Synthesis of monomeric acridine derived nucleic acid intercalators

Article abstract of DOI:10.1515/znb-2005-0113

A series of antiviral compounds consisting of an intercalating acridine derived part, a spacer region and a reactive EDTA-derived conjugate was synthesized in an easy sequence. In the presence of ascorbate a reduction of the phage-titer of MS2 phages by several logarithmic decades was achieved.

Full text of DOI:10.1515/znb-2005-0113

Synthesis of Monomeric Acridine Derived Nucleic Acid Intercalators
Rene´ Csuk, Christian Raschke, Gunnar Go¨the, and Stefan Reißmann
Institut fu¨r Organische Chemie, Martin-Luther-Universita¨t Halle-Wittenberg,
Kurt-Mothes-Str. 2, D-06120 Halle (Saale), Germany
Reprint requests to Prof. Dr. R. Csuk. E-mail: csuk@chemie.uni-halle.de
Z. Naturforsch. 60b, 83 – 88 (2005); received June 9, 2004
A series of antiviral compounds consisting of an intercalating acridine derived part, a spacer region
and a reactive EDTA-derived conjugate was synthesized in an easy sequence. In the presence of
ascorbate a reduction of the phage-titer of MS2 phages by several logarithmic decades was achieved.
Key words: Acridine, Antivirals, Intercalators, Fenton Mechanism
Introduction
The blood supply in industrialized countries is safer
than ever. However, blood is a natural vehicle for the
transmission of infectious agents. In recent years, nu-
merous pathogens have emerged in Europe, the United
States and worldwide with the potential to affect the
safety of the blood supply. Although the movement
of transfusable blood and blood components between
countries is relatively uncommon, infectious agents
can cross international borders, however, through mi-
gration or travel. Finally, variant forms of recognized
pathogens can potentially affect the safety of the blood
supply as well.
To address this target in a more general way, one
may think of disrupting the ability of the genetic
blueprints in DNA and RNA to be expressed and
reproduced. Thus, it seems of interest to synthesize
(small) molecules that prevent viruses, bacteria and
other pathogens from causing infections and also pre-
vent the proliferation of white blood cells which are as-
sociated with a variety of adverse transfusion reactions.
Scheme 1. a) EtOH, H2SO4; b) PLE; c) NMM, isobutyl chlo-
roformate.
order to improve the intercalating ability of such com-
pounds, we planned to access molecules possessing a
substituted acridine-derived intercalating part being at-
tached to an EDTA derived metal complexation region
that should be able to bind Fe(II) and Fe(III).
The major blood components used for transfusion –
platelets, plasma, and red blood cells – do not contain
nuclear DNA or RNA, and therefore retain their bio-
logical utility after an inactivation treatment. This may
be achieved either by UV mediated crosslinking of the
nucleic acid or by damaging the DNA/RNA by the ac-
tion of radicals. The latter may be the result of an in-
tracellular metal-catalyzed Fenton process [1, 2].
Reaction of the amines 1 – 10 [3] with EDTA-
triethylester (11) (that was conveniently prepared even
on a larger preparative scale from the corresponding
tetra-ethylester 12 [4, 5] by selective enzymatic mono-
deesterification [6] using pig liver esterase (PLE))
using the mixed-anhydride method (isobutyl chlo-
roformate/ N-methyl-morpholine N-oxide (NMM))
Quite recently, the first compounds consisting of an furnished the corresponding triethylesters 13 – 22.
intercalator and a reactive centre have been investi- Saponification of the esters was performed by their
gated [1] and they showed some promising results. In treatment with aq. sodium hydroxide to afford the
c
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R. Csuk et al. · Synthesis of Monomeric Acridine Derived Nucleic Acid Intercalators
given in ppm, J in Hz), IR spectra (film or KBr pellet) on a
Perkin-Elmer 298 instrument, MS spectra were taken either
on a MAT311A or a Varian-112S instrument; for elemen-
tal analysis a Foss-Heraeus Vario EL instrument was used.
TLC was performed on silica gel (Merck 5554, detection by
dipping in a solution containing 10% sulfuric acid (400 ml),
ammonium molybdate (20 g) and cerium(IV) sulfate (20 mg)
followed by heating to 150 ◦C. All reactions were performed
under dry argon. Compounds 13 – 22 and 23 – 32 are mem-
bers of homologous series and thus similar in their spectra;
only representative values are therefore given.
The effect of the compounds on bacteriophages was tested
on bacteriophages MS2 grown on its E.coli ATCC15597
host. Log-phase host bacteria for bacteriophage proparga-
tion were grown in tryptic soy broth (TSB, Sigma Chemi-
cals) on an orbital shaker at room temperature until turbid.
Fig. 1. Inactivation of phage MS2 as a function of spacer
length (5 mmol Na ascorbate, 100 µmol of compounds, 3
equiv. Fe3+ loaded, incubation time 4 h, 25 ◦C).
Before use, 100 µl of culture was inoculated into fresh TSB
conta◦ining 0.0025% CaCl2. The cultures were incubated in
a 37 C shaking water bath for 4 h until the log-phase was
reached. Fresh bacteriophages were cultured from freezer
stocks. Log-phase host bateria and phages were mixed at a
multiplicity of infection of approximately 1 in 5 ml TSB.
The culture was kept on ice for 15 min to facilitate adsorp-
tion of the phages to the host cells, then incubated overnight
at 37 ◦C, the phage culture was filtered through a 0.2 µm cel-
lulose acetate syringe filter to remove host bacteria and then
stored at 4 ◦C. Typical yields were 1×1010 PFU/ml. Cultur-
able counts of phage were performed by mixing 100 µl of
phage suspension and 100 µl of host culture in 4 ml molten
TSB top agar (containing 0.7% agar). The top agar was vor-
texed gently, then poured on TSB plates; the plates were in-
target compounds 23 – 32. These target molecules
were treated with a threefold molar excess of Fe3+
,
lyophilized and incubated with the phages in Tris-
buffer in the presence of sodium ascorbate [1].
As shown in Fig. 1, the length of the spacer exhibits
significant effects on the reduction of the phage titer of
MS2 phages. Best results were obtained for a spacer
length of n = 6 giving rise to a reduction of the phage
titer of MS2 phages by > 6 logarithmic decades. Ad-
ditional screening revealed that the inactivation of this
virus depends both on the concentration and tempera-
ture, the time of incubation as well as on the concen-
tration of ascorbate. Increased activity with increased
concentration of ascorbate as well as the observation
that no activity is associated with these compounds in
the absence of ascorbate allows a triggering of the ac-
◦
cubated at 37 C. Bacteriophages from freeze stocks were
diluted into buffer (30 mM Tris, 150 mM KCl, pH 8.3) to a
final population density of approximately 1 × 109 PFU/ml.
Dilutions were prepared in phosphate-buffered saline solu-
tion. Initial as well post-exposure culturable counts were per-
formed in triplicate. The counts were divided by the mean
tivity by the addition of ascorbic acid. In addition, the unexposed control counts to normalize the data and then log-
transformed.
phage was reduced by ◦at least 5 – 6 logarithmic lev-
els (27, 100 µmol, 25 C, 4 h incubation) in plasma,
Ethylenediaminetetraacetic acid triethyl ester (11)
erythrocyte concentrate, in whole blood and in throm-
To a solution of EDTA (15.0 g, 51.3 mmol) in ethanol
(600 ml) conc. sulfuric acid (10 ml) was added and the mix-
ture was heated under reflux for 6 h. After cooling to room
temperature the reaction mixture was neutralized by the care-
ful addition of NaHCO3 and the solvents were removed un-
der reduced pressure. The residue was suspended in water
(200 ml) and extracted with chloroform (2 × 150 ml). The
combined organic phases were dried (Na2SO4), the solvent
was evaporated and EDTA-tetraethyl es◦ter 12 (14.5 g, 70%)
bocyte concentrate. No significant antipathogenic ac-
tivity, however, was found upon incubating the MS2
phages with acridine, Fe(III)-loaded EDTA in the pres-
ence of ascorbate. Additional screening is presently
performed in our laboratories.
Experimental Section
General
The melting points are uncorrected (Reichert hot stage mi- was obtained. M.p. 30 – 32 ◦C (lit.: 34 C [4], 32 C [5]). –
croscope), NMR spectra (internal Me4Si) were recorded us- IR (KBr): ν = 2982s, 2940m, 1736s, 1614w, 1447m, 1421m,
ing either a Bruker AM250 or a Varian XL300 instrument (δ 1368s, 1348s, 1190s, 1030s, 862w, 749m cm−1. – 1H NMR
◦
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85
3
(500 MHz, CDCl3): δ = 4.12 (q, JH,H = 7.10 Hz, 8H, 171.0 (C=O), 170.9 (C=O), 156.0 (q), 150.9 (q), 148.2 (q),
4×CH2), 3.56 (s, 8H, 4×CH2), 2.87 (s, 4H, 2×CH2), 1.23 146.0 (q), 135.0 (q), 130.1 (CH), 127.0 (CH), 125.6 (CH),
(t, 3JH,H = 7.10 Hz, 12H, 4×CH3). – 13C NMR (125 MHz, 124.6 (CH), 123.2 (CH), 116.5 (q), 113.8 (q), 99.8 (CH),
CDCl3): δ = 171.3 (CO), 60.4 (C-2), 55.2 (C-3), 52.3 (C-4), 60.7 (CH2), 60.6 (CH2), 58.5 (CH2(4”)), 55.8 (OMe), 55.6
14.2 (C-1). – MS (ESI, 4.1 kV, 8 µl/min, N2, methanol): (CH2(5”)), 54.6 (CH2(3”)), 52.9 (CH2(1”)), 52.5 (CH2(2”)),
m/z = 405 (19%) [MH]+, 427 (100%) [MNa]+].
52.3 (CH2(1’)), 40.0 (CH2(2’)), 14.2 (CH3). – MS (ESI,
4.1 kV, 8 µl/min, N2, methanol): m/z = 660 (100%)
[M(35Cl)H]+, 661 (34%) [M(37Cl)H]+. – Analysis for
C32H42ClN5O8 (660.17): calcd. C 58.22, H 6.41, N 10.61;
found C 58.01, H 6.55, N 10.42.
To an emulsion of 12 (20.0 g, 49.0 mmol) in water
(1500 ml) containing Na2HPO4 (39.2 g, 276 mmol) and
KH2PO4 (1.16 g, 8.5 mmol) at 27 ◦C PLE (BioChem-
ica, 2.5 ml (3500 units)) were added and the mixture was
stirred for 8 h. After extraction with hexane (2×50 ml, dis-
carded) the aqu. layer was extracted with dichloromethane
(5 × 100 ml) and the combined organic phases were dried
(MgSO4), and the solvent was evaporated to yield 11 (14.5 g,
78%) as a viscous oil [6]. IR (KBr): ν = 2983m, 2938m,
1738s, 1634m, 1378s, 1199s, 1097s, 1028m, 865w, 734w,
588w cm−1. – 1H NMR (500 MHz, CDCl3): δ = 4.15 (m,
3JH,H = 7.10 Hz, 6 H, 3×CH2), 3.53 (s, 4H, 2×CH2), 3.47
(s, 2H, CH2), 3.45 (s, 2H, CH2), 2.84 (s, 4H, CH2), 1.24
Ethyl 2-((2-{[3-(9-{6-chloro-2-methoxyacridinyl}amino)-
propyl]amino}-2-oxoethyl){2-[bis(2-ethoxy-2-oxoethyl)-
amino]ethyl}amino)-acetate (14)
As described for 13 from 2 (1.0 g, 2.6 mmol), 11
(1.16 g, 3.1 mmol) and isobutyl chloroformate (423 mg,
3.10 mmol) 14 (730 mg, 42%) was obtained as an or-
ange coloured viscous oil. RF (methanol/ethyl acetate 1:6)
0.63. – MS (ESI, 4.1 kV, 8 µl/min, N2, methanol): m/z =
674 (100%) [M(35Cl)H]+, 676 (34%) [M(37Cl)H]+. – Anal-
ysis for C33H44ClN5O8 (674.20): calcd. C 58.79, H 6.58,
N 10.39; found C 58.51, H 6.78, N 10.14.
3
(t, JH,H = 7.10 Hz, 9H, 3×CH3). – 13C NMR (125 MHz,
CDCl3): δ = 173.0 (CO), 170.9 (CO), 170.6 (CO), 70.0 (C-
7), 60.8 (C-6), 57.0 (C-5), 56.1 (C-4), 54.6 (C-3), 52.5 (C-
1), 51.7 (C-2), 14.1 (C-8,9). – MS (ESI, 4.1 kV, 8 µl/min,
N2, methanol): m/z = 377 (39%) [MH]+, 399 (100%)
[MNa]+, 415 (28%) [MK]+, 791 (93%) [M2K]+. – HRMS
for C16H29N2O8: calcd. 376.14857; found 376.14859.
Ethyl 2-((2-{[4-(9-{6-chloro-2-methoxyacridinyl}amino)-
butyl]amino}-2-oxoethyl){2-[bis(2-ethoxy-2-oxoethyl)-
amino]ethyl}amino)-acetate (15)
As described for 13 from 2 (1.0 g, 2.5 mmol), 11
(1.12 g, 3.0 mmol) and isobutyl chloroformate (408 mg,
2.99 mmol) 15 (1.35 g, 79%) was obtained as a highly vis-
cous, orange-coloured oil. RF (methanol/ethyl acetate 1:6)
0.63. – MS (ESI, 4.1 kV, 8 µl/min, N2, methanol): m/z =
688 (100%) [M(35Cl)H]+, 690 (45%) [M(37Cl)H]+. – Anal-
ysis for C34H46ClN5O8 (688.23): calcd. C 59.34, H 6.74,
N 10.18; found C 59.13, H 6.94, N 10.03.
Ethyl 2-((2-{[2-(9-{6-chloro-2-methoxyacridinyl}amino)-
ethyl]amino}-2-oxoethyl){2-[bis(2-ethoxy-2-oxoethyl)-
amino]ethyl}amino)-acetate (13)
To an ice cold solution of 11 (410 mg, 1.09 mmol) and
NMM (1 ml) in DMF/ethyl acetate (1:1, 50 ml) isobutyl
chloroformate (147 mg, 1.08 mmol) was added and stir-
ring at this temperature continued for another hour. Then 1
(336 mg, 0.90 mmol) and NMM (2 ml) were added and the
mixture stirred for 12 h at room temperature, the solvents
were removed under diminished pressure and the residue
was subjected to chromatography (silica gel, methanol/ethyl
acetate 1:9) to afford 13 (355 mg, 60%) as a highly vis-
cous orange oil. RF (methanol/ethyl acetate 1:6) 0.63. –
UV/vis (methanol): λmax(logε) = 287 nm (4.64). – IR (film):
ν = 3305m, 2981m, 2361w, 1738s, 1634s, 1608m, 1564s,
1523s, 1440s, 1372m, 1347m, 1241s, 1199s, 1030s cm−1. –
1H NMR (400 MHz, CDCl3): δ = 8.83 (br m, 1H, NH), 8.15
Ethyl 2-((2-{[5-(9-{6-chloro-2-methoxyacridinyl}amino)-
pentyl]amino}-2-oxoethyl){2-[bis(2-ethoxy-2-oxoethyl)-
amino]ethyl amino)-acetate (16)
As described for 13 from 4 (0.8 g, 1.92 mmol), 11
(866 mg, 2.3 mmol) and isobutyl chloroformate (315 mg,
2.31 mmol) 16 (785 mg, 58%) was obtained as a highly vis-
cous, orange-coloured oil. RF (methanol/ethyl acetate 1:6)
0.63. – MS (ESI, 4.1 kV, 8 µl/min, N2, methanol): m/z = 702
(100%) [M(35Cl)H]+, 704 (55%) [M(37Cl)H]+. – Analysis
for C35H48ClN5O8 (702.25): calcd. C 59.86, H 6.89, N 9.97;
found C 59.64, H 7.02, N 9.75.
3
4
(d, JH,H = 9.32 Hz, 1H, 8-H), 8.01 (d, JH,H = 2.07 Hz,
3
1H, 5-H), 7.95 (d, JH,H = 9.32 Hz, 1H, 4-H), 7.43 (d,
4JH,H = 2.59 Hz, 1H, 1-H), 7.35 (dd, JH,H = 9.32 Hz,
3
4JH,H = 2.59 Hz, 1H, 3-H), 7.18 (dd, JH,H = 9.32 Hz,
3
Ethyl 2-((2-{[6-(9-{6-chloro-2-methoxyacridinyl}amino)-
hexyl]amino}-2-oxoethyl){2-[bis(2-ethoxy-2-oxoethyl)-
amino]ethyl)amino)-acetate (17)
4JH,H = 2.07 Hz, 1H, 7-H), 4.08 – 3.99 (m, 8H, 3×CH2),
3.99 (s, 3H, OCH3), CH2(1’)), 3.78 – 3.73 (m, 2H, CH2(2’)),
3.37 (s, 4H, 2×CH2(5”)), 3.31 (s, 2H, CH2(4”)), 3.20 (s, 2H,
CH2(3”)), 2.66 (s, 4H, 2×CH2(1”, 2”)), 1.21 – 1.16 (m, 9H,
As described for 13 from 5 (550 mg, 1.28 mmol), 11
3×CH3). – 13C NMR (125 MHz, CDCl3): δ = 174.3 (C=O), (578 mg, 1.54 mmol) and isobutyl chloroformate (210 mg,
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R. Csuk et al. · Synthesis of Monomeric Acridine Derived Nucleic Acid Intercalators
1.54 mmol) 17 (649 mg, 71%) was obtained as a highly vis- ysis for C38H54ClN5O8 (744.34): calcd. C 61.32, H 7.31,
cous, orange-coloured oil. RF (methanol/ethyl acetate 1:6) N 9.41; found C 61.05, H 7.55, N 9.31.
0.63. – UV/vis (methanol): λmax(logε) = 285 nm (4.66). –
IR (film): ν = 3307m, 2934m, 2858m, 1738s, 1667s, 1633s,
Ethyl 2-((2-{[9-(9-{6-chloro-2-methoxyacridinyl}amino)-
1606m, 1521m, 1466m, 1437m, 1372m, 1238s, 1199s,
nonyl]amino}-2-oxoethyl){2-[bis(2-ethoxy-2-oxoethyl)-
1
1137m, 1030m cm−1. – H NMR (500 MHz, CDCl3): δ =
amino]ethyl}amino)-acetate (20)
8.04 (d, 3JH,H = 9.32 Hz, 1H, 8-H), 8.04 (d, 4JH,H = 2.07 Hz,
As described for 13 from 8 (247 mg, 0.52 mmol), 11
(235 mg, 0.62 mmol) and isobutyl chloroformate (85 mg,
0.62 mmol) 20 (258 mg, 65%) was obtained as a highly vis-
cous, orange-coloured oil. RF (methanol/ethyl acetate 1:6)
0.63. – MS (ESI, 4.1 kV, 8 µl/min, N2, methanol): m/z =
758 (100%) [M(35Cl)H]+, 760 (43%) [M(37Cl)H]+. – Anal-
ysis for C39H56ClN5O8 (758.36): calcd. C 61.77, H 7.44,
N 9.24; found C 61.57, H 7.64, N 8.96.
3
1H, 5-H), 8.00 (br m, 1H, NH), 7.96 (d, JH,H = 9.32 Hz,
3
4
1H, 4-H), 7.37 (dd, JH,H = 9.32 Hz, JH,H = 2.59 Hz,
3
4
1H, 3-H), 7.27 (dd, JH,H = 9.32 Hz, JH,H = 2.07 Hz,
4
1H, 7-H), 7.26 (d, JH,H = 2.59 Hz, 1H, 1-H), 4.12 (q,
3JH,H = 7.10 Hz, 2H, CH2), 4.11 (q, JH,H = 7.10 Hz,
3
4H, 2×CH2), 3.95 (s, 3 H, OCH3), 3.74 – 3.70 (m, 2H,
CH2(1’)), 3.50 (s, 4H, 2×CH2(5”)), 3.39 (s, 2H, CH2(4”)),
3.27 (s, 2H, CH2(3”)), 3.27 – 3.23 (m, 2H, CH2(6’)), 2.82 –
2.74 (m, 4H, 2×CH2(1”,2”)), 1.80 – 1.74 (m, 2H, CH2(2’)),
1.56 – 1.44 (m, 4H, 2×CH2(3’,5’)), 1.40 – 1.35 (m, 2H,
Ethyl 2-((2-{[10-(9-{6-chloro-2-methoxyacridinyl}amino)-
decyl]amino}-2-oxoethyl){2-[bis(2-ethoxy-2-oxoethyl)-
amino]ethyl}amino)-acetate (21)
As described for 13 from 9 (1.0 g, 2.1 mmol), 11
(930 mg, 2.47 mmol) and isobutyl chloroformate (337 mg,
2.47 mmol) 21 (1.21 g, 76%) was obtained as a highly vis-
cous, orange-coloured oil. RF (methanol/ethyl acetate 1:6)
0.63. – MS (ESI, 4.1 kV, 8 µl/min, N2, methanol): m/z =
772 (100%) [M(35Cl)H]+, 774 (70%) [M(37Cl)H]+. – Anal-
ysis for C40H58ClN5O8 (772.39): calcd. C 62.20, H 7.57,
N 9.07; found C 61.96, H 7.69, N 8.86.
3
CH2(4’)), 1.24 – 1.17 (t, JH,H = 7.10 Hz, 9H, 3×CH3). –
13C NMR (125 MHz, CDCl3): δ = 171.33 (C=O),
171.27 (C=O), 170.0 (C=O), 155.9 (q), 150.3 (q), 139.6 (q),
135.2 (q), 131.9 (CH), 125.7 (CH), 124.6 (CH), 124.4 (CH),
124.2 (CH), 117.4 (q), 115.1 (q), 99.7 (CH), 60.7 (CH2),
60.5 (CH2), 58.7 (CH2(4”)), 56.0 (CH2(3”)), 55.6 (OMe),
54.9 (CH2(5”)), 53.0 (CH2(1”)), 52.3 (CH2(2”)), 50.2
(CH2(1’)), 38.6 (CH2(6’)), 31.4 (CH2), 29.5 (CH2), 26.35
(CH2), 26.28 (CH2), 14.2 (CH3). – MS (ESI, 4.1 kV,
8 µl/min, N2, methanol): m/z = 716 (100%) [M(35Cl)H]+,
718 (45%) [M(37Cl)H]+. – Analysis for C36H50ClN5O8
(716.28): calcd. C 60.37, H 7.04, N 9.78; found C 59.92,
H 7.21, N 9.61.
Ethyl 2-((2-{[12-(9-{6-chloro-2-methoxyacridinyl}amino)-
dodecyl]amino}-2-oxoethyl){2-[bis(2-ethoxy-2-oxoethyl)-
amino]ethyl}amino)-acetate (22)
As described for 13 from 10 (0.6 g, 1.17 mmol), 11
(528 mg, 1.4 mmol) and isobutyl chloroformate (192 mg,
1.41 mmol) 22 was obtained as a highly viscous, orange-
coloured oil. RF (methanol/ethyl acetate 1:6) 0.63. – MS
(ESI, 4.1 kV, 8 µl/min, N2, methanol): m/z = 801 (100%)
[M(35Cl)H]+, 803 (74%) [M(37Cl)H]+. – Analysis for
C42H62ClN5O8 (800.44): calcd. C 63.02, H 7.81, N 8.75;
found C 62.94, H 8.02, N 8.49.
Ethyl 2-((2-{[7-(9-{6-chloro-2-methoxyacridinyl}amino)-
heptyl]amino}-2-oxoethyl){2-[bis(2-ethoxy-2-oxoethyl)-
amino]ethyl}amino)-acetate (18)
As described for 13 from 6 (1.0 g, 2.3 mmol), 11
(1.02 g, 2.7 mmol) and isobutyl chloroformate (369 mg,
2.7 mmol) 18 (710 mg, 43%) was obtained as a highly vis-
cous, orange-coloured oil. RF (methanol/ethyl acetate 1:6)
0.63. – MS (ESI, 4.1 kV, 8 µl/min, N2, methanol): m/z =
730 (100%) [M(35Cl)H]+, 732 (43%) [M(37Cl)H]+. – Anal-
ysis for C37H52ClN5O8 (730.31): calcd. C 60.85, H 7.18,
N 9.59; found C 60.65, H 7.34, N 9.34.
2-[{2-[(2-{[2-(9-{6-Chloro-2-methoxyacridinyl}amino)-
ethyl]amino}-2-oxoethyl)(carboxymethyl)amino]ethyl}-
(carboxymethyl)amino] acetic acid (23)
Ethyl 2-((2-{[8-(9-{6-chloro-2-methoxyacridinyl}amino)-
octyl]amino}-2-oxoethyl){2-[bis(2-ethoxy-2-oxoethyl)-
amino]ethyl}amino)-acetate (19)
A solution of 13 (335 mg, 0.51 mmol) in ethanol/water
5:1, 50 ml) containing NaOH (142 mg, 3.55 mmol) was
stirred at room temperature overnight. After neutralization
As described for 13 from 7 (330 mg, 0.72 mmol), 11 with aq. hydrochloric acid (10%), the solvents were removed
(325 mg, 0.86 mmol) and isobutyl chloroformate (118 mg, under reduced pressure 23 (485 mg, 43 wt-% NaCl by anal-
0.86 mmol) 19 (298 mg, 56%) was obtained as a highly vis- ysis) was obtained as a red coloured amorphous solid. An
cous, orange-coloured oil. RF (methanol/ethyl acetate 1:6) analytically pure sample was obtained after dialysis fol-
0.63. – MS (ESI, 4.1 kV, 8 µl/min, N2, methanol): m/z = lowed by repeated chromatography (RP18, methanol/water
744 (100%) [M(35Cl)H]+, 746 (63%) [M(37Cl)H]+. – Anal- and acetonitrile/water). UV/vis (methanol): λmax(logε) =
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R. Csuk et al. · Synthesis of Monomeric Acridine Derived Nucleic Acid Intercalators
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286 nm (4.45). – IR (KBr): ν = 3324s, 1582s, 1526m, phous solid. An analytically pure sample was obtained af-
1403s, 1338m, 1256m, 1172w, 1114w, 1027w cm−1. – ter dialysis followed by repeated chromatography (RP18,
3
1H NMR (500 MHz, D2O / CD3OD): δ = 7.53 (d, JH,H
=
methanol/water and acetonitrile/water). – MS (ESI, 4.1 kV,
9.31 Hz, 1H, 8-H), 7.27 (d, JH,H = 9.31 Hz, 1H, 4-H), 8 µl/min, N2, methanol): m/z = 616 (100%) [M(35Cl)-H]−,
3
3
4
7.22 (s, 1H, 5-H), 7.02 (dd, JH,H = 9.31 Hz, JH,H
=
=
618 (55%) [M(37Cl)-H]−. – Analysis for C29H36ClN5O8
(618.09): calcd. C 56.36, H 5.87, N 11.33; found C 56.09,
3
4
2.59 Hz, 1H, 3-H), 6.88 (dd, JH,H = 9.31 Hz, JH,H
2.07 Hz, 1H, 7-H), 6.64 (s, 1H, 1-H), 3.68 (s, 3H, OCH3), H 5.99, N 11.06.
3.56 – 3.53 (m, 2H, CH2(1’)), 3.29 – 3.24 (m, 2H, CH2(2’)),
2-[{2-[(2-{[6-(9-{6-Chloro-2-methoxyacridinyl}amino)-
2.80 (s, 4H, 2×CH2(5”)), 2.72 (s, 2H, CH2(4”)), 2.70
(s, 2H, CH2(4”)), 2.22 – 2.16 (m, 2H, CH2(1”)), 2.10 –
2.04 (m, 2H, CH2(1”)). – 13C NMR (100 MHz, CD3OD):
δ = 178.6 (C=O), 178.3 (C=O), 174.3 (C=O), 154.6 (q),
150.5 (q), 145.9 (q), 143.4 (q), 135.4 (q), 127.9 (CH),
125.1 (CH), 124.39 (CH), 124.38 (CH), 123.4 (CH),
115.7 (q), 113.2 (q), 99.9 (CH), 58.73 (CH2(5”)), 58.72
(CH2(4”)), 58.5 (CH2(3”)), 55.6 (OMe), 52.0 (CH2(1”)),
51.8 (CH2(2”)), 48.6 (CH2(1’)), 39.9 (CH2(2’)). – MS
(ESI, 4.1 kV, 8 µl/min, N2, methanol): m/z = 574 (60%)
[M(35Cl)-H]−, 576 (100%) [M(37Cl)-H]−. – Analysis for
C26H30ClN5O8 (576.01): calcd. C 54.21, H 5.25, N 12.16;
found C 54.01, H 5.50, N 12.00.
hexyl]amino}-2-oxoethyl)(carboxymethyl)amino]ethyl}-
(carboxymethyl)amino] acetic acid (27)
As described for 23, from 17 (595 mg, 0.83 mmol)
27 (838 mg, 40 wt-% NaCl) was obtained as a red,
amorphous solid. An analytically pure sample was ob-
tained after dialysis followed by repeated chromatogra-
phy (RP18, methanol/water and acetonitrile/water). UV/vis
(methanol): λmax(logε) = 287 nm (4.56). – IR (KBr): ν =
3424s, 2932m, 1589s, 1501m, 1404s, 1326m, 1245s, 1178w,
1120m, 1092m, 1031w cm−1. – 1H NMR (500 MHz,
CD3OD): δ = 7.27 (d, 3JH,H = 9.31 Hz, 1H, 8-H), 7.03 (d,
4
3JH,H = 9.31 Hz, 1H, 4-H), 6.96 (d, JH,H = 2.07 Hz, 1H,
3
4
5-H), 6.94 (dd, JH,H = 9.31 Hz, JH,H = 2.59 Hz, 1H, 3-
2-[{2-[(2-{[3-(9-{6-Chloro-2-methoxyacridinyl}amino)-
propyl]amino}-2-oxoethyl)(carboxymethyl)amino]ethyl}-
(carboxymethyl)amino] acetic acid (24)
H), 6.76 (dd, JH,H = 9.31 Hz, 4JH,H = 2.07 Hz, 1H, 7-H),
6.37 (d, JH,H = 2.59 Hz, 1H, 1-H), 3.55 (s, 3H, OCH3),
3
4
3.31 (br s, 4H, 2×CH2(5”)), 3.14 (s, 2H, CH2(4”)), 3.12 –
3.07 (m, 2H, CH2(1’)), 3.07 – 3.02 (m, 2H, CH2(6’)), 3.05
(s, 2H, CH2(3”)), 2.85 – 2.77 (m, 2H, CH2(1”)), 2.72 – 2.66
(m, 2H, CH2(2”)), 1.41 – 1.31 (m, 4H, 2 x CH2(2’,5’)), 1.15 –
1.07 (m, 4H, 2×CH2(3’,4’)). – 13C NMR (125 MHz, D2O):
δ = 178.8 (C=O), 178.7 (C=O), 173.5 (C=O), 154.0 (q),
151.0 (q), 142.7 (q), 139.5 (q), 136.6 (q), 124.9 (CH),
124.6 (CH), 124.5 (CH), 123.0 (CH), 121.4 (CH), 113.3 (q),
110.7 (q), 100.4 (CH), 58.6 (CH2(5”)), 58.3 (CH2(4”)), 57.7
(CH2(3”)), 55.3 (OMe), 52.1 (CH2(1”)), 51.4 (CH2(2”)),
48.2 (CH2(1’)), 38.9 (CH2(6’)), 29.8 (CH2), 28.2 (CH2),
25.7 (CH2), 25.6 (CH2). – MS (ESI, 4.1 kV, 8 µl/min, N2,
methanol): m/z = 630 (45%) [M(35Cl)-H]−, 632 (100%)
[M(37Cl)-H]−. – Analysis for C30H38ClN5O8 (632.12):
calcd. C 57.00, H 6.06, N 11.08; found C 56.77, H 6.29,
N 10.87.
As described for 23, from 14 (722 mg, 1.07) 24 (1.04 g,
42 wt-% NaCl) was obtained as a red, amorphous solid.
An analytically pure sample was obtained after dialysis fol-
lowed by repeated chromatography (RP18, methanol/water
and acetonitrile/water). – MS (ESI, 4.1 kV, 8 µl/min, N2,
methanol): m/z = 588 (100%) [M(35Cl)-H]−, 590 (30%)
[M(37Cl)-H]−. – Analysis for C27H32ClN5O8 (590.04):
calcd. C 54.96, H 5.47, N 11.87; found C 54.74, H 5.61,
N 11.62.
2-[{2-[(2-{[4-(9-{6-Chloro-2-methoxyacridinyl}amino)-
butyl]amino}-2-oxoethyl)(carboxymethyl)amino]ethyl}-
(carboxymethyl)amino] acetic acid (25)
As described for 23, from 15 (940 mg, 1.37 mmol)
25 (1.34 g, 42 wt-% NaCl) was obtained as a red, amor-
phous solid. An analytically pure sample was obtained af-
ter dialysis followed by repeated chromatography (RP18,
methanol/water and acetonitrile/water). – MS (ESI, 4.1 kV,
8 µl/min, N2, methanol): m/z = 602 (100%) [M(35Cl)-H]−,
604 (28%) [M(37Cl)-H]−. – Analysis for C28H34ClN5O8
(604.06): calcd. C 55.68, H 5.67, N 11.59; found C 55.41,
H 5.89, N 11.32.
2-[{2-[(2-{[7-(9-{6-Chloro-2-methoxyacridinyl}amino)-
heptyl]amino}-2-oxoethyl)(carboxymethyl)amino]ethyl}-
(carboxymethyl)amino] acetic acid (28)
As described for 23, from 18 (698 mg, 0.96 mmol)
28 (1.03 g, 43 wt-% NaCl) was obtained as a red, amor-
phous solid. An analytically pure sample was obtained af-
ter dialysis followed by repeated chromatography (RP18,
methanol/water and acetonitrile/water). – MS (ESI, 4.1 kV,
8 µl/min, N2, methanol): m/z = 644 (60%) [M(35Cl)-H]−,
646 (100%) [M(37Cl)-H]−. – Analysis for C31H40ClN5O8
2-[{2-[(2-{[5-(9-{6-Chloro-2-methoxyacridinyl}amino)-
pentyl]amino}-2-oxoethyl)(carboxymethyl)amino]ethyl}-
(carboxymethyl)amino acetic acid (26)
As described for 23 from 16 (773 mg, 1.10 mmol) 26 (646.15): calcd. C 57.63, H 6.24, N 10.84; found C 57.41,
(1.10 g, 41 wt-% NaCl) was obtained as a red, amor- H 6.39, N 10.63.
Unauthenticated
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88
R. Csuk et al. · Synthesis of Monomeric Acridine Derived Nucleic Acid Intercalators
2-[{2-[(2-{[8-(9-{6-Chloro-2-methoxyacridinyl}amino)-
octyl]amino-2-oxoethyl)(carboxymethyl)amino]ethyl}-
(carboxymethyl)amino] acetic acid (29)
phous solid. An analytically pure sample was obtained af-
ter dialysis followed by repeated chromatography (RP18,
methanol/water and acetonitrile/water). – MS (ESI, 4.1 kV,
8 µl/min, N2, methanol): m/z = 686 (100%) [M(35Cl)-H]−,
688 (15%) [M(37Cl)-H]−. – Analysis for C34H46ClN5O8
(688.22): calcd. C 59.34, H 6.74, N 10.18; found C 59.05,
H 6.92, N 9.95.
As described for 23, from 19 (252 mg, 0.34 mmol) 29
(351 mg, 40 wt-% NaCl) was obtained as a red, amor-
phous solid. An analytically pure sample was obtained af-
ter dialysis followed by repeated chromatography (RP18,
methanol/water and acetonitrile/water). – MS (ESI, 4.1 kV,
8 µl/min, N2, methanol): m/z = 658 (100%) [M(35Cl)-H]−,
660 (60%) [M(37Cl)-H]−. – Analysis for C32H42ClN5O8
(660.17): calcd. C 58.22, H 6.41, N 10.61; found C 57.99,
H 6.70, N 10.38.
2-[{2-[(2-{[12-(9-{6-Chloro-2-methoxyacridinyl}amino)-
dodecyl]amino}-2-oxoethyl)(carboxymethyl)amino]ethyl}-
(carboxymethyl)amino] acetic acid (32)
As described for 23 from 22 (411 mg, 0.51 mmol) 32
(560 mg, 38 wt-% NaCl) was obtained as a red, amor-
phous solid. An analytically pure sample was obtained af-
ter dialysis followed by repeated chromatography (RP18,
methanol/water and acetonitrile/water). – MS (ESI, 4.1 kV,
8 µl/min, N2, methanol): m/z = 714 (100%) [M(35Cl)-H]−,
716 (30%) [M(37Cl)-H]−. – Analysis for C36H50ClN5O8
(716.28): calcd. C 60.37, H 7.04, N 9.78; found C 60.11,
H 7.28, N 9.59.
2-[{2-[(2-{[9-(9-{6-Chloro-2-methoxyacridinyl}amino)-
nonyl]amino}-2-oxoethyl)(carboxymethyl)amino]ethyl}-
(carboxymethyl)amino] acetic acid (30)
As described for 23, from 20 (206 mg, 0.27 mmol) 30
(285 mg, 39 wt-% NaCl) was obtained as a red, amor-
phous solid. An analytically pure sample was obtained af-
ter dialysis followed by repeated chromatography (RP18,
methanol/water and acetonitrile/water). – MS (ESI, 4.1 kV,
8 µl/min, N2, methanol): m/z = 672 (90%) [M(35Cl)-H]−,
674 (100%) [M(37Cl)-H]−. – Analysis for C33H44ClN5O8
(674.20): calcd. C 58.79, H 6.58, N 10.39; found C 58.51,
H 6.81, N 10.07.
Acknowledgements
Thanks for helpful discussions and parts of the biologi-
cal screening are due to Dr. H. Knoller and Dr. H.-J. Neu-
mann (Fresenius Hemocare); additional screening has been
performed by Bioscreen Ltd. We like to thank Dr. R. Kluge
for the ESI-MS spectra and Dr. D. Stro¨hl for taking the NMR
spectra.
2-[{2-[(2-{[10-(9-{6-Chloro-2-methoxyacridinyl}amino)-
decyl]amino}-2-oxoethyl)(carboxymethyl)amino]ethyl}-
(carboxymethyl)amino] acetic acid (31)
As described for 23, from 21 (669 mg, 0.87 mmol) 31
(922 mg, 39 wt-% NaCl) was obtained as a red, amor-
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Unauthenticated
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