Pracinostat (SB939) 929016-96-6_HDAC_DNA Damage_Signaling Pathways_Products

Pracinostat (formerly SB-939) is a novel, potent and orally bioavailable pan-HDAC (histone deacetylase) inhibitor with potential anticancer activity. As opposed to class III HDACs, it shows comparatively higher selectivity (more than 1000-fold) for class I, class II, and class IV HDACs. Prostate carcinomas, colon cancer, ovarian cancer, acute myeloid leukemia (AML), B cell lymphoma, and other cancer cell lines are among those against which it exhibits strong anti-proliferative activity in vitro. In order to treat acute myelocytic leukemia (AML) and T-cell lymphoma, pracinostat was given FDA Orphan Drug status in March 2014.

Physicochemical Properties

Molecular Formula

C20H30N4O2

Molecular Weight

358.48

Exact Mass

358.236

Elemental Analysis

C, 67.01; H, 8.44; N, 15.63; O, 8.93

CAS #

929016-96-6

Related CAS #

929016-96-6

PubChem CID

49855250

Appearance

White to light brown solid powder

Density

1.1±0.1 g/cm3

Index of Refraction

1.568

LogP

4.45

Hydrogen Bond Donor Count

Hydrogen Bond Acceptor Count

Rotatable Bond Count

Heavy Atom Count

Complexity

453

Defined Atom Stereocenter Count

SMILES

C(N1C(CCCC)=NC2C=C(C=CC1=2)/C=C/C(=O)NO)CN(CC)CC

InChi Key

JHDKZFFAIZKUCU-ZRDIBKRKSA-N

InChi Code

InChI=1S/C20H30N4O2/c1-4-7-8-19-21-17-15-16(10-12-20(25)22-26)9-11-18(17)24(19)14-13-23(5-2)6-3/h9-12,15,26H,4-8,13-14H2,1-3H3,(H,22,25)/b12-10+

Chemical Name

(E)-3-[2-butyl-1-[2-(diethylamino)ethyl]benzimidazol-5-yl]-N-hydroxyprop-2-enamide

Synonyms

SB939; SB-939; SB 939

HS Tariff Code

2934.99.9001

Storage

Powder-20°C 3 years

4°C 2 years

In solvent -80°C 6 months

-20°C 1 month

Shipping Condition

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

Biological Activity

Targets

HDAC10 ( IC50 = 40 nM ); HDAC3 ( IC50 = 43 nM ); HDAC5 ( IC50 = 47 nM ); HDAC1 ( IC50 = 49 nM ); HDAC4 ( IC50 = 56 nM ); HDAC9 ( IC50 = 70 nM ); HDAC11 ( IC50 = 93 nM ); HDAC2 ( IC50 = 96 nM ); HDAC7 ( IC50 = 137 nM ); HDAC8 ( IC50 = 140 nM ); HDAC6 ( IC50 = 1008 nM ); MBLAC2 ( IC50 < 10 nM )

ln Vitro

In vitro activity: Pracinostat (SB939) is a strong new hydroxamate-based inhibitor of HDACs class I, II, and IV.It has no effect on the class III isoenzyme SIRT I, but it inhibits the isolated enzymes with Ki values of 19 to 48 nM for class I, 16 to 247 nM for class II, and 43 nM for class IV. The HCT-116 colon cancer cell line and the HL-60 acute myeloid leukemia cell line are both affected by SB939; their IC50 values are 0.48 μM and 70 nM, respectively. At concentrations as high as 100 μM, SB939 does not impede the proliferation of normal human dermal fibroblasts[1]. CYP2C19 is inhibited by pracinostat (SB939, compound 3), with an IC50 of 5.78 μM. With IC50s of 0.48 ± 0.21, 0.56 ± 0.08, 0.48 ± 0.27, and 0.34 ± 0.06, SB939 exhibits strong activities against A2780, COLO 205, HCT-116, and PC-3 cell lines[2]. In JAK2^V617F and FLT-ITD cell lines, precinostat downregulates JAK and FLT3 signaling. When combined with pacritinib, precinostat exhibits synergy. In vitro synergy between pacritinib and pranistat on apoptosis and STAT signaling is demonstrated. In JAK2^V617F or FLT3-ITD AML cell lines, precinostat synergistically inhibits the growth of several AML subtypes and is a potent inhibitor of AML subtype proliferation when used alone[3].

ln Vivo

Pracinostat (SB939, 25-100 mg/kg) significantly inhibits the growth of HCT-116 xenografts at different doses. Tumor tissue is where SB939 preferentially assembles. In the Apcmin genetic colon cancer mouse model, SB939 (50 or 75 mg/kg) demonstrates anti-tumor activities[1]. Mice carrying MV4-11 xenografts exhibit a significant reduction in tumor growth inhibition (TGI) when given pracinostat (25 or 50 mg/kg per day for 21 days). The reduction is 59 and 116%, respectively. In two separate in vivo models of human AML, the combination of pracistat (75 mg/kg, q.o.d.) and pracitinib is effective and synergistic. When it comes to AML-induced plasma cytokines, growth factors, and chemokines, prancinotide and pacritinib work in concert[3].

Enzyme Assay

All recombinant HDAC enzymes are expressed in S*BIO through cloning, except for SIRT1. The assay buffer (25 mM Tris-HCl, pH 7.5; 137 mM NaCl; 2.7 mM KCl, 1 mM MgCl2, and 1 mg/mL BSA), various concentrations of SB939, and the fluorogenic deacetylase substrate Flour de LysTM are all included in the reaction mix, which has a total reaction volume of 33 μL. The mixture is then incubated at room temperature for two hours. After adding 16 μL of Flour de LysTM developer, incubate for an extra 10 minutes. With a microplate reader, the light emission is measured at 460 nm. To generate IC50 values, use the XLfit software.

Cell Assay

Prior to treating with SB939, cells are seeded at a predefined optimal density in 96-well plates during the log growth phase, and they are allowed to rest for either 24 hours (for adherent cells) or 2 hours (for suspension cells). All the experiments are conducted in triplicates for 96 hours using 1% solvent. For adherent cells, the CyQUANT Cell Proliferation Assay Kit is used, and for suspension cells, the CellTiter96 Aqueous One solution cell proliferation kit. The total volume used in the experiments is 100 μL, and the concentrations of SB939 are diluted nine times in serial order to get from 100 μM to 1.5 nM. The XLfit software is utilized to ascertain the IC50 [1].

Animal Protocol

Standard rodent diet is fed to both male ApcMin/+ mice and female C57BL/6 mice. Mice with the verified mutation who are between 16 and 20.5 weeks old and score positively in the hemocult assay are selected for the study. Mice receive intraperitoneal injections (i.p.) of 40 mg/kg 5-FU once daily for five days of treatment, followed by a nine-day recovery period and five more days of treatment. The injection volume is 200 μL per 20 g body weight. Treatment with SB939 is administered orally once daily at 50 or 75 mg/kg for a continuous 21 days. The small intestine, caecum, and colon are removed on the final day of treatment; they are then cut into segments and spread flat on plastic film in a formaldehyde bath after being fixed with repeated injections of 4% PBS-buffered formaldehyde into the gut lumen. Under a dissection microscope, tumor load is measured. The samples are evaluated and analyzed while blinded[1].

ADME/Pharmacokinetics

Absorption, Distribution and Excretion
Oral bioavailability in mice is 34%.

References

[1]. SB939, a novel potent and orally active histone deacetylase inhibitor with high tumor exposure and efficacy in mouse models of colorectal cancer. Mol Cancer Ther. 2010 Mar;9(3):642-52.

[2]. 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 Jul 14;54(13):4694-720.

[3]. The oral HDAC inhibitor pracinostat (SB939) is efficacious and synergistic with the JAK2 inhibitor pacritinib (SB1518) in preclinical models of AML. Blood Cancer J. 2012 May;2(5):e69.

[4]. Target deconvolution of HDAC pharmacopoeia reveals MBLAC2 as common off-target. Nat Chem Biol. 2022 Aug;18(8):812-820.

Additional Infomation

Pracinostat is a hydroxamic acid that is N-hydroxyacrylamide which is substituted at position 3 by a 2-butyl-1-[2-(diethylamino)ethyl]-1H-benzimidazol-5-yl group (the E isomer). An orally available pan-histone deacetylase inhibitor with demonstrated activity in the treatment of advanced solid tumours. It has a role as an EC 3.5.1.98 (histone deacetylase) inhibitor, an antineoplastic agent, an apoptosis inducer and an antimalarial. It is an olefinic compound, a hydroxamic acid, a benzimidazole and a tertiary amino compound.
Pracinostat is a novel HDAC inhibitor with improved in vivo properties compared to other HDAC inhibitors currently in clinical trials, allowing oral dosing. Data demonstrate that Pracinostat is a potent and effective anti-tumor drug with potential as an oral therapy for a variety of human hematological and solid tumors.
Pracinostat is an orally available, small-molecule histone deacetylase (HDAC) inhibitor with potential antineoplastic activity. Pracinostat inhibits HDACs, which may result in the accumulation of highly acetylated histones, followed by the induction of chromatin remodeling; the selective transcription of tumor suppressor genes; the tumor suppressor protein-mediated inhibition of tumor cell division; and, finally, the induction of tumor cell apoptosis. This agent may possess improved metabolic, pharmacokinetic and pharmacological properties compared to other HDAC inhibitors.

Drug Indication
For the treatment of various forms of cancer.
Treatment of acute myeloid leukaemia

Mechanism of Action
Inhibition of HDAC activity allows for the accumulation of acetyl groups on the histone lysine residues resulting in an open chromatin structure and transcriptional activation. In vitro, SB939 causes the accumulation of acetylated histones and induces cell cycle arrest and/or apoptosis of some transformed cells. The mechanism of the antineoplastic effect of SB939 has not been fully characterized.

Pharmacodynamics
SB939 is a novel compound with superior pharmaceutical, metabolic and pharmacokinetic properties. SB939 has demonstrated excellent in vivo anti-tumour activity in various animal models with dose proportional pharmacodynamic effects. The pharmacokinetics and pharmacodynamic attributes of SB939 explain and differentiate it as the best in class HDAC inhibitor.

Solubility Data

Solubility (In Vitro)

DMSO: 72~250 mg/mL (200.8~697.4 mM)

Water: <1 mg/mL

Ethanol: ~27 mg/mL (~75.3 mM)

Solubility (In Vivo)

Solubility in Formulation 1: ≥ 2.75 mg/mL (7.67 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 27.5 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.

Solubility in Formulation 2: ≥ 2.08 mg/mL (5.80 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

Solubility in Formulation 3: ≥ 2.08 mg/mL (5.80 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

Solubility in Formulation 4: 1% DMSO+30% polyethylene glycol+1% Tween 80: 30mg/mL

(Please use freshly prepared in vivo formulations for optimal results.)

Preparing Stock Solutions		1 mg	5 mg	10 mg
	1 mM	2.7896 mL	13.9478 mL	27.8956 mL
	5 mM	0.5579 mL	2.7896 mL	5.5791 mL
	10 mM	0.2790 mL	1.3948 mL	2.7896 mL

*Note: Please select an appropriate solvent for the preparation of stock solution based on your experiment needs. For most products, DMSO can be used for preparing stock solutions (e.g. 5 mM, 10 mM, or 20 mM concentration); some products with high aqueous solubility may be dissolved in water directly. Solubility information is available at the above Solubility Data section. Once the stock solution is prepared, aliquot it to routine usage volumes and store at -20°C or -80°C. Avoid repeated freeze and thaw cycles.

PeptideDB

Pracinostat (SB939) 929016-96-6

CAS No.: 929016-96-6

Physicochemical Properties

Biological Activity

Solubility Data