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Letters to the Editor
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Figure 3. TOP2 inhibitor dexrazoxane displays anti-myeloma properties and combinatorial activity with lenalidomide. (A) Immunoblot showing TOP2B expres- sion in MM.1S cells treated with dimethyl sulfoxide (DMSO) or dexrazoxane (DXZ) (20 mM or 50 mM) across 24, 48 and 72 hours. Tubulin expression is provided as a loading control. The experiment is representative of three biological replicates. (B) Bar graphs demonstrating Nicoletti cell cycle profiling of MM.1S and MM.1S resistant (MM1Sres) cells exposed to DMSO, lenalidomide (LEN) (2 mM), DXZ (20 mM) or combined LEN/DXZ (combo). The plot represents an aggregate of three independent experiments with three technical replicates each. Error bars represent mean ± standard error of the mean of n=3 independent experi- ments. v: significant vs. vehicle (DMSO); l: significant vs. LEN; d: significant vs. DXZ (P<0.05 or less). (C) Heatmaps representing the variation of zero interaction potency (ZIP) synergy score in MM.1S and MM.1Sres cells treated with increasing concentration of LEN and DXZ in combination. The average percentage of PI negative cells (viable) in each drug combination of three independent experiments was employed to compute synergy. The R package synergyfinder v2.2.4 was employed to perform the analysis. (D) Average viable cell count at 3 and 7 days for MM.1S and MM.1Sres treated with DMSO, LEN (2 mM), DXZ (5 mM or 20 mM depending on the cell line) or combined LEN/DXZ (combo). (E) Immunoblot showing expression of IKZF1, IKZF3 after 24h of treatment with DMSO, LEN (2 mM), DXZ (20 μM) or combined treatment and expression of IRF4 and MYC after 72 hours of treatment in the same conditions. HSP90 is provided as loading control. The experiment is representative of three biological replicates. (F) Viable cell count at 3 and 7 days of OPM2, RPMI-8226 and JJN3 cells treated with DMSO, LEN (2 mM), DXZ (20 mM) or combo. (G) Percentage of PI negative (viable) OPM2 cells treated with DMSO, LEN (2 mM), DXZ (20 mM) or combo and assessed by flow cytometry. (H) Percentage of PI negative (viable) RPMI-8226 cells treated with DMSO, LEN (2 mM), DXZ (5 mM) or combo and assessed by flow cytometry. (I) Percentage of PI negative (viable) JJN3 cells treated with DMSO, LEN (2 mM), DXZ (20 mM) or combo and assessed by flow cytometry. In (D), (F), (G), (H) and (I), error bars represent mean ± standard error of the mean of n=3 independent experiments. v: significant vs. vehicle (DMSO); l: significant vs. LEN; d: signif- icant vs. DXZ (P<0.05 or less).
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