SK1-I , BML 258
Sphingosine kinase 1 (SphK1) inhibitor; antiproliferative
- D-erythro-Pent-1-enitol, 1,2,4-trideoxy-4-(methylamino)-1-(4-pentylphenyl)-, (1E)-
||(2R,3S,4E)-N-methyl-5-(4′-pentylphenyl)-2-aminopent-4-ene-1,3-diol . HCl
||C17H27NO2 . HCl
- Originator Enzo Biochem; Virginia Commonwealth University
- Developer Enzo Biochem
- Class Antineoplastics; Small molecules
- Mechanism of Action Sphingosine kinase inhibitors
- Preclinical Autoimmune hepatitis; Haematological malignancies; Liver cancer; Solid tumours
- 07 May 2019 Preclinical trials in Liver cancer in USA (unspecified route)
- 03 Dec 2018 SK1 I is available for licensing as of 03 Dec 2018. http://www.enzo.com/
- 03 Dec 2018 Enzo Biochem has patent pending for SK1 I worldwide
SK1 I, a small molecule that specifically inhibits sphingosine kinase 1, is being developed by Enzo Biochem for the treatment of cancer and autoimmune diseases. Preclinical development is underway for the treatment of solid tumours, liver cancer, haematological malignancies and autoimmune hepatitis in the US.
As at December 2018, Enzo Biochem seeks partners for the development of SK1
SK1-I is a sphingosine analog and a sphingosine competitive inhibitor specific for sphingosine kinase 1 (SK1), with ki~10µM and excellent water solubility. It is not to be confused with SKI-I, 5-naphthalen-2-yl-2H-pyrazole-3-carboxylic acid (2-hydroxy-naphthalen-1-ylmethylene)-hydrazide, CAS 306301-68-8, a noncompetitive inhibitor of both SK1 and SK2 with poor water solubility (K.J. French, et al., 2006; N.J. Pyne and S. Pyne, 2010). SK1-I does not inhibit SK2, PKCα, PKCδ, PKA, AKT1, ERK1, EGFR, CDK2, IKKβ or CamK2β. Not only does it decrease sphingosine-1-phosphate levels, it also causes an accumulation of its proapoptotic precursor ceremide. Inhibits tumor cell growth in vitro and in vivo.
This patent was granted in July 30, 2019 and set to expire on October 24, 2038. Claims methods for synthesizing the compound (2R,3S,4E)-N-methyl-5-(4′-pentylphenyl)-2-aminopent-4-ene-1,3-diol (also known as SK1-I and BML-258 (as HCl salt)) and its intermediates.
(2R,3S,4E)-N-methyl-5-(4′-pentylphenyl)-2-aminopent-4-ene-1,3-diol, also known as SK1-I and BML-258 (as HCl salt), is a pharmaceutical inhibitor of sphingosine kinase 1 initially described in Paugh et al., Blood. 2008 Aug. 15; 112(4): 1382-1391. An existing method for synthesizing SK1-I is disclosed in U.S. Pat. No. 8,314,151.
The invention provides methods and intermediate compounds for synthesizing the compound (2R,3 S,4E)-N-methyl-5-(4′-pentylphenyl)-2-aminopent-4-ene-1,3-diol, also known as SK1-I, and related compounds. The structure of SK1-I is shown below.
A step-wise synthesis of SK1-I according to the invention is exemplified as follows.
N-Boc-(D)-Serine Methyl Ester
To an ice-cooled suspension of the (D)-Serine methyl ester hydrochloride (62.24 g, 0.4 mol) in dichloromethane (600.0 mL), triethylamine (40.4 g, 0.4 mol) was added. After the mixture was stirred for 30 min, Boc anhydride (96.0 g, 0.44 mol) in dichloromethane (100 mL) was added dropwise with vigorous stirring over 30 min. The reaction mixture was stirred for 16 hours at room temperature. Water (600 mL) was added. The organic layer was separated. The aqueous layer was extracted with 2×200 mL of dichloromethane. The combined organic layer was washed with water (2×400 mL) and dried (Na 2
). The solution was filtered, concentrated under reduced pressure to give an oil 93.36 g (˜100% yield), which was used directly in the next step without further purification.
Protection of N-Boc-(D)-Serine Methyl Ester
Boc-Serine methyl ester from above (93.0 g, 0.42 mol) and catalyst p-toluenesulfonic acid (9.3 g) were dissolved in dichloromethane (500 mL) and 2,2-dimethoxypropane (500 mL). The mixture was stirred at room temperature for 20 hours with a drying tube. Saturated sodium bicarbonate (600.0 mL) was added. The mixture was then stirred vigorously for 30 min. The organic layer was separated, washed with bicarbonate (2×400.0 mL), water (400.0 mL), saturated NaCl (400.0 mL) and dried (Na 2
). The solution was filtered and concentrated under vacuum to give 87.22 g oil (84% yield for two steps), which was used directly in the next step without further purification.
To a cooled solution of the ester (87.0 g, 0.336 mol) in anhydrous toluene (690.0 mL, −78° C., acetone/dry ice bath), DIBAL in toluene (1.49 M in toluene, 392 mL, 585.0 mmol) was added dropwise under argon in such a way that the internal temperature did not rise above −70° C. After the addition, the reaction mixture was stirred for an additional 4 hours at −78° C. Methanol (128 mL) was added to the mixture to quench the reaction. The mixture was poured slowly into an aqueous solution of Rochelle salt (potassium sodium tartrate tetrahydrate; 1.2 M, 660 g/1949 mL water) with vigorous stirring. The mixture was stirred at room temperature until clear separation into two layers. The aqueous layer was extracted with diethyl ether (2×300.0 mL). The combined organic layer was washed with water (2×800 mL) and brine (800 mL), then dried with anhydrous Na 2
. The solvent was evaporated under vacuum to give aldehyde as a pale yellow oil (68.59 g, 89%), which was used without further purification.
Addition of 4-Pentylphenyl Acetylene to the Above Aldehyde
To a cooled (−20° C.) solution of 4-n-pentylphenylacetylene (51.68 g, 300 mmol) in dry THF (400 mL), n-BuLi solution (2.5 M in hexane, 120 mL, 300 mmol) was added dropwise under argon. After 2 hours, the mixture was cooled to −78° C., followed by the addition of HMPA (hexmethylphosphoramide, 64.5 g, 360 mmol). After the mixture was stirred at −78° C. for an additional 30 mins, methyl (R)-(+)-3-(t-butoxycarbonyl)-2,2-dimethyl-4-oxazolidinecarboxaldehyde (58.0 g, 248.3 mmol) in anhydrous THF (tetrahydrofuran; 100 mL) was added dropwise (maintaining the temperature below −60° C.). The mixture was stirred for an additional 5 hours at −78° C., then quenched by saturated ammonium chloride solution (1000 mL). The aqueous layer was extracted with ethyl ether (3×400 mL). The combined organic layer was washed with 0.5 N HCl (2×400 mL) and brine (400 mL), then dried with anhydrous sodium sulfate. The solvent was removed under vacuum to give a yellow oil (104.04 g, ˜100% yield), which was used without further purification.
Deprotection of the Above Oxazolidine
To an ice cooled solution of Boc-oxazolidine (103.0 g, 257.0 mmol) in methanol (1000 mL), was added conc. HCl (43.5 mL, pre-cooled to 0° C.). The mixture was stirred at room temperature overnight and then extracted with hexane (3×400 mL). The pH of the methanol solution was adjusted with solid sodium bicarbonate to 8.0. Boc anhydride (53.94 g, 245.92 mmol) was added and the mixture was stirred at room temperature for 1-4 hours until the disappearance of formed intermediate free amine. The solvent was removed under vacuum. The residue was redissolved in water (300 mL) and diethyl ether (300 mL). The ethyl ether layer was dried with anhydrous sodium sulfate and then evaporated to give a brown oil (87.54 g, 94%), which was used without further purification.
Reduction of the Above Alcohol
To an ice-cooled solution of the above acetylene (87.0 g, 241.0 mmol) in THF (800 mL), Red-Al (Sodium bis(2-methoxyethoxy)aluminum dihydride; 60% w/w in toluene, 392 mL; 1.205 mol) was added dropwise over 1 hour under argon with stirring. The solution was then stirred at room temperature for 36 hours. The reaction mixture was cooled in an ice bath and then poured carefully into a pre-cooled solution of Rochelle salt in water (700 g in 2200 mL of water). The mixture was vigorously stirred until two layers were visible and well separated. The aqueous layer was extracted with 2×600 mL of toluene. The combined toluene layer was washed with water (2×800 mL) and saturated sodium chloride (800 mL) and dried (Na 2
). The solvent was removed under vacuum to give a yellowish semi solid, which was recrystallized with hexane (200 mL) to give a white solid 43.3 g (purity: >98%; yield: 49%)
Deprotection to SK1-I (BML-258)
To a solution of Boc protected amine (15 g, 41.3 mmol) in anhydrous THF (300 mL), DIBAL (25% w/w in toluene, 1.49 M, 278 mL, 413 mmol) was added at room temperature under argon. The mixture was refluxed until the starting material disappeared. The mixture was cooled to room temperature and poured into Rochelle salt (340 g/1000 mL water) containing sodium hydroxide (50 g, ˜5%). The mixture was stirred vigorously for 1 hour. The aqueous layer was extracted with ethyl acetate (2×500 mL). The combined organic layer was washed with water (1000 mL) and brine (1000 mL) and dried with anhydrous sodium sulfate. The solvent was removed under vacuum to afford yellowish oil, which turned into a pale solid after storing at −20° C. overnight. To a cold solution (ice bath) of this solid in ethyl ether (400 mL), was added 1M HCl in ethyl ether (50 mL). The white precipitate was collected by filtration and washed with ethyl ether (2×50 mL), and then dried under vacuum to give product as a white solid (8.11 g, 63% yield).
claiming sphingosine pathway modulating compounds for the treatment of cancers, assigned to Enzo Biochem Inc , naming different team
Sphingosine- 1 -phosphate (SIP) was discovered to be a bioactive signaling molecule over 20 years ago. Studies have since identified two related kinases, sphingosine kinase 1 and 2 (a/k/a sphingosine kinase “type I” and “type II” respectively, and SphKl and SphK2 respectively), which catalyze the phosphorylation of sphingosine to SIP. Extracellular SIP can bind to and activate each of five S IP-specific, G protein-coupled receptors (designated S IPR1-5) to regulate cellular and physiological processes in an autocrine or paracrine manner. Selective inhibitors of each of sphingosine kinase 1 and 2, as well as both nonselective and selective agonists of SlPRs, have been developed and are known in the art.
Product Literature References
Sphingosine kinase 1 activation by estrogen receptor α36 contributes to tamoxifen resistance in breast cancer
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TP53 is required for BECN1- and ATG5-dependent cell death induced by sphingosine kinase 1 inhibition
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A novel E2F/Sphingosine kinase 1 axis regulates anthracycline response in squamous cell carcinoma
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Inhibition of Sphingosine kinase 1 in doxorubicin-treated SCC cells and in vivo., Abstract
Inhibition of Sphingosine Kinase 1 Ameliorates Angiotensin II-induced Hypertension and Inhibits Transmembrane Calcium Entry via Store-Operated Calcium Channel
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Inhibition of Sphingosine kinase 1, Abstract
K63-linked polyubiquitination of transcription factor IRF1 is essential for IL-1-induced production of chemokines CXCL10 and CCL5.
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Inhibition of Sphingosine kinase 1 in primary human astrocytes and mice, Abstract
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LRIG1 modulates aggressiveness of head and neck cancers by regulating EGFR-MAPK-SPHK1 signaling and extracellular matrix remodeling
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Role of sphingosine kinase 1 and sphingosine-1-phosphate in CD40 signaling and IgE class switching
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Inhibition of Sphingosine kinase 1 in human tonsil B cells, mouse splenic B cells and in mice, Abstract
Sphingosine kinase-1 enhances resistance to apoptosis through activation of PI3K/Akt/NF-κB pathway in human non–small cell lung cancer
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Targeting sphingosine kinase 1 inhibits Akt signaling, induces apoptosis, and suppresses growth of human glioblastoma cells and xenografts
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A selective sphingosine kinase 1 inhibitor integrates multiple molecular therapeutic targets in human leukemia
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General Literature References
Sphingosine-1-phosphate and cancer
: N.J. Pyne & S. Pyne; Nat. Rev. Cancer 10
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Antitumor Activity of Sphingosine Kinase Inhibitors
: K.J. French, et al.; J. Pharmacol. Exp. Ther. 318
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; Full Text
/////////SK1-I , SK1I , SK1 I , BML 258, Enzo Biochem, Virginia Commonwealth, Preclinical, solid tumours, liver cancer, haematological malignancies, autoimmune hepatitis,