Abstract: The missing linking: BCNU is a chemotherapy drug that generates an ethylene bridge between N(1) of deoxyguanosine and N(3) of deoxycytidine. No synthesis of a DNA containing this moiety has been reported until now. A new strategy using a photocaged nucleobase that, when released, generates a highly reactive intermediate which cross-links the opposing DNA strand in a manner analogous to BCNU (see scheme, NBOC=ortho-nitrobenzyloxycarbonyl).
Abstract: Guanidino-modified phthalocyanines are evaluated in vitro (polymerase-stop assays and FRET) and in cultured cells as G4-DNA ligands and modulators of gene transcription.
Abstract: A new family of G-quadruplex ligands termed "amido phthalocyanines" (APcs) was synthesized by reacting variable amino acids with tetraamino zinc phthalocyanine. Variation in the number of methylene units separating the APc scaffold from terminal ammonium groups systematically modulated ammonium pK(a) values that, in turn, mediated APc aggregation and DNA binding. Certain APcs exhibited nearly 1000-fold enhancements in fluorescence quantum yield upon binding G-quadruplex DNA under physiological conditions of pH and ionic strength. G-quadruplexes derived from the c-myc and c-kit promoters and the human telomeric repeat were evaluated for APc affinity and specificity using two complementary and direct fluorescence binding assays that revealed apparent dissociation constants ranging from 20 to 200 nM. Approximately 500-fold lower affinities for duplex and single-stranded DNAs were observed. Interestingly, APc-quadruplex binding was relatively insensitive to ionic strength (0.03-1 M KCl) but highly dependent on the pH of the solution. Our results provide a mechanism for the "turn-on" fluorescence properties exhibited by these compounds that will assist in future rational design of new G-quadruplex-specific fluorescent probes and drug candidates.
Abstract: A new family of cationic phthalocyanines containing four guanidinium groups was synthesized in pyridine-HCl at 120 degrees C; under these conditions zinc was removed from both the starting materials and products to reveal a new synthetic route to metal-free phthalocyanines.
Abstract: EIN -Fluoreszenz und AUS -Transkriptionsregulierung treten auf, wenn das Guanidinium-modifizierte Zinkphthalocyanin Zn-DIGP an eine G-Quadruplex-DNA des c-Myc-Promotors bindet (siehe Bild). Die Gleichgewichtsdissoziationskonstante Kd ist kleiner 2 nM, was diese Wechselwirkung zur stärksten bis dato beschriebenen Bindungsinteraktion einer G-Quadruplexstruktur mit einem kleinen Molekül macht.
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Abstract: The invention relates to porphyrins, porphyrazines and phthalocyanines carrying a guanidine substituent. These compounds are obtained by treating a porphyrin, porphyrazine or phthalocyanine, respectively, carrying an amino substituent with cyanamide or a carbodiimide in an ionic liquid at elevated temperature. Described are valuable diagnostic, photoelectronic, and therapeutic uses, e.g. as photosensitizing agents in photodynamic therapy, fluorescent and magnetic probes for diagnosis, or to transport other molecules through the plasma membranes of living cells. Furthermore, the present invention relates to a method of removing a metal from metalloporphyrins, metalloporphyrazines and metallophthalocyanines by heating the zinc(II)-containing compounds in an ionic liquid and a proton source at temperatures between 60°C and 200°C
Abstract: DNA interstrand crosslinks (ICLs) are highly cytotoxic lesions formed by a variety of important anti-tumor agents. Despite the clinical importance of ICLs, the mechanisms by which these lesions are repaired in mammalian cells have so far remained elusive. One of the obstacles in the study of ICL repair has been the limited availability of suitable methods for the synthesis of defined site-specific ICLs. We report here the synthesis of a site-specific ICL containing an ethylene-bridged G-T base pair based on the incorporation of a crosslink precursor containing a selectively reactive group on one strand using solid-phase synthesis. 3-(2-chloroethyl)thymidine was incorporated into oligonucleotides and underwent ICL formation upon annealing to a complementary strand by reacting with the base opposite to the modified T residue. A strong preference for ICL formation with a G residue opposite the reactive T was observed. Detailed characterization of the reaction product revealed that the alkylation reaction occurred with the O-6 group of G and a mechanism accounting for this preference is proposed. These G-T crosslinks introduced here will be useful for studies of ICL repair.
Abstract: Aromatic amino and nitro compounds are potent carcinogens found in the environment that exert their toxic effects by reacting with DNA following metabolic activation. One important adduct is N-(deoxyguanosin-8-yl)-2-acetylaminofluorene (dG-AAF), which has been extensively used in studies of the mechanisms of DNA repair and mutagenesis. Despite the importance of dG-AAF adducts in DNA, an efficient method for its incorporation into DNA using solid-phase synthesis is still missing. We report the development of a modified 'ultra-mild' DNA synthesis protocol that allows the incorporation of dG-AAF into oligonucleotides of any length accessible by solid-phase DNA synthesis with high efficiency and independent of sequence context. Key to this endeavor was the development of improved deprotection conditions (10% diisopropylamine in methanol supplemented with 0.25 M of beta-mercaptoethanol) designed to remove protecting groups of commercially available 'ultra-mild' phosphoramidite building blocks without compromising the integrity of the exquisitely base-labile acetyl group at N8 of dG-AAF. We demonstrate the suitability of these oligonucleotides in the nucleotide excision repair reaction. Our synthetic approach should facilitate comprehensive studies of the mechanisms of repair and mutagenesis induced by dG-AAF adducts in DNA and should be of general use for the incorporation of base-labile functionalities into DNA.
Abstract: The Me3SiC(1) bond of the bis-(trimethylsilyl)ethynylated anhydroalditol 2 is selectively cleaved with BuLi to yield 3/4, while AgNO2/KCN in MeOH cleaves the Me3SiC(2) bond, leading to 5 (Scheme 1). Both Me3Si groups are removed with NaOH in MeOH ( 7), the (i-Pr)3Si group is selectively cleaved with HCl in aq. MeOH ( 6); all silyl substituents are removed with Bu4NF ( 8). Acetolysis transformed 9 into 13, which was desilylated to 14, while thiolysis of 9 led to a mixture 11/12. The tetraacetate 14 has also been obtained from 9 via 10. Oxidative dimerisation of either 3 or 5, or of a mixture 3/5 yields only the homodimers 15 and 16 (Scheme 2); treatment of 16 with AgNO2/KCN yielded 17, deprotection proceeding much more slowly than the cleavage of the Me3SiC(2) group of 2. The iodoalkyne 20, required for the cross-coupling with 5 according to Cadiot-Chodkiewicz, was prepared by deprotection of 3/4 to 18, methoxymethylation (19), and iodination. Cross-coupling yielded mostly 21, besides the homodimer 22. Similarly, cross-coupling of 20 and 23 (obtained from 5) led to 24 and 22. The structure of 24 was established by X-ray analysis (Fig.), showing a C(6)-C(5) distance of 5.2 Ã…. The conditions for deprotecting 2 were applied to 21, and led to 25 (AgNO2/KCN), 26 (aq. NaOH), 27 (Bu4NF), and 29 (HCl/MeOH; Scheme 3). Attempted deprotection of the propargylic-ether moiety with BuLi, however, failed. The dimer 27 was further deprotected to 28. Acetolytic (Ac2O/Me3SiOTf) debenzylation of the dimer 30, obtained from 10, gave 31 (83%) which was deacetylated to 32 (Scheme 4). Cross-coupling of 5 and the bromoalkyne 33, obtained from 10, yielded 34; again, acetolysis proceeded well, leading to 35. The cellobiose derivative 38 was prepared from the lactone 36 via 37. The glycosidic linkage of 38 proved resistant to the conditions of acetolysis, leading to 39. Acetolysis of the benzylated thiophene 40 (from 30 with Na2S) yielded the octaacetate 41, but proceeded in substantially lower yields (50%).
Abstract: NaSMe in toluene leads to regioselective de-C-silylation of the bis[(trimethylsilyl)ethynyl]saccharide 2, but to decomposition of butadiynes such as 1 or 12. We have, therefore, combined the known reagent-controlled, regioselective desilylation of 2 and of 12 (AgNO2/KCN) with a substrate-controlled regioselective de-C-silylation, based on C-silyl groups of different size. This combination was studied with the fully protected 3 which was mono-desilylated to 4 or to 5 (Scheme 1). Triethylsilylation of 5 ( 6) was followed by removal of the Me3Si group ( 7), introduction of a (t-Bu)Me2Si group ( 8) and removal of the Et3Si group yielded 9; these high-yielding transformations proceed with a high degree of selectivity. Iodination of 4 gave 10. The latter was coupled with 5 to the homodimer 11 and the heterodimer 12, which was desilylated to 13. The second building block for the tetramer was obtained by coupling 14 (from 7) with 5, leading to 15 and 16. Removal of the Me3Si group ( 17) and iodination led to 18 which was coupled with 13 to the homotetramer 20 and the heterotetramer 19 (Scheme 2). Deprotection of 19 gave 21, which was, on the one hand, iodinated to 22, and, on the other hand, protected by the (t-Bu)Me2Si group ( 23). Removal of the Et3Si group ( 24) and coupling afforded the homooctamer 26 and the heterooctamer 25. Yields of iodination, silylation, and desilylation were consistently high, while heterocoupling proceeded in only 50-55%. Cleavage of the (i-Pr)3SiC and MeOCH2O groups of 11 ( 27), 15 ( 28), 20 ( 29) and 26 ( 30) proceeded in high yields (Scheme 3). Complete deprotection in two steps of the heterocoupling products 16 ( 31 32), 19 ( 33 34), and 25 ( 35 36) gave the unprotected dimer 32, tetramer 34, and octamer 36 in high yields (Scheme 4). Only the dimer 32 is soluble in H2O; the 1H-NMR spectra of 32, 34, and 36 in (D6)DMSO (relatively low concentration) show no signs of association.
Abstract: It is proposed to study the influence of interresidue H-bonds on the structure and properties of polysaccharides by comparing them to a series of systematically modified oligosaccharide analogues where some or all of the glycosidic O-atoms are replaced by buta-1,3-diyne-1,4-diyl groups. This group is long enough to interrupt the interresidue H-bonds, is chemically versatile, and allows a binomial synthesis. Several approaches to the simplest monomeric unit required to make analogues of cellulose are described. In the first approach, allyl -D-galactopyranoside (1) was transformed via 2 and the tribenzyl ether 3 into the triflate 4 (Scheme 2). Substitution by cyanide ( 5-7) followed by reduction with DIBAH led in high yield to the aldehyde 9, which was transformed into the dibromoalkene 10 and the alkyne 11 following the Corey-Fuchs procedure (Scheme 3). The alkyne was deprotected via 12 or directly to the hemiacetal 13. Oxidation to the lactone 14, followed by addition of lithium (trimethylsilyl)acetylide Me3SiCCLi/CeCl3 ( 15) and reductive dehydroxylation afforded the disilylated dialkyne 16. The large excess of Pd catalyst required for the transformation 11 13 was avoided by deallylating the dibromoalkene 10 ( 17 18), followed by oxidation to the lactone 19, addition of Me3SiCCLi to the anomeric hemiketals 20 (-D/-D 7:2), dehydroxylation to 21, and elimination to the monosilylated dialkyne 22 (Scheme 3). In an alternative approach, treatment of the epoxide 24 (from 23) with Me3SiCCLi/Et2AlCl according to a known procedure gave not only the alkyne 27 but also 25, resulting from participation of the MeOCH2O group (Scheme 4). Using Me3Al instead of Et2AlCl increased the yield and selectivity. Deprotection of 27 ( 28), dibenzylation ( 29), and acetolysis led to the diacetate 30 which was partially deacetylated ( 31) and oxidized to the lactone 32. Addition of Me3SiCCLi/TiCl4 afforded the anomeric hemiketals 33 (-D/-D 3:2) which were deoxygenated to the dialkyne 34. This synthesis of target monomers was shortened by treating the hydroxy acetal 36 (from 27) with (Me3SiCC)3Al (Scheme 5): formation of the alkyne 37 (70%) by fully retentive alkynylating acetal cleavage is rationalised by postulating a participation of HOC(3). The sequence was further improved by substituting the MeOCH2O by the (i-Pr)3SiO group (Scheme 6); the epoxide 38 (from 23); yielded 85% of the alkyne 39 which was transformed, on the one hand, via 40 into the dibenzyl ether 29, and, on the other hand, after C-desilylation ( 41) into the dialkyne 42. Finally, combined alkynylating opening of the oxirane and the 1,3-dioxolane rings of 38 with excess Et2Al CCSiMe3 led directly to the monomer 43 which is thus available in two steps and 77% yield from 23 (Scheme 6).
Abstract: Phenyl beta-methoxyacrylates, linked to an aromatic ring via an olefinic bridge, have been identified as novel, potentially inexpensive, antimalarial agents. The compounds are believed to exert their activity by inhibition of mitochondrial electron transport at the cytochrome bc(1) complex. A series of compounds have been synthesized to define structure-activity relationships affecting antimalarial activity. It was found that the beta-methoxyacrylate was required ortho to the linker and the optimal bridge was (E,E)-butadiene. Compounds in which the second aromatic ring was ortho-substituted or ortho,para-disubstituted gave optimal potency. Several compounds were identified with potency that is superior to that of chloroquine both in culture and in a murine malaria model.
Abstract: The invention relates to the use of $g(b)-alkoxyacrylates of formulae (IA) or (IB) wherein R, R?1¿, R?6¿, R?7¿, R?8¿, X and Z are as defined herein, as therapeutically active substances, especially in the treatment or prophylaxis of malaria and to medicaments containing these substances.
Abstract: Highly regioselective (>90%) MOM-protection of 3-hydroxy-5-phenyl-isoxazole, followed by elaboration in 4-position via directed ortho-metalation and mild deprotection with cold methanolic HCl provided ready access to a series of zwitterionic isoxazole derivatives.
Abstract: Aromatic amino and nitro compounds are potent carcinogens found in the environment that exert their toxic effects by reacting with DNA following metabolic activation. One important adduct is N-(deoxyguanosin-8-yl)-2-acetylaminofluorene (dG-AAF), which has been extensively used in studies of the mechanisms of DNA repair and mutagenesis. Despite the importance of dG-AAF adducts in DNA, an efficient method for its incorporation into DNA using solid-phase synthesis is still missing. We report the development of a modified 'ultra-mild' DNA synthesis protocol that allows the incorporation of dG-AAF into oligonucleotides of any length accessible by solid-phase DNA synthesis with high efficiency and independent of sequence context. Key to this endeavor was the development of improved deprotection conditions (10% diisopropylamine in methanol supplemented with 0.25 M of beta-mercaptoethanol) designed to remove protecting groups of commercially available 'ultra-mild' phosphoramidite building blocks without compromising the integrity of the exquisitely base-labile acetyl group at N8 of dG-AAF. We demonstrate the suitability of these oligonucleotides in the nucleotide excision repair reaction. Our synthetic approach should facilitate comprehensive studies of the mechanisms of repair and mutagenesis induced by dG-AAF adducts in DNA and should be of general use for the incorporation of base-labile functionalities into DNA.
Abstract: It is proposed to study the influence of intra- and interresidue H-bonds
on the structure and properties of polysaccharides by comparing them to a
series of systematically modified oligosaccharide analogues where some or
all of the O-C(l) units are replaced by a buta-l,3-diyne-l,4-diyl group.
This group is long enough to interrupt the interresidue H-bonds and
chemically versatile; it allows a binomial synthesis.
Several variations of the synthesis of the simplest monomeric unit that is
required for the synthesis of one of the cellulose analogues are described.
In the first variant, treatment of the epoxide 105 with LiMe3SiCsCLi/Et2-
A1C1 according to a known procedure gave the desired alkyne 106 besides
107, resulting from a neighbouring group participation of the MeOCH20
group. Using Me3Al instead of Et2AlCl increased the yield and selectivity
of the ring opening. Deprotection of 106 (—> 111), dibenzylation (-> 115),
and acetolysis led to the diacetate 116 which was partially deacetylated (—>
117) and oxidised to the lactone 118. Addition of (Me3SiOCLi)/TiCl4
afforded the anomeric hemiketals 119 (a-D/$-D = 3:2) which were
deoxygenated to the dialkyne 120. This synthesis of the monomers was
shortened by treating the hydroxyacetal 129 (from 106) with (Me3Si-
OQ3AI (Scheme 30); formation of the alkyne 130 (70%) by retentive
alkynylating acetal cleavage is rationalised by postulating a participation
of HOC(3). The synthesis was further improved by substituting the MeOCH2O
by the (iPr^SiO group (Scheme 32); the epoxide 131 from 104
yielded 85% of the alkyne 132 which was transformed, on the one hand,
via 133 into the dibenzyl ether 115 and, on the other hand, after C-desilylation
(—> 134) into the dialkyne 135. Finally, combined alkynylating
opening of the oxirane and the 1,3-dioxolane rings of 131 with excess
-219-
Et2A10CSiMe3 led directly to the monomer 137 which is thus available
in two steps and 77% yield from 104 (Scheme 33).
The Me3Si-C(l) bond of the bis-trimethylsilylethynylated anhydroalditol
137 was regioselectively cleaved with BuLi to yield 144/145, while
AgN02/KCN in MeOH cleaved the Me3Si-C(2') bond, leading to 135.
Both Me3Si groups were removed with NaOH in MeOH (-» 147), the (i-
Pr)3Si group was selectively cleaved with HC1 in aq. MeOH (-» 146); all
silyl substituents were removed with BU4NF (— 148). Oxidative
dimerisation of either 144 or 135, or of a mixture of 144/146 yielded only
the homodimers 149 and 150; treatment of 150 with AgN02/KCN yielded
151, deprotection proceeding much more slowly than the cleavage of the
C(2')Me3Si group of 137.
The iodoalkyne 154, required for the cross coupling with 135 according
to Cadiot-Chodkiewicz and Wityak-Chan, was prepared by deprotection of
144/145 to 152, methoxymethylation (-> 153), and iodination. Crosscoupling
yielded mostly 156, besides the homodimer 155. Similarly, cross
coupling of 154 and 159 (obtained from 145) led to 160 and 155. The
structure of 160 was established by X-ray analysis, showing a C(6)-C(5')
distance of 5.2 A. The conditions for desilylating 137 were applied to 156,
and led regioselectively to 161 (AgN02/KCN), 162 (aq. NaOH), 164
(BU4NF), and 163 (HCl/MeOH). Attempted deprotection of the
propargylic ether moiety with BuLi, however, failed. The dimer 164 was
further deprotected to 165.
Cleavage of the benzyloxy groups was then studied. Acetolysis (AC2O-
/Me3SiOTf) transformed 169 into 174, which was desilylated to 175, while
thiolysis of 169 led to a mixture of 171 and 172. The tetraacetate 175 has
also been obtained from 169 via 170. Acetolytic debenzylation of the
dimer 176, obtained from 170, gave 177 (83%), which was deacetylated to
178. Cross coupling of the alkyne 135 and the bromoalkyne 179, obtained
from 170, yielded 180; again, acetolysis proceeded well, leading to 181.
The cellobiose derivative 184 was prepared from the lactone 182 via 183.
The glycosidic linkage of 184 proved resistant to the conditions of aceto¬
lysis, leading to 185. Acetolysis of the benzylated thiophene 186 (from 176
with Na2S) yielded the octaacetate 187, but proceeded in substantially
lower yields (50%). NaSMe in toluene leads to regioselective de-C-
220-
silylation of the bis(trimethylsilyl (Me3Si) ethynyl) saccharide 137, but to
decomposition of butadiynes such as 156 or 200.
We have therefore combined the reagent-controlled, regioselective
desilylation of 137 and of 200 (AgNC^/KCN) with a substrate controlled
regioselective de-C-silylation, based on C-silyl groups of different size.
This combination was studied with the fully protected 188 which was
mono-desilylated to 189 or to 159. Triethylsilyation of 159 (-» 195) was
followed by removal of the Me3Si group (-»196). Introduction of a tert.-
butyldimethylsilyl (TBDMS) group (—> 197) and removal of the triethylsilyl
(TES) group yielded 198; these high-yielding transformations proceed
with a high degree of selectivity.
Iodination of 189 gave 190; this was coupled with 159 to the homodimer
199 and the heterodimer 200, which was desilylated to 204. The second
building block for the tetramer was obtained by coupling 201 (from 196)
with 159, leading to 202 and 203. Removal of the Me3Si group (-> 206)
and iodination led to 207 which was coupled with 204 to the homotetramer
208 and the heterotetramer 209. Deprotection of 209 gave 210, which was,
on the one hand, iodinated to 211, and, on the other hand, protected as the
C-TBDMS alkyne 212. Removal of the TES group and coupling afforded
the homooctamer 214 and the heterooctamer 215. Yields of iodination,
silylation, and desilylation were consistently high, while heterocoupling
proceeded in only 50-64%.
Cleavage of the (iPrbSiO- and MeOCFhO- groups of 199 (-> 216),
202 (-» 217), 208 (-> 218) and 214 (-» 219) proceeded in high yields.
Complete deprotection in two steps of the heterocoupling products 203 (-*
220 -> 165), 209 (-» 221 -> 222), and 215 (-» 223 -» 224) gave the
unprotected dimer 165, tetramer 222, and octamer 224 in high yields. Only
the dimer 165 is soluble in H2O; the !H-NMR spectra of 165, 222, and 224
in D6-DMSO (relatively low concentration) show no signs of
