Dear Colleagues,

Heparanase is an endoglycosidase that cleaves the heparan sulfate (HS) side chains of HS proteoglycans (HSPG). Compelling evidence ties heparanase to all aspects of cancer progression, namely, tumor initiation, growth, metastasis, and chemoresistance, supporting the notion that heparanase is a master regulator of cancer aggressiveness and crosstalk with the tumor microenvironment. Heparanase upregulation has also been identified in inflammatory diseases, atherosclerosis, fibrosis, diabetes, diabetic nephropathy, and viral infection.

Unlike the intense research effort devoted to exploring the significance of heparanase in human diseases, very little attention has been given to heparanase 2 (=Hpa2), a close homolog of heparanase that lacks HS-degrading activity. Notably, Hpa2 retains the capacity to bind heparin/HS and, importantly, exhibits an even higher affinity toward HS than heparanase, thus competing for HS binding and inhibiting heparanase enzymatic activity. Hpa2 exhibits antitumor, anti-angiogenic and anti-inflammatory protective effects due in part to its heparanase-inhibiting activity, and to an elevation of endoplasmic reticulum (ER) stress that leads to apoptotic cell death. It appears that Hpa2 functions as a natural inhibitor of the heparanase enzyme and that the heparanase/Hpa2 ratio regulates to a large extent the balance between disease progression and suppression.

This Special Issue aims at exploring the multifaceted activities of heparanase and heparanase-2 in cancer and inflammation and the related clinical implications and future directions.

Prof. Dr. Israël Vlodavsky
Prof. Dr. Jin-Ping Li
Guest Editors

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• heparanase
• tumor microenvironment
• extracellular matrix
• tumor progression
• metastasis

Title: In vivo transduction of HPSE2 and LRIG2, genes implicated in urofacial syndrome, into the mouse bladder
Authors: Filipa M Lopes1, Celine Grenier1, Benjamin W Jarvis1, Adrian Lene1, Simon N Waddington2, Adrian Woolf1, Neil Roberts1
Affiliation: 1. Division of Cell Matrix Biology and Regenerative Medicine, School of Biological Sciences, The University of Manchester 2. Maternal & Fetal Medicine, EGA Institute for Women's Health, Faculty of Population Health Sciences, UCL
Abstract: Background. HPSE2, coding for heparanase 2, is a heparan sulfate binding protein with high sequence conservation with the endoglycosidase heparanase, but lacking enzymatic function. Causative pathogenic variants have been reported in HPSE2 in some patients with urofacial syndrome (UFS), a devastating early-onset bladder disease. Pathogenic variants have also been discovered in leucine-rich repeats and immunoglobulin-like domains 2 (LRIG2), in other individuals with UFS. Mouse models of both forms of UFS indicate an underlying neuropathy in the bladder. Knowing the implicated genes and the affected tissue opens the possibility of using gene therapy to cure mouse models of the disease. In this study, we aim to take the first steps to developing a treatment for UFS, by using viral vectors to deliver healthy human copies of HPSE2 and LRIG2 to neurons in the mouse bladder. Methods. Vectors contained either a nerve-specific synapsin (SYN1) promoter or a global CAG promoter to drive human HPSE2 or LRIG2 expression. Adeno-associated virus 9 (AAV9) vector was tested in N2a neuroblastoma cells. In vivo delivery into wildtype neonatal mice was achieved by superficial temporal vein injection. Transgene expression was detected using Basescope on sectioned bladders from injected and control mice, to detect either vector transduction or transgene expression. Results. Therapeutic human transgenes were successfully transduced into neuroblastoma cells by the AAV vectors. Next, vectors were delivered to wildtype mice by systemic injection. Mice tolerated vector dose escalation without loss of viability, weight and behaviour remaining normal to adulthood. Bladder body and outflow were analysed for endogenous and human transgene transcripts and the viral genome. The human transgenes were detected in mice injected with the relevant vector, and transcript abundance in bladder neurons correlated with the vector dose. At low vector dose, sparse human HPSE2 transcripts were observed in bladder neuron cell bodies. At high dose, abundant transcripts were observed in the autonomic nerve cell bodies, and also greater intensity of staining was observed in the bladder smooth muscle detrusor. Similarly, at low dose the SYN1 promoter drove low levels of human LRIG2 in bladder neurons, while at high dose abundant neuronal LRIG2 was detected. For other relevant organs, in the liver, highly abundance vector was detected, correlating with transgene expression. In the kidney, low or absent vector and transgene expression were observed. Summary. Here, we present initial findings for the first study of vector mediated gene therapy for a disease congenital that affects the lower urinary tract. We provide, to our knowledge, the first ever description of transduction and therapeutic transgene expression in autonomic nerves. We confirm that our AAV9-transgene vectors are able to safely transduce bladder neurons and drive expression of HPSE2 and LRIG2. Next, we will utilise this strategy in mutant mice to ameliorate the voiding dysfunction and causative neurophysiological defects. If successful, we aim to translate this to clinical use, and intend for our strategy will become a template for other models of genetic bladder disease. Highlights. We show, for the first time, that adeno viral vectors are able to delivery therapeutic human transgenes to autonomic nerves in the mouse bladder. This opens the possibility of using gene therapy to treat genetic diseases of the urinary tract such as urofacial syndrome, which is caused by mutations in HPSE2 and LRIG2.