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Unlike the intense research effort devoted to exploring the significance of heparanase in human diseases, very little attention was given to its close homolog, heparanase 2 (Hpa2). The emerging role of Hpa2 in a rare autosomal recessive congenital disease called urofacial syndrome (UFS), clearly indicates that Hpa2 is not a pseudogene but rather a gene coding for an important protein. Hpa2 lacks the heparan sulfate (HS)-degrading activity typical of heparanase, yet exhibits high affinity to HS, affinity that is 10-fold higher than that of heparanase. The consequences of this high-affinity interaction of Hpa2 with plasma membrane HSPG has not been explored yet. Here, we used highly purified Hpa2 protein to examine this aspect. We provide evidence that cells adhere to and spread on dishes coated with Hpa2. We also show that cell migration is attenuated markedly by exogenous addition of Hpa2 to primary and transformed cells, a function that agrees with the anti-cancer properties of Hpa2. Interestingly, we found that exogenous addition of Hpa2 also disrupts the morphology of cell colonies, resulting in cell scattering. This implies that under certain conditions and experimental settings, Hpa2 may exhibit pro-tumorigenic properties. We further developed a panel of anti-Hpa2 monoclonal antibodies (mAb) and show that these properties of Hpa2 are prevented by some of the newly-developed mAb, thus providing new molecular tools to better appreciate the significance of Hpa2 in health and disease. [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]

The endoglycosidase heparanase specifically cleaves the heparan sulfate (HS) side chains on proteoglycans, an activity that has been implicated strongly in tumor metastasis and angiogenesis. Heparanase-2 (Hpa2) is a close homolog of heparanase that lacks intrinsic HS-degrading activity but retains the capacity to bind HS with high affinity. In head and neck cancer patients, Hpa2 expression was markedly elevated, correlating with prolonged time to disease recurrence and inversely correlating with tumor cell dissemination to regional lymph nodes, suggesting that Hpa2 functions as a tumor suppressor. The molecular mechanism associated with favorable prognosis following Hpa2 induction is unclear. Here we provide evidence that Hpa2 overexpression in head and neck cancer cells markedly reduces tumor growth. Restrained tumor growth was associated with a prominent decrease in tumor vascularity (blood and lymph vessels), likely due to reduced Id1 expression, a transcription factor highly implicated in VEGF-A and VEGF-C gene regulation. We also noted that tumors produced by Hpa2- overexpressing cells are abundantly decorated with stromal cells and collagen deposition, correlating with a marked increase in lysyl oxidase expression. Notably, heparanase enzymatic activity was unimpaired in cells overexpressing Hpa2, suggesting that reduced tumor growth is not caused by heparanase regulation. Moreover, growth of tumor xenografts by Hpa2-overexpressing cells was unaffected by administration of a mAb that targets the heparin-binding domain of Hpa2, implying that Hpa2 function does not rely on heparanase or heparan sulfate. Cancer Res; 76(9); 2791-801. [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]

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]