Abstract
Abstract 4808
The antileukemic activity of histone deacetylase inhibitors (HDACi) has driven the search for epigenetic drugs with higher substrate specificity. Most of the currently used HDACi target class I, II and IV, with some bearing a class preference but only very few being selective in inhibiting specific HDACs, such as the HDAC6-selective inhibitor tubacin. HDAC6 inhibition leads to acetylation of non-histone proteins such as Hsp90 and alpha-tubulin. As it was recently demonstrated that HDAC6 is overexpressed in AML cells, we sought to investigate the effect of selective HDAC6 inhibition by the novel hydroxamic acid derivate ST80 in myeloid cell lines.
ST80 (selective HDAC6 inhibitor, 30-fold higher enzyme inhibition as compared to HDAC1) and the non-selective novel hydroxamic acid derivates ST13 (pan-HDACi) and ST34 (pan-HDACi with preference for class I) had previously been tested in enzymatic assays for their HDAC inhibitory potential (Scott et al. Mol Cancer Res. 6:1250-8, 2008). Cell lines HL60, Kasumi-1, NB4, THP1, U937 and K562 were treated with 10 nM to 30 μM of these three drugs. Viability and growth inhibition were determined using trypan blue staining. Acetylation of histone H3, H4 and alpha-tubulin and HDAC6 expression were determined by Western blot and quantified by densitometry. Tubulin-selective acetylation was calculated as the ratio of tubulin acetylation vs. H4 acetylation (ac-tubulin:ac-H4 quotient).
At 1 μm, ST80, ST13 and ST34 all acetylated tubulin (8-, 8- and 2-fold in NB4, 11-, 14- and 3.4-fold in HL60, respectively, after a 12 h treatment). However, the calculated ac-tubulin:ac-H4 ratio of ST80 was 15- and 8-fold higher in NB4 and 9-fold higher in HL60 when compared to ST13 and ST34. The inhibitory concentration (IC) 50 (cell growth) of ST80 in the six myeloid cell lines ranged from 2.8 μM (NB4) to 5.1 μM (Kasumi-1) after 48 h treatment. Median cell viability of all 6 cell lines at 48 h was 93.7 % (range: 87.0 - 96.8 %) at 1 μm and 90.3 % (65.7 - 95.7 %) at 5 μm of ST80. HDAC6 protein levels were strongly variable between cell lines; however, growth inhibition by ST80 was independent of HDAC6 expression.
The novel hydroxamic acid derivate ST80 shows antileukemic activity in myeloid cell lines at low micromolar concentrations, which affect cell viability only modestly. The degree of relative tubulin acetylation by ST80 indicates a selective HDAC6 inhibitory activity in myeloid leukemias. The favorable ratio of ST80 growth inhibition vs. cytotoxicity warrants combination studies of this drug with other compounds.
No relevant conflicts of interest to declare.
Author notes
Asterisk with author names denotes non-ASH members.
