NL2010036C2

NL2010036C2 - Verapamil like compounds.

Info

Publication number: NL2010036C2
Application number: NL2010036A
Authority: NL
Inventors: Renske Raaphorst; Gert Luurtsema; Albert Dirk Windhorst; Adriaan Lammertsma
Original assignee: Stichting Tech Wetenschapp; Stichting Vu Vumc; Univ Groningen; Groningen Acad Ziekenhuis
Priority date: 2012-12-21
Filing date: 2012-12-21
Publication date: 2014-06-24
Also published as: WO2014098593A9; WO2014098593A1

Abstract

The invention is directed to a compound, a salt or a solvate thereof according to (I) wherein R1 is an optionally branched fluoroalkyl group or [18F]fluoroalkyl group, R2, R3, R4 and R5 are eacn independently a methyl, [3H]methyl, fluoroalkyl or [18F]fluoroalkyl group, and R6 is a (R)-cyano group or a (S)-cyano group; or wherein R1 is a methyl group, and R3 is a R3 is a fluoroalkyl group or a [18F]fluoroalkyl group, R2, R4 and R5 are each independently a methyl, [3H]methyl, fluoroalkyl or [18F]fluoroalkyl group, and R6 is a (R)-cyano group or a (S)-cyano group; or wherein if R1 is hydrogen, at least one of R2, R3, R4 is an optionally branched fluoroalkyl group or [18F]fluoroalkyl group.

Description

VERAPAMIL LIKE COMPOUNDS

The present invention relates to verapamil like compounds, their use as a P-glycoprotein inhibitor or substrate, a radiopharmaceutical formulation comprising specific 5 verapamil like compounds, compound, a radiopharmaceutical formulation comprising specific verapamil like compounds for use in vivo diagnostic or in vivo imaging method, a method for the in vivo diagnosis or in vivo imaging of a P-glycoprotein related disease in a subject and a method to prepare the novel verapamil like compounds.

Positron emission tomography (PET) is a nuclear medicine imaging technique that 10 produces images of functional processes of the body. Radiotracers are used in PET as diagnostic tools and to image tissue concentration of molecules of interest.

P-glycoprotein (P-gp) is an ABC efflux transporter mainly expressed in the blood brain barrier (Nature 1984; 309:626-628) (Science 1983, 221:1285-1288). It transports xenobiotic and structurally diverse compounds out of the cell. Overexpression of P-gp leads 15 to decreased uptake of pharmaceuticals aimed at the brain. Furthermore, this protein is involved in diseases like epilepsy, Alzheimer's disease and Parkinson's disease. Imaging this particular protein would give insight into its role in healthy and diseased state.

A known tracer for P-gp is [Hcjverapamil (J Nucl Med 1996; 37; 1571-1575). It was utilized as a calcium channel blocker drug, but was discovered to be a P-gp substrate as well. 20 It is labeled with carbon-11 and is employed in the clinic as a diagnostic tool (Eur J Clin Pharmacol 2001; 56:827-829).

A disadvantage of the P-gp [Hcjverapamil compound is that it has a very short half-life of only 20 minutes. This short half-life limits the application of this compound.

The object of this invention is to provide an alternative for verapamil. This is 25 achieved by the following compound. Compound or a salt or solvate thereof according to 2 CC^'SX; wherein R^ is hydrogen, an optionally branched fluoroalkyl group, an optionally branched [18F]fluoroalkyl group or a methyl group, 5 R2, R3, R4 and R5 are each independently a methyl, [8H]methyl, fluoroalkyl or [l8F]fluoroalkyl group, and

Rg is a (R)-cyano group or a (S)-cyano group.

The compound according to this invention can be referred to as a verapamil like compound. The fluoroalkyl analogs of verapamil according to the above may be provided in 10 an unlabeled and radiolabeled state. The radiolabeled compounds may advantageously be used as a radiopharmaceutical for use as a tracer for P-gp with a longer half-life than the existing tracers. The fluorine-18 labeled P-gp tracer is preferred, since the half-life of fluorine-18 is 110 minutes compared to 20 min for carbon-11. This is more practical and allows transport of the radiolabeled compound to hospitals which do not have their own 15 cyclotron capacities. The tritium labeled compounds according to the present invention may advantageously be used for in vitro studies to investigate the interaction with P-gp at low concentration levels, better mimicking the in vivo situation than with unlabeled compounds. A first group of preferred verapamil like compounds are compounds wherein R^ is a fluoroalkyl group or more preferably a [l8F]fluoroalkyl group. Preferably the alkyl group has 20 1 to 3 carbon atoms of which methyl or ethyl are especially suited. Preferably R2, R3, R4 and R5 are a methyl group. If R^ is a fluoroalkyl group R2, R4 and R5 may be all a methyl group and R3 is a [8H]methyl group.

The non radiolabeled compounds according to this first group may advantageously be used as reference compounds. Examples of such preferred compounds are illustrated in 25 the attached Figures as compounds 1: (S)- and (/?)-Fluoroethylverapamil, wherein R^ is a fluoroethyl group and R2, R3, R4 and R5 are each a methyl group; compounds 2: (5)- and (/?)-Fluoromethylverapamil, wherein R^ is a fluoromethyl group and R2, R3, R4 and R5 are each a methyl group.

3

Examples of suitable radiolabeled compounds according to this First Group are compound 5: (S)- and (/?)-[18F]fluoroethylverapamil, wherein is a [18F]fluoroethyl group and R2, R3, R4 and R5 are each a methyl group; compound 6, (S)- and (/?)-[l8F]fluoromethylverapamil, wherein R^ is a [l8F]fluoromethyl group and R2, R3, R4 and R5 5 are each a methyl group.

The radiolabeled compounds are preferably prepared starting from a suitable precursor. These precursors are actually the more important compounds as they are the compounds which are stored and used to prepare the radiolabeled compounds hours before their actual use. These compounds should have a molecular structure which enables one to 10 easily, preferably by means of one synthesis step, prepare the desired radiolabeled compound. Applicants found that compounds 9 and 10, wherein R^ is a fluoromethyl or fluoroethyl group and R2, R4 and R5 are a methyl group and R3 is hydrogen are suited precursors for preparing compounds 7 and 8 respectively by means of a methylation. Preferred precursor compounds are compounds wherein R^ is hydrogen and 15 wherein group R^ is substituted by the desired fluoroalkyl group by means of an alkylation. Compounds 11, wherein R^ is hydrogen, R2, R4 and R5 are a methyl group and R3 is hydrogen are suited intermediate compounds to prepare compounds 9 and 10, which compounds 9 and 10 are in turn suitable precursors to prepare compounds 7 and 8 respectively.

20 For a second group of compounds R^ is a methyl group. Preferably R2, R4 and R5 are a methyl groups and R3 is a fluoroalkyl group or a more preferably a [l8F]fluoroalkyl group. Suitably the alkyl group has 1 to 3 carbon atoms, suitably methyl or ethyl. Examples are compound 3, (S)- and (/?)-0-fluoroethylverapamil, wherein R^, R2, R4 and R5 are each a methyl group and R3 is a fluoroethyl group; compound 4, (5)- and [R)-0-25 fluoromethylverapamil, wherein R^, R2, R4 and R5 are each a methyl group and R3 is a fluoromethyl group. Examples of radiolabeled compounds are compound 12, (S)- and [R)-0-[18F]fluoroethylverapamil, wherein R^, R2, R4 and R5 are each a methyl group and R3 is a - [18F]fluoroethyl group and compound 13, (5)- and (/?)-0-[18F]fluoromethylverapamil, wherein R^, R2, R4 and R5 are each a methyl group and R3 is a -[l8F]fluoromethyl group.

4

The above radiolabeled compounds are preferably prepared from a precursor compound wherein R3 is hydrogen. This hydroxyl group may be substituted by the desired fluoroalkyl group by means of alkylation. An example of a suitable precursor compound is compound 14, (S)- and (/?)-desmethylverapamil, wherein R^, R2, R4 and R5 are each a 5 methyl group and R3 is hydrogen.

The radio labelled compounds and non-radio labelled compounds according to the present invention may be purified according to those methods known to the person skilled in the art, for example by means of HPLC purification or Solid Phase Extraction (SPE). The HPLC purification is preferable carried out on a preparative HPLC column packed with 10 reverse phase material such as, but not limited to, C18, C18-EPS or C8, a mobile phase consisting of a mixture of methanol, ethanol or acetonitrile mixed with water or water containing buffer like, but not limited to, ammonium dihydrogen phosphate or an acid like phosphoric acid or trifluoracetic acid. The Solid Phase Extraction (SPE) is preferably performed on a Seppak® like, but not limited to, C18, tC18, Silica or an Oasis Seppak®. The 15 compound is preferably eluted from the Seppak® with a solvent suitable for injection in vivo, like ethanol.

The above treated compounds may be formulated to a desired formulation for their intended use. For example the collected HPLC fraction from the preparative HPLC, containing a compound according to the invention may be diluted with water or water 20 containing such as, but not limited to, sodium hydroxide or hydrogen chloride. The diluted fraction as prepared is trapped on a Seppak® like, but not limited to, C18, tC18, Silica or an Oasis Seppak® and the compound is preferably eluted from the Seppak® with a solvent suitable for injection in vivo, like ethanol. The obtained eluate is preferable diluted with pharmaceutically acceptable buffers such as, but not limited to 0.9% sodium chloride, 25 sodiumdihydrogenphosphate 7.09 mM in 0.9 % sodiumchloride or citrate buffer, pharmaceutically acceptable solubilisers such as, but not limited to, ethanol, tween or phospholipids and/or with pharmaceutically acceptable stabilizers or antioxidants such as, but not limited to, ascorbic acid, gentisic acid or p-aminobenzoic acid.

The invention is also directed to the salts and solvates of the compounds described 30 above. Suitable salts according to the invention, include physiologically acceptable acid addition salts such as those derived from mineral acids, but not limited to, hydrochloric, 5 hydrobromic, phosphoric, metaphosphoric, nitric or sulphuric acids or those derived from organic acids such as, but not limited to, tartaric, fumaric, malonic, citric, benzoic, trifluoroacetic, lactic, glycolic, gluconic, methanesulphonic or p-toluenesulphonic acids.

Applicants found that the compounds according to the invention may be 5 advantageously be used as a P-glycoprotein inhibitor or substrate. More specifically the compounds may be used as part of a pharmaceutical formulation for use in diagnostics of/studying epilepsy, Alzheimer's disease and Parkinson's disease.

The invention is especially directed to a radiopharmaceutical formulation comprising the above described radiolabeled compounds. More especially the invention is 10 directed to a radiopharmaceutical formulation comprising the above described radiolabeled [18p] compounds for use in vivo diagnostic or in vivo imaging method. The diagnostic or imaging method of the P-glycoprotein will provide insight into diseases where this protein plays a role, like for example epilepsy, Alzheimer's disease and Parkinson's disease. The imaging method may be positron emission tomography (PET). The invention is thus also 15 directed to a method for the in vivo diagnosis or in vivo imaging of a P-glycoprotein related disease in a subject, preferably a human, comprising administration of a radiolabeled [18F]compound according to the invention or a formulation comprising such a a radiolabeled [l^Fjcompound.

The invention shall be illustrated by means of the following non-limiting examples. 20 Example 1

CN CN

In a solution of 5-((3,4-dimethoxyphenethyl)amino)-2-(3,4-dimethoxyphenyl)-2-isopropylpentanenitrile (100 mg, 0.227 mmol) (J Med Chem 1999, 42,1687-1697) in 2 ml DMF, l-bromo-2-fluoroethane (0.068 ml, 0.908 mmol) and potassium carbonate (125 mg, 25 0.908 mmol) were added and stirred at 65°C overnight. Consumption of the starting amine was observed on TLC (MeOH:DCM 5:95, v/v) and the reaction mixture was diluted with 20 ml ethyl acetate and the mixture was washed with water (3 x 20 ml), brine (20 ml) and dried over sodium sulfate. The volatiles were removed by rotary evaporation and the remainder was purified by flash column chromatography (MeOH:DCM = 1:99, v/v) to afford 5-((3,4- 6 dimethoxyphenethyl)(2-fluoroethyl)amino)-2-(3,4-dimethoxyphenyl)-2-isopropylpentanenitrile. 1H-NMR (250 MHz, CDCI3) δ: 6.76 (m, 6H), 4.53 (m, 1H), 4.34 (m, 1H), 3.86 (m, 12H), 2.77 (m, 1H), 2.66 (m, 4H), 2.52 (t, 2H,7= 6.8), 2.06 (m, 2H), 1.82 (m, 1H), 1.49 (m, 1H), 1.17 (d, 3H,V= 6.63), 0.78 (d, 3H,J= 6.63) 13C-NMR (62.9 MHz, CDCI3) δ: 5 148.908,148.718,148.171,147.208,132.849,130.593,121.468,120.466,118.596,111.967, 111.084,110.950,109.478, 83.983, 81.313, 77.507, 77.000, 76.490, 56.386, 37.914, 35.414, 33.015, 23.400, 18.910, 18.558

Example 2 10 ^*Υ)Η

To a solution of 5-bromo-2-(3,4-dimethoxyphenyl)-2-isopropylpentanenitrile (60 mg, 0.176 mmol) in 1 ml DMF 4-(2-aminoethyl)-2-methoxyphenol (44.2 mg, 0.265 mmol) and potassium carbonate (36.6 mg, 0.265 mmol) were added. The reaction was stirred overnight at room temperature. The mixture was diluted with diethyl ether and washed with water, 15 brine and dried over Na2S04. The volatiles were removed by rotary evaporation and the crude remainder was purified by flash column chromatography (MeOH:DCM= 5:95, v/v) to afford 2-(3,4-dimethoxyphenyl)-5-((4-hydroxy-3-methoxyphenethyl)amino)-2-isopropylpentanenitrile. ^-NMR (250 MHz, CDCI3) δ: 6.83 (m, 4H), 6.65 (m, 2H), 3.86 (m, 9H), 2.80 (b, 4H), 2.66 (t, 2H, J= 6.3), 2.08 (m, 3H), 1.89 (dt, 1H, J= 4.5 12.4), 1.59 (m, 1H), 20 1.17 (d, 3H, J= 6.63), 0.78 (d, 3H, J= 6.63) 13C-NMR (125.78 MHz, CDCI3) δ: 148.955,148.226, 146.505, 144.065, 131.235, 130.408, 121.321, 121.132, 118.608, 114.384, 111.195,111.018, 109.457, 55.964, 55.849, 55.811, 50.772, 49.080, 37.852, 35.507, 25.570, 18.928, 18.554

Example 3

F

Y H Y / 25 YY^ YY^

To a solution of 2-(3,4-dimethoxyphenyl)-5-((4-hydroxy-3-methoxyphenethyl)amino)-2-isopropylpentanenitrile (16 mg, 0.038 mmol) in 1 ml acetonitrile l-bromo-2-fluoroethane (8.38 pi, 0.113 mmol) and DIPEA (9.83 μΙ, 0.056 mmol) were added. The reaction was 7 brought to 70°C and was stirred overnight. TLC analysis (MeOH:DCM=5:95) still showed remaining starting material and extra l-bromo-2-fluoroethane (10 μΙ, 0.134 mmol) was added and the reaction was stirred at 70°C for additional 6 hours. The volume was reduced by rotary evaporation. The crude product was dissolved in ethyl acetate, washed with sat.

5 NaHC03 and dried over Na2S04. The volatiles were removed by rotary evaporation and the remaining crude product was purified by gradient flash column chromatography (MeOH:DCM=2:98 to 5:95, v/v) to afford 2-(3,4-dimethoxyphenyl)-5-((2-fluoroethyl)(4-hydroxy-3-methoxyphenethyl)amino)-2-isopropylpentanenitrile. 1H-NMR (250 MHz, CDCI3) δ: 6.83 (m, 4H), 6.63 (m, 2H), 5.47 (b, 1H), 4.53 (m, 1H), 4.34 (m, 1H), 3.87 (m, 9H), 2.77 (q, 10 1H, J= 4.8), 2.63 (m, 4H), 2.51 (t, 2H, J= 6.8), 2.06 (m, 2H), 1.81 (dt, 1H, J= 4.5 12.4), 1.50 (m, 2H), 1.17 (d, 3H, J= 6.63), 0.78 (d, 3H, J= 6.63)

Example 4

CN CN

15 [18F]2-bromofluoroethane (Bioorganic & Medicinal Chemistry 2009,17, 7441-7448) was added to a suspension of 5-((3,4-dimethoxyphenethyl)amino)-2-(3,4-dimethoxyphenyl)-2-isopropylpentanenitrile (5 mg) and potassium carbonate (5 mg) in 1 ml DMF. The obtained mixture was brought to 100 °C for 30 minutes and cooled to 20 °C. The reaction was quenched with 1 ml acetonitrile and purified by HPLC (Kromasil 100-10-C18 250*22 mm 20 column) concentrated on a tC-18 Seppak® and formulated in a phosphate buffer/saline mixture. Yield 10-30% (corrected for decay), purity > 95%, specific activity > 20 GBq/pmol

Claims

A compound or a salt or a solvate thereof, in accordance with 5 Y wherein R 1 is hydrogen, an optionally branched fluoroalkyl group, an optionally branched [18 F] fluoroalkyl group or a methyl group, R 2, R 3, R 4 and R 5 are each independently methyl, a [3 H] methyl, a fluoroalkyl, or a [18 F] fluoroalkyl group, and R e is a (f?) cyano group or a fS ') cyano group.

A compound according to claim 1, wherein R 1 is a fluoroalkyl group or an [18 F] fluoroalkyl group, wherein the alkyl group comprises 1 to 3 carbon atoms. 15

A compound according to claim 2, wherein the alkyl group is methyl or ethyl.

A compound according to claim 2 or claim 3, wherein R 1 is a [18 F] fluoroalkyl group.

A compound according to claim 2 or claim 3, wherein R 2, R 3, R 4 and R 5 are a methyl group, or wherein R 2, R 4 and R 5 are a methyl group and R 3 is a hydrogen.

A compound according to claim 2 or claim 3, wherein R 2, R 4 and R 5 are a methyl group Λ and R 3 is a [H] methyl group. 25

The compound of claim 1, wherein R 1 is hydrogen or a methyl group.

A compound according to claim 7, wherein R 1 is a methyl group, R 2, R 4 and R 5 are methyl groups, and R 3 is a fluoroalkyl group or an [18 F] fluoroalkyl group, wherein the alkyl group comprises 1 to 3 carbon atoms.

The compound of claim 8, wherein the alkyl group is methyl or ethyl.

The compound of claim 8 or claim 9, wherein R 3 is a [18 F] fluoroalkyl group.

11. A compound according to any one of claims 1-8, for use as a P-glycoprotein inhibitor or substrate.

A radiopharmaceutical formulation comprising the compound of claim 4, 6, or 10.

A radiopharmaceutical formulation comprising the compound of claim 4 or 10 for use in in vivo diagnostic methods or in in vivo imaging methods. Method for the in vivo diagnosis or for the in vivo imaging of a P-2 glycoprotein-related disorder in a subject, preferably a human, comprising administering a compound according to claim 4 or claim 10, then as many as 4 of a formulation according to claim 13.