Retrospective evaluation of toceranib phosphate (Palladia®) toxicity in cats.

The purpose of this study was to describe the toxicity profile of toceranib phosphate in tumour bearing cats. Medical records were reviewed from seven institutions. Patients with incomplete medical records and those receiving concurrent chemotherapy or NSAIDs (non-steroidal anti-inflammatory) were excluded. Fifty-five cats met the inclusion criteria. Carcinoma was diagnosed in 55% of cases. Median oral toceranib dose was 2.7 mg kg-1 and was most commonly administered on Monday, Wednesday and Friday. Thrombocytopenia (16.3%) and neutropenia (9.1%) were the most common haematologic toxicities. Azotemia (14.5%) and alanine aminotransferase (ALT) elevations (7.2%) were the most frequently encountered biochemical alterations. Gastrointestinal (GI) toxicity was seen in 21.8% of cats, and was lower than previously reported in dogs. The results of this study showed that treatment of cats with toceranib is well-tolerated and toxicity is uncommon. Additional studies to define a more structured dosing schedule and to evaluate the efficacy of toceranib in the treatment of feline cancers are needed.

Prognostic markers for canine melanocytic neoplasms: a comparative review of the literature and goals for future investigation.

Many studies have evaluated various prognostic markers for canine melanocytic neoplasms either as primary or secondary goals; however, design, methodology, and statistical validation vary widely across these studies. The goal of this article was to evaluate and compare published canine melanocytic neoplasm studies in relation to the principals established in the Recommended Guidelines for the Conduct and Evaluation of Prognostic Studies in Veterinary Oncology. Based on this evaluation, we determined which parameters currently have the most statistically supported validity for prognostic use in canine melanocytic neoplasia. This information can also be used as part of evidence-based prospective evaluations of treatment regimens. Additionally, we highlight areas in which the current data are incomplete and that warrant further evaluation. This article represents an initiative of the American College of Veterinary Pathologists' Oncology Committee and has been reviewed and endorsed by the World Small Animal Veterinary Association.

Xenogeneic murine tyrosinase DNA vaccine for malignant melanoma of the digit of dogs.

BACKGROUND: Malignant melanoma of dogs is a highly aggressive neoplasm and is the 2nd most common digit tumor. Metastatic disease is a common sequela for which few effective treatment options exist. Studies show that xenogeneic tyrosinase DNA vaccination yields immune responses and prolongation of survival in dogs with oral malignant melanoma. OBJECTIVES/HYPOTHESIS: Describe clinical findings and tumor characteristics of a cohort of dogs with digit malignant melanoma, and evaluate the prognostic utility of a proposed staging system. Determine if a novel xenogeneic DNA vaccine is safe and potentially effective for treatment of dogs with digit melanoma. ANIMALS: Fifty-eight dogs with digit malignant melanoma treated at the Animal Medical Center between 2004 and 2007. METHODS: Retrospective, medical records review of dogs with digit melanoma treated with xenogeneic DNA vaccine. RESULTS: Overall median survival time (MST) for dogs treated with loco-regional control and xenogeneic DNA vaccine was 476 days with a 1-year survival rate of 63%. MST for dogs presenting with metastasis was 105 days versus 533 days for dogs presenting without metastasis (P < .0001). Forty-eight percent of the dogs in the latter group were alive at 2 and 3 years. A proposed staging system proved prognostic with stages I-IV dogs surviving >952, >1,093, 321, and 76 days, respectively. CONCLUSIONS AND CLINICAL IMPORTANCE: The xenogeneic murine tyrosinase DNA vaccine was safe and appears effective when used in conjunction with local and regional disease control. The proposed staging system was prognostic in this study and future studies might benefit from utilizing this staging system.

Recommended guidelines for the conduct and evaluation of prognostic studies in veterinary oncology.

There is an increasing need for more accurate prognostic and predictive markers in veterinary oncology because of an increasing number of treatment options, the increased financial costs associated with treatment, and the emotional stress experienced by owners in association with the disease and its treatment. Numerous studies have evaluated potential prognostic and predictive markers for veterinary neoplastic diseases, but there are no established guidelines or standards for the conduct and reporting of prognostic studies in veterinary medicine. This lack of standardization has made the evaluation and comparison of studies difficult. Most important, translating these results to clinical applications is problematic. To address this issue, the American College of Veterinary Pathologists' Oncology Committee organized an initiative to establish guidelines for the conduct and reporting of prognostic studies in veterinary oncology. The goal of this initiative is to increase the quality and standardization of veterinary prognostic studies to facilitate independent evaluation, validation, comparison, and implementation of study results. This article represents a consensus statement on the conduct and reporting of prognostic studies in veterinary oncology from veterinary pathologists and oncologists from around the world. These guidelines should be considered a recommendation based on the current state of knowledge in the field, and they will need to be continually reevaluated and revised as the field of veterinary oncology continues to progress. As mentioned, these guidelines were developed through an initiative of the American College of Veterinary Pathologists' Oncology Committee, and they have been reviewed and endorsed by the World Small Animal Veterinary Association.

ABCB1-1Delta polymorphism can predict hematologic toxicity in dogs treated with vincristine.

BACKGROUND: Dogs that harbor the naturally occurring ABCB1-1Delta polymorphism experience increased susceptibility to avermectin-induced neurological toxicosis as a result of deficient P-glycoprotein function. Whether or not the ABCB1-1Delta polymorphism affects susceptibility to toxicity of other P-glycoprotein substrate drugs has not been studied. HYPOTHESIS: Dogs that possess the ABCB1-1Delta mutation are more likely to develop hematologic toxicity associated with vincristine than ABCB1 wild-type dogs. ANIMALS: Thirty-four dogs diagnosed with lymphoma were included in this study. METHODS: Cheek swab samples were obtained from dogs diagnosed with lymphoma that were to be treated with vincristine. DNA was extracted from cheek swabs and the ABCB1 genotype was determined. Hematologic adverse drug reactions were recorded for each dog and graded according to the Veterinary Comparative Oncology Group's criteria for adverse event reporting (Consensus Document). In order to avoid possible bias, ABCB1 genotype results for a particular patient were not disclosed to oncologists until an initial adverse event report had been submitted. RESULTS: Dogs heterozygous or homozygous for the ABCB1-1Delta mutation were significantly more likely to develop hematologic toxicity, specifically neutropenia (P= .0005) and thrombocytopenia (P= .0001), after treatment with vincristine than ABCB1 wild-type dogs. CONCLUSIONS AND CLINICAL IMPLICATIONS: At currently recommended dosages (0.5-0.7 mg/M(2)), vincristine is likely to cause hematologic toxicity in dogs with the ABCB1-1Delta mutation, resulting in treatment delays and unacceptable morbidity and mortality. Assessing the ABCB1-1Delta genotype before vincristine administration and decreasing the dosage may prevent toxicity and treatment delays resulting from neutropenia or thrombocytopenia.

Use of imiquimod 5% cream (Aldara) in cats with multicentric squamous cell carcinoma in situ: 12 cases (2002-2005).

Multicentric squamous cell carcinoma in situ (MSCCIS) is a variant of squamous cell carcinoma in cats, commonly referred to as Bowen's-like disease. Imiquimod 5% cream (Aldara) is a novel immune response modifier (IRM) that has been reported as a successful treatment for Bowen's disease in humans. The purpose of this study was to describe clinical findings, treatment protocols and survival in cats with MSCCIS treated with imiquimod 5% cream and to examine the effects of imiquimod 5% cream in cats with MSCCIS. The expression of papillomavirus group-specific antigen in the study population was also determined. From review of medical records, 12 cats were identified with a histologic diagnosis of MSCCIS and treatment with imiquimod 5% cream. Initial lesions responded to imiquimod 5% cream in all cats. Most cats (75%) developed new lesions. New lesions also responded to imiquimod 5% cream in all cats treated. Five cats (41%) had side effects suspected to be associated with the use of imiquimod 5% cream, including local erythema (25%), increased liver enzymes and neutropenia (8%), and partial anorexia and vomiting (8%). Kaplan-Meier median treatment duration and median survival time probabilities for cats in this study were 1189 days, respectively. A time to failure model was generated as many cats were censored from analysis well before the aforementioned projected median. This model resulted in a shorter median survival time of 243 days. No patient-related, tumour-related or treatment-related prognostic variables were identified. No expression for papilloma group-specific antigen was found. Imiquimod 5% cream appears to be well tolerated in the majority of cats, and further studies are warranted to further examine its usefulness in cats with this disease.

Evaluation of P-glycoprotein expression in feline lymphoma and correlation with clinical outcome.

P-glycoprotein (Pgp) is a transmembrane protein pump involved in drug resistance in canine and human lymphoma. There are no published clinical studies evaluating Pgp expression in feline lymphoma. The purpose of this study is to evaluate the level of Pgp expression in feline lymphoma and correlate it with clinical outcome. Two human Pgp monoclonal antibodies, C219 and C494, were used to detect Pgp expression in tissue samples from 63 cats with lymphoma. Demographic results appear comparable to recently published feline lymphoma studies. The Kaplan-Meier median remission and survival times were 164 and 571 days, respectively. Fourteen cats had positive expression of Pgp using MAb C219, and 40 were positive with C494. Variables statistically associated with survival included bone marrow involvement, stage, substage, and use of radiation therapy as a part of treatment. Pgp expression as assessed by MAb C219 and C494 is not predictive of remission or survival time in cats with lymphoma.

Treatment of canine mast cell tumours with vinblastine, cyclophosphamide and prednisone: 35 cases (1997-2004).

The purpose of this retrospective study was to determine the efficacy and toxicity of a combined protocol of vinblastine, cyclophosphamide and prednisone (VCP) in 35 dogs with mast cell tumours (MCTs). Eleven dogs had measurable disease (group 1) and 24 dogs had incompletely excised MCT or were at high risk for metastasis (group 2). Five patients in group 1 achieved complete response, two partial responses, two stable diseases and two progressive diseases. The median progression-free survival time (PFST) for group 1 and 2 dogs was 74 and 865 days, respectively. The median overall survival time (OST) for group 1 and 2 dogs was 145 and >2092 days, respectively. Significant negative multivariate prognostic factors included macroscopic disease and reduced vinblastine (VBL) treatments for PFST, and presence of MCT in bone marrow analysis, Patnaik grade III MCT and reduced VBL treatments for OST. Toxicity was infrequent and self-limiting. This study suggests that the VCP protocol should be considered as an option in the treatment of MCT in dogs.

Clinicopathological findings and results of bone marrow aspiration in dogs with cutaneous mast cell tumours: 157 cases (1999-2002).

Bone marrow aspiration for routine staging of canine cutaneous mast cell tumour is not consistently performed, and the overall incidence of bone marrow infiltration and predictive value of the complete blood count (CBC) is unknown. This study evaluated a series of 157 dogs presented for cutaneous mast cell tumours in which a CBC and bone marrow aspiration were performed. The incidence of bone marrow infiltration at initial staging was low at 2.8%, and 4.5% overall. Factors significantly associated with bone marrow infiltration included increased age, leucocytosis, anaemia, neutrophilia, monocytosis, eosinophilia, thrombocytopenia, being purebred and staging at the time of recurrent or progressive disease. Our study suggests that a bone marrow sample is not indicated for routine staging but maybe indicated for those dogs with mast cell tumours having either an abnormal haemogram (neutrophilia, monocytosis, eosinophilia, basophilia, anaemia and thrombocytopenia) or presenting for tumour regrowth, progression or new occurrence.

Response of canine cutaneous epitheliotropic lymphoma to lomustine (CCNU): a retrospective study of 46 cases (1999-2004).

BACKGROUND: Epitheliotropic lymphoma (ELSA) is an uncommon cutaneous canine malignancy of T lymphocytes. A consensus regarding the therapeutic standard of care is lacking, warranting evaluation of chemotherapeutic agents traditionally employed against canine nodal lymphoma in the treatment of ELSA. HYPOTHESIS: The purpose of this retrospective, multi-institutional study was to evaluate the efficacy of 1-(2-chloroethyl)-3-cyclohexyl-l-nitrosourea (CCNU) in the treatment of ELSA. ANIMALS: Forty-six dogs with adequate follow-up and treatment response information. METHODS: All cases were diagnosed histopathologically. Immunohistochemisty (CD3, CD79a) was performed on 42/46 samples. RESULTS: Presenting skin lesions included generalized scales (25/46), plaques or nodules (22/46), mucocutaneous lesions (14/ 46), and corneal involvement (1/46). Lymph node involvement and Sézary syndrome were documented in 7 and 2 dogs, respectively. The median number of CCNU treatments was 4 (range, 1-11), with a median starting dose of 60 mg/m(2) (range, 30-95). Of the 46 dogs, 15 achieved complete remission, 23 achieved partial remission, 5 had stable disease, and 3 had progressive disease, for an overall response rate of 83%. The median number of treatments to achieve a response was 1 (range, 1-6). The overall median duration of response was 94 days (range, 22-282). Sixteen dose reductions were required because of neutropenia (10/46), thrombocytopenia (1/46), anemia (1/46), increased liver enzyme activity (3/46), or unspecified reasons (1/46). CONCLUSIONS AND CLINICAL IMPLICATIONS: Given the high response rate and well tolerated protocol, prospective studies are warranted to investigate the utility of CCNU alone or in multi-agent protocols for the treatment of ELSA.

Prognostic significance of intratumoral microvessel density in canine soft-tissue sarcomas.

The prognosis of canine soft-tissue sarcomas (STS) has traditionally been based on histologic grading. We have recently demonstrated the prognostic value of cellular proliferation markers in canine STS. Another method of predicting the behavior of neoplasms is intratumoral microvessel density (IMD), which is a measure of tumor angiogenesis. The prognostic significance of IMD has been documented in many human neoplasms and in a limited number of canine and feline neoplasms. To evaluate the prognostic value of IMD in canine STS, we studied 57 STS and compared IMD with histologic features, histologic grade, cellular proliferation, metastatic propensity, and survival. Using immunohistochemistry, the STS were labeled with anti-factor VIII-related antigen (FVIII-RA) and anti-CD31 antibodies to determine 3 IMD parameters: mean microvessel density, high microvessel density, and microvessel area. Using FVIII-RA and CD31, increasing IMD was statistically associated with increasing histologic grade, necrosis scores, and mitotic scores. Higher FVIII-RA IMD values were significantly associated with higher median argyrophilic nucleolar organizing region (AgNOR) values (as previously investigated) and increased metastatic propensity. Fibrosarcomas appear to be the least vascularized of STS. There is no correlation between IMD and survival. Our results indicate that IMD is of prognostic value for histologic grade, histologic features, cellular proliferation (based on AgNOR), and metastatic propensity of canine STS, specifically when using FVIII-RA as the endothelial marker. Assessing histologic grading, cellular proliferation, and IMD of canine STS at the time of diagnosis could therefore provide better prognostic information for the veterinary clinician.

P-glycoprotein expression in lamina propria lymphocytes of duodenal biopsy samples in dogs with chronic idiopathic enteropathies.

P-glycoprotein (p-gp) is a transmembrane protein functioning as a drug-efflux pump in the intestinal epithelium. Human patients with inflammatory bowel disease (IBD) who fail to respond to treatment with steroids express high levels of p-gp in lamina propria lymphocytes. The purpose of this study was to investigate p-gp expression in duodenal biopsy samples of dogs with chronic enteropathies and to evaluate the expression of p-gp after treatment with a known inducer of p-gp (prednisolone). Duodenal biopsy samples from 48 dogs were evaluated immunohistochemically with the mouse monoclonal antibody C219 for expression of p-gp in lamina propria lymphocytes. Biopsy samples were available from 15 dogs after treatment with prednisolone and 16 dogs after dietary therapy alone ("elimination diet"). Treatment with prednisolone resulted in an increase in p-gp expression (P=0.005). In contrast, dietary treatment alone produced no significant change in p-gp expression (P=0.59). A low p-gp score before initiation of steroid treatment was significantly associated with a positive response to treatment (P=0.01). These results indicate that lamina propria lymphocyte expression of p-gp is upregulated after prednisolone treatment in dogs with IBD, and that mucosal expression of p-gp may be of value in predicting the response to therapy.

Development of a xenogeneic DNA vaccine program for canine malignant melanoma at the Animal Medical Center.

INTRODUCTION: Canine malignant melanoma (CMM) is an aggressive neoplasm treated with surgery and/or fractionated RT; however, metastatic disease is common and chemoresistant. Preclinical and clinical studies by our laboratory and others have shown that xenogeneic DNA vaccination with tyrosinase family members can produce immune responses resulting in tumor rejection or protection and prolongation of survival. These studies provided the impetus for development of a xenogeneic DNA vaccine program in CMM. MATERIALS AND METHODS: Cohorts of three dogs each received increasing doses of xenogeneic plasmid DNA encoding either human tyrosinase (huTyr; 100/500/1500 mcg), murine GP75 (muGP75; 100/500/1500 mcg), murine tyrosinase (muTyr; 5 dogs each at 100/500 mcg), muTyr+/-HuGM-CSF (9 dogs at 50 mcg muTyr, 3 dogs each at 100/400/800 mcg HuGM-CSF, or 3 dogs each at 50 mcg muTyr with 100/400/800 mcg HuGM-CSF), or 50 mcg MuTyr intramuscularly biweekly for a total of four vaccinations. RESULTS: The Kaplan-Meier median survival time (KM MST) for all stage II-IV dogs treated with huTyr, muGP75 and muTyr are 389, 153 and 224 days, respectively. Preliminarily, the KM MST for stage II-IV dogs treated with 50 mcg MuTyr, 100/400/800 mcg HuGM-CSF or combination MuTyr/HuGM-CSF are 242, 148 and >402 (median not reached) days, respectively. Thirty-three stage II-III dogs with loco-regionally controlled CMM across the xenogeneic vaccine studies have a KM MST of 569 days. Minimal to mild pain was noted on vaccination and one dog experienced vitiligo. We have recently investigated antibody responses in dogs vaccinated with HuTyr and found 2- to 5-fold increases in circulating antibodies to human tyrosinase. CONCLUSIONS: The results of these trials demonstrate that xenogeneic DNA vaccination in CMM: (1) is safe, (2) leads to the development of anti-tyrosinase antibodies, (3) is potentially therapeutic, and (4) is an attractive candidate for further evaluation in an adjuvant, minimal residual disease Phase II setting for CMM.

Canine hepatic neuroendocrine carcinoma: an immunohistochemical and electron microscopic study.

Ten dogs with neuroendocrine carcinoma of the liver were selected for inclusion in the study. Clinical signs were anorexia (7), vomiting (5), polydipsia/polyuria (3), icterus (2), lethargy (2), weight loss (2), paresis (1), ataxia (1), weakness (1), collapse (1), and urinary tract infection (1). Hematologic and biochemical abnormalities included anemia (2/8), leukocytosis (4/8), high liver enzyme activity (serum alkaline phosphatase, 7/9; alanine transaminase, 7/9; aspartate transaminase, 8/9), and high total bilirubin (6/9). Grossly, the tumors were diffuse, involving all liver lobes in six dogs, and two dogs had various-sized nodules in addition to diffuse involvement. Histologically, there were eight tumors with solid or trabecular pattern (group A), one tumor with cords or rows of neoplastic cells (group B), and one tumor with multiple rosette-like structures (group C). Immunohistochemical studies revealed that all 10 neoplasms were positive for at least one of the endocrine markers used: neuron-specific enolase (NSE; 8/10), synaptophysin (5/10), and chromogranin-A (3/10). A panel of NSE, chromagranin-A, and synaptophysin detected 100% of the tumors in our series. Electron microscopy confirmed the diagnosis by the presence of intracytoplasmic neurosecretory granules in the two examined cases. Our results show that neuroendocrine markers commonly used in humans can be used for the diagnosis of hepatic neuroendocrine carcinoma in dogs, preferably a panel of synaptophysin, chromagranin-A, and NSE because chromogranin-A alone is not as useful in dogs as in humans.

Evaluation of intracavitary mitoxantrone and carboplatin for treatment of carcinomatosis, sarcomatosis and mesothelioma, with or without malignant effusions: A retrospective analysis of 12 cases (1997-2002)*.

Abstract Carcinomatosis, sarcomatosis and mesothelioma, with or without malignant effusions, are difficult to treat and generally carry a poor prognosis. The purpose of this study was two-fold; first, to determine the prognosis for dogs with carcinomatosis, sarcomatosis, or mesothelioma, with or without malignant effusions; second, to evaluate the safety and efficacy of treatment with intracavitary (IC) carboplatin and mitoxantrone in dogs with these syndromes. Nineteen dogs were evaluated. Seven were untreated and 12 were treated with IC chemotherapy (mitoxantrone and/or carboplatin), and multiple factors were analysed for significance with respect to survival time. The median survival time (MST) for untreated dogs was 25 days, whereas the MST for treated dogs was 332 days (Log Rank, P < 0.0001). Treatment with IC chemotherapy was well tolerated. This study suggests that IC chemotherapy with mitoxantrone and/or carboplatin is an effective treatment for dogs with carcinomatosis, sarcomatosis or mesothelioma, with or without malignant effusion.

Immunohistochemical detection of HER-2/neu expression in spontaneous feline mammary tumours.

Mammary gland tumours (MGT) are the third most common tumours in the cat. At least 85% are malignant and metastasis is common. The HER-2/neu protooncogene encodes a 185-kDa transmembrane tyrosine receptor kinase protein. Approximately 25-30% of human MGT demonstrate HER-2/neu protein overexpression in the malignant cells, and overexpression has been associated with an increased metastatic propensity and a decreased prognosis. No reports have been published, to date, investigating the expression of Her-2/neu in cats or cats with spontaneous mammary tumours. Based on the increased percentage of malignant mammary cancers in cats, compared to that in dogs, and the correlation of an increased malignancy and a decreased prognosis with Her-2 overexpression in human mammary cancer, we hypothesized that cats with spontaneous malignant mammary adenocarcinoma overexpress Her-2/neu in the neoplastic mammary epithelial cells. Thirty cats with MGT were assayed for Her-2/neu immunohistochemical expression. The median percentage of cells from feline MGT expressing Her-2/neu by utilizing the Dako polyclonal and CB11 monoclonal antibodies was 85 and 92.5, respectively. Her-2/neu expression intensity grades 2 and 3 consistent with the overexpression by utilizing the Dako polyclonal and CB11 monoclonal antibodies were observed in 90 and 76.7% of cats with MGT, respectively. The level of overexpression concordance across the two antibodies was 70%. The results from this study suggest that Her-2/neu overexpression is common in cats with spontaneous MGT, and therefore appears to represent an excellent model for Her-2/neu-overexpressing human breast cancer.

Characterization of spindle cell component of ferret (Mustela putorius furo) adrenal cortical neoplasms - correlation to clinical parameters and prognosis.

Adrenal cortical epithelial tumours are common in ferrets. A variant tumour type with prominent spindle cell proliferation has been identified. We characterized these variant tumours with light microscopy and immunohistochemical analysis and correlate these features to clinical parameters and prognosis. We classified 24 ferret adrenal cortical masses with recognizable spindle cell proliferation obtained from the AMC and AFIP databases, based on percentage of spindle cells present and features of malignancy. These masses were separated into hyperplastic nodules and adenomas (both with 1-24% spindle cells), 'mixed' adenomas (>or=25% spindle cells), adenocarcinomas (1-24%) and 'mixed' adenocarcinomas (>or=25% spindle cells). Tumours were evaluated immunohistochemically for smooth muscle actin (SMA) and estrogen receptor (ER) expression. Disease-free interval (DFI) and survival time (ST) were calculated using Kaplan-Meier product limit method. Of 24 cases of spindle cell variant adrenal tumours, one was a hyperplastic nodule, 10 were adenomas, three were 'mixed' adenomas, six were adenocarcinomas and four were 'mixed' adenocarcinomas. The proliferative spindle cell cytoplasm was SMA-positive (smooth muscle myocyte origin). ER positivity, seen in nine of 24 cases, was restricted to adenocarcinomas, 'mixed' adenomas and 'mixed' adenocarcinomas. DFI and ST were significantly reduced in 'mixed' adenocarcinomas or tumours with ER expression. DFI was significantly reduced in tumours with marked smooth muscle. The spindle cell component of these variant adrenal tumours is smooth muscle in origin. The presence of abundant smooth muscle, a more malignant histologic grade ('mixed' adenocarcinomas) and ER expression are significantly positively correlated to both decreased DFI and decreased ST.

Prognostic factors in dogs with urinary bladder carcinoma.

Medical records and biopsy specimens were retrospectively reviewed from 25 dogs diagnosed with unresectable urinary bladder carcinoma and treated with chemotherapy. Our intention was to identify clinical, histologic, and immunohistochemical indicators of prognosis. Immunohistochemical stains for P-glycoprotein, glutathione-S-transferase pi, and factor VIII-related antigen were applied to archived tissue. There were more spayed female dogs than castrated male dogs (76% versus 24%). Transitional cell carcinoma was the most common tumor (88%, n = 22), followed by undifferentiated carcinoma (8%, n = 2) and squamous cell carcinoma (4%, n = 1). Overall median survival was 251 days. Histologic diagnosis and immunohistochemical characteristics did not correlate with prognosis. Spayed females survived significantly longer than castrated males (358 days versus 145 days, P = .042). Dogs that received either doxorubicin or mitoxantrone in addition to a platinum-based chemotherapeutic (either cisplatin or carboplatin) lived significantly longer than those that received only a platinum compound (358 days versus 132 days, P = .042).

An N-myristoylated protein kinase C-alpha pseudosubstrate peptide that functions as a multidrug resistance reversal agent in human breast cancer cells is not a P-glycoprotein substrate.

Protein kinase C-alpha (PKC-alpha) activation is an important contributing factor in human breast cancer MCF-7 MDR cell drug resistance. We recently reported the use of N-myristoylated PKC-alpha pseudosubstrate peptides with potent PKC-alpha inhibitory activity as reversal agents of drug resistance in MCF-7 MDR cells. The peptides potently inhibit phosphorylation of the PKC-alpha substrates P-glycoprotein (P-gp), raf kinase and PKC-alpha itself in MCF-7 MDR cells in association with a severalfold induction of intracellular uptake of P-gp substrate chemotherapeutics and a statistically significant twofold increase in cellular chemosensitivity. We now report that the N-myristoylated PKC-alpha pseudosubstrate peptide N-myristoyl-RFARKGALRQKNV (P3) is not a P-gp substrate in MCF-7 MDR cells based on a comparison of the cellular uptake of [125I]-radiolabeled P3 in MCF-7 MDR vs MCF-7 WT cells. The extent of cellular uptake of the radiolabeled peptide in the drug-resistant cell line MCF-7 MDR was either greater than or equivalent to the uptake in the parental drug-sensitive MCF-7 WT cell line over a time course of 30 min to 6 h, and across a peptide concentration range of 25-100 microM. Additionally, treatment of the MCF-7 MDR cells with verapamil (VPL), a known P-gp efflux inhibitor, had no effect on the cellular accumulation of radiolabeled P3. Our results provide direct evidence that the N-myristoylated pseudosubstrate peptide is taken up equivalently by drug-sensitive and MDR cancer cells and therefore has potential value as an MDR reversal agent that operates by a novel mechanism.