3.1 Patient selection
The medical records from client-owned dogs with high-risk cMCTs treated with VBL-based chemotherapy (including VCP and VP protocol) or imatinib at National Taiwan University Veterinary Hospital (NTUVH) Animal Cancer Treatment Center from 2011 to 2019 were retrospectively reviewed. The patients were grouped into the VBL group or the imatinib group. High-risk MCT was defined as histopathologically diagnosed Patnaik grade III or Kiupel high-grade MCT [3, 5], or histopathologically diagnosed Patnaik grade II or cytologically diagnosed MCT with the presence of cytologically confirmed nodal or visceral metastatic disease. Other criteria for inclusion in the study included measurable gross lesion and available medical records. Patients treated with concurrent antineoplastic treatment other than steroids, including chemotherapy, target therapy, and RT, were excluded. Prior chemotherapy or target therapy was allowed if more than three weeks had elapsed since the last treatment or progressive disease observed.
The following data were recorded for each patient: the institution of treatment, age at the time of diagnosis, breed, sex, body weight, tumor location, the sum of the longest diameter of target lesions, previous treatment, clinical stage and substage, and histological grade. Treatment information was also collected, including the number of doses, dose interval, dosage, response, and adverse events.
3.2 Clinical stage
All patients underwent clinical staging according to the WHO clinical staging system for MCTs [1] (Table 1). The results of physical examination, complete blood count,
serum biochemistry panel, blood smear, thoracic radiography, abdominal ultrasound, fine-needle aspiration of the regional lymph node, and/or fine-needle aspiration of the spleen were included. Cases with honeycomb appearance of the spleen would also be classified as stage 4 even without cytological confirmation.
3.3 Tumor grade
Tumors were classified as Grade I, Grade II, or Grade III according to the Patnaik grading system, or classified as low-grade or high-grade according to the Kiupel grading system [5] by the pathologist in School of Veterinary Medicine, National Taiwan University.
3.4 Treatments
3.4.1 VBL group
The patients in the VBL group received either VCP protocol or VP protocol, and the regimens depended on clinicians’ preference. In VCP protocol, VBL (Vinblastine Injection, Hospira) was administered as a rapid IV bolus at 2 mg/m2 to 3 mg/m2 every three weeks (on day 1 of the 21-day protocol). CTX (Endoxan, Baxter) was administered at 200 mg/m2 to 250 mg/m2 every three weeks orally (over day 8 to day 9 of the 21-day protocol). In VP protocol, VBL was administered as a rapid IV bolus at 2 mg/m2 to 3 mg/m2 every one to two weeks. Prednisolone was administered orally at a dosage range of 1 mg/kg/day to 2 mg/kg/day, then tapered over several weeks on a clinician-dependent basis.
3.4.2 Imatinib group
Imatinib (Gleevec, Novartis) was administered orally at a target dosage of 10 mg/kg once daily. Prednisolone was administered orally at a dosage range from 1 mg/kg/day to 2 mg/kg/day, then tapered over several weeks on a clinician-dependent basis.
3.5 Response
Antitumor responses were evaluated according to the Response Evaluation Criteria in Solid Tumors (RECIST) criteria [40]. Target lesions were measured with calipers at each recheck, and the sum of the longest diameters of each target lesion would be recorded in the case of multiple tumors. Responses were calculated by the formula listed below:
tumor response = [(post-treatment measurement – treatment measurement) / pre-treatment measurement] x 100%, then categorized as complete remission (CR; the disappearance of all target lesions and all lymph nodes <10 mm short axis), partial remission (PR; >30% but <100% decrease in the sum of diameters of target lesions), progressive disease (PD; at least a 20% increase in the sum of diameters of target lesions or appearance of a new lesion), stable disease (SD; neither sufficient shrinkage to qualify for PR nor sufficient increase to qualify for PD).
3.6 Toxicity
Adverse events were evaluated based on the results of the blood exam and patient history at each recheck. They were attributed to the study treatment if they occurred or increased in severity during or after exposure to the drug while no evidence of disease progression, showing dose-response patterns in the individual patient, and resolved with drug interruption [41]. Treatment-related adverse events were then graded from 1 to 5
based on the Veterinary Cooperative Oncology Group Common Terminology Criteria for Adverse Events (VCOG-CTCAE) [42] (Table 3).
3.7 Statistical analysis
To compare the demographic distribution and the tumor features between groups, we have conducted the following tests. Pearson’s chi-square test was used in categorical data including breed, sex, stage, substage, had received previous steroid treatment or not, previous systemic treatment, local recurrence or not, and tumor location. If the expected value of a given cell in the comparison was less than five, Fisher’s exact test was substituted for Pearson’s chi-square test. The Mann-Whitney U test compared continuous data including body weight, age, the sum of the longest diameter of target lesions, and median dosage of prednisolone.
ORR, CBR and PFI were defined as below: The ORR was defined as the percentage of patients that had experienced CR or PR The CBR was defined as the percentage of patients that had experienced CR, PR, or SD [40]. PFI was calculated from the date of treatment initiation to the date of PD.
To compare ORR and CBR between groups, Pearson’s chi-square test was used, but Fisher’s exact test was used instead if the expected value of a given cell in the comparison was less than five. Median PFI was calculated by the Kaplan-Meier method. Cases were censored if they had not developed PD at the time of data analysis or treatment withdrawn due to reasons unrelated to disease progression. The difference of median PFI between groups was assessed with the log-rank test.
The incidence of adverse events between groups was analyzed by Pearson’s chi-square test or by Fisher’s exact test if the expected value of a given cell in the comparison
was less than five.
Univariate analysis to assess potential prognostic factors in PFI was performed by the Kaplan-Meier method with the log-rank test. Factors included age, breed, sex, body weight, the sum of the longest diameter of the target lesions, local recurrence or not, tumor location, the median dosage of prednisolone, stage, substage, had received previous steroid treatment or not, previous systemic treatment, and objective response to the treatment. Factors found to be significant (P < 0.05) in univariate analysis were then included in multivariate analysis using the Cox proportional hazards model.
Statistical significance was established as P < 0.05. All statistical analyses were performed with SPSS v. 25 software.