Robert Kawęcki

Interaction of flavonoid topoisomerase I and II inhibitors with DNA oligomers

The binding affinities of flavonoids, genistein and quercetin, to DNA oligomers have been established by means of studying by NMR the diffusion coefficients of these compounds with and without the presence of DNA. Genistein was found to bind very weakly, Ka = 1.54 × 102 M−1, as compared to quercetin, Ka = 5.75 × 103 M−1 and luteolin, Ka = 2.17 × 104 M−1 (as reported in the literature). In the case of genistein a hydrogen bond between the NHF,B cytidine protons of the edge base pair and a genistein hydroxyl is proposed, based on the shape of the DOSY spectrum. MP2 and DFT calculations of genistein show a 9 kcal mol−1 excess energy of the planar conformation as compared with a twisted one. For this reason intercalation of the planar genistein into the base pairs is rather unlikely.

Solution and solid state 13C NMR and X-ray studies of genistein complexes with amines. Potential biological function of the C-7, C-5, and C-4′-OH groups

Parent genistein and its new amine complexes with morpholine and piperazine were studied comparatively in the solid and liquid states by X-ray crystallography and 13C and 15N NMR spectroscopy. Biochanine A and its complexes were used as reference. Secondary deuterium isotope effects on 13C chemical shifts in solution were studied in parent isoflavones and their morpholine and piperazine complexes to aid in evaluation of the electronic distribution in both systems. In addition, to quantify the extent of proton transfer as well as to establish strong hydrogen bonding of the 7-OH group in a morpholine complex, proton transfer from the 7-OH group to the piperazine nitrogen atom was also confirmed by 13C NMR in the solid state and by X-ray studies. The effect of 7-OH deprotonation yields a high frequency shift of 7–8 ppm on the C-7 carbon atom of the piperazine complex whereas it is as large as 12 ppm in the morpholine complex in the solid. The former trend is confirmed from solution state concentration studies which also show that the isoflavones have a strong tendency to form complexes with bases. Depending on the pKa difference between the isoflavones and the base this leads either to proton transfer and ion-pair formation or, in the case of a larger pKa difference, to a hydrogen bonded ion pair. The concentration studies show formation of a 1 : 1 genistein–piperazine complex in DMSO. Addition of water leads to formation of solvent separated ions. The C-5 OH group is involved in strong intramolecular hydrogen bonding leading to a pseudo aromatic ring extending the aromatic part of the drug pharmacophore. The analysis also suggests the way that both the C-7 and C-4′ hydroxyl group of genistein may participate in stabilising the ternary inhibitor complexes of tyrosine-specific kinases or DNA topoisomerase II.

Binding of Topotecan to a Nicked DNA Oligomer in Solution

Topotecan (TPT) is in clinical use as an antitumor agent. It acts by binding to the covalent complex formed between nicked DNA and topoisomerase I, and inserts itself into the single-strand nick, thereby inhibiting the religation of the nick and acting as a poison. A crystal structure analysis of the ternary complex has shown how the drug binds (B. L. Staker, K. Hjerrild, M. D. Feese, C. A. Behnke, A. B. Burgin, L. Stewart, Proc. Natl. Acad. Sci. U.S.A., 2002, 99, 15 387-15 392), but has left a number of unanswered questions. Herein, we use NMR spectroscopy and molecular modeling to show that the solution structure of a complex of TPT with nicked natural DNA is similar, but not identical to the crystal conformation, and that other geometries are of very low population. We also show that the lactone form of TPT binds approximately 40 times more strongly than the ring-opened carboxylate.

New, recoverable and highly effective sulfinyl chiral auxiliary

A facile synthesis of a bicyclic 8-menthylsulfinate is described. The reaction of this sultine with organometallic compounds leads to γ-hydroxysulfoxides, while the reaction with lithium amide affords 8-menthylsulfinamide. The chiral efficiency of the 8-menthylsulfinyl auxiliary was tested in the reaction of sulfinimines with Grignard reagents. A highly stereoselective addition of ethylmagnesium bromide to 8-menthylsulfinimine was observed (98% de).

Facile synthesis of homochiral derivatives of 10-bornane sulfinates, sulfinamides and sulfinimines

Abstract A convenient route for the synthesis of enantiopure 10-bornanesulfinic acid derivatives from (1 S )-camphorsulfonyl chloride is described. An interesting epimerization at sulfur in five-membered sultines is observed, leading to diastereoisomerically pure ( S S )-10-isobornyl sulfinates. A large difference in solubility in organic solvents allowed ( R S )-2-oxo-10-bornanesulfinamide to be obtained as the major diastereoisomer. Cyclization of 2-oxo-10-bornanesulfinamides gave five-membered sulfinimines.

Chiral Thiophene Sulfonamide—A Challenge for VOA Calculations

Two enantiomers of 2-methyl-N-(1-thien-2-ylethyl)propane-2-sulfonamide (TSA) were synthesized, and their VCD, ROA, IR, and Raman spectra were registered. The solved (S)-TSA X-ray structure shows a disorder connected to the presence of two TSA conformers differing by a slight rotation of the thiophene ring. Two molecules in the unit cell of the monoclinic P21 crystal form a net of NH···OS and C*H···OS hydrogen bonds. Out of a series of computational levels tested to interpret the spectra, the B3LYP functional combined with the def2TZVP basis set satisfactorily reproduces the experimental VCD and ROA spectra. To simulate the VCD spectra of TSA enantiomers in KBr pellets, dimers and tetramers, with two different positions of the thiophene ring, were considered. The VCD spectra measured in CDCl3 are completely different from those taken in KBr due to the conformational freedom of TSA in chloroform. Seven TSA conformers fall into two groups of opposite configurations at the pyramidal N atom forming the additional stereogenic center. However, the barriers between conformers in each group are lower than the energy of thermal motions at 300 K. Thus, all conformers, but the most stable in each group, are likely to be metastable states. The calculated IR, VCD, Raman, and ROA spectra of the conformers depend not only on the type of stereogenic N atom but also on the thiophene ring rotation. Yet, they are likely to coexist because of low barriers between them. Three approaches were tested to reproduce the chiroptical spectra in solution using PCM and hybrid solvation models. As a consequence, it was found that a model in which all conformers contribute to the spectra with equal population factors seems to best reproduce the experimental data. Such a result suggests that in a dissolved state in 300 K TSA occurs in a very shallow potential well and all of its conformers coexist.

A novel isoquinoline alkaloid, DD-carboxypeptidase inhibitor, with antibacterial activity isolated from Streptomyces sp. 8812. Part II: Physicochemical properties and structure elucidation.

A novel antimicrobial agent labeled JS-1, being a member of isoquinoline alkaloids, of molecular formula C10H9NO4 was isolated from the culture broth of Streptomyces sp. 8812. In this study, we present the structure based on physicochemical and spectroscopic NMR investigations and on quantum chemical structure modeling. The structure of a molecule suggests the biosynthetic path starting from 3′-hydroxy tyrosine. The synthesis was undertaken and it resulted in NMR data that fully agree with the presented analysis.

Spontaneous 2′-deoxyguanosine alkylation by a new generation of topoisomerase I inhibitors of the camptothecin family

We present NMR and LC-MS evidence that 7-ethyl-9-(N-morpholinyl)methyl-10-hydroxycamptothecin spontaneously binds covalently to 2′-deoxyguanosine under near-physiological conditions. The 2-NH2 is alkylated selectively independent of conditions, either in pure water at pH 6 or in 25 mM NaCl/25 mM potassium phosphate at pH 7. This is the first documented case of the spontaneous covalent binding of 2′-deoxyguanosine by a potential topoisomerase I inhibitor. This type of camptothecin derivative (a 9-alkylaminomethyl-substituted derivative of the parent SN38, 7-ethyl-10-hydroxycamptothecin) may potentially be active in vivo because it belongs to the class of camptothecin derivatives (including Camptosar or Hycamtin) used clinically as chemotherapeutic agents for the treatment of cervical, colon, and breast cancer.

DOSY NMR and MALDI-TOF evidence of covalent binding the DNA duplex by trimethylammonium salts of topotecan upon near UV irradiation

Using DOSY NMR and MALDI‐TOF MS techniques, we present evidence that quaternary trimethylammonium salts of topotecan, [TPT‐NMe3]+ X− (X = CF3SO3, HCOO), bind covalently the natural DNA oligomer upon near UV irradiation in water under physiological conditions. It is shown that formate salt is very reactive at pH 7 and requires short irradiation time. This weak irradiation at 365 nm paves the way for a new application of TPT derivatives in clinical use, which can dramatically increase the therapeutic effects of a medicine. Copyright

Genistein Binding Mode to Doubly Nicked Dumbbell DNA. Dynamic and Diffusion Ordered NMR Study

New genistein derivatives were synthesized, which are fairly well soluble in water, with respect to parent genistein, and thus facilitate study of the interaction with dumbbell DNA dodecamer, mimicking the biological target for topoisomerase II inhibitors. A pulsed field gradient spin echo NMR experiment was used to check the binding and to estimate the association constants and its pH dependence of genistein with dumbbell DNA. Experimental restraints based on nuclear Overhauser spectroscopy spectra were used to calculate the NMR structure in solution in case of 6,8-disubstituted genistein with dimethylaminomethyl groups and were used in molecular modeling calculations. The structure is dynamic, and 10 molecular dynamics runs yield a family of conformations that essentially differ in a depth of inclusion of genistein into a nick. The paper experimentally shows evidence for binding, intercalation in the nick is proposed as a mode of genistein binding, and a model of the event is provided.