Abstract 4102: The involvement of EMT in heparanase promoted bone-metastasis in multiple myeloma

Multiple myeloma (MM), a hematologic malignancy, preferentially grows in bone marrow and frequently metastasizes between bones. Our previous studies have demonstrated that heparanase (HPSE), an enzyme overexpressed in MM cells, promotes MM bone-metastasis and progression. However, the mechanism of HPSE action is unclear. Epithelial-mesenchymal-transition (EMT) is a process giving epithelial cells the features of mesenchymal cells which has been shown to be critical in the metastatic process of epithelial-origin tumors. Although MM is not an epithelial-derived malignancy, MM cells indeed express epithelial marker E-cadherin and mesenchymal marker Vimentin, and have the ability to adhere to and interact with the extracellular matrix (ECM). In the present study, we determined the role of EMT in HPSE-induced MM bone metastasis via: (1) Using Western blot to evaluate the expression of EMT markers E-cadherin, Vimentin and Fibronectin in MM cells that were transfected with empty vector (HPSE-low) or HPSE cDNA (HPSE-high), as well as MM cells treated with recombinant human HPSE (rHPSE) for 48hrs; (2) staining HPSE and EMT markers on tumors established by HPSE-low or HPSE-high cells in SCID mice (n = 10), and on bone marrow biopsies of 35 MM patients; (3) knocking down Vimentin in HPSE-high-luc MM cells by Vimentin shRNA (VIM k/d-luc cells) and testing bone homing of VIM k/d-luc cells in SCID mice by i.v. and s.c. injection of these cells. Tumor growth and bone-metastasis was determined by measuring human kappa levels (a soluble marker of MM cells) in mouse sera and weekly bioluminescent imaging. Western blots demonstrated HPSE-high and rHPSE treated MM cells expressed significantly higher levels of the mesenchymal markers Vimentin and Fibronectin, and lower levels of the epithelial marker E-cadherin, compared to HPSE-low or PBS treated MM cells. Significantly increased expression of Vimentin and decreased expression of E-cadherin was revealed in tumors formed by HPSE-high cells compared with the tumors formed by HPSE-low cells. Immunostaining of bone marrow biopsies of MM patients indicated HPSE and Vimentin expression in MM cells have a significant positive correlation (rs = 0.414, p = 0.014). Finally, both SCID-i.v. injection and SCID-s.c. injection models showed that knocking down mesenchymal marker Vimentin in HPSE-high MM cells successfully blocked HPSE-induced bone metastasis and inhibited tumor growth, compared with mice bearing shRNA control HPSE-high MM cells. In conclusion, our studies demonstrated (1) EMT-like features are a novel biomarker of poor prognosis in MM; (2) HPSE promotes MM metastasis to bone and disease progression, at least in part, through inducing EMT-like features in MM cells; (3) Inhibition of Vimentin expression in MM cells impairs HPSE-induced tumor growth and bone metastasis. Note: This abstract was not presented at the meeting. Citation Format: Qianying Pan, Juan Li, Patrick D. Rowan, Timothy N. Trotter, Yang Yang. The involvement of EMT in heparanase promoted bone-metastasis in multiple myeloma. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4102. doi:10.1158/1538-7445.AM2015-4102