LIPOPHOSPHONOXINS OF SECOND GENERATION, AND THEIR USE

Abstract

Lipophosphonoxins of general Formula I are described, in which R.sub.1 is C8-C22, preferably C10-C18 and more preferably C12-C16 alkyl, or hexadecyloxypropyl, tetradecyloxypropyl, tetradecyloxyethyl or hexadecyloxyethyl, R.sub.2 is uracil, thymine, or cytosine, and R.sub.3 is selected from a group which contains compounds of general formulas II and III.

Claims

1: Lipophosphonoxins of general formula I, ##STR00013## wherein R.sub.1 is selected from C8-C22 alkyl, hexadecyloxypropyl, tetradecyloxypropyl, tetradecyloxyetyl, hexadecyloxyetyl; R.sub.2 is selected from uracil, thymine, cytosine; and R.sub.3 is selected from the group consisting of compounds of formulas II-V: ##STR00014## wherein R.sub.4 is H, CH.sub.2NH.sub.2 or CH.sub.2OH, R.sub.5 is H, NH.sub.2 or OH, R.sub.6 is H, NH.sub.2 or OH, R.sub.7 is H, CH.sub.2NH.sub.2 or CH.sub.2OH, R.sub.8 is H, CH.sub.2NH.sub.2 or CH.sub.2OH, R.sub.9 is H, NH.sub.2 or OH, R.sub.10 is H, NH.sub.2 or OH, R.sub.11 is H, NH.sub.2 or OH, R.sub.12 is H, CH.sub.2NH.sub.2 or CH.sub.2OH, R.sub.13 is NH.sub.2 or NH—CH(NH.sub.2)NH, R.sub.14 is NH.sub.2 or NH—CH(NH.sub.2)NH, R.sub.15 is NH.sub.2 or NH—CH(NH.sub.2)NH, R.sub.16 is NH.sub.2 or NH—CH(NH.sub.2)NH, whereas at least one of R.sub.5 and R.sub.6 groups must be NH.sub.2 or at least one of R.sub.4 and R.sub.7 groups must be CH.sub.2NH.sub.2, and whereas at least one of R.sub.9, R.sub.10 and R.sub.11 groups must be NH.sub.2 or at least one of R.sub.8 and R.sub.12 groups must be CH.sub.2NH.sub.2; and their pharmaceutically acceptable salts and/or hydrates.

2: Lipophosphonoxins of general formula I according to claim 1, or pharmaceutically acceptable salts and/or hydrates and/or mixtures of such compounds, for use as a medicament.

3: Lipophosphonoxins of general formula I according to claim 1 or their diastereomers, or pharmaceutically acceptable salts and/or hydrates and/or mixtures of such compounds, for use as an antibacterial agent.

4: Antibacterial drug, characterized in that it contains at least one lipophosphonoxin of general formula I according to claim 1, or a diastereomer, or a pharmaceutically acceptable salt and/or hydrate, and/or a mixture of such compounds as the active ingredient.

5: Disinfectant for other than therapeutic purposes and/or selective culture medium characterized in that it contains at least one lipophosphonoxin of general formula I according to claim 1, or its diastereomer, or a pharmaceutically acceptable salt and/or hydrate, and/or mixture of such compounds, as the active ingredient.

6: A method of treatment of bacterial infections comprising the step of administering the lipophosphonoxins of Formula I according to claim 1, or their diastereomers, or pharmaceutically acceptable salts and hydrates, and/or mixtures of such compounds to the subject in need of such treatment.

7: A method of preparation of a disinfectant comprising the step of providing lipophosphonoxins of Formula I according to claim 1, or their diastereomers, or pharmaceutically acceptable salts and/or hydrates, and/or mixtures of such compounds.

8: A method of preparation of selective cultivation medium for in vitro cultures comprising the step of providing lipophosphonoxins of Formula I according to claim 1, or their diastereomers, or pharmaceutically acceptable salts and/or hydrates, and/or mixtures of such compounds.

Description

EXAMPLES

List of Abbreviations

[0036] DCM dichloromethane [0037] TPSCl triisopropylbenzenesulfonylchloride [0038] IR infrared spectrum [0039] HR-ESI high-resolution electrospray ionisation mass spectrum [0040] HR-EI high-resolution electroimpact ionisation mass spectrum [0041] n-BuOH n-butylalcohol [0042] DMTr dimethoxytrityl [0043] THF tetrahydrofuran [0044] EC.sub.50 median active (effective) concentration (causing 50% of maximum effect) [0045] IC.sub.50 inhibitory concentration (causing 50% of the maximum inhibitory effect) [0046] rt room temperature [0047] MIC minimum inhibitory concentration [0048] MBC minimal bactericidal concentration

Example 1

Hexadecyl-uridine-5′-yl-2-N-bis (3-aminopropyl)-2-aminoethyl phosphonate

[0049] ##STR00003##

[0050] A mixture of bis boc-N-1-(3-aminopropyl)propane-1,3-diamine (0.53 g, 1.5 mmol) (prepared according to J. Med. Chem. 2014, 57 (22), 9409-9423) and hexadecyl-2′,3′-isopropylidenuridin-5′-yl-vinylphosphonate (0.6 g, 1 mmol) (prepared according to J. Med. Chem. 2011, 54(22), 7884-7898) in n-BuOH (10 ml) was stirred overnight at 105° C. The reaction mixture was concentrated in vacuum and the isopropylidene-protected intermediate was purified by chromatography on silica gel using a linear gradient of ethanol in chloroform (0-10%). The resulting solid was dissolved in 0.5 mol.Math.l.sup.−1 HCl in methanol (40 ml) and the mixture was stirred for 12 hours at room temperature. The reaction mixture was concentrated to about half volume on rotary evaporator and added to cold ethyl acetate (20 ml). The solid obtained was filtered and dried. This resulted in the desired product as an amorphous solid in 74% yield (0.56 g, 0.74 mmol).

[0051] .sup.1H NMR (500.0 MHz, CD.sub.3OD): 0.90 (m, 6H, CH.sub.3(CH.sub.2).sub.14CH.sub.2O); 1.24-1.43 (m, 52H, CH.sub.3(CH.sub.2).sub.13CH.sub.2CH.sub.2O); 1.71 (m, 4H, CH.sub.3(CH.sub.2).sub.13CH.sub.2CH.sub.2O); 2.15-2.25 (bm, 8H, NCH.sub.2CH.sub.2CH.sub.2NH.sub.2); 2.52-2.67 (m, 4H, PCH.sub.2CH.sub.2N); 3.10 (t, 8H, J.sub.vic=7.5, NCH.sub.2CH.sub.2CH.sub.2NH.sub.2); 3.35-3.42 (bm, 8H, NCH.sub.2CH.sub.2CH.sub.2NH.sub.2); 3.44-3.52 (bm, 4H, PCH.sub.2CH.sub.2N); 4.10-4.21 (m, 8H, H-3′,4′, CH.sub.3(CH.sub.2).sub.14CH.sub.2O); 4.27 (dd, 1H, J.sub.2′,3′=5.4, J.sub.2′,1′=4.2, H-2′); 4.28 (dd, 1H, J.sub.2′,3′=5.3, J.sub.2′,1′=3.9, H-2′); 4.34 (ddd, 1H, J.sub.gem=11.6, J.sub.H,P=7.5, J.sub.5′b,4′=5.4, H-5′b); 4.39 (dd, 2.sub.H,P=7.6, J.sub.5′,4′=4.3, H-5′); 4.43 (ddd, 1H, J.sub.gem=11.6, J.sub.H,P=7.3, J.sub.5′a,4′=2.9, H-5′a); 5.77 (d, 2H, J.sub.5,6=8.0, H-5); 5.84 (d, 1H, J.sub.1′,2′=4.2, H-1′); 5.85 (d, 1H, =3.9, H-1′); 7.74 (d, 1H, J.sub.6,5=8.0, H-6); 7.75 (d, 1H, J.sub.6,5=8.0, H-6).

[0052] .sup.13C NMR (125.7 MHz, CD.sub.3OD): 14.45 (CH.sub.3(CH.sub.2).sub.14CH.sub.2O); 21.33 (d, J.sub.C,P=140.8, PCH.sub.2CH.sub.2N); 21.37 (d, J.sub.C,P=141.1, PCH.sub.2CH.sub.2N); 23.28 (NCH.sub.2CH.sub.2CH.sub.2NH.sub.2); 23.73, 26.57, 30.32, 30.47, 30.68, 30.75, 30.76, 30.80 (CH.sub.3(CH.sub.2).sub.13CH.sub.2CH.sub.2O); 31.56 (d, J.sub.C,P=5.9, CH.sub.3(CH.sub.2).sub.13CH.sub.2CH.sub.2O); 31.55, 31.56 (d, J.sub.C,P=5.9, CH.sub.3(CH.sub.2).sub.13CH.sub.2CH.sub.2O); 33.07 (CH.sub.3(CH.sub.2).sub.13CH.sub.2CH.sub.2O); 37.87 (NCH.sub.2CH.sub.2CH.sub.2NH.sub.2); 48.58 (NCH.sub.2CH.sub.2P); 51.09 (NCH.sub.2CH.sub.2CH.sub.2NH.sub.2); 67.37 (d, J.sub.C,P=6.1, CH.sub.2-5′); 68.37 (d, J.sub.C,P=6.8, CH.sub.3(CH.sub.2).sub.14CH.sub.2O); 68.56 (d, J.sub.C,P=6.8, CH.sub.3(CH.sub.2).sub.14CH.sub.2O); 70.81, 70.90 (CH-3′); 74.61, 74.65 (CH-2′); 83.37 (d, J.sub.C,P=6.0, CH-4′); 83.39 (d, J.sub.C,P=6.2, CH-4′); 92.14, 92.26 (CH-1′); 103.17, 103.21 (CH-5); 143.00, 143.04 (CH-6); 152.22, 152.28 (C-2); 165.96, 165.97 (C-4). .sup.31P{.sup.1H} NMR (202.3 MHz, CD.sub.3OD): 27.67; 28.13.

[0053] IR v.sub.max(KBr) 3426 (s, vbr), 3047 (m, vbr), 2640 (m, vbr, sh), 2090 (w, vbr, sh), 1700 (vs, sh), 1681 (vs), 1467 (m), 1429 (w), 1390 (w), 1261 (w, br), 1206 (s), 1080 (w, sh), 1060 (m), 1021 (m, br), 1002 (m), 764 (vw, sh).

[0054] HR-ESI C.sub.33H.sub.65O.sub.8N.sub.5P (M+H).sup.+ calculated 690.45653, found 690.45656.

Example 2

Pentadecyl-uridine-5′-yl-2-N-bis(3-aminopropyl)-2-aminoethyl phosphonate

[0055] ##STR00004##

[0056] The compound in Example 2 was prepared by the same procedure as the one in Example 1 from bis boc-N-1-(3-aminopropyl)propane-1,3-diamine (0.53 g, 1.5 mmol) and pentadecyl-2′,3′-isopropylidenuridine-5′-yl-vinylphosphonate (prepared according to J. Med. Chem. 2011, 54(22), 7884-7898) (0.63 g, 1 mmol) in 75% yield (0.56 g, 0.75 mmol).

[0057] .sup.1H NMR (500.0 MHz, CD.sub.3OD): 0.90 (m, 6H, CH.sub.3(CH.sub.2).sub.13CH.sub.2O-A,B); 1.24-1.43 (m, 48H, CH.sub.3(CH.sub.2).sub.12CH.sub.2CH.sub.2O-A,B); 1.71 (m, 4H, CH.sub.3(CH.sub.2).sub.12CH.sub.2CH.sub.2O-A,B); 2.15-2.23 (bm, 8H, NCH.sub.2CH.sub.2CH.sub.2NH.sub.2-A,B); 2.52-2.62 (m, 4H, PCH.sub.2CH.sub.2N-A,B); 3.09 (t, 8H, J.sub.vic=7.5, NCH.sub.2CH.sub.2CH.sub.2NH.sub.2); 3.32-3.42 (bm, 8H, NCH.sub.2CH.sub.2CH.sub.2NH.sub.2); 3.43-3.51 (bm, 4H, PCH.sub.2CH.sub.2N); 4.11-4.20 (m, 8H, H-3′,4′-A,B, CH.sub.3(CH.sub.2).sub.13CH.sub.2O-A,B); 4.26 (dd, 1H, =5.3, J.sub.2′,1′=4.1, H-2′-A); 4.28 (dd, 1H, =5,1, =3.9, H-2′-B); 4.30-4.45 (m, 4H, H-5′-A,B); 5.764 (d, 1H, J.sub.5,6=8.0, H-5-A); 5.766 (d, 1H, J.sub.5,6=8.0, H-5-B); 5.83 (d, 1H, J.sub.1′,2′=4.1, H-1′-A); 5.84 (d, 1H, J.sub.1′,2′=3.9, H-1′-B); 7.73 (d, 1H, J.sub.6,5=8.0, H-6-B); 7.74 (d, 1H, J.sub.6,5=8.0, H-6-A). .sup.13C NMR (125.7 MHz, CD.sub.3OD): 14.45 (CH.sub.3(CH.sub.2).sub.13CH.sub.2O-A,B); 21.30 (d, J.sub.C,P=140.7, PCH.sub.2CH.sub.2N-A); 21.35 (d, J.sub.C,P=140.9, PCH.sub.2CH.sub.2N—B); 23.34 (NCH.sub.2CH.sub.2CH.sub.2NH.sub.2-A,B); 23.74, 26.58, 30.33, 30.48, 30.69, 30.75, 30.77, 30.79; 30.81 (CH.sub.3(CH.sub.2).sub.12CH.sub.2CH.sub.2O-A,B); 31.56 (d, J.sub.C,P=5.9, CH.sub.3(CH.sub.2).sub.12CH.sub.2CH.sub.2O-A); 31.57 (d, J.sub.C,P=5.9, CH.sub.3(CH.sub.2).sub.12CH.sub.2CH.sub.2O—B); 33.08 (CH.sub.3(CH.sub.2).sub.12CH.sub.2CH.sub.2O-A,B); 37.87 (NCH.sub.2CH.sub.2CH.sub.2NH.sub.2-A,B); 48.51 (NCH.sub.2CH.sub.2P-A,B); 51.12 (NCH.sub.2CH.sub.2CH.sub.2NH.sub.2-A,B); 67.41 (d, J.sub.C,P=6.3, CH.sub.2-5′-A); 67.44 (d, J.sub.C,P=6.1, CH.sub.2-5′-B); 68.36 (d, J.sub.C,P=6.8, CH.sub.3(CH.sub.2).sub.13CH.sub.2O—B); 68.56 (d, J.sub.C,P=6.8, CH.sub.3(CH.sub.2).sub.13CH.sub.2O-A); 70.84 (CH-3′-B); 70.91 (CH-3′-A); 74.59 (CH-2′-A); 74.63 (CH-2′-B); 83.35 (d, J.sub.C,P=6.1, CH-4′-A,B); 92.31 (CH-1′-A); 92.40 (CH-1′-B); 103.13 (CH-5-A); 103.18 (CH-5-B); 143.03 (CH-6-A); 143.07 (CH-6-B); 152.20 (C-2-A); 152.27 (C-2-B); 165.98 (C-4-A); 165.99 (C-4-B).

[0058] .sup.31P{.sup.1H} NMR (202.3 MHz, CD.sub.3OD): 27.65 (A); 28.10 (B).

[0059] IR v.sub.max(KBr) 3050 (s, vbr, sh), 3411 (s, br), 2645 (m, br), 2924 (vs), 2854 (vs), 2563 (m, br), 2035 (w, br), 1975 (w, br, sh), 1690 (vs, br), 1624 (m), 1520 (m, br, sh), 1466 (s), 1408 (m), 1386 (m), 1266 (s), 1233 (s, br, sh), 1075 (s, sh), 1055 (s), 1035 (s, br, sh), 997 (s), 822 (m), 764 (w), 721 (w).

[0060] HR-ESI C.sub.32H.sub.63O.sub.8N.sub.5P (M+H).sup.+ calculated 676.44088, found 676.44092.

Example 3

Tetradecyl-uridine-5′-yl-2-N-bis (3-aminopropyl)-2-aminoethyl phosphonate

[0061] ##STR00005##

[0062] The compound in Example 3 was prepared by the same procedure as the one in Example 1 from boc-N-1-(3-aminopropyl)propane-1,3-diamine (0.62 g, 1.86 mmol) and tetradecyl-2′,3′-isopropylidenuridine-5′-yl-vinylphosphonate (prepared according to J. Med. Chem. 2011, 54(22), 7884-7898) (0.76 g, 1.33 mmol) in 65% yield (0.64 g, 0.87 mmol).

[0063] .sup.1H NMR (500.0 MHz, CD.sub.3OD): 0.90 (m, 6H, CH.sub.3(CH.sub.2).sub.12CH.sub.2O-A,B); 1.24-1.43 (m, 44H, CH.sub.3(CH.sub.2).sub.11CH.sub.2CH.sub.2O-A,B); 1.71 (m, 4H, CH.sub.3(CH.sub.2).sub.12CH.sub.2CH.sub.2O-A,B); 2.16-2.26 (bm, 8H, NCH.sub.2CH.sub.2CH.sub.2NH.sub.2-A,B); 2.54-2.65 (m, 4H, PCH.sub.2CH.sub.2N-A,B); 3.10 (t, 8H, J J.sub.vic=7.5, NCH.sub.2CH.sub.2CH.sub.2NH.sub.2); 3.35-3.42 (bm, 8H, NCH.sub.2CH.sub.2CH.sub.2NH.sub.2); 3.44-3.52 (bm, 4H, PCH.sub.2CH.sub.2N); 4.08-4.22 (m, 8H, H-3′,4′-A,B, CH.sub.3(CH.sub.2).sub.12CH.sub.2O-A,B); 4.27 (dd, 1H, J.sub.2′,3′=5.4, J.sub.2′,1′=4.2, H-2′-B); 4.28 (dd, 1H, J.sub.2′,3′=5.3, J.sub.2′,1′=3.9, H-2′-A); 4.34 (ddd, 1H, J.sub.gem=11.5, J.sub.H,P=7.4, J.sub.5′b,4′=5.4, H-5′b-B); 4.39 (dd, 2H, J.sub.H,P=7.6, J.sub.5′,4′=4.2, H-5′-A); 4.43 (ddd, 1H, J.sub.gem=11.6, J.sub.H,P=7.2, J.sub.5′a,4′=2.9, H-5′a-B); 5.78 (d, 2H, J.sub.5,6=8.0, H-5-A,B); 5.84 (d, 1H, J.sub.1′,2′=4.2, H-1′-B); 5.85 (d, 1H, J.sub.1′,2′=3.9, H-1′-A); 7.74 (d, 1H, J.sub.6,5=8.0, H-6-A); 7.75 (d, 1H, J.sub.6,5=8.0, H-6-B).

[0064] .sup.13C NMR (125.7 MHz, CD.sub.3OD): 14.45 (CH.sub.3(CH.sub.2).sub.12CH.sub.2O-A,B); 21.33 (d, J.sub.C,P=140.7, PCH.sub.2CH.sub.2N—B); 21.38 (d, J.sub.C,P=141.1, PCH.sub.2CH.sub.2N-A); 23.28 (NCH.sub.2CH.sub.2CH.sub.2NH.sub.2-A,B); 23.73, 26.57, 30.32, 30.48, 30.68, 30.74, 30.76, 30.78, 30.79, 30.81 (CH.sub.3(CH.sub.2).sub.11CH.sub.2CH.sub.2O-A,B); 31.55 (d, J.sub.C,P=5.8, CH.sub.3(CH.sub.2).sub.11CH.sub.2CH.sub.2O—B); 31.56 (d, J.sub.C,P=5.9, CH.sub.3(CH.sub.2).sub.11CH.sub.2CH.sub.2O-A); 33.07 (CH.sub.3(CH.sub.2).sub.11CH.sub.2CH.sub.2O-A,B); 37.87 (NCH.sub.2CH.sub.2CH.sub.2NH.sub.2-A,B); 48.59 (NCH.sub.2CH.sub.2P-A,B); 51.09 (NCH.sub.2CH.sub.2CH.sub.2NH.sub.2-A,B); 67.36 (d, J.sub.C,P=6.2, CH.sub.2-5′-A); 67.37 (d, J.sub.C,P=6.2, CH.sub.2-5′-B); 68.37 (d, J.sub.C,P=6.8, CH.sub.3(CH.sub.2).sub.12CH.sub.2O—B); 68.56 (d, J.sub.C,P=6.8, CH.sub.3(CH.sub.2).sub.12CH.sub.2O-A); 70.81 (CH-3′-A); 70.90 (CH-3′-B); 74.61 (CH-2′-B); 74.66 (CH-2′-A); 83.37 (d, J.sub.C,P=5,9, CH-4′-A); 83.39 (d, J.sub.C,P=6.1, CH-4′-B); 92.12 (CH-1′-B); 92.24 (CH-1′-A); 103.17 (CH-5-B); 103.21 (CH-5-A); 142.99 (CH-6-B); 143.04 (CH-6-A); 152.23 (C-2-B); 152.28 (C-2-A); 165.96 (C-4-B); 165.97 (C-4-A).

[0065] .sup.31P{.sup.1H} NMR (202.3 MHz, CD.sub.3OD): 27.66 (B); 28.12 (A).

[0066] IR v.sub.max(CHCl.sub.3) 3415 (s, vbr), 3045 (s, vbr, sh), 2924 (vs), 2854 (s), 2644 (m, vbr), 2563 (m, vbr), 2028 (w, vbr), 1972 (w, vbr, sh), 1690 (vs, br), 1624 (m), 1520 (w, br, sh), 1465 (s), 1407 (m), 1386 (m), 1266 (s), 1232 (s, sh), 1072 (s, sh), 1054 (s), 1015 (s, sh), 996 (s), 823 (m), 763 (w), 721 (w).

[0067] HR-ESI C.sub.31H.sub.61O.sub.8N.sub.5P (M+H).sup.+ calculated 662.42523, found 662.42502.

Example 4

Tridecyl-uridine-5′-yl-2-N-bis (3-aminopropyl)-2-aminoethyl phosphonate

[0068] ##STR00006##

[0069] The compound in Example 4 was prepared by the same procedure as the one in Example 1 from boc-N-1-(3-aminopropyl)propane-1,3-diamine (0.95 g, 2.85 mmol) and tridecyl-2′,3′-isopropylidenuridine-5′-yl-vinylphosphonate (prepared according to J. Med. Chem. 2011, 54(22), 7884-7898) (1.32 g, 2.38 mmol) in 65% yield (1.11 g, 1.54 mmol).

[0070] .sup.1H NMR (500.0 MHz, CD.sub.3OD): 0.90 (m, 6H, CH.sub.3(CH.sub.2).sub.11CH.sub.2O); 1.24-1.43 (m, 40H, CH.sub.3(CH.sub.2).sub.10CH.sub.2CH.sub.2O); 1.68-1.75 (m, 4H, CH.sub.3(CH.sub.2).sub.10CH.sub.2CH.sub.2O); 2.18 (bm, 8H, NCH.sub.2CH.sub.2CH.sub.2NH.sub.2); 2.55 (m, 4H, PCH.sub.2CH.sub.2N); 3.09 (t, 8H, J.sub.vic=7.4, NCH.sub.2CH.sub.2CH.sub.2NH.sub.2); 3.35 (m, 8H, NCH.sub.2CH.sub.2CH.sub.2NH.sub.2); 3.45 (m, 4H, PCH.sub.2CH.sub.2N); 4.09-4.20 (m, 8H, H-3′,4′, CH.sub.3(CH.sub.2).sub.11CH.sub.2O); 4.26 (dd, 1H, J.sub.2′3′=5.2, J.sub.2′,1′=4.0, H-2′); 4.28 (dd, 1H, J.sub.2′,3′=5.1, J.sub.2′,1′=3.9, H-2′); 4.33 (ddd, 1H, J.sub.gem=11.4, J.sub.H,P=7.6, J.sub.5′b,4′=5.3, H-5′b); 4.38 (dd, 2H, J.sub.H,P=7.6, J.sub.5′,4′=4.2, H-5′); 4.43 (ddd, 1H, J=11.4, J.sub.H,P=7.4, J.sub.5′a,4′=2.9, H-5′a); 5.759, 5.761 (2×d, 2×1H, J.sub.5,6=8.1, H-5); 5.83 (d, 1H, J.sub.1′,2′=4.0, H-1′); 5.84 (d, 1H, J.sub.1′,2′=3.9, H-1′); 7.72, 7.73 (2×d, 2×1H, J.sub.6,5=8.1, H-6).

[0071] .sup.13C NMR (125.7 MHz, CD.sub.3OD): 14.44 (CH.sub.3(CH.sub.2).sub.11CH.sub.2O); 20.34 (d, J.sub.C,P=141.8, PCH.sub.2CH.sub.2N); 23.40 (NCH.sub.2CH.sub.2CH.sub.2NH.sub.2); 23.74; 26.58; 30.32; 30.48; 30.68; 30.74; 30.77, 30.78, 30.80 (CH.sub.3(CH.sub.2).sub.10CH.sub.2CH.sub.2O); 31.57, 31.58 (d, J.sub.C,P=6.0, CH.sub.3(CH.sub.2).sub.10CH.sub.2CH.sub.2O); 33.08 (CH.sub.3(CH.sub.2).sub.9CH.sub.2CH.sub.2N); 37.91 (NCH.sub.2CH.sub.2CH.sub.2NH.sub.2); 48.66 (PCH.sub.2CH.sub.2N); 51.17 (NCH.sub.2CH.sub.2CH.sub.2NH.sub.2); 67.41, 67.47 (d, J.sub.C,P=6.3, CH.sub.2-5′); 68.34, 68.55 (d, J.sub.C,P=6.9, CH.sub.3(CH.sub.2).sub.10CH.sub.2CH.sub.2O); 70.87; 70.93 (CH-3′); 74.59; 74.63 (CH-2′); 83.35, 83.36 (d, J.sub.C,P=6.2, CH-4′); 92.40, 92.47 (CH-1′); 103.13, 103.19 (CH-5); 143.03, 143.07 (CH-6); 152.20, 152.28 (C-2); 165.95, 165.96 (C-4).

[0072] .sup.31P{.sup.1H} NMR (202.3 MHz, CD.sub.3OD): 27.57, 28.01.

[0073] IR v.sub.max(KBr) 3424 (s, br), 3047 (br, sh), 2925 (vs), 2854 (s), 2642 (m, br), 2562 (w, br), 2030 (vw, vbr), 1975 (vw, vbr), 1690 (vs), 1465 (m), 1406 (m), 1385 (m), 1266 (m), 1232 (m, sh), 1075 (m, sh), 1054 (m, br), 1035 (m, vbr), 996 (m), 821 (w), 764 (w), 721 (w). HR-ESI C.sub.30H.sub.59O.sub.8N.sub.5P (M+H).sup.+ calculated 648.409583, found 648.409712.

Example 5

Dodecyl-uridine-5′-yl-2-N-bis (3-aminopropyl)-2-aminoethyl phosphonate

[0074] ##STR00007##

[0075] The compound in Example 5 was prepared by the same procedure as the one in Example 1 from boc-N-1-(3-aminopropyl)propane-1,3-diamine (0.53 g, 1.5 mmol) and dodecyl-2′,3′-isopropylidenuridine-5′-yl-vinylphosphonate (prepared according to J. Med. Chem. 2011, 54(22), 7884-7898) (0.55 g, 1 mmol) in 41% yield (0.29 g, 0.41 mmol).

[0076] .sup.1H NMR (600.1 MHz, CD.sub.3OD): 0.90 (m, 6H, CH.sub.3(CH.sub.2).sub.10CH.sub.2O); 1.25-1.42 (m, 36H, CH.sub.3(CH.sub.2).sub.9CH.sub.2CH.sub.2O); 1.71 (m, 4H, CH.sub.3(CH.sub.2).sub.9CH.sub.2CH.sub.2O); 2.22 (bm, 8H, NCH.sub.2CH.sub.2CH.sub.2NH.sub.2); 2.60 (m, 4H, PCH.sub.2CH.sub.2N); 3.11 (t, 8H, J.sub.vic=7.4, NCH.sub.2CH.sub.2CH.sub.2NH.sub.2); 3.39 (m, 8H, NCH.sub.2CH.sub.2CH.sub.2NH.sub.2); 3.48 (m, 4H, PCH.sub.2CH.sub.2N); 4.08-4.22 (m, 8H, H-3′,4′, CH.sub.3(CH.sub.2).sub.10CH.sub.2O); 4.27 (dd, 1H, J.sub.2′,3′=5.5, =4.2, H-2′); 4.29 (dd, 1H, J.sub.2′,3′=5.4, =4.0, H-2′); 4.34 (ddd, 1H, J.sub.gem=11.4, J.sub.H,P=7.5, J.sub.5′b,4′=5.4, H-5′b); 4.39 (dd, 2H, J.sub.H,P=7.5, J.sub.5′4′=4.3, H-5′); 4.44 (ddd, 1H, J=11.4, J.sub.H,P=7.3, J.sub.5′a,4′=2.9, H-5′a); 5.78 (d, 2H, J.sub.5,6=8.1, H-5); 5.84 (d, 1H, J.sub.1′,2′=4.2, H-1′); 5.85 (d, 1H, J.sub.1′,2′=4,0, H-1′); 7.73, 7.74 (2×d, 2×1H, J.sub.6,5=8.1, H-6).

[0077] .sup.13C NMR (150.9 MHz, CD.sub.3OD): 14.43 (CH.sub.3(CH.sub.2).sub.10CH.sub.2O); 20.38, 21.42 (d, J.sub.C,P=140.9, PCH.sub.2CH.sub.2N); 23.28 (NCH.sub.2CH.sub.2CH.sub.2NH.sub.2); 23.71; 26.56; 30.30; 30.46; 30.66; 30.72; 30.74, 30.76 (CH.sub.3(CH.sub.2).sub.9CH.sub.2CH.sub.2O); 31.55, 31.56 (d, J.sub.C,P=5.9, CH.sub.3(CH.sub.2).sub.9CH.sub.2CH.sub.2O); 33.05 (CH.sub.3(CH.sub.2).sub.9CH.sub.2CH.sub.2N); 37.90 (NCH.sub.2CH.sub.2CH.sub.2NH.sub.2); 48.65 (PCH.sub.2CH.sub.2N); 51.13 (NCH.sub.2CH.sub.2CH.sub.2NH.sub.2); 67.35, 67.37 (d, J.sub.C,P=6.2, CH.sub.2-5′); 68.39, 68.57 (d, J.sub.C,P=6.8, CH.sub.3(CH.sub.2).sub.9CH.sub.2CH.sub.2O); 70.83; 70.92 (CH-3′); 74.61; 74.66 (CH-2′); 83.40, 83.42 (d, J.sub.C,P=6.1, CH-4′) 92.13, 92.25 (CH-1′); 103.19, 103.23 (CH-5); 142.98, 143.02 (CH-6); 152.23, 152,28 (C-2); 165.93, 165.94 (C-4).

[0078] .sup.31P{.sup.1H} NMR (202.3 MHz, CD.sub.3OD): 27.60, 28.05.

[0079] IR v.sub.max(KBr) 3391 (s, br), 3000 (vs, vbr), 2925 (vs), 2854 (vs), 2645 (s, br), 2563 (m, br), 2031 (w, br), 1692 (vs, br), 1623 (m), 1575 (w, sh), 1515 (m, br, sh), 1465 (s), 1408 (m), 1385 (m), 1267 (s), 1233 (s, sh), 1077 (s, br, sh), 1058 (s, br, sh), 1036 (s, br), 998 (s, br), 823 (m), 763 (w), 721 (w).

[0080] HR-ESI C.sub.29H.sub.57O.sub.8N.sub.5P (M+H).sup.+ calculated 634.39393, found 634.39398.

Example 6

Pentadecyl-uridine-5′-yl-2-N-bis(3-aminoethyl)-2-aminoethyl phosphonate

[0081] ##STR00008##

[0082] The compound in Example 6 was prepared by the same procedure as the one in Example 1 from boc-N-1-(2-aminoetyl)ethane-1,2-diamine (0.2 g, 0.66 mmol) (prepared according to J. Med. Chem. 2014, 57 (22), 9409-9423) and pentadecyl-2′,3′-isopropylidenuridine-5′-yl-vinylphosphonate (prepared according to J. Med. Chem. 2011, 54(22), 7884-7898) (0.29 g, 0.5 mmol) in 50% yield (0.19 g, 0.25 mmol).

[0083] .sup.1H NMR (500.0 MHz, CD.sub.3OD): 0.90 (m, 6H, CH.sub.3(CH.sub.2).sub.13CH.sub.2O); 1.23-1.44 (m, 48H, CH.sub.3(CH.sub.2).sub.12CH.sub.2CH.sub.2O); 1.70 (m, 4H, CH.sub.3(CH.sub.2).sub.12CH.sub.2CH.sub.2O); 2.16-2.29 (m, 4H, PCH.sub.2CH.sub.2N); 2.83-3.00 (m, 12H, NCH.sub.2CH.sub.2NH.sub.2, PCH.sub.2CH.sub.2N); 3.09-3.17 (bm, 8H, NCH.sub.2CH.sub.2NH.sub.2); 4.06-4.20 (m, 8H, H-3′,4′, CH.sub.3(CH.sub.2).sub.13CH.sub.2O); 4.24-4.43 (m, 6H, H-2′,5′); 5.755 (d, 1H, J.sub.5,6=8.0, H-5); 5.757 (d, 1H, J.sub.5,6=8.0, H-5); 5.808 (d, 1H, J.sub.1′,2′=3.7, H-1′); 5.812 (d, 1H, J.sub.1′,2′=3.9, H-1′); 7.71 (d, 1H, J.sub.6,5=8.0, H-6); 7.72 (d, 1H, J.sub.6,5=8.0, H-6).

[0084] .sup.13C NMR (125.7 MHz, CD.sub.3OD): 14.45 (CH.sub.3(CH.sub.2).sub.13CH.sub.2O-A,B); 22.78 (d, J.sub.C,P=138.3, PCH.sub.2CH.sub.2N); 22.84 (d, J.sub.C,P=138.2, PCH.sub.2CH.sub.2N); 23.74, 26.64, 30.34, 30.48, 30.70, 30.75, 30.77, 30.79; 30.81 (CH.sub.3(CH.sub.2).sub.12CH.sub.2CH.sub.2O); 31.57 (d, J.sub.C,P=6.0, CH.sub.3(CH.sub.2).sub.12CH.sub.2CH.sub.2O); 31.59 (d, J.sub.C,P=6.0, CH.sub.3(CH.sub.2).sub.12CH.sub.2CH.sub.2O); 33.08 (CH.sub.3(CH.sub.2).sub.12CH.sub.2CH.sub.2O); 37.90, 37.95 (NCH.sub.2CH.sub.2NH.sub.2); 47.34 (NCH.sub.2CH.sub.2P); 51.47, 51.53 (NCH.sub.2CH.sub.2NH.sub.2); 66.78 (d, J.sub.C,P=6.3, CH.sub.2-5′); 66.95 (d, J.sub.C,P=6.6, CH.sub.2-5′); 67.89 (d, J.sub.C,P=6.9, CH.sub.3(CH.sub.2).sub.13CH.sub.2O); 67.93 (d, J.sub.C,P=6.9, CH.sub.3(CH.sub.2).sub.13CH.sub.2O); 70.81, 70.86 (CH-3′); 74.54, 74.63 (CH-2′); 83.34 (d, J.sub.C,P=6.2, CH-4′); 83.39 (d, J.sub.C,P=6.2, CH-4′); 92.66, 92.68 (CH-1′); 103.04, 103.05 (CH-5); 143.05, 143.09 (CH-6); 152.17, 152.20 (C-2); 165.99, 165.99 (C-4).

[0085] .sup.31P{.sup.1H} NMR (202.3 MHz, CD.sub.3OD): 33.66.

[0086] v.sub.max(KBr) 3423 (s, vbr), 3018 (s, vbr, sh), 2924 (vs), 2854 (vs), 2650 (m, vbr, sh), 2560 (m, vbr), 2032 (vw, vbr), 1946 (vw, vbr), 1691 (s, br), 1626 (m), 1466 (s), 1406 (m), 1387 (m), 1266 (m), 1237 (m, br, sh), 1074 (m, sh), 1052 (m, sh), 1021 (s, br), 1000 (m, br, sh), 822 (w), 767 (w), 722 (w).

[0087] HR-ESI C.sub.30H.sub.59O.sub.8N.sub.5P (M+H).sup.+ calculated 648.40958, found 648.40969.

Example 7

Tetradecyl-uridine-5′-yl-2-N-bis(3-aminoethyl)-2-aminoethyl phosphonate

[0088] ##STR00009##

[0089] The compound in Example 7 was prepared by the same procedure as the one in Example 1 from boc-N-1-(2-aminoetyetyl)ethane-1,2-diamine (0.4 g, 1.32 mmol) and tetradecyl-2′,3′-isopropylidenuridine-5′-yl-vinylphosphonate (0.63 g, 1.1 mmol) (prepared according to J. Med. Chem. 2011, 54(22), 7884-7898) in 37% yield (0.27 g, 0.41 mmol).

[0090] .sup.1H NMR (500.0 MHz, CD.sub.3OD): 0.90 (m, 6H, CH.sub.3(CH.sub.2).sub.12CH.sub.2O-A,B); 1.13-1.46 (m, 44H, CH.sub.3(CH.sub.2).sub.11CH.sub.2CH.sub.2O-A,B); 1.71 (m, 4H, CH.sub.3(CH.sub.2).sub.11CH.sub.2CH.sub.2O-A,B); 2.28-2.49 (m, 4H, PCH.sub.2CH.sub.2N-A,B); 3.03-3.23 (m, 12H, NCH.sub.2CH.sub.2NH.sub.2-A,B, PCH.sub.2CH.sub.2N-A,B); 3.23-3.33 (bm, 8H, NCH.sub.2CH.sub.2NH.sub.2-A,B); 4.06-4.22 (m, 8H, H-3′,4′-A,B, CH.sub.3(CH.sub.2).sub.13CH.sub.2O-A,B); 4.24-4.44 (m, 6H, H-2′,5′-A,B); 5.76 (d, 2H, J.sub.5,6=8.0, H-5-A,B); 5.83 (d, 2H, J.sub.1′,2′=4.0, H-1′-A,B); 7.73 (d, 1H, J.sub.6,5=8.0, H-6-B); 7.74 (d, 1H, J.sub.6,5=8.0, H-6-A).

[0091] .sup.13C NMR (125.7 MHz, CD.sub.3OD): 14.45 (CH.sub.3(CH.sub.2).sub.12CH.sub.2O-A,B); 22.35 (d, J.sub.C,P=140.4, PCH.sub.2CH.sub.2N—B); 22.39 (d, J.sub.C,P=141.7, PCH.sub.2CH.sub.2N-A); 23.74, 26.62, 30.34, 30.49, 30.69, 30.75, 30.77, 30.79, 30.80, 30.81 (CH.sub.3(CH.sub.2).sub.11CH.sub.2CH.sub.2O-A,B); 31.56 (d, J.sub.C,P=6.0, CH.sub.3(CH.sub.2).sub.11CH.sub.2CH.sub.2O-A); 31.58 (d, J.sub.C,P=6.0, CH.sub.3(CH.sub.2).sub.11CH.sub.2CH.sub.2O—B); 33.08 (CH.sub.3(CH.sub.2).sub.11CH.sub.2CH.sub.2O-A,B); 37.07 (NCH.sub.2CH.sub.2NH.sub.2-A,B); 48.03 (NCH.sub.2CH.sub.2P); 51.48 (NCH.sub.2CH.sub.2NH.sub.2—B); 51.51 (NCH.sub.2CH.sub.2NH.sub.2-A); 66.95 (d, J.sub.C,P=6.5, CH.sub.2-5′-B); 67.05 (d, J.sub.C,P=6.3, CH.sub.2-5′-A); 68.08 (d, J.sub.C,P=7.3, CH.sub.3(CH.sub.2).sub.12CH.sub.2O)—B); 68.15 (d, J.sub.C,P=7.4, CH.sub.3(CH.sub.2).sub.12CH.sub.2O-A); 70.78 (CH-3′-B); 70.86 (CH-3′-A); 74.55 (CH-2′-A); 74.64 (CH-2′-B); 83.36 (d, J.sub.C,P=6.0, CH-4′-B); 83.41 (d, J.sub.C,P=6.2, CH-4′-A); 92.46 (CH-1′-A); 92.51 (CH-1′-B); 103.11 (CH-5-A,B); 143.07 (CH-6-B); 143.09 (CH-6-A); 152.21 (C-2-A); 152.25 (C-2-B); 165.95 (C-4-A); 165.98 (C-4-B).

[0092] .sup.31P{.sup.1H} NMR (202.3 MHz, CD.sub.3OD): 31.69 (A,B).

[0093] IR v.sub.max(KBr) 3427 (s, br), 3000 (s, vbr), 2956 (s), 2924 (vs), 2854 (s), 2560 (m, vbr), 2040 (vw, vbr), 1691 (s), 1466 (m), 1407 (w), 1387 (w), 1267 (m), 1235 (m, br, sh), 1073 (m, sh), 1051 (m, sh), 1018 (m), 1003 (m, sh), 824 (w), 766 (w, sh), 721 (vw).

[0094] HR-ESI C.sub.29H.sub.57O.sub.8N.sub.5P (M+H).sup.+ calculated 634.39393, found 634.39391.

Example 8

Pentadecyl-uridine-5′-yl-2-N-bis(3-guanidinoethyl)-2-aminoethyl phosphonate

[0095] ##STR00010##

[0096] A mixture of 1H-pyrazol-1-carboxamidinuhydrochloride (0.24 g, 1.67 mol), the compound from Example 2 (0.5 g, 0.67 mmol) and dietylisopropylamine (0.57 mlL, 3.35 mmol) in DMF (10 ml) was stirred under argon at rt overnight. The solvent was evaporated and the product was obtained after reverse phase chromatography using a linear gradient of methanol in water (10-100%), evaporation and reprecipitation with ethyl acetate (50 ml) from a solution in 0.5 mol.Math.l.sup.−1 HCl in methanol (20 ml) in 64% yield (0.36 g, 0.43 mmol) as an amorphous solid.

[0097] .sup.1H NMR (600.1 MHz, CD.sub.3OD): 0.90 (m, 6H, CH.sub.3(CH.sub.2).sub.13CH.sub.2O); 1.25-1.43 (m, 48H, CH.sub.3(CH.sub.2).sub.12CH.sub.2CH.sub.2O); 1.68-1.75 (m, 4H, CH.sub.3(CH.sub.2).sub.12CH.sub.2CH.sub.2O); 2.04-2.13 (bm, 8H, NCH.sub.2CH.sub.2CH.sub.2NH); 2.48-2.60 (m, 4H, PCH.sub.2CH.sub.2N); 3.26-3.37 (m, 16H, NCH.sub.2CH.sub.2CH.sub.2NH); 3.40-3.49 (m, 4H, PCH.sub.2CH.sub.2N); 4.10-4.20 (m, 8H, H-3′,4′, CH.sub.3(CH.sub.2).sub.12CH.sub.2O); 4.25-4.29 (m, 2H, H-2′); 4.33 (ddd, 1H, J.sub.gem=11.4, J.sub.H,P=7.5, J.sub.5′b,4′=5.3, H-5′b); 4.38 (dd, 2H, J.sub.H,P=7.5, J.sub.5′,4′=4.2, H-5′); 4.43 (ddd, 1H, J=11.4, J.sub.H,P=7.1, J.sub.5′a,4′=2.9, H-5′a); 5.760, 5.763 (2×d, 2×1H, J.sub.5,6=8.0, H-5); 5.83 (d, 1H, J.sub.1′,2′=4.2, H-1′); 5.84 (d, 1H, J.sub.1′,2′=3.9, H-1′); 7.724, 7.727 (2×d, 2×1H, J.sub.6,5=8.0, H-6).

[0098] .sup.13C NMR (150.9 MHz, CD.sub.3OD): 14.43 (CH.sub.3(CH.sub.2).sub.13CH.sub.2O); 21.36 (d, J.sub.C,P=139.6, PCH.sub.2CH.sub.2N); 23.73 (NCH.sub.2CH.sub.2CH.sub.2NH.sub.2); 24.82, 26.59, 30.30, 30.31, 30.47, 30.68, 30.74, 30.76, 30.78, 30.79 (CH.sub.3(CH.sub.2).sub.12CH.sub.2CH.sub.2O); 31.56 (d, J.sub.C,P=5.9, CH.sub.3(CH.sub.2).sub.12CH.sub.2CH.sub.2O); 33.07 (CH.sub.3(CH.sub.2).sub.12CH.sub.2CH.sub.2N); 39.70 (NCH.sub.2CH.sub.2CH.sub.2NH); 48.40 (d, J.sub.C,P=5.4, PCH.sub.2CH.sub.2N); 51.66 (NCH.sub.2CH.sub.2CH.sub.2NH); 67.35 (d, J.sub.C,P=6.4, CH.sub.2-5′); 67.43 (d, J.sub.C,P=6.3, CH.sub.2-5′); 68.33 (d, J.sub.C,P=6.9, CH.sub.3(CH.sub.2).sub.12CH.sub.2CH.sub.2O); 68.53 (d, J.sub.C,P=6.7, CH.sub.3(CH.sub.2).sub.12CH.sub.2CH.sub.2O); 70.84, 70.88 (CH-3′); 74.62, 74.68 (CH-2′); 83.38, 83.43 (2×d, J.sub.C,P=6.1, CH-4′); 92.37, 92.41 (CH-1′); 103.14, 103.19 (CH-5); 143.02, 143.03 (CH-6); 152.19, 152.25 (C-2); 158.69 (C-guanidine); 165.964, 165.970 (C-4).

[0099] .sup.31P{.sup.1H} NMR (202.3 MHz, CD.sub.3OD): 27.79, 28.23.

[0100] IR v.sub.max(KBr) 3424 (vs, vbr), 3260 (s, br, sh), 3183 (s, br), 2700 (w, vbr), 1694 (s, sh), 1671 (s), 1646 (s, sh), 1624 (s, sh), 1466 (m), 1387 (w), 1268 (m), 1223 (m, br), 1076 (w, sh), 1056 (m), 1036 (m, vbr), 1000 (m), 762 (w, br), 721 (w).

[0101] HR-ESI C.sub.34H.sub.67O.sub.8N.sub.9P (M+H).sup.+ calculated 760.48447, found 760.48452.

Example 9

Tetradecyl-uridine-5′-yl-2-N-bis(3-guanidinoethyl)-2-aminoethyl phosphonate

[0102] ##STR00011##

[0103] The compound in Example 9 was prepared by the same procedure as the compound in Example 8 from 1H-pyrazole-1-carboxamidinehydrochloride (1.8 g, 12.25 mmol), the compound from Example 3 (3 g, 4.1 mmol) and diethylisopropylamine (4, 2 ml, 24.6 mmol) in DMF (40 ml) in 59% yield (2.06 g, 2.41 mmol) as an amorphous solid.

[0104] .sup.1H NMR (500.0 MHz, CD.sub.3OD): 0.90 (m, 6H, CH.sub.3(CH.sub.2).sub.12CH.sub.2O-A,B); 1.25-1.43 (m, 44H, CH.sub.3(CH.sub.2).sub.11CH.sub.2CH.sub.2O-A,B); 1.68-1.75 (m, 4H, CH.sub.3(CH.sub.2).sub.11CH.sub.2CH.sub.2O-A,B); 2.04-2.13 (bm, 8H, NCH.sub.2CH.sub.2CH.sub.2NH-A,B); 2.52-2.64 (m, 4H, PCH.sub.2CH.sub.2N-A,B); 3.32-3.37 (m, 16H, NCH.sub.2CH.sub.2CH.sub.2NH-A,B); 3.42-3.50 (m, 4H, PCH.sub.2CH.sub.2N-A,B); 4.11-4.20 (m, 8H, H-3′,4′-A,B, CH.sub.3(CH.sub.2).sub.12CH.sub.2O-A,B); 4.26 (dd, 1H, J.sub.2′,3′=5.3, J.sub.2′,1′=4.2, H-2′-B); 4.27 (dd, 1H, J.sub.2′,3′=5,2, J.sub.2′,1′=3.9, H-2′-A); 4.33 (ddd, 1H, J.sub.gem=11.4, J.sub.H,P=7.5, J.sub.5b′,4′=5.3, H-5′b-B); 4.38 (dd, 2H, J.sub.H,P=7.5, J.sub.5′,4′=4.2, H-5′-A); 4.43 (ddd, 1H, J=11.4, J.sub.H,P=7.1, J.sub.5′a,4′=2.9, H-5′a-B); 5.77 (d, 2H, J.sub.5,6=8.1, H-5-A,B); 5.84 (d, 1H, J.sub.1′,2′=4.2, H-1′-B); 5.85 (d, 1H, J.sub.1′,2′=3.9, H-1′-A); 7.747 (d, 1H, J.sub.6,5=8.1, H-6-A); 7.752 (d, 1H, J.sub.6,5=8.1, H-6-B).

[0105] .sup.13C NMR (125.7 MHz, CD.sub.3OD): 14.47 (CH.sub.3(CH.sub.2).sub.12CH.sub.2O); 21.32 (d, J.sub.C,P=141.3, PCH.sub.2CH.sub.2N-A,B); 23.76 (NCH.sub.2CH.sub.2CH.sub.2NH.sub.2-A,B); 24.76, 26.60, 30.33, 30.51, 30.71, 30.77, 30.79, 30.81, 30.83, 30.84 (CH.sub.3(CH.sub.2).sub.11CH.sub.2CH.sub.2O-A,B); 31.56 (d, J.sub.C,P=6,1, CH.sub.3(CH.sub.2).sub.11CH.sub.2CH.sub.2O-A,B); 33.09 (CH.sub.3(CH.sub.2).sub.11CH.sub.2CH.sub.2N-A,B); 39.64 (NCH.sub.2CH.sub.2CH.sub.2NH-A,B); 48.38 (PCH.sub.2CH.sub.2N-A,B); 51.56 (NCH.sub.2CH.sub.2CH.sub.2NH-A,B); 67.31 (d, J.sub.C,P=6.5, CH.sub.2-5′-B); 67.37 (d, J.sub.C,P=6.5, CH.sub.2-5′-A); 68.31 (d, J.sub.C,P=6.8, CH.sub.3(CH.sub.2).sub.11CH.sub.2CH.sub.2O-A); 68.51 (d, J.sub.C,P=6.8, CH.sub.3(CH.sub.2).sub.11CH.sub.2CH.sub.2O—B); 70.78 (CH-3′-A); 70.84 (CH-3′-B); 74.63 (CH-2′-B); 74.70 (CH-2′-A); 83.37 (d, J.sub.C,P=5.6, CH-4′-B); 83.42 (d, J.sub.C,P=5.6, CH-4′-B); 92.13 (CH-1′-B); 92.20 (CH-1′-A); 103.13 (CH-5-B); 103.17 (CH-5-A); 143.00 (CH-6-B); 143.03 (CH-6-A); 152.19 (C-2-B); 152.24 (C-2-A); 158.62 (C-guanidine-A,B); 165.99 (C-4-B); 166.00 (C-4-A),

[0106] .sup.31P{.sup.1H} NMR (202.3 MHz, CD.sub.3OD): 27.83 (P-B); 28.28 (P-A).

[0107] IR v.sub.max(KBr) 3320 (s, vbr), 3260 (s, vbr), 3155 (s, vbr), 2925 (s), 2854 (s), 2710 (m, vbr), 2604 (m), 2502 (m, vbr, sh), 1669 (vs, vbr), 1622 (vs, sh), 1465 (s), 1407 (m), 1379 (s), 1265 (s), 1235 (s, br, sh), 1075 (s, br, sh), 1045 (s), 1016 (s, br), 1002 (s, sh), 822 (m), 720 (w), 580 (m, vbr), 490 (m, br, sh).

[0108] HR-ESI C.sub.33H.sub.65O.sub.8N.sub.9P (M+H).sup.+ calculated 746.46882, found 746.46902.

Example 10

Tetradecyl-uridine-5′-yl-(3-aminopyrrolidin-1-N-yl) ethyl phosphonate

[0109] ##STR00012##

[0110] The compound in Example 10 was prepared by the same procedure as the one in Example 1 from 3-boc-3-aminopyrrolidine (0.51 g, 2.75 mmol) and tetradecyl-2′,3′-isopropylidenuridine-5′-yl-vinylphosphonate (1.31 g, 2.3 mmol) (prepared according to J. Med. Chem. 2011, 54(22), 7884-7898) in 26% yield (0.384 g, 0.59 mmol).

[0111] .sup.1H NMR (500.0 MHz, CD.sub.3OD): 0.90 (m, 6H, CH.sub.3(CH.sub.2).sub.12CH.sub.2O-A,B); 1.24-1.43 (m, 44H, CH.sub.3(CH.sub.2).sub.11CH.sub.2CH.sub.2O-A,B); 1.65-1.73 (m, 4H, CH.sub.3(CH.sub.2).sub.11CH.sub.2CH.sub.2O-A,B); 1.94-2.02 (bm, 2H, H-4b-pyrrolidine-A,B); 2.12-2.25 (bm, 6H, H-4b-pyrrolidine-A,B, PCH.sub.2CH.sub.2N-A,B); 2.89-3.02 (bm, 4H, PCH.sub.2CH.sub.2N-A,B); 3.24 (bdd, 2H, J.sub.gem=12.2, J.sub.2b,3=3.5, H-2b-pyrrolidine-A,B); 3.29-3.36 (m, 4H, H-2a,5b-pyrrolidine-A,B); 3.43-3.49 (m, 2H, H-5a-pyrrolidine-A,B); 3.62 (bm, 2H, H-3-pyrrolidine-A,B); 4.04-4.17 (m, 8H, H-3′,4′-A,B, CH.sub.3(CH.sub.2).sub.12CH.sub.2O-A,B); 4.21-4.39 (m, 6H, H-2′,5′-A,B); 5.73 (d, 2H, J.sub.5,6=8.0, H-5-A,B); 5.83 (d, 2H, J.sub.1′,2′=3.9, H-1′-A,B); 7.71 (d, 1H, J.sub.6,5=8.0, H-6-B); 7.72 (d, 1H, J.sub.6,5=8.0, H-6-A).

[0112] .sup.13C NMR (125.7 MHz, CD.sub.3OD): 14.44 (CH.sub.3(CH.sub.2).sub.12CH.sub.2O-A,B); 23.74 (CH.sub.3(CH.sub.2).sub.11CH.sub.2CH.sub.2O-A,B); 26.08 (d, J.sub.C,P=139.9, PCH.sub.2CH.sub.2N-A,B); 26.65, 26.66, 30.29, 30.30, 30,48, 30.67, 30.68, 30,72, 30,73, 30,77, 30.78, 30.79, 30.81 (CH.sub.3(CH.sub.2).sub.11CH.sub.2CH.sub.2O-A,B); 31.03 (CH.sub.2-4-pyrrolidine-A,B); 31.57 (b, J.sub.C,P=6.1, CH.sub.3(CH.sub.2).sub.11CH.sub.2CH.sub.2O-A); 31.58 (d, J.sub.C,P=6.1, CH.sub.3(CH.sub.2).sub.11CH.sub.2CH.sub.2O—B); 33.08 (CH.sub.3(CH.sub.2).sub.11CH.sub.2CH.sub.2O-A,B); 42.13 (d, J.sub.C,P=2.5, NCH.sub.2CH.sub.2P-A,B); 45.29 (CH.sub.2-5-pyrrolidine-A,B); 50.72, 50.79 (CH.sub.2-2-pyrrolidine-A,B); 57.60 (CH-3-pyrrolidine-B); 57.63 (CH-3-pyrrolidine-A); 66.55 (d, J.sub.C,P=6.6, CH.sub.2-5′-A); 66.63 (d, J.sub.C,P=6.5, CH.sub.2-5′-B); 67.68 (d, J.sub.C,P=6.9, CH.sub.3(CH.sub.2).sub.12CH.sub.2O—B); 67.78 (d, J.sub.C,P=6.9, CH.sub.3(CH.sub.2).sub.12CH.sub.2O-A); 70.84 (CH-3′-A,B); 74.83 (CH-2′-A); 74.86 (CH-2′-B); 83.48 (d, J.sub.C,P=6.4, CH-4′-A); 83.51 (d, J.sub.C,P=6.3, CH-4′-B); 92.16 (CH-1′-A); 92.24 (CH-1′-B); 102.97 (CH-5-A,B); 142.75 (CH-6-A); 142.78 (CH-6-B); 152.16 (C-2-A); 152.17 (C-2-B); 165.98 (C-4-A); 165.99 (C-4-B).

[0113] .sup.31P{.sup.1H} NMR (202.3 MHz, CD.sub.3OD): 32.16 (A); 32.36 (B).

[0114] IR v.sub.max(CHCl.sub.3) 3415 (s, vbr), 3051 (s, br), 2924 (vs), 2854 (vs), 2755 (m, vbr, sh), 2455 (w, vbr), 2030 (vw, vbr), 1970 (vw, vbr), 1693 (vs, br), 1464 (s), 1405 (m, sh), 1385 (m), 1261 (s, br), 1224 (m), 1075 (s, sh), 1053 (s), 1036 (s, sh), 1019 (s, sh), 996 (s), 822 (m), 766 (w), 721 (w).

[0115] HR-ESI C.sub.29H.sub.54O.sub.8N.sub.4P (M+H).sup.+ calculated 617.36738, found 617.36742.

[0116] Antibacterial Activity

[0117] Antibacterial activity was measured using a standard microdilution method, showing the minimum inhibitory concentration (MIC) of the test sample which results in inhibition of bacterial growth. Disposable microtiter plates were used for the tests. Samples are dissolved in the brain-heart infusion broth (HiMedia Laboraties Pvt. Ltd., Czech Republic), and Mueller Hinton broth (HiMedia Laboraties, see above) at a final concentration ranging from 200 μg/ml to 1.5625 μg/ml. Plates were inoculated with a standard amount of test bacteria—inoculum density in the hole corresponds to 10.sup.5-6 CFU/ml (colony forming units/ml). MIC values are read after 24/48 hours of incubation at 37° C. as the minimum inhibitory concentration of the test substance at which the growth of bacteria is inhibited. Minimal bactericidal concentration (MBC) is defined as the minimum concentration of the sample needed to achieve irreversible inhibition, therefore killing the bacteria after a defined time of incubation. The MBC was determined by inoculation method. 10 μl from the wells in a microplate with a defined concentration of test substance is taken with an applicator, and inoculated onto the surface of blood agar (Trios, Czech Republic) and Sabouraud agar (Trios, CR). The MBC was determined as the lowest concentration that inhibited visible growth of the bacteria used.

[0118] Standard reference bacterial strains (Escherichia coli CCM 3954, Pseudomonas aeruginosa CCM 3955, Enterococcus faecalis CCM 4224, Staphylococcus aureus CCM 4223) were obtained from the Czech Collection of Microorganisms (CCM) at Masaryk University in Brno. Streptococcus agalactiae, Bacillus subtilis were obtained from the University Hospital Olomouc. The tested microorganisms were maintained in cryobanks (ITEST plus, Czech Republic) at −80° C.

TABLE-US-00001 TABLE 1 Minimum inhibitory concentrations of lipophosphonoxins of the present invention against a panel of reference bacterial strains MIC μg/ml Pseudomonas Enterococcus Staphylococcus Compound Escherichia coli aeruginosa faecalis aureus Streptococcus from example CCM 3954 CCM 3955 CCM 4224 CCM 4223 Bacillus subtilis agalactiae 1 3.125 6.25 50 12.5 0.78 3.125 2 6.25 3.125 50 6.25 1.56 3.125 3 6.25 0.78 25 6.25 0.78 3.125 4 25 3.125 50 12.5 3.125 6.25 5 25 3.125 100 25 3.125 12.5 6 1.56 1.56 12.5 6.25 0.78 3.125 7 12.5 3.125 100 25 6.25 6.25 8 0.78 0.78 25 3.125 0.39 1.56 9 3.125 3.125 12.5 6.25 1.56 3.125 10 3.125 3.125 6.25 12.5 1.56 3.125

TABLE-US-00002 TABLE 2 Minimum inhibitory concentrations of lipophosphonoxins of the present invention against a panel of reference bacterial strains MIC μg/ml Burkholderia Compound Salmonella Acinetobacter Stenotrophomonas multivorans ATCC from example Enteritidis S2-25 baumanii A3-08 matophilia S2-23 BAA-247 2 3.125 6.25 3.125 12.5 3 6.25 25 50 200 4 50 50 50 50 7 12.5 25 6.25 12.5 8 1.56 6.25 3.125 100 9 3.125 12.5 12.5 100

TABLE-US-00003 TABLE 3 Minimum inhibitory concentrations of some of lipophosphonoxins of the present invention against a panel of resistant bacterial strains MIC μg/ml Compound E. coli P. aeruginosa S. aureus S. haemolyticus E. faecium S. epidermidis from example 16702 16575 MRSA 4591 16568 VanA 419ana 8700B 1 3.125 3.125 25 3.125 25 3.125 2 6.25 3.125 12.5 3.125 25 6.25 3 3.125 1.56 6.25 1.56 100 1.56 4 50 25 25 6.25 100 12.5 5 25 12.5 50 12.5 200 12.5 6 1.56 1.56 6.25 1.56 100 1.56 7 6.25 6.25 12.5 3.125 100 3.125 8 0.78 1.56 3.125 1.56 50 1.56 9 3.125 6.25 6.25 3.125 50 3.125 10 3.125 3.125 25 6.25 50 3.125 *Multidrug-resistant bacterial strains isolated from clinical specimens from patients in University Hospital Olomouc: MRSA—methicillin-resistant Staphylococcus aureus 4591, Staphylococcus haemolyticus (a fluoroquinolone-resistant strain) 16568, Enterococcus faecium (vancomycin-resistant strain) VanA 419/ana, Staphylococcus epidermidis (methicilin-resistant strain) 8700/B

[0119] In all cases, the value of the minimum inhibitory concentration (MIC) which is the concentration of test substance in the medium, which inhibited 100% of the growth of the tested bacteria, was equal to the minimum bactericidal concentration (MBC) which is the concentration at which 100% of the the tested bacteria were killed. The MBC value was tested so that the bacteria tested for MIC were inoculated into a medium, which did not contain an inhibitor, and were monitored for growth.

[0120] Benefits of Lipophosphonoxins of the Second Generation:

[0121] Compared to LPPO of the first generation (J. Med. Chem. 2011, 54(22), 7884-7898, CZ PV 2011-312, EP2527351), the LPPO of the second generation show a much broader spectrum of antibacterial activity. Surprisingly, they are mainly effective against clinically important gram-negative bacterial strains and against harmful multiresistant bacterial strains occurring in the hospital environment.

[0122] According to the OECD404 test for skin irritation in rabbits, LPPO, specifically the compound of Example 3, is not an irritant.

[0123] Maximum tolerated dose in mice was very high, for the compound of Example 3 and oral administration the maximum tolerated dose was 1500 mg/kg of bodyweight.

[0124] The mechanism of action of LPPO of the second generation consists in the selective disruption of the bacterial cell membrane.

[0125] LPPO are well soluble in water.

[0126] LPPO exhibit high stability at a wide pH range (1-8).

[0127] Resistance formation against LPPO is very unlikely, since LPPO directly target the cell membrane, which is crucial for the life of the bacteria.

INDUSTRIAL APPLICABILITY

[0128] As antibacterial agents, lipophosphonoxins of this invention can be used as active ingredients of pharmaceutical compositions for the treatment of even very resistant bacterial infections, as ingredients of disinfectants and/or of selective culture media.