WO2021038306A1 - Selenophenochromene phosphonic acids, preparation and use as antiproliferative agents - Google Patents

Abstract

The present invention relates to a novel selenophenochromene phosphonic acids as antiproliferative agents, as well as methods of their manufacturing and use in different pharmaceutical compositions for the treatment or prevention of various diseases and disorders by administration of such substances.

Description

Selenophenochromene phosphonic acids, preparation and use as antiproliferative agents FIELD OF THE INVENTION [1] The present invention relates to a novel to anti-cancer compounds, synthesis thereof, and methods of using same. The present invention discloses novel 2H-selenopheno[3,2- h]chromene phosphonic acid derivatives, a process of the manufacture and the use of the disclosed compounds for treatment of cancer. BACKGROUND OF THE INVENTION [2] Cancer is the second leading cause of death globally, and is responsible for an estimated 9.6 million deaths in 2018 (World Health Organization data). Globally, about 1 in 6 deaths is due to cancer. Tumour growth is a complex multistage process. Occurrence and progressive tumour growth is dependent on both the properties of cancer cells, and on the state of immunological reactivity. This determines the diversity of approaches of cancer therapy using one or several basic methods: surgery, radiotherapy, chemotherapy and immunotherapy. [3] The 2H-chromene system is a structural feature of many natural compounds and pharmacological agents possessing, among other properties, anti-cancer [1], anti-HIV [2], antibacterial activity [3]. [4] Many coumarin derivatives as medicines with substantial activity in vitro and in vivo were discovered (Psoralen, Angelicin, Xanthotoxin, Bergapten, Nodakenetin, etc). Imperatorin shows the ability to inhibit tumour growth. [4,5] Osthole inhibits the migration and invasion of breast cancer cells and effectively blocks matrix metalloproteinases promoter and enzyme activity. [6,7] It was found that phosphonic derivatives of coumarins exhibit very high alkylating activity and good cytotoxicity on NALM-6 (human leukemia). [8] [5] Recently we claimed 2H-selenopheno[3,2-h]chromenes as antimetastatic agents in treatment of carcinomas and melanoma. These derivatives exhibit low acute toxicity and almost completely (98 %) prevent mammary carcinoma 4T1 metastasis formation in vivo (BALB/c female mice) and melanoma B16-F10 metastasis in lung by 82 % without any visual side effects. [9] [6] Unfortunately, the number of cancer diagnosis in past few years has increased almost twice. In clinical practice, the treatment of cancer involves a wide range of chemotherapy drugs. Besides, most of them exhibit various side effects, high toxicity and moderate selectivity. Therefore, a new generation of selective, low toxic anticancer agent is one of the main tasks in medicinal chemistry and pharmaceutical industries. THE PRESENT INVENTION [7] We have surprisingly determined that certain selenophenochromene phosphonic acids exhibit ability to suppress cancer cell growth and therefore may act as anticancer agents. These substances may be administered in the form of a pharmaceutical composition, wherein they are present together with one or more pharmaceutically acceptable diluents, carriers, or excipients. OBJECTS OF THE INVENTION [8] It is an object of the present invention to provide novel pharmaceutical compounds with anti-cancer properties, useful for treatment of primary cancers, methods for manufacturing of disclosed compounds and the treatment of various cancers by administration of such substances. SUMMARY OF THE INVENTION [9] We disclosed compounds selected from those of Formula I

R¹ represents H or alkyl; R² represents a halogen atom; R³ represents C_1-4-alkyl-N-heterocyclyl; [10] As used herein, the term “alkyl” refers to a straight or branched hydrocarbon chain, containing the indicated number of carbon atoms. For example, C₁-C₆ alkyl indicates that the alkyl group may have from 1 to 6 (inclusive) carbon atoms. [11] The term “heterocyclyl” as used herein refers to a nonaromatic, saturated or partially unsaturated 3-10 membered monocyclic, 8-12 membered bicyclic, or 11-14 membered tricyclic ring system having 1-3 heteroatoms if monocyclic, 1-6 heteroatoms if bicyclic, or 1-9 heteroatoms if tricyclic, said heteroatoms selected from O, N, S, Si and P (e.g., carbon atoms and 1-3, 1-6, or 1-9 heteroatoms of O, N, S, Si and P if monocyclic, bicyclic, or tricyclic, respectively). [12] Any ring atom can be substituted (e.g., with one or more substituents). Heterocyclyl groups can contain fused rings, which are rings that share one or more common atoms. Examples of heterocyclyl groups include, but are not limited to, radicals of tetrahydrofuran, tetrahydrothiophene, tetrahydropyran, piperidine, piperazine, morpholine, pyrroline, pyrimidine, pyrrolidine, indoline, tetrahydropyridine, dihydropyran, thianthrene, pyran, benzopyran, xanthene, phenoxathiin, phenothiazine, furazan, lactones, lactams such as azetidinones and pyrrolidinones, sultams, sultones, and the like. [13] As used herein, the term "halogen" refers to fluorine, chlorine, bromine and iodine. [14] Embodiments of the present disclosure encompass any racemic, optically-active, polymorphic, tautomeric, or stereoisomeric form or mixture thereof, of a compound of the disclosure, which possesses the useful properties described herein. [15] In cases where compounds are sufficiently basic or acidic to form stable nontoxic acid or base salts, use of the compounds as pharmaceutically acceptable salts may be appropriate. Examples of pharmaceutically acceptable salts within the scope of embodiments of the present disclosure include organic acid addition salts formed with acids which form a physiological acceptable anion and inorganic salts. [16] Specific compounds of Formula I within the present invention include but are not limited to: Diethyl (7-bromo-8-((dimethylamino)methyl)-2-oxo-2H-selenopheno[3,2-h]chromen-3- yl)phosphonate; Diethyl (7-bromo-2-oxo-8-(piperidin-1-ylmethyl)-2H-selenopheno[3,2-h]chromen-3- yl)phosphonate hydrochloride; Diethyl (7-bromo-8-(morpholinomethyl)-2-oxo-2H-selenopheno[3,2-h]chromen-3- yl)phosphonate hydrochloride; Diethyl (7-bromo-8-((4-methylpiperazin-1-yl)methyl)-2-oxo-2H-selenopheno[3,2-h]chromen- 3-yl)phosphonate; Diethyl (7-bromo-2-oxo-8-(piperazin-1-ylmethyl)-2H-selenopheno[3,2-h]chromen-3- yl)phosphonate hydrochloride; Diethyl (8-((1,4-diazepan-1-yl)methyl)-7-bromo-2-oxo-2H-selenopheno[3,2-h]chromen-3- yl)phosphonate hydrochloride; (7-Bromo-8-((dimethylamino)methyl)-2-oxo-2H-selenopheno[3,2-h]chromen-3- yl)phosphonic acid hydrochloride; (7-Bromo-2-oxo-8-(piperidin-1-ylmethyl)-2H-selenopheno[3,2-h]chromen-3-yl)phosphonic acid hydrochloride; (7-Bromo-8-(morpholinomethyl)-2-oxo-2H-selenopheno[3,2-h]chromen-3-yl)phosphonic acid hydrochloride; (7-Bromo-8-((4-methylpiperazin-1-yl)methyl)-2-oxo-2H-selenopheno[3,2-h]chromen-3- yl)phosphonic acid hydrochloride. DETAILED DESCRIPTION OF THE INVENTION [17] Searching for compounds with ability to suppress cancer growth we unexpectedly discovered that 2H-selenopheno[3,2-h]chromene phosphonic acid derivatives of Formula I exhibit antiproliferative activity on various tumor cell lines. It was surprisingly also that these compounds were highly selective against cancer cells without significant cytotoxicity against normal 3T3 cells (mouse embryo fibroblasts), widely used for in vitro estimation of LD₅₀. [18] Our finding is unexpected, because it is well known, that high toxicity and low selectivity against cancer cells versus normal cells are typical for selenium containing molecules. [19] We discovered that cytotoxic activity is typical for a number of compounds for 2H- selenopheno[3,2-h]chromene phosphonic acids and their esters, especially if there are substituents present in positions 7 and 8 of this scaffold. We found that the most appropriate substituents for R₂ in position 7 of compounds according to Formula 1 are halogens, as well as for R₃ in the position 8 most preferable were C_1-4alkyl-N-heterocyclyl-groups. [20] Phosphonic acid or ester moiety will be preferred in position 3 of compounds represented by Formula I. [21] We discovered also, that substituents R₁ could be preferably selected from group of substituents consisting of H, C₁-C₆ hydrocarbon group. [22] As used herein, the term “hydrocarbon” refers to a cyclic, branched, or straight chain alkyl group, an alkenyl group, or an alkynyl group. Hydrocarbon-groups can either be unsubstituted or substituted with one or more substituents. If the hydrocarbon is a branched structure having substituent(s) thereon, then the substitution may be on either the hydrocarbon backbone or on the branch; alternatively, the substitutions may be on the hydrocarbon backbone and on the branch. [23] In cases where compounds are sufficiently basic or acidic to form stable nontoxic acid or base salts, use of the compounds as pharmaceutically acceptable salts may be appropriate. Examples of pharmaceutically acceptable salts within the scope of embodiments of the present disclosure include organic acid addition salts formed with acids which form a physiological acceptable anion and inorganic salts. [24] The term “pharmaceutically acceptable” refers here to the therapeutically active non- toxic salt forms, which the compounds of Formula I are able to form. The latter can conveniently be obtained by treating the base form with such appropriate acids as inorganic acids such as hydrochloric acid, hydrobromic acid; sulfuric acid; nitric acid; phosphoric acid and the like; or organic acids such as acetic, propanoic, hydroxyacetic, 2-hydroxypropanoic, oxopropanoic, oxalic, malonic, succinic, maleic, fumaric, malic, tartaric, methanesulfonic, benzenesulfonic, 4-methylbenzenesulfonic, 2-hydroxybenzoic, and like acids. Conversely, the salt may be converted to the free base by treatment with alkali. [25] For therapeutic use, the compounds of Formula I can be in the form of a solvate. [26] 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, 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. [27] 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. [28] 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. [29] 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. [30] 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. 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. [31] 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. [32] General procedure of compounds preparation of Formula I (Scheme 1) Diethyl (7-hydroxy-2-oxo-2H-chromen-3-yl)phosphonate (2) was obtained by reflux of 7- hydroxycoumarin (1) and diethyl phosphite in the presence of Mg(NO₃)₂ ^. 6 H₂O and AgNO₃ under argon atmosphere in dry MeCN. Desired 2 isolated in 40% yield as a pale yellow amorphous solid. 3-(Diethoxyphosphoryl)-2-oxo-2H-chromen-7-yl trifluoromethanesulfonate (3) prepared by treating of 2 with Tf₂O in dry DCM. Derivative 3 was obtained in almost quantitative yield as a pale brown amorphous solid. For triple bond introduction in position 7 Sonogashira protocol have been utilized. 7-substituted-2-oxo-2H-chromene-3-phosphonic acid esters (II) prepared by reaction of 3 with terminal acetylenes in the presence of a catalytic amount of tetrakis(triphenylphosphine)palladium(0) and copper iodide in dry DMF / triethylamine at 20 ^oC or slightly elevated temperature (up to 40 ^oC) under Ar atmosphere. Desired products II were isolated by flash chromatography on silica gel. [33] Surprisingly, we have found that the treatment of ethynyl chromenes II with in situ prepared selenium(IV) bromide led to the formation of selenopheno[3,2-h]chromene phosphonic acid esters (I-1–I-10). Reaction performed by dissolving of selenium dioxide in concentrated hydrobromic acid followed by addition of ethynyl chromene II in dioxane; the mixture was stirred at rt for 20 hours. Then the reaction mixture was diluted with EtOAc and sat. aq. NaHCO₃ and stirred for 30 min. The crude product was purified by silica gel column chromatography using a mixture of DCM/MeOH as an eluent to give a free base form of the corresponding cyclization product. The corresponding hydrochlorides I-1 – I-4 were obtained by addition of 1 M HCl solution in Et₂O to a cooled (ice/water bath) solution. Removal of volatile matters under reduced pressure and drying under vacuum afforded products as greenish-yellow amorphous solids in 60-70 % yield. [34] Derivatives I-5 and I-6 were isolated by dilution of the reaction mixture with ethanol till homogeneous greenish-yellow solution was obtained. The obtained solution was passed through an ion-exchange column containing Amberlite IRA-400 resin (counter ion Cl-). The crude material was pre-purified by reversed phase flash chromatography and pure products were obtained by direct phase flash chromatography using a mixture of DCM/EtOH (slightly acidified by 1M HCl in dry Et₂O). After evaporation of the solvents, pure I-5 and I-6 were obtained. Phosphonic acids I-7 – I-10 were obtained from the corresponding esters adding of concentrated HCl and refluxing for 5h. After evaporation of the solvent, the solids were dried under vacuum to give the corresponding products as a pale brown amorphous solid in quantitative yield. [35] Scheme 1. General procedure towards compounds of Formula I.

[36] Reaction conditions: A–HP(O)(OEt)₂ (2.0 equiv.), Mg(NO₃)₂ . 6H₂O (0.5 equiv.), AgNO₃ (5.0 mol-%), MeCN, 100 °C, Ar, 6h; B – Tf₂O (1.5 equiv.), Et₃N (2.0 equiv.), DCM, 0 °C, 2h; C – the corresponding propargylamine derivative (1.4 equiv.), (PPh₃)₄Pd (6.0 mol-%), Pd(OAc)₂ (8.0 mol-%), CuI (8.0 mol-%), Et₃N (3.0 equiv.), DMF, 40 °C, Ar, 1h; D – SeO₂ (3.0 equiv.), 48 % HBr/dioxane, r.t., 24 h; E – conc. HCl, reflux, 5 h. Examples [37] 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. [38] Hereinafter, ”DMF” is defined as N,N-dimethylformamide, ”DMSO” as dimethyl sulfoxide, ”HCl” as hydrochloric acid, ”MeCN” as acetonitrile, ”DIEA” as diisopropylethylamine, ”EtOAc” as ethyl acetate, ”rt” as room temperature. Intermediate 2 [39] Diethyl (7-hydroxy-2-oxo-2H-chromen-3-yl)phosphonate (2). To a mixture of 7- hydroxycoumarin (1) (5g, 30.8 mmol), diethyl phosphite (8.51 g, 61.6 mmol), Mg(NO₃)₂ ^. 6 H₂O (3.95 g, 15.4 mmol), and AgNO₃ (262 mg, 1.54 mmol) under argon atmosphere was added dry MeCN (60 ml) and the resulting solution was stirred at 100 °C for 6 h. After cooling to r.t., the reaction mixture was diluted with EtOAc (500 ml) and filtered. The volatile matters were removed under reduced pressure, and the crude product was purified by silica gel column chromatography using mixture of PE/EtOAc (5:1 ® 0:1) as an eluent to give 2 (3.7 g) in 40 % yield as a pale yellow amorphous solid. [40] ¹H NMR (DMSO-d₆, 400 MHz): d = 11.04 (br s, 1H; OH), 8.51 (d, 1H; 4-CH), 7.77 (d, 1H, 1H; 5-CH), 6.84 (dd, 1H; 6-CH), 6.74 (d, 1H; 8-CH), 4.15-4.01 (m, 4H; 2 × CH₂), 1.25 ppm (t, 6H; 2 × CH₃). Intermediate 3 [41] 3-(Diethoxyphosphoryl)-2-oxo-2H-chromen-7-yl trifluoromethanesulfonate (3). To a cooled (0 °C) solution of 2 (200 mg, 0.671 mmol) and Et₃N (0.19 ml, 1.34 mmol) in dry DCM (20 ml) dropwise was added solution of Tf₂O (285 mg, 1.01 mmol) in dry DCM (2.00 ml) and the reaction mixture was stirred at the same temperature for 2 h. Then, ice (40 g) was added, the cooling bath was removed, and the resulting mixture was stirred at r.t. till the ice melted. Additional amount of DCM (50 ml) was added and the stirring was continued for 15 min. The organic phase was separated, and the aqueous phase was extracted with DCM (2 × 50 ml). Combined organic phases were washed with brine (40 ml), dried over anhydrous Na₂SO₄ and the solvents were removed by evaporation under reduced pressure. The crude product was purified by silica gel column chromatography using mixture of PE/EtOAc (1:0 ® 1:3) as an eluent to give 3 (270 mg) in 93 % yield as a pale brown amorphous solid. Mp = 95-96 °C; [42] ¹H NMR (CDCl₃, 400 MHz): d = 8.51 (d, 1H; 4-CH), 7.70 (d, 1H; 5-CH), 7.30 (d, 1H; 8-CH), 7.27 (dd, 1H; 5-CH), 4.36-4.19 (m, 4H; 2 × CH₂), 1.38 ppm (t, 6H; 2 × CH₃); [43] ¹³C NMR (CDCl₃, 100.6 MHz): d = 156.9 (d), 155.8 (d), 152.3 (d), 151.7 (d), 131.0 (s), 120.3 (d), 118.2 (s), 117.8 (d), 117.8 (d), 110.5 (s), 63.7 (d), 16.4 ppm (d); ¹⁹F NMR (CDCl₃, 376.21 MHz): d = -72.5 ppm (s); [44] ³¹P NMR (CDCl₃, 161.91 MHz): d = 9.43 ppm (m). MS (EI, 70 eV): m/z (%) = 431 (100) [M + 1]⁺. HRMS (ESI): calcd. for C₁₄H₁₅F₃O₈PS⁺ [M + H]⁺ 431.0172; found 431.0176. Elem. analysis: C₁₄H₁₄F₃O₈PS (430.01): calcd. C 39.08, H 3.28, S 7.45; found C 39.25, H 3.25, S 7.47. Intermediates IIa-f [45] To a mixture of (PPh₃)₄Pd (241 mg, 0.209 mmol), Pd(OAc)₂ (63 mg, 0.279 mmol), and CuI (53 mg, 0.279 mmol) under argon atmosphere was added dry DMF (10 ml). The resulting suspension was purged with argon for 15 min at r.t. and afterwards heated at 40 °C for additional 15 min. Then, solution of 3 (3.49 mmol), the corresponding propargylamine derivative (4.89 mmol), and triethylamine (1.48 ml, 10.5 mmol) in dry DMF (20 ml, purged with argon) was added and the reaction mixture was stirred at 40 °C for 3 h. After cooing to r.t., the reaction mixture was diluted by EtOAc (800 ml). Then, water (150 ml) and conc. aq. NH₃ (2 ml) were added and the resulting mixture was stirred at r.t. for 30 min. The aqueous phase was separated and the organic phase was washed with water (4 × 150 ml) and brine (150 ml), dried over anhydrous Na₂SO₄, and concentrated under reduced pressure. The crude product was purified by silica gel column chromatography using a mixture of DCM/MeOH as an eluent to give the corresponding coupling product IIa-f in 62-76 % yield. [46] Diethyl (7-(3-(dimethylamino)prop-1-yn-1-yl)-2-oxo-2H-chromen-3- yl)phosphonate (II-1). Eluent: DCM/MeOH 1:0 ® 100:7; 62 % yield; [47] ¹H NMR (CDCl₃, 400 MHz): d = 8.47 (d, 1H; 4-CH), 7.51 (d, 1H; 5-CH), 7.40-7.38 (m, 1H; 8-CH), 7.36 (dd, 1H; 6-CH), 4.35-4.19 (m, 4H; 2 × OCH₂CH₃), 3.51 (s, 2H; CH₂NMe₂), 2.38 (s, 6H; N(CH₃)₂), 1.41-1.36 ppm (m, 6H; 2 × OCH₂CH₃); [48] ¹³C NMR (CDCl₃, 100.6 MHz): d =157.8 (d), 154.9 (d), 152.4 (d), 129.2 (m), 129.0 (s), 128.0 (d), 119.4 (d), 117.7 (d), 117.4 (d), 90.4 (s), 83.8 (s), 63.3 (d), 48.5 (s), 44.2 (s), 16.2 ppm (d); [49] ³¹P NMR (CDCl₃, 161.91 MHz): d = 10.59 ppm (m); HRMS (ESI): calcd. for C₁₈H₂₃NO₅P⁺ [M + H]⁺ 364.1308; found 364.1321. [50] Diethyl (2-oxo-7-(3-(piperidin-1-yl)prop-1-yn-1-yl)-2H-chromen-3- yl)phosphonate (II-2). Eluent: DCM/MeOH 1:0 ® 20:1; 67 % yield; [51] ¹H NMR (CDCl₃, 400 MHz): d = 8.43 (d, 1H; 4-CH), 7.48 (d, 1H; 5-CH), 7.35-7.33 (m, 1H; 8-CH), 7.32 (dd, 6-CH), 4.32-4.15 (m, 4H; 2 × OCH₂CH₃), 3.48 (s, 2H; 3’’-CH₂), 2.61-2.46 (m, 4H; 2’,6’-CH₂), 1.68-1.56 (m, 4H; 3’,5’-CH₂), 1.47-1.38 (m, 2H; 4’-CH₂), 1.35 ppm (t, 6H; 2 × OCH₂CH₃); [52] ¹³C NMR (CDCl₃, 100.6 MHz): d = 157.9 (d), 154.9 (d), 152.5 (d), 129.4 (d), 129.0 (s), 128.1 (d), 119.5 (d), 117.7 (d), 117.4 (d), 90.9 (s), 83.6 (s), 63.4 (d), 53.6 (s), 48.4 (s), 25.8 (s), 23.8 (s), 16.3 ppm (d); [53] ³¹P NMR (CDCl₃, 161.91 MHz): d = 10.57 ppm (s); HRMS (ESI): calcd. for C₂₁H₂₇NO₅P⁺ [M + H]⁺ 404.1621; found 404.1629. [54] Diethyl (7-(3-morpholinoprop-1-yn-1-yl)-2-oxo-2H-chromen-3-yl)phosphonate (II-3). Eluent: DCM/MeOH 1:0 ® 100:3; 76 % yield; [55] ¹H NMR (DMSO-d₆, 400 MHz): d = 8.64 (d, 1H; 4-CH), 7.93 (d, 1H; 5-CH), 7.52- 7.50 (m, 1H; 8-CH), 7.45 (dd, 1H; 6-CH), 4.20-4.06 (m, 4H; 2 × OCH₂CH₃), 3.65-3.59 (m, 4H; 2’,6’-CH₂), 3.58 (s, 2H; 3’’-CH₂), 2.57-2.51 (m, 4H; 3’,5’-CH₂), 1.27 ppm (t, 6H; 2 × OCH₂CH₃); [56] ¹³C NMR (DMSO-d₆, 100.6 MHz): d = 157.3 (d), 154.4 (d), 152.6 (d), 130.3 (s), 127.9 (m), 127.7 (m), 118.6 (m), 117.8 (d), 117.1 (d), 90.2 (s), 83.9 (s), 66.0 (s), 62.5 (d), 51.7 (s), 46.9 (s), 16.1 ppm (d); [57] ³¹P NMR (DMSO-d₆, 161.91 MHz): d = 10.80 ppm (s). HRMS (ESI): calcd. for C₂₀H₂₅NO₆P⁺ [M + H]⁺ 406.1414; found 406.1429. [58] Diethyl (7-(3-(4-methylpiperazin-1-yl)prop-1-yn-1-yl)-2-oxo-2H-chromen-3- yl)phosphonate (II-4). Eluent: DCM/MeOH 1:0 ® 10:1; 74 % yield; [59] ¹H NMR (CDCl₃, 400 MHz): d = 8.46 (d, 1H; 4-CH), 7.49 (d, ³J_(H,H) = 7.9 Hz, 1H; 5- CH), 7.38-7.32 (m, 2H; 6,8-CH), 4.35-4.18 (m, 4H; 2 × OCH₂CH₃), 3.57 (s, 2H; 3’’-CH₂), 2.80-2.41 (m, 8H; 2’,3’,5’,6’-CH₂), 2.32 (s, 3H; NCH₃), 1.38 ppm (t, 6H; 2 × OCH₂CH₃); [60] ¹³C NMR (CDCl₃, 100.6 MHz): d = 157.9 (d), 154.9 (d), 152.6 (d), 129.2 (m), 129.0 (s), 128.2 (d), 119.6 (d), 117.8 (d), 117.5 (d), 90.2 (s), 84.0 (s), 63.4 (d), 54.9 (s), 52.1 (s), 47.7 (s), 46.0 (s), 16.3 ppm (d); [61] ³¹P NMR (CDCl₃, 161.91 MHz): d = 10.57 ppm (s); HRMS (ESI): calcd. For C₂₁H₂₈N₂O₅P⁺ [M + H]⁺ calcd. 419.1730, found 419.1724. [62] Tert-Butyl 4-(3-(3-(diethoxyphosphoryl)-2-oxo-2H-chromen-7-yl)prop-2-yn-1- yl)piperazine-1-carboxylate (II-5). Eluent: DCM/MeOH 1:0 ® 200:8; 60 % yield; [63] ¹H NMR (CDCl₃, 400 MHz): d = 8.46 (dd, 1H; 4-CH), 7.51 (d, 1H; 5-CH), 7.38-7.36 (m, 1H; 8-CH), 7.34 (dd, 1H; 6-CH), 4.35-4.18 (m, 4H; 2 × OCH₂CH₃), 3.57 (s, 2H; 3’’- CH₂), 3.53-3.47 (m, 4H, 3’,5’-CH₂), 2.61-2.54 (m, 4H, 2’,6’-CH₂), 1.47 (s, 9H, C(CH₃)₃), 1.38 ppm (td, 6H; 2 × OCH₂CH₃); HRMS (ESI): calcd. For C₂₅H₃₄N₂O₇P⁺ [M + H]⁺ calcd. 505.2098, found 505.2116. [64] tert-butyl 4-(3-(3-(diethoxyphosphoryl)-2-oxo-2H-chromen-7-yl)prop-2-yn-1-yl)- 1,4-diazepane-1-carboxylate (II-6). Eluent: DCM/MeOH 1:0 ® 100:4; 87 % yield; [65] ¹H NMR (CDCl₃, 400 MHz): d = 8.45 (dd, 1H; 4-CH), 7.49 (d, 1H; 5-CH), 7.36-7.34 (m, 1H; 8-CH), 7.32 (dd, 1H; 6-CH), 4.34-4.17 (m, 4H; 2 × OCH₂CH₃), 3.63 (s, 2H; 3’’- CH₂), 3.59-3.42 (m, 4H, 2’’’,7’’’-CH₂), 2.82-2.68 (m, 4H; 3’’’,5’’’-CH₂), 1.95-1.82 (m, 2H, 6’’’-CH₂), 1.45 (s, 9H, C(CH₃)₃), 1.37 ppm (td, 6H; 2 × OCH₂CH₃); HRMS (ESI): calcd. For C₂₆H₃₆N₂O₇P⁺ [M + H]⁺ calcd. 519.2255, found 519.2274. [66] General method for preparation of selenopheno[3,2-h]chromenes (I-1- I-4). SeO₂ (0.333 g, 3.00 mmol) was dissolved in 48 % aq. HBr (1.29 ml) and stirred at r.t. for 15 min. Then, solution of the corresponding alkyne (1.00 mmol) in dioxane (15 ml) was added dropwise and the reaction mixture was stirred at r.t. for 20 h. After dilution of the reaction mixture by EtOAc (300 ml), sat. aq. NaHCO₃ (200 ml) was added and the resulting mixture was stirred at r.t. for 30 min. The organic phase was separated and the aqueous phase was extracted with EtOAc (2 × 100 ml). Combined organic phases were washed with brine (100 ml), dried over anhydrous Na₂SO₄ and concentrated under reduced pressure. The crude product was purified by silica gel column chromatography using a mixture of DCM/MeOH as an eluent to give a free base form of the corresponding cyclization product II. The corresponding hydrochlorides II were obtained by addition of 1 M HCl solution in Et₂O (3.0 equiv.) to a cooled (ice/water bath) solution of the corresponding free base form in dry DCM and stirring at the same temperature for 3 h. Removal of volatile matters under reduced pressure and drying under vacuum afforded II as greenish-yellow amorphous solids in 60- 70 % yield. Example 1. [67] Diethyl (7-bromo-8-((dimethylamino)methyl)-2-oxo-2H-selenopheno[3,2- h]chromen-3-yl)phosphonate hydrochloride (I-1). Eluent: DCM/MeOH200:1 ® 200:3; 60 % yield; [68] ¹H NMR (DMSO-d₆, 400 MHz): d = 10.65 (br s, 1H; NH), 8.81 (d, 1H; 4-CH), 8.09 (d, 1H; 5-CH), 7.86 (d, 1H; 6-CH), 4.86-4.68 (m, 2H; NCH₂), 4.23-4.07 (m, 4H; 2 × OCH₂CH₃), 2.96-2.72 (m, 6H; N(CH₃)₂), 1.28 ppm (t, 6H; 2 × OCH₂CH₃); [69] ¹³C NMR (free base) (100.6 MHz, CDCl₃): d = 157.8 (d), 154.0 (d), 152.7 (d), 152.6 (s), 146.1 (s), 125.8 (d), 125.7 (s), 121.1 (d), 116.1 (d), 113.3 (d), 105.9 (s), 63.4 (d), 60.4 (s), 46.1 (s), 16.3 ppm (d); [70] ³¹P NMR (free base) (161.91 MHz, CDCl₃): d = 11.13 ppm (m). HRMS (ESI): calcd. for C₁₈H₂₂BrNO₅PSe⁺ [M + H]⁺ calcd. 521.9579, found 521.9586. C₁₈H₂₁BrNO₅PSe ^. 1.8 HCl (586.83): calcd. C 36.84, H 3.92, N 2.39; found C 36.88, H 3.91, N 2.39. Example 2. [71] Diethyl (7-bromo-2-oxo-8-(piperidin-1-ylmethyl)-2H-selenopheno[3,2-h]chromen- 3-yl)phosphonate hydrochloride (I-2). Eluent: DCM/MeOH200:1 ® 100:1; 70 % yield; [72] ¹H NMR (CDCl₃, 400 MHz): d = 12.9 (br s, 1H; NH), 8.61 (d, 1H; 4-CH), 7.80 (d, 1H; 5-CH), 7.63 (d, 1H; 6-CH), 4.65 (m, 2H), 4.37-4.21 (m, 4H; 2 × OCH₂CH₃), 3.63-3.53 (m, 2H), 2.87-2.74 (m, 2H), 2.33-2.18 (m, 2H), 1.99-1.86 (m, 3H), 1.50-1.38 (m, 1H), 1.39 ppm (t, 6H; 2 × OCH₂CH₃); [73] ¹³C NMR (free base) (100.6 MHz, CDCl₃): d = 157.8 (d), 154.6 (s), 154.1 (d), 152.7 (d), 146.5 (s), 125.7 (m), 125.6 (s), 120.8 (m), 115.8 (d), 113.1 (d), 104.8 (s), 63.3 (d), 59.5 (s), 55.2 (s), 26.0 (s), 23.9 (s), 16.3 ppm (d); [74] ³¹P NMR (free base) (161.91 MHz, CDCl3): d = 11.29 ppm (m). HRMS (ESI): calcd. forC₂₁H₂₆BrNO₅PSe⁺ [M + H]⁺ calcd. 561.9892, found 561.9887. C₂₁H₂₅BrNO₅PSe ^. 1.3 HCl (608.66): calcd. C 41.44, H 4.36, N 2.30; found C 41.47, H 4.28, N 2.36. Example 3. [75] Diethyl (7-bromo-8-(morpholinomethyl)-2-oxo-2H-selenopheno[3,2-h]chromen-3- yl)phosphonate hydrochloride (I-3). Eluent: DCM/MeOH200:1 ® 100:1; 63 % yield; [76] ¹H NMR (DMSO-d₆, 400 MHz): d = 8.79 (d, 1H; 4-CH), 8.06 (d, 1H; 5-CH), 7.85- 7.77 (m, 1H; 6-CH), 4.92-4.40 (m, 2H), 4.23-4.07 (m, 4H; 2 × OCH₂CH₃), 3.95-3.64 (m, 4H), 3.36-2.86 (m, 4H), 1.28 ppm (t, 6H; 2 × OCH₂CH₃); [77] ¹³C NMR (free base) (100.6 MHz, CDCl₃): d = 157.7 (d), 153.9 (d), 152.6 (d), 151.8 (s), 146.2 (s), 125.8 (s), 125.7 (d), 121.0 (d), 116.2 (d), 113.3 (d), 106.0 (s), 67.0 (s), 63.3 (d), 59.2 (s), 54.1 (s), 16.3 ppm (d); [78] ³¹P NMR (free base) (161.91 MHz, CDCl3): d = 13.32 ppm (s). HRMS (ESI): calcd. for C₂₀H₂₄BrNO₆PSe⁺ [M + H]⁺calcd. 563.9684, found 563.9702. C₂₀H₂₃BrNO₆PSe ^. 1.3 HCl (610.64): calcd. C 39.34, H 4.01, N 2.29; found C 39.43, H 3.85, N 2.31. Example 4. [79] Diethyl (7-bromo-8-((4-methylpiperazin-1-yl)methyl)-2-oxo-2H-selenopheno[3,2- h]chromen-3-yl)phosphonate hydrochloride (I-4). Eluent: DCM/MeOH 1:0 ® 40:1; 48 % yield; [80] ¹H NMR (DMSO-d₆, 400 MHz): d = 10.81 (br s, 1H; NH), 8.77 (d, 1H; 4-CH), 8.03 (d, 1H; 5-CH), 7.74 (d, 1H; 6-CH), 4.21-4.08 (m, 4H; 2 × OCH₂CH₃), 4.09 (s, 2H), 3.47-3.37 (m, 2H), 3.22-3.08 (m, 4H), 2.86-2.73 (m, 5H), 1.28 ppm (t, 6H; 2 × OCH₂CH₃); [81] ¹³C NMR (free base) (100.6 MHz, CDCl₃): d = 157.8 (d), 154.0 (d), 153.0 (s), 152.7 (d), 146.3 (m), 125.7 (d), 125.7 (s), 120.9 (d), 116.1 (d), 113.2 (d), 105.5 (s), 63.3 (d), 58.7 (s), 55.1 (s), 53.6 (s), 45.9 (s), 16.3 ppm (d); [82] ³¹P NMR (161.91 MHz, DMSO-d₆): d = 10.97 ppm (s). HRMS (ESI): calcd. for C₂₁H₂₇BrN₂O₅PSe⁺ [M + H]⁺ calcd. 577.0001, found 577.0006. [83] General method for preparation of selenopheno[3,2-h]chromenes (I-5, I-6). SeO₂ (0.333 g, 3.00 mmol) was dissolved in 48 % aq. HBr (1.29 ml) and stirred at r.t. for 15 min. Then, solution of the corresponding alkyne II-5, II-6 (1.00 mmol) in dioxane (15 ml) was added dropwise and the reaction mixture was stirred at r.t. for 48 h. Then, reaction mixture was diluted with EtOH, till homogeneous greenish-yellow solution was obtained. The obtained solution was passed through an ion-exchange column containing Amberlite IRA-400 resin (counter ion Cl-) and, subsequently, the volatile matters were removed under reduced pressure. The crude material was pre-purified by reversed phase flash chromatography (30 % MeCN in H₂O (pH 4 by HCl)) and pure products were obtained by direct phase flash chromatography using a mixture of DCM/EtOH (slightly acidified by 1M HCl in dry Et₂O). After evaporation of the solvents, pure I-5 and I-6 were obtained in 15 % and 40 % yield respectively. Example 5. [84] Diethyl (7-bromo-2-oxo-8-(piperazin-1-ylmethyl)-2H-selenopheno[3,2- h]chromen-3-yl)phosphonate hydrochloride (I-5). Eluent: DCM/EtOH(HCl) 100:1 ® 10:1; 15 % yield; [85] ¹H NMR (CDCl₃, 400 MHz): d = 8.60 (d, 1H; 4-CH), 7.73 (d, 1H; 5-CH), 7.59 (d, 1H; 6-CH), 4.36-4.19 (m, 4H; 2 × OCH₂CH₃), 3.92 (s, 2H, 1’-CH₂), 3.19-3.13 (m, 4H, 3’’,5’’- CH₂), 2.91-2.84 (m, 4H, 2’’,6’’-CH₂), 1.39 ppm (td, 6H; 2 × OCH₂CH₃); HRMS (ESI): calcd. for C₂₀H₂₅BrN₂O₅PSe⁺ [M + H]⁺ calcd. 562.9844, found 562.9854. C₂₀H₂₄BrN₂O₅PSe ^. 1.0 HCl (598.72): calcd. C 40.12, H 4.21, N 4.68; found C 40.41, H 4.14, N 4.59. Example 6. [86] Diethyl (8-((1,4-diazepan-1-yl)methyl)-7-bromo-2-oxo-2H-selenopheno[3,2- h]chromen-3-yl)phosphonate hydrochloride (I-6). Eluent: DCM/EtOH(HCl) 100:1 ® 5:1; 40 % yield; [87] ¹H NMR (CDCl₃, 400 MHz): d = 9.68 (br s, 1H), 8.60 (d, 1H; 4-CH), 7.72 (d, 1H; 5- CH), 7.59 (d, 1H; 6-CH), 4.37-4.19 (m, 4H; 2 × OCH₂CH₃), 4.07 (s, 2H, 1’-CH₂), 3.51-3.32 (m, 4H, 3’’,5’’-CH₂), 3.20-3.10 (m, 2H), 3.05-2.97 (m, 2H), 2.25-2.16 (m, 2H, 6’’-CH₂), 1.39 ppm (td, 6H; 2 × OCH₂CH₃); HRMS (ESI): calcd. for C₂₁H₂₇BrN₂O₅PSe⁺ [M + H]⁺ calcd. 577.0001, found 577.0012. C₂₁H₂₆BrN₂O₅PSe ^. 1.1 HCl (616.40): calcd. C 40.92, H 4.43, N 4.54; found C 40.81, H 4.39, N 4.45. [88] General method for preparation of selenopheno[3,2-h]chromenes (I-7 – I-10). To the corresponding phosphonate ester I-1 – I-4 (1.00 mmol) was added conc. HCl (20 ml) and the reaction mixture was stirred under reflux for 5 h. After cooling to r.t., the volatile matters were removed under reduced pressure. Water (10 ml) was added to the crude product and the resulting suspension was stirred at r.t. for 5 h. After evaporation of the solvent, the product was dried under vacuum to give the corresponding product I-7 – I-10 as a pale brown amorphous solid in quantitative yield. Example 7. [89] (7-Bromo-8-((dimethylamino)methyl)-2-oxo-2H-selenopheno[3,2-h]chromen-3- yl)phosphonic acid hydrochloride (I-7). [90] ¹H NMR (DMSO-d₆, 400 MHz): d = 8.63 (d, 1H; 4-CH), 8.04 (d, 1H; 5-CH), 7.82 (d, 1H; 6-CH), 4.75 (s, 2H; NCH₂), 2.84 ppm (s, 6H; N(CH₃)₂); [91] ³¹P NMR (161.91 MHz, DMSO-d₆): d = 4.68 ppm (m). HRMS (ESI): calcd. for C₁₄H₁₄BrNO₅PSe⁺ [M + H]⁺ calcd. 465.8953, found 465.8953. C₁₄H₁₃BrNO₅PSe ^. 1.0 HCl ^. 0.7 H₂O (514.16): calcd. C 32.70, H 3.02, N 2.72; found C 32.50, H 3.04, N 2.69. Example 8. [92] (7-Bromo-2-oxo-8-(piperidin-1-ylmethyl)-2H-selenopheno[3,2-h]chromen-3- yl)phosphonic acid hydrochloride (I-8). [93] ¹H NMR (CD₃OD, 300 MHz): d = 8.60 (d, 1H; 4-CH), 7.94 (d, 1H; 5-CH), 7.90 (d, 1H; 6-CH),3.70-3.56 (m, 2H), 3.27-2.12 (m, 2H), 2.09-1.47 ppm (m, 6H); [94] ³¹P NMR (161.91 MHz, DMSO-d₆): d = 4.67 ppm (s). HRMS (ESI): calcd. for C₁₇H₁₈BrNO₅PSe⁺ [M + H]⁺ calcd. 505.9266, found 505.9264. C₁₇H₁₇BrNO₅PSe ^. 1.8 HCl (570.79): calcd. C 35.77, H 3.32, N 2.45; found C 35.72, H 3.38, N 2.35. Example 9. [95] (7-Bromo-8-(morpholinomethyl)-2-oxo-2H-selenopheno[3,2-h]chromen-3- yl)phosphonic acid hydrochloride (I-9). [96] ¹H NMR (CD₃OD, 300 MHz): d = 8.60 (d, 1H; 4-CH), 7.95 (d, 1H; 5-CH), 7.90 (d, 1H; 6-CH), 4.92 (s, 2H), 4.14-3.78 (m, 4H), 3.58-3.43 ppm (m, 4H); [97] ³¹P NMR (161.91 MHz, DMSO-d₆): d = 4.78 ppm (s). HRMS (ESI): calcd. for C₁₆H₁₆BrNO₆PSe⁺ [M + H]⁺calcd. 507.9058, found 507.9065. C₁₆H₁₅BrNO₆PSe ^. 1.3 HCl (554.53): calcd. C 34.66, H 2.96, N 2.53; found C 34.70, H 2.89, N 2.56. Example 10. [98] (7-Bromo-8-((4-methylpiperazin-1-yl)methyl)-2-oxo-2H-selenopheno[3,2- h]chromen-3-yl)phosphonic acid hydrochloride (I-10). [99] ¹H NMR (DMSO-d₆, 400 MHz): d = 8.61 (d, 1H; 4-CH), 7.99 (d, 1H; 5-CH), 7.71 (d, 1H; 6-CH), 4.06 (s, 3H; NCH₃), 3.46-3.38 (m, 2H), 3.21-3.07 (m, 4H), 2.82-2.70 ppm (m, 4H); [100] ³¹P NMR (161.91 MHz, DMSO-d₆): d = 4.97 ppm (s). HRMS (ESI): calcd. for C₁₇H₁₉BrN₂O₅PSe⁺ [M + H]⁺ calcd. 520.9375, found 520.9376. C₁₇H₁₈BrN₂O₅PSe ^. 1.8 HCl ^. 0.3 H₂O (591.20): calcd. C 33.41, H 3.36, N 4.58; found C 33.36, H 3.37, N 4.45. [101] Antiproliferative activity in vitro Anticancer activity of 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), SH-SY5Y (human neuroblastoma), CCL-8 (mouse sarcoma), MH-22A (mouse hepatoma), and HepG2 (human hepatocellular carcinoma) were cultured in standard medium DMEM (Dulbecco’s modified Eagle’s medium) (“Sigma”) supplemented with 10% heat-inactivated fetal bovine serum (“Sigma”). About 2-9^.10⁴ cells/mL (depending on line nature) were placed in 96-well plates immediately after compounds were added to the wells. Untreated cells were used as a control. The plates were incubated for 72 h, 37 ^oC, 5 % CO₂. The number of surviving cells was determined using 3- (4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolinium bromide (MTT). MTT-test: after incubating culture medium was removed and 200 µL fresh medium with 10mM HEPES was added in each well of the plate, than 20 µL MTT (2mg/mL in HBSS) was added. After incubation (3 hr., 37 ^oC, 5 % CO₂), the medium with MTT was removed and 200 µL DMSO were added at once to each sample. The samples were tested at 540 nm on Anthos HT II photometer. [102] In parallel, the borderline concentration relevant to the highest tolerated dose has been determined for each compound using the NIH 3T3 (mouse embryo fibroblasts) cell line and the basal cytotoxicity obtained has then been used to predict the starting doses for in vivo acute oral LD₅₀ values in rodents. [103] Basal cytotoxicity test: the Neutral Red Uptake (NRU) assay was performed according to the standard protocol of Stokes modified by the NICEATM-ECVAM validation study. The NRU cytotoxicity assay procedure is based on the ability of viable cells to incorporate and bind Neutral Red, a supravitally dye. Balb/c 3T3 (Mouse Swiss Albino embryo fibroblast) cells (9000 cells/well) were placed into 96-well plates for 24 h in Dulbecco's modified Eagle's (DMEM) medium containing 5 % fetal bovine serum. The cells were then exposed to the test compounds over a range of seven concentrations (1000, 316, 100, 31, 10, 3, 1 µg/ml) for 24 h. Untreated cells were used as a control. After 24 h, the medium was removed from all plates. Then, 250 µL of Neutral Red solution were added (0.05 mg/mL NR in DMEM 24 h pre-incubated at 37 °C and then filtered before use through 0.22- µm syringe filter). Plates were incubated for 3 h and then cells were washed three times with PBS. The dye within viable cells was released by extraction with a mixture of acetic acid, ethanol and water (1:50:49). Absorbance of Neutral Red was measured using a spectrophotometer multiplate reader (TECAN, Infinite M1000) at 540 nm. The optical density (OD) was calculated using the formula: OD (treated cells)*100/OD (control cells). The IC₅₀ values were calculated using the programme Graph Pad Prism® 3.0. [104] Estimation of LD₅₀ from IC₅₀ values: Data from the in vitro tests were used for estimating the starting dose for acute oral systemic toxicity tests in rodents. The in vivo starting dose is an estimated LD₅₀ value calculated by inserting the in vitro IC₅₀ value into a regression formula: log LD₅₀ (mM/kg) = 0.439 log IC₅₀ (mM) + 0.621. The value is recalculated to mg/kg and compounds are evaluated in accordance with four toxicity categories: category 1: LD₅₀ £ 5 mg/kg (highly toxic); category 2: 5 < LD₅₀ £ 50 mg/kg (moderately toxic); category 3: 50 < LD₅₀ £ 300 mg/kg (slightly toxic); category 4: 300 < LD₅₀ £ 2000 mg/kg (practically non-toxic). [105] The results of cell culture-based studies are summarized in Table 1. In opposite to previously claimed selenophenochromenones, tested compounds with few exceptions showed medium to high cytotoxicity against malignant tumor cells. Notably, all derivatives are medium to low toxic to normal NIH 3T3 cells according to basal cytotoxicity test (LD₅₀ > 542 mg/kg) what is very surprisingly for Se-containing compounds. [106] Derivative I-3 is able to suppress human breast adenocarcinoma cell growth (IC₅₀=7.5 µM). Dimethylaminomethyl selenophenochromene phosphonic acid I-7 showed impressive cytotoxic effect on MDA-MB-435s (human melanoma, IC₅₀=1.6 µM) and HT-1080 (human fibrosarcoma, IC₅₀=9.3 µM) cell lines, although, its corresponding ester I-1 showed only moderate activity (IC₅₀=29÷114 µM) on studied cancer cell lines. Notably, all the rest malignant cells were not sensitive to I-7. The introduction of morpholyl (I-8), piperidyl (I-9), and methylpiperazyl (I-10) substituents instead of dimethylamino moiety led to dramatic decrease in cytotoxic effect. The performed tests of cytotoxicity of synthesized compounds of Formula I on cell lines clearly demonstrated, that compounds of this application are highly active in treatment of cancer of different localizations. This unexpected discovery together with unprecedentedly high selectivity of compounds towards cancer cells makes these compounds very promising as anti-cancer medicines. Table 1. In vitro cytotoxicity of selenopheno[h]chromenes I-1 – I-10 on 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), SH-SY5Y (human neuroblastoma), CCL-8 (mouse sarcoma), 3T3 (mouse embryo fibroblasts), MH-22A (mouse hepatoma), HepG2 (human hep ll l i ) C Basal cytotoxicity Estimated* LD₅₀, mM/kg mg/kg _I-1 7 40±1 70±15 114±11 2.6±0.2 1530 _I-2 6 37±1 7±7 78±7 1.2±0.06 734.5 _I-3 6 40±1 77±6 120±6 3.0±0.1 1818 I-4 6 40±1 39±8 73±11 2.5±0.01 1726 I-5 7 89±2 27±1 72±7 0.9±0.04 542 I-6 5 27±1 14±2 16±2 1.0±0.1 648 I-7 1 >194 >194 >194 >3.8 >2000 I-8 2 59±4 >180 53±5 1.8±0.09 980 I-9 > ne ne ne 3.8±0.05 2158 I-10 n ne ne ne 4.7±0.02 2871

Literature [1] Z. Xiao, S. L. Morris-Natschke, K.-H. Lee, Strategies for the Optimization of Natural Leads to Anticancer Drugs or Drug Candidates, Med Res Rev. 36 (2016) 32–91. [2] A. K. Ghosh, H. L. Osswald, G. Prato, Recent Progress in the Development of HIV-1 Protease Inhibitors for the Treatment of HIV/AIDS, J Med Chem. 59 (2016) 5172–5208. [3] F. J. Reen, J. A. Gutiérrez-Barranquero, M. L. Parages, F. O Gara, Coumarin: a novel player in microbial quorum sensing and biofilm formation inhibition. Appl Microbiol Biotechnol. 102 (2018) 2063-2073. [4] E. Kozioł, K. Skalicka-Woźniak, Imperatorin-pharmacological meaning and analytical clues: profound investigation. Phytochem Rev. 15 (2016) 627-649; [5] D. Bądziul, J. Jakubowicz-Gil, R, Paduch, K. Głowniak, A. Gawron, Combined treatment with quercetin and imperatorin as a potent strategy for killing HeLa and Hep-2 cells. Mol Cell Biochem. 392 (2014) 213-27. [6] Z. R. Zhang, W. N. Leung, H. Y. Cheung, C. W. Chan, Osthole: A Review on Its Bioactivities, Pharmacological Properties, and Potential as Alternative Medicine. Evid Based Complement Alternat Med. (2015) 2015:919616. [7] J. J. Li, W. L. Chen, J. Y. Wang, Q. W. Hu, Z. P. Sun, S. Zhang, S. Liu, X. H. Han, Wenshen Zhuanggu formula effectively suppresses breast cancer bone metastases in a mouse Xenograft model. Acta Pharmacol Sin. 38 (2017) 1369-1380. [8] E. Budzisz, E. Brzezinska, U. Krajewska, M. Rozalski, Cytotoxic effects, alkylating properties and molecular modelling of coumarin derivatives and their phosphonic analogues, Eur. J. Med. Chem., 38 (2003) 597-603. [9] P.Arsenjans, J.Vasiljeva, I.Domracheva, I.Shestakova, I.Kalvins. WO2018015788 (A1)., 25.01.2018.

Claims

CLAIMS 1. A compound of Formula I:

R¹ represents H or alkyl; R² represents a halogen atom; R³ represents C_1-4-alkyl-N-heterocyclyl; its optical isomers, polymorphs and pharmaceutically acceptable acid addition salts and hydrates and solvates thereof. 2. The compound according to Claim 1 for use as medicament for the treatment of disease in which inhibition of cell proliferation (melanoma, carcinoma, sarcoma, neuroblastoma, hepatoma) is required. 3. The compound according to Claim 1, wherein the compound is selected from the group consisting of: Diethyl (7-bromo-8-((dimethylamino)methyl)-2-oxo-2H-selenopheno[3,2-h]chromen- 3-yl)phosphonate; Diethyl (7-bromo-2-oxo-8-(piperidin-1-ylmethyl)-2H-selenopheno[3,2-h]chromen-3- yl)phosphonate hydrochloride; Diethyl (7-bromo-8-(morpholinomethyl)-2-oxo-2H-selenopheno[3,2-h]chromen-3- yl)phosphonate hydrochloride; Diethyl (7-bromo-8-((4-methylpiperazin-1-yl)methyl)-2-oxo-2H-selenopheno[3,2- h]chromen-3-yl)phosphonate; Diethyl (7-bromo-2-oxo-8-(piperazin-1-ylmethyl)-2H-selenopheno[3,2-h]chromen-3- yl)phosphonate hydrochloride; Diethyl (8-((1,4-diazepan-1-yl)methyl)-7-bromo-2-oxo-2H-selenopheno[3,2- h]chromen-3-yl)phosphonate hydrochloride; (7-Bromo-8-((dimethylamino)methyl)-2-oxo-2H-selenopheno[3,2-h]chromen-3- yl)phosphonic acid hydrochloride; (7-Bromo-2-oxo-8-(piperidin-1-ylmethyl)-2H-selenopheno[3,2-h]chromen-3- yl)phosphonic acid hydrochloride; (7-Bromo-8-(morpholinomethyl)-2-oxo-2H-selenopheno[3,2-h]chromen-3- yl)phosphonic acid hydrochloride; (7-Bromo-8-((4-methylpiperazin-1-yl)methyl)-2-oxo-2H-selenopheno[3,2- h]chromen-3-yl)phosphonic acid hydrochloride and pharmaceutically acceptable enantiomers, diastereomers, racemates, and salts thereof. 4. The compound accordin

g to Claim 1-3, wherein the compound has the structure:

5. A process for the synthesis of a compound of Formula I:

wherein: R¹ represents H or alkyl; R² represents a halogen atom; R³ represents C_1-4-alkyl-N-heterocyclyl; comprising reacting an intermediate of Formula II with selenium(IV) bromide:

wherein: R¹ represents alkyl; R² represents C_1-4alkyl-N-heterocyclyl. 6. A process for the synthesis of an intermediate of Formula II comprising treatment of an intermediate of Formula 3:

wherein: R¹ represents alkyl; with terminal acetylene containing N-heterocyclyl moiety in the presence of a catalytic amount of tetrakis(triphenylphosphine)palladium(0) and copper iodide.