SLAMF1 regulation of chemotaxis and autophagy determines CLL patient response
Cinzia Bologna, Roberta Buonincontri, Sara Serra, Tiziana Vaisitti, Valentina Audrito, Davide Brusa, Andrea Pagnani, Marta Coscia, Giovanni D’Arena, Elisabetta Mereu, Roberto Piva, Richard R. Furman, Davide Rossi, Gianluca Gaidano, Cox Terhorst, Silvia Deaglio
Cinzia Bologna, Roberta Buonincontri, Sara Serra, Tiziana Vaisitti, Valentina Audrito, Davide Brusa, Andrea Pagnani, Marta Coscia, Giovanni D’Arena, Elisabetta Mereu, Roberto Piva, Richard R. Furman, Davide Rossi, Gianluca Gaidano, Cox Terhorst, Silvia Deaglio
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Research Article Oncology

SLAMF1 regulation of chemotaxis and autophagy determines CLL patient response

Abstract

Chronic lymphocytic leukemia (CLL) is a variable disease; therefore, markers to identify aggressive forms are essential for patient management. Here, we have shown that expression of the costimulatory molecule and microbial sensor SLAMF1 (also known as CD150) is lost in a subset of patients with an aggressive CLL that associates with a shorter time to first treatment and reduced overall survival. SLAMF1 silencing in CLL-like Mec-1 cells, which constitutively express SLAMF1, modulated pathways related to cell migration, cytoskeletal organization, and intracellular vesicle formation and recirculation. SLAMF1 deficiency associated with increased expression of CXCR4, CD38, and CD44, thereby positively affecting chemotactic responses to CXCL12. SLAMF1 ligation with an agonistic monoclonal antibody increased ROS accumulation and induced phosphorylation of p38, JNK1/2, and BCL2, thereby promoting the autophagic flux. Beclin1 dissociated from BCL2 in response to SLAMF1 ligation, resulting in formation of the autophagy macrocomplex, which contains SLAMF1, beclin1, and the enzyme VPS34. Accordingly, SLAMF1-silenced cells or SLAMF1lo primary CLL cells were resistant to autophagy-activating therapeutic agents, such as fludarabine and the BCL2 homology domain 3 mimetic ABT-737. Together, these results indicate that loss of SLAMF1 expression in CLL modulates genetic pathways that regulate chemotaxis and autophagy and that potentially affect drug responses, and suggest that these effects underlie unfavorable clinical outcome experienced by SLAMF1lo patients.

Authors

Cinzia Bologna, Roberta Buonincontri, Sara Serra, Tiziana Vaisitti, Valentina Audrito, Davide Brusa, Andrea Pagnani, Marta Coscia, Giovanni D’Arena, Elisabetta Mereu, Roberto Piva, Richard R. Furman, Davide Rossi, Gianluca Gaidano, Cox Terhorst, Silvia Deaglio

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Figure 4

SLAMF1 cross-linking induces ROS accumulation, activates JNK1/2 and p38 kinases, and phosphorylates BCL2.

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SLAMF1 cross-linking induces ROS accumulation, activates JNK1/2 and p38 ...
(A and B) SLAMF1 cross-linking induces ROS accumulation, as measured by flow cytometry in CLL cells (A) or Mec-1 cells (B) by using CM-H2DCFDA or CellRox dies, respectively. The histogram on the left shows results from a representative experiment. The bar graph in A derives from 8 independent experiments using primary cells from 8 different patients. Silencing of gp91-phox or p40-phox of the NOX2 enzyme complex by siRNA transfection impairs ROS accumulation following SLAMF1 cross-linking in Mec-1 cells (B). The Western blot confirms silencing of both molecules. Results from 5 independent experiments, each performed in triplicate. (C) Ligation of SLAMF1 on primary CLL cells induces time-dependent tyrosine phosphorylation of JNK1/2 and p38. A representative blot is shown, together with cumulative data obtained from 8 independent experiments from 8 different patients. The fold-increase represents the increase in band intensity over the untreated condition. (D) Ligation of SLAMF1 induces a rapid and robust phosphorylation of BCL2 at S70 in primary CLL cells, as shown with a specific mAb. The bar graph shows cumulative data from 7 different patients. (E) Confocal microscopy showing p-BCL2 accumulation (green) following SLAMF1 ligation in primary CLL cells. The cytoskeleton was visualized by AlexaFluor 568–conjugated phalloidin; nuclei were counterstained with DAPI. Original magnification, ×63; zoom factor of 2. Representative image from 5 independent experiments. ***P ≤ 0.001, **P ≤ 0.01, and *P ≤ 0.05, as calculated by Friedman’s test followed by Dunn’s multiple comparison.