Rowan J. Milner

Cisplatin: a review of toxicities and therapeutic applications.

Cisplatin is a platinum chemotherapeutic used in a variety of malignancies. The antineoplastic activity occurs from DNA cross-links and adducts, in addition to the generation of superoxide radicals. Nephrotoxicity is the most well-known and potentially most clinically significant toxicity. Unfortunately, the mechanism for cisplatin nephrotoxicity has not been completely elucidated; however, many theories have been developed. Other toxicities include gastrointestinal, myelosuppression, ototoxicity and neurotoxicity. Saline diuresis is currently the most accepted way to prevent cisplatin nephrotoxicity. Research has focused on pharmaceuticals and enzyme/molecular alterations as alternatives to long-term diuresis. No agents have currently been identified that can protect from all toxicities. Cisplatin has shown activity against osteosarcoma, transitional cell carcinoma, squamous cell carcinoma (SCC), melanoma, mesothelioma, carcinomatosis and germinal cell tumours in the dog. In the cat, cisplatin cannot be utilized because of fulminant pulmonary oedema that occurs at standard doses. Intralesional cisplatin has been utilized in horses for the treatment of SCC and sarcoids.

Radical excision with five-centimeter margins for treatment of feline injection-site sarcomas: 91 cases (1998–2002)

OBJECTIVE To evaluate outcomes of radical excision of feline injection-site sarcomas (ISS) via assessment of local recurrence and metastasis rates, survival times, and complications associated with surgery. DESIGN Retrospective case series. ANIMALS 91 cats with ISS. PROCEDURES Medical records of cats that had radical excision of ISS without adjunctive treatment were reviewed. Information extracted included sex, type of surgical procedure, histologic tumor grade, tumor diameter, time from tumor detection to definitive surgery, complications associated with surgery, whether tumors recurred locally or metastasized, and survival times. Diagnosis of ISS was histologically confirmed, and additional follow-up was performed. RESULTS Overall median survival time was 901 days. Thirteen of 91 (14%) cats had local tumor recurrence; 18 (20%) cats had evidence of metastasis after surgery. Median survival time of cats with and without recurrence was 499 and 1,461 days, respectively. Median survival time of cats with and without metastasis was 388 and 1,528 days, respectively. Tumor recurrence and metastasis were significantly associated with survival time, whereas other examined variables were not. Major complications occurred in 10 cats, including 7 with incisional dehiscence. CONCLUSIONS AND CLINICAL RELEVANCE Radical excision of ISS resulted in a metastasis rate similar to rates reported previously; the local recurrence rate appeared to be substantially less than rates reported after less aggressive surgeries, with or without adjuvant treatment. Major complication rates were similar to rates reported previously after aggressive surgical resection of ISS. Radical excision may be a valuable means of attaining an improved outcome in the treatment of feline ISS.

Comparison of Canine Cardiac Troponin I Concentrations as Determined by 3 Analyzers

BACKGROUND Recent interest in cardiac biomarkers has led to the validation of several commercial analyzers for cardiac troponin I (cTnI) evaluation in dogs; however, these analyzers have not been standardized. HYPOTHESIS It was hypothesized that canine plasma cTnI concentrations as determined by 3 different analyzers would be similar. ANIMALS Twenty-three dogs with cardiac disease were studied. METHODS Reconstituted purified canine free cTnI was diluted with canine plasma to 8 concentrations (0.01, 0.1, 0.78, 1.56, 3.13, 6.25, 12.5, and 25 ng/mL), for analysis by 3 analyzers, the Biosite Triage Meter, the Dade-Behring Stratus, and the Beckman-Coulter Access AccuTnI. Plasma samples from 23 dogs with cardiac disease were also analyzed for cTnI concentrations on all analyzers. RESULTS Troponin I concentrations in sick dogs were <0.05-5.72 ng/mL (Biosite), 0.02-11.1 ng/mL (Access), and 0.02-9.73 ng/mL (Stratus). Analyzer results were highly correlated with each other (r = 0.97 to 1.0 for purified dilutions, r = 0.61 to 0.89 for samples from dogs); however, the limits of agreement were wide for both purified dilutions and clinical samples. Recovery was highest for the Access (334-1467%) and lowest for the Biosite (38-60%); Stratus 52-233%. Analyzer variability was lowest for the Access (1.2-10.4%) and highest for the Stratus (4.8-33.6%); Biosite 2.8-16.5%. CONCLUSIONS AND CLINICAL IMPORTANCE Results from this study suggest that although canine cTnI values obtained from the Biosite, Stratus, and Access analyzers are closely correlated, they cannot be directly compared with each other. In the absence of a gold standard none of the analyzers can be considered more correct than the others.

Radiation therapy for canine appendicular osteosarcoma.

Radiation therapy (RT) for the management of canine appendicular osteosarcoma (OSA) can be described as either palliative- or curative intent. Palliative RT uses coarsely fractionated external beam RT or radiopharmaceuticals to provide relief of pain and lameness associated with OSA while resulting in minimal, if any, radiation-induced acute adverse effects. Limb amputation and chemotherapy are considered (together) the standard of care for curative-intent treatment of canine appendicular OSA. When limb amputation is not possible, RT can be used for limb sparing and is supplemented with chemotherapy for presumed micrometastatic disease. Fractionated tumour irradiation with curative intent appears to be ineffective and local disease control can more likely be achieved when stereotactic radiosurgery or intra-operative extracorporeal irradiation is combined with strict case selection and adjunctive chemotherapy. The availability of limb-sparing RT is limited by experience and availability of specialised equipment. When planned and administered appropriately, radiation-associated adverse effects are often mild and self-limiting.

Immunophenotypic and cytomorphologic subclassification of T-cell lymphoma in the boxer breed.

The boxer breed is at high risk for developing lymphoma and, in contrast to the general canine population, is predisposed to the T-cell variant of the disease. The purpose of this study was to more accurately classify lymphoma in this breed. Clinical, cytomorphologic and immunophenotypic data were examined in 43 boxers with lymphoma. Twenty-five cases were collected prospectively and a further 18 cases were obtained retrospectively. Lymphomas were classified as multicentric (n=29), mediastinal (n=6) and intestinal (n=8). Of the 40 immunophenotyped samples, 34 (85%) were T-cell, 5 (12.5%) were B-cell and 1 was a non-B-cell non-T-cell lymphoma. Immunophenotypic subtyping was done on prospectively collected T-cell lymphoma samples (n=22) to differentiate CD4 (helper) from CD8 (cytotoxic) T-cell origin as well as to determine the T-cell receptor (TCR) expression (TCRalphabeta vs. TCRdeltagamma). Phenotypic expression was CD4+ (n=12), double negative (DN) (n=6), double positive (DP) (n=1) and CD8+ (n=1), respectively, while two samples had no interpretable result. 20/22 samples were TCRalphabeta+ with only 1 sample being TCRdeltagamma+ and 1 with no interpretable result. Cytomorphologic analysis was done on the same 22 samples using the World Health Organization (WHO) classification scheme. According to this scheme, 17/22 samples were classified as lymphoblastic, 2/22 as large cell peripheral T-cell lymphoma-not otherwise specified (PTCL-NOS), 2/22 as large granular lymphoma (LGL) high-grade and 1/22 as small lymphocytic. The results of this study indicate that lymphoma in the boxer breed is a disease comprised predominantly of TCRalphabeta+, CD4+ (helper) T-cells with lymphoblastic (high-grade) morphology.

Frameless stereotactic radiosurgery for the treatment of primary intracranial tumours in dogs

Stereotactic radiosurgery (SRS) is a procedure that delivers a single large radiation dose to a well-defined target. Here, we describe a frameless SRS technique suitable for intracranial targets in canines. Medical records of dogs diagnosed with a primary intracranial tumour by imaging or histopathology that underwent SRS were retrospectively reviewed. Frameless SRS was used successfully to treat tumours in 51 dogs with a variety of head sizes and shapes. Tumours diagnosed included 38 meningiomas, 4 pituitary tumours, 4 trigeminal nerve tumours, 3 gliomas, 1 histiocytic sarcoma and 1 choroid plexus tumour. Median survival time was 399 days for all tumours and for dogs with meningiomas; cause-specific survival was 493 days for both cohorts. Acute grade III central nervous system toxicity (altered mentation) occurred in two dogs. Frameless SRS resulted in survival times comparable to conventional radiation therapy, but with fewer acute adverse effects and only a single anaesthetic episode required for therapy.

THE EFFECT OF THE BISPHOSPHONATE ALENDRONATE ON VIABILITY OF CANINE OSTEOSARCOMA CELLS IN VITRO

Abstract The objective of this study was to determine the effect of alendronate on the viability of canine osteosarcoma cells and nonneoplastic canine cells. The sample population was composed of canine osteosarcoma tumor cells. Osteosarcoma cells and canine fibroblasts were maintained in culture under standard conditions. The MTT assay for cell viability was performed after 24, 48, and 72 h of incubation with alendronate (0.001 to 1000 μM) or no drug (control). Plates were set up so that each concentration and the control had a sample number of 8. The optical density (OD) of each well was measured at 540 nm using an enzyme-linked immunosorbent assay microplate reader. The percent viability was determined for each concentration and for each incubation time. After 24 h of incubation of POS (parent osteosarcoma) and HMPOS cells with alendronate, there was no significant difference in mean OD at any drug concentration when compared with control samples. A significant concentration- and time-dependent reduction in mean OD of osteosarcoma cells was observed after 48 and 72 h of incubation, with alendronate concentrations ranging from 10 to 1000 μM. The lowest percent cell viability observed in treated cells was 35%. Conversely, alendronate did not significantly affect mean OD in fibroblasts, and the lowest percent cell viability observed was 76%. Our data indicate that alendronate may have the potential to inhibit canine osteosarcoma tumor growth. It will be important to determine the clinical relevance of these in vitro findings. If similar findings are observed in vivo, use of alendronate may also be indicated as an adjuvant to existing chemotherapeutic protocols.SummaryMosquito densonucleosis viruses (MDVs) have the potential for use as biocontrol agents. To facilitate densovirus production, the Aedes albopictus mosquito cell line C6/36 was adapted to two commercially available serum-free protein-free media (SFPFM), Sf-900 II and Drosophila-SFM. Cells adapted more slowly to growth in Sf-900 II medium, but once adapted, they grew more rapidly and appeared healthier than cells growing in Drosophila-SFM. Cells that were adapted to growth in each of these SFPFM were tested for their ability to be transfected and infected with MDVs. The Sf-900 II-adapted cell line survived transfection and showed infection rates comparable with cells growing in L15 supplemented Cells adapted to each of these SFPFM were adapted to growth in spinner flasks. Cells in Sf-900 II grew substantially better in spinner flasks than cells in Drosophila-SFM media. Cells grown in Sf-900 II could be frozen and, when thawed, could support the production of densonucleosis viruses in spinner flasks.

Canine Anatomic Phantom for Preclinical Dosimetry in Internal Emitter Therapy

The majority of investigational studies of new diagnostic and therapeutic radiopharmaceuticals use murine animal models for preclinical assessments of pharmacokinetics and organ radiation dosimetry. Although mice and rats are widely available and relatively inexpensive, their smaller organ anatomy relative to that of humans can lead to considerable differences in organ dosimetry, thus complicating extrapolations of dose–response relationships to human patients. Nonhuman primates circumvent these problems in many respects but are increasingly becoming expensive and limited because of ethical considerations. With the recent completion of the dog genome project and the recognition of many similarities between canine and human cancers, dogs are increasingly being considered in cancer research and drug development. The main objective of this study was to construct a 3-dimensional computational phantom of a large dog on the basis of whole-body multislice CT data. Methods: A female hound cross underwent whole-body contrast-enhanced CT at a 2-mm slice thickness. On completion of the scan, the dog was euthanized, and the entire skeleton was harvested for a subsequent microCT investigation. The CT data were imported into a computational software program and used to create a polygon-mesh phantom of the entire animal. All of the major organs and bones were semiautomatically segmented and tagged to the CT slices. The phantom data were imported into a second software program and transformed to a nonuniform rational basis-spline surface phantom, allowing easy alteration of the phantom to simulate dogs of smaller or larger statures. A voxel-based version of the canine phantom was created by use of an in-house routine for subsequent import into the EGSnrc radiation transport code for photon and β-particle organ dosimetry. Results: The resulting voxel-based version of the canine phantom had a total body mass of 26.0 kg and a total body tissue mass (exclusive of wall organ content) of 24.5 kg. Although this University of Florida (UF) canine phantom displayed a total body mass intermediate between those of the Oak Ridge National Laboratory (ORNL) 5-y and 10-y stylized human phantoms of the MIRDOSE and OLINDA software codes, considerable differences were noted in organ photon cross-doses. For example, ratios of the specific absorbed fraction Φ(lungs ← liver)UF Dog to Φ(lungs ← liver)ORNL 5-y ranged from ∼30 at 10 keV to ∼3.5 at 1 MeV. Corresponding ratios of Φ(lungs ← liver)UF Dog to Φ(lungs ← liver)ORNL 10-y ranged from ∼6 at 10 keV to ∼1.3 at 1 MeV. Conversely, values of Φ(kidneys ← spleen) and Φ(liver ← spleen) were noted to be much lower (factors of 2–4) and much higher (factors of 2–15), respectively, in the canine phantom than in the ORNL human phantoms. These differences were attributed more to organ shape and position within the torso than to organ mass, because many of the canine organs closely approximated their counterparts volumetrically in the stylized pediatric human phantoms. Conclusion: The use of canine models, particularly in spontaneously occurring malignancies such as osteosarcoma, for preclinical testing of antineoplastic agents offers significant advantages over current murine models. However, the development of canine-specific technology is critical to the optimization of these studies. The UF canine dosimetry phantom described here aims to solve problems that could stem from the use of current human dosimetry models during radiopharmaceutical research.

Efficacy and toxicity of BOPP and LOPP chemotherapy for the treatment of relapsed canine lymphoma.

Mechlorethamine (Mustargen), Oncovin) (vincristine), procarbazine and prednisone (MOPP) chemotherapy is useful for relapsed canine lymphoma. This study evaluates the efficacy of MOPP after substitution of CCNU (lomustine, LOPP protocol) or BCNU (carmustine, BOPP protocol) for mechlorethamine in 60 dogs with relapsed lymphoma. Seven of 14 (50%) dogs treated with BOPP responded, for a median of 129.5 days for complete responders (range 9-354 days) and a median of 140 days for partial responders (range 4-276 days). Twenty-three of 44 (52%) dogs treated with LOPP responded for a median of 112 days for complete responders (range 48-250 days) and a median of 84.5 days for partial responders (range 69-290 days). Two dogs receiving a combination of LOPP and BOPP partially responded for 28 and 163 days, respectively. With BOPP chemotherapy, nine dogs (20.5%) and seven dogs (50%) had one or more episodes of Grade II or higher neutropenia and thrombocytopenia, respectively. Seven dogs (50%) had one or more episodes of Grade II or higher gastrointestinal toxicity. While receiving LOPP chemotherapy, 28 dogs (63.6%) and 17 dogs (38.6%) had one or more episodes of Grade II or higher neutropenia and thrombocytopenia, respectively. Seventeen dogs (38.6%) had one or more episodes of Grade II or higher gastrointestinal toxicity. Overall, there were 17 non-fatal treatment-related episodes of sepsis requiring hospitalization. Eight dogs (13%) died or were euthanized because of treatment-related sepsis and/or chemotherapy-related complications. Severe haematologic toxicity, coupled with the improved response duration observed in dogs receiving reduced doses during B/L-OPP rescue, underscores the need for protocol optimization.

MRI Measurement of Bone Marrow Cellularity for Radiation Dosimetry

The current gold standard for measuring marrow cellularity is the bone marrow (BM) biopsy of the iliac crest. This measure is not predictive of total marrow cellularity, because the biopsy volume is typically small and fat fraction varies across the skeleton. MRI and localized MR spectroscopy have been demonstrated as noninvasive means for measuring BM cellularity in patients. The accuracy of these methods has been well established in phantom studies and in the determination of in vivo hepatic fat fractions but not for in vivo measurement of BM cellularity. Methods: Spoiled gradient-echo in vivo images of the femur, humerus, upper spine, and lower spine were acquired for 2 dogs using a clinical 3-T MRI scanner. Single-peak iterative decomposition of water and fat with echo asymmetry and least squares (SP-IDEAL) was used to derive BM fat fractions. Stimulated-echo acquisition mode spectra were acquired in order to perform multipeak IDEAL with precalibration (MP-IDEAL). In vivo accuracy was validated by comparison with histology measurements. Histologic fat fractions were derived from adipocyte segmentation. Results: Bland–Altman plots demonstrated excellent agreement between SP-IDEAL and histology, with a mean difference of −0.52% cellularity and most differences within ±2% cellularity, but agreement between MP-IDEAL and histology was not as good (mean difference, −7% cellularity, and differences between 5% and −20%). Conclusion: Adipocyte segmentation of histology slides provides a measure of volumetric fat fraction (i.e., adipocyte volume fraction [AVF]) and not chemical fat fraction, because fat fraction measured from histology is invariant to the relative abundances of lipid chemical species. In contrast, MP-IDEAL provides a measure of chemical fat fraction, thus explaining the poor agreement of this method with histology. SP-IDEAL measures the relative abundance of methylene lipids, and this measure is shown to be equivalent to AVF. AVF provides the appropriate parameter to account for patient-specific cellularity in BM mass predictive equations and is consistent with current micro-CT–based models of skeletal dosimetry.