Osteosarcoma (OS) is the most common malignant bone tumor in dogs. Despite aggressive surgical management and systemic chemotherapy, 90% of dogs still die due to chemotherapeutic-resistant metastatic disease. As chemotherapy remains the backbone for treatment of OS metastases, the development of combinational treatments with novel targeted molecular therapeutics and conventional chemotherapy represents a potential strategy to enhance therapeutic response. Exportin 1 (XPO1, also known as CRM1) is a chaperone protein responsible for the export of >200 target proteins out of the nucleus. Dysregulation of XPO1 activity is documented in a number of human cancers and XPO1 expression has been associated with the development of chemotherapy resistance. Prior studies in canine melanoma and mammary carcinoma cell lines have demonstrated that XPO1 is a relevant target for therapeutic intervention and recent phase I and II clinical trials evaluating a novel, orally bioavailable Selective Inhibitor of Nuclear Export (SINE) Verdinexor in dogs with spontaneous cancer demonstrate anti-tumor activity against non-Hodgkin lymphoma. In the present study, we sought to characterize the expression of XPO1 in primary canine OS tumor samples, OS cell lines and normal osteoblasts and evaluate the in vitro efficacy of Verdinexor alone or in combination with doxorubicin in canine OS cell lines. Real time PCR and Western blotting was performed to assess XPO1 transcript and protein expression in normal canine osteoblast cells, canine OS cell lines and primary OS tumor tissues. Canine OS cell lines and a subset of primary OS tumors had increased expression of XPO1 mRNA and protein compared to normal canine osteoblasts. All canine OS cell lines exhibited dose-dependent growth inhibition and increased caspase 3,7 activity in response to low nanomolar concentrations of Verdinexor (IC50 concentrations ranging from 21-74 nM). Notably, growth inhibition of normal canine osteoblast cell lines treated with Verdinexor was only observed at high micromolar concentrations (IC50 21 µM). The combination of Verdinexor and doxorubicin resulted in potent inhibition of cell viability and demonstrated synergetic activity in three canine OS cell lines. Concordantly, OS cell lines showed increased DNA damage as assessed by γH2AX immunofluorescence following combination treatment with Verdinexor and doxorubicin compared to either single-agent. These findings demonstrate that Verdinexor has biologic activity against canine OS cell lines at physiologically relevant doses and suggest that XPO1 inhibition in combination with standard doxorubicin treatment offers promising potential for chemotherapeutic intervention in canine OS.
Gain an understanding of the functions of Exportin 1 (XPO1) in cancer pathogenesis and its dysregulation in canine cancers.
Review XPO1 inhibition as a therapeutic target and discuss novel, orally bioavailable selective inhibitors of nuclear export (SINE) in veterinary oncology.
Discuss recent data investigating XPO1 dysregulation in canine osteosarcoma and biological activity of Verdinexor against canine osteosarcoma cell lines.