Drugs in Clinical Pipeline: Tefinostat

Tefinostat [Cyclopentyl (S)-2-cyclohexyl-2-((4-(8-(hydroxyamino)-8-oxooctanamido) benzyl) amino)acetate] is a novel monocyte/macrophage-targeted histone deacetylase inhibitor (HDACi) that is cleaved to an active acid, CHR-2847, by an intracellular esterase (human carboxylesterase-1, hCE-1), found only in cells of monocytoid lineage and hepatocytes. 

Tefinostat and its acid CHR-2845 had similar 50% inhibitory concentrations (IC₅₀) when tested against the HDAC activity of HeLa cell nuclear extracts that contain multiple HDAC isoforms (113 nM for Tefinostat, 160 nM for CHR-2847, respectively).

CHR-2847, being a charged molecule, cannot readily leave cells and, hence, selectively accumulates and is active within hCE1- expressing cells, resulting in a 20- to 100-fold increase in the anti-proliferative potency of Tefinostat for monocytic, over non-monocytic, tumour cells. The Esterase Sensitive Motif (ESM) technology used to design Tefinostat is widely applicable and an increase in potency against monocytic tumour cell lines has been demonstrated with a variety of targets [1, 2].

Tefinostat is a much better substrate for hCE1 than for the related, and ubiquitously expressed, hCE2, as evidenced by the rate of production of CHR-2847 when hydrolysed by the two enzymes: thus hCE1 produces CHR-2847 at 4400 pg/ml/ min whereas hCE2 hydrolyses tefinostat much more slowly with a rate less than 200 pg/ml/min.                                

The activity of Tefinostat is as follows:

IC₅₀ (HDAC HeLa cell nuclear extract assay) = 113 nM

Common Name: Tefinostat

Synonyms: CHR-2845; CHR 2845; CHR2845

IUPAC Name: Cyclopentyl (S)-2-cyclohexyl-2-((4-(8-(hydroxyamino)-8-oxooctanamido) benzyl)amino)acetate

CAS Number: 914605-76-8

Mechanism of Action: HDAC Inhibitor; Histone Deacetylase Inhibitor

Indication: Various Cancers

Development Stage: Phase I/II

Company: Chroma Therapeutics

Phase I Study  [3]

In a first-in-human, standard 3 + 3 dose escalating trial of oral, once daily Tefinostat was conducted to determine the safety, tolerability, pharmacokinetic and pharmacodynamic profile of tefinostat in relapsed/refractory haematological diseases. 

Eighteen patients were enrolled at doses of 20-640 mg. Plasma concentrations of Tefinostat exceeded those demonstrated to give in vitro anti-proliferative activity. Flow cytometric pharmacodynamic assays demonstrated monocyte-targeted increases in protein acetylation, without corresponding changes in lymphocytes. 

Dose-limiting toxicities (DLTs) were not observed and dose escalation was halted at 640 mg without identification of the maximum tolerated dose (MTD). Drug-related toxicities were largely Common Toxicity Criteria for Adverse Events grade 1/2 and included nausea, anorexia, fatigue, constipation, rash and increased blood creatinine. 

a: A patient with chronic monomyelocytic leukaemia achieved a bone marrow response, with no change in peripheral monocytes. 

b: An acute myeloid leukaemia type M2 patient showed a more than 50% decrease in bone marrow blasts and clearance of peripheral blasts. 

In conclusion, Tefinostat produces monocyte-targeted HDACi activity and is well tolerated, without the DLTs, e.g. fatigue, diarrhoea, thrombocytopenia, commonly seen with non-targeted HDACi. The early signs of efficacy and absence of significant toxicity warrant further evaluation of Tefinostat in larger studies.

References:

1. Ossenkoppele, G. J.; et. al. A phase I first-in-human study with tefinostat-a monocyte/macrophage targeted histone deacetylase inhibitor-in patients with advanced haematological malignancies. Br J Haematol 2013, 162(2), 191-201.

2. Needham, L.A.; et. al.  Drug targeting to monocytes and macrophages using esterase-sensitive chemical motifs. J Pharmacol Exp Ther 2011, 339, 132-142. 
3. Ossenkoppele, G. J.; et. al. A phase I first-in-human study with tefinostat - a monocyte/macrophage targeted histone deacetylase inhibitor-in patients with advanced haematological malignancies. Br J Haematol 2013, 162(2), 191-201.
4. ClinicalTrials.gov Safety and Tolerability of CHR-2845 to Treat Haematological Diseases or Lymphoid Malignancies (CHR-2845-001). NCT00820508 (retrieved 28-09-2015)

Drugs in Clinical Pipeline: MC1568

MC1568 [(E)-3-(5-((E)-3-(3-fluorophenyl)-3-oxoprop-1-en-1-yl)-1-methyl-1H-pyrrol-2-yl)-N-hydroxyacrylamide], one of the only known selective inhibitors of class IIa histone deacetylase (HDAC) enzymes to be documented in the literature till date. It was identified from a series of novel (aryloxopropenyl)-pyrrolyl hydroxyamides which are structurally related to aroyl-pyrrolyl-hydroxy-amides (APHAs) and were highly selective against the class II (class IIa) histone deacetylase homologue HD1-A. The compound has also been the starting point for a number of structure-activity relationship and molecular modelling studies.

MC1568 was tested against maize HD1-B and HD1-A two mammalian class I and class II (IIa) HDAC homologues. It inhibited HD1-B and HD1-A with IC₅₀ value 38.8 ± 1.16 and 0.22 ± 0.01 uM, respectively. The selectivity index was 176.4 in favour of class IIa. Moreover, its maize HD2 50% inhibitory activity (IC₅₀) was found to be 22.0 ± 1.32 uM.

Being the most selective compound in the maize HD1-B/ HD1-A system, MC1568 was evaluated against human HDAC1 and HDAC4 in comparison with SAHA as reference drug. Human breast cancer ZR-75.1 cell lysates were immunoprecipitated with antibodies against HDAC1 and HDAC4, and inhibitory assays were performed on such immunoprecipitates (IPs) with MC1568 (5 µM) and SAHA (5 µM). Data reported clearly show that MC1568 lacked any inhibitory activity against human HDAC1 (% inhibition of 5 uM = 0%) but was effective in inhibiting human HDAC4 enzyme (% inhibition of 5 uM = 54.9 %) [1].

The activity of MC1568 is as follows:

% Inhibition of HDAC1 @ 5 uM = 0 %

% Inhibition of HDAC4 @ 5 uM = 54.9 %

Common Name: MC1568

Synonyms: MC1568; MC 1568; MC-1568

IUPAC Name: (E)-3-(5-((E)-3-(3-fluorophenyl)-3-oxoprop-1-en-1-yl)-1-methyl-1H-pyrrol-2-yl)-N-hydroxyacrylamide

CAS Number: 852475-26-4

Mechanism of Action: HDAC Inhibitor; Histone Deacetylase Inhibitor; HDAC4 Inhibitor

Indication: Various Cancers

Development Stage: Investigational

Company: Italian University

References:

1. Mai, M.; et. al. Class II (IIa)-selective histone deacetylase inhibitors. 1. Synthesis and biological evaluation of novel (aryloxopropenyl)pyrrolyl hydroxyamides. J Med Chem 2005, 48(9), 3344-3353.

Drugs in Clinical Pipeline: Givinostat

Givinostat [(6-((diethylamino)methyl)naphthalen-2-yl)methyl(4-(hydroxycarbamoyl) phenyl)carbamate] is an orally active, hydroxamic-containing histone deacetylase (HDAC) inhibitor, with broad anti-inflammatory, anti-angiogenic and anti-neoplastic properties.

Givinostat is pan-HDAC inhibitor showing potent inhibition of both Class I (IC₅₀ HDAC 1, 2, 3 and 8 = 198, 325, 157 and 854 nM, respectively) and Class II HDACs (IC₅₀ 4, 5, 6, 7, 9, 10 and 11 = 1059, 532, 524, 541, 315, 340 and 292 nM, respectively). Furthermore, it reduces caspase-1 activity in human peripheral blood mononuclear cells (PBMC) and the secretion of IL-1β and other cytokines at 25-100 nM. At concentrations greater than 200 nM, Givinostat is toxic in vitro [1].

Givinostat has a dual anti-leukemic activity, since it induces apoptosis of both multiple myeloma (MM) and acute myelogenous leukemia (AML) cells and inhibits the production of IL-6 and VEGF by mesenchymal stromal cells (MSCs), two soluble factors crucial for leukemia growth and drug resistance. 

In 2010, Givinostat was granted orphan drug designation in the European Union for the treatment of systemic juvenile idiopathic arthritis and polycythaemia vera.

The activity of Givinostat is as follows:

IC50 (HDAC1 enzyme assay) = 198 nM

IC50 (HDAC2 enzyme assay) = 325 nM

IC50 (HDAC3 enzyme assay) = 157 nM

IC50 (HDAC8 enzyme assay) = 854 nM

IC50 (HDAC4 enzyme assay) = 1059 nM

IC50 (HDAC5 enzyme assay) = 532 nM

IC50 (HDAC7 enzyme assay) = 524 nM

IC50 (HDAC9 enzyme assay) = 541 nM

IC50 (HDAC6 enzyme assay) = 315 nM

IC50 (HDAC10 enzyme assay) = 340 nM

IC50 (HDAC11 enzyme assay) = 292 nM

Common Name: Givinostat

Synonyms: Gavinostat; ITF-2357; ITF 2357; ITF2357

IUPAC Name: (6-((diethylamino)methyl)naphthalen-2-yl)methyl (4-(hydroxycarbamoyl) phenyl)carbamate

CAS Number: 497833-27-9 

Mechanism of Action: HADC Inhibitor; pan-HADC Inhibitor; Histone Deacetylase Inhibitor

Indication: Various Cancers; Solid Tumors

Development Stage: Phase II

Company: Italfarmaco

DNA is tightly wrapped around nuclear histones and maintained in a state of deacetylation by histone deacetylases (HDACs), silencing gene expression. In humans, there are 18 HDACs divided into classes based on their dependence on zinc for enzyme activity. Class I (HDAC 1, 2,3, and 8) and Class II (HDAC 4, 5, 6, 7, 9, 10 and 11) are Zn-dependent enzymes, whereas class III (sirtuins 1-7) are NAD+ dependent. Although HDAC’s deacetylate the highly conserved N-terminal lysines on nuclear histones, HDACs also target cytosolic proteins such as transcription factors and proteins that regulate cell proliferation, migration and death (4). HDAC inhibitors hyperacetylate signal transducers and activators of transcription (STAT) and NFkβ, both of which are associated with decreased inflammation [1].

Low nonapoptotic concentrations of Givinostat reduce pro-inflammatory cytokine production in primary cells in vitro and exhibit anti-inflammatory effects in vivo. In lipopolysaccharide (LPS)-stimulated cultured human peripheral blood mononuclear cells (PBMCs), Givinostat reduced by 50% the release of tumor necrosis factor-alpha (TNFα) at 10 to 22 nM, the release of intracellular interleukin (IL)-1α at 12 nM, the secretion of IL-1β at 12.5 to 25 nM, and the production of interferon-gamma (IFNγ) at 25 nM. There was no reduction in IL-8 in these same cultures. Using the combination of IL-12 plus IL-18, IFNγ and IL-6 production was reduced by 50% at 12.5 to 25 nM, independent of decreased IL-1 or TNFα. There was no evidence of cell death in LPS-stimulated PBMCs at 100 nM Givinostat, using assays for DNA degradation, annexin V, and caspase-3/7. By Northern blotting of PBMCs, there was a 50% to 90% reduction in LPS-induced steady-state levels of TNFα and IFNγ mRNA but no effect on IL-1β or IL-8 levels. Real-time PCR confirmed the reduction in TNFα RNA by Givinostat. Oral administration of 1.0 to 10 mg/kg Givinostat to mice reduced LPS-induced serum TNFα and IFNγ by more than 50%. Anti-CD3-induced cytokines were not suppressed by Givinostat in PBMCs either in vitro or in the circulation in mice. In concanavalin-A-induced hepatitis, 1 or 5 mg/kg of oral Givinostat significantly reduced liver damage [2].

Givinostat inhibited production of growth and angiogenic factors by bone marrow stromal cells, in particular IL-6 and vascular endothelial growth factor (VEGF). Givinostat induced apoptosis in 8/9 MM and 6/7 AML cell lines, as well as 4/4 multiple myeloma (MM) and 18/20 acute myelogenous leukemia (AML) freshly isolated cases, with a mean IC₅₀ of 0.2 uM. Givinostat activated the intrinsic apoptotic pathway, upregulated p21 and downmodulated Bcl-2 and Mcl-1. The drug induced hyperacetylation of histone H3, H4 and tubulin. When studied in more physiological conditions, Givinostat was still strongly cytotoxic for the interleukin-6 (IL-6)-dependent MM cell line CMA-03, or for AML samples maximally stimulated by co-culture on mesenchymal stromal cells (MSCs), but not for the MSCs themselves. Interestingly, Givinostat inhibited the production of IL-6, VEGF and interferon-italic gamma by MSCs by 80-95%. Finally, the drug significantly prolonged survival of severe combined immunodeficient mice inoculated with the AML-PS in vivo passaged cell line already at the 10 mg/kg oral dose [3].

References:
1. Li, S.; et. al. Specific inhibition of histone deacetylase 8 reduces gene expression and production of proinflammatory cytokines in vitro and in vivo. Journal of Biological Chemistry 2014 DOI: http://www.jbc.org/cgi/doi/10.1074/jbc.M114.618454
2. Leoni, F.; et. al. The histone deacetylase inhibitor ITF2357 reduces production of pro-inflammatory cytokines in vitro and systemic inflammation in vivo. Mol Med 2005, 11(1-12), 1-15.
3. Rambaldi, A.; et. al. The histone deacetylase inhibitor ITF2357 has anti-leukemic activity in vitro and in vivo and inhibits IL-6 and VEGF production by stromal cells. Leukemia 2007, 21, 1892-1900.

Drugs in Clinical Pipeline: Compound 7

Compound 7 [tert-butyl (4-(3-((7-(hydroxyamino)-7-oxoheptyl)carbamoyl)isoxazol-5-yl)phenyl)carbamate] a picomolar inhibitor of Histone Deacetylases 6 (HDAC6 IC₅₀ ~ 2 pM) was identified in a series of hydroxamate based HDAC inhibitors. The series of inhibitors contain a phenylisoxazole as the CAP group that has been synthesized using nitrile oxide cycloaddition chemistry.

The activity of Compound 7 is as follows:

IC₅₀ (HDAC1 enzyme assay) = 271 ± 23 nM

IC₅₀ (HDAC2 enzyme assay) = 252 ± 10 nM

IC₅₀ (HDAC3 enzyme assay) = 0.42 ± 0.08 nM

IC₅₀ (HDAC8 enzyme assay) = 6851 ± 707 nM

IC₅₀ (HDAC6 enzyme assay) = 0.002 nM

IC₅₀ (HDAC10 enzyme assay) = 90.7 ± 12 nM

Common Name: Compound 7

Synonyms:  -

IUPAC Name: tert-butyl (4-(3-((7-(hydroxyamino)-7-oxoheptyl)carbamoyl)isoxazol-5-yl) phenyl)carbamate

CAS Number: 1045792-66-2

SMILES: O=C(OC(C)(C)C)NC1=CC=C(C2=CC(C(NCCCCCCC(NO)=O)=O)=NO2)C=C1

Mechanism of Action: HDAC Inhibitor; HDAC6 Inhibitor; Histone Deacetylases 6 Inhibitor

Indication: Various Cancers; Anti-tumor Therapy

Development Stage: Investigational

Company: University of Illinois and Mayo Clinic

The design of HDAC inhibitors as novel therapeutics for the treatment of a wide range of disorders, including cancers, neurodegenerative diseases, and even malaria has generated a lot of interest. These compounds owe their action to their ability to reactivate silenced genes by modulating the condensation status of DNA. The post-translational acetylation status of chromatin is determined by the competing activities of two classes of enzymes, histone acetyltransferases (HATs) and histone deacetylases (HDACs), which control the acetylation of lysine residues making up the histones. In general, HATs function to acetylate lysine groups in nuclear histones, resulting in neutralization of the charges on the histones and a more open, transcriptionally active chromatin structure, while the HDACs function to deacetylate and suppress transcription (the positively charged lysine amine group interacts with the negatively charge DNA organophosphate groups to cause compaction of the chromatin structure). A shift in the balance of acetylation on chromatin may result in changes in the regulation of patterns of gene expression. Many cancers are associated with aberrant transcriptional activity and HDACs and HATs associate with transcription activators and repressors, these enzymes have been identified as attractive targets for cancer therapy. Indeed, inhibitors of the HDACs have been shown to block tumor cell growth and induce differentiation and cell death [1].

HDAC6 plays an important role in deacetylation of tubulin, hence is a huge interest in the researcher and pharma community [2]. Using X-ray crystal structure of HDAC7 (2PQP) template a HDAC6 homology model was built. Preliminary investigations of other class I and class II HDAC homology models indicate that the loop areas surrounding the CAP residues show significant differences among the various HDACs and may eventually account for the enzyme selectivity that has been observed [1].

The enzyme selectivity of compound 7 was explained on the basis of the following drug-target interactions:

1. The ZBG (zinc-binding group) hydroxamic acid group and the six methylene groups bind deep inside the narrow binding pocket.

2. The CAP region of the bound ligand appears to interact with only one side of the protein.

3. The carbonyl group of the Boc group might interact with His499, which may be important in positioning the CAP residue on the surface of the HDAC protein. Such interactions are absent with the other inhibitors included in this study and may thus explain the higher potency observed for this compound.

Compound 7 was tested against five transformed cell lines as well as against two normal cell lines. Compounds 7 is active against both the Mia Paca-2 and Panc04.03 cell lines at the 100 nM level and at the 200-300 nM level against HupT3 [1].

References:

1. Kozikowski, A. P.; et. al. Use of the nitrile oxide cycloaddition (NOC) reaction for molecular probe generation: a new class of enzyme selective histone deacetylase inhibitors (HDACIs) showing picomolar activity at HDAC6. J Med Chem 2008, 51(15), 4370-4373.

2. Bulinski, J. C.; et. al. HDAC6 deacetylation of tubulin modulates dynamics of cellular adhesions. Journal of Cell Science 2007, 120, 1469-1479.

Drugs in Clinical Pipeline: Rocilinostat

Rocilinostat [2-(diphenylamino)-N-(7-(hydroxyamino)-7-oxoheptyl)pyrimidine-5-carboxamide] is novel, orally bioavailable, selective and specific inhibitor of histone deacetylase 6 (HDAC6) with potential antineoplastic activity. 

Rocilinostat, a hydroxamic acid derivative demonstrated potent and selective inhibitory activity against HDAC6, with an enzymatic IC₅₀ value of ~ 5 nM. Rocilinostat is 12-, 10-, and 11-fold less active against HDAC1, HDAC2, and HDAC3 (class I HDACs, IC₅₀ = 58, 48 and 51 nM), respectively. Rocilinostat has minimal activity (IC₅₀ greater than 1 µM) against HDAC4, HDAC5, HDAC7, HDAC9, HDAC11, Sirtuin1, and Sirtuin2, and has slight activity against HDAC8 (IC₅₀ = 0.1 µM) [1].

Acetylon Pharmaceuticals is developing Rocilinostat and it is evaluating Ricolinostat in several clinical studies for the treatment of multiple myeloma and lymphoma. Rocilinostat was chosen from several lead candidates because of its properties that favor drug development. These criteria included at least 10-fold selectivity against HDAC6 compared with class 1 HDACs, minimal activity against other HDAC enzymes, and a lack of significant activity against an extensive panel of receptors, transporters, and enzymes, including kinases. Additional criteria included suitable oral bioavailability in rodents and nonrodents, cellular permeability, metabolic stability, and an appropriate in vitro safety profile with minimal drug-drug interaction, minimal potential for QTc prolongation (hERG channel), and no significant genotoxic signal in mammalian cells. Further, experimental data show that Rocilinostat was less toxic against PBMCs and T cells isolated from healthy volunteers compared with SAHA.

The activity of Rocilinostat is as follows:

IC₅₀ (HDAC6 enzyme assay) = 4.7 nM

IC₅₀ (HDAC2 enzyme assay) = 48 nM

IC₅₀ (HDAC3 enzyme assay) = 51 nM

IC₅₀ (HDAC1 enzyme assay) = 58 nM

IC₅₀ (HDAC8 enzyme assay) = 100 nM

IC₅₀ (HDAC7 enzyme assay) = 1400 nM

IC₅₀ (HDAC4 enzyme assay) = 7000 nM

IC₅₀ (HDAC5 enzyme assay) = 5000 nM

IC₅₀ (HDAC9 enzyme assay) = greater than 10 uM

IC₅₀ (HDAC11 enzyme assay) = greater than 10 uM

IC₅₀ (Sirtuin 1 enzyme assay) = greater than 10 uM

IC₅₀ (Sirtuin 2 enzyme assay) = greater than 10 uM

Common Name: Rocilinostat

Synonyms: ACY-1215; ACY-1215; ACY-1215

IUPAC Name: 2-(diphenylamino)-N-(7-(hydroxyamino)-7-oxoheptyl)pyrimidine-5-carboxamide

CAS Number: 1316214-52-4

Mechanism of Action: HDAC Inhibitor; HDAC6 Inhibitor; Histone Deacetylase 6 Inhibitor

Indication: Multiple Myeloma and Lymphoma

Development Stage: Phase I/II

Company: Acetylon Pharmaceuticals

Pre-Clinical Characterisation

Unlike all other HDACs, HDAC6 has substrate specificity for α-tubulin because of its α-tubulin deacetylase domain. Rocilinostat induces potent acetylation of α-tubulin at very low doses and triggers acetylation of lysine on histone H3 and histone H4 only at higher doses, confirming its specific inhibitory effect on HDAC6 activity. This specific inhibition was also observed in patient multiple myeloma (MM) cells, in which Rocilinostat increased acetylated a-tubulin after 4 hours of treatment. However, after prolonged exposure or with significantly higher concentrations of Rocilinostat, it is possible that the low level of class 1 HDACs (HDAC1, HDAC2, and HDAC3) inhibition by Rocilinostat may also contribute to MM cell cytotoxicity and to potent inhibition of HDAC6.

References:

1. Santo, L.; et. al. Preclinical activity, pharmacodynamic, and pharmacokinetic properties of a selective HDAC6 inhibitor, ACY-1215, in combination with bortezomib in multiple myeloma. Blood 2012, 119(11), 2579-2589.

Drugs in Clinical Pipeline: Abexinostat

Abexinostat [3-(dimethylaminomethyl)-N-[2-[4-(hydroxycarbamoyl)phenoxy]ethyl]-1-benzofuran-2-carboxamide] is a novel, second-generation phenyl hydroxamic acid-based, orally bioavailable HDAC inhibitor that has shown to have in vitro and in vivo activity  against a broad array of cancers, including hematopoietic malignancies and bone and soft-tissue sarcomas. Abexinostat inhibited pure recombinant HDAC1 with a Ki of 0.007 micromol/L, and also inhibited the other HDAC isozymes HDAC2, HDAC3/SMRT, HDAC6, HDAC8, and HDAC10 in the nanomolar range. Treatment of cultured tumor cell lines grown in vitro with Abexinostat resulted in the accumulation of acetylated histone and acetylated tubulin, resulting in an inhibition of tumor cell growth and the induction of apoptosis [1]. Abexinostat is being developed by Pharmacyclics Inc.

Common Name: Abexinostat

Synonyms: CRA-024781; CRA 024781; CRA024781; PCI-24781; PCI24781; PCI 24781

IUPAC Name: 3-(dimethylaminomethyl)-N-[2-[4-(hydroxycarbamoyl)phenoxy]ethyl]-1-benzofuran-2-carboxamide

CAS Number: 783355-60-2; 783356-67-2 (hydrochloride)

Mechanism of Action: HDAC Inhibitor; Histone Deacetylase Inhibitor

Indication: Various Cancers

Development Stage: Phase II

Company: Pharmacyclics Inc

It has also shown good tolerability and activity in Phase I and II clinical trials against lymphoma, as well as against solid tumors in Phase-I trials. Additionally, it acts as a potent radiosensitizing agent and is synergistic with cytotoxic chemotherapy, such as doxorubicin in preclinical models [2].

References:

1. Buggy, J. J.; et. al. CRA-024781: a novel synthetic inhibitor of histone deacetylase enzymes with antitumor activity in vitro and in vivo. Mol Cancer Ther 2006, 5(5), 1309-1317.

2. Sholler, G. S.; et. al. PCI-24781 (abexinostat), a novel histone deacetylase inhibitor, induces reactive oxygen species-dependent apoptosis and is synergistic with bortezomib in neuroblastoma. J Cancer Ther Res 2013, 2, 21.

Drugs in Clinical Pipeline: Resminostat

Resminostat [(E)-3-(1-((4-((dimethylamino)methyl)phenyl)sulfonyl)-1H-pyrrol-3-yl)-N-hydroxyacrylamide] is an orally bioavailable inhibitor of histone deacetylases (HDACs) with potential antineoplastic activity.

The inhibition of HDACs by resminostat results in an accumulation of highly acetylated histones, followed by an abduction of chromatin remodeling, inhibition of tumor suppressor genes transcription and cell division, and finally tumor cell apoptosis.

Resminostat is a potent inhibitor of HDACs 1, 3 and 6 [IC50 = 43-72 nmol/l] representing HDAC classes I and II and induces hyperacetylation of histone H4 in MM cells. Low micromolar concentrations of resminostat abrogated cell growth and strongly induced apoptosis (IC50 = 2.5-3 micromol/l in 3 out of 4 MM cell lines) in MM cell lines as well as primary MM cells. At 1 micromol/l, resminostat inhibited proliferation and induced G0/G1 cell cycle arrest in 3 out of 4 MM cell lines accompanied with decreased levels of cyclin D1, cdc25a, Cdk4 and pRb as well as upregulation of p21. Resminostat decreased phosphorylation of 4E-BP1 and p70S6k indicating an interference with Akt pathway signalling [1].

Resminostat is developed by 4SC in Europe and its Japanese development partner Yakult Honsha in Asia-has been investigated to date in a broad clinical Phase I/II programme in the four indications of liver cancer (hepatocellular carcinoma, HCC), Hodgkin Lymphoma (HL), colorectal cancer (CRC), and non-small-cell lung cancer (NSCLC). In all trials, the compound showed good safety and tolerability as well as promising anti-tumour activity both as monotherapy and in combination with other cancer therapies.

In preclinical studies, Resminostat has been shown to effectively inhibit epithelial-mesenchymal transition (EMT). EMT, which may be promoted through the administration of certain conventional cancer therapies, leads to the formation of particularly aggressive tumour cells, which ultimately may result in greater proliferation of cancer cells in patients and the patients' death. HDAC inhibitors modify the three-dimensional chromatin DNA structure of tumour cells and can trigger cell differentiation, which can ultimately result in programmed cell death (apoptosis). HDAC inhibitors therefore offer a mechanism of action that has the potential to halt tumour progression and induce tumour regression. Furthermore, Resminostat-due to its epigenetic mode of action-can develop an additional synergetic effect in combined treatments with other traditional cancer therapies and also fight the development of resistance to other cancer medications [2].

References:

1. Mandl-Weber, S.; et. al. The novel inhibitor of histone deacetylase resminostat (RAS2410) inhibits proliferation and induces apoptosis in multiple myeloma (MM) cells. Br J Haematol 2010, 149(4), 518-528.
2. Resminostat

Drugs in Clinical Pipeline: 4SC-202

4SC-202 [(E)-N-(2-aminophenyl)-3-(1-((4-(1-methyl-1H-pyrazol-4-yl)phenyl)sulfonyl) -1H-pyrrol-3-yl)acrylamide] is an oral anti-cancer compound and possesses a unique mode of action consisting of both epigenetic regulation and cancer stem cell targeting.  4SC-202 is a selective inhibitor of LSD1 and HDACs 1, 2, and 3 with a unique combination of anti-cancer mode of actions, namely epigenetic regulation and targeting of cancer stem cells.

Drugs in Clinical Pipeline: CUDC-101

CUDC-101 [7-(4-(3-ethynylphenylamino)-7-methoxyquinazolin-6-yloxy)-N-hydroxyheptanamide] is a novel molecule which simultaneously inhibits histone deacetylase (HDAC) and the receptor kinases epidermal growth factor receptor (EGFR) and human epidermal growth factor receptor 2 (HER2) in cancer cells - all of which are overexpressed in many cancers. CUDC-101, currently being developed by Curis, Inc., and was in Phase I trail for the common type of especially aggressive head and neck cancer that is not caused by the human papilloma virus (HPV-) but rather by tobacco or alcohol.

Strengthened by its integrated histone deacetylase inhibition, CUDC-101 synergistically blocked key regulators of EGFR/HER2 signaling pathways, also attenuating multiple compensatory pathways, such as AKT, HER3, and MET, which enable cancer cells to escape the effects of conventional EGFR/HER2 inhibitors.  It potently blocks the receptor tyrosine kinases EGFR (aka HER1) and HER2 (IC50s = 2.4 and 16.4 nM, respectively). CUDC-101 also inhibits the activity of class I and class II HDACs at nanomolar concentrations (e.g., IC50s = 4.5, 12.6, 13.2, and 11.4 nM for HDAC1, 2, 4, and 5, respectively). It has only weak effects on over 60 other kinases when tested at 5 µM [1].

Cancer cells that have acquired resistance to single-target EGFR inhibitors through upregulation of AXL or loss of E-cadherin remain sensitive to CUDC-101, which inhibits MET- and AXL-mediated signaling, restores E-cadherin expression, and reduces cell migration. CUDC-101 also efficiently inhibited the proliferation of MET-overexpressing non-small cell lung cancer and gastric cancer cell lines and inhibited the migration and invasion of invasive tumor cells. Taken together, these results suggest that coupling HDAC and HER2 inhibitory activities to an EGFR inhibitor may potentially be effective in overcoming drug resistance and preventing cancer cell migration [2].

Phase I

In the phase 1 trial CUDC-101 was combined with the standard of care for treatment of head and neck cancer, which includes the chemotherapy drug cisplatin and radiation. Specifically, the study was performed in 12 medium- to high-risk head and neck cancer patients. At 18 months median follow up, one patient's cancer had worsened, two had died, and nine remained free of disease. Testing of blood and tumor samples showed that CUDC-101 had indeed inhibited the action of EGFR, HDAC and HER2. Although the MTD was identified, a high rate of dose-limiting toxicity (DLT)-independent discontinuation of CUDC-101 suggests a need for alternate schedules or routes of administration [4].

References:

1. Lai, C. J.; et. al. CUDC-101, a multitargeted inhibitor of histone deacetylase, epidermal growth factor receptor, and human epidermal growth factor receptor 2, exerts potent anticancer activity. Cancer Res 2010, 70(9), 3647-3656.
2. Wang, J.; et. al. Potential advantages of CUDC-101, a multitargeted HDAC, EGFR, and HER2 inhibitor, in treating drug resistance and preventing cancer cell migration and invasion. Mol Cancer Ther 2013, 12(6), 925-936.
3. Cai, X.; et. al. Discovery of 7-(4-(3-ethynylphenylamino)-7-methoxyquinazolin-6-yloxy)-N-hydroxyheptanamide (CUDc-101) as a potent multi-acting HDAC, EGFR, and HER2 inhibitor for the treatment of cancer. J Med Chem 2010, 53(5), 2000-2009.
4. Galloway, T. J.; et. al. A Phase I Study of CUDC-101, a Multitarget Inhibitor of HDACs, EGFR, and HER2, in Combination with Chemoradiation in Patients with Head and Neck Squamous Cell Carcinoma. Clin Cancer Res 2015, 21(7), 1566-1573.

Chidamide

Common name: Chidamide; CS055; HBI-8000; Epidaza

Trademarks: Epidaza

Molecular Formula: C22H19FN4O2

CAS Registry Number: 743420-02-2

CAS Name: N-(2-amino-5-fluorophenyl)-4-[[[1-oxo-3-(3-pyridinyl)-2-propen-1-yl]amino]methyl]-benzamide

Molecular Weight: 390.416

SMILES: O=C(NCC1=CC=C(C(NC2=CC(F)=CC=C2N)=O)C=C1)/C=C/C3=CC=CN=C3

InChI Key: WXHHICFWKXDFOW-BJMVGYQFSA-N

InChI: 1S/C22H19FN4O2/c23-18-8-9-19(24)20(12-18)27-22(29)17-6-3-16(4-7-17)14-26-21(28)10-5-15-2-1-11-25-13-15/h1-13H,14,24H2,(H,26,28)(H,27,29)?/b10-5+

Activity: HDAC Inhibitor; Cancer Drug; Histone Deacetylase Inhibitor; HDAC-1, 2,3,10 Inhibitor; Treatment for Peripheral T-cell Lymphomas; Treatment for PTCL

Status: Launched 2014 (China)

Originator: Shenzhen Chipscreen Biosciences Ltd

Chiamide synthesis: US7244751B2

Procedure:

Step a: To a suspension of 0.33 g (2.01 mmol) of N,N'-carbonyldiimidazole in tetrahydrofunan (10 ml) is added drop-wise a solution of 0.30 g (2.01 mmol) of 3-pyridineacrylic acid at 0 °C. Then, the mixture is stirred at room temperature for 3 hours and added drop-wise to a separately prepared 2.0 ml (2.00 mmol) of 1N aqueous sodium hydroxide solution including 0.30 g (2.00 mmol) of 4-aminomethylbenzoic acid, followed by stirring at room temperature for 8 hours. The reaction mixture is evaporated under vacuum. To the residue is added a saturated solution of sodium chloride (2 ml), then the mixture is neutralized with concentrated hydrochloric acid to pH 5. The deposited white solid is collected by filtration, washed with ice-water, and then dried to give 4-[N-(Pyridin-3-ylacryloyl)aminomethyl]benzoic acid (0.46 g, 82%). HRMS calcd for C16H14N2O3: 282.2988. Found: 282.2990. MA calcd for: C16H14N2O3: C, 68.07%; H, 5.00%; N, 9.92%. Found: C, 68.21%; H, 5.03%; N, 9.90%.

Step b: To a suspension of 0.29 g (1.78 mmol) of N,N'-carbonyldiimidazole in tetrahydrofunan (15 ml) is added 0.50 g (1.78 mmol) of 4-[N-(Pyridin-3-ylacryloyl)aminomethyl]benzoic acid, followed by stirring at 45 °C. for 1 hour. After cooling, the reaction mixture is added to a separately prepared tetrahydrofiman (10 ml) solution including 0.28 g (2.22 mmol) of 4-fluoro-1,2-phenylenediamine and 0.20 g (1.78 mmol) of trifluoroacetic acid at room temperature. After reaction at room temperature for 24 hours, the deposited white solid is collected by filtration, washed with tetrahydrofunan, and then dried to give N-(2-amino-4-fluorophenyl)-4-[N-(Pyridin-3-ylacryloyl)aminomethyl]benzamide (0.40 g, 57%). 1H NMR (300 MHz, DMSO-d6): dppm: 4.49 (2H, d), 4.84 (2H, br.s), 6.60 (1H, t), 6.80 (2H, m),696 (1H, t), 7.18 (1H, d), 7.42 (2H, d), 7.52 (1H, d), 7.95 (2H, d), 8.02 (1H, d), 8.56 (1H, d), 8.72 (1H, br. t), 8.78 (1H, s), 9.60 (1H, br.s). IR (KBr) cm1: 3310, 1655, 1631, 1524, 1305, 750. HRMS calcd for C22H19N4O2F: 390.4170. Found: 390.4172. MA calcd for C22H19N4O2F: C, 67.68%; H, 4.40%; N, 14.35%. Found: C, 67.52%; H, 4.38%; N, 14.42%.

Activity:

Chidamide is an orally bioavailable, low-nanomolar inhibitor of cancer-associated histone deacetylase (HDAC) enzymes with favorable pharmacology and tolerability profiles relative to existing benzamide and non-benzamide HDAC inhibitors. It targets specifically the subtype 1, 2, 3 of Class I and subtype 10 of Class IIb HDAC and is being studied in multiple clinical trials as a single agent or in combination with chemotherapeutic agents for the treatment of various hematological and solid cancers. In clinically administrated concentrations, it demonstrated a unique epigenetic mechanism of actions against tumor cell development, involving preferential induction of growth arrest and apoptosis in blood and lymphoid-derived tumor cells, activation of NK-mediated and CD8-mediated antigen-specific cellular anti-tumor immunity, differentiation of tumor stem cells, reversal of drug-resisting tumor cells and epithelia to mesenchymal transition, which are hallmarks of treatment resistance, tumor cell metastasis and recurrence.

Chidamide was shown to enhance the cytotoxic effect of human peripheral mononuclear cells ex vivo on K562 target cells, accompanied by the upregulation of proteins involved in NK cell functions. Furthermore, the expression of a number of genes involved in immune cell-mediated antitumor activity was observed to be upregulated in peripheral white blood cells from two T-cell lymphoma patients who responded to chidamide administration [1].

In a reported study, researchers investigated the effects of Chidamide on proliferation, differentiation and apoptosis in human leukaemia cell lines and primary myeloid leukaemia cells. The results showed that 

1. At low concentrations ( less than 1 µM), Chidamide induced G1 arrest.
2. At moderate concentrations (0.5 µM-2 µM), Chidamide induced differentiation, as determined by the increased expression of the myeloid differentiation marker CD11b. 
3. At relatively high concentrations (2 µM-4 µM), Chidamide potently induced caspase-dependent apoptosis. 
4. Co-treatment with the ROS (reactive oxygen species) scavengers N-acetyl-L-cysteine or Tiron blocked Chidamide-induced cell differentiation and apoptosis, suggesting an essential role for ROS in these effects. Cytochrome c release and ROS-mediated mitochondrial dysfunction are involved in Chidamide-induced apoptosis of leukaemia. 
5. In addition to cell lines, Chidamide also exhibits therapeutic effects in human primary leukaemia cells. 
6. Moreover, daily oral Chidamide treatment of nude mice bearing HL60 cell xenografts suppressed tumour growth, induced tumour cell apoptosis and prolonged the survival of tumour-bearing mice. 
In conclusion, findings demonstrate that Chidamide is a novel HDACi with potential chemotherapeutic value in several haematological malignancies, especially leukaemia [2].

References:
1. Ning, Z. Q.; et. al. Chidamide (CS055/HBI-8000): a new histone deacetylase inhibitor of the benzamide class with antitumor activity and the ability to enhance immune cell-mediated tumor cell cytotoxicity. Cancer Chemother Pharmacol 2012, 69(4), 901-909. (activity)
2. Gong, K.; et. al. CS055 (Chidamide/HBI-8000), a novel histone deacetylase inhibitor, induces G1 arrest, ROS-dependent apoptosis and differentiation in human leukaemia cells. Biochem J 2012, 443(3), 735-746. (activity)

3. Hu, W.; et. al. N-(2-amino-5-fluorophenyl)-4-[N-(Pyridin-3-ylacryloyl) aminomethyl ]benzamide or other derivatives for treating cancer and psoriasis. US7244751B2
4. Lu, X.; et. al. Crystal form of chidamide, preparation method and use thereof. WO2014082354A1
5. Yin, Z.-H.; et. al. Synthesis of chidamide,a new histone deacetylase (HDAC) inhibitor. Chin J New Drugs 2004, 13(6), 536-538. (starts with basic raw materials)

Panobinostat

Common name: Panobinostat; LBH-589; LBN-589; LBH589; NVP-LBH589; Farydak
Trademarks:  Farydak
Molecular Formula: C21H23N3O2 
CAS Registry Number: 404950-80-7
CAS Name: (2E)-N-hydroxy-3-[4-({[2-(2-methyl-1H-indol-3-yl)ethyl]amino}methyl)phenyl]
acrylamide
Molecular Weight: 349.426
SMILES: CC1=C(C2=CC=CC=C2N1)CCNCC3=CC=C(C=C3)C=CC(=O)NO
InChI Key: FPOHNWQLNRZRFC-ZHACJKMWSA-N
InChI: InChI=1S/C21H23N3O2/c1-15-18(19-4-2-3-5-20(19)23-15)12-13-22-14-17-8-6-16(7-9-17)10-11-21(25)24-26/h2-11,22-23,26H,12-14H2,1H3,(H,24,25)/b11-10+
Activity: Histone Deacetylase Inhibitor; HDAC Inhibitor; pan-HDAC Inhibitior; Treatment of  Multiple Myeloma; Treatment of  Pancreatic Cancer Cells
Status: Launched 2015
Originator: Novartis

Panobinostat synthesis: J Med Chem 2011, 54(13), 4694–4720

Panobinostat synthesis: US7989639B2

References:

1. Wang, H.; et. al. Discovery of (2E)-3-{2-Butyl-1-[2-(diethylamino)ethyl]-1H-benzimidazol-5-yl}-N-hydroxyacrylamide (SB939), an Orally Active Histone Deacetylase Inhibitor with a Superior Preclinical Profile. J Med Chem 2011, 54(13), 4694–4720. (synthesis in suppl. sheets)

2. Parker, D. J.; et. al. Process for making salts of N-hydroxy-3-[4-[[[2-(2-methyl-1H-indol-3-yl)ethyl]amino]methyl]phenyl]-2E-2-propenamide. US7989639B2

3. Bair, K. W.; et. al. Hydroxamate derivatives useful as deacetylase inhibitors. WO2002022577A2 (gives an insight to synthetic route, but doesnt provide the desired molecule)