Downregulation of Cell Cycle and Checkpoint Genes by Class I HDAC Inhibitors Limits Synergism with G2/M Checkpoint Inhibitor MK-1775 in Bladder Cancer Cells

Abstract

Since genes encoding epigenetic regulators are often mutated or deregulated in urothelial carcinoma (UC), they represent promising therapeutic targets. Specifically, inhibition of Class-I histone deacetylase (HDAC) isoenzymes induces cell death in UC cell lines (UCC) and, in contrast to other cancer types, cell cycle arrest in G2/M. Here, we investigated whether mutations in cell cycle genes contribute to G2/M rather than G1 arrest, identified the precise point of arrest and clarified the function of individual HDAC Class-I isoenzymes. Database analyses of UC tissues and cell lines revealed mutations in G1/S, but not G2/M checkpoint regulators. Using class I-specific HDAC inhibitors (HDACi) with different isoenzyme specificity (Romidepsin, Entinostat, RGFP966), cell cycle arrest was shown to occur at the G2/M transition and to depend on inhibition of HDAC1/2 rather than HDAC3. Since HDAC1/2 inhibition caused cell-type-specific downregulation of genes encoding G2/M regulators, the WEE1 inhibitor MK-1775 could not overcome G2/M checkpoint arrest and therefore did not synergize with Romidepsin inhibiting HDAC1/2. Instead, since DNA damage was induced by inhibition of HDAC1/2, but not of HDAC3, combinations between inhibitors of HDAC1/2 and of DNA repair should be attempted.

Multipathway regulation induced by 4‐(phenylsulfonyl)morpholine derivatives against triple‐negative breast cancer

“…Suppression of the genes involved in the regulation of G2/M affected cell cycle by G2/M checkpoint arrest. [ 40 ] Inhibition of E2F transcriptional activation induced G1 cell‐cycle arrest. [ 41 ] Inhibition of epithelial–mesenchymal transition (EMT), another downregulated gene set, could suppress tumor‐initiating and metastatic potential.…”

Quantitative Acetylomics Uncover Acetylation-Mediated Pathway Changes Following Histone Deacetylase Inhibition in Anaplastic Large Cell Lymphoma

“…Additionally, in SAHA samples, “cell cycle” could be observed among downregulated pathways. HDACi are known to induce cell cycle arrest [ 42 , 43 , 44 , 45 ]. In the Hdac1 KO SC sample, downregulated pathways were different as compared to the HDACi treatments and affected oncogenic signaling such as “PTEN Regulation” and “PIP3 activates AKT signaling”.…”

“…The interconnectedness of the proteins with increased acetylation detected in MS-275 treated samples can be observed in Figure 5 E, where chromatin modifiers and proteins involved in homology directed repair and cell cycle are highlighted. It was shown before that HDACi treatment can induce G/M2 cell cycle arrest [ 42 , 43 , 44 , 45 ] and that HDACs are involved in DDR [ 49 ]. Proteins involved in cell proliferation and DNA replication included POLD3 (K429ac), RPA3 (K33ac), PCNA (K80ac) and NPM1 (K141ac, K227ac) ( Figure 5 D).…”

“…Co-staining for TP53BP1 revealed a prominent overlap with γH2AX foci following DNA damage in untreated and HDACi pre-treated irradiated samples, suggesting that acetylation of TP53BP1 on K1357 did not affect TP53BP1 recruitment to sites of DNA damage. In addition, we detected a higher number of mitotic cells upon HDACi treatment, with chromosomes with highly enriched γH2AX, which might be due to G2/M phase arrest following treatment and irradiation [ 42 , 43 , 44 , 45 ] ( Supplementary Figure S3A,B ). Co-immunofluorescence for TP53BP1 revealed that γH2AX positive chromosomes were devoid of TP53BP1 staining.…”

New synergistic combination therapy approaches with HDAC inhibitor quisinostat, cisplatin or PARP inhibitor talazoparib for urothelial carcinoma

“…[14][15][16] We also reported earlier that rather HDAC1 and HDAC 2 are better suited targets than HDAC 8 and 3. 13,17 Since romidepsin targets mainly HDAC1 and 2 we had characterized its effect on UC cells extensively earlier. 16 In this study, we characterized the impact of the second generation HDACi quisinostat on UC cells.…”