Revisiting Ki-67 Assessment in Canine Mast Cell Tumours: From Manual Hotspot to Automated Global Analysis

Canine mast cell tumours (MCTs) show highly variable behaviour, and Ki-67 is an established prognostic indicator. Conventional Ki-67 assessment is manual and restricted to small hotspot areas, limiting reliability. This study presents a semi-automated whole-tumour tissue section (global) Ki-67 analysis workflow, outlines its limitations, and examines correlations with hotspot counts and clinical outcome. A total of 309 canine MCTs were assessed using a deep-learning-assisted quantification with commercial software. Global Ki-67 metrics were correlated with hotspot Ki-67 counts and histomorphologic tumour grades, as supported by clinical follow-up data from 68 dogs. The defined analytic workflow enabled an overall feasible global Ki-67 assessment in canine MCTs. The region-of-interest (ROI) definition required frequent manual adjustments, whereas Ki-67 quantification was fully automated and rapid. Global Ki-67 metrics correlated with manual hotspot counts, with Ki-67-positive cell density on average twice as high in tumour hotspots compared with whole tumour sections, with differences ranging up to 38-fold. Exploratory survival analyses suggested promising predictive power, warranting validation in a robust survival study. With established digital pathology tools, global whole-tumour assessment of Ki-67 and other biomarkers is feasible. It should become the new standard for defining robust prognostic and predictive markers in canine mast cell and other tumours.