Tzahi Neuman

Heparanase Is Essential for the Development of Diabetic Nephropathy in Mice

Diabetic nephropathy (DN) is the major life-threatening complication of diabetes. Abnormal permselectivity of glomerular basement membrane (GBM) plays an important role in DN pathogenesis. Heparanase is the predominant enzyme that degrades heparan sulfate (HS), the main polysaccharide of the GBM. Loss of GBM HS in diabetic kidney was associated with increased glomerular expression of heparanase; however, the causal involvement of heparanase in the pathogenesis of DN has not been demonstrated. We report for the first time the essential involvement of heparanase in DN. With the use of Hpse-KO mice, we found that deletion of the heparanase gene protects diabetic mice from DN. Furthermore, by investigating the molecular mechanism underlying induction of the enzyme in DN, we found that transcription factor early growth response 1 (Egr1) is responsible for activation of heparanase promoter under diabetic conditions. The specific heparanase inhibitor SST0001 markedly decreased the extent of albuminuria and renal damage in mouse models of DN. Our results collectively underscore the crucial role of heparanase in the pathogenesis of DN and its potential as a highly relevant target for therapeutic interventions in patients with DN.

A Harmonization Study for the Use of 22C3 PD-L1 Immunohistochemical Staining on Ventana’s Platform

Introduction Immunotherapy is a novel treatment for lung cancer. Pembrolizumab (Merck Sharp and Dohme, Kenilworth, NJ) is a monoclonal antibody against programmed cell death 1 that has been approved for use with NSCLC together with a companion diagnostic by Dako (Carpinteria, CA). Ventanas BenchMark XT (Ventana Medical Systems, Tucson, AZ) is a widely used immunohistochemical (IHC) platform. However, data on its reliability and reproducibility with the 22C3 antibody are scant. Methods We performed a comprehensive calibration of 22C3 programmed cell death ligand 1 (PD‐L1) staining on the BenchMark XT platform using Dako’s prediluted 22C3 anti–PD‐L1 primary antibody with two of Ventana’s detection systems. Forty‐one random cases of NSCLC were then independently evaluated by two pathologists. Each case was scored using Dako‐ or Ventana‐stained slides. The scores obtained with the two 22C3 Ventana assays were compared with those obtained using the Dako 22C3 IHC platform. Results The Dako IHC platform stratified eight, seven, and 26 cases as being strongly positive, weakly positive, and negative for PD‐L1, respectively, whereas 36 of 41 cases (87.8%) had the same results with Ventana’s UltraView 22C3 protocol (Pearson’s correlation score 0.91, p < 0.0001). Moreover, 35 of 41 cases (85.3%) had the same results with Ventana’s OptiView 22C3 protocol (Pearson’s correlation score 0.89, p < 0.0001). Conclusions The results of this study demonstrate that the same PD‐L1 IHC algorithm can be reliably applied to Ventanas BenchMark XT platform. Furthermore, we were able to detect all of the strongly positive cases with high interobserver and intraobserver agreement by using the Ventana platform. These findings suggest that the Ventana platform can be used to stratify patients for pembrolizumab‐based immunotherapy.

The Inhibitor of Apoptosis Protein Livin (ML-IAP) Plays a Dual Role in Tumorigenicity

The inhibitor of apoptosis protein (IAP) family can inhibit apoptosis induced by a variety of stimuli. We and others previously described the IAP Livin (ML-IAP). We found that Livin is unique among the IAP members as, on a strong apoptotic stimulus, it is specifically cleaved by caspases to produce a truncated protein with paradoxical proapoptotic activity (tLivin). We also showed that Livin encodes two splicing variants, termed Livin alpha and beta, with diverse antiapoptotic effects in vitro. In this study, we compared the Livin isoforms in vivo. An animal model was established and the effects of Livin alpha and beta on the initiation and development of tumors were compared. In the animal model, Livin alpha promotes tumor initiation in comparison with control. Interestingly, the growth of tumors originating from cells expressing Livin beta was inhibited. In these tumors, Livin beta was cleaved and produced a high level of the proapoptotic tLivin beta that repressed tumor development. When we eliminated the proapoptotic effect of Livin beta by point mutations, the resulting antiapoptotic Livin beta mutants contributed to tumor progression. In terms of mechanism, we show that Livin beta tumors develop only in mice lacking natural killer (NK) cell activity. Thus, from the animal model, we can conclude that Livin plays a major role in tumorigenicity and that NK cells induce cleavage of Livin to its proapoptotic truncated protein that in turn inhibits tumor growth. Therefore, Livin and tLivin may serve as potential targets for cancer therapy.

Cytotoxic chemotherapy-induced odontalgia: a differential diagnosis for dental pain.

INTRODUCTION Peripheral neurotoxicity and neuropathic pain are well-known complications of several anti-cancer chemotherapeutic agents. Such pain might cause an impairment of the patients quality of life and is a common limiting factor of anti-cancer chemotherapy. Neurotoxicity in orofacial structures has been previously described as diffuse jaw pain or numbness. Currently, localized pulpal pain is not listed as a possible complication of cytotoxic therapy. The aim of this report was to suggest cytotoxic-induced neurotoxicity as a differential diagnosis for toothache during anti-cancer therapy. METHODS We described the diagnostic process in a patient suffering from severe pulpal pain in apparently intact teeth during cytotoxic therapy. A non-Hodgkins lymphoma patient complained of 2 episodes of excruciating dental pain evoked by mouth breathing, which caused nocturnal awakenings. RESULTS Both episodes developed immediately after administrations of cyclophosphamide as part of an anti-cancer chemotherapy protocol. Clinical parameters and radiographic characteristics eliminated other possible dental and facial etiologies. Pulp extirpation (pulpectomy) resulted in immediate pain relief. In both episodes, cytologic evaluation of the extirpated pulp tissue failed to show inflammation or an infiltration of lymphoma cells. CONCLUSIONS This case presented a circumstantial relation between the clinical presentation of dental pain, with associated significant impairment of the patients quality of life, and the timing of administrations of high-dose cyclophosphamide. It suggests that chemotherapy-induced toxicity might manifest as pulpitis-like toothache, which might present a diagnostic challenge for the dental practitioner.

NKp46 Receptor-Mediated Interferon-γ Production by Natural Killer Cells Increases Fibronectin 1 to Alter Tumor Architecture and Control Metastasis

Summary Natural killer (NK) cells are innate lymphoid cells, and their presence within human tumors correlates with better prognosis. However, the mechanisms by which NK cells control tumors in vivo are unclear. Here, we used reflectance confocal microscopy (RCM) imaging in humans and in mice to visualize tumor architecture in vivo. We demonstrated that signaling via the NK cell receptor NKp46 (human) and Ncr1 (mouse) induced interferon‐&ggr; (IFN‐&ggr;) secretion from intratumoral NK cells. NKp46‐ and Ncr1‐mediated IFN‐&ggr; production led to the increased expression of the extracellular matrix protein fibronectin 1 (FN1) in the tumors, which altered primary tumor architecture and resulted in decreased metastases formation. Injection of IFN‐&ggr; into tumor‐bearing mice or transgenic overexpression of Ncr1 in NK cells in mice resulted in decreased metastasis formation. Thus, we have defined a mechanism of NK cell‐mediated control of metastases in vivo that may help develop NK cell‐dependent cancer therapies. Graphical Abstract Figure. No Caption available. HighlightsNKp46 expression on NK cells controls growth of melanoma and carcinoma metastasesIn the absence of NKp46, tumor architectural properties indicate an aggressive phenotypeNK cell NKp46‐mediated IFN‐&ggr; production controls tumor structure via FN1 inductionIFN‐&ggr; treatment or Ncr1 overexpression in tumor‐bearing mice decreases tumor metastases &NA; NK cells defend against various pathogens and tumors, but the mechanisms by which they control tumor metastases are not clear. Here, Glasner et al. show that NK cells prevent tumor metastases in vivo by editing tumor architecture via NKp46‐mediated IFN‐&ggr; production that leads to upregulation of extracellular matrix protein FN1 in the tumor.

Asymptomatic pancreatic perivascular epithelial cell tumor (PEComa) in a male patient: report and literature review.

CONTEXT Perivascular epithelial cell tumors (PEComas) are a family of rare mesenchymal neoplasms which share cellular, immunohistochemical and ultrastructural characteristics but are found in different visceral and soft tissue sites. PEComas of the pancreas are extremely rare neoplasms. CASE REPORT We describe a 49-year-old male who was incidentally diagnosed with a pancreatic mass. Endoscopic ultrasound-guided biopsy suggested a PEComa. An uneventful pylorus-preserving pancreaticoduodenectomy was thus performed. The tumor was a solid well-circumscribed mass in the pancreatic head with dilatation of the main pancreatic duct. Histopathology revealed a well-circumscribed and vascularized neoplasm, measuring 32x27x30 mm, composed of epithelioid smooth muscle cells with clear cytoplasm rich in glycogen. The tumor exhibited immunoreactivity to alpha-smooth muscle actin and to melanoma-associated antigen HMB-45. CONCLUSIONS Although rare, pancreatic PEComas should be included in the differential diagnosis of a pancreatic mass. Currently, the paucity of cases published makes it impossible to predict the behavior and prognosis of these tumors or to advocate an optimal therapy.

Malignant Appendiceal GIST: Case Report and Review of the Literature

IntroductionGastro-intestinal stromal tumors (GISTs) of the appendix are a rare entity. To date, only a handful has been described in the literature, all of which have been of the benign type.Case ReportWe present the first reported case of a malignant appendiceal GIST. The tumor was discovered when the patient presented with a peri-appendiceal abscess which appeared suspicious on CT. The abscess was drained and managed medically. The patient responded to antibiotic treatment but subsequent CT and biopsy confirmed the diagnosis of appendiceal GIST, and the patient was started on treatment with imatinab mesylate.DiscussionOne week after initiation of therapy, the patient returned with frank peritonitis necessitating surgery. Abdominal exploration revealed an appendiceal GIST locally invading and perforating adjacent bowel. We describe the complex presentation and course of the case as well as a literature review of the appendiceal GISTs and the current approach to treatment.

Induction of heparanase by HPV E6 oncogene in head and neck squamous cell carcinoma

High‐risk human papillomavirus (HPV)‐positive head and neck squamous cell carcinomas (HNSCCs) are highly invasive; however the identity of downstream effectors responsible for their aggressive phenotype remains underinvestigated. Here, we report that HPV‐mediated up‐regulation of heparanase enzyme can provide mechanistic explanation for augmented invasiveness of HPV‐positive HNSCCs. Heparanase is the sole mammalian enzyme (endo‐β‐d‐glucuronidase) degrading heparan sulphate glycosaminoglycan, key polysaccharide of the extracellular matrix. Cleavage of heparan sulphate by heparanase leads to disassembly of extracellular barriers, enabling local invasion and metastatic spread of the tumour, and releases heparan sulphate‐bound growth factors from the extracellular depots. Heparanase is tightly implicated in head and neck cancer progression; yet, molecular mechanisms underlying transcriptional activation of the heparanase gene in HNSCC are largely unknown. We found that HPV16 oncogene E6 is capable of inducing overexpression of heparanase in HNSCC. Notably, radiation treatment dose‐dependently suppresses E6‐induced heparanase expression in vitro. Our results provide the first evidence for a functional involvement of HPV in heparanase induction in head and neck tumourigenesis and, given ongoing clinical testing of several heparanase‐inhibiting compounds, offer important avenue for future therapeutic exploration in HNSCC, as well as other HPV‐associated malignancies (i.e. cervical carcinoma).

Heparanase is preferentially expressed in human psoriatic lesions and induces development of psoriasiform skin inflammation in mice

Heparanase is the sole mammalian endoglycosidase that selectively degrades heparan sulfate, the key polysaccharide associated with the cell surface and extracellular matrix of a wide range of tissues. Extensively studied for its capacity to promote cancer progression, heparanase enzyme was recently implicated as an important determinant in several inflammatory disorders as well. Applying immunohistochemical staining, we detected preferential expression of heparanase by epidermal keratinocytes in human psoriatic lesions. To investigate the role of the enzyme in the pathogenesis of psoriasis, we utilized heparanase transgenic mice in a model of 12-O-tetradecanoyl phorbol 12-myristate 13-acetate-induced cutaneous inflammation. We report that over-expression of the enzyme promotes development of mouse skin lesions that strongly recapitulate the human disease in terms of histomorphological appearance and molecular/cellular characteristics. Importantly, heparanase of epidermal origin appears to facilitate abnormal activation of skin-infiltrating macrophages, thus generating psoriasis-like inflammation conditions, characterized by induction of STAT3, enhanced NF-κB signaling, elevated expression of TNF-α and increased vascularization. Taken together, our results reveal, for the first time, involvement of heparanase in the pathogenesis of psoriasis and highlight a role for the enzyme in facilitating abnormal interactions between immune and epithelial cell subsets of the affected skin. Heparanase inhibitors (currently under clinical testing in malignant diseases) could hence turn highly beneficial in psoriatic patients as well.

Extra domain-A fibronectin is necessary for the development of nasal remodeling in chronic allergen-induced rhinitis

BACKGROUND Extra domain A-containing fibronectin (EDA-FN) is necessary for the development of allergen-induced lower airway fibrosis. The pathogenesis of fibrosis in allergic rhinitis has not been well studied. OBJECTIVES To determine whether EDA-fibronectin is necessary for the development of nasal remodeling in a murine model of chronic allergic rhinitis and in human allergic rhinitis. METHODS EDA(-/-) and wild-type (WT) C57Bl/6 mice were sensitized intraperitoneally and then challenged with inhaled ovalbumin (OVA) or saline for 2 and 5 weeks. Clinical signs of rhinitis and histological analysis of nasal tissue were evaluated. Immunohistological staining for EDA-FN was performed in human tissue of inferior nasal conchae from patients with allergic rhinitis and controls. RESULTS After 2 weeks of allergen exposure, only goblet cell hyperplasia and perivascular eosinophilia were observed. After 5 weeks, goblet cell number, thickening of the subepithelial layer, and extent and area of collagen deposition were increased in the nasal tissue of WT OVA (ovalbumin)-challenged mice as compared with saline controls (P < .0001, P < .0001, P = .018, and P = .03, respectively). Clinical signs of rhinitis were observed only in WT OVA-challenged mice. In the EDA(-/-) mice exposed to OVA, collagen deposition, collagen area, and subepithelial thickness showed no increase and were similar to saline control mice, whereas goblet cell hyperplasia was similar to WT OVA-challenged mice. EDA-FN expression was prominent in inferior conchae from patients with allergic rhinitis but was absent in control patients. CONCLUSION EDA-containing fibronectin is necessary for the development of nasal tissue fibrotic remodeling process in both murine and human allergic rhinitis.