Fig. 5: Inhibiting the secretion of exosomes in 5TGM1 mice with GW4869 reduces tumor load when combined with standard-of-care treatment bortezomib.

a Relative viability of MC3T3-E1 was measured by a Cell Titer Glo assay. Cells were cultured in 5TGM1 concentrated conditioned medium (CCM), 5TGM1 sEVs (100 µg/ml); 5TGM1 CCM collected after treatment of 5TGM1 cells with GW4869 (10 µM) for 24 h (GW4869), reducing exosome secretion and finally a combination of 5TGM1 GW4869 CCM and 5TGM1 sEVs (100 µg/ml) (GW4869 + 5TGM1 sEVs). Bars represent mean ± SD. Experiments were repeated at least three times. * = p < 0.05, *** = p < 0.001. b Western blot analysis of 5TGM1 CCM after treatment for 24 h with an indicated dosage of GW4869, showing a reduction in exosomal markers TSG101 and CD81. c 6-week old mice were inoculated with 1 million 5TGM1 cells on day 1. Treatment with bortezomib (0.6 mpk, two times per week, SC) and/or GW4869 (2.5 mpk, three times per week, IP) started the next day. N = 10 for each treatment group. At day 25 all mice were sacrificed. d Spleen, serum and bone marrow (BM) were collected for analysis of M-protein (protein electrophoresis) and BM plasmacytosis. e Quantitative µCT analysis was performed for naive and vehicle mice. For mice from the treatment groups, six mice with representative tumor load for the rest of the group were selected for µCT analysis. Shown are cortical bone volume (Ct. BV/TV) and trabecular bone volume (Tb. BV/TV). * = p < 0.05, ** = p < 0.01. f Levels of serum circulating type I collagen degradation product (Collagen C-terminal telopeptide or CTX) was measured by the CTX ELISA kit (cf. supplementary materials) in all treatment groups. Serum was diluted 1:10. * = p < 0.05, *** = p < 0.001