15-EEZE, and gemcitabine regressed established pancreatic tumors and prolonged survival without overt toxicity. Thus, 20-HETE critically regulates Ras activation, inflammation, and pancreatic tumor growth and metastasis. Our studies offer a mechanistic rationale for pharmacologically targeting 20-HETE as a novel approach to treat pancreatic cancer. Citation Format: Yael Gus-Brautbar, Julia Piwowarski, Jessica A. Casper, Diane R. Bielenberg, Jennifer Cheng, Birgitta Schmidt, Bruce D. Hammock, Darryl C. Zeldin, Mark W. Kieran, Dipak Panigrahy. Targeting the eicosanoid 20-HETE suppresses pancreatic cancer growth and metastasis through regulation of inflammation. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 5133.

Abstract 766: Control of lung cancer with aspirin-triggered stimulation of resolution

Abstract Objective: To establish aspirin-triggered resolvins as a novel treatment to enhance current cancer therapies through the resolution of inflammation and clearance of cancer therapy-induced tumor cell debris. Background: Inflammation in the tumor microenvironment is now recognized as a strong promoter of tumor growth. Substantial epidemiological evidence suggests that aspirin, which suppresses inflammation, also reduces the risk of cancer. However, the mechanism by which aspirin reduces cancer risk and improves survival remains unclear, and toxicity has limited its clinical use. Low-dose aspirin stimulates the endogenous production of aspirin-triggered (AT) resolvins, novel omega-3 fatty acid-derived pro-resolving lipid autacoid mediators. We recently discovered that resolvins have potent anti-tumor activity, making resolvins potential candidates to mediate the anti-cancer activity of aspirin. Thus, we hypothesize that aspirin's anti-cancer activity is mediated in part by AT-resolvins via novel anti-inflammatory and pro-resolution mechanisms. Methods: Flow cytometry confirmed apoptotic and necrotic lung tumor cell debris following chemotherapy (etoposide) or targeted therapy (erlotinib) treatment in vitro. Macrophage phagocytosis was quantified via Relative Fluorescent Units (RFU). Cytokine levels were quantified via ELISA. Results: AT-resolvins counter therapy-induced debris-stimulated tumor growth and spontaneous lung metastases via enhancing endogenous clearance of tumor debris via macrophages and counter-regulating macrophage cytokine production. Pharmacological and genetic antagonism of the AT-resolvin D1 receptor (ALX/FPR2) neutralized aspirin's anti-cancer activity. Conclusions: AT-resolvins may provide a novel pathway and molecular mechanism to explain how aspirin reduces cancer risk so broadly. These anti-inflammatory and pro-resolving lipid autacoid mediators may complement current therapies for lung cancer with minimal toxicity. Citation Format: Molly M. Gilligan, Megan L. Sulciner, Yael Gus-Brautbar, Sesquile Ramon, Romain A. Colas, Sui Huang, Charles N. Serhan, Dipak Panigrahy. Control of lung cancer with aspirin-triggered stimulation of resolution. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 766.

Dietary linoleic acid-induced alterations in pro- and anti-nociceptive lipid autacoids: Implications for idiopathic pain syndromes?

BackgroundChronic idiopathic pain syndromes are major causes of personal suffering, disability, and societal expense. Dietary n-6 linoleic acid has increased markedly in modern industrialized populations over the past century. These high amounts of linoleic acid could hypothetically predispose to physical pain by increasing the production of pro-nociceptive linoleic acid-derived lipid autacoids and by interfering with the production of anti-nociceptive lipid autacoids derived from n-3 fatty acids. Here, we used a rat model to determine the effect of increasing dietary linoleic acid as a controlled variable for 15 weeks on nociceptive lipid autacoids and their precursor n-6 and n-3 fatty acids in tissues associated with idiopathic pain syndromes.ResultsIncreasing dietary linoleic acid markedly increased the abundance of linoleic acid and its pro-nociceptive derivatives and reduced the abundance of n-3 eicosapentaenoic acid and docosahexaenoic acid and their anti-nociceptive monoepoxide derivatives. Diet-induced changes occurred in a tissue-specific manner, with marked alterations of nociceptive lipid autacoids in both peripheral and central tissues, and the most pronounced changes in their fatty acid precursors in peripheral tissues.ConclusionsThe present findings provide biochemical support for the hypothesis that the high linoleic acid content of modern industrialized diets may create a biochemical susceptibility to develop chronic pain. Dietary linoleic acid lowering should be further investigated as part of an integrative strategy for the prevention and management of idiopathic pain syndromes.

Dietary linoleic acid-induced alterations in pro- and anti-nociceptive lipid autacoids: Implications for idiopathic pain syndromes?

Dietary linoleic acid-induced alterations in pro- and anti-nociceptive lipid autacoids: Implications for idiopathic pain syndromes?

Pro-resolving mediators produced from EPA and DHA: Overview of the pathways involved and their mechanisms in metabolic syndrome and related liver diseases

A novel genus of pro-resolving lipid mediators endogenously generated from omega-3 polyunsaturated fatty acids has been identified in exudates obtained during the resolution phase of acute inflammation. The term specialized pro-resolving mediators (SPM) has been coined for these lipid mediators, comprising four novel chemical mediator families designated resolvins of the E series (if derived from eicosapentaenoic acid) and resolvins of the D series, protectins and maresins (if generated from docosahexaenoic acid). These SPM act not only as “stop-signals” of inflammatory response, but also as facilitators of the ability of macrophages to clear apoptotic cells (efferocytosis) and migrate to peripheral lymph nodes (efflux), thus, expediting their removal from sites of inflammation. In this review, we provide an overview of the current efforts to elucidate the structure-function, biosynthesis and actions of these omega-3-derived SPM in the context of inflammatory diseases. We specifically highlight the role of these SPM as endogenous counter-regulators of the persistent inflammatory status present in adipose tissue of obese individuals and describe the potential therapeutic impact of these bioactive lipid autacoids on the prevention of hepatic co-morbidities associated with obesity and the metabolic syndrome.