Role of Sphingosine -1-Phosphate Receptor-1 (S1PR1) in Classical Hodgkin Lymphoma.
Michael J Kluk, Bonnie Wang, Kieran Ryan, Scott J Rodig, Teresa Sanchez, Jeffery L Kutok. Brigham and Women's Hospital, Boston, MA; Beth Israel Deaconess Medical Center, Boston, MA
Background: S1PR1 belongs to the G-protein coupled receptor family (S1PR1-5) that is important in modulation of lymphocyte trafficking and is a target for therapeutic immunosuppression. Notably, S1PR1 promotes migration while S1PR2 inhibits migration. The S1PR family ligand is sphingosine -1-phosphate (S1P), a sphingolipid highly abundant in plasma. Despite considerable knowledge of S1PR1 biology in normal lymphocytes, little is known about S1PR1 in lymphoid malignancies. Therefore, given the contiguous lymph node pattern of spread in classical Hodgkin lymphoma (cHL), we sought to study the role S1PR1 might play in the pathogenesis of cHL.
Design: IHC for S1PR1 (Santa Cruz) was performed on primary cHL cases and cHL cell lines. Quantitative PCR for S1PR transcripts was performed on KM-H2 cells. In vitro migration was assessed with a Boyden chamber-based migration assay (Neuro Probe) using S1PR agonists (S1P and FTY720-P) (Cayman Chemical), S1PR1 antagonist (VPC-44116; Dr. K. Lynch) and S1PR2 antagonist (JTE013; Cayman Chemical).
Results: IHC for S1PR1 in primary cHL revealed positive staining of Hodgkin Reed Sternberg (HRS) cells in 14/15 cases. Most cases showed strong to moderate staining in HRS cells, with only one case showing no S1PR1 staining. IHC for S1PR1 in cHL cell lines showed membranous and/or cytoplasmic positivity in all lines tested (KM-H2, L-428, L-1236 and SUP-HD1). Real time RT-PCR of KM-H2 cells showed ∼10 fold higher levels of S1PR1 than S1PR2. In contrast, S1PR3, S1PR4 and S1PR5 were expressed at extremely low levels. S1P induced a 5 fold average increase (5 +/- 2; n=5) in KM-H2 cell migration with maximal response at 1-10nM S1P. FTY720-P, a potent agonist for S1PR1 but not S1PR2, also strongly induced migration with maximal responses at 1nM. Pretreatment of KH-H2 cells with 0.1uM VPC 44116 (S1PR1 antagonist) blocked S1P-induced migration whereas 0.1uM JTE-013 (S1PR2 antagonist) enhanced S1P-induced migration. These results correlated with impairment of S1P-induced AKT phosphorylation by VPC 44116 but not by JTE-013.
Conclusions: These findings demonstrate that the pro-migratory receptor, S1PR1, is expressed in primary cHL and cHL cell lines. In KM-H2 cells, the increased expression of S1PR1 compared to S1PR2 permits S1P-induced migration and AKT phosphorylation, which are abrogated by the S1PR1, but not the S1PR2, antagonist. Taken together, these data suggest that S1PR1 signaling promotes HRS cell motility and may be important in cHL dissemination.
Tuesday, March 1, 2011 8:00 AM
Platform Session: Section B, Tuesday Morning