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Letters to the Editor
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Figure 3. Ricolinostat promotes the activity of S63845 independent of HDAC6 inhibition. (A) MM.1S cells were either treated with panobinostat or ricolinostat for 24 hours (h), then whole-cell lysates were blotted for the indicated proteins. (B) Single-cell clones of MM.1S cells harboring a Tet-on miR-E vector expressing either a short hairpin RNA (shRNA) targeting Renilla (control) or an shRNA targeting HDAC6 were exposed to 0.3 μg/mL doxycycline for 48 h and whole-cell lysates were blotted for the indicated proteins. Western blots are representative for three independent experiments. (C and D) Cells were pretreated with doxy- cycline for 48 h and viability was assessed after an additional 48-h treatment with increasing concentrations of S63845 as indicated in the Figure. Results show the mean +/- standard deviation (SD) of three independent experiments performed in triplicates. (E and J) CD138 purified plasma cells sorted from patients with multiple myeloma (MM) were exposed to S63845, panobinostat or ricolinostat the respective combinations for 20 h (E and H) or 10 h (I and J). (E and H) Apoptotic cells were determined via Annexin V/7AAD positive staining. (I and J) Whole-cell lysates of primary MM cells were blotted for the indicated proteins. Short-term exposure of primary patient samples was chosen to avoid spontaneous cell death. (K) Proposed model of the underlying mechanism of the observed synergism between MCL-1i and HDACi in MM. In S63845 treated cells BCL-XL is capable to sequester BAK, hence inhibiting the apoptotic signaling cascade resulting in diminished MM cell death. In combination with HDACi, BCL-XL protein is downregulated and thereby BAK, released by MCL-1i, can be activated, can oligomerize and can insert into the MOMP which in turn releases cytochrome c and activates initiator as well as effector caspases, leading to PARP cleavage and cell death.
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