Objective: To evaluate in vitro effects of gemcitabine alone and in combination with carboplatin on canine transitional cell carcinoma (TCC) cell lines. Sample: In vitro cultures of 5 canine TCC cell lines. Procedures: Cells were treated with gemcitabine, carboplatin, or a combination of both at various concentrations. Cell proliferation was assessed via a fluorescence-based microplate cell proliferation assay. Cell cycle was evaluated via propidium iodide staining, and apoptosis was assessed by measurement of caspase 3 and 7 enzymatic activity. Synergy between gemcitabine and carboplatin was quantified via combination index analyses. Results: Treatment of 5 canine TCC cell lines with gemcitabine or carboplatin decreased cell proliferation, increased apoptosis, and induced cell cycle arrest. Cell cycle arrest and apoptosis were markedly increased when cell lines were treated with both gemcitabine and carboplatin simultaneously or sequentially. Order of administration during sequential treatment did not consistently affect cell proliferation results in TCC cell lines. When TCC cell lines were treated with gemcitabine and carboplatin in combination at therapeutically relevant concentrations (gemcitabine concentration, < 10μM; carboplatin concentration, < 250μM), a significant decrease in cell proliferation was observed, compared with cell proliferation following treatment with gemcitabine or carboplatin alone. In combination, the effects of gemcitabine and carboplatin were synergistic in 3 of 5 cell lines and additive in the other 2. Conclusions and Clinical Relevance: Gemcitabine had antitumor effects on canine TCC cells in vitro, and the combination of gemcitabine and carboplatin had synergistic activity at biologically achievable concentrations.

Objective-To establish the maximum tolerated dose of Clostridium novyi-NT spores in tumor-bearing dogs and evaluate spore germination within tumors and tumor response. Animals-6 client-owned dogs. Procedures-A standard dose-escalation study was planned, with maximum tolerated dose defined as the highest dose at which 0 or 1 of 6 dogs had dose-limiting toxicoses (DLT). Dogs received 1 dose of C novyi-NT spores IV. Toxicoses were graded and interventions performed according to specific guidelines. Grade 3 or higher toxicosis or any toxicosis combination that substantially affected patient status was considered DLT. Clinical response was measured by use of response evaluation criteria in solid tumors at 28 days. Results-The first 2 dogs had DLT. The dose was decreased. Two of the next 4 dogs had DLT; therefore, dose administration was stopped because the study endpoint had been reached. The most common toxicosis was fever (n = 6 dogs). Two dogs developed abscesses (1 within a nasal carcinoma and 1 splenic abscess) attributable to C novyi-NT infection; both required surgical intervention. Clostridium novyi-NT was cultured from 1 of 6 tumors. Five dogs were available for response assessment (4 had stable disease; 1 had progressive disease). Conclusions and Clinical Relevance-Results indicated that C novyi-NT can germinate within tumors of dogs. Toxicosis, although common and sometimes severe, was manageable with treatment. Further studies in dogs with superficial tumors may allow for continued dose escalation and provide information for use in clinical trials in veterinary and human oncology.

Objective: To determine expression of microRNA (miRNA) in urinary bladder samples obtained from dogs with grossly normal urinary bladders, inflammatory bladder disease, or transitional cell carcinoma (TCC) and in cells of established canine TCC cell lines. Sample: Samples of grossly normal bladders (n = 4) and bladders from dogs with inflammatory bladder disease (13) or TCC (18), and cells of 5 established canine TCC cell lines. Procedures: Expression of 5 miRNAs (miR-34a, let-7c, miR-16, miR-103b, and miR-106b) that target p53, Rb, or Bcl-2 protein pathways was determined for bladder samples and cells via quantitative real-time PCR assay. Effects of cisplatin (5μM) on proliferation and miRNA expression of cells were determined. Results: Expression of miR-34a and miR-106b was significantly higher in TCC samples than it was in samples of grossly normal bladders. Expression of miR-34a, miR-16, miR-103b, and miR-106b was higher in TCC samples than it was in bladder samples from dogs with inflammatory bladder disease. Cells of established canine TCC cell lines that had the lowest growth after cisplatin treatment had increased miR-34a expression after such treatment. Conclusions and Clinical Relevance: Findings of this study indicated results of miRNA expression assays can be used to distinguish between samples of grossly normal bladders and bladders of dogs with inflammatory bladder disease or TCC. This finding may have clinical relevance because currently available diagnostic tests cannot be used to differentiate these tissues, and inflammatory bladder disease and TCC are both prevalent in dogs. Validation of miRNA expression assays as diagnostic tests may be warranted.

Objective: To determine the accuracy of 3-D and 2-D ultrasonography for quantification of tumor volume in dogs with transitional cell carcinoma (TCC) of the urinary bladder. Animals: 10 dogs with biopsy-confirmed TCC. Procedures: The urinary bladder of each dog was distended with saline (0.9% NaCl) solution (5.0 mL/kg), and masses were measured via 3-D and 2-D ultrasonography. Masses were also measured via 3-D ultrasonography after bladders were distended with 2.5 and 1.0 mL of saline solution/kg. Subsequently, the bladder was deflated and distended with CO2 (5.0 mL/kg); CT was performed after IV contrast medium administration. Tumor volumes were calculated via 3-D ultrasonography, 2-D ultrasonography, and CT (reference method) and compared via ANOVA, Deming regression, and Bland-Altman plots. Repeated-measures ANOVA was used to assess effects of bladder distension on 3-D tumor volume measurements. Repeatability of measurements was estimated via the coefficient of variation for each method. Results: Repeatability was considered good for all 3 methods. There was no significant difference in tumor volume measurements obtained via 3-D ultrasonography at different degrees of urinary bladder distension. Results of Deming regression and Bland-Altman plots indicated excellent agreement between tumor volume measurement with 3-D ultrasonography and CT, but not between 2-D ultrasonography and CT. Conclusions and Clinical Relevance: Tumor volume in dogs with TCC of the urinary bladder was accurately measured via 3-D ultrasonography. Use of 3-D ultrasonography can provide a less expensive and more practical method for monitoring response to treatment than CT and was more accurate than 2-D ultrasonography.