STIM1 at the plasma membrane as a new target in progressive chronic lymphocytic leukemia.

Background: Dysregulation in calcium (Ca ²⁺ ) signaling is a hallmark of chronic lymphocytic leukemia (CLL). While the role of the B cell receptor (BCR) Ca ²⁺ pathway has been associated with disease progression, the importance of the newly described constitutive Ca ²⁺ entry (CE) pathway is less clear. In addition, we hypothesized that these differences reflect modifications of the CE pathway and Ca ²⁺ actors such as Orai1, transient receptor potential canonical (TRPC) 1, and stromal interaction molecule 1 (STIM1), the latter being the focus of this study.
Methods: An extensive analysis of the Ca ²⁺ entry (CE) pathway in CLL B cells was performed including constitutive Ca ²⁺ entry, basal Ca ²⁺ levels, and store operated Ca ²⁺ entry (SOCE) activated following B cell receptor engagement or using Thapsigargin. The molecular characterization of the calcium channels Orai1 and TRPC1 and to their partner STIM1 was performed by flow cytometry and/or Western blotting. Specific siRNAs for Orai1, TRPC1 and STIM1 plus the Orai1 channel blocker Synta66 were used. CLL B cell viability was tested in the presence of an anti-STIM1 monoclonal antibody (mAb, clone GOK) coupled or not with an anti-CD20 mAb, rituximab. The Cox regression model was used to determine the optimal threshold and to stratify patients.
Results: Seeking to explore the CE pathway, we found in untreated CLL patients that an abnormal CE pathway was (i) highly associated with the disease outcome; (ii) positively correlated with basal Ca ²⁺ concentrations; (iii) independent from the BCR-PLCγ2-InsP ₃ R (SOCE) Ca ²⁺ signaling pathway; (iv) supported by Orai1 and TRPC1 channels; (v) regulated by the pool of STIM1 located in the plasma membrane (STIM1 _PM ); and (vi) blocked when using a mAb targeting STIM1 _PM . Next, we further established an association between an elevated expression of STIM1 _PM and clinical outcome. In addition, combining an anti-STIM1 mAb with rituximab significantly reduced in vitro CLL B cell viability within the high STIM1 _PM CLL subgroup.
Conclusions: These data establish the critical role of a newly discovered BCR independent Ca ²⁺ entry in CLL evolution, provide new insights into CLL pathophysiology, and support innovative therapeutic perspectives such as targeting STIM1 located at the plasma membrane.