Review paper: Cancer chemopreventive compounds and canine cancer.

Canine cancer has become more prevalent in recent years because of increased life expectancy and greater attention to the health of pets. The range of cancers seen in dogs is as diverse as that in human patients, and despite more intensive therapeutic interventions, fatality rates remain unacceptably high in both species. Chemoprevention is therefore an important means of confronting this disease. Because domestic pets share our environment, greater cross-application and study of the protumorigenic and antitumorigenic factors in our shared environment will benefit all species, leading to the development of new families of less toxic antitumorigenic compounds based on novel and established molecular targets. Currently, the most interesting cancer preventive agents are nonsteroidal anti-inflammatory drugs, peroxisome proliferator-activated receptor-gamma ligands, and dietary compounds. This article provides an overview of what is known about how these agents affect molecular signaling in neoplastic disease, with reference to reported application and/or study in dogs where available.

Antigen and antibody testing for the diagnosis of blastomycosis in dogs.

BACKGROUND: Early diagnosis and treatment are associated with an improved prognosis in blastomycosis. The diagnosis of blastomycosis may be missed by cytology, histopathology, culture, or serology. An enzyme immunoassay (EIA) for detection of Blastomyces dermatitidis galactomannan antigen in body fluids has been used for rapid diagnosis of blastomycosis in humans. HYPOTHESIS: Measurement of Blastomyces antigen in urine or serum by the MVista Blastomyces antigen EIA is more sensitive than measurement of anti-Blastomyces antibodies for diagnosis of blastomycosis in dogs. METHODS: Serum and urine samples from 46 dogs with confirmed blastomycosis were tested for Blastomyces antigen and serum was tested for anti-Blastomyces antibodies. RESULTS: The sensitivity for the detection of antigen in urine was 93.5% and it was 87.0% in serum. The sensitivity of antibody detection by agar gel immunodiffusion (AGID) was 17.4% and it was 76.1% by EIA. Antigen and antibody decreased during itraconazole treatment. CONCLUSIONS AND CLINICAL IMPORTANCE: Antigen detection is a more sensitive test for diagnosis of blastomycosis than antibody testing by AGID, the only commercially available method. Antigen concentrations decreased with treatment.

Effects of L-asparaginase on plasma amino acid profiles and tumor burden in cats with lymphoma.

BACKGROUND: L-Asparaginase (Elspar(a)), is an Escherichia coli-derived enzyme that depletes lymphoma cells of asparagine, inhibiting protein synthesis and resulting in cell death. The single agent response rate in cats with lymphoma and impact of L-asparaginase on plasma amino acid concentrations is unknown. HYPOTHESES: L-Asparaginase significantly reduces plasma asparagine concentrations and has demonstrable efficacy against untreated lymphoma in cats. ANIMALS: Thirteen cats with confirmed lymphoma (LSA) of any anatomic site were given 1 dose 400 IU/kg IM) of L-asparaginase for initial LSA treatment. METHODS: Plasma collected at 0, 2, and 7 days after L-asparaginase therapy was assayed for ammonia, asparagine, aspartic acid, glutamine, and glutamic acid concentrations. Cats were restaged 7 days later to assess tumor response. RESULTS: Eight cats had T-cell LSA, 4 cats had B-cell LSA, and 1 cat's immunophenotype was unknown. Two complete and 2 partial responses to L-asparaginase were seen. Four cats had stable disease, and 5 cats had progressive disease. Ammonia and aspartic acid concentrations were increased from baseline at 2 and 7 days posttreatment. Asparagine concentrations were decreased from baseline at 2 days but not 7 days posttreatment. Glutamic acid concentrations were increased at day 2 compared to day 7 posttreatment but not compared to baseline. Glutamine concentrations were unchanged. CONCLUSIONS AND CLINICAL IMPORTANCE: L-asparaginase significantly reduced asparagine concentrations within 2 days of treatment, but this effect was lost within 7 days. The apparent overall response rate of feline LSA to L-asparaginase in this study was 30%.

Treatment of Disseminated Aspergillosis with Posaconazole in 10 Dogs.

BACKGROUND: Few effective treatments for disseminated Aspergillus infections in dogs are available. Posaconazole has potent and broad-spectrum activity against Aspergillus spp., but its use has not yet been sufficiently evaluated in dogs. HYPOTHESIS/OBJECTIVES: The aim of this study was to determine the safety and efficacy of posaconazole for the treatment of naturally occurring disseminated Aspergillus infections in dogs. ANIMALS: Ten client-owned dogs with disseminated aspergillosis. METHODS: Prospective, nonrandomized, noncontrolled study with posaconazole administered to dogs at dosage of 5 mg/kg p.o. q12h. The primary veterinarian or the veterinary specialist caring for the dogs provided patient data. RESULTS: The treatment response for dogs with disseminated disease while receiving posaconazole was defined as clinical remission (n = 4) and clinical improvement (n = 6). There was a high rate of relapse during treatment or after cessation of treatment in both groups, and most dogs died or were euthanized due to progressive disease. Excluding 1 dog concurrently treated with terbinafine that remains alive 5 years after diagnosis, the mean survival time for dogs was 241 days (range 44-516 days). Three other dogs lived >1 year after starting treatment. No clinically relevant adverse events or increases in serum liver enzyme activity occurred during treatment with posaconazole. CONCLUSIONS AND CLINICAL IMPORTANCE: Posaconazole appears to be safe and well-tolerated for treatment of disseminated Aspergillus infections in dogs. Long-term survival >1 year is possible with prolonged treatment, but relapse is common.

Serum granulocyte colony-stimulating factor (G-CSF) and interleukin-1 (IL-1) concentrations after chemotherapy-induced neutropenia in normal and tumor-bearing dogs.

Hematopoiesis is regulated by complex interactions of hematopoietic growth factors known as colony-stimulating factors and interleukins. We used sensitive bioassays to quantitate serum granulocyte colony-stimulating factor (G-CSF) and interleukin-1 (IL-1) concentrations in normal and tumor-bearing dogs following administration of myelosuppressive chemotherapy (vincristine, doxorubicin, cyclophosphamide). Serum G-CSF and IL-1 increased during the neutrophil nadir in 13 of the 16 dogs. Serum G-CSF concentrations were significantly increased in normal and in tumor-bearing dogs on neutropenic compared to non-neutropenic days. Serum IL-1 concentrations increased significantly on neutropenic days in normal dogs but not in tumor-bearing dogs. A marked neutrophilia was observed in normal dogs, but not in tumor-bearing dogs, following the increases in serum G-CSF and IL-1 concentrations (days 7, 8, and 9, p < 0.05). Normal dogs produced significantly more G-CSF on neutropenic days compared to dogs with lymphoma. On non-neutropenic days, serum IL-1 concentrations were significantly increased in dogs with lymphoma and in dogs with nonlymphoid malignancies compared to normal dogs. These results suggest an important role for G-CSF and IL-1 in hematopoietic recovery after chemotherapy-induced myelosuppression and document an altered hematopoietic regulation in animals with malignancy compared to normal subjects.

Masitinib mesylate for metastatic and non-resectable canine cutaneous mast cell tumours.

Masitinib mesylate is a tyrosine kinase inhibitor approved for the treatment of gross, non-metastatic grade II and III canine mast cell tumours (MCTs). This study evaluated the use of masitinib as a frontline and rescue agent for metastatic and non-metastatic canine MCTs. Identification of toxicities and prognostic factors in these dogs was of secondary interest. Twenty-six dogs were included in this study. The overall response rate to masitinib was 50%. The median survival time for dogs that responded to masitinib was 630 days versus 137 days for dogs that did not respond (P = 0.0033). Toxicity was recorded in 61.5% of treated dogs, but the majority of adverse events were mild and self-limiting. Response to masitinib, not tumour grade, stage or location, was the most significant prognostic factor for survival in dogs with MCTs.

The frequency of micronuclei in lymphocytes of dogs with osteosarcoma: a predictive variable for tumor response during cisplatin chemotherapy.

To our knowledge, there are no features identifiable at the time of diagnosis or during treatment that can assist the clinician in predicting the response to cisplatin therapy in dogs with osteosarcoma. In this study, we describe a direct relationship between the percentage of G0 lymphocytes containing micronuclei following exposure to one dose of cisplatin in vivo and tumor response in dogs with osteosarcoma. The response of tumors to chemotherapy is thought to be a function of the drug's pharmacological properties (e.g., peak plasma concentration and elimination half-life); however, a relationship between platinum DNA adduct levels in leukocyte DNA and tumor response has been observed by others, suggesting that clinical resistance to platinum drugs is attributable to DNA repair functions of the host, and thus the degree of cytotoxicity is similar across all cell types. Our results support this hypothesis. Those dogs receiving cisplatin chemotherapy and having a micronuclear frequency of greater than 10% had median remission and survival times of 68.3 and 79.0 weeks, respectively, whereas those dogs with a micronuclear frequency of less than 10% had median remission and survival times of 14.1 and 17.9 weeks, respectively.

Separation of feline bone marrow cells by counterflow centrifugal elutriation. Identification and isolation of presumptive early and late myeloid/erythroid progenitors.

Counterflow centrifugal elutriation (CCE) has been used to separate nucleated cells from mammalian bone marrow on the basis of size with the resultant isolation of hematopoietic cells in varying stages of lineage development. We examined the feasibility of identifying and isolating such cells from feline bone marrow. CCE was performed with a Beckman J6MI centrifuge and a Sanderson chamber, using a fixed rotor speed of 3000 rpm and collection of cells at (1) 16-, (2) 21-, (3) 25-, (4) 32 ml/min, and (5) a rotor off fraction. Recovery of the total input cells in four replicate experiments averaged 86%, with the maximum number of recovered cells in fraction 4. Analysis by flow cytometry and monoclonal antibodies revealed mononuclear cells in fractions 1 and 2 and early and late differentiating myeloid/erythroid cells in fractions 2 through 5. T lymphocytes and alloreactivity in a mixed lymphocyte reaction (MLR) were restricted to fractions 1 and 2; removal of T cells and MLR activity was accomplished by immunomagnetic depletion. In vitro cultures for clonogenic cells revealed CFU-GM and BFU-E colonies in fractions 2 through 5, with fraction 4 containing the greatest absolute number of myeloid colonies and fractions 3 and 4 the majority of the erythroid colonies. More important, in examining the plating efficiency for clonogenic cells in the different fractions it was found that this increased significantly in fractions 2 and 3 when the culture time was extended from 7 to 14 days; in contrast, fractions 4 and 5 reached their maximum plating efficiency within 7 days with no further increase on day 14. We interpret these findings to indicate the presence of late differentiating progenitors in the large-cell size fractions 4 and 5, while the smaller mononuclear cells in fractions 2 and 3 represent an earlier, more primitive population of hematopoietic cells requiring an extended time in culture for full colony development.

Amputation and dexniguldipine as treatment for canine appendicular osteosarcoma.

The biological behavior of osteosarcoma in dogs is similar to that in humans and the dog has been suggested as a model for the disease in humans. Because occult metastatic disease is common at presentation, systemic therapy is necessary. The dihydropyridine, dexniguldipine hydrochloride (B859-35), is a potent inhibitor of protein-kinase-C(PKC)-stimulated cell proliferation and has shown therapeutic activity in experimentally induced neuroendocrine hamster lung tumors and in a mammary cancer cell line. In human osteosarcoma cell lines, PKC activity can be down-regulated, resulting in increased sensitivity to cisplatin. Since these results supported the involvement of PKC inhibitors in the therapeutic management of osteosarcoma, we performed a prospective, randomized clinical trial using dogs with naturally occurring appendicular osteosarcoma to determine the therapeutic potential of dexniguldipine. Dogs received either no drug treatment (control group, n = 8), standard treatment (e.g., cisplatin, n = 14), or dexniguldipine treatment (n = 14) following amputation. Dexniguldipine- and cisplatin-treated dogs had a longer median remission duration and survival time than untreated dogs (P < 0.05); however, dexniguldipine-treated dogs had a shorter survival time than cisplatin-treated dogs (P < 0.05). The results of this study demonstrate that dexniguldipine has significant activity in the inhibition of canine osteosarcoma micrometastases. The identification of a tumor model that may be responsive to this class of antiproliferative agents warrants further clinical investigation to determine the optimum dosage of dexniguldipine and the role it may have in the therapeutic management of canine osteosarcoma.

Isolation and functional studies on feline bone marrow derived macrophages.

In this report, we describe an in vitro culture method for feline bone marrow cells, which yields large numbers of quiescent macrophages after 14 days of culture. The bulk of the cultured cell population consists of macrophages as assessed by morphology, macrophage specific cytochemistry, and phagocytosis. The remaining cells were lymphocytes, bone marrow stromal cells, fibroblasts and occasional polymorphonuclear leukocytes. While resting cells produced no detectable interleukin 1, stimulation with lipopolysaccharide (LPS) induced the production of biologically active interleukin 1. After 6 h LPS stimulation, mRNA for tumor necrosis factor alpha and interleukin 1 beta was detectable. The absence of mRNA in unstimulated cells indicates cultured macrophages were not activated until stimulated by LPS or plastic adherence. This approach provides a useful means to measure potential modulatory effects by virus infections or other agents upon feline macrophage gene expression.

Treatment of coccidioidomycosis osteomyelitis with itraconazole in a horse. A brief report.

Itraconazole, a tricyclic azole effective against a number of deep mycotic diseases, was used to treat a Quarter Horse filly with coccidioidomycosis. The horse was almost normal after 90 days of treatment. Five months after discontinuing itraconazole treatment, the filly had severe neck pain and neurologic signs from recurrence of coccidioidomycosis and was treated with itraconazole for an additional 6 months. Her clinical condition improved to almost normal and the filly has remained normal for 2 years. There was no evidence of drug toxicity.

Efficacy of a feline leukemia virus vaccine in a natural exposure challenge.

A commercial feline leukemia virus (FeLV) vaccine was evaluated in a natural exposure system. All kittens were negative for FeLV antigen on two enzyme-linked immunosorbent assay (ELISA) tests and one indirect immunofluorescence antibody (IFA) test before vaccination or exposure. Twenty-three kittens were vaccinated subcutaneously at nine and 12 weeks of age. The vaccinated kittens and 14 unvaccinated littermates were housed in an infected environment starting at 14 weeks. The kittens were exposed for 24 weeks by living in a large room with one feline leukemia virus-positive, asymptomatic adult cat for each five kittens. Sixty-four percent of the unvaccinated kittens and 70% of the vaccinated kittens became infected as determined by ELISA. Forty-three percent of unvaccinated kittens and 39% of vaccinated kittens died. There was no difference between the infection and mortality of vaccinated kittens that developed antibodies to anti-FeLV glycoprotein 70-envelope antigen and those that did not. Consideration should be given to evaluation of feline leukemia virus vaccines using "street" virus in a natural exposure system.

Concurrent infection with Toxoplasma gondii and feline leukemia virus. Antibody response and oocyst production.

Four adult cats (two testing positive and two negative for feline leukemia virus FeLV) were fed Toxoplasma gondii tissue cysts collected from the brains of mice. Two control cats (1 FeLV+, 1 FeLV-) were not fed cysts. The cats infected with T. gondii shed thousands of oocysts but remained clinically and physically normal, with hemograms and clinical chemistry values essentially unchanged irrespective of their FeLV status. Infection with FeLV did not increase the duration of oocyst shedding. At necropsy no significant lesions were found. T. gondii antibodies were detected by three serologic tests in the cats fed tissue cysts. The time necessary for an antibody response to T. gondii was not altered by the FeLV infection. Indirect hemagglutination (IHA) was the least reliable of the serologic tests studied; it detected antibodies later in the infection, and titers were less than in the other tests. Latex agglutination (LA) detected antibodies a few days before IHA, but titers were less than in modified direct agglutination (MAT). MAT detected antibodies earliest in the infection and also measured antibodies in aqueous humor and cerebrospinal fluid.

Basophilic leukemia in a dog.

Basophilic leukemia with thrombocytosis was diagnosed in a 4-year-old Shih Tzu. This diagnosis was based on cytochemical staining and cytologic examination of blood and bone marrow smears. Hydroxyurea, an inhibitor of DNA synthesis, at a dose of 50 mg/kg PO bid induced hematologic remission after 7 days of treatment. Adverse effects observed included pruritus, erythema of the ventral abdomen, generalized alopecia, and possibly, diabetes mellitus. The dog remained in remission for 21 months before becoming lethargic, at which time the owners requested euthanasia but did not allow a necropsy.

Radiotherapy of metastatic seminoma in the dog. Case reports.

Four dogs with metastatic seminoma were treated with cesium 137 teleradiotherapy. Minimum total tumor dose ranged from 17 to 40 gray (Gy) and was usually given through bilateral opposing sublumbar ports in eight to ten fractions, with three fractions given weekly. The tumor regressed in all four dogs. The first dog (case 1) was free of tumor and died of non-tumor related causes at 57 months. The second dog (case 2) was free of tumor but was euthanatized at 37 months for a limb fracture. The third dog (case 3) was euthanatized for undertermined pulmonary disease 43 months after radiotherapy. The fourth dog (case 4) was euthanatized 6 months following radiotherapy because of transitional cell carcinoma and renal failure. No evidence of seminoma was found at necropsy. Radiotherapy was shown to be effective treatment for seminoma with regional metastasis.

Treatment of blastomycosis with itraconazole in 112 dogs.

One hundred twelve client-owned dogs with blastomycosis were treated with itraconazole, 5 or 10 mg/kg/d. The first group of 70 dogs treated in 1987 and 1988 received 10 mg/kg/d (group 1), and the second group of 42 dogs treated after October 1988 received 5 mg/kg/d (group 2). Even though the groups were treated at different times, the dogs were similar in age and gender distribution, number of sites involved, and percent and severity of pulmonary involvement. The proportion of dogs cured with a 60-day course of itraconazole was similar for both groups (53.6% versus 54.3%) and for a second historical control group treated with amphotericin B (57%); the recurrence rate was also similar, 20%, 21.4%, and 20%, respectively. Dogs treated with itraconazole had similar mortality rates (25.7% at 5 mg/kg/d; 25% at 10 mg/kg/day) to those treated with amphotericin B (23%). Seventeen of the 23 dogs that died (74%), did so during the first week of treatment; these early deaths were usually attributed to respiratory failure. The only site of infection that was significantly associated with failure (death or recurrence) was the brain. There was a marked difference in survival times between dogs without lung disease or with mild lung disease compared with dogs with moderate or severe lung disease. Serum itraconazole concentrations reached steady state by 14 days of treatment. Dogs receiving 5 mg/kg/d of itraconazole (group 2) had mean serum concentrations of 3.55 +/- 2.81 mg/mL (range, 0.67 to 10.8 micrograms/mL), whereas dogs receiving 10 micrograms/kg/d (group 1) had mean concentrations of 13.46 +/- 8.49 micrograms/mL (range, 1.8 to 28 micrograms/mL) (P < or = .001). There was no association between cure and serum itraconazole concentrations. Dogs in group 1 had significantly more adverse effects than dogs in group 2 (P = .046). Anorexia was the most common adverse effect, occurring in 14.9% of dogs in group 1. Only 8% of dogs in group 2 had adverse effects. Serum concentrations of itraconazole were positively correlated with serum alkaline phosphatase and alanine aminotransferase activities. Our findings indicate that itraconazole administered at a dose of 5 mg/kg/d is the drug of choice for blastomycosis in dogs.

Itraconazole for the treatment of histoplasmosis in cats.

Eight cats with histoplasmosis were treated with itraconazole at 5 mg/kg per dose PO bid. There were multiple sites of infection, and 2 of the cats had hypercalcemia that was attributed to the histoplasmosis. All 8 cats were eventually cured, but 2 cats experienced recurrences of disease after completion of therapy, requiring 2 to 3 additional months of itraconazole. There were no clinically relevant adverse effects during treatment. Although a limited number of cats were treated, the study suggests that itraconazole is a well-tolerated and effective drug for the treatment of histoplasmosis in the cat.

Treatment of mycotic rhinitis with itraconazole in three horses.

Itraconazole, a third-generation azole, was evaluated for treatment of resistant nasal mycotic infections in horses. Two horses with Aspergillus spp nasal granulomas and 1 horse with Conidiobolus coronatus nasal infection were treated with itraconazole (3 mg/kg PO bid). One of the horses with nasal aspergillosis was also treated by surgical resection of the nasal septum. The treatment time for the horses ranged from 3 to 4.5 months. No adverse effects were noted in any of the horses during the treatment period. Peak and trough serum itraconazole concentrations were < 0.5 micrograms/mL in all 3 horses. Itraconazole (3 mg/kg PO bid) appears to be effective in the treatment of nasal Aspergillus spp infections in horses because the fungal infection was eliminated in both horses. One horse still had excessive nasal sounds during exercise and was retired from training, whereas the other horse returned to normal. The nasal C. coronatus infection appeared resistant to itraconazole treatment in the affected horse because the granulomas were still present after 4.5 months of treatment.

Investigation of the effects of hyperthyroidism on renal function in the cat.

This study evaluated the effects of thyroxine on renal function in the cat. Baseline serum thyroxine (T4) concentrations, clinicopathologic data (complete blood count [CBC], serum chemistry panel, urinalysis), and nuclear medicine determinations of glomerular filtration rate (GFR), effective renal plasma flow (ERPF), and effective renal blood flow (ERBF) were measured in 10 normal adult cats. Cats were then injected with thyroxine (T4) (50 micrograms/kg SQ) daily for 30 d to induce hyperthyroidism. Clinicopathologic and nuclear medicine studies were repeated at 30 d. Cats injected with thyroxine had significant increases in T4, GFR, and ERBF and significant declines in serum creatinine and blood urea nitrogen (BUN) values. Administration of high doses of exogenous thyroxine to cats results in significant stimulation of renal function.

Feline skin lymphoma: characterization of tumor and identification of tumor-stimulating serum factor(s).

A feline leukemia virus-negative skin lymphoma was characterized as a T-lymphocyte neoplasm, using the guinea pig erythrocyte rosetting technique. The lymphoma cells responded well to phytohemagglutinin compared with normal feline lymphocytes which did not respond. Serum factor(s) was found in serum of a cat with lymphoma that was highly stimulating to autologous tumor cells, but not to normal cat lymphocytes.