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HPSE2, the gene‐encoding heparanase 2 (Hpa2), is mutated in urofacial syndrome (UFS), a rare autosomal recessive congenital disease attributed to peripheral neuropathy. Hpa2 lacks intrinsic heparan sulfate (HS)‐degrading activity, the hallmark of heparanase (Hpa1), yet it exhibits a high affinity toward HS, thereby inhibiting Hpa1 enzymatic activity. Hpa2 regulates selected genes that promote normal differentiation, tissue homeostasis, and endoplasmic reticulum (ER) stress, resulting in antitumor, antiangiogenic, and anti‐inflammatory effects. Importantly, stress conditions induce the expression of Hpa2, thus establishing a feedback loop, where Hpa2 enhances ER stress which, in turn, induces Hpa2 expression. In most cases, cancer patients who retain high levels of Hpa2 survive longer than patients bearing Hpa2‐low tumors. Experimentally, overexpression of Hpa2 attenuates the growth of tumor xenografts, whereas Hpa2 gene silencing results in aggressive tumors. Studies applying conditional Hpa2 knockout (cHpa2‐KO) mice revealed an essential involvement of Hpa2 contributed by the host in protecting against cancer and inflammation. This was best reflected by the distorted morphology of the Hpa2‐null pancreas, including massive infiltration of immune cells, acinar to adipocyte trans‐differentiation, and acinar to ductal metaplasia. Moreover, orthotopic inoculation of pancreatic ductal adenocarcinoma (PDAC) cells into the pancreas of Hpa2‐null vs. wild‐type mice yielded tumors that were by far more aggressive. Likewise, intravenous inoculation of cancer cells into cHpa2‐KO mice resulted in a dramatically increased lung colonization reflecting the involvement of Hpa2 in restricting the formation of a premetastatic niche. Elucidating Hpa2 structure–activity‐relationships is expected to support the development of Hpa2‐based therapies against cancer and inflammation. [ABSTRACT FROM AUTHOR]

Cell surface proteoglycans are important constituents of the glycocalyx and participate in cell–cell and cell–extracellular matrix (ECM) interactions, enzyme activation and inhibition, and multiple signaling routes, thereby regulating cell proliferation, survival, adhesion, migration, and differentiation. Heparanase, the sole mammalian heparan sulfate degrading endoglycosidase, acts as an "activator" of HS proteoglycans, thus regulating tissue hemostasis. Heparanase is a multifaceted enzyme that together with heparan sulfate, primarily syndecan-1, drives signal transduction, immune cell activation, exosome formation, autophagy, and gene transcription via enzymatic and nonenzymatic activities. An important feature is the ability of heparanase to stimulate syndecan-1 shedding, thereby impacting cell behavior both locally and distally from its cell of origin. Heparanase releases a myriad of HS-bound growth factors, cytokines, and chemokines that are sequestered by heparan sulfate in the glycocalyx and ECM. Collectively, the heparan sulfate–heparanase axis plays pivotal roles in creating a permissive environment for cell proliferation, differentiation, and function, often resulting in the pathogenesis of diseases such as cancer, inflammation, endotheliitis, kidney dysfunction, tissue fibrosis, and viral infection. [ABSTRACT FROM AUTHOR]

Heparanase, the sole heparan sulfate (HS)-degrading endoglycosidase, regulates multiple biological activities that enhance tumor growth, metastasis, angiogenesis, and inflammation. Heparanase accomplishes this by degrading HS and thereby regulating the bioavailability of heparin-binding proteins; priming the tumor microenvironment; mediating tumor–host crosstalk; and inducing gene transcription, signaling pathways, exosome formation, and autophagy that together promote tumor cell performance and chemoresistance. By contrast, heparanase-2, a close homolog of heparanase, lacks enzymatic activity, inhibits heparanase activity, and regulates selected genes that promote normal differentiation, endoplasmic reticulum stress, tumor fibrosis, and apoptosis, together resulting in tumor suppression. The emerging premise is that heparanase is a master regulator of the aggressive phenotype of cancer, while heparanase-2 functions as a tumor suppressor. [ABSTRACT FROM AUTHOR]

High heparanase expression is associated with enhanced tumor growth, angiogenesis, and metastasis in many types of cancer. However, the mechanisms driving high heparanase expression are not fully understood. In the present study, we discovered that drugs used in the treatment of myeloma upregulate heparanase expression. Frontline anti-myeloma drugs, bortezomib and carfilzomib activate the nuclear factor-kappa B (NF-κB) pathway to trigger heparanase expression in tumor cells. Blocking the NF-κB pathway diminished this chemotherapy-induced upregulation of heparanase expression. Activated NF-κB signaling was also found to drive high heparanase expression in drug resistant myeloma cell lines. In addition to enhancing heparanase expression, chemotherapy also caused release of heparanase by tumor cells into the conditioned medium. This soluble heparanase was taken up by macrophages and triggered an increase in TNF-α production. Heparanase is also taken up by tumor cells where it induced expression of HGF, VEGF and MMP-9 and activated ERK and Akt signaling pathways. These changes induced by heparanase are known to be associated with the promotion of an aggressive tumor phenotype. Importantly, the heparanase inhibitor Roneparstat diminished the uptake and the downstream effects of soluble heparanase. Together, these discoveries reveal a novel mechanism whereby chemotherapy upregulates heparanase, a known promoter of myeloma growth, and suggest that therapeutic targeting of heparanase during anti-cancer therapy may improve patient outcome. [ABSTRACT FROM AUTHOR]

Abstract: Heparanase has generated substantial interest as therapeutic target for antitumor therapy, because its activity is implicated in malignant behavior of cancer cells and in tumor progression. Increased heparanase expression was found in numerous tumor types and correlates with poor prognosis. Heparanase, an endoglucuronidase responsible for heparan sulfate cleavage, regulates the structure and function of heparan sulfate proteoglycans, leading to disassembly of the extracellular matrix. The action of heparanase is involved in multiple regulatory events related, among other effects, to augmented bioavailability of growth factors and cytokines. Inhibitors of heparanase suppress tumor growth, angiogenesis and metastasis by modulating growth factor-mediated signaling, ECM barrier function and cell interactions in the tumor microenvironment. Therefore, targeting heparanase has potential implications for anti-tumor, anti-angiogenic and anti-inflammatory therapies. Current approaches for heparanase inhibition include development of chemically modified heparins, small molecule inhibitors and neutralizing antibodies. The available evidence supports the emerging utility of heparanase inhibition as a promising antitumor strategy, specifically in rational combination with other agents. The recent studies with compounds designed to block heparanase (e.g., modified heparins) provide a rational basis for their therapeutic application and optimization. [Copyright &y& Elsevier]

Abstract: Heparanase, the sole mammalian endoglycosidase degrading heparan sulfate, is causally involved in cancer metastasis, angiogenesis, inflammation and kidney dysfunction. Despite the wide occurrence and impact of heparan sulfate proteoglycans in vascular biology, the significance of heparanase in vessel wall disorders is underestimated. Blood vessels are highly active structures whose morphology rapidly adapts to maintain vascular function under altered systemic and local conditions. In some pathologies (restenosis, thrombosis, atherosclerosis) this normally beneficial adaptation may be detrimental to overall function. Enzymatic dependent and independent effects of heparanase on arterial structure mechanics and repair closely regulate arterial compliance and neointimal proliferation following endovascular stenting. Additionally, heparanase promotes thrombosis after vascular injury and contributes to a pro-coagulant state in human carotid atherosclerosis. Importantly, heparanase is closely associated with development and progression of atherosclerotic plaques, including stable to unstable plaque transition. Consequently, heparanase levels are markedly increased in the plasma of patients with acute myocardial infarction. Noteworthy, heparanase activates macrophages, resulting in marked induction of cytokine expression associated with plaque progression towards vulnerability. Together, heparanase emerges as a regulator of vulnerable lesion development and potential target for therapeutic intervention in atherosclerosis and related vessel wall complications. [Copyright &y& Elsevier]

Development of compounds that target both heparanase and selectins is emerging as a promising approach for cancer therapy. Selectins are vascular cell adhesion molecules that mediate tumor cell interactions with platelets, leukocytes, and the vascular endothelium. Heparanase is an endoglycosidase that degrades heparan sulfate in the tumor microenvironment, cell surfaces, and vessel wall. Acting together, these molecules facilitate tumor cell arrest, extravasation, and metastasis. Here, we report the preparation of novel semisynthetic sulfated tri mannose C-C-linked dimers (STMCs) endowed with heparanase and selectin inhibitory activity. The P-selectin specificity of the STMC was defined by the anomeric linkage of the C-C bond. This STMC hexasaccharide is an effective inhibitor of P-selectin in vivo. We show that selective inhibition of heparanase attenuates metastasis in B16-BL6 melanoma cells, expressing high levels of this endoglycosidase, but has no effect on the metastasis of MC-38 carcinoma cells that express little or no heparanase activity. P-selectin-specific STMC attenuated metastasis in both animal models, indicating that inhibition of tumor cell interaction with the vascular endothelium is critical for cancer dissemination. Thus, the small size, the stability of the C-C bond, and the chemically defined structure of the newly generated STMCs make them superior to heparin derivatives and signify STMCs as valuable candidates for further evaluation. [ABSTRACT FROM AUTHOR]

Mammalian heparanase, an endoglycosidase-degrading heparan sulfate, is synthesized as a latent 65 kDa precursor that undergoes proteolytic processing, primarily by cathepsin-L, yielding 8 kDa and 50 kDa subunits that heterodimerize to form a highly active enzyme. Enhanced heparanase expression in human tumors correlates with metastatic potential, tumor vascularity, and reduced postoperative survival of cancer patients, attributed to enzymatic and nonenzymatic activities of the heparanase protein. Urinary and plasma levels of heparanase are elevated in cancer patients and suppressed in response to effective anticancer treatments. These observations and the anticancerous effect of heparanase gene silencing and of heparanase-inhibiting molecules suggest that the enzyme is a promising target for anticancer drug development. [ABSTRACT FROM AUTHOR]

Heparanase is an endo-β-D-glucuronidase involved in extracellular matrix remodeling and degradation and implicated in tumor metastasis, angiogenesis, inflammation, and autoimmunity. The enzyme is synthesized as a latent 65-kDa protein and is processed in the lysosomal compartment to an active 58-kDa heterodimer, where it is stored in a stable form. In contrast, its heparan sulfate substrate is localized extracellularly, suggesting the existence of mechanisms that trigger heparanase secretion. Here we show that secretion of the active enzyme is mediated by the protein kinase A and C pathways. Moreover, secretion of active heparanase was observed upon cell stimulation with physiological concentrations of adenosine, ADP, and ATP, as well as by the noncleavable ATP analogue adenosine 5′-O-(thiotriphosphate). Indeed, heparanase secretion was noted upon cell stimulation with a specific P2Y1 receptor agonist and was inhibited by P2Y receptor antagonists. The kinetics of heparanase secretion resembled the secretion of cathepsin D, a lysosomal enzyme, indicating that the secreted heparanase is of lysosomal origin. We suggest that secretion of active heparanase is initiated by extracellular cues activating the protein kinase A and C signaling pathways. The secreted enzyme(s) then facilitate cell invasion associated with cancer metastasis, angiogenesis, and inflammation. [ABSTRACT FROM AUTHOR]

The immunomodulating capacity of heparin led us to test the effect of the synthetic heparin-mimicking and low anticoagulant compound RG-13577 on the course of experimental autoimmune encephalomyelitis (EAE) and central nervous system (CNS) inflammation. EAE was induced in SJL mice by inoculation with whole mouse spinal cord homogenate. RG-13577, delivered intraperitoneally, inhibited the clinical signs of acute EAE and markedly ameliorated inflammation in the spinal cord, primarily by inhibiting heparanase activity in lymphocytes and astrocytes and thus impairing lymphocyte traffic. RG-13577 treatment was effective when started on day of disease induction or day 7 after induction. The low molecular weight heparin, enoxaparin, tested under the same conditions, exerted only a minor insignificant inhibitory effect. RG-13577 also inhibited the tyrosine phosphorylation of several proteins, particularly Erk1 and Erk2 of the MAP kinase signaling pathways associated with inflammation and cell proliferation. RG-13577 blocked the activity of sPLA2 and inhibited CNS PGE2 production both in vivo and in vitro. [Copyright &y& Elsevier]

Cleavage of heparan sulphate proteoglycans (HSPGs) affects the integrity and functional state of tissues and thereby fundamental normal and pathological phenomena involving cell migration and response to changes in the extracellular microenvironment. Heparanase, degrading heparan sulphate (HS) at specific intrachain sites, is synthesized as a latent ∼65 kDa protein that is processed at the N-terminus into a highly active ∼50 kDa form. The heparanase enzyme is preferentially expressed in human tumours and its overexpression in low-metastatic tumour cells confers a highly invasive phenotype in experimental animals. Heparanase also releases angiogenic factors and accessory fragments of HS from the tumour microenvironment and induces an angiogenic response in vivo. Heparanase may thus facilitate tumour cell invasion, vascularization and survival in a given microenvironment, all critical events in cancer progression. These observations, the anticancerous effect of heparanase-inhibiting molecules, and the unexpected identification of a single predominant functional heparanase suggest that the enzyme is a promising target for drug development. [Copyright &y& Elsevier]

Heparan sulfate proteoglycans interact with many extracellular matrix constituents, growth factors and enzymes. Degradation of heparan sulfate by endoglycosidic heparanase cleavage affects a variety of biological processes. We have purified a 50-kDa heparanase from human hepatoma and placenta, and now report cloning of the cDNA and gene encoding this enzyme. Expression of the cloned cDNA in insect and mammalian cells yielded 65-kDa and 50-kDa recombinant heparanase proteins. The 50-kDa enzyme represents an N-terminally processed enzyme, at least 100-fold more active than the 65-kDa form. The heparanase mRNA and protein are preferentially expressed in metastatic cell lines and specimens of human breast, colon and liver carcinomas. Low metastatic murine T-lymphoma and melanoma cells transfected with the heparanase cDNA acquired a highly metastatic phenotype in vivo, reflected by a massive liver and lung colonization. This represents the first cloned mammalian heparanase, to our knowledge, and provides direct evidence for its role in tumor metastasis. Cloning of the heparanase gene enables the development of specific molecular probes for early detection and treatment of cancer metastasis and autoimmune disorders. [ABSTRACT FROM AUTHOR]

Heparanase is regarded as a promising target for anticancer drugs and Ronepastat is one of the most promising heparanase inhibitors insert in clinical study for Multiple Myeloma Therapy. To improve its pharmacokinetic/pharmacodynamic profile, as well to have an antidote able to neutralize its activity in case of over dosages or intolerance, a new class of its derivatives was obtained inserting non-carbohydrate moieties of different length between the polysaccharide chain and biotin or its derivatives. In vitro these novel derivatives maintain the anti-heparanase activity without induced toxicity. The newly synthesized compounds retained the ability to attenuate the growth of CAG myeloma tumors in mice with potency similar, or in one case even higher than that of the reference compound Roneparstat as well as inhibited metastatic dissemination (lung colonization) of murine B16-F10 melanoma cells in vivo. Image 1 • Heparanase is regarded as a promising target for anticancer drugs and Ronepastat is one of one of its most promising inhibitors. • A new class of biotinylated N-acetyl-gs-heparin have been synthetized through a linker of different functional groups. • Compounds were characterized by NMR and evaluation of molecular weight. • "in vitro" and "in vivo" tests showed that the chemical modifications introduced do not hinder their biological activity. [ABSTRACT FROM AUTHOR]

Heparanase (Hpa1) is expressed by tumor cells and cells of the tumor microenvironment and functions to remodel the extracellular matrix (ECM) and regulate the bioavailability of ECM-bound factors that support tumor growth. Heparanase expression is upregulated in human carcinomas, sarcomas, and hematological malignancies, correlating with increased tumor metastasis, vascular density, and shorter postoperative survival of cancer patients, and encouraging the development of heparanase inhibitors as anti-cancer drugs. Among these are heparin/HS mimetics, the only heparanase-inhibiting compounds that are being evaluated in clinical trials. We have synthesized dicarboxylated oxy-heparins (DCoxHs) containing three carboxylate groups per split residue (DC-Hep). The resulting lead compound (termed XII) was upscaled, characterized, and examined for its effectiveness in tumor models. Potent anti-tumorigenic effects were obtained in models of pancreatic carcinoma, breast cancer, mesothelioma, and myeloma, yielding tumor growth inhibition (TGI) values ranging from 21 to 70% and extending the survival time of the mice. Of particular significance was the inhibition of spontaneous metastasis in an orthotopic model of breast carcinoma following resection of the primary tumor. It appears that apart from inhibition of heparanase enzymatic activity, compound XII reduces the levels of heparanase protein and inhibits its cellular uptake and activation. Heparanase-dependent and -independent effects of XII are being investigated. Collectively, our pre-clinical studies with compound XII strongly justify its examination in cancer patients. [ABSTRACT FROM AUTHOR]

Abstract Emergence of the field of exosome biology has opened an exciting door to better understand communication between cells. These tiny nanovesicles act as potent regulators of biological function by delivering proteins, lipids and nucleic acids from the cell of origin to target cells. Recently, several enzymes including membrane-type 1 matrix metalloproteinase (MT1-MMP), insulin-degrading enzyme (IDE), sialidase and heparanase, among others, were localized on the surface of exosomes secreted by various cell types. These exosomal surface enzymes retain their activity and can degrade their natural substrates present within extracellular spaces. To date, enzymes on exosome surfaces have been associated with the mobilization of growth factors, degradation of extracellular matrix macromolecules and destruction of amyloid β plaques. This review focuses on the emerging role of exosomal surface enzymes and how this mechanism of remodeling within the extracellular space may regulate disease progression as related to cancer, inflammation and Alzheimer's disease. Highlights • Exosomes regulate cell communication by transferring proteins, lipids and nucleic acids between cells • New evidence demonstrates that enzymatically active proteases and glycosidases are present on the surface of some exosomes • These enzymes on exosome surfaces can degrade the ECM, liberate growth factors and alter cell adhesion and invasion • Enzymatic functions of exosomes have implications in the progression of cancer, inflammation and Alzheimer's disease [ABSTRACT FROM AUTHOR]

Multiple myeloma (MM) is a plasma cell malignancy that is accompanied by hypercalcemia, renal failure, anemia, and lytic bone lesions. Heparanase (HPSE) plays an important role in supporting and promoting myeloma progression, maintenance of plasma cell stemness, and resistance to therapy. Previous studies identified functional single nucleotide polymorphisms (SNPs) located in the HPSE gene. In the present study, 5 functional HPSE SNPs and 11 novel HPSE2 SNPs were examined. A very significant association between two enhancer (rs4693608 and rs4693084), and two insulator (rs4364254 and rs4426765) HPSE SNPs and primary paraskeletal disease (PS) was observed. SNP rs657442, located in intron 9 of the HPSE2 gene, revealed a significant protective association with primary paraskeletal disease and lytic bone lesions. The present study demonstrates a promoting (HPSE gene) and protective (HPSE2 gene) role of gene regulatory elements in the development of paraskeletal disease and bone morbidity. The effect of signal discrepancy between myeloma cells and normal cells of the tumor microenvironment is proposed as a mechanism for the involvement of heparanase in primary PS. We suggest that an increase in heparanase-2 expression can lead to effective suppression of heparanase activity in multiple myeloma accompanied by extramedullary and osteolytic bone disease. [ABSTRACT FROM AUTHOR]

Heparanase is the sole endoglucuronidase that degrades heparan sulfate in the cell surface and extracellular matrix (ECM). Several studies have reported the localization of heparanase in the cell nucleus, but the functional role of the nuclear enzyme is still obscure. Subjecting mouse embryonic fibroblasts (MEFs) derived from heparanase knockout (Hpse-KO) mice and applying transposase-accessible chromatin with sequencing (ATAC-seq), we revealed that heparanase is involved in the regulation of chromatin accessibility. Integrating with genome-wide analysis of chromatin states revealed an overall low activity in the enhancer and promoter regions of Hpse-KO MEFs compared with wild-type (WT) MEFs. Western blot analysis of MEFs and tissues derived from Hpse-KO vs. WT mice confirmed reduced expression of H3K27ac (acetylated lysine at N-terminal position 27 of the histone H3 protein). Our results offer a mechanistic explanation for the well-documented attenuation of inflammatory responses and tumor growth in Hpse-KO mice. [ABSTRACT FROM AUTHOR]

The emerging role of heparanase in tumor initiation, growth, metastasis, and chemoresistance is well recognized and is encouraging the development of heparanase inhibitors as anticancer drugs. Unlike the function of heparanase in cancer cells, very little attention has been given to heparanase contributed by cells composing the tumor microenvironment. Here we used a genetic approach and examined the behavior and function of macrophages isolated from wild-type (WT) and heparanase-knockout (Hpa-KO) mice. Hpa-KO macrophages express lower levels of cytokines (e.g., TNFα, IL1-β) and exhibit lower motility and phagocytic capacities. Intriguingly, inoculation of control monocytes togetherwith Lewis lung carcinoma (LLC) cells into Hpa-KO mice resulted in nearly complete inhibition of tumor growth. In striking contrast, inoculating LLC cells together with monocytes isolated from Hpa-KO mice did not affect tumor growth, indicating that heparanase is critically required for activation and function of macrophages. Mechanistically, we describe a linear cascade by which heparanase activates Erk, p38, and JNK signaling in macrophages, leading to increased c-Fos levels and induction of cytokine expression in a manner that apparently does not require heparanase enzymatic activity. These results identify heparanase as a key mediator of macrophage activation and function in tumorigenesis and cross-talk with the tumor microenvironment. [ABSTRACT FROM AUTHOR]

• Hpa2 attenuates sarcoma tumor growth. • Tumor growth inhibition by Hpa2 involves increased stress conditions (ER stress, hypoxia, JNK phosphorylation), induction of p63, and attenuation of stemness. • In leiomyosarcoma biopsies, nuclear localization of Hpa2 was associated with low stage tumors. This finding opens a new direction in Hpa2 research. The pro-tumorigenic properties of heparanase are well documented and established. In contrast, the role of heparanase 2 (Hpa2), a close homolog of heparanase, in cancer is not entirely clear. In carcinomas, Hpa2 is thought to attenuate tumor growth, possibly by inhibiting heparanase enzymatic activity. Here, we examine the role of Hpa2 in sarcoma, a group of rare tumors of mesenchymal origin, accounting for approximately 1% of all malignant tumors. Consistently, we found that overexpression of Hpa2 attenuates tumor growth while Hpa2 gene silencing results in bigger tumors. Mechanistically, attenuation of tumor growth by Hpa2 was associated with increased tumor stress conditions, involving ER stress, hypoxia, and JNK phosphorylation, leading to increased apoptotic cell death. In addition, overexpression of Hpa2 induces the expression of the p53 family member, p63 which, in sarcoma, functions to attenuate tumor growth. Moreover, we show that Hpa2 profoundly reduces stem cell characteristics of the sarcoma cells (stemness), most evident by failure of Hpa2 cells to grow as spheroids typical of cancer stem cells. Likewise, expression of CD44, a well-established cancer stem cell marker, was prominently decreased in Hpa2 cells. CD44 is also a cell surface receptor for hyaluronic acid (HA), a nonsulfated glycosaminoglycan that is enriched in connective tissues. Reduced expression of CD44 by Hpa2 may thus represent impaired cross-talk between Hpa2 and the extracellular matrix. Clinically, we found that Hpa2 is expressed by leiomyosarcoma tumor biopsies. Interestingly, nuclear localization of Hpa2 was associated with low-stage tumors. This finding opens a new direction in Hpa2 research. [ABSTRACT FROM AUTHOR]

Abstract: Heparanase is an endoglycosidase which cleaves heparan sulfate (HS) and hence participates in degradation and remodeling of the extracellular matrix (ECM). The enzyme also releases angiogenic factors from the ECM and thereby induces an angiogenic response in vivo. Heparanase is preferentially expressed in human tumors and its over-expression in tumor cells confers an accelerated growth and invasive phenotype in experimental animals. In contrast, heparanase gene silencing is associated with a marked inhibition of tumor progression. Heparanase upregulation correlates with increased tumor vascularity and poor postoperative survival of cancer patients. Studies on relationships between structure and the heparanase-inhibiting activity of nonanticogulant heparins systematically differing in their O-sulfation patterns, degrees of N-acetylation, and glycol-splitting of nonsulfated uronic acid residues, have permitted to select effective inhibitors of the enzymatic activity of heparanase. N-acetylated, glycol-split heparins emerged as highly effective and specific inhibitors of heparanase and tumor growth and metastasis. Several observations support the involvement of heparanase in haemostasis. A marked induction of tissue factor (TF) was noted in response to heparanase over-expression in tumor-derived cell lines and heparanase over-expressing transgenic mice. A direct correlation was also found between heparanase and TF expression levels in leukemia patients. TF induction was even more pronounced upon exogenous addition of heparanase to primary endothelial cells that do not normally express TF, and this induction was associated with enhanced coagulation. These and other results indicate that pro-heparanase is rapidly tethered on cell surfaces, partially depending on cell surface heparan sulfate, generating a local procoagulant effect. In addition, pro-heparanase can reverse the anti-coagulant effect of unfractionated heparin and the Factor Xa inhibitory activity of low molecular weight heparin (LMWH). These effects were also demonstrated in plasma derived from patients treated with LMWH. The pro-coagulant effects of pro-heparanase were also exerted by a peptide corresponding to its major functional heparin-binding domain. Heparanase pro-coagulant activities suggest its possible role as a natural regulator of heparinoid anti-coagulant activities, and point to a possible use of this molecule or its heparin binding domain as antidote for heparinoid therapies. [Copyright &y& Elsevier]

In this study, superparamagnetic iron oxide nanoparticles (SPIONs) were engineered with an organic coating composed of low molecular weight heparin (LMWH) and bovine serum albumin (BSA), providing heparin-based nanoparticle systems (LMWH@SPIONs). The purpose was to merge the properties of the heparin skeleton and an inorganic core to build up a targeted theranostic nanosystem, which was eventually enhanced by loading a chemotherapeutic agent. Iron oxide cores were prepared via the co-precipitation of iron salts in an alkaline environment and oleic acid (OA) capping. Dopamine (DA) was covalently linked to BSA and LMWH by amide linkages via carbodiimide coupling. The following ligand exchange reaction between the DA-BSA/DA-LMWH and OA was conducted in a biphasic system composed of water and hexane, affording LMWH@SPIONs stabilized in water by polystyrene sulfonate (PSS). Their size and morphology were investigated via dynamic light scattering (DLS) and transmission electron microscopy (TEM), respectively. The LMWH@SPIONs' cytotoxicity was tested, showing marginal or no toxicity for samples prepared with PSS at concentrations of 50 µg/mL. Their inhibitory activity on the heparanase enzyme was measured, showing an effective inhibition at concentrations comparable to G4000 (N-desulfo-N-acetyl heparin, a non-anticoagulant and antiheparanase heparin derivative; Roneparstat). The LMWH@SPION encapsulation of paclitaxel (PTX) enhanced the antitumor effect of this chemotherapeutic on breast cancer cells, likely due to an improved internalization of the nanoformulated drug with respect to the free molecule. Lastly, time-domain NMR (TD-NMR) experiments were conducted on LMWH@SPIONs obtaining relaxivity values within the same order of magnitude as currently used commercial contrast agents. [ABSTRACT FROM AUTHOR]

The kidney glomerular filtration barrier (GFB) is enriched with heparan sulfate (HS) proteoglycans, which contribute to its permselectivity. The endoglycosidase heparanase cleaves HS and hence appears to be involved in the pathogenesis of kidney injury and glomerulonephritis. We have recently reported, nonetheless, that heparanase overexpression preserved glomerular structure and kidney function in an experimental model of Adriamycin-induced nephropathy. To elucidate mechanisms underlying heparanase function in podocytes—key GFB cells, we utilized a human podocyte cell line and transgenic mice overexpressing heparanase. Notably, podocytes overexpressing heparanase (H) demonstrated significantly higher survival rates and viability after exposure to Adriamycin or hydrogen peroxide, compared with mock-infected (V) podocytes. Immunofluorescence staining of kidney cryo-sections and cultured H and V podocytes as well as immunoblotting of proteins extracted from cultured cells, revealed that exposure to toxic injury resulted in a significant increase in autophagic flux in H podocytes, which was reversed by the heparanase inhibitor, Roneparstat (SST0001). Heparanase overexpression was also associated with substantial transcriptional upregulation of autophagy genes BCN1, ATG5, and ATG12, following Adriamycin treatment. Moreover, cleaved caspase-3 was attenuated in H podocytes exposed to Adriamycin, indicating lower apoptotic cell death in H vs. V podocytes. Collectively, these findings suggest that in podocytes, elevated levels of heparanase promote cytoprotection. [ABSTRACT FROM AUTHOR]

Breast cancer (BC) and obesity are two heterogeneous conditions with a tremendous impact on health. BC is the most commonly diagnosed neoplasm and the leading cause of cancer-related mortality among women, and the prevalence of obesity in women worldwide reaches pandemic proportions. Obesity is a significant risk factor for both incidence and worse prognosis in estrogen receptor positive (ER+) BC. Yet, the mechanisms underlying the association between excess adiposity and increased risk/therapy resistance/poorer outcome of ER+, but not ER−negative (ER−), BC are not fully understood. Tumor-promoting action of obesity, predominantly in ER + BC patients, is often attributed to the augmented production of estrogen in 'obese' adipose tissue. However, in addition to the estrogen production, expression levels of ER represent a key determinant in hormone-driven breast tumorigenesis and therapy response. Here, utilizing in vitro and in vivo models of BC, we show that macrophages, whose adverse activation by obesogenic substances is fueled by heparanase (extracellular matrix-degrading enzyme), are capable of upregulating ER expression in tumor cells, in the setting of obesity-associated BC. These findings underscore a previously unknown mechanism through which interplay between cellular/extracellular elements of obesity-associated BC microenvironment influences estrogen sensitivity—a critical component in hormone-related cancer progression and resistance to therapy. [ABSTRACT FROM AUTHOR]

Heparanase is elevated in various pathological conditions, primarily cancer and inflammation. To investigate the significance and involvement of heparanase in liver fibrosis, we compared the susceptibility of wild-type (WT) and heparanase-overexpressing transgenic (Hpa-tg) mice to carbon tetrachloride (CCL4)-induced fibrosis. In comparison with WT mice, Hpa-tg mice displayed a severe degree of tissue damage and fibrosis, including higher necrotic tendency and intensified expression of smooth muscle actin. While damage to the WT liver started to recover after the acute phase, damage to the Hpa-tg liver was persistent. Recovery was attributed, in part, to heparanase-stimulated autophagic activity in response to CCL4, leading to increased apoptosis and necrosis. The total number of stellate cells was significantly higher in the Hpa-tg than the WT liver, likely contributing to the increased amounts of lipid droplets and smooth muscle actin. Our results support the notion that heparanase enhances inflammatory responses, and hence may serve as a target for the treatment of liver damage and fibrosis. [ABSTRACT FROM AUTHOR]

Heparin is a polysaccharide expressed in animal connective tissue-type mast cells. Owing to the special pentasaccharide sequence, heparin specifically binds to antithrombin (AT) and increases the inhibitory activity of AT towards coagulation enzymes. Heparin isolated from porcine intestinal mucosa has an average molecular weight of 15 kDa, while heparins recovered from rat skin and the peritoneal cavity were 60–100 kDa and can be fragmented by the endo-glucuronidase heparanase in vitro. In this study, we have examined heparin isolated from in vitro matured fetal skin mast cells (FSMC) and peritoneal cavity mast cells (PCMC) collected from wildtype (WT), heparanase knockout (Hpa-KO), and heparanase overexpressing (Hpa-tg) mice. The metabolically 35S-labeled heparin products from the mast cells of WT, Hpa-KO, and Hpa-tg mice were compared and analyzed for molecular size and AT-binding activity. The results show that PCMC produced heparins with a size similar to heparin from porcine intestinal mast cells, whilst FSMC produced much longer chains. As expected, heparanase overexpression resulted in the generation of smaller fragments in both cell types, while heparins recovered from heparanase knockout cells were slightly longer than heparin from WT cells. Unexpectedly, we found that heparanase expression affected the production of total glycosaminoglycans (GAGs) and the proportion between heparin and other GAGs but essentially had no effect on heparin catabolism. [ABSTRACT FROM AUTHOR]

Heparanase activity plays a decisive role in cell dissemination associated with cancer metastasis. Cellular uptake of heparanase is considered a pre-requisite for the delivery of latent 65-kDa heparanase to lysosomes and its subsequent proteolytic processing and activation into 8- and 50-kDa protein subunits by cathepsin L. Heparan sulfate proteoglycans, and particularly syndecan, are instrumental for heparanase uptake and activation, through a process that has been shown to occur independent of rafts. Nevertheless, the molecular mechanism underlying syndecan-mediated internalization outside of rafts is unclear. Here, we examined the role of syndecan-1 cytoplasmic domain in heparanase processing, utilizing deletion constructs lacking the entire cytoplasmic domain (Delta), the conserved (C1 or C2), or variable (V) regions. Heparanase processing was markedly increased following syndecan-1 over-expression; in contrast, heparanase was retained at the cell membrane and its processing was impaired in cells over-expressing syndecan-1 deleted for the entire cytoplasmic tail. We have next revealed that conserved domain 2 (C2) and variable (V) regions of syndecan-1 cytoplasmic tail mediate heparanase processing. Furthermore, we found that syntenin, known to interact with syndecan C2 domain, and α actinin are essential for heparanase processing. [ABSTRACT FROM AUTHOR]

• Expression of Hpa2 is markedly induced by stress. • Hpa2 induction by stress is mediated by ATF3. • High levels of Hpa2 and ATF3 are associated with longer survival of cancer patients. Activity of heparanase, endoglycosidase that cleaves heparan sulfate side chains in heparan sulfate proteoglycans, is highly implicated in tumor progression and metastasis. Heparanase inhibitors are therefore being evaluated clinically as anti-cancer therapeutics. Heparanase 2 (Hpa2) is a close homolog of heparanase that lacks HS-degrading activity and functions as an endogenous inhibitor of heparanase. As a result, Hpa2 appears to attenuate tumor growth but mechanisms that regulate Hpa2 expression and determine the ratio between heparanase and Hpa2 are largely unknown. We have recently reported that the expression of Hpa2 is induced by endoplasmic reticulum (ER) and proteotoxic stresses, but the mechanism(s) underlying Hpa2 gene regulation was obscure. Here we expand the notion that Hpa2 is regulated by conditions of stress. We report that while ER and hypoxia, each alone, resulted in a 3-7 fold increase in Hpa2 expression, combining ER stress and hypoxia resulted in a noticeable, over 40-fold increase in Hpa2 expression. A prominent induction of Hpa2 expression was also quantified in cells exposed to heat shock, proteotoxic stress, lysosomal stress, and chemotherapy (cisplatin), strongly implying that Hpa2 is regulated by conditions of stress. Furthermore, analyses of the Hpa2 gene promoter led to the identification of activating-transcription-factor 3 (ATF3) as a transcription factor that mediates Hpa2 induction by stress, thus revealing, for the first time, a molecular mechanism that underlies Hpa2 gene regulation. Induction of Hpa2 and ATF3 by conditions of stress that often accompany the rapid expansion of tumors is likely translated to improved survival of cancer patients. [ABSTRACT FROM AUTHOR]

Heparanase is an endo‐β‐glycosidase that cleaves heparan sulfate chains at the level of the glycosidic bond of selected glucuronic acid residues. Heparin fragments, that can compete with heparan sulfate for binding to heparanase, have been reported as weak heparanase inhibitors. To reinforce their inhibitory activity, a known β‐glucuronidase inhibitor has been introduced at their reducing end to directly interfere with the active site of the enzyme. We report here the synthesis of such oligosaccharides, displaying D‐glucaro‐δ‐lactam at their reducing end. Thanks to the trichloroacetimidate glycosylation method, we could devise experimental conditions allowing O‐glycosylation of the lactam ring, and we thus chemically prepared trisaccharides that were further elongated and sulfated using enzymatic methods. Heparanase inhibitory activity was not significantly improved by the presence of the D‐glucaro‐δ‐lactam ring. [ABSTRACT FROM AUTHOR]

Heparanase functions as a heparan sulfate-degrading enzyme and as a ligand for an unidentified signaling receptor(s). Here, several reactions involved in the activation of the PI3K-AKT pathway by latent heparanase were characterized. Protein suppression using specific siRNAs revealed that heparanase-induced phosphorylation of AKT at Ser-473 was RICTOR-mTOR-dependent, whereas ILK and PAK1/2 were dispensable. p110α was the PI3K catalytic isoform preferred by heparanase for AKT activation and cell proliferation because the p110α inhibitor YM024 blocked these processes. Heparanase-induced AKT phosphorylation was low in mouse embryonic fibroblast cells expressing a RAS interaction-defective p110α compared with wild type cells, indicating that RAS has an important role in the PI3K-AKT activation. The response to heparanase was also inefficient in suspension cultures of several cell lines, suggesting a requirement of integrins in this pathway. Adhesion via either αVβ3 or α5β1 promoted heparanase-induced AKT phosphorylation, and a stronger effect was seen when both integrins were engaged. Simultaneous inhibition of FAK and PYK2 using a chemical inhibitor, or suppression of their expression, inhibited heparanase-induced AKT activation and cell proliferation. Stimulation of cells with heparanase enhanced their resistance against oxidative stress- or growth factor starvation-induced apoptosis. These results demonstrate that there is an intimate cross-talk between the heparanase receptor(s) and integrins during induction of the prosurvival PI3K-AKT pathway by heparanase. [ABSTRACT FROM AUTHOR]

Abstract: Heparanase is an endo-β-glucuronidase that cleaves heparan sulfate side chains presumably at sites of low sulfation, activity that is strongly implicated with cell invasion associated with cancer metastasis, a consequence of structural modification that loosens the extracellular matrix barrier. In addition, heparanase exerts pro-adhesive properties, mediated by clustering of membrane heparan sulfate proteoglycans (i.e., syndecans) and activation of signaling molecules such as Akt, Src, EGFR, and Rac in a heparan sulfate-dependent and -independent manner. Activation of signaling cascades by enzymatically inactive heparanase and by a peptide corresponding to its substrate binding domain not only increases cell adhesion but also facilitates cancer cell growth. This notion is supported by preclinical and clinical settings, encouraging the development of anti-heparanase therapeutics. Here, we summarize recent progress in heparanase research emphasizing the molecular mechanisms that govern its pro-tumorigenic and pro-adhesive properties. Pro-adhesive properties of the heparanase homolog, heparanase 2 (Hpa2), are also discussed. Enzymatic activity-independent function of proteases (i.e., matrix metalloproteinases) is discussed in the context of cell adhesion and tumor progression. Collectively, these examples suggest that enzyme function exceeds beyond the enzymatic aspect, thus significantly expanding the scope of the functional proteome. Cross-talk with matrix metalloproteinases and the role of heparanase in pathological settings other than cancer are also described. [ABSTRACT FROM AUTHOR]

Heparanase is a mammalian endo-β-D-glucuronidase capable of cleaving HS side chains at a limited number of sites, activity that is strongly implicated in tumor metastasis, neovascularization, inflammation, and autoimmunity. Clinically, up-regulation of heparanase mRNA and protein expression has been documented in a variety of primary human tumors, correlating with reduced postoperative survival and increased lymph node and distant metastasis, thus providing strong clinical support for the prometastatic feature of the enzyme and making it an attractive target for the development of anticancer and anti-inflammatory drugs. Screening a panel of monoclonal antibodies for their ability to inhibit heparanase enzymatic activity, we noted that one hybridoma, 6F8, exhibited the opposite effect and significantly enhanced heparanase activity. Here, we provide evidence that antibody 6F8 enhances the activity of recombinant and cellular heparanase, facilitates invasion of tumor-derived cells in vitro, and improves wound healing in a mouse punch model in vivo. These results support a role of heparanase in the course of wound healing and, moreover, suggest that monoclonal antibodies can be applied clinically for the enhancement, rather than inhibition, of certain enzymes. [ABSTRACT FROM AUTHOR]

Abstract: The endoglycosidase heparanase is the predominant enzyme that degrades heparan sulfate side chains of heparan sulfate proteoglycans, activity that is strongly implicated in tumor metastasis. Apart of its well characterized enzymatic activity, heparanase was noted to exert also enzymatic-independent functions. Among these is the induction of Akt/PKB phosphorylation noted in endothelial- and tumor-derived cells. Protein domains of heparanase required for signaling were not identified to date, nor were identified heparanase binding proteins/receptors capable of transmitting heparanase signals. Here, we examined the possible function of mannose 6-phosphate receptor (MPR) and low-density lipoprotein-receptor related protein (LRP), recently implicated in cellular uptake of heparanase, as heparanase receptors mediating Akt phosphorylation. We found that heparanase addition to MPR- and LRP-deficient fibroblasts elicited Akt activation indistinguishable from control fibroblasts. In contrast, disruption of lipid rafts abrogated Akt/PKB phosphorylation following heparanase addition. These results suggest that lipid raft-resident receptor mediates heparanase signaling. [Copyright &y& Elsevier]

Abstract: Heparanase is an endoglycosidase which cleaves heparan sulfate (HS) and hence participates in degradation and remodeling of the extracellular matrix (ECM). Heparanase is preferentially expressed in human tumors and its over-expression in tumor cells confers an invasive phenotype in experimental animals. The enzyme also releases angiogenic factors from the ECM and thereby induces an angiogenic response in vivo. Heparanase upregulation correlates with increased tumor vascularity and poor post-operative survival of cancer patients. Heparanase is synthesized as a 65kDa inactive precursor that undergoes proteolytic cleavage, yielding 8 and 50kDa protein subunits that heterodimerize to form an active enzyme. Human heparanase is localized primarily within late endosomes and lysosomes and occasionally on the cell surface and within the cell nucleus. Transcriptional activity of the heparanase promoter is stimulated by demethylation, early growth response 1 (EGR1) transcription factor, estrogen, inflammatory cytokines and inactivation of p53. N-acetylated glycol-split species of heparin as well as siRNA heparanase gene silencing inhibit tumor metastasis and angiogenesis in experimental models. These observations and the unexpected identification of a single functional heparanase, suggest that the enzyme is a promising target for anti-cancer and anti-inflammatory drug development. Heparanase exhibits also non-enzymatic activities, independent of its involvement in ECM degradation and changes in the extracellular microenvironment. For example, cell surface expression of heparanase elicits a firm cell adhesion, reflecting an involvement in cell–ECM interaction. Heparanase enhances Akt signaling and stimulates PI3K- and p38-dependent endothelial cell migration and invasion. It also promotes VEGF expression via the Src pathway. The enzyme may thus activate endothelial cells and elicits angiogenic and survival responses. Studies with heparanase over-expressing transgenic mice revealed that the enzyme functions in normal processes involving cell mobilization, HS turnover, tissue vascularization and remodeling. In this review, we summarize the current status of heparanase research, emphasizing molecular and cellular aspects of the enzyme, including its mode of processing and activation, control of heparanase gene expression, enzymatic and non-enzymatic functions, and causal involvement in cancer metastasis and angiogenesis. We also discuss clinical aspects and trategies for the development of heparanase inhibitors. [Copyright &y& Elsevier]

Abstract: Heparanase is an endo-β-glucuronodase involved in cleavage of heparan sulfate side chains, activity that is strongly implicated in cell dissemination associated with tumor metastasis and inflammation. Heparanase is first synthesized as a latent 65kDa precursor that is converted into an active enzyme upon proteolytic processing. Previously, we have reported that elevation of the lysosomal pH results in complete inhibition of heparanase processing, suggesting that lysosomal protease(s) and acidic pH conditions are required for heparanase processing. Here, we adopted a cell fractionation approach and provide evidence that incubation of the pro-enzyme with lysosome/endosome, but not with cytoplasmic fractions resulted in processing and activation of the 65kDa latent heparanase. Moreover, while the water soluble lysosome/endosome fraction exhibited no apparent processing activity, heparanase processing by the water insoluble lysosome/endosome membrane fraction was readily detected and exhibited the expected pH dependency. [Copyright &y& Elsevier]

Tumor extracellular matrix (ECM) operates in a coordinated mode with cancer and stroma cells to evoke the multistep process of metastatic potential. The remodeled tumor-associated matrix provides a point for direct or complementary therapeutic targeting. Here, we cover and critically address the importance of ECM networks and their macromolecules in cancer. We focus on the roles of key structural and functional ECM components, and their degradation enzymes and extracellular vesicles, aiming at improving our understanding of the mechanisms contributing to tumor initiation, growth, and dissemination, and discuss potential new approaches for ECM-based therapeutic targeting and diagnosis. Modified extracellular matrix in the tumor microenvironment dictates cancer cell properties, metastatic potential, and formation of premetastatic niches. Proteoglycans and hyaluronan in provisional matrix modulate the bioavailability of matrix-bound effectors, cancer cell signaling, angiogenesis, and tumor spread. The outside-in interactions of integrins with tumor matrix, the migratory tracks for cancer cells created by fibrillar collagen fiber alignment, and the intense trafficking of extracellular vesicles, impact tumor progression. Proteolytic (matrix metalloproteinases and cathepsins) and glycosidic (heparanase and hyaluronidases) enzymes mediate protumorigenic interactions and stemness, associated with therapeutic resistance. Heparanase promotes the aggressive phenotype of cancer, while heparanase-2 functions as a tumor suppressor. Interference with exosome biogenesis, miRNAs functions, provisional matrix and premetastatic niche formation, may drive future cancer therapeutics. [ABSTRACT FROM AUTHOR]

We report that gastric cancer patients exhibiting high levels of heparanase 2 (Hpa2) survive longer. Similarly, mice administrated with gastric carcinoma cells engineered to overexpress Hpa2 produced smaller tumors and survived longer than mice administrated with control cells. These beneficial effects were found to associate with increased phosphorylation of AMP-activated protein kinase (AMPK) that play an instrumental role in cell metabolism and is situated at the center of a tumor suppressor network. We also found that MG132, an inhibitor of the proteasome that results in proteotoxic stress, prominently enhances Hpa2 expression. Notably, Hpa2 induction by MG132 appeared to be mediated by AMPK, thus establishing a loop that feeds itself where Hpa2 enhances AMPK phosphorylation that, in turn, induces Hpa2 expression, possibly leading to attenuation of gastric tumorigenesis. Heparanase is highly implicated in tumor metastasis due to its capacity to cleave heparan sulfate and, consequently, remodel the extracellular matrix underlying epithelial and endothelial cells. In striking contrast, only little attention was given to its close homolog, heparanase 2 (Hpa2), possibly because it lacks heparan sulfate-degrading activity typical of heparanase. We subjected sections of gastric carcinoma to immunostaining and correlated Hpa2 immunoreactivity with clinical records, including tumor grade, stage and patients' status. We over-expressed Hpa2 in gastric carcinoma cell lines and examined their tumorigenic properties in vitro and in vivo. We also evaluated the expression of Hpa2 by gastric carcinoma cells following inhibition of the proteasome, leading to proteotoxic stress, and the resulting signaling responsible for Hpa2 gene regulation. Here, we report that gastric cancer patients exhibiting high levels of Hpa2 survive longer. Similarly, mice administrated with gastric carcinoma cells engineered to over-express Hpa2 produced smaller tumors and survived longer than mice administrated with control cells. This was associated with increased phosphorylation of AMP-activated protein kinase (AMPK), a kinase that is situated at the center of a tumor suppressor network. We also found that MG132, an inhibitor of the proteasome that results in proteotoxic stress, prominently enhances Hpa2 expression. Notably, Hpa2 induction by MG132 appeared to be mediated by AMPK, and AMPK was found to induce the expression of Hpa2, thus establishing a loop that feeds itself where Hpa2 enhances AMPK phosphorylation that, in turn, induces Hpa2 expression, leading to attenuation of gastric tumorigenesis. These results indicate that high levels of Hpa2 in some tumors are due to stress conditions that tumors often experience due to their high rates of cell proliferation and high metabolic demands. This increase in Hpa2 levels by the stressed tumors appears critically important for patient outcomes. [ABSTRACT FROM AUTHOR]

Defective amyloid-β (Aβ) clearance from the brain is a major contributing factor to the pathophysiology of Alzheimer's disease (AD). Aβ clearance is mediated by macrophages, enzymatic degradation, perivascular drainage along the vascular basement membrane (VBM) and transcytosis across the blood–brain barrier (BBB). AD pathology is typically associated with cerebral amyloid angiopathy due to perivascular accumulation of Aβ. Heparan sulfate (HS) is an important component of the VBM, thought to fulfill multiple roles in AD pathology. We previously showed that macrophage-mediated clearance of intracortically injected Aβ was impaired in the brains of transgenic mice overexpressing heparanase (Hpa-tg). This study revealed that perivascular drainage was impeded in the Hpa-tg brain, evidenced by perivascular accumulation of the injected Aβ in the thalamus of Hpa-tg mice. Furthermore, endogenous Aβ accumulated at the perivasculature of Hpa-tg thalamus, but not in control thalamus. This perivascular clearance defect was confirmed following intracortical injection of dextran that was largely retained in the perivasculature of Hpa-tg brains, compared to control brains. Hpa-tg brains presented with thicker VBMs and swollen perivascular astrocyte endfeet, as well as elevated expression of the BBB-associated water-pump protein aquaporin 4 (AQP4). Elevated levels of both heparanase and AQP4 were also detected in human AD brain. These findings indicate that elevated heparanase levels alter the organization and composition of the BBB, likely through increased fragmentation of BBB-associated HS, resulting in defective perivascular drainage. This defect contributes to perivascular accumulation of Aβ in the Hpa-tg brain, highlighting a potential role for heparanase in the pathogenesis of AD. [ABSTRACT FROM AUTHOR]

• We report that Hpa2 attenuates head and neck tumor growth, associating with induction of Sox2 expression. • We provide evidence that silencing of Sox2 resulted in bigger tumors endowed with reduced cytokeratin levels, whereas smaller tumors were developed by cells overexpressing Sox2, suggesting that in head and neck carcinoma, Sox2 functions to inhibit tumor growth. • Hpa2-null cells engineered by Crispr/Cas 9, produced bigger tumors vs control cells, and rescue of Hpa2 attenuated tumor growth. • These results strongly imply that Hpa2 functions as a tumor suppressor in head and neck cancer, involving Sox2 upregulation mediated, in part, by the high-affinity interaction of Hpa2 with heparan sulfate. The pro-tumorigenic properties of heparanase are well documented, and heparanase inhibitors are being evaluated clinically as anti-cancer therapeutics. In contrast, the role of heparanase 2 (Hpa2), a close homolog of heparanase, in cancer is largely unknown. Previously, we have reported that in head and neck cancer, high levels of Hpa2 are associated with prolonged patient survival and decreased tumor cell dissemination to regional lymph nodes, suggesting that Hpa2 functions to restrain tumorigenesis. Also, patients with high levels of Hpa2 were diagnosed as low grade and exhibited increased expression of cytokeratins, an indication that Hpa2 promotes or maintains epithelial cell differentiation and identity. To reveal the molecular mechanism underlying the tumor suppressor properties of Hpa2, and its ability to induce the expression of cytokeratin, we employed overexpression as well as gene editing (Crispr) approaches, combined with gene array and RNAseq methodologies. At the top of the list of many genes found to be affected by Hpa2 was Sox2. Here we provide evidence that silencing of Sox2 resulted in bigger tumors endowed with reduced cytokeratin levels, whereas smaller tumors were developed by cells overexpressing Sox2, suggesting that in head and neck carcinoma, Sox2 functions to inhibit tumor growth. Notably, Hpa2-null cells engineered by Crispr/Cas 9, produced bigger tumors vs control cells, and rescue of Hpa2 attenuated tumor growth. These results strongly imply that Hpa2 functions as a tumor suppressor in head and neck cancer, involving Sox2 upregulation mediated, in part, by the high-affinity interaction of Hpa2 with heparan sulfate. [ABSTRACT FROM AUTHOR]

• Our findings indicate that Hpa2 attenuates pancreatic tumor growth and highlight a feedback mechanism by which Hpa2 enhances ER stress which, in turn, induces Hpa2 expression. This results in consistent and severe ER stress, leading to increased apoptotic cell death and attenuated tumor growth. Thus, compounds designed to elicit ER stress may turn beneficial therapeutics in pancreatic cancer. • Hpa2 attenuates pancreatic tumor growth by exerting a feedback mechanism where Hpa2 enhances ER stress that, in turn, induces Hpa2 expression. • This results in consistent and severe ER stress, leading to increased apoptotic cell death and attenuated tumor growth. • Compounds designed to elicit ER stress may turn beneficial therapeutics in pancreatic cancer. While the pro-tumorigenic properties of the ECM-degrading heparanase enzyme are well documented, the role of its close homolog, heparanase 2 (Hpa2), in cancer is largely unknown. We examined the role of Hpa2 in pancreatic cancer, a malignancy characterized by a dense fibrotic ECM associated with poor response to treatment and bad prognosis. We show that pancreatic ductal adenocarcinoma (PDAC) patients that exhibit high levels of Hpa2 survive longer than patients with low levels of Hpa2. Strikingly, overexpression of Hpa2 in pancreatic carcinoma cells resulted in a most prominent decrease in the growth of tumors implanted orthotopically and intraperitoneally, whereas Hpa2 silencing resulted in bigger tumors. We further found that Hpa2 enhances endoplasmic reticulum (ER) stress response and renders cells more sensitive to external stress, associating with increased apoptosis. Interestingly, we observed that ER stress induces the expression of Hpa2, thus establishing a feedback loop by which Hpa2 enhances ER stress that, in turn, induces Hpa2 expression. This leads to increased apoptosis and attenuated tumor growth. Altogether, Hpa2 emerges as a powerful tumor suppressor in pancreatic cancer. [ABSTRACT FROM AUTHOR]

Further, factor VIII (FVIII), a potent and key factor in the coagulation process, is greatly increased in ICU patients with COVID-19.16 In this setting, it has been proposed that the systemic hyperinflammation observed in severe COVID-19 is comparable to a cytokine release syndrome (CRS), or cytokine storm. The coronavirus disease 2019 (COVID-19) pandemic, a viral illness caused by the severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2),1 has produced at the time of this writing nearly 33 million cases of infection, with over a million deaths in 235 countries,2 causing an unprecedented burden on healthcare systems and a severe global socioeconomic crisis. COVID-19-induced endotheliitis: emerging evidence and possible therapeutic strategies Most reported cases are asymptomatic or present with mild symptoms; however, 7-26% of hospitalised patients experience severe disease, often requiring admission to intensive care units (ICUs), with progressive multiple organ dysfunction and high mortality.3-5 Such differences in clinical outcomes have led physicians to initiate diverse pharmacological therapies at various stages of the disease, generating challenges as to the most appropriate therapeutic choice for COVID-19. [Extracted from the article]

Heparanase is the predominant enzyme that cleaves heparan sulfate, the main polysaccharide in the extracellular matrix. While the role of heparanase in sustaining the pathology of autoimmune diabetes is well documented, its association with metabolic syndrome/type 2 diabetes attracted less attention. Our research was undertaken to elucidate the significance of heparanase in impaired glucose metabolism in metabolic syndrome and early type 2 diabetes. Here, we report that heparanase exerts opposite effects in insulin-producing (i.e., islets) vs. insulin-target (i.e., skeletal muscle) compartments, sustaining or hampering proper regulation of glucose homeostasis depending on the site of action. We observed that the enzyme promotes macrophage infiltration into islets in a murine model of metabolic syndrome, and fosters β-cell-damaging properties of macrophages activated in vitro by components of diabetogenic/obese milieu (i.e., fatty acids). On the other hand, in skeletal muscle (prototypic insulin-target tissue), heparanase is essential to ensure insulin sensitivity. Thus, despite a deleterious effect of heparanase on macrophage infiltration in islets, the enzyme appears to have beneficial role in glucose homeostasis in metabolic syndrome. The dichotomic action of the enzyme in the maintenance of glycemic control should be taken into account when considering heparanase-targeting strategies for the treatment of diabetes. [ABSTRACT FROM AUTHOR]

Previous studies have shown that high-density lipoprotein(HDL) is mitogenic for low-density vascular endothelial cell cultures. In contrast, low-density lipoprotein (LUL) at physiological concentration is cytotoxic for this cell type [Tauber et at. (1980) J. Clin. Invest. 66, 696-709]. In order to relate these observations to the physico-chemical events occurring after the interaction of these lipoproteins with the cells, we have studied the binding, internalization, and degradation of HDL as a function of cell density and organization and have compared the results to those obtained with LDL. Cultured vascular endothelial cells bind HDL to an extent which, on a molar basis, is similar to that of LDL. Since an 80% loss of the LDL receptor sites in cells that were preexposed to LDL was not associated with a detectable decrease in HDL binding, it is likely that HDL and LDL binding sites exist on the cell surface as distinct entities. A 5-10-fold molar excess of LDL over 125I-HDL was almost as effective as a similar excess of unlabeled HDL in reducing the binding of 125I-HDL, indicating that LDL can compete with HDL for HDL- binding sites. In contrast, 125I-LDL binding and uptake were reduced by less than 20% in the presence of a 50-fold molar excess of unlabeled HDL. Both uptake and degradation of HDL were 10-20-fold lower than those of LDL. HDL degradation was only slightly or not inhibited by chloroquine at a concentration which almost fully inhibited the degradation of LDL. Confluent and highly organized endothelial cell cultures bind HDL and LDL particles to an extent which is half of the amount bound to sparse or subconlluent cultures. However, the formation of a resting endothelial cell monolayer was associated with only a twofold decrease in the uptake of HDL, as compared to a 10-20-fold decrease in the uptake and degradation of LDL. In both sparse and confluent endothelial cultures, uptake via fluid pinocytosis and the associated non-specific adsorptive endocytosis could account for most of the HDL uptake. The different pathways of internalization of HDL venus that of LDL could explain the lack of toxicity of MDL when added at high concentration to sparse and actively growing cultures. [ABSTRACT FROM AUTHOR]

On March 11, 2020, the World Health Organization declared its assessment of coronavirus disease 2019 (COVID‐19) as a global pandemic. However, specific anti‐severe acute respiratory syndrome‐coronavirus 2 (SARS‐CoV‐2) drugs are still under development, and patients are managed by multiple complementary treatments. We performed a retrospective analysis to compare and evaluate the effect of low molecular weight heparin (LMWH) treatment on disease progression. For this purpose, the clinical records and laboratory indicators were extracted from electronic medical records of 42 patients with COVID‐19 (21 of whom were treated with LMWH, and 21 without LMWH) hospitalized (Union Hospital of Huazhong University of Science and Technology) from February 1 to March 15, 2020. Changes in the percentage of lymphocytes before and after LMWH treatment were significantly different from those in the control group (P = 0.011). Likewise, changes in the levels of D‐dimer and fibrinogen degradation products in the LMWH group before and after treatment were significantly different from those in the control group (P = 0.035). Remarkably, IL‐6 levels were significantly reduced after LMWH treatment (P = 0.006), indicating that, besides other beneficial properties, LMWH may exert an anti‐inflammatory effect and attenuate in part the "cytokine storm" induced by the virus. Our results support the use of LMWH as a potential therapeutic drug for the treatment of COVID‐19, paving the way for a subsequent well‐controlled clinical study. [ABSTRACT FROM AUTHOR]

• Mice overexpressing heparanase (Hpa-tg) were applied to reveal the contribution of host heparanase to tumor growth and metastasis. • Xenograft tumors transplanted in Hpa-tg mice exhibited accelerated tumor progression and shorter survival time. • Accelerated tumor growth was attributed to increased levels of pro-tumorigenic factors in the plasma and bone marrow of the Hpa-tg mice. • Host heparanase impaired the overall anti-tumor activity of immune T-cell subpopulations. • Heparanase mediates the pro-tumorigenic crosstalk between the tumor, tumor microenvironment, immune cells and host factors. Heparanase, the sole heparan sulfate degrading endoglycosidase, regulates multiple biological activities that enhance tumor growth, angiogenesis and metastasis. Much of the impact of heparanase on tumor progression is related to its function in mediating tumor-host crosstalk, priming the tumor microenvironment to better support tumor growth and metastasis. We have utilized mice over-expressing (Hpa-tg) heparanase to reveal the role of host heparanase in tumor initiation, growth and metastasis. While in wild type mice tumor development in response to DMBA carcinogenesis was restricted to the mammary gland, Hpa-tg mice developed tumors also in their lungs and liver, associating with reduced survival of the tumor-bearing mice. Consistently, xenograft tumors (lymphoma, melanoma, lung carcinoma, pancreatic carcinoma) transplanted in Hpa-tg mice exhibited accelerated tumor growth and shorter survival of the tumor-bearing mice compared with wild type mice. Hpa-tg mice were also more prone to the development of metastases following intravenous or subcutaneous injection of tumor cells. In some models, the growth advantage was associated with infiltration of heparanase-high host cells into the tumors. However, in other models, heparanase-high host cells were not detected in the primary tumor, implying that the growth advantage in Hpa-tg mice is due to systemic factors. Indeed, we found that plasma from Hpa-tg mice enhanced tumor cell migration and invasion attributed to increased levels of pro-tumorigenic factors (i.e., RANKL, SPARC, MIP-2) in the plasma of Hpa-Tg vs. wild type mice. Furthermore, tumor aggressiveness and short survival time were demonstrated in wild type mice transplanted with bone marrow derived from Hpa-tg but not wild type mice. These results were attributed, among other factors, to upregulation of pro-tumorigenic (i.e., IL35+) and downregulation of anti-tumorigenic (i.e., IFN-γ+) T-cell subpopulations in the spleen, lymph nodes and blood of Hpa-tg vs. wild type mice and their increased infiltration into the primary tumor. Collectively, our results emphasize the significance of host heparanase in mediating the pro-tumorigenic and pro-metastatic interactions between the tumor cells and the host tumor microenvironment, immune cells and systemic factors. [ABSTRACT FROM AUTHOR]

Although cancer cells use heparanase for tumor metastasis, favourable effects of heparanase have been reported in the management of Alzheimer's disease and diabetes. Indeed, we previously established a protective function for heparanase in the acutely diabetic heart, where it conferred cardiomyocyte resistance to oxidative stress and apoptosis by provoking changes in gene expression. In this study, we tested if overexpression of heparanase can protect the heart against chemically induced or ischemia/reperfusion (I/R) injury. Transcriptomic analysis of Hep-tg hearts reveal that 240 genes related to the stress response, immune response, cell death, and development were altered in a pro-survival direction encompassing genes promoting the unfolded protein response (UPR) and autophagy, as well as those protecting against oxidative stress. The observed UPR activation was adaptive and not apoptotic, was mediated by activation of ATF6α, and when combined with mTOR inhibition, induced autophagy. Subjecting wild type (WT) mice to increasing concentrations of the ER stress inducer thapsigargin evoked a transition from adaptive to apoptotic UPR, an effect that was attenuated in Hep-tg mouse hearts. Consistent with these observations, when exposed to I/R, the infarct size and markers of apoptosis were significantly lower in the Hep-tg heart compared to WT. Finally, UPR and autophagy inhibitors reduced the protective effects of heparanase overexpression during I/R. Our data suggest that the mechanisms that underlie the role of heparanase in promoting cell survival could be uniquely beneficial to the heart by providing protection against cellular stresses, and could be useful for exploitation as a therapeutic target for the treatment of heart disease. Unlabelled Image • Cardiac overexpression of heparanase upregulated pro-survival and catabolism genes. • These gene changes contributed to activation of adaptive UPR and autophagy. • Activation of ATF6α is a key initiator of adaptive UPR. • Heparanase promotes autophagy by UPR activation and mTOR inhibition. • These adaptations protected the heart against chemical and I/R induced injury. [ABSTRACT FROM AUTHOR]

Abstract It is now well recognized that heparanase, an endo -β-D-glucuronidase capable of cleaving heparan sulfate (HS) side chains at a limited number of sites, promotes tumorigenesis by diverse mechanisms. Compelling evidence strongly implies that heparanase is a viable target for cancer therapy, thus encouraging the development of heparanase inhibitors as anti-cancer therapeutics. Here, we examined the efficacy and mode of action of PG545, an HS-mimetic heparanase inhibitor, in human lymphoma. We found that PG545 exhibits a strong anti-lymphoma effect, eliciting lymphoma cell apoptosis. Notably, this anti-lymphoma effect involves ER stress response that was accompanied by increased autophagy. The persistent ER stress evoked by PG545 is held responsible for cell apoptosis because apoptotic cell death was attenuated by an inhibitor of PERK, a molecular effector of ER stress. Importantly, PG545 had no such apoptotic effect on naïve splenocytes, further encouraging the development of this compound as anti-lymphoma drug. Surprisingly, we found that PG545 also elicits apoptosis in lymphoma cells that are devoid of heparanase activity (i.e., Raji), indicating that the drug also exerts heparanase-independent function(s) that together underlie the high potency of PG545 in preclinical cancer models. Highlights • Compelling evidence implies that heparanase is a viable target for cancer therapy. • We examined the mode of action of PG545, an HS-mimetic heparanase inhibitor. • PG545 exhibits a strong anti-lymphoma effect, eliciting lymphoma cell apoptosis. • This anti-lymphoma effect involves ER stress response accompanied by increased autophagy. • PG545 has no such effects on naïve splenocytes, supporting its development as anti-lymphoma drug. • PG545 also elicits apoptosis in lymphoma cells that are devoid of heparanase activity. [ABSTRACT FROM AUTHOR]

Vaccinia virus (VACV), like many other viruses, binds to cell surface heparan sulfate (HS) prior to infecting cells. Since HS is ubiquitously expressed extracellularly, it seemed likely that VACV-HS interaction may impede virus spread, with host heparanase, the only known mammalian endoglycosidase that can degrade HS, potentially overcoming this problem. In support of this hypothesis, we found that, compared to wild type, mice deficient in heparanase showed a 1–3 days delay in the spread of VACV to distant organs, such as ovaries, following intranasal inoculation, or to ovaries and spleen following intramuscular inoculation. These delays in spread occurred despite heparanase deficiency having no effect on VACV replication at inoculation sites. Subsequent in vitro studies revealed that heparanase treatment released VACV from HS expressing, but not HS deficient, infected cell monolayers. Collectively these data suggest that VACV relies on host heparanase to degrade HS in order to spread to distant sites. • Poxviruses, like several other viruses, use cell surface heparan sulfate (HS) as a co-receptor during infection. • Heparanase is the only known mammalian endoglycosidase that can degrade HS. • Spread of HS binding viruses like vaccinia virus (VACV), is aided by host derived heparanase. • Heparanase treatment releases VACV from HS expressing, but not HS deficient, cell monolayers infected with VACV. • Targetting heparanase could be a novel approach to controlling HS-dependent viral infection. [ABSTRACT FROM AUTHOR]