Role of notch pathway in EV-mediated crosstalk between multiple myeloma and tumor microenvironment

INTRODUCTION Multiple myeloma (MM) is an incurable cancer characterized by a strong tropism to bone marrow (BM) and an important involvement of Notch pathway. Recently, extracellular vesicles (EVs) have been reported as mediators in creating a supportive milieu for MM. These evidences make us to hypothesize that it may have a role in EVs mediated cross-talk. EXPERIMENTAL MODEL We established two MM cell lines that stably retain the doxycycline-inducible pTRIPZ vector containing either scrambled or anti Jagged1/Jagged2 shRNA, and a bone marrow mesenchimal stromal cell (BM-MSC) line (HS5) that expresses shRNAs for Notch1 and 2 and the scrambled control. EVs were isolated by ultracentrifugation and used for functional assays. RESULTS Notch1/2 KD in BM-MSC results in a decrease in the release of EVs; BMSC-EV derived from Notch1/2 KD cells showed a reduced ability to induce Bortezomib resistance and to stimulate migration of MM cells. On the other side, MM-EVs are able to activate Notch in HS5, increasing the production of pro-tumoral factor SDF1; the effect is lost when EVs are isolated from MM cells where Jag1/2 were silenced. Moreover EVs released by co-cultures of BMSCs and MM cells silenced for Jag1/2 display a reduced ability to increase osteoclastogenesis compared with EVs from the control culture. CONCLUSION We prove that BM-MSC and MM derived EVs show biological functions in accordance with Notch pathway activation status and their role as mediators of tumor progression and relapse.