Studies of cannabinoid receptor 1 in mantle cell lymphoma

Mantle cell lymphoma (MCL) is a malignant B-cell lymphoma that affects older individuals and has a male predominance. MCL has one of the worst prognoses among lymphomas and currently there is a search for a curative therapy. This thesis focuses on the possibility to induce cannabinoid receptor mediated cell death in MCL and other malignant lymphomas.

* The effects of cannabinoids on cell fate were investigated in MCL cell lines and patient samples. Nanomolar doses of cannabinoids did not induce growth of MCL cells. Instead, micromolar doses induced cell death and decreased viability and growth of MCL cells expressing the cannabinoid receptors CB1 and CB2 while control cells lacking expression of CB1 remained unaffected. Interestingly, at micromolar doses the cannabinoid receptor agonist anandamide and the antagonist SR141716 additively decreased viability.

* Signaling in MCL after treatment with the cannabinoids Win55 and methanandamide was investigated. The signaling was mediated via both CB1 and CB2 since blocking with nanomolar doses of antagonists prevented apoptosis. The signaling was mediated via de novo synthesis of the second messenger ceramide which caused phosphorylation of the MAP-kinase p38. This was followed by a disruption of the mitochondrial membrane potential and subsequently apoptosis. Signaling via the CB1 and CB2 receptors in MCL led to cell death while normal B-cells were spared.

* The expression of cannabinoid receptors in other B-cell lymphomas was investigated. Using quantitative real-time PCR it was found that 80% of the selected lymphoma samples expressed CB1 and/or CB2. The expression was confirmed at protein level by Western Blot and immunohistochemistry. Further, methanandamide treatment induced cell death in B-CLL cell lines. Importantly, we showed that methanandamide treatment reduced tumor burden in a MCL xenograft mouse model. Thus the cannabinoid system could be a potential target in malignant lymphoma.

* The ceramide metabolism in connection to cannabinoid treatment was studied. Pharmacological inhibition of the enzymes serine palmitoyl transferase, ceramide synthase or dihydroceramide desaturase prior to cannabinoid treatment led to disruption of ceramide synthesis and cell death in MCL. An upregulation of ceramide synthase 3 and 6 mRNA was observed after treatment with cannabinoids. Moreover, the effect of cannabinoid treatment on viability and cell death was potentiated by inhibiting the ceramide metabolizing enzymes sphingosine kinase-1 and glucosylceramide synthase.

In conclusion, MCL and a large percentage of other B-cell lymphomas express functional cannabinoid receptors that can mediate cell death specifically in malignant cells. Cannabinoid receptor mediated cell death in MCL could be potentiated by modulation of ceramide metabolism.

List of scientific papers

I. Flygare J, Gustafsson K, Kimby E, Christensson B, Sander B (2005). "Cannabinoid receptor ligands mediate growth inhibition and cell death in mantle cell lymphoma." FEBS Lett 579(30): 6885-9
https://pubmed.ncbi.nlm.nih.gov/16337199

II. Gustafsson K, Christensson B, Sander B, Flygare J (2006). "Cannabinoid receptor-mediated apoptosis induced by R(+)-methanandamide and Win55,212-2 is associated with ceramide accumulation and p38 activation in mantle cell lymphoma." Mol Pharmacol 70(5): 1612-20. Epub 2006 Aug 25
https://pubmed.ncbi.nlm.nih.gov/16936228

III. Gustafsson K, Wang X, Severa D, Eriksson M, Kimby E, Merup M, Christensson B, Flygare J, Sander B (2008). "Expression of cannabinoid receptors type 1 and type 2 in non-Hodgkin lymphoma: growth inhibition by receptor activation." Int J Cancer 123(5): 1025-33
https://pubmed.ncbi.nlm.nih.gov/18546271

IV. Gustafsson K, Sander B, Bielawski J, Hannun YA, Flygare J (2008). "Role for de novo synthesis of ceramide in cannabinoid-induced cytotoxicity in Mantle Cell Lymphoma." Journal of Biological Chemistry (Submitted)