Selective HDAC3 Inhibitors with Potent In Vivo Antitumor Efficacy against Triple-Negative Breast Cancer.

HDAC3 modulation shows promise for breast cancer, including triple-negative cases. Novel pyrazino-hydrazide-based HDAC3 inhibitors were designed and synthesized. Lead compound 4i exhibited potent HDAC3 inhibition (IC50 = 14 nM) with at least 121-fold selectivity. It demonstrated strong cytotoxicity against triple-negative breast cancer cells (IC50: 0.55 µM for 4T1, 0.74 µM for MDA-MB-231) with least normal cell toxicity. Metabolically stable 4i displayed a superior pharmacokinetic profile. A dose-dependent therapeutic efficacy of 4i was observed in a tumor-bearing mouse model. The biomarker analysis with tumor tissues displayed enhanced acetylation on Ac-H3K9, Ac-H3K27, and Ac-H4K12 compared to Ac-tubulin and Ac-SMC3 indicating HDAC3 selectivity of 4i in vivo. The immunoblotting study with tumor tissue showed upregulation of apoptotic proteins caspase-3, caspase-7, and cytochrome c and the downregulation of proliferation markers Bcl-2, CD44, EGFR, and Ki-67. Compound 4i represents a promising candidate for targeted breast cancer therapy, particularly for cases with triple-negative breast cancer.

HDAC8 as an emerging target in drug discovery with special emphasis on medicinal chemistry.

HDAC8 catalyzes the deacetylation of both histones and nonhistone proteins. The abnormal expression of HDAC8 is associated with various pathological conditions causing cancer and other diseases like myopathies, Cornelia de Lange syndrome, renal fibrosis, and viral and parasitic infections. The substrates of HDAC8 are involved in diverse molecular mechanisms of cancer such as cell proliferation, invasion, metastasis and drug resistance. Based on the crystal structures and the key residues at the active site, HDAC8 inhibitors have been designed along the canonical pharmacophore. This article details the importance, recent advancements, and the structural and functional aspects of HDAC8 with special emphasis on the medicinal chemistry aspect of HDAC8 inhibitors that will help in developing novel epigenetic therapeutics.

Selective inhibition of histone deacetylase 3 by novel hydrazide based small molecules as therapeutic intervention for the treatment of cancer.

A promising hydrazide based small molecule lead as a potent and selective histone deacetylase 3 (HDAC3) inhibitor has been developed from a small series of synthesized novel chemical entities. The lead compound (4e) displayed high HDAC3 inhibitory potency (IC50 = 15.41 nM) and a minimum of 18-fold selectivity over other HDAC isoforms. It also exhibited potent cytotoxicity against several cancer cell lines with minimal toxicity against normal cell lines tested. Compound 4e also enhanced acetylation levels on H3K9, H4K12 and H3K27 both in vitro and in vivo. It also induced cell cycle arrest at the G2/M phase in B16F10 and 4T1 cells. It caused significant apoptosis and upregulated the expression of caspase-3, caspase-7, cytochrome c and downregulated the expression of BCL2 in tumour tissue. In addition, the downregulation of CD44, EGFR and Ki-67 suggested the potential of compound 4e in reducing cell proliferation and metastasis in mice. Further, a marked decrease in the tumour volume was observed with no general toxicity in the major organs when treated with 4e in the 4T1-Luc xenograft mouse model. Therefore, compound 4e is a promising candidate selectively targeting HDAC3 with a significant antitumour activity that can be evaluated further in preclinical and clinical evaluation.

Estrogenic Activity of Tetrazole Derivatives Bearing Bisphenol Structures: Computational Studies, Synthesis, and In Vitro Assessment.

The classification of bisphenol A (BPA) as an industrial endocrine disruptor has led to a ban of this ubiquitous critical starting material from food and medical applications. Thus, scientists worldwide are researching to develop non-ER binding starting compounds to fulfill unmet market needs. In line with this trending research topic, the current paper highlights the development of tetrazole derivatives bearing a bisphenol structure (TbB) as a novel weak binder or potential inactive to the estrogen receptor (ER) and androgen receptor (AR). The structure and ligand-based approach supported by binding affinity analysis, electrostatic complementarity, ADMET prediction, and in silico studies identified TbBs as privileged substitutes for BPA. Five TbB ligands were successfully synthesized and subjected to biological testing comprising radioligand competitive binding and functional cellular assays. The measured IC50 value for BPA was found to be 0.24 µM, whereas all the inhibitions were less than 15% for the two TbB ligands, 223-2 and 223-3. As these TbB ligands did not meet the established acceptance criteria of 50% inhibition, they are considered as extremely weak binders to ERα. Steric clashes, the desolvation effect, and the increased total polar surface area (TPSA) of TbB ligands in the hydrophobic binding site are hypothesized to be possible reasons for low binding. Modeling studies complemented by bioassays highlight TbB compounds as privileged prospective BPA replacements. However, more research on TbB ligand toxicity is needed to understand and substantiate that the adverse effects on the hormonal system, for example, via metabolic activation, are not elicited.

Design, synthesis and structure-activity relationship studies of novel spirochromanone hydrochloride analogs as anticancer agents.

Aim: Literature reports suggest spirochromanone derivatives exhibit anticancer activity. Methodology: The authors designed and synthesized 18 spirochromanone derivatives (Csp 1-18). The compounds were characterized and evaluated for anticancer activity against human breast cancer (MCF-7) and murine melanoma (B16F10) cell lines. Results: The anticancer activity ranged from 4.34 to 29.31 µm. The most potent compounds, Csp 12 and Csp 18, were less toxic against the human embryonic kidney (HEK-293) cell line and ∼ two/∼fourfold selective toward MCF-7 than B16F10 in comparison to the reference, BG-45. Csp 12 caused 28.6% total apoptosis, leading to significant cytotoxicity, and arrested the G2 phase of the cell cycle in B16F10 cells. A molecular docking study of Csp 12 exhibited effective binding at the active site of the epidermal growth factor receptor kinase domain. Conclusion: This study highlights the importance of spirochromanones as anticancer agents.

Design, synthesis and binding mode of interaction of novel small molecule o-hydroxy benzamides as HDAC3-selective inhibitors with promising antitumor effects in 4T1-Luc breast cancer xenograft model.

Histone deacetylase 3 (HDAC3) is one of the most promising targets to develop anticancer therapeutics. In continuation of our quest for selective HDAC3 inhibitors, a series of small molecules having o-hydroxy benzamide as the novel zinc binding group (ZBG) has been introduced for the first time that can be able to produce good HDAC3-selectivity over other HDACs. The most promising HDAC3 inhibitors, 11a and 12b, displayed promising in vitro anticancer activities with less toxicity to normal kidney cells. These compounds significantly upregulate histone acetylation and induce apoptosis with a G2/M phase arrest in B16F10 cells. Compound 11a exhibited potent antitumor efficacy in 4T1-Luc breast cancer xenograft mouse model in female Balb/c mice. It also showed significant tumor growth suppression with no general toxicity and extended survival rates post-tumor resection. It significantly induced higher ROS generation, leading to apoptosis. No considerable toxicity was noticed in major organs isolated from the compound 11a-treated mice. Compound 11a also induced the upregulation of acH3K9, acH4K12, caspase-3 and caspase-7 as analyzed by immunoblotting with treated tumor tissue. Overall, HDAC3 selective inhibitor 11a might be a potential lead for the clinical translation as an emerging drug candidate.

Synthesis, biological evaluation, and molecular docking analysis of novel linker-less benzamide based potent and selective HDAC3 inhibitors.

A series of novel linker-less benzamides with different aryl and heteroaryl cap groups have been designed, synthesized, and screened as potent histone deacetylase (HDAC) inhibitors with promising anticancer activity. Two lead compounds 5e and 5f were found as potent and highly selective HDAC3 inhibitors over other Class-I HDACs and HDAC6. Compound 5e bearing a 6-quinolinyl moiety as the cap group was found to be a highly potent HDAC3 inhibitor (IC50 = 560 nM) and displayed 46-fold selectivity for HDAC3 over HDAC2, and 33-fold selectivity for HDAC3 over HDAC1. The synthesized compounds possess antiproliferative activities against different cancer cell lines and significantly less cytotoxic to normal cells. Molecular Docking studies of compounds 5e and 5f reveal a similar binding mode of interactions as CI994 at the HDAC3 active site. These observations agreed with the in vitro HDAC3 inhibitory activities. Significant enhancement of the endogenous acetylation level on H3K9 and H4K12 was found when B16F10 cells were treated with compounds 5e and 5f in a dose-dependent manner. The compounds induced apoptotic cell death in Annexin-V/FITC-PI assay and caused cell cycle arrest at G2/M phase of cell cycle in B16F10 cells. These compounds may serve as potential HDAC3 inhibitory anticancer therapeutics.

Design, synthesis and biological evaluation of 7-(5-((substituted - amino)-methyl)-thiophen-2-yl)-spiro-[chroman-2,4'-piperidin]-4-one hydrochloride analogues as anticancer agents.

A series of thirty-one novel 7-(5-((amino)-methyl)-thiophen-2-yl)-spiro-[chroman-2,4'-piperidin]-4-one hydrochloride analogues (Cst 1 - 31) have been designed, synthesized and characterized by 1H NMR, 13C NMR and MS spectral analysis. Here, we evaluated the anticancer potential and biological results of low-molecular-weight bridgehead oxygen and nitrogen-containing spirochromanones on proliferation and apoptosis of the human breast cancer cell line (MCF-7) and Murine melanoma (B16F10). The anticancer activity ranged from 2.9 to 35.0 µM. The most potent compounds Cst-22, Cst-24 and Cst-31 were found to be less toxic against human embryonic kidney (HEK-293) cell lines. Cst-24 and Cst-31 were found to be causing significant cytotoxicity through apoptotic cell death and also G2 phase arrest of cell cycle in B16F10 cells. In-silico ADME prediction stidies of the titled compounds were found within the rules outlined, and these compounds may not face any pharmacokinetic associated issues in the mere future upon developmental stage. These conjugates may serve as a lead for the discovery of potential anticancer drug candidate with better therapeutic profile.

PT3: A Novel Benzamide Class Histone Deacetylase 3 Inhibitor Improves Learning and Memory in Novel Object Recognition Mouse Model.

The importance of HDAC3 in transcriptional regulation of genes associated with long-term memory is well established. Here, we report a novel HDAC3 inhibitor, PT3, with an excellent blood-brain barrier permeability and ability to enhance long-term memory in mouse model of novel object recognition (NOR). PT3 exhibited higher selectivity for HDAC3 over HDAC1, HDAC6, and HDAC8 compared to the reference compound CI994. PT3 has significant distribution into the brain tissue with Cmax at 0.5 h and t1/2 of 2.5 h. Treatment with PT3 significantly improved the discrimination index in C57/BL6 mice in the NOR model. Brain tissue analysis of mice treated with PT3 for NOR test showed significant increase in H3K9 acetylation in hippocampus. Gene expression analysis by RT-qPCR of the hippocampus tissue revealed upregulation of CREB 1, BDNF, TRKB, Nr4a2, c-fos, PKA, GAP 43, PSD 95 and MMP9 expression in mice treated with PT3. Similar to the phenotype observed in the in vivo experiment, we found upregulation of H3K9 acetylation, CREB 1, BDNF, TRKB, Nr4a2, c-fos, PKA, GAP 43 and MMP9 expression in mouse neuronal (N2A) cells treated with PT3. Thus, our preclinical studies identify PT3 as a potential HDAC3 selective inhibitor that crosses the blood-brain barrier and improves the long-term memory formation in C57/BL6 mice. We propose PT3 as a candidate with therapeutic potential to treat age-related memory loss as well as other disorders with declined memory function like Alzheimer's disease.

HDAC6 as privileged target in drug discovery: A perspective.

HDAC6, a class IIB HDAC isoenzyme, stands unique in its structural and physiological functions. Besides histone modification, largely due to its cytoplasmic localization, HDAC6 also targets several non-histone proteins including Hsp90, α-tubulin, cortactin, HSF1, etc. Thus, it is one of the key regulators of different physiological and pathological disease conditions. HDAC6 is involved in different signaling pathways associated with several neurological disorders, various cancers at early and advanced stage, rare diseases and immunological conditions. Therefore, targeting HDAC6 has been found to be effective for various therapeutic purposes in recent years. Though several HDAC6 inhibitors (HDAC6is) have been developed till date, only two ACY-1215 (ricolinostat) and ACY-241 (citarinostat) are in the clinical trials. A lot of work is still needed to pinpoint strictly selective as well as potent HDAC6i. Considering the recent crystal structure of HDAC6, novel HDAC6is of significant therapeutic value can be designed. Notably, the canonical pharmacophore features of HDAC6is consist of a zinc binding group (ZBG), a linker function and a cap group. Significant modifications of cap function may lead to achieve better selectivity of the inhibitors. This review details the study about the structural biology of HDAC6, the physiological and pathological role of HDAC6 in several disease states and the detailed structure-activity relationships (SARs) of the known HDAC6is. This detailed review will provide key insights to design novel and highly effective HDAC6i in the future.

Discovery of 1,2,3-triazole based quinoxaline-1,4-di-N-oxide derivatives as potential anti-tubercular agents.

A series of thirty one novel 2-(((1-(substituted phenyl)-1H-1,2,3-triazol-4-yl)methoxy)carbonyl)-3-methylquinoxaline-1,4-dioxide (7a-l), 3-(((1-(substituted phenyl)-1H-1,2,3-triazol-4-yl)methoxy)carbonyl)-6-chloro-2-methylquinoxaline-1,4-dioxide (8a-l) and 2-(((1-(substituted phenyl)-1H-1,2,3-triazol-4-yl)methoxy)carbonyl)-6,7-dichloro-3-methylquinoxaline-1,4-dioxide (9a-g) analogues were synthesized, characterized using various analytical techniques and single crystal was developed for the compounds 8 g and 9f. Synthesized compounds were evaluated for in vitro anti-tubercular activity against Mycobacterium tuberculosis H37Rv strain and two clinical isolates Spec. 210 and Spec. 192. The titled compounds exhibited minimum inhibitory concentration (MIC) ranging from 30.35 to 252.00 µM. Among the tested compounds, 8e, 8 l, 9c and 9d exhibited moderate activity (MIC = 47.6 - 52.0 µM) and 8a exhibited significant anti-tubercular activity (MIC = 30.35 µM). Furthermore, 8e, 8 l, and 9d were found to be less toxic against human embryonic kidney, HEK 293 cell lines. Finally, a docking study was also performed using MTB DNA Gyrase (PDB ID: 5BS8) for the significantly active compound 8a to know the exact binding pattern within the active site of the target enzyme.

Structure-activity relationship of human carbonic anhydrase-II inhibitors: Detailed insight for future development as anti-glaucoma agents.

Human carbonic anhydrase-II (hCA-II) is the most dominant physiologic isoform amongst the sixteen reported hCA isoforms. Because of its high availability in the different anatomical, and cellular sites of the eye like retina and lens, it plays a more prominent role in the regulation of intraocular pressure than the other twelve catalytically active hCA isoforms. This isoform is also located in the brain, kidney, gastric mucosa, osteoclasts, RBCs, skeletal muscle, testes, pancreas, lungs, etc. Earlier, hCA-II inhibitors were designed based on the sulfonamides e.g. acetazolamide, dichlorphenamide, methazolamide, ethoxzolamide, etc. and they were used systemically in antiglaucoma therapy. Many successful attempts have been made by the researchers in order to design more potent and effective inhibitors by incorporating various moieties in sulphonamides. Some novel scaffolds like chalcones, thiophenes, organotellurium compounds, dithiocarbamate, selenide, and 2-benzylpyrazine, etc. were also designed as hCA-II inhibitors and their inhibitory efficacy was proved in the nanomolar range. In order to obtain relevant information from the insights of their structure-activity relationship, the reported hCA-II inhibitors from the year 1989 to 2019 were critically analysed. It gave a complete insight into the relationship between their structure-activity and hCA-II inhibition. The broad spectrum of our investigation may help researchers to summarize all the crucial structural information required for the development of more potent hCA-II inhibitors for glaucoma.

Design, synthesis and biological evaluation of 1,2,3-triazole based 2-aminobenzimidazoles as novel inhibitors of LasR dependent quorum sensing in Pseudomonas aeruginosa.

Bacteria regulate their phenotype, growth and population via a signalling pathway known as quorum sensing. In this process, bacteria produce signalling molecules (autoinducers) to recognize their population density. Inhibiting this quorum sensing signalling pathway is one of the potential methods to treat bacterial infection. 2-Aminobenimdazoles are reported to be the strongest inhibitors of quorum sensing against wild-type P. aeruginosa. 1,2,3-Triazole based acyl homoserine lactones are found to be good inhibitors of the quorum sensing LasR receptor. Hence, in our current study, forty 1,2,3-triazole based 2-aminobenzimdazoles were synthesized and characterized using IR, NMR, MS and elemental analysis. A single crystal was developed for N-(1H-benzo[d]imidazol-2-yl)-2-(4-nonyl-1H-1,2,3-triazol-1-yl)acetamide (6d). All final compounds were screened for in vitro quorum sensing inhibitory activity against Pseudomonas aeruginosa. The quorum sensing inhibitory activity was determined in the LasR expressing P. aeruginosa MH602 reporter strain by measuring green fluorescent protein production. Among the title compounds, N-(1H-benzo[d]imidazol-2-yl)-2-(4-(4-chlorophenyl)-1H-1,2,3-triazol-1-yl)acetamide (6i) exhibited good quorum sensing inhibitory activity of 64.99% at 250 µM. N-(1H-Benzo[d]imidazol-2-yl)-2-(4-(4-nitrophenyl)-1H-1,2,3-triazol-1-yl)acetamide (6p) exhibited the most promising quorum sensing inhibitory activity with 68.23, 67.10 and 63.67% inhibition at 250, 125 and 62.5 µM, respectively. N-(1H-Benzo[d]imidazol-2-yl)-2-(4-(4-(trifluoromethyl)phenyl)-1H-1,2,3-triazol-1-yl)acetamide (6o) and N-(5,6-dimethyl-1H-benzo[d]imidazol-2-yl)-2-(4-(4-(trifluoromethyl)phenyl)-1H-1,2,3-triazol-1-yl)acetamide (7l) also exhibited 64.25% and 65.80% quorum sensing inhibition at 250 µM. Compound 6p, the most active quorum sensing inhibitor, also displayed low cytotoxicity at the tested concentrations (25, 50 and 100 µM) against normal human embryonic kidney cell lines. Finally, a docking study using Schrodinger Glide elucidated the possible putative binding mode of the significantly active compound 6p at the active site of the target LasR receptor (PDB ID: 2UV0).