Glioma Cell Secretion: a Driver of Tumor Progression and a Potential Therapeutic Target Damian A

Published OnlineFirst October 17, 2018; DOI: 10.1158/0008-5472.CAN-18-0345

Cancer Review Research

Glioma Cell Secretion: A Driver of Tumor Progression and a Potential Therapeutic Target Damian A. Almiron Bonnin1,2, Matthew C. Havrda1,2, and Mark A. Israel1,2,3

Abstract

Cellular secretion is an important mediator of cancer progres- ple oncogenic pathologies. In this review, we describe tumor cell sion. Secreted molecules in glioma are key components of secretion in high-grade glioma and highlight potential novel complex autocrine and paracrine pathways that mediate multi- therapeutic opportunities. Cancer Res; 78(21); 6031–9. 2018 AACR.

Introduction Glioma-Secreted Molecules Impact Disease Glial cells in the central nervous system (CNS) provide trophic Progression support for neurons (1). In glial tumors, this trophic support is Glioma cells modify their microenvironment by introducing dysregulated creating a pro-oncogenic microenvironment medi- diverse molecules into the extracellular space (Table 1). To exem- ated by a heterogeneous array of molecules secreted into the plify the pro-oncogenic role that secreted molecules can have on – extracellular space (2 15). The glioma secretome includes pro- glioma pathology, we review the functional impact of specific teins, nucleic acids, and metabolites that are often overexpressed cytokines, metabolites, and nucleic acids on glioma biology. By in malignant tissue and contribute to virtually every aspect of describing some of the potent antitumorigenic effects observed in – cancer pathology (Table 1; Fig. 1; refs. 2 15), providing a strong preclinical therapeutic studies targeting tumor cell secretion, we – rationale to target the cancer cell secretory mechanisms. also highlight how blocking secreted molecules might be of fi Although the speci c mechanisms regulating secretion in therapeutic impact in gliomas, as well as other tumors. malignant cells remain to be fully characterized, there is significant evidence that the secretory mechanisms themselves are Cytokines altered during oncogenesis (8, 16–36). Well-known mediators Cytokines are essential mediators of cellular signaling (2, 13, of secretion, like the ADP-ribosylation factors (ARF) and the small 15). In glioma, secreted cytokines, including IL1b, IL6, and IL8, Rab GTPase proteins (RAB) have been reported to be dysregulated create a state of chronic inflammation that promotes the malig- in glioma and several other tumors (17, 20, 22, 28, 31, 32, 37–41). nant phenotype (15). These cytokines are associated with poor These proteins facilitate secretion of pro-oncogenic molecules prognosis for patients with high-grade gliomas (HGG; refs. 2, 13, (28, 42) and their inhibition diminishes multiple aspects of 15). Both in vitro and in vivo studies demonstrate that targeting cancer pathology including cellular proliferation, survival, and these mediators of inflammation inhibits important aspects of invasion (20, 22, 28, 32, 37–39, 41, 42), while showing no signs glioma pathology including angiogenesis, proliferation, and inva- of obvious toxicities in animal models (18, 19, 21, 26, 36, 43–46). sion (2, 13, 15). It has also been shown that cytokines, like IL6, This reliance of cancer cells on secretory pathways is exemplified IL8, EGF, and TGFb, promote resistance to antineoplastic therapy by the unfolded protein response (UPR; ref. 18). UPR activation is in glioma (15, 24, 47, 48), breast (49), and prostate cancer (50). In thought to be crucial for oncogenic progression (18), and agents melanoma, secretion has been identified as an important mech- inhibiting the UPR have shown potent antitumorigenic effects in anism facilitating the emergence of drug resistance via the acti- models of glioma, multiple myeloma, and pancreatic cancer (18). vation of the AKT pathway (51). Importantly, these cytokines also Tumor cell secretory "addiction" describes the dependence of facilitate the maintenance of cancer stem cells (Table 1; ref. 15), tumor cells on secretory pathways like the UPR (18), and suggest which are largely refractory to therapy, and play an important role a potential therapeutic window to target these pathways. The in cancer progression (52). functional impact of secreted molecules and secretory pathways Platelet-derived growth factor (PDGF), one of the best charac- on glioma biology underscores the potential therapeutic implica- terized cytokines in HGGs and other cancers (2, 13, 53–55), is the tions of targeting the tumor cell secretion (Table 1). ideal example to illustrate the functional impact of cytokines on cancer biology. Autocrine PDGF signaling was determined to play 1Department of Molecular and Systems Biology, Geisel School of Medicine at an important role in malignant transformation (2), and mouse Dartmouth, Hanover, New Hampshire. 2Norris Cotton Cancer Center, Geisel models of HGG demonstrate that PDGF signaling is sufficient for School of Medicine at Dartmouth, Lebanon, New Hampshire. 3Departments of tumor initiation and progression (13, 54). Dysregulated PDGF Medicine and Pediatrics, Geisel School of Medicine at Dartmouth, Hanover, New signaling activates MAPK-ERK and PI3K-AKT, two nodal points Hampshire. critical for cell proliferation, resistance to apoptosis, and invasion Corresponding Author: Mark A. Israel, M.D., Norris Cotton Cancer Center at (10, 12, 56). PDGF-mediated PI3K/AKT activation has also been Dartmouth, One Medical Center Drive, Lebanon, NH 03756. Phone: 603-653- shown to regulate glucose metabolism facilitating the Warburg 3611; Fax: 603-653-9003; E-mail: [email protected] effect in HGGs (12). In vitro and in vivo studies confirm PDGF's doi: 10.1158/0008-5472.CAN-18-0345 recognized function enhancing tumor angiogenesis by stimulat- 2018 American Association for Cancer Research. ing endothelial cell migration and promoting endothelial cell

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Table 1. Glioma-secreted molecules and the hallmarks of cancer they affect Secreted molecules functional in glioma Impacted hallmark of cancer Platelet-derived growth factor (2, 116), hepatocyte growth factor (56, 117), insulin-like growth factor (56, 118), Sustaining proliferative transforming growth factor a (119), adrenomedullin (120), epidermal growth factor receptor variant III (121), signaling sphingosine-1-phosphate (122) microRNA-17 (14, 123), microRNA-19 (14, 123), microRNA-21 (14, 123), microRNA-24 (123, 124), microRNA-26a (14, 123), Evading growth suppression microRNA-221/222 (123, 124) Transforming growth factor b2 (125), interleukin-10 (126), kynurenine (11), lactate dehydrogenase (14, 127), osteopontin (128) Avoiding immune destruction Vascular endothelial growth factor (43), basic fibroblast growth factor (5, 14), interleukin-6 (15, 129), interleukin-8 (15, 130), Enabling replicative C-X-C motif chemokine ligand 12 (131, 132), gremlin 1 (133), sema 3C (134), periostin (135, 136), sphingosine-1-phosphate immortality/ stemness (122), telomerase reverse transcriptase transcript (14), transferrin (137) Tumor necrosis factor a (138), interleukin-1b (138), interleukin-6 (15, 129), interleukin-8 (15, 139), glutamate (140) Tumor-promoting inflammation Transforming growth factor a (6, 119), hepatocyte growth factor (117), EGF (6), periostin (135, 136), osteopontin (128), C-X-C Activating invasion and motif chemokine ligand 12 (131), glial cell–derived neurotrophic factor (141), urokinase-type plasminogen activator metastasis (142), protease nexin 1 (143), metalloproteinase 2 (144), metalloproteinase 9 (144), autotaxin (145), kynurenine (11), glutamate (146), versican (147), laminins (148), metastasis-associated lung adenocarcinoma noncoding RNA (14, 149), sphingosine-1-phosphate (6, 122), microRNA-20a (14, 123), microRNA-21 (14, 123) Vascular endothelial growth factor (14), TGFb2 (125, 150), fibroblast growth factor (14, 150), hepatocyte growth factor Inducing angiogenesis (117, 150), epidermal growth factor (6, 150), interleukin-6 (15, 129), interleukin-8 (15, 138), C-X-C motif chemokine ligand 12 (151), angiogenin (14, 152), platelet-derived growth factor (116, 150) Kynurenine (4, 11), human endogenous retrovirus retrotransponson (3, 153), long interspersed nuclear element 1 Genome instability retrotransponson (3, 154), arthrobacter luteus (Alu) retrotransponson (3, 155) Vascular endothelial growth factor (14, 56), fibroblast growth factor (5, 14, 56), epidermal growth factor (6, 56), interleukin-6 Resisting cell death (15, 129), Sema 3C (134), microRNA-21 (14, 123), microRNA-92 (14, 123) Platelet derived growth factor (12, 116, 156), vascular endothelial growth factor (14, 156), fibroblast growth factor (14, 156), Deregulating cellular energetics hepatocyte growth factor (117, 156), epidermal growth factor (6, 156)

proliferation (2). Consistent with these findings, disrupting PDGF of disease progression (62). In glioma and other tumors, signaling markedly reduces angiogenesis, tumor growth, and nucleic acids can be secreted in extracellular vesicles (EV) and invasion in multiple mouse models of glioma (2, 10, 12, 13). deliveredtonearbycells(3,7,9, 14). EVs can carry mutated or amplified oncogene sequences, mRNA, transposable elements, Metabolites ormiRNAs(7,9,14).EVsfromHGGcontainanarrayofpro- Malignant cells reprogram their metabolism to meet the bio- oncogenic miRNAs such as miR-19b, miR-20, and miR-21 that energetic and biosynthetic demands of tumor growth (57). A promote HGG progression (14). Importantly, genetic material consequence of this altered metabolism is the aberrant produc- contained in these EVs, such as mRNA and miRNAs, can be tion of metabolites that can have profound effects on tumor delivered to nearby cells where they remain functional (7, 9, biology (57). Studies suggest that several metabolites, including 14). Glioma-derived cells incubated in medium supplemented well-known molecules like glutamate and 2-hydroxyglutarate, are with EVs from patient-derived glioblastoma cells exhibit involved in glioma progression (58, 59). increased proliferation when compared with glioma cells incu- A recently described, secreted pro-oncogenic metabolite in bated in normal, untreated growth medium (14). Although glioma is kynurenine (4, 11). Kynurenine is an intermediate in further experimentation is required to demonstrate specific L-tryptophan catabolism and an endogenous aryl hydrocarbon effects of each nucleic acid carried within EVs, it is highly likely receptor (AhR) agonist (4, 11). At levels secreted by glioma cells, that they are, at least in part, responsible for these EV-mediated but not at levels secreted by normal cells, kynurenine promotes effects in glioma. genomic instability of HGG-derived cells (4). AhR activation by Studies have demonstrated that EVs derived from glioma cells secreted kynurenine upregulates the trans-lesion synthesis poly- can transform fibroblasts and epithelial cells (8). Consistent with merase, hpol k (4). The dysregulated overexpression of this these findings, targeting proteins required for EV biogenesis polymerase, which normally mediates the DNA damage tolerance effectively blocks EV-induced oncogenic transformation (8). pathway, increases genomic instability and promotes tumorigen- These findings suggest that DNA transfer in EVs could transform esis (4, 60). Such an autocrine pathway mediating increased nearby cells contributing to the high degree of heterogeneity and genomic instability may contribute to the exceptionally high polyclonality observed in human HGGs (61, 63). levels of genomic heterogeneity characterizing HGG (61). Kynurenine also has been shown to have immunosuppressive Molecular Mechanisms of Secretion effects, to enhance cellular motility and to promote survival of HGG cells (11). Inhibition of kynurenine secretion in glioma Classical secretory pathway increases the lysis of glioma cells by alloreactive blood mononu- Malignant cells utilize the classical secretory pathway. In this clear cells, decreases cellular migration in vitro, and markedly pathway, proteins with a signal peptide are recognized by the decreases clonogenic survival of glioma cells (11). Consistent signal recognition particle and then inserted into the endoplas- with these observations, kynurenine-secreting tumors grow sig- mic reticulum (ER; ref. 64). In the ER, translation is completed nificantly more rapidly and have a higher proliferation index than and the newly synthesized proteins are transported through the nonsecreting tumors in immunocompetent mice (11). Golgi apparatus to the plasma membrane in a process mediated by numerous proteins including RABs and ARFs (64). Nucleic acids Accumulating evidence indicates that RABs and ARFs are of Secreted nucleic acids are commonly found in the blood of significant importance for cancer progression (17, 20, 28, 37, 65, patients with cancer and they have been shown to be indicative 66). Aberrant expression of RAB proteins, a family of more than

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Glioma Secretion Drives Tumor Progression

Sustaining proliferative Evading growth signaling suppressors

Deregulating Avoiding immune cellular energetics destruction Membrane Classical transport proteins secretion

Type II Type IV or or ABC Golgi transporter Golgi Bypass based Enabling Resisting secretion replicative cell death Nucleus ER immortality

Type III or auto- Exosomes Multivesicular body phagosome based secretion

Type I or pore Microvesicles based secretion Genome instability Tumor-promoting & mutation inﬂammation

Inducing Activating invasion angiogenesis & metastasis

Figure 1. Schematic representation of the impact of glioma secretion on the hallmarks of cancer. Pro-oncogenic molecules can be transported through the plasma membrane by classical secretory mechanisms, nonclassical secretory mechanisms (type I–IV; refs. 5, 15, 16, 19, 21, 25, 26, 29, 33, 72–81, 83), microvesicles, and exosomes (3, 8, 9, 14, 28, 32, 36). Molecules secreted by these mechanisms have been reported to contribute to each of the hallmarks of cancer (red arrows; refs. 2–15), as deﬁned by Hanahan and Weinberg (114).

70 members, is observed in glioma and other cancers (22, 28, 31, in vitro, and this oncogenic RAB35 was sufficient to drive vesicular 32, 40, 67, 68). In HGG RAB3A, RAB34, and RAB27A expression transport in the absence of stimuli, suggesting that dysregulated levels correlate with survival and pathologic grade (22, 31, 40). vesicular trafficking can contribute to oncogenesis (70). RAB3A, which regulates vesicular transport in neurons (69), is Like the RAB family of proteins, ARFs and their associated associated with increased proliferation and chemo- and radio- guanine nucleotide exchange factors (GEF) and GTPase-activating resistance of cultured glioma cells and tumor formation and proteins (GAP) are emerging as novel mediators of disease pro- glioma growth in nude mice (22). A recent study has identified gression in gliomas, as well as other tumors (17, 20, 36–39, RAB35 as a novel activator of PI3K in multiple cells from kidney, 65, 66). Dysregulated ARF6 has been reported to be a key driver colon, prostate, and cervical cancer. Somatic RAB35 mutations of tumor cell invasion in several cancer types including glioma found in human tumors were capable of transforming cells (17, 20, 65, 66). A glioma study has shown that ARF6 knockdown

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by siRNA markedly decreases invasion of intracranial brain xeno- useful in the clinic due to the side effects they cause (19). Multiple grafts in nude mice (20). ARF6 has also been shown to orchestrate strategies to prevent such drug toxicities are currently under the activity of the oncogenic guanine nucleotide–binding protein investigation (26). G(q) subunit alpha (GNAQ) in uveal melanoma by mediating the The type III pathway of nonclassical secretion relies on autop- vesicular transport of GNAQ and b-catenin (71). Consistent with hagosomes (72), double-membraned vesicles that enclose cellu- this finding, targeting ARF6 with the small-molecule inhibitor, lar material destined to be degraded or secreted (25). Secretory NAV-2729, reduces melanoma proliferation and tumorigenesis autophagosomes carry cytokines including IL1b, and IL18 (25, in vivo (71). 72), which are well-characterized mediators of glioma progression (15, 75, 76). The process of autophagosome formation is Nonclassical secretory pathway tightly controlled by highly conserved autophagy-related genes A substantial proportion of cancer secretion is mediated by (ATG; refs. 25, 72). These genes, as well as the process of autop- the nonclassical secretory pathway (29). This pathway is hagy, can have pro-oncogenic or antioncogenic effects in HGG induced by cellular stress (72), a characteristic of transformed (30, 79, 80). For example, targeting ATG7 or ATG13 has been cells (18, 73). There are four types of nonclassical pathway shown to result in markedly decreased KRAS-driven HGG growth secretion, which may play a role in cancer: type I or pore- in vivo (80), while inhibiting ATG5 enhances the resistance of mediated secretion, type II or ABC transporter-mediated secre- cultured HGG cells to temozolomide, a drug routinely used in the tion, type III or endosome/autophagosome-mediated secretion, treatment of glioblastoma (30). The mechanisms behind this dual and type IV or Golgi-bypass secretion (72). role of ATGs in HGG remain to be elucidated (79). Proteins secreted by the type I pathway rely on the formation of The type IV nonclassical pathway mediates the secretion of pores in the plasma membrane. The formation of these pores can proteins during ER stress that bypass the Golgi enroute to the be mediated by either the cytoplasmic protein to be secreted or by plasma membrane following synthesis (72). The IRE1a protein of inflammation (72). Secretion of FGF2, a widely studied pro- the UPR activates this pathway in response to ER stress (18, 81), oncogenic protein of importance in glioma (5), is a well-known which is provoked by many conditions common to malignant example of the former mechanism (72). FGF2 binds to phospha- tissues including hypoxia, nutrient deprivation, increased meta- tidylinositol 4, 5-bisphosphate (PI(4,5)P2) on the plasma mem- bolic activity, and high levels of proliferation (18, 73, 82). Human brane and oligomerizes promoting the formation of a lipidic pore glioma cells expressing a nonfunctional mutant of IRE1a dem- through which secretion occurs (72). Alternatively, pore forma- onstrate markedly decreased intracranial tumor growth in nude tion can be mediated by inflammation (72, 74), which charac- mice (82). Golgi bypass in the type IV pathway is mediated by an terizes the glioma microenvironment (75). During inflammation, ER-sorting machinery that includes heat shock proteins (HSP; ref. gasdermin D, gasdermin A, and gasdermin A3 are cleaved by 72). The role of HSPs in HGGs has been well described (33, 83). caspase-1 releasing the N-terminal half of these proteins: gasder- For example, HSP70, which is upregulated in HGGs (33, 83), min-N, which binds PIP2 on the plasma membrane and forms 16- increases tumor formation, survival, and chemoresistance in fold-symmetry in the pore (72, 74). These pores then mediate the glioblastoma (33, 83). secretion of IL1b (74), a cytokine known to promote further inflammation and drive glioma growth (75, 76). While a role Extracellular vesicle formation and secretion for the gasdermin protein family in glioma has not been pro- Many different terms are used in the literature to describe EVs posed, members of this family are aberrantly expressed in several including microvesicles, exosomes, oncosomes, microparticles, tumor types (16) and promote brain metastasis of tumors from and ectosomes (84, 85). Even although there is considerable outside the CNS (77). debate in the field as to the proper usage of these terms (84, 85), The type II nonclassical pathway relies on the ATP-binding EVs can be broadly classified into two main groups based on cassette (ABC) transporters (72), which are well-described med- their biogenesis: microvesicles and exosomes (36). Microvesi- iators of chemoresistance in glioma as well as other tumor types cles are derived from the plasma membrane, while exosomes (27, 78). These transporters facilitate chemoresistance by medi- originate from multivesicular endosomal compartments (36). ating the efflux of chemotherapeutic agents from malignant cells However, experimental procedures designed to isolate each of (27, 78). Overexpression of the ABC transporter subfamily B these vesicle types, including centrifugation protocols, com- member 1, which is also known as the multidrug resistance mercial kits, and filters,arebasedonthesizeandweightof protein 1 (MDR1), mediates resistance to chemotherapeutic these vesicles, and not on their intracellular origin (86, 87). agents in aggressive gliomas (27, 78). Inhibiting MDR1, in vitro Consequently, material isolated by these experimental techni- or in vivo, increases the sensitivity of HGG cells to vincristine and ques contain a mixture of different types of EVs (86, 87), and temozolomide, two antineoplastic agents commonly used in the thus it is important to consider investigational findings in the management of patients with glioblastoma (78). Furthermore, light of these experimental limitations. MDR1 inhibitors have been shown to lead to increased concen- In cancer, EVs transport proteins, lipids, and nucleic acid (36) trations of chemotherapy agents in the brain when used as and can promote multiple aspects of cancer progression (3, 9, 14, adjuvants (19, 21). MDR1 is also expressed in endothelial cells 32, 36, 66), and even induce malignant transformation (8, 88). of the blood–brain barrier (19), where it pumps metabolic waste The small Ras homolog gene family member A (RhoA) GTPase products out of the CNS as well as enhancing the efflux of regulates EV biogenesis in glioma cells (8, 36). Studies demon- chemotherapy agents and decreasing their availability in brain strate that RhoA facilitates the activation of Rho kinase (ROCK) tumors (19). Thus, clinical therapies that effectively inhibited and subsequently the Lim kinase (LIMK; refs. 8, 36). LIMK these transporters could potentially have the added advantage of phosphorylates cofilin causing a reorganization of actin fibers increasing the CNS availability of chemotherapy agents active in and microvesicle shedding (8, 36). Inhibition of these proteins glioma (21), although to date, MDR1 inhibitors have not been has been shown to markedly reduce microvesicle release in

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glioma-derived cells (8). Inhibition of the RhoA–ROCK–LIMK EI, which inhibits Sec61-mediated protein translocation in pathway reduces cellular proliferation and migration in glioma the classical secretory pathway (44), has potent antioncogenic (8, 35). Pharmacologic inhibition of this pathway by the ROCK activity (46, 89). Preclinical studies demonstrated that EI treat- inhibitor, Y-27632, suppresses migration and proliferation of ment causes a significant reduction of invasion and cellular glioma-derived cells. Similarly, silencing of LIMK reduced growth proliferation of a non–smallcelllungcarcinoma–derived cell of a breast cancer xenograft in nude mice (8). Consistent with line, while increasing cell death by apoptosis (46, 89). In vivo,EI these observations, RhoA, which mediates the activation of ROCK treatment resulted in reduced xenograft growth in nude mice and LIMK as discussed previously (8, 36), is highly expressed in with no signs of toxicity (46). Other Sec61 inhibitors, like HGGs and its expression is correlated with pathologic grade (34). apratoxin (104) and cotransins (105), have also been shown RAB and ARF family members, which were discussed in previous to have significant antitumorigenic activity in preclinical stud- sections, have also been implicated in EV formation in gliomas as ies (106–108). However, these agents, as well as other Sec61 well as other types of cancer (32, 36, 66). inhibitors, like decatransin (109) and mycolactone (110), remain to be studied in the context of HGG, and they have Therapeutic Opportunities yet to be studied in clinical trials for any tumor type. These pharmacologic inhibitors of secretion represent a relatively Targeting secretion in glioma unexplored therapeutic opportunity for tumors, like HGG, that The concept of tumor cell–secretory addiction describes the rely on secretory pathways, as discussed above. reliance of cancer cells on secreted molecules and their secretory Another potential target to inhibit cancer-associated secretion machinery (18). Studies described above demonstrate the signif- is the UPR (18). Treatment of HGG cells with epigallocatechin icance of this concept in HGG (Table 1). By describing some of the gallate, which inhibits GRP78, significantly increases the cytotoxic essential functions of glioma-secreted molecules and the mechan- effect of temozolomide on HGG (90). This drug is currently being isms facilitating their secretion in glioma pathology (Fig. 1), we evaluated as an adjuvant for treatment of colorectal cancer in a have sought to underscore the potential therapeutic benefits of phase I clinical trial (111). Similarly, inhibition of the glucose- targeting secretion in HGGs. Targeting proteins of the secretory regulated protein 78 (GRP78), a key regulator of the UPR path- mechanism in glioma has been shown to inhibit key aspects of way, by the novel fusion protein EGF-SubA results in substantial glioma biology including tumor cell proliferation and survival growth reduction of HGG xenografts in nude mice (45). (Table 1; Fig. 1; refs. 8, 17–23, 26–28, 32, 34–36, 38, 39, 44– Botulinum neurotoxin–based pharmacologic agents represent 46, 89, 90). Furthermore, targeting secretion seems to selectively another promising strategy to inhibit secretion (112). These affect transformed cells, while sparing normal cells, as evidenced drugs, which are known as targeted secretion inhibitors (TSI), by the lack of toxicities reported in multiple animal experiments can target specific cells and cleave SNARE proteins (112). TSIs testing agents that inhibit secretion (18, 19, 21, 26, 36, 43–46). contain a targeting domain, a transport domain, and a blocking This suggests that targeting proteins of the secretory machinery to domain. The targeting domain can be engineered to bind classic disturb the secretion of molecules that enhance tumor pathology glioblastoma markers, such as EGFR (113). The transport domain may be a promising treatment strategy. facilitates the entry of this agent into the cytosol, where the Brefeldin A, a known secretion inhibitor (23), and EHT-1864, enzymatic blocking domain cleaves SNARE proteins to inhibit which inhibits Rac1 (91), a known mediator of secretion (92, 93), protein secretion (112). Even although TSIs have yet to be dramatically reduce VEGF secretion, decrease GSC self-renewal, explored in the context of cancer, their characteristics discussed and inhibit tumor growth in a mouse model of HGG (54). The above make them well suited for cancer therapeutics. antitumorigenic effects of these agents also have been observed in several other cancer types (38, 39, 41). Multiple high-throughput drug screens have identified novel small-molecule inhibitors that Conclusion effectively target mediators of cellular secretion (94). Two small- Cancer cell secretion is a vital process contributing to glioma molecule inhibitors of secretion that have been studied in cancer progression. Numerous studies have characterized the autocrine are AMF-26 (37) and Eeyarestatin I (EI; ref. 44). Like Brefeldin A, and paracrine pathways that facilitate many key aspects of tumor AMF-26 targets the ARF1-guanine nucleotide exchange factor progression described as hallmarks of cancer by Hanahan and (GEF) interaction, which prevents the assembly of the vesicle Weinberg (Table 1; Fig. 1; ref. 114). The capacity of cancer cells to coating protein complex I (COPI) assembly and consequently secrete a wide range of different soluble factors with redundant inhibits protein secretion (37). AMF-26 has potent antitumor functions (Table 1) could also explain cancer resistance to anti- activity (37). Oral administration of AMF-26 resulted in complete neoplastic therapies. In HGG, studies have shown that when regression of human breast cancer xenografts in nude mice, and signaling from a specific secreted factor is blocked, an alternative was nontoxic to animals (37) providing further evidence of the secreted factor can maintain the oncogenic functions of the potential of such drugs to have an acceptable therapeutic index. secreted factor that was blocked (54, 115). These secreted factors Early preclinical studies also suggest that other pharmacologic have also been observed to promote resistance to radiation and agents that result in the inhibition of ARF1-mediated COPI chemotherapy (15, 24, 47–51). Therefore, targeting the secretory assembly, like Exo2 (8), LM11 (95), and Golgicide A (96), have mechanisms of cancer cells could potentially reduce simulta- significant antitumorigenic activity in breast and prostate cancer neously the levels of multiple pro-oncogenic secreted factors and models (97–99). However, these agents, as well as other agents consequently diminish cancer drug resistance and increase patient that block ER-to-Golgi vesicular transport, including Exo1 (100), survival. However, despite accumulating evidence demonstrating LG186 (100), CI-976 (101), dispergo (102), and FLI-06 (103), the importance of these secretory mechanisms in glioma biology have not been tested in glioblastoma and their clinical value for (8, 16–36), there are currently no clinically available therapies cancer therapeutics remains largely unexplored. targeting these mechanisms.

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As pharmacologic agents targeting secretory mechanisms Disclosure of Potential Conflicts of Interest emerge, it will become important to cautiously consider their No potential conflicts of interest were disclosed. safety. Although several drugs targeting secretory mechanisms have been tested in animal models with no observable toxicities Acknowledgments (18, 19, 21, 26, 36, 43–46), inhibiting such an essential cellular Support was generously provided by the Theodora B. Betz Foundation function as secretion poses obvious risks. Many cell types, and (toM.A.Israel),theJordanandKyra Memorial Foundation (to M.A. Israel), and the Andrea Clark Nelson Medical Research Endowment especially neurons, glial cells, and immune cells, require secretion (to M.A. Israel). to function properly. Much like current chemotherapeutic drugs, dose levels and dosing schedules are likely to require careful evaluation to target secretion from malignant cells, while sparing Received February 2, 2018; revised May 30, 2018; accepted August 14, 2018; secretion from normal cells. published first October 17, 2018.

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www.aacrjournals.org Cancer Res; 78(21) November 1, 2018 6039

Glioma Cell Secretion: A Driver of Tumor Progression and a Potential Therapeutic Target

Damian A. Almiron Bonnin, Matthew C. Havrda and Mark A. Israel

Cancer Res 2018;78:6031-6039. Published OnlineFirst October 17, 2018.

Updated version Access the most recent version of this article at: doi:10.1158/0008-5472.CAN-18-0345

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