Epigenetic regulation in muscle-invasive urothelial carcinoma of the bladder in the dog, a translational model of human cancer.

Muscle-invasive urothelial carcinoma (MIUC) is the most common type of bladder malignancy in dogs, but the treatments used in the clinic are relatively ineffective for most of them. Dogs represent a naturally- occurring model for human MIUC and the advances in veterinary oncology could benefit human oncology as well. The field of epigenetics presents unique opportunities for new cancer therapeutics or biomarkers, as epigenetic modification of key genes can regulate tumor initiation and progression. This review summarizes the existing literature on epigenetic changes in canine MIUC as compared to human MIUC and provides suggestions for future studies that could benefit both human and canine patients

Morphological Changes of Gingiva in Streptozotocin Diabetic Rats

Gingivitis and periodontitis are chronic bacterial diseases of the underlying and surrounding tooth tissues. Diabetes mellitus is responsible for tooth deprivation both by decay and periodontal disease. The streptozotocin-induced diabetes results in a diabetic status in experimental animals similar to that observed in diabetes patients. The aim of the study was to investigate the relationship between the gingival lesions and the microangiopathy changes in streptozotocin-induced diabetes mellitus. Forty male Wistar rats were divided into two groups (control and experimental). Diabetes mellitus was induced by 45 mg/kg IV streptozotocin. The histological investigation of the marginal gingival and the relevant gingival papilla showed inflammation of the lamina propria and the squamous epithelium as well as marked thickness of the arteriole in the diabetic group, but no changes were observed in the control group. The results suggested a probable application of a routine gingival histological investigation in diabetic patients in order to control the progress of disease complications. It may be concluded that histological gingival investigation can be used as a routine assay for the control of the diabetic disease and prevention of its complications

Alteration in expression and subcellular localization of the androgen receptor- regulated FAM111A protease is associated with emergence of castration resistant prostate cancer

The androgen receptor (AR) is a pivotal regulator of growth and survival of prostate cancer (PCa) and the majority of lethal castration-resistant prostate cancers (CRPC) remain reliant on AR signaling. PCa exhibits variability in progression and responses to treatment suggesting genetic heterogeneity. Two independent studies identified PCa predisposing single nucleotide polymorphisms (SNPs) within the FAM111A protease gene, but the mechanistic basis of this association remained elusive. Our in vitro and in vivo studies uncovered that AR represses FAM111A in castration sensitive and resistant cells via an AR binding site within the FAM111A gene. FAM111A levels are significantly lower in matched castration-resistant than in castration-sensitive cells and xenografts, and lower in metastatic lesions than in primary tumors. We discovered that FAM111A is AR-repressed in castration sensitive PCa xenograft and multiple PCa cells. Additionally, FAM111A subcellular localization changes dramatically with acquisition of castration resistance, where in castration sensitive cells FAM111A is predominantly in the nucleoli, but with castration resistance it becomes more dispersed in the nucleus and in the cytoplasm. FAM111A depletion in castration sensitive and resistant cells enhances the efficacy of PARP1 inhibitors olaparib and niraparib, consistent with its role in DNA repair. Moreover, FAM111A depletion reduces AR target gene prostate specific antigen (PSA) and transmembrane serine protease 2 (TMPRSS2) transcription, indicating that FAM111A modulates AR-dependent gene expression forming a FAM111A-AR co-regulatory loop in PCa. Our studies argue that AR-dependent FAM111A regulation modulates PCa gene expression, acquisition of castration resistance, and sensitivity to agents that target DNA damage repair

Untargeted Metabolomics Identify a Panel of Urinary Biomarkers for the Diagnosis of Urothelial Carcinoma of the Bladder, as Compared to Urolithiasis with or without Urinary Tract Infection in Dogs

Urothelial carcinoma (UC), the most common urologic cancer in dogs, is often diagnosed late because the clinical signs are shared by other non-malignant lower urinary tract disorders (LUTD). The urine-based BRAFV595E test for UC is highly effective only in certain breeds; hence additional non-invasive biomarkers of UC are needed. Here, urine from dogs with UC (n = 27), urolithiasis (n = 8), or urolithiasis with urinary tract infection (UTI) (n = 8) were subjected to untargeted metabolomics analyses, using GC-TOF-MS for primary metabolites, QTOF-MS for complex lipids, and HILIC-QTOF MS for secondary and charged metabolites. After adjusting for age and sex, we identified 1123 known metabolites that were differentially expressed between UC and LUTD. Twenty-seven metabolites were significant (1.5 ≤ log2FC ≤ −1.5, adjusted p-value < 0.05); however, 10 of these could be attributed to treatment-related changes. Of the remaining 17, 6 (hippuric acid, N-Acetylphenylalanine, sarcosine, octanoylcarnitine, N-alpha-methylhistamine, glycerol-3-galactoside) discriminated between UC and LUTD (area under the ROC curve > 0.85). Of the 6 metabolites, only hippuric acid and N-alpha-methylhistamine were discriminatory in both male (n = 20) and female (n = 23) dogs, while sarcosine was an effective discriminator in several breeds, but only in females. Further investigation of these metabolites is warranted for potential use as non-invasive diagnostic biomarkers of dogs with UC that present with LUTD-related clinical signs

Comparative Metabolomics and Transcriptomics in mouse and dog models of Urological Cancers- Testing Toxicity and Efficacy of Novel Compounds

In this dissertation, I utilized multi-omics to elucidate different mechanisms of therapeutic resistance in aggressive forms of lower urinary tract neoplasms: human muscle-invasive urothelial carcinoma of the bladder and human castration resistant prostate cancer. I assessed the toxicity and efficacy of new small molecules with anti-tumor properties that were developed at the University of California at Davis. Finally, since the dog is a naturally occurring model of human cancers, I summarized similarities and differences between canine and human bladder cancer and identified a panel of potential urinary biomarkers for the early diagnosis of canine bladder cancer. In order to discover more effective cancer therapeutics, more efficient and representative in vitro and in vivo models are needed. Dogs represent a naturally occurring model of cancer. Dogs are immunocompetent animals that share risk factors, pathophysiological features, clinical signs and response to chemotherapeutics with human cancer patients. Muscle-invasive urothelial carcinoma (MIUC) is the most common urologic cancer in dogs, but often diagnosed late because the clinical signs are shared by other non-malignant lower urinary tract disorders (LUTD). Here, I used urine from dogs with MIUC or other LUTD and subjected them to untargeted metabolomic analyses to discover a biomarker for early MIUC diagnosis. After adjusting for age and sex, I identified 1123 known metabolites that were differentially expressed between UC and controls. Six metabolites (hippuric acid, N-acetylphenylalanine, sarcosine, octanoylcarnitine, N-alpha-methylhistamine, glycerol-3-galactoside) discriminated between UC and LUTD (Area under the ROC curve (AUC) >0.85, 1.5≤log2FC≤-1.5 and adj.p.value<0.05). Further investigation of these metabolites may lead to their potential use as non-invasive diagnostic biomarkers for dogs with MIUC that present with LUTD-related clinical signs.The dog is a naturally occurring model for human MIUC. MIUC is the most advanced type of bladder malignancy in humans. Cisplatin (CDDP) is the mainstay of neoadjuvant and adjuvant chemotherapy to treat muscle-invasive bladder cancer (MIBC) in patients undergoing radical cystectomy (RC), but most patients eventually develop chemoresistance. ErbB3, a member of the EGFR family of receptors, is frequently deregulated in CDDP-resistant tumors. I found that CDDP affects ErbB3 signaling by paradoxically increasing the levels of Heregulin, ErbB3 receptor’s ligand, while reducing ErbB3 phosphorylation at Y1289 and protein levels at the plasma membrane in CDDP-resistant cells. The combination of cisplatin and seribantumab, a monoclonal antibody targeting ErbB3, increased apoptosis and growth arrest in S-phase in CDDP-resistant and sensitive MIUC cells. Overall, the anti-ErbB3 antibody MM-121 shows anti-tumorigenic properties in cisplatin-sensitive and resistant cells, indicating a possible use in MIUC. MIUC has a higher predilection in men than women (approximately 3:1), which has attracted interest in the role of the androgen receptor (AR) in this malignancy. AR consists of the N-terminal domain (NTD), the DNA-binding domain (DBD), the hinge region (HR) and the ligand binding domain (LBD), where androgens bind to. Androgen deprivation therapy (ADT) is the mainstay for PCa, but it is not effective for low molecular weight (LMW) AR splice variants that lack the LBD. These LMW AR splice variants can function independently of androgen stimulation and lead to ADT-resistant form of prostate cancer. However, the AR is expressed, and is functional in other cancers as well. Here I report a novel, LMW AR variant, named AR-v19, that is missing the LBD and contains 15 additional amino acids encoded by intron 3 sequences. I detected AR-v19 in primary BlCa tumors and patient-derived xenografts as well as in CWR-22Rv1 PCa cells. AR-v19 depletion afflicted mitochondrial respiration by decreasing maximal respiration and spare respiratory capacity in UMUC-3 BlCa cells. Our collaborators developed a novel small molecule C15 which targets the DBD of the AR which is well-conserved among the AR splice variants. C15 exhibited low toxicity. Since AR-v19 was also expressed in CWR-22Rv1 derived xenografts, C15 was effective in improving the overall survival of Balb/c nude mice bearing CWR-22Rv1-derived xenografts. C15 holds promise as a new therapeutic option for AR variants that have lost the LBD but retained the DBD, including AR-v19. As mentioned above, ADT is the mainstay for PCa, however soon the patients become resistant, leading to the development of castration-resistant prostate cancer (CRPC), for which new therapeutics are urgently needed. For this reason, I assessed the safety, toxicity and cell signaling of another small molecule developed in UCDavis, LLS80, in CRPC cell lines and cell line-derived xenografts. LLS80, is an inhibitor of a protein called galectin-1 (gal-1) which is upregulated in PCa and is associated with a worse clinical prognosis. LLS80 caused a reduction in PCa cell viability alone and in combination with the AR inhibitor enzalutamide in gal-1 expressing cells as well as in Balb/c nude mice bearing CWR-Rv1 and CWR-R1-derived xenografts. Transcriptomic analysis of LLS80-treated CWR-Rv1 xenografts, identified 57 genes differentially expressed, including the downregulation of various AR targets and markers of differentiation towards a sub-lethal form of CRPC called Neuroendocrine PCa. Overall, LLS80 holds promise as a new therapeutic that may effectively block PCa progression towards the Neuroendocrine phenotype following AR inhibition

EFFECT OF FORCEPS SIZE AND MODE OF ORIENTATION ON ENDOSCOPIC SMALL-BOWEL BIOPSY EVALUATION

Endoscopy is increasingly being used to obtain duodenal biopsy specimens in suspected small intestinal malabsorption. We have prospectively evaluated the effect of standard and jumbo biopsy forceps, as well as the mode of orientation of the specimens (naked eye or stereomicroscopy), on duodenal biopsy weight, length, depth, and orientation in 18 consecutive patients. A pair of biopsy specimens was obtained from each patient by each type of forceps in random order. After they had been weighed, one biopsy specimen from each pair was oriented stereomicroscopically and all four were blindly evaluated by two pathologists. The biopsy specimens obtained with the jumbo forceps were significantly larger (15.9 +/- 6.9 mg, mean +/- SD) and longer (0.6 + 0.2 cm) than those obtained with the standard forceps (8.0 +/- 1.3 mg, 0.4 +/- 0.2 cm, respectively; p < 0.001). Seventy-two percent of the jumbo biopsy specimens that were oriented with stereomicroscopy included a minimum of four villi in a row, as compared to 44% of the eye-oriented jumbo specimens and less than 39% of the standard specimens, irrespective of the mode of orientation (p = 0.02). These results indicate that the jumbo forceps is superior to the standard, because it produces a larger duodenal mucosal specimen, usually suitable for optimal histologic evaluation when oriented with stereomicroscopy

Nifedipine-induced histological changes in the parotid glands of hypertensive rats

Nifedipine is a widely used anti-anginal and anti-hypertensiveagent. It is associated with significant gingival changes attributedmore to collagen hyperplasia than to enhancement of proteinsynthesis. We investigated the influence of nifedipine on morphologicalchanges in the parotid glands of rats in a model of hypertension.Twenty-eight male Wistar rats (8-10 weeks; 200 ± 15 g) were dividedinto four groups (A-D). Hypertension was induced by surgical meansin groups C and D. Animals in groups B and D were treated with nifedipine(0.85 mg/kg) via a gastroesophageal catheter the day after surgery(experimental day-1) for 2 weeks. A significant difference was observedbetween the control group and nifedipine group and betweenthe control group and hypertension group with regard to the weightof the parotid gland and its surface area. Histological findings demonstratedchanges in the parotid glands of hypertensive animals withmild vessel dilatation and infiltration of inflammatory cells. These histologicalfindings seemed to be due more to changes in venous functionthan to alterations in gland architecture