Anti-Cancer Effects of Endocannabinoids in Canine Non-Hodgkin Lymphoma

Saba Omer¹, Maninder Sandey², Mohammed Mansour¹, Satyanarayana Pondugula¹ , Sindhu Ramesh⁴, Muralikrishnan Dhanasekaran⁴, Brad Matz³, Dawn Boothe¹

¹ Department of Anatomy, Physiology, & Pharmacology, ²Department of Pathobiology,

³Department of Clinical Sciences, College of Veterinary Medicine, Auburn University, Auburn, Alabama, USA

⁴Department of Drug Discovery and Development, Harrison School of Pharmacy, Auburn University, Auburn, Alabama, USA

Abstract

The role of the endocannabinoid system in cancer physiology is currently a matter of active debate.  However, generally, the endocannabinoid system is upregulated in cancer tissues compared with non-tumor tissues which suggest the pro-tumorigenic nature of the system. On the other hand, in the last two decades, many in vitro and in vivo studies have shown to induce cancer cell death on activation of cannabinoid receptors with endogenous and exogenous cannabinoids. These contrasting observations indicate further need for investigations to understand the exact role of the endocannabinoid system in cancer, if it is one of antitumor effects, the precise mechanisms by which cannabinoids accomplish this. The purpose of this research was to explore, for the first time, the effects of cannabinoids on canine non-Hodgkin lymphoma (NHL) cell lines and activated PBMCs. Specifically, the goal was to study in detail the effects of endocannabinoids, AEA, and 2AG, on canine NHL cell viability. Non-Hodgkin lymphoma is one of the leading causes of cancer-associated mortality in humans and canines. In this study, we have used canine B cell type (1771 and CLBL1) and T cell type (CL1) NHL cell lines and healthy canine activated PMBCs. Cells were cultured in RPMI, and the expression of endocannabinoid receptors (CB1 and CB2) was studied using conventional endpoint PCR and quantitative PCR. To study the effects of AEA and 2-AG, cells were treated at concentrations from 100 nM to 50µM for 24 and 48 hours. The vehicle was used as a control. Cell viability was assessed using colorimetric MTT assay (3-(4,5-Dimethylthiazol-2-yl)-2,5 Diphenyltetrazolium Bromide) and oxidative stress, inflammation, mitochondrial function, and apoptotic markers of cell death were assessed using spectrophotometric and fluorometric analysis. Data were analyzed using one-way ANOVA, student t-test, and regression analysis with Prism® and SAS® statistical software.  Our results suggest that canine B cell lymphoma shows higher expression of cannabinoid receptors compare to canine T cell lymphoma and healthy canine PBMCs and endocannabinoids have an anti-proliferative and pro-apoptotic effect on canine NHL cells. These results support the need for further studies providing evidence of efficacy against canine lymphomas. 

Acknowledgements

We are thankful to Dr. Steven Suter for sharing Canine lymphoma cell lines with us. We are also thankful to the following students for helping us with biochemical analysis Suhrud Pathak, Dylan Bowen and Rishi Nadar.

Keywords

Lymphoma, Cannabinoid, Anti-cancer