Deuterated analogues of selenophenochromenes, synthesis thereof, and methods of using same agents

Abstract

The present invention relates to a novel cancer curing deuterated selenopheno [h] chromene derivatives, as well as methods of their manufacturing and use in different pharmaceutical compositions for the treatment of cancer by administration of such substances.

Claims

1. A compound of formula (I) ##STR00006## wherein R represents C.sub.1-C.sub.7 alkyl group containing deuterium.

2-4. (canceled)

5. The compound according to claim 1 wherein the compound is selected from the group consisting of: Methyl-d3 7-bromo-8-((4-methylpiperazin-1-yl)methyl)-2-oxo-2H-selenopheno[3,2-h]chromene-3-carboxylate; Ethyl-d5 7-bromo-8-((4-methylpiperazin-1-yl)methyl)-2-oxo-2H-selenopheno[3,2-h]chromene-3-carboxylate; and Butyl-d9 7-bromo-8-((4-methylpiperazin-1-yl)methyl)-2-oxo-2H-selenopheno[3,2-h]chromene-3-carboxylate.

6. The compound according to claim 5, wherein the compound has the structure: ##STR00007##

7. The compound according to claim 5, wherein the compound has the structure: ##STR00008##

8. A pharmaceutical composition comprising the compound according to claim 1, and a pharmaceutically acceptable carrier.

9. The pharmaceutical composition according to claim 8, wherein the composition is a mono-phasic pharmaceutical composition suitable for parenteral or oral administration consisting essentially of a therapeutically effective amount of the compound of formula I, and a pharmaceutically acceptable carrier.

10. The compound of claim 1, wherein R is C.sub.1-C.sub.4 alkyl group containing deuterium.

11. The compound of claim 5, wherein the compound has the structure: ##STR00009##

12. A method of treating cancer, comprising administering a therapeutically effective amount of the compound of claim 1 to a patient in need thereof.

13. The method of claim 12, wherein the cancer is breast, sarcoma, fibrosarcoma, or lung cancer.

14. The method of claim 12, wherein the compound is administered in conjunction with one or more chemotherapeutic agents, surgery, chemotherapy, radiation, immunotherapy, or combinations thereof.

15. The method of claim 12, wherein R is C.sub.1-C.sub.4 alkyl group containing deuterium.

16. The method of claim 15, wherein the compound is selected from the group consisting of: methyl-d3 7-bromo-8-((4-methylpiperazin-1-yl)methyl)-2-oxo-2H-selenopheno[3,2-h]chromene-3-carboxylate; ethyl-d5 7-bromo-8-((4-methylpiperazin-1-yl)methyl)-2-oxo-2H-selenopheno[3,2-h]chromene-3-carboxylate; and butyl-d9 7-bromo-8-((4-methylpiperazin-1-yl)methyl)-2-oxo-2H-selenopheno[3,2-h]chromene-3-carboxylate.

17. The method of claim 12, wherein the compound is ##STR00010##

18. The method of claim 12, wherein the compound is ##STR00011##

19. The method of claim 12, wherein the compound is ##STR00012##

Description

DETAILED DESCRIPTION OF THE INVENTION

[0010] Searching for anticancer compounds with improved stability and anticancer activity we unexpectedly discovered that deuterated 2H-selenopheno[3,2-h]chromene derivatives of Formula I exhibit higher stability and cytotoxic effect against cancer cells compared with non-deuterated analogue.

[0011] Pharmaceutical compositions in accordance with embodiments of the disclosure may be prepared by combining the disclosed compounds with a solid or liquid pharmaceutically acceptable carrier and, optionally, with pharmaceutically acceptable adjuvants and excipients employing standard and conventional techniques. Solid form compositions include powders, tablets, dispersible granules, capsules, cachets and suppositories. A solid carrier may be at least one substance that may also function as a diluent, flavoring agent, solubilizer, lubricant, suspending agent, binder, tablet disintegrating agent, and encapsulating agent. Inert solid carriers include magnesium carbonate, magnesium stearate, talc, sugar, lactose, pectin, dextrin, starch, gelatin, cellulosic materials, low melting wax, cocoa butter, and the like. Liquid form compositions include solutions, suspensions and emulsions. For example, there may be provided solutions of the compounds disclosed herein dissolved in water and water-propylene glycol systems, optionally containing suitable conventional coloring agents, flavoring agents, stabilizers, and/or thickening agents.

[0012] In an embodiment, a pharmaceutical composition may be provided employing conventional techniques in unit dosage form containing effective or appropriate amounts of one or more active component. In embodiments, the quantity of active component (compound) in a pharmaceutical composition and unit dosage form thereof may be varied or adjusted widely depending upon the particular application, the potency of the particular compound and the desired concentration. In an embodiment, the quantity of active component may range from 0.5% to 90% by weight of the composition.

[0013] In embodiments, in therapeutic use for treating, ameliorating, preventing, or combating cancer in animals, the compounds or pharmaceutical compositions thereof may be administered orally, parenterally, topically, and/or by inhalation at a dosage to obtain and maintain a concentration or blood-level of active component in the animal undergoing treatment that is therapeutically effective.

[0014] In an embodiment, such a therapeutically effective amount of dosage of active component may be in the range of about 0.1 to about 100 mg/kg, more preferably about 3.0 to about 50 mg/kg, of body weight/day. It is to be understood that the dosages may vary depending upon the requirements of the patient, the severity, type, stage, grade, or location of the cancer being treated, and the particular compound being used.

[0015] Also, it is to be understood that the initial dosage administered may be increased beyond the above upper level in order to rapidly achieve the desired blood-level or the initial dosage may be smaller than the optimum and the daily dosage may be progressively increased during the course of treatment depending on the particular situation. If desired, the daily dose also may be divided into multiple doses for administration, for instance, two to four times per day.

[0016] Scheme 1 describes the preparation of compounds of Formula I of the present invention. All of the final compounds of the present invention can be prepared by procedures described in these charts or by procedures analogous thereto, which procedures would be well known to one of ordinary skill in organic chemistry. All of the variables used in the scheme are as defined below or as in the claims.

General Procedure of Compounds Preparation of Formula 1 (Scheme 1)

[0017] ##STR00002##

Examples

[0018] Preparation of the disclosed compounds of the present invention is described in the following examples, which are intended as an illustration of and not a limitation upon the scope of the invention.

Hereinafter, “DMF” is defined as N,N-dimethylformamide, “DMSO” as dimethyl sulfoxide, “HCl” as hydrochloric acid, “HOBt” as hydroxybenzotriazole, “EDC HCl” as 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride “DCM” as dichloromethane, “rt” as room temperature.

General Method for Preparation of Deuterated selenopheno[3,2-h]chromenes (I)

[0019] A round bottom flask (100 mL), equipped with a magnetic stirrer and septum, was charged with 7-bromo-8-((4-methylpiperazin-1-yl)methyl)-2-oxo-2H-selenopheno[3,2-h]chromene-3-carboxylic acid dihydrochloride (1, 350 mg, 0.63 mmol), hydroxybenzotriazole (96 mg, 0.63 mmol) and dry DMF (15 mL). Subsequently, after the addition of N-methylmorpholine (134 μL, 1.25 mmol) and appropriated deuterated alcohol (2.51 mmol), 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (300 mg, 1.57 mmol) was added in one portion. Reaction mixture was stirred overnight, then DMF was evaporated under reduced pressure. Crude residue was purified by flash chromatography on silica gel using DCM/MeOH mixture as eluent. After being purified I-1 was dissolved in DCM and converted to its hydrochloride salt by adding 1M HCl solution in diethyl ether. Products I-2 and I-3 were converted to HCl salts before purification.

Example 1

[0020] Methyl-d3 7-bromo-8-((4-methylpiperazin-1-yl)methyl)-2-oxo-2H-selenopheno[3,2-h]chromene-3-carboxylate dihydrochloride (I-1)

##STR00003##

Yield, 69%. M.p. >200 ° C.

[0021] .sup.1H NMR (400 MHz, Chloroform-d) δ 8.66 (s, 1H), 7.69 (d, 1H), 7.56 (d, 1H), 3.84 (s, 2H), 2.87-2.39 (m, 8H), 2.34 (s, 3H). .sup.13C NMR (101 MHz, CDCl.sub.3) δ 163.9, 156.4, 153.7, 152.8, 150.0, 146.7, 126.1, 125.7, 121.1, 116.4, 113.2, 105.6, 58.8, 55.2, 53.7, 46.0. .sup.77Se NMR (76 MHz, CDCl.sub.3) δ 574.6. HRMS (ESI) calcd for C.sub.19H.sub.16D.sub.3BrN.sub.2O.sub.4Se [M+H].sup.+ 501.9954, found 501.9965.

Example 2

[0022] Ethyl-d5 7-bromo-8-((4-methylpiperazin-1-yl)methyl)-2-oxo-2H-selenopheno[3,2-h]chromene-3-carboxylate dihydrochloride (I-2)

##STR00004##

Yield, 92%. M.p. >200 ° C.

[0023] .sup.1H NMR (400 MHz, Chloroform-d) δ 8.57 (s, 1H), 7.61 (d, 1H), 7.50 (d, 1H), 3.83 (s, 2H), 2.97-2.46 (m, 8H), 2.42 (s, 3H). .sup.13C NMR (101 MHz, CDCl.sub.3) δ 163.0, 156.2, 152.9, 152.5, 149.3, 146.4, 126.0, 125.5, 120.9, 116.6, 113.2, 105.8, 77.5, 58.6, 54.9, 53.0, 45.5. .sup.77Se NMR (76 MHz, CDCl.sub.3) δ 572.8. HRMS (ESI) calcd for C.sub.20H.sub.16D.sub.5BrN.sub.2O.sub.4Se [M+H].sup.+ 518.0237, found 518.0250.

Example 3

[0024] Butyl-d9 7-bromo-8-((4-methylpiperazin-1-yl)methyl)-2-oxo-2H-selenopheno[3,2-h]chromene-3-carboxylate dihydrochloride (I-3)

##STR00005##

Yield, 80%. M.p. >200 ° C.

[0025] .sup.1H NMR (400 MHz, Chloroform-d) δ 8.60 (s, 1H), 7.69 (d, 1H), 7.57 (d, 1H), 3.84 (s, 2H), 2.87-2.38 (m, 8H), 2.33 (s, 3H). .sup.13C NMR (101 MHz, CDCl.sub.3) δ 163.2, 156.2, 153.5, 152.7, 149.3, 146.5, 125.9, 125.7, 120.9, 116.8, 113.1, 105.5, 58.8, 55.2, 53.7, 46.0. .sup.77Se NMR (76 MHz, CDCl.sub.3) δ 574.2. HRMS (ESI) calcd for C.sub.22H.sub.16D.sub.9BrN.sub.2O.sub.4Se [M+H].sup.+ 550.0801, found 550.0808.

Metabolic Stability In Vitro

[0026] Assay: the human hepatoma cells HepG2 (ATCC HB-8065) were grown in Eagle's Minimum Essential Medium supplemented with 10% fetal calf serum. The cells were harvested at 80% confluence. The hepatocytes were scraped, two time washed in Hank's Balanced Salt Solution (HBSS) buffer pH-7.4 and suspended in homogenization buffer composed of 10 mM Tris-HCl, 10 mM KCl, 0.15 mM MgCl.sub.2 pH 6.7. The cells were homogenized on ice by homogenizer LabGEN7 (Cole Palmer) at 5000 rpm 15 sec. Homogenized cells in HBSS with compound (protein concentration is 95 μg/ml) incubate 37° C. After 10, 30, 60, 120, 180, and 240 min collect 1 ml sample and add 0.1 ml 40% TCA, then centrifugation at 1200 g for 10 min. Supernatant collected and analyzed.

[0027] Results: The stability of deuterated selenopheno[h]chromenes I-1-I-3 compared with non-deuterated analogue methyl 7-bromo-8-((4-methylpiperazin-1-yl)methyl)-2-oxo-2H-selenopheno[3,2-h]chromene-3-carboxylate dihydrochloride (I-0) was tested in vitro using lysate of human hepatoma cells HepG2. The results are presented in FIG. 1 (x-axis—minutes of incubation, y-axis—percent of remained ester). Surprisingly, I-3 showed the highest stability to hydrolysis, 24% of deuterated butyl ester remained after 240 minutes of incubation. Notably, I-0 almost completely was hydrolyzed.

[0028] Anticancer activity of deuterated selenopheno[h]chromenes was tested in vitro using cytotoxicity assay. Thus, monolayer tumor cell lines MDA-MB-435s (human melanoma), MCF-7 (human breast adenocarcinoma, estrogen-positive), MES-SA (human uterus sarcoma), HT-1080 (human fibrosarcoma), A549 (human lung carcinoma), GM08402 (human fibroblast, apparently healthy), 3T3 (mouse embryo fibroblasts), H9C2 (rat cardiomyocytes) were cultured in standard medium DMEM (Dulbecco's modified Eagle's medium) (“Sigma”) supplemented with 10% heat-inactivated fetal bovine serum (“Sigma”). The cell viability was assessed by addition of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolinium bromide (MTT). Briefly, cells were seeded (2−6×10.sup.4 cells/ml) in 96-well plates and allowed to attach for 24 h.

[0029] Tested compound solutions were prepared and serially diluted to obtain appropriate concentrations. Cells were treated with different concentrations (0.032-100 μM) and incubated for 48 h at 37° C., 5% CO.sub.2 Then the culture medium was removed and medium containing 0.2 mg/ml MTT was added. After 3 h (37° C., 5% CO.sub.2), the MTT-containing medium was removed, and 200 μl of dimethyl sulfoxide (DMSO) was immediately added to each sample. The absorbance was assessed at 540 nm on a Tecan multiplate reader Infinite 1000 (Austria). The half-maximal inhibitory concentration (IC.sub.50) of each compound was calculated using Graph Pad Prism® 3.0.

[0030] The results of cell culture-based studies are summarized in Table 1. In general, tested compounds showed medium or low cytotoxicity against malignant tumor cells. However, the introduction of deuterated substituent led to significant increase in cytotoxic effect on triple negative breast carcinoma (MDA-MB-231), sarcoma (MES-SA), fibrosarcoma (HT-1080) and lung carcinoma cell lines (A549) IC.sub.50 increased up to 29 μM. This unexpected discovery together with increase in stability of compounds I-1-I-3 towards cancer cells makes these compounds very promising as anticancer medicines.

TABLE-US-00001 TABLE 1 In vitro cytotoxicity of deuterated selenopheno[h]chromenes on monolayer tumor cell lines. Cytotoxicity IC.sub.50, μM Nr. MDA-MB-231 MCF-7 MES-SA HT-1080 A549 GM08402 3T3 H9C2 I-0 95 ± 4  54 ± 6 56 ± 1 58 ± 5 >100 77 ± 16 100 ± 5  92 ± 12 I-1 84 ± 13 82 ± 5 40 ± 7 70 ± 6 105 ± 7 89 ± 16  66 ± 11 82 ± 14 I-2 90 ± 15 46 ± 4  56 ± 15 59 ± 6 >100 94 ± 8  64 ± 9 73 ± 4  I-3 35 ± 8  70 ± 3 29 ± 3 31 ± 3  40 ± 13 62 ± 3  34 ± 7 30 ± 4 

References

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