Canine and Human Insulinoma : prognostic factors, druggable genes and cancer stem cells

Insulinoma (INS), which causes clinical signs associated with hypoglycaemia, is the most common pancreatic neuroendocrine tumour (pNET) of dogs and humans. Ten tot fifteen percent of human INS metastasise to regional lymph nodes and the liver, and these are referred to as ‘malignant INS’. Surgical excision of malignant INS is rarely complete and malignant INS generally respond poorly to traditional chemotherapeutic agent regimens. Therefore, recurrence is likely, leading to decreased survival times in affected individuals. In the dog, the biological course of INS resembles that of malignant INS in humans, since >95% of these canine tumours metastasise. In the dog, as in humans, surgery along with optional post-operative medical therapy, remains the recommended approach when possible. However, the prognosis for canine INS is still poor, because primary INS is frequently inoperable and/or micrometastases are missed during surgery. Therefore, novel approaches are needed to improve diagnosis, therapy and prognosis for both canine and human INS. Comparative oncology aims to utilise spontaneous tumours in pet animals as sophisticated models for the study of human cancer biology and therapy. Since the release of the canine genome in 2005, cancer in dogs has been repeatedly emphasised as an excellent model for humans. Naturally-occurring cancer in dogs and humans share similarities in histology, tumour biology, and response to conventional therapies. Regarding the close resemblance to human malignant INS, canine INS forms an interesting study model for human INS. Novel targets for human and canine INS therapy may be identified by uncovering the pathways underlying tumourigenesis of canine INS. The aims of this study were to: 1.Develop a novel surgical technique for resection of canine insulinomas and abdominal lymph node metastases; 2.Establish reliable prognostic biomarkers for canine insulinomas that facilitate optimal patient management; 3.Identify novel druggable targets in canine and human insulinomas. It was demonstrated that partial pancreatectomy using a bipolar vessel-sealing device (BVSD) was a safe and viable alternative to conventional methods of pancreatectomy for canine INS. BVSD pancreatectomy in dogs with INS significantly decreased operative and hospitalisation times and was not associated with more clinical complications than conventional pancreatectomy. Tumour size, TNM stage, stromal fibrosis and the Ki67 index proved to be useful parameters to predict clinical behaviour of canine INS. Novel druggable genes in canine and human INS were identified, using quantitative real-time PCR, and microarray analysis. Using flow cytometric analysis, athymic mice and zebrafish embryo xenografts assays, CD90 was identified as insulinoma cancer stem cell marker with the potential to serve as a druggable target

Use of a bipolar vessel-sealing device in resection of canine insulinoma

OBJECTIVES: To describe partial pancreatectomy using a bipolar vessel-sealing device (BVSD) and compare this novel technique to the conventional suture-fracture (SF) method for canine insulinoma. METHODS: Pre-, intra- and postoperative data of eight dogs with insulinoma, which underwent resection using the BVSD (LigaSure V), were prospectively collected and compared with those of eight randomly selected case-matched patients that underwent resection using the conventional SF technique. RESULTS: Mean surgical time was significantly (P=0·022) shorter in the BVSD (107 ±9 minutes) than in the SF (135 ±22 minutes) group. The BVSD technique was negatively associated with surgical time and duration of the hospitalisation period. Neither technique caused intraoperative complications, such as bleeding, collateral damage to adjacent tissues or problems with sealing or suturing the pancreatic tissue. Three dogs in the SF group and none in the BVSD group developed postoperative clinical signs associated with pancreatitis. CLINICAL SIGNIFICANCE: BVSD is a safe and viable alternative to conventional methods of pancreatectomy for canine insulinoma. It provides the possibility to remove insulinomas in the pancreatic limbs and corpus with relative ease. BVSD pancreatectomy in dogs with insulinoma significantly decreases operative and hospitalisation times and is not associated with more clinical complications than SF pancreatectom

Expression of insulin-likegrowthfactor-1 by canineinsulinomas and their metastases

Abstract The long-term prognosis after surgical resection of canineinsulinoma is poor. Signs of hypoglycaemia often recur soon after surgery because tumour tissue has only been resected partially and/or functional (micro-)metastases were present. Using quantitative real-time PCR, the expression of 16 target genes was compared between primary canineinsulinomas and their corresponding metastases. There was significantly higher expression of genes encoding for growth hormone (GH) and insulin-likegrowthfactor-1 (IGF-1) in metastases compared to their primary tumours. Immunohistochemical examination of proteins of the GH–IGF-1 axis revealed expression of GH, IGF-1 and GH receptor (GHR) in both primary insulinomas and metastases. Immunohistochemical staining for IGF-1 was significantly higher in metastases compared to primary tumours. Keywords Canine; Insulinoma; β-cell tumour; Growth hormone; Insulin-likegrowthfacto

Expression of insulin-likegrowthfactor-1 by canineinsulinomas and their metastases

Abstract The long-term prognosis after surgical resection of canineinsulinoma is poor. Signs of hypoglycaemia often recur soon after surgery because tumour tissue has only been resected partially and/or functional (micro-)metastases were present. Using quantitative real-time PCR, the expression of 16 target genes was compared between primary canineinsulinomas and their corresponding metastases. There was significantly higher expression of genes encoding for growth hormone (GH) and insulin-likegrowthfactor-1 (IGF-1) in metastases compared to their primary tumours. Immunohistochemical examination of proteins of the GH–IGF-1 axis revealed expression of GH, IGF-1 and GH receptor (GHR) in both primary insulinomas and metastases. Immunohistochemical staining for IGF-1 was significantly higher in metastases compared to primary tumours. Keywords Canine; Insulinoma; β-cell tumour; Growth hormone; Insulin-likegrowthfacto

Identification of Novel Targets for Lung Cancer Therapy Using an Induced Pluripotent Stem Cell Model

RATIONALE: Despite extensive studies, the genetic and epigenetic mechanisms that mediate initiation and progression of lung cancers have not been fully elucidated. Previously, we have demonstrated that via complementary mechanisms, including DNA methylation, polycomb repressive complexes, and noncoding RNAs, cigarette smoke induces stem-like phenotypes that coincide with progression to malignancy in normal respiratory epithelia as well as enhanced growth and metastatic potential of lung cancer cells. OBJECTIVES: To further investigate epigenetic mechanisms contributing to stemness/pluripotency in lung cancers and potentially identify novel therapeutic targets in these malignancies, induced pluripotent stem cells were generated from normal human small airway epithelial cells. METHODS: Lung induced pluripotent stem cells were generated by lentiviral transduction of small airway epithelial cells of OSKM (Yamanaka) factors (octamer-binding transcription factor 4 [Oct4], sex-determining region Y box 2 [SOX2], Kruppel-like factor 4 [KLF4], and MYC proto-oncogene, bHLH transcription factor [MYC]). Western blot, real-time polymerase chain reaction, and chromatin immunoprecipitation sequencing analysis were performed. RESULTS: The lung induced pluripotent stem cells exhibited hallmarks of pluripotency, including morphology, surface antigen and stem cell gene expression, in vitro proliferation, and teratoma formation. In addition, lung induced pluripotent stem cells exhibited no chromosomal aberrations, complete silencing of reprogramming transgenes, genomic hypermethylation, upregulation of genes encoding components of polycomb repressive complex 2, hypermethylation of stem cell polycomb targets, and modulation of more than 15,000 other genes relative to parental small airway epithelial cells. Additional sex combs like-3 (ASXL3), encoding a polycomb repressive complex 2-associated protein not previously described in reprogrammed cells, was markedly upregulated in lung induced pluripotent stem cell as well as human small cell lung cancer lines and specimens. Overexpression of the additional sex combs like-3 gene correlated with increased genomic copy number in small cell lung cancer lines. Knock-down of the additional sex combs like-3 gene inhibited proliferation, clonogenicity, and teratoma formation by lung induced pluripotent stem cells and significantly diminished in vitro clonogenicity and growth of small cell lung cancer cells in vivo. CONCLUSIONS: Collectively, these studies highlight the potential utility of this lung induced pluripotent stem cell model for elucidating epigenetic mechanisms contributing to pulmonary carcinogenesis and suggest that additional sex combs like-3 is a novel target for small cell lung cancer therapy