MS-275 derivative

Histone Deacetylases inhibitors (HDACi) modulate the acetylation profile of lysines on the histone tails to facilitate DNA accessibility to transcription factors. While the phenotypes caused by HDAC inhibition on cancer cells have been studied extensively, the nuclear geometry and expression signatures modulated by these enzymes have not been well understood.This work attempts to understand the functional implication of HDAC inhibitor treatment (NaB and MS275) on cervical cancer cells. We observed an increase in nuclear area upon HDAC inhibition correlating with an increase in expression of active histone marks and lamins and a decrease in levels of repressive epigenetic marks. Our transcriptomic sequencing of HeLa cells treated individually with these inhibitors have identified dysregulation of nucleoporins affecting the nucleocytoplasmic exchange and nucleo-cytoplasmic transport through the nuclear pores. These act in concert with the increase in acetylation due to HDAC inhibition and contribute to the increase in nuclear area. In order to derive clinical implications of the observed mechano signalling genes and epigenetic factors differentially expressed in HeLa cells, we verified these on a TCGA cervical cancer cohort of 148 patients and observed an upregulation of various nucleoporins in cervical cancer patients. Interestingly, the low expression of LMNA and high expression of NUP58 were associated with lower survival rate in the cohort. These signatures have also been validated on Indian cervical cancer tissues. This novel and intricate mechanism of modulating epigenetic regulation and nuclear architecture changes by HDAC inhibition can be utilized for designing targetted epigenetic therapy for cervical cancer.

Histone deacetylase (HDAC) density and activity are altered in different brain disorders. Antipsychotic drugs (APs) might modulate HDAC activity in brains of schizophrenia subjects.

HDAC activity assay amenable for enzyme kinetics and HDAC inhibitor (HDACi) screening studies in postmortem human brain samples.

The optimization and characterization work involved several steps. The nucleosolic subcellular fraction and total protein amount needed for an optimal HDAC activity on Boc-Lys(Ac)-AMC substrate were characterized. Signal-to-noise ratio (1.8) and Z-score values (0.82) were indicators of the assay quality. Inhibition studies with non-selective (belinostat, vorinostat, valproic acid) and selective (apicidin, MS275, romidepsin, tacedinaline and EX527) HDACis showed that the optimized assay detected class I HDAC activity. The obtained IC50 values were similar to those previously reported, proving the assay reliability. We used the optimized assay to study the effect of APs on HDAC activity. Inhibition studies with APs in postmortem human brain, together with enzyme kinetic studies in brains of rats chronically treated with APs observed no modulation of class I HDAC activity.

This study describes the optimization of a reliable and cost efficient HDAC activity assay for its use in postmortem human brain samples. The assay does not depend on antibody specificity and it is valid for enzyme kinetic studies and for the screening of new potential class I HDACis.

We optimized and characterized an assay to measure HDAC activity in postmortem human brain samples. We did not observe any modulatory effect of APs on HDAC activity.