Canine osteosarcoma (OSA) is the most common primary bone tumour in dogs. Chemotherapy delays metastasis, yet most dogs succumb within one year of diagnosis. Recent studies show that certain chemotherapeutics are sequestered within acidic endosomes, leading to chemoresistance. Furthermore, the high metabolic demand of cancer cells results in a preferential glycolytic shift that leads to tumor acidosis.
Amiloride, a potassium-sparing diuretic, is among a class of proton pump inhibitors prescribed by veterinary cardiologists for refractory heart failure. In this study, we hypothesized that amiloride sensitizes canine OSA cells to carboplatin and doxorubicin by reducing their sequestration within acidic endosomes.
Assessments of cell viability and apoptosis were performed in three OSA cell lines after single agent or combination treatment, and the synergism of each combination was evaluated with combination index (CI) calculations. Metabolic profiling of extracellular acidification rates (ECAR) and oxygen consumption rates (OCR) was performed, and immunoblotting was used to evaluate apoptotic protein expression.
Amiloride sensitized canine OSA cells to doxorubicin (CI < 1.0), but not carboplatin (CI ≥ 1.0) in combination therapy. Consistent with these findings, combination treatment with doxorubicin significantly increased late apoptosis (P < 0.0001), but decreased necrosis (P < 0.001) in OSA cells compared to treatment with doxorubicin alone. Cells treated with amiloride upregulated proteins involved in p53-mediated apoptosis and downregulated Akt-specific survival. Energetic stress tests further demonstrated metabolic switching to a less glycolytic phenotype with significant decreases in ECAR (P = 0.021).
The well-known safety profile of amiloride and its likelihood of synergism with doxorubicin may justify drug repurposing for clinical trial evaluation in canine OSA.
Ontario Veterinary College, University of Guelph
Thursday, June 14
4:30 PM – 4:45 PM
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