Epigenetics of canine lung cancer development

Abstract

Background: Human lung cancer is a leading cause of cancer related mortality and has significant economic impact. Cigarette smoking and exposure to second-hand smoke are a common cause of lung cancer development. Lung cancer is fairly uncommon in dogs despite sharing similar environments, being exposed to second-hand smoke, and having similar respiratory physiology. Understanding the cause of apparent species protection could result in new prevention and treatment avenues for man and dog. One way of investigating species differences is investigation the methylome and protein expression of common cancer related genes in the dog and comparing those to know findings in man. We elected to study of the methylation status of a panel of canine genes known to be deregulated in human cancer including CDA, DLX-1-3', DLX1-5', FOXB2, HOXA9, HOXB5, and PPAR-[gamma]. During the investigation of these genes in canine lung cancer, and interesting methylation profile was discovered for PPAR-[gamma]. PPAR-[gamma] is a ligand-dependent transcription factor that plays important roles in cellular proliferation and differentiation. It has been implicated as a tumor suppressor in many solid tumors including human prostate, breast, colon, and lung cancer. Our initial findings of an altered methylome of PPAR-[gamma] prompted further investigation of the tissue distribution of PPAR-[gamma] in normal canine lung, canine lung cancer, and metastatic to lung cancer. We also went on to investigate the role of DNA methylation on control of gene expression. The protein was studied using immunohistochemistry (IHC) and DNA methylation was studied using combined bisulfite restriction analysis (COBRA), and methylation-specific PCR (MSP). Results: PPAR-[gamma] is expressed in all large conducting airways, particularly in goblet cells and bronchial glands, in the canine lung. The protein is also expressed in interstitial macrophages. PPAR-[gamma] is expressed in 33% of canine non-small cell lung cancer (NSCLC) cases and 66% of metastatic osteosarcoma (OSA) cases. There is a significant loss of 5' PPAR-[gamma] methylation from normal lung to primary lung cancer and metastatic OSA (p=0.0002), however altered PPAR-[gamma] promoter methylation at the interrogated locus does not appear to be associated with changes in protein expression. Conclusions: PPAR-[gamma] protein is expressed in normal canine lung tissue, canine primary lung cancer, and metastatic OSA. Confirmation of PPAR-[gamma] protein expression in tumor-bearing dogs supports the investigation of PPAR-[gamma] agonists in this subset of veterinary patients. These results are the first to describe epigenetic marks and protein localization of PPAR-[gamma] among different lung pathologies in the dog.