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J. He et al.
proteins. As shown in Figure 4 and Online Supplementary Figure S5, most of these proteins showed the same changes in the presence and absence of Q-VD-Oph.
Chidamide decreases the MM tumor burden and prevents myeloma-associated bone loss in a mouse model of disseminated myeloma
Two murine models were employed to examine the dual anti-tumor and bone-protective effects of chidamide. First, as shown in Figure 5A, a significant decrease in tumor volume was observed in chidamide-treated NOD/SCID mice compared with that observed in the vehicle group. Consistent with the Western blot results from the cell lines, immunohistochemical staining revealed decreased expression of Bcl-xL, CDK4, CDK6 and Myc and increased levels of cleaved caspase-3 (Online
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Supplementary Figure S6). Next, NCG mice were intra- venously injected with RPMI-8226-luc cells to establish the murine model of myeloma-induced bone destruction. Compared with the vehicle group, chidamide clearly con- trolled the tumor burden, as measured by serum levels of human Ig λ light chain secreted by RPMI-8226-luc cells (***P<0.001, Figure 5B) and the bioluminescence in vivo imaging (Figure 5B). Then, serum levels of the bone resorption marker Carboxy-terminal telopeptide 1 (CTX- I) were measured and were found to be significantly diminished in chidamide-treated mice (**P<0.01, Figure 5C); in addition, a significantly increased serum level of amino-terminal propeptide (PINP), the bone formation marker, was observed (***P<0.001, Figure 5C). Consistent with these findings, micro-CT three-dimensional recon- structed images at the metaphysis of distal femurs
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Figure 2. MM cells are vulner- able to chidamide. (A) The cytotoxicity of chidamide toward MM cell lines was examined using flow cytome- try (Annexin V/PI), and all data are summarized as the means±SEM (n=3). (B) The expression of apoptosis-asso- ciated proteins, including PARP-1 and cleaved caspas- es 3, 7, 8, and 9, in chi- damide-treated ARP-1 and MM.1s cells was detected. (C) The effect of chidamide on apoptosis in MM.1s cells after 48h with or without co- incubation with the pan-cas- pase inhibitor Q-VD-Oph. After co-incubation with Q-VD-Oph, chidamide-induced apoptosis was inhibited. (D) Cell cycle arrest was examined using flow cytometry, and ARP-1 cells underwent G1 phase arrest in response to incuba- tion with chidamide regard- less of co-incubation with Q- VD-Oph. (E) Chidamide could significantly induce + primary CD138 cell apoptosis after incubation for 48h (n=5, *P<0.05). (E) Chidamide could significantly induce pri- mary CD138+ cell apoptosis after incubation for 48h (n=5). PI: propidium iodide; MM: multiple myeloma.
haematologica | 2018; 103(8)