1993

1) A. Kraszewski, M. Sobkowski & J. Stawiński
"Studies on Reactions of Nucleoside H-Phosphonates with Bifunctional Reagents. 1. Reaction with Amino Alcohols"
J. Chem. Soc. Perkin Trans. 1, 1699-1704, 1993.
Abstract: The reaction of nucleoside H-phosphonates with amino alcohols in the presence of condensing agents has been studied. Depending on the coupling procedure used and the length of the polymethylene bridge in the amino alcohols, different H-phosphonate diesters, cyclic phosphoramidite derivatives or phosphite triesters can be produced. Oxidation of these species with iodine under various experimental conditions has been studied and a general procedure for synthesis of nucleoside alkyl phosphodiesters has been developed.

  

1994

2) G. Ott, L. Arnold, J. Smrt, M. Sobkowski, S. Limmer, H.P. Hofmann & M. Sprinzl
"The chemical synthesis of biochemically active oligoribonucleotides using dimethylaminomethlene protected purine H-phosphonates"
Nucleosides Nucleotides, 13 (5), 1069-1085, 1994.
Abstract: Dimethylaminomethylene was applied as the protecting group for the exocyclic amino groups of adenosine and guanosine in the automated chemical synthesis of oligoribonucleotides on a polymer bound support. The dimethylaminomethylene protecting group can be removed at room temperature under conditions where the concomitant loss of the 2'-protection group can be excluded. The transformation of 2'-O-(t-butyldimethylsilyl)-5'-O-(4,4'-dimethoxytrityl) protected nucleosides to 3'-H-phosphonates yields synthons, well suited for the automated chemical synthesis of oligoribonucleotides. Using these H- phosphonate monomers, a coupling time of two minutes is sufficient to obtain average coupling yields of more than 98 %. Synthesized RNA is recognized as a substrate in an enzymatic reaction, forms the expected secondary structures and is suitable for NMR structural investigations.

3) J. Jankowska, M. Sobkowski, J. Stawiński & A. Kraszewski
"Studies on aryl H-phosphonates. 1. An efficient method for the preparation of deoxyribo-and ribonucleoside 3'-H-phosphonate monoesters by transesterification of diphenyl H-phosphonate"
Tetrahedron Lett., 35 (20), 3355-3358, 1994.
Abstract: A convenient method for the preparation of deoxyribonucleoside and ribonucleoside 3'-H-phosphonate monoesters via transesterification of diphenyl H- phosphonate with suitable protected nucleosides in pyridine is described.

4) M. Sobkowski, J. Stawiński, A. Sobkowska & A. Kraszewski
"Studies on reactions of nucleoside H-phosphonates with bifunctional reagents. 2. Stability of nucleoside H-phosphonate diesters in the presence of amino alcohols"
J. Chem. Soc. Perkin Trans. 1, 1803-1808, 1994.
Abstract: H-Phosphonate diesters undergo transesterification with amino alcohols to afford as primary products the mixed and the symmetrical H-phosphonate esters. Alcohols react similarly but only in the presence of an external base or in a basic solvent. The rate and the course of transesterification strongly depend on the reaction conditions, the reactivity of the H-phosphonate diester used and the nature of the amino alcohol.

  

1995

5) M. Sobkowski, J. Stawiński & A. Kraszewski
"Studies on reactions of nucleoside H-phosphonate diesters with bifunctional reagents. 4. Chemoselectivity during oxidative coupling of nucleoside H-phosphonate diesters with amino alcohols controlled by protonation of the amino function"
Tetrahedron Lett., 36 (13), 2295-2298, 1995.
Alternate link: ICM   Biblioteka Wirtualna Nauki
Abstract: A Kraszewski, Polish Acad Sci, Inst Bioorgan Chem, Noskowskiego 12-14, PL-61704 Poznan, Poland The course of oxidative coupling of H-phosphonate diesters with amino alcohols, promoted by iodine, can be controlled by protonation of the amine function of amino alcohols. Based on this phenomenon, a rapid and efficient method for synthesis of aminoalkyl phosphotriesters or hydroxyalkyl phosphoramidates was developed.

6) A. Kers, I. Kers, J. Stawiński, M. Sobkowski & A. Kraszewski
"Studies on aryl H-phosphonates. 2. A general method for the preparation of alkyl H-phosphonate monoesters"
Synthesis, 4, 4427-4430, 1995.
Abstract: A simple and efficient synthetic method for the preparation of alkyl H-phosphonate monoesters has been developed. The method consists of transesterification of commercially available diphenyl H-phosphonate with an appropriate alcohol under mild conditions, followed by ammonolysis of the in situ formed dialkyl and alkyl phenyl H-phosphonates.  

7) A. Sobkowska, M. Sobkowski, J. Stawiński & A. Kraszewski
"Studies on aryl H-phosphonates. Synthesis of nucleoside N-alkylphosphonamidates"
Nucleosides Nucleotides, 14 (3-5), 703-706, 1995.
Abstract: The aminolysis of aryl nucleoside H-phosphonate diesters with various amines was studied. The new simple and efficient method of synthesis of nucleoside phosphonamidates is described.

8) M. Sobkowski, J. Stawiński, A. Sobkowska & A. Kraszewski
"Studies on reactions of nucleoside H-phosphonates with bifunctional reagents. 3. Further studies on transesterification of nucleoside H-phosphonate diesters with amino alcohols"
Nucleosides Nucleotides, 14 (3-5), 839-842, 1995..
Abstract: Reactions of 5'-O-(4,4'-dimethoxytrityl)thymidin-3'-yl 2-cyanoethyl phosphonate with alcohols, amines and amino alcohols in pyridine are described. Transesterification was found to be faster than beta-elimination of 2- cyanoethyl group.

  

1996

9) J. Stawiński, A. Kraszewski & M. Sobkowski
"Exploring Reactions of Nucleoside H-phosphonates with Bifunctional Reagents"
Phos. Sulf. Sil., 109-110, 261-264, 1996.
Abstract: Studies on reactions of nucleoside H-phosphonates with various amino alcohols showed that: (i) condensations of H-phosphonate monoesters with amino alcohols proceed with a complete chemoselectivity producing H-phosphonate diesters exclusively; (ii) H-phosphonate diesters undergo transesterification with amino alcohols and afford various products depending on the reaction conditions; (iii) the course of the oxidative coupling of nucleoside H-phosphonate diesters with amino alcohols can be controlled by protonation of the amino function, and thus the reaction can be steered to afford aminoalkyl phosphotriesters or hydroxyalkyl phosphoramidates.

10) M. Sobkowski, J. Stawiński & A. Kraszewski
"Aryl H-phosphonates. Part 5. A Simple Method for the Synthesis of Aminoalkyl H-phosphonate Monoesters via Transesterification of Difenyl H-phosphonate with Amino Alcohols"
Collection Czechoslov. Chem. Commun., 61, S238-S241, 1996.
Abstract: A new method for the synthesis of N-dimethoxytritylated aminoalkyl H-phosphonate monoesters is described. It consists of transesterification of diphenyl H-phosphonate with an amino alcohol, hydrolysis of the produced di(aminoalkyl) H-phosphonate diester to the corresponding monoester, followed by its tritylation, and silica gel purification of the final product..

11) A. Kers, I. Kers, J. Stawiński, M. Sobkowski & A. Kraszewski
"Studies on aryl H-phosphonates. 3. Mechanistic investigations related to the disproportionation of diphenyl H-phosphonate under anhydrous basic conditions"
Tetrahedron, 52 (29), 9931-9944, 1996.
Alternate link: ICM   Biblioteka Wirtualna Nauki
Abstract: Diphenyl H-phosphonate undergoes under anhydrous reaction conditions a base-promoted disproportionation to triphenyl phosphite and phenyl H-phosphonate. On the basis of P-31 NMR data the most likely mechanism for this transformation was proposed. In order to substantiate these findings and to get a deeper insight into the chemistry of aryl H-phosphonate esters, we carried out also some studies on activation of phenyl and diphenyl H-phosphonates with various condensing agents. We found that aryl vs alkyl esters of phosphonic acid often follow different reaction pathways during the activation, and this can most likely be traced back to higher electrophilicity of the phosphorus centre and to higher reactivity of the P-H bonds in aryl H-phosphonate derivatives.

12) J. Cieślak, M. Sobkowski, A. Kraszewski & J. Stawiński
"Aryl H-phosphonates. 4. A new method for internucleotide bond formation based on transesterification of aryl nucleoside H-phosphonate diesters"
Tetrahedron Lett., 37 (26), 4561-4564, 1996.
Alternate link: ICM   Biblioteka Wirtualna Nauki
Abstract: Under mild reaction conditions nucleoside aryl H- phosphonate diesters undergo fast and efficient transesterification with suitably protected nucleosides, affording dinucleoside (3'-5') H-phosphonate diesters.

13) J. Cieślak, J. Jankowska, A. Kers, I. Kers, A. Sobkowska, M. Sobkowski, J. Stawiński & A. Kraszewski
"Synthetic Applications of Aryl H-phosphonates in Nucleotide Chemistry"
Collection Czechoslov. Chem. Commun., 61, S242-S245, 1996.
Abstract: Aryl nucleoside H-phosphonate diesters were found to be versatile synthetic intermediates for the preparation of various functionalized alkyl nucleoside H-phosphonates, dinucleoside H-phosphonates, and nucleoside H-phosphonamidates.

14) A. Kers, I. Kers, A. Kraszewski, M. Sobkowski, T. Szabó, M. Thelin, R. Zain & J. Stawiński
"Nucleoside Phosphonates. Development of Synthetic Methods and Reagents"
Nucleosides Nucleotides, 15 (1-3), 361-378, 1996.

   

1997

15) A. Sobkowska, M. Sobkowski, J. Cieślak, A. Kraszewski,  I. Kers & J. Stawiński
"Aryl H-Phosphonates. 6. Synthetic Studies on the Preparation of Nucleoside N-Alkyl-H-phosphonamidates"
J. Org. Chem., 62 (14), 4791-4794, 1997.
Abstract: Various approaches to the synthesis of nucleoside H-phosphonamidates have been investigated. Direct couplings of nucleoside H-phosphonates with amines have been hampered by extensive reactions of the condensing agents with amines. Preactivation of nucleoside H-phosphonates with pivaloyl chloride or chlorophosphates, followed by the addition of amines, notably diminished these side reactions. The most efficient and versatile route to nucleoside N-alkyl H-phosphonates was found to be aminolysis of the in situ-produced aryl nucleoside H-phosphonates with appropriate amines.

  

1998

16) M. Sobkowski, A. Kraszewski & J. Stawiński
"The Reactions of H-Phosphonates with Bifunctional Reagents. Part V. Functionalization of Support-Bound Oligonucleotides and Synthesis of Non-Radioactive Hybridization Probes"
Nucleosides Nucleotides, 17 (1-3), 253-267, 1998.
Abstract: Three methods for the functionalization of oligonucleotides with aminoalkyl moieties have been developed and their efficiencies were evaluated in the preparation of non-radioactive hybridization probes.

17) J. Jankowska, A. Sobkowska, J. Cieślak, M. Sobkowski, A. Kraszewski, J. Stawiński & D. Shugar
"Nucleoside H-phosphonates. 18. Synthesis of unprotected nucleoside 5'-H-phosphonates and nucleoside 5'-H-phosphonothioates and their conversion into the 5'-phosphorothioate and 5'-phosphorodithioate monoesters"
J. Org. Chem., 63 (23), 8150-8156, 1998.
Abstract: A simple and efficient protocol for the preparation of unprotected nucleoside 5'-H-phosphonates and nucleoside 5'- H-phosphonothioates via a one-step deprotection of suitable precursors with methylamine has been developed. The synthetic utility of the unprotected nucleotide derivatives was demonstrated by converting them under mild conditions to the corresponding nucleoside 5'- phosphorothioate and nucleoside 5'-phosphorodithioate monoesters. Factors affecting oxidation of H-phosphonate, H-phosphonothioate, and phosphite derivatives with elemental sulfur are also discussed.
 

Chapter 1) M. Sobkowski & A. Kraszewski
"Oligonukleotydowe sondy molekularne o detekcji nieradioizotopowej"
in "Na pograniczu chemii i biologii" vol. 1, pp. 17-80, Wydawnictwo Naukowe UAM, Poznań 1998

 
 

1999

Conf. 1) H. Seliger, M. Hinz, P. Jaisankar, M. Sobkowski, R. Ditz & F. Eisenbeiss
"A polymer support with unusually high loading capacity for large-scale oligonucleotide synthesis"
Poster presented at: IBC's 6th International Conference on Oligo-Therapeutics / IBC's 3rd International Conference on Oligo-Technologies, May 5-6, 1999, La Jolla, CA, USA
Abstract: The production of multikilogram quantities of synthetic oligonucleotides for therapeutic purposes calls for the development of new methodology, not just the scale-up of existing techniques. The use of supports with very high loading is particularly attractive, since the required amount of solid phase support per unit production of oligonucleotide is significantly reduced, as is the cost of disposal or regeneration. This likewise reduces the expenditure for apparatus and consumables. based on earlier experiments with macroporous PVA Merckogel resins, we have developed a new supportsystem with loading capacity ranging between 100 and 1000 micromol nucleoside/g. Applying standard phosphoramidite chemistry, oligonucleotides with typically up to 20 bases, but also longer sequences, could be prepared with yields of chain elongation averaging up to 99%. Even assuming only 97% average yield of chain extension, this constitutes a ca. 8-fold higher oligonucleotide output per unit support weight with respect to conventional 50 micromol/g supports, and ca. 3-fold with respect to 200 micromol/g "high-load" supports. Calculation of the composition of the support system after synthesis shows, that with loading around 700 to 800 micromol/g the material obtained after the preparation of a 20-mer consists of ca. 80% oligonucleotide and only 20% of polymeric support material. Applications of such supports, as well as studies on different reaction conditions and activating agents will be reported..

Conf. 2) J. Cieślak, J. Jankowska, M. Sobkowski, A. Kers, I. Kers, J. Stawiński & A. Kraszewski
"The Reactions of Aryl Nucleoside H-phosphonates with O-, N-, and S-nucleophiles"
Abstract: Aryl nucleoside H-phosphonates, derivatives of controlled reactivity, have been developed as useful synthetic intermediates for the preparation of variety of nucleotide analogues containing P-O, P-N and P-S bonds.
Collection Symp. Series 2, 63-68, 1999
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Conf. 3) H. Seliger, M. Hinz, M. Sobkowski, R. Ditz & F. Eisenbeiss
"High load support material for oligonucleotide synthesis"
Poster presented at: "Biotechnica", October 5-7, 1999, Hanover, Germany
Abstract: The synthesis of large amounts of oligonucleotides is still rather expensive, thus hampering the development and use of therapeutic oligo-nucleotides. The support material accounts for a large percentage of these costs. An increase in loading of the support material will still maintaining high coupling efficiency would not only diminish direct costs of the support material but also indirect costs by allowing a smaller design of the synthesizer. Together with Merck KGaA, a new support material, based on Merckogel OR 1000000 has been developed, meeting the requirements of large scale oligonucleotide synthesis nearly ideally.

Conf. 4) H. Seliger, M. Sobkowski, M. Hinz, P. Jaisankar, R. Ditz, F. Eisenbeiss, H. Knoller, R. Zimmermann, A. Rück, K. Heckelsmiller, R. Steiner & R. Masantschek
"Some Recent Contributions to Antisense Research. International Symposium on Drug Regulation of Gene Expression"
Communication presented at "International Symposium on Drug Regulation of Gene Expression" Bressanone, Italy, 1999.

  

2000

Conf. 5) H. Seliger, M. Hinz, R.K. Pandey, J. Simanowski, M. Sobkowski, S. Hahner, F. Hillenkamp, R. Ditz & F. Eisenbeiss
"Bead Technology for Oligonucleotide Libraries and Large-Scale Synthesis"
Poster presented at: "Millennium Conference on Nucleic Acid Therapeutics", 8-11 January, 2000, Clearwater Beach, Florida, USA
Abstract:  Particle Systems with unusually high loading capacity are the basis (1.) for combinatorial libraries of synthetic oligonucleotides used in the discovery of diagnostic and therapeutic lead structures and (2.) for the large-scale preparation of therapeutic oligonucleotides. After having scanned a number of commercial beads for these purposes, our work currently concentrates on Tentagel^(R) (Rapp Polymers, Tübingen) and Merckogel^(R) (Merck KGaA, Darmstadt). Both polymers are of spherical shape, can be obtained with quite uniform size, are mechanically stable and are produced/functionalized with hydroxy or amino groups.
Ad (1.) Beads of size around 30 microm diameter with nucleotide loading around 200 micromol/g were selected for the preparation of libraries. Oligonucleotide synthesis was done according to the "split/couple/combine" procedure¹ with yields of chain extension around 98%. Specificity tests of e.g. octanucleotide libraries were done routinely by hybridization to fluorescently labeled oligonucleotides, selection of fluorescent particles, followed by dehybridization and detection of the sequence off single beads by MALDI-TOF mass spectroscopy. This procedure has been extended to screening high-affinity oligonucleotide-protein interactions. As an example, the binding of thrombin to an octanucleotide library was analyzed and will be dis-cussed in relation to previous lead structures generated through solution aptamer techniques².
Ad (2.) Beads, that (a) are characterized by having unusually high loading capacity and (b) nevertheless give near-quantitative yields of chain extension, are particularly advantageous for the scale-up of oligonucleotide synthesis. Our efforts are concentrated on Merckogel^(R), a macroporous polyvinylacetate resin, functionalized via partial hydrolysis, which meets these requirements better than any other currently available particle system. Up to capacities of 500-600 micromol nucleotide/g the chainextension on such resins proceeds with 98-99% average. The productivity of such a high-load system is demonstrated by the fact, that after preparation of a 20mer the immobilized oligodeoxynucleotide comprises ca. 60% of the total weight of the carrier. The loading capacities can even be increased to 1000 micromol/g and beyond, however, resulting in decreased synthesis efficiency. Since the Merckogel^(R) shows similar swelling in all solvents used during the phosphoramidite cycle, these characteristics may contribute to a significant reduction of the volume of reaction vessels as well as the cost of chemicals and equipment during the large-scale production of therapeutic oligonucleotides.

1. A. Furka, F. Sebestyen, M. Agesdom, G. Dibo, Abstracts, l4th Internat. Congress Biochem. 1988, 47, Int. J. Peptide Protein Res. 37, 487-491 (1991); K.S. Lam, S.E. Salmon, E.M. Hersh, W.J. Hruby, W.M. Kasmierski, R.J. Knapp, Nature 354, 82-84 (1991).
2. L.C. Bock, L. Griffin, J.A. Latham, E.H. Vermaas, J.J. Toole, Nature 335, 564-566 (1992); K.Y. Wang, S. McCurdy, R.G. Shea, 5. Swaminathan, P.H. Bolton, Biochemistry 32, 1899- 1904 (1993).

Conf. 6) M. Sobkowski, M. Hinz, R. Ditz, F. Eisenbeiss, M. Rimmler & H. Seliger
"A high-load polymer support for the production of oligonucleotides of therapeutic interest"
Proceedings of XIV International Round Table "Nucleosides, Nucleotides and Their Biological Applications", San Francisco 2000, p. 128 (Abstract 205)
Abstract:  The synthesis of large quantities of synthetic oligonucleotides for therapeutic applications requires the development of a technical-scale methodology. Supports with very high loading are essential, if the expenditure for the preparation, regeneration or disposal of the support material, as well as the cost of apparatus and consumables are to be minimized. We have developed high-load support materials based on macroporous polyvinylacetate resins (MerckogelR), which almost ideally meet the requirements for large-scale oligonucleotide synthesis. The MerckogelR supports are featured by the following characteristics:-Procedures have been developed for controlled partial hydrolysis without disruption of the macroporous structure.-The nucleoside capacity can be regulated by variation of the loading conditions, yielding reproducibly up to 900 micromol/g.-The support material is compatible with standard oligonucleotide chain extension chemistry and workup procedures.-The supports have nearly equal swelling properties in various apolar solvents used during the oligonucleotide elongation cycle.-Standardized protocols have been developed to achieve 98-99% average cycle yields in the preparation of 15-30 base oligonucleotides with carriers loaded up to 350 micromol/g; similar yields have been obtained in experiments with support capacities of 500 micromol/g and more.

18) M. Sobkowski, M. Wenska, A. Kraszewski & J. Stawiński
"Studies on reactions of nucleoside H-phosphonates with bifunctional reagents. Part VI. Reaction with diols"
Nucleosides Nucleotides Nucleic Acids 19 (10-12), 1487-1503, 2000.
Abstract: Reactions of nucleoside H-phosphonates with various diols using different types of condensing agents have been studied. Depending on the coupling procedure and the length of a polymethylene chain of the diol, acyclic H-phosphonate diesters or cyclic phosphite triesters were formed. The course of oxidation with iodine to produce cyclic nucleoside alkyl phosphotriesters or hydroxyalkyl nucleoside phosphodiesters can be controlled by the amount of water present in the reaction medium.

  

2001

Pat. 1)  H. Seliger, M. Sobkowski & M. Hinz
"Large Scale-Synthese modifizierter Oligonukleotide"
("Intermediate for oligonucleotide synthesis comprises partially hydrolysed cross-linked vinyl acetate copolymer loaded with nucleotide derivative")
Deutsches Patent DE 10051726 A, 2001.
Title page.pdf

19) J. Cieślak, M. Sobkowski, J. Jankowska, M. Wenska, M. Szymczak, B. Imiołczyk, I. Zagórowska, D. Shugar, J. Stawiński & A. Kraszewski
"Nucleoside phosphate analogues of biological interest, and their synthesis via aryl nucleoside H-phosphonates as intermediates"
Acta Biochim.Pol. 48 (2), 429-442, 2001.
Abstract: This review presents a brief account of the chemistry and mechanistic aspects of aryl H-phosphonates, and selected applications of this class of compounds as intermediates in the synthesis of a wide range of biologically important analogues of nucleoside phosphates, and oligonucleotides, in which the phosphate moieties are replaced by other structurally related groups. The aryl nucleoside H-phosphonates, compounds of controlled reactivity, have proven to be more versatile and superior to various mixed anhydrides as synthetic intermediates, particularly for preparation of nucleotide analogues bearing P-N or P-S bonds in various configurational arrangements at the phosphate moiety.

20) M. Wenska, J. Jankowska, M. Sobkowski, J. Stawiński & A. Kraszewski
"A new method for the synthesis of nucleoside 2',3'-O,O-cyclic phosphorodithioates via aryl cyclic phosphites as intermediates"
Tetrahedron Letters 42 (45), 8055-8058, 2001.
Abstract: The reaction of 5'-O-protected ribonucleosides with tri(4-nitrophenyl) phosphite in the presence of pyridine furnished rapid formation of the corresponding 4-nitrophenyl 2',3'-O,O-cyclic phosphites which upon sulfhydrolysis, followed by sulfurization of the resultant cyclic H-phosphonothioate and removal of the 5'-O-protecting groups, afforded nucleoside 2',3'-O,O-cyclic phosphorodithioates in high yields.

  

2002

21) J. Cieślak, J. Jankowska, M. Sobkowski, M. Wenska, J. Stawiński & A. Kraszewski
"Aryl H-phosphonates. Part 13. A new, general entry to aryl nucleoside phosphate and aryl nucleoside phosphorothioate diesters"
J.Chem.Soc.Perkin Trans. 1, 31-37, 2002.
Abstract: The reaction of nucleoside H-phosphonate monoesters with phenols in the presence of a condensing agent, followed by oxidation of the in-situ-generated aryl nucleoside H-phosphonate diesters with iodine/water or with elemental sulfur, provides a new, one-pot, efficient entry to nucleoside phosphate or nucleoside phosphorothioate diesters bearing diverse aryl moieties.

Conf. 7) M. Sobkowski, J. Cieślak, J. Jankowska, J. Stawiński & A. Kraszewski
"Dinucleoside aryl phosphorothioates as building blocks for large scale synthesis of chimeric oligonucleotide analogues"
In: Collection Symposium Series (Z. Tocik and M. Hocek, Eds.), 5, 283-289, 2002. ISBN 80-86241-16-5

   

2003

22) M. Bollmark, T. Johansson, M. Kullberg, J. Nilsson, J. Stawiński, J. Cieślak, J. Jankowska, M. Sobkowski, M. Szymczak, M. Wenska & A. Kraszewski
"Developing synthetic methods for bioactive phosphorus compounds using H-phosphonate chemistry: A progress report"
Nucleosides Nucleotides Nucleic Acids 22 (5-8), 617-621, 2003.
Abstract: In this paper a short account of our recent research concerning the development, of new synthetic methods and reagents for the preparation of nucleotides and their analogues, is given.

Conf. 8) A. M. Awad, M. Sobkowski, M. Hinz & H. Seliger.
"Novel Antisense Oligonucleotides With Enhanced Endonuclease Stability"
Poster presented at IBC's 4th Annual Conference EuroTIDES, November 11-12 2003, Berlin, Germany

  

2004

23) A. M. Awad, M. Sobkowski & H. Seliger.
"Enzymatic and Hybridization Properties of Oligonucleotide Analogs Containing Novel Phosphoramidate Internucleotide Linkages"
Nucleosides Nucleotides Nucleic Acids 23 (5), 777-787, 2004.
Abstract: In line with the paradigm, that antisense oligonucleotides should contain minimal structural modifications, in order to minimize the risk of toxicity and antigenicity, we describe here the preparation and the properties of oligonucleotides modified to contain, in addition to phosphodiester bonds, a small number of phosphoramidate internucleotide linkages substituted with aminoethoxyethyl groups in order to convey protection against exo- and endonucleases. Prolonged stability was, in fact, found in model experiments with respective enzymes, as well as in studies done in human blood serum. Regardless of number and position of phosphoramidate linkages, the modified oligonucleotides showed only a slight decrease of Tm in hybridization studies with complementary oligonucleotides.
 

Conf. 9) Z. Wang, I. Cedillo, D. L. Cole, Y. S. Sanghvi, M. Hinz, W. Prukała, M. Sobkowski, H. Seliger, M. Rimmler, R. Ditz & J. Hoffmeyer.
"OligoPrep: A New PVA Support for Oligonucleotide Synthesis at Large Scale"
 In: "Innovation & Perspectives in Solid Phase Synthesis & Combinatorial Libraries 2004" (Proceedings of the 8th International Symposium, London), Mayflower Worldwide Ltd,  R. Epton, ed., pp 118-122, 2004.  ISBN 0-9515735-5-1
Abstract: OligoPrep(TM) was developed as an ideal support for automated solid-phase synthesis of therapeutic phosphorothioate oligonucleotides. Swelling properties, loading of the first nucleoside, and capping and detritylation steps were carefully studied and optimized on the new support. The beads proved to be chemically and mechanically stable under synthesis conditions at a loading of ~250 micromol/g. Synthesis of 20-mer phosphorothioate oligonucleotides at 1 mmol using phosphoramidite chemistry was successful at high product yield and purity.

  

2005

24) T. Johansson, J. Nilsson, M. Kullberg, G. Laven, M. Sobkowski, A. Szymańska, M. Szymczak, A. Kraszewski & J. Stawiński.
"Developing synthetic methods for bioactive phosphorus compounds using H-phosphonate chemistry: A progress report"
Nucleosides Nucleotides Nucleic Acids 24, (5-7), 353-357, 2005
Abstract: In this paper a short account of our recent basic studies aiming towards development of new synthetic methods for the preparation of nucleotide analogues using H-phosphonate chemistry is presented.

25) M. Sobkowski, J. Jankowska, J. Stawiński & A. Kraszewski
"Aryl Nucleoside H-phosphonates as a Tool for Investigation of Stereospecificity During Coupling"
Nucleosides Nucleotides Nucleic Acids 24, (5-7), 887-890, 2005
Abstract: It was found that in stereoselective condensations of ribonucleoside 3'-H-phosphonates with alcohols, the major diastereomer of the produced H-phosphonate diesters is formed from the minor diastereomer of the intermediate phosphonic-pivalic anhydride.

26) M. Sobkowski, J. Jankowska, J. Stawiński & A. Kraszewski
"A Cautionary Note on the Use of the ³¹P NMR Spectroscopy in Stereochemical Correlation Analysis"
Nucleosides Nucleotides Nucleic Acids 24, (5-7), 1033-1036, 2005
Abstract: Stereoselectivity in condensation of protected ribonucleoside 3'-H-phosphonates with hydroxylic components was investigated using ³¹P NMR spectroscopy. The correlation between absolute configuration at the phosphorus centre and the chemical shifts of the produced H-phosphonate diesters and the corresponding phosphorothioates, was studied.

27) M. Sobkowski, J. Stawiński & A. Kraszewski
"A proposal for a new stereochemical notation for P-chiral nucleotide analogues and related compounds"
Nucleosides Nucleotides Nucleic Acids 24 (9), 1301-1309, 2005.
Abstract: A new stereochemical notation for P-chiral nucleotide analogues and related compounds is proposed. In this notation, the names of configurations, designated as D_P and L_P, are derived from a geometrical relationship, rather than from priority rules, of substituents at the phosphorus centre. This new stereochemical description offers clear advantages over the CIP R/S nomenclature, particularly when used for comparing the influence of absolute configuration at the phosphorus centre on physico-chemical and biological properties of oligonucleotide analogues or in stereochemical correlation analysis of P-chiral nucleotide derivatives.

28) M. Sobkowski, J. Jankowska, J. Stawiński & A. Kraszewski
"Stereochemistry of internucleotide bond formation by the H-phosphonate method. 1. Synthesis and ³¹P NMR analysis of 16 diribonucleoside (3'-5')-H-phosphonates and the corresponding phosphorothioates"
Nucleosides Nucleotides Nucleic Acids 24 (10-12),  1469-1484, 2005
Abstract: Sixteen diribonucleoside (3'-5')-H-phosphonates were synthesized via condensation of the protected ribonucleoside 3'-H-phosphonates with nucleosides, and the influence of a nucleoside sequence on the observed stereoselectivity was analyzed. ³¹P NMR spectroscopy was used to evaluate a relationship between chemical shift and absolute configuration at the phosphorus centre of the H-phosphonate diesters as well as of the corresponding phosphorothioate diesters. Although for the most cases such correlation was found, there was however several exceptions to the rule where the relative positions of resonances arising from R_P and S_P diastereomers were reversed.
 

Conf. 10) M. Sobkowski, J. Jankowska, J. Stawiński & A. Kraszewski
"Stereochemistry of internucleotide bond formation by the H-phosphonate method. 2. Transesterification of aryl ribonucleoside H-phosphonate diesters with alcohols"
In: Collection Symposium Series (M. Hocek, Ed.), 7, 183-187, 2005. ISBN 80-86241-25-4
Abstract: Aryl ribonucleoside 3'-H-phosphonates were used as model compounds in investigations of stereochemistry of an internucleotide bond formation. For all four ribonucleoside 3'-H-phosphonates studied the major diastereomers of the produced H-phosphonate diesters were found to be formed from the minor diastereomers of nucleoside aryl H-phosphonate diesters. These studies indicate the importance of an equilibrium between substrate diastereomers and can be relevant to the stereoselectivity observed in condensation reactions of ribonucleoside H-phosphonates promoted by condensing agents, e.g. pivaloyl chloride.
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Conf. 11) M. Sobkowski, J. Stawiński & A. Kraszewski
"A proposal of D_P/L_P notation for nucleotide analogues with a chiral phosphorus centre"
In: Collection Symposium Series (M. Hocek, Ed.), 7, 467-469, 2005. ISBN 80-86241-25-4
Abstract: A proposal of a new notation for P-chiral nucleotide analogues is outlined. In contrast to the absolute R_P/S_P convention, the new D_P/L_P system is based on a geometrical relationship between substituents at the phosphorus atom, and thus appears to be particularly convenient for structural analyses of physical, chemical, and biological properties of nucleotide and oligonucleotide analogues.
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2006

29) M. Sobkowski, J. Stawiński & A. Kraszewski
"A proposal for a convenient notation of P-chiral nucleotide analogues. Part 2. Dinucleoside monophosphate analogues"
Nucleosides Nucleotides Nucleic Acids 25 (12), 1363-1375, 2006.
Abstract: A configuration of ligands around a phosphorus atom in P-chiral dinucleoside monophosphate analogues can be described using D_P/L_P stereochemical notation, which allows immediate correlation between the notation of configuration and the actual spatial arrangement of phosphorus ligands. The area of applications of this new stereochemical nomenclature covers dinucleoside units bridged by virtually any type of tri- and tetra-coordinated phosphorus moieties, i.e. phosphorothioates, phosphoramidates, phosphoramidites, boranophosphates, methanephosphonates, H-phosphonates, and many others.

30) M. Sobkowski, J. Stawiński & A. Kraszewski
"A proposal for a convenient notation of P-chiral nucleotide analogues. Part 3. Compounds with One Nucleoside Residue and Non-Nucleosidic Derivatives"
Nucleosides Nucleotides Nucleic Acids 25 (12), 1377-1389, 2006
Abstract: Recently, we have proposed a new D_P/L_P stereochemical notation for P-chiral dinucleoside monophosphate analogues that permits simple correlation between spatial arrangement of the substituents and the configuration at the phosphorus center. As an extension of this work, we present here applications of the D_P/L_P notation to derivatives containing only one nucleoside unit
(e.g., alkyl nucleoside phosphodiesters, nucleoside phosphomonoesters, cyclic phosphate derivatives, nucleoside di-, and triphosphates) and to nonnucleosidic phosphorus compounds.

Conf. 12) M. Sobkowski
"Stereochemistry of internucleotide bond formation by the H-phosphonate method"
Oral presentation at "Chemiedozententagung", Hamburg, Germany, 2006.

2007

Chapter 2) M. Sobkowski, J. Stawiński & A. Kraszewski
"A Convenient Stereochemical Notation for P-Chiral Nucleotide Analogs"
In: Current Protocols in Nucleic Acid Chemistry. APPENDIX 1E
John Wiley and Sons, Inc.
Abstract: The D_P/L_P convention is a stereochemical notation for P-chiral nucleotide analogs and related compounds. In contrast to the absolute R_P/S_P notation, the D_P/L_P system is based on a geometrical relationship between substituents in a dinucleoside monophosphate skeleton. The D_P/L_P notation is a convenient alternative to the R_P/S_P notation for stereochemical correlation analysis of physical, chemical, and biological properties of nucleotide and oligonucleotide analogs bearing any type of tri- or tetra-coordinated phosphorus moiety.

31) M. Sobkowski, J. Stawiński & A. Kraszewski
"Stereochemistry of internucleotide bond formation by the H-phosphonate method. 3. Investigations on a mechanism of stereoselectivity induction"
Tetrahedron: Asymmetry 18 (19), 2336-2348, 2007. 
Abstract: The stereochemistry of condensation of ribonucleoside H-phosphonate monoesters with hydroxylic components has been investigated using 31P NMR spectroscopy. It was found that the reactions owe their stereoselectivity to a dynamic kinetic asymmetric transformation. The relative rates of the equilibration and esterification of the reactive species were evaluated. Absolute configurations of the compounds involved in the reaction pathways were tentatively assigned on the basis of the chemical shifts of their 31P NMR signals. The correctness of the experimental data interpretation was validated for the H-phosphonic-pivalic mixed anhydrides and H-phosphonate active aryl esters.

 

2008

Conf. 13) M. Sobkowski
"Diverse chemoselectivity during acylation of nucleosides"
In: Collection Symposium Series (M. Hocek, Ed.), 10, 277-281, 2008.
Abstract: The acylation of nucleosides with acid chlorides under very mild conditions (diluted solution in neutral solvents containing a few equivalents of an amine) was investigated. The rate and the sites of the reaction were found to depend strongly on the kind of amine used. In the presence of DMAP acylation occurred chemoselectively on the hydroxy groups of the ribose moiety, while a mixture of triethylamine and pyridine promoted acylation of the thymine residue.
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32) M. Sobkowski, J. Stawiński & A. Kraszewski
"Stereochemistry of internucleotide bond formation by the H-phosphonate method. Part 6. Optimization of the reaction conditions towards highest stereoselectivity"
Tetrahedron: Asymmetry 19 (21), 2508-2518, 2008. 
Abstract: The condensations of ribonucleoside 3'-H-phosphonates with simple alcohols and nucleosides are known to be stereoselective reactions that favour formation of D_P(S)_P diastereomers of the produced H-phosphonate diesters without engaging any chiral auxiliaries. We have investigated various reaction conditions in order to attain the highest stereoselectivity of the condensation. With an optimal choice of the solvents, reagent concentrations, temperature, and the condensing agent used, the diastereomeric excess of the D_P(S)_P isomers of dinucleoside H-phosphonates was enhanced from the initial 50 - 70% to ca. 85%.

2009

33) M. Sobkowski, J. Stawiński & A. Kraszewski
"The role of nucleophilic catalysis in chemistry and stereochemistry of ribonucleoside H-phosphonate condensation" (Part 4 of the series "Stereochemistry of internucleotide bond formation by the H-phosphonate method")
New Journal of Chemistry 33, 164-170, 2009
Abstract: Efficiency and stereoselectivity of condensations of ribonucleoside 3'-H-phosphonates with alcohols were investigated as a function of amines used as bases. It was found that irrespective of the presence or absence of nucleophilic catalysts, the Dynamic Kinetic Asymmetric Transformation (DYKAT) was the major factor responsible for the stereoselective formation of the D_P(S)_P isomers of the H-phosphonate diesters, and a mechanistic rationalization of this observation was proposed. Studies on the reactions carried out in the presence of various bases led to conclusion that certain sterically hindered pyridines, e.g. 2,6-lutidine, may act as nucleophilic catalysts in the condensation of ribonucleoside 3'-H-phosphonates with alcohols.

34) M. Sobkowski, J. Stawiński & A. Kraszewski
"A proposal for a convenient notation of P-chiral nucleotide analogues. Part 4. A relationship between the D_P/L_P  notation and stereochemistry of reactions"
Nucleosides Nucleotides Nucleic Acids 28 (1), 29-42, 2009
Abstract: Recently, we have proposed a new D_P/L_P stereochemical notation for P-chiral dinucleoside monophosphate analogues based on a structural relationship between compounds. As an extension of this work, we present here applications of the D_P/L_P notation for tracking stereochemistry of reaction pathways involving H-phosphonate, phosphoramidite, phosphorotriester, and other intermediates frequently met in the nucleotide chemistry.

35) J. Romanowska, A. Szymańska-Michalak, M. Pietkiewicz, M. Sobkowski, J. Boryski, J. Stawiński & A. Kraszewski
"A New, Efficient Entry to Non-Lipophilic H-Phosphonate Monoesters - Preparation of Anti-HIV Nucleotide Analogues"
Letters in Organic Chemistry 6 (6), 496-499, 2009
Abstract: Crystalline ammonium (9H-fluoren-9-yl)methyl H-phosphonate was prepared by a new, simple method and it was used as the reagent of choice for the introduction of an H-phosphonate monoester functionality into non-lipophilic anti-HIV nucleoside analogues.

2010

36) M. Sobkowski
"A convenient protection for 4-oxopyrimidine moiety in nucleosides by the pivaloyl group"
Collection of Czechoslovak Chemical Communications 75 (1), 33-57, 2010
Abstract: Application of the pivaloyl group as a protection for the N3 position of thymidine and uridine was investigated. Pivaloylation of thymidine is a very rapid reaction proceeding under mild conditions with excellent regioselectivity for sugar or thymine moiety, depending on the amines used. Several pivaloylated thymidine derivatives were obtained by treatment of unprotected thymidine with pivaloyl chloride under various experimental conditions. Stability of the N3-pivaloyl protecting group under basic and acidic conditions was evaluated and the conditions for its selective removal were found.

37) M. Sobkowski, J. Stawiński & A. Kraszewski
"Stereochemistry of internucleotide bond formation by the H-phosphonate method. 7. Stereoselective formation of ribonucleoside (R_P)- and (S_P)-3'-H-phosphonothioate monoesters"
Tetrahedron: Asymmetry 21 (4), 410-419, 2010
Abstract: Ribonucleoside 3'-H-phosphonothioate monoesters exist in a form of R_P and S_P diastereomers. In order to obtain them in good yields and in high stereochemical purity, stereoselective strategies for their preparation were investigated. For the synthesis of the R_P isomer a stereoselective sulfhydrolysis of an activated nucleoside H-phosphonate was developed, while the monoesters having having S_P configuration were prepared by asymmetric transformation of diastereomeric mixtures of nucleoside 3'-H-phosphonothioates using either a condensation  with 9-fluorenemethanol, followed by its beta-elimination, or via pivaloylation-hydrolysis reaction sequence. A tentative assignment of absolute configurations of the obtained diastereomers of 3'-H-phosphonothioate esters was done via a stereochemical correlation analysis.

38) M. Sobkowski
"Chemistry and stereochemistry of internucleotide bond formation by the H-phosphonate method" (Perspective)
New Journal of Chemistry 34 (5), 854-869, 2010
Abstract: The appropriately protected ribonucleoside 3'-H-phosphonates react with nucleosides and alcohols in the presence of condensing agents under mild conditions, with a notable stereoselectivity. Chemistry and stereochemistry of this reaction were investigated using ³¹P NMR spectroscopy. A plausible mechanism for the asymmetric induction observed was identified as Dynamic Kinetic Asymmetric Transformation (DYKAT) operating due to different esterification rates of rapidly equilibrating P-epimers of the reactive intermediates. This diverse reactivity was tentatively attributed to steric demands of the H-phosphonic moiety located in the vicinity of bulky protecting group in the 2' position. The role of nucleophilic and base catalysis was analysed and the absolute configuration of the intermediates involved in the reaction pathways was tentatively assigned using ³¹P NMR stereochemical correlation analysis. With an optimal choice of the reaction conditions the diastereomers of dinucleoside H-phosphonate diesters could be obtained usually in >90:10 ratio.

39) M. Sobkowski, J. Stawiński & A. Kraszewski
"Stereochemistry of internucleotide bond formation by the H-phosphonate method. 5. The role of Bronsted and H-bonding base catalysis in ribonucleoside H-phosphonate condensation - chemical and stereochemical consequences"
Nucleosides Nucleotides Nucleic Acids 29 (8), 628-645, 2010
Abstract: Efficiency and stereoselectivity of condensations of ribonucleoside 3'-H-phosphonates with ethanol promoted by pivaloyl chloride were investigated as a function of tertiary amines used. Side reactions leading to an increased demand for the condensing agent were identified as derived from an attack of the pivalate anion at carbonyl centres of reactive pivaloyl derivatives. The conditions that secured quantitative yields of H-phosphonate diester condensations were assessed. Several tertiary amines promoted condensations with stereoselectivity higher than that observed for pyridine derivatives. A correlation between diastereoselectivity of the product formation and Bronsted and H-bonding basicities of the amine used was found.

2011

40) M. Sobkowski, J. Jankowska, J. Stawiński & A. Kraszewski
"Unusual stereochemistry of esterification of uridine 3'-H-phosphonothioate"
Phosphorus, Sulfur and Silicon 186 (4), 952-955, 2011
Abstract: According to ³¹P NMR correlation analysis, reactive derivatives of uridine 3'-H-phosphonothioate react with O-nucleophiles probably with retention of configuration.

41) J. Romanowska, M. Sobkowski, A. Szymańska-Michalak, K. Kołodziej, A. Dąbrowska, A. Lipniacki, A. Piasek, Z.M. Pietrusiewicz, M. Figlerowicz, A. Guranowski, J. Boryski, J. Stawiński & A. Kraszewski
"Aryl H-Phosphonates 17: (N-Aryl)phosphoramidates of Pyrimidine Nucleoside Analogues and Their Synthesis, Selected Properties, and Anti-HIV Activity"
J. Med. Chem., 54 (19), 6482-6491, 2011
Abstract: New synthetic protocol for the preparation of nucleoside 5'-(N-aryl)phosphoramidate monoesters 4 was developed. It consisted of a condensation of the corresponding nucleoside 5'-H-phosphonates with aromatic- or heteroaromatic amines promoted by diphenyl phosphorochloridate, followed by oxidation of the produced H-phosphonamidates with iodine/water. 5'-(N-Aryl)phosphoramidate monoesters derived from 3'-azido-3'-deoxythymidine (AZT) or 2',3'-dideoxyuridine (ddU) nucleosides and various aromatic and heteroaromatic amines were evaluated as potential anti-HIV drugs. If was found that these compounds act most likely as pronucleotides, and some of them have therapeutic indices superior to those of the reference AZT.

2012

Conf. 14) J. Romanowska, K. Kołodziej, M. Sobkowski & A. Kraszewski
"Nukleozydo (N-arylo)amidofosforany - nowe wysoce aktywne i nietoksyczne pronukleotydy anty-HIV"
Poster presented at "55. Zjazd PTChem i SITPChem" conference, September 16-20, 2012, Białystok, Poland
Conference proceedings, p. 199

2013

Conf. 15) K. Kołodziej & M. Sobkowski
"Aryl nucleoside phosphoramidates as new anti-HIV prodrugs"
BioTechnologia, 94 (1), 80, 2013

Conf. 16) M. Materna & M. Sobkowski
"Stereochemistry of phosphorylating agents comprising nucleophilic catalytic groups"
BioTechnologia, 94 (1), 86, 2013

Conf. 17) J. Romanowska, M. Sobkowski & A. Kraszewski
"New type of anti-HIV phosphoramidate prodrugs"
BioTechnologia, 94 (1), 94, 2013

Conf. 18) M. Materna, M. Sobkowski & J. Stawiński
"Stereochemia czynników fosforylujących zawierających nukleofilowe grupy katalityczne"
Poster presented at "56. Zjazd PTChem i SITPChem" conference, September 16-20, 2013, Siedlce, Poland
Conference proceedings, p. 185

Conf. 19) K. Kołodziej, M. Sobkowski, J. Stawiński & A. Kraszewski
"Nukleozydo (N-arylo)amidofosforany, potencjalne pronukleotydy anty-HIV"
Poster presented at "56. Zjazd PTChem i SITPChem" conference, September 16-20, 2013, Siedlce, Poland
Conference proceedings, p. 402

Conf. 20) M. Sobkowski
"Diastereoselective esterification of ribonucleoside 3'-H-phosphonates"
Oral presentation at "Towards a New RNA Word" conference, November 12-14, 2013, Poznań, Poland

2014

Conf. 21) M. Materna, J. Stawiński & M. Sobkowski
"N-Oxides as reagents in nucleotide chemistry"
In: Collection Symposium Series (M. Hocek, Ed.), 14, 322-323, 2014.
Abstract: N-Oxides were investigated as oxidants of H-phosphonates. These compounds are powerful nucleophiles and are able to substitute leaving groups at phosphorus centers rapidly. The produced putative ammonio intermediates of type P-O-N⁺R₃ collapse rapidly with the O-N bond fission making the N-oxide efficient donors of the oxygen atom to reactive H-phosphonates.

Conf. 22) M. Materna, J. Stawiński & M. Sobkowski
"Trialkylamine N-Oxides as Reagents in Nucleotide Chemistry"
In: Collection Symposium Series (M. Hocek, Ed.), 14, 369-372, 2014.
Abstract: Trialkylamine N-oxides were introduced to the chemistry of phosphorus and nucleotide derivatives. These compounds are powerful nucleophiles and are able to substitute leaving groups in phosphorus centers rapidly. The putatively produced ammonio intermediates of type P-O-N⁺R₃ collapse rapidly with O-N bond fission making the N-oxide investigated efficient donors of the oxygen atom to reactive H-phosphonates and halogenophosphates. Synthetic applications of these reactions are outlined.

Conf. 23) K. Kołodziej, A. Kraszewski, J. Stawiński & M. Sobkowski

"The Loss of TEAH⁺ Cation From Certain Nucleotide Analogues"

In: Collection Symposium Series (M. Hocek, Ed.), 14, 373-374, 2014.

Abstract: During chromatographic purification and evaporation of solvents, triethylammonium salts of phosphate esters and amides may loose the cation yielding new species differing in chemical and physical properties.

 

Conf. 24) J. Romanowska, M. Rożniewska, M. Sobkowski, A. Szymańska-Michalak, A. Dąbrowska, A. Lipniacki, A Piasek, J. Boryski, J. Stawiński & A. Kraszewski
"Wspolczesne koncepcje pronukleotydow anty-HIV"
Poster presented at "57. Zjazd PTChem i SITPChem" conference, September 14-18, 2014, Częstochowa, Poland
Conference proceedings, p. 197

Conf. 25) M. Materna, J. Stawiński & M. Sobkowski
"Trialkylamine N-oxides in nucleotide chemistry"
Oral presentation at "XXI Round Table on Nucleosides, Nucleotides and Nucleic Acids" conference, August 24-28, 2014, Poznań, Poland
Conference proceedings, p. OP17

Conf. 26) M. Materna, J. Stawiński & M. Sobkowski
"N-Oxides in the synthesis of nucleotide analogues"
Poster presented at "XXI Round Table on Nucleosides, Nucleotides and Nucleic Acids" conference, August 24-28, 2014, Poznań, Poland
Conference proceedings, p. 37

Conf. 27) M. Materna, J. Stawiński & M. Sobkowski
"Oxidative dehalogenation of halogenophosphates by N-oxides in the synthesis of nucleotide analogues"
Poster presented at "XXI Round Table on Nucleosides, Nucleotides and Nucleic Acids" conference, August 24-28, 2014, Poznań, Poland
Conference proceedings, p. 38

2015

42) K. Kołodziej, J. Romanowska, J. Stawiński, A. Kraszewski & M. Sobkowski
"The case of triethylammonium cation loss during purification of certain nucleotide analogues. A cautionary note"
Analytical & Bioanalytical Chemistry, 407 (6), 1775-1780, 2015.
Open Access

Abstract: Nucleotides, their analogues, and other phosphate esters and phosphoramidates often contain triethylammonium cation as a counterion. We found that this may be lost during chromatographic purification or concentration of solutions, yielding products in acidic forms or containing sub-stoichiometric amounts of the counterion. This in turn may be detrimental e.g. due to possible decomposition of a compound or inaccurate sample preparation. Correlations between the structure of studied compounds and their susceptibility for cation loss were analyzed. Modifications in preparative techniques were developed to obtain the studied compounds with stoichiometric anion:cation ratios. 

43) M. Sobkowski,A. Kraszewski & J. Stawiński
"Recent Advances in H-Phosphonate Chemistry. Part 1. H-Phosphonate Esters: Synthesis and Basic Reactions"
Topics in Current Chemistry, 361 (Phosphorus Chemistry II), 137-177, 2015.

Abstract: This review covers recent progress in the preparation of H-phosphonate mono- and diesters, basic studies on mechanistic and stereochemical aspects of this class of phosphorus compounds, and their fundamental chemistry in terms of transformation of P–H bonds into P-heteroatom bonds. Selected recent applications of H-phosphonate derivatives in basic organic phosphorus chemistry and in the synthesis of biologically important phosphorus compounds are also discussed.

44) M. Sobkowski, A. Kraszewski & J. Stawiński
"Recent Advances in H-Phosphonate Chemistry. Part 2. Synthesis of C-Phosphonate Derivatives"
Topics in Current Chemistry, 361 (Phosphorus Chemistry II), 179-216, 2015.

Abstract: This chapter provides an overview of recent advances in the development of new methods and protocols for the formation of the P–C bond using H-phosphonate diesters as starting materials. Various chemical and stereochemical aspects of the transition metal-catalyzed cross-coupling and organocatalyst-promoted reactions which are relevant to the synthesis of structurally diverse C-phosphonate derivatives are surveyed. 

45) K. Kołodziej, J. Romanowska, J. Stawiński, J. Boryski, A. Dąbrowska, A. Lipniacki, A. Piasek, A. Kraszewski M. Sobkowski
"Aryl H-Phosphonates 18. Synthesis, properties, and biological activity of 2',3'-dideoxynucleoside (N-heteroaryl)phosphoramidates of increased lipophilicity"
European Journal of Medicinal Chemistry, 100, 77-88, 2015.

Abstract: Recently, AZT (N-pyridyl)phosphoramidates were reported as a new type of potential anti-HIV therapeutics. In continuation of that work, here we present new (N-heteroaryl)phosphoramidate derivatives of antiviral 2',3'-dideoxynucleosides containing other types of N-heteroaryl moieties, particularly those with higher lipophilicity. The present studies comprise mechanistic investigations using ³¹P NMR correlation analysis, which permitted improvements in the synthetic procedures. The obtained compounds were tested in biological systems to establish their cytotoxicity and anti-HIV activity. The results were analyzed with respect to possible correlations between biological and physico-chemical properties of the phosphoramidates studied, to get some insight into their antiviral mode of action.

Conf. 28) M. Materna, J. Stawiński & M. Sobkowski
"Zastosowanie N-tlenków w syntezie analogów nukleotydów"
Poster presented at "58. Zjazd PTChem i SITPChem" conference, September 21-25, 2015, Gdańsk, Poland
Conference proceedings, p. 162

Conf. 29) M. Materna, J. Stawiński & M. Sobkowski
"N-Oxides in the synthesis of nucleotide analogues"
Poster presented at "Recent Advances in Nucleic Acids Therapeutics" conference, October 15-16, 2015, Łódź, Poland
Conference proceedings, p. P6

Pat. 2) A. Kraszewski, J. Romanowska, M. Sobkowski, A. Szymańska-Michalak, J. Stawiński, J. Boryski, A. Lipniacki & A. Piasek
"Nucleotide analogue, method of synthesis of nucleotide analogue, use of nucleotide analogue, antiviral pro-nucleotide, pharmaceutical composition"
Patent US9206209
2016

46) M. Materna, J. Stawiński, A. Kiliszek, W. Rypniewski & M. Sobkowski
"Oxyonium phosphobetaines – unusually stable nucleophilic catalyst-phosphate complexes formed from H-phosphonates and N-oxides"
RSC Advances, 6 (18), 14448-14451, 2016.
Abstract: Aryl H-phosphonates react with N-oxides to form previously unknown stable zwitterionic oxyonium phosphates comprising an ⁻O–P–O–N⁺Z atom system. Their structures were confirmed i.a. by X-ray crystal structure analysis, and some mechanisms were proposed for their formation. Stability during storage and reactivity toward nucleophiles point to their possible synthetic applications.

Conf. 30) M. Materna, J. Stawiński & M. Sobkowski
"Nucleoside oxyonium phosphobetaines – reactive phosphate complexes in new synthetic strategies for biologically active nucleotide analogues"
Poster presented at "XXII Round Table on Nucleosides, Nucleotides and Nucleic Acids" conference, July 18-25, 2016, Paris, France
Conference proceedings, p. 234

Conf. 31) M. Materna, J. Stawiński & M. Sobkowski
"Applications of N-oxides in the chemistry of nucleotide analogues"
Poster presented at "XXII Round Table on Nucleosides, Nucleotides and Nucleic Acids" conference, July 18-25, 2016, Paris, France
Conference proceedings, p. 252
2017

Conf. 32) M. Sobkowski, M. Materna & J. Stawiński
"Nowa chemia N-tlenków amin i zwiazków fosforu"
Invited oral presentation at "60. Zjazd PTChem" conference, September 17-21, 2017, Wrocław, Poland
Conference proceedings, p. 278
2018

Conf. 33) M. Sobkowski, J. Romanowska, K. Kołodziej, M. Rachwalak, T. Jakubowski, J. Gołębiewska, J. Stawiński & A. Kraszewski
"Nowe strategie pronukleotydowe w zwalczaniu wirusa HIV"
Invited oral presentation at "XI Ogólnopolskie Sympozjum Chemii Organicznej" conference, April 8-11, 2011, Warszawa, Poland
Conference proceedings, p. W11
2019

47) J. Romanowska, K. Kołodziej, M. Sobkowski, M. Rachwalak, T. Jakubowski, J. Gołębiewska, A. Kraszewski, J. Boryski, A. Dąbrowska & J. Stawiński
"Aryl H-phosphonates. 19. New anti-HIV pronucleotide phosphoramidate diesters containing amino- and hydroxypyridine auxiliaries"
European Journal of Medicinal Chemistry, 164, 47-58, 2019.
Abstract: We have designed a new type of AZT and ddU phosphoramidate diesters containing various combinations of 2-, 3-, 4-aminopyridine and 2-, 3-, 4-hydroxypyridine moieties attached to the phosphorus center, as potential anti-HIV pronucleotides. Depending on the pKa values of the aminopyridines and the hydroxypyridines used, alternative synthetic strategies based on H-phosphonate chemistry were developed for their preparation. Synthetic aspects of these transformations and the biological activity of the synthesized compounds are discussed.
2020

48) M. Rachwalak, M. Rozniewska, J. Golebiewska, T. Jakubowski, M. Sobkowski & J. Romanowska
"A practical synthesis of nucleoside 5'-diphosphates from nucleoside 5'-H-phosphonate monoesters"
Synthetic Communications, 50, 3836-3844, 2020.
Abstract: A simple and convenient synthetic protocols based onH-phosphonate chemistry have been developed for the preparation of nucleoside 5'-diphosphates. It consists of oxidation of the silylated H-phosphonate monoesters in pyridine with iodine to produce the corresponding N-pyridiniumphosphonate intermediates, followed by their reactions with orthophosphoric acid used as a nucleophile. The added value of the method is that substrates for the reaction, H-phosphonate monoesters, can be prepared in situ from the corresponding nucleosides and used for the next step without isolation. The procedure is simple and applicable to the preparation of various diphosphates of ribo- and deoxyribonucleosides, and their analogues.

49) A. Kraszewski, M. Sobkowski & J. Stawinski
"H-Phosphonate Chemistry in the Synthesis of Electrically Neutral and Charged Antiviral and Anticancer Pronucleotides"
Frontiers in Chemistry, 8, art. no. 595738, 2020.
Open Access
Abstract: In this review a short account of our work on the synthesis and biological activity of electrically neutral and charged anti-HIV and anticancer pronucleotides, presented on the background of the contemporary research in this area, is given.
2021

49) M. Rachwalak, J. Romanowska, M. Sobkowski & J. Stawinski
"Nucleoside Di- and Triphosphates as a New Generation of Anti-HIV Pronucleotides. Chemical and Biological Aspects"
Applied Sciences, 50, art. no. 2248, 2021.
Open Access
Abstract: This review provides a short account of the chemical synthesis of nucleoside di- and triphosphates on a historical background, together with the use of this class of compounds as potential pronucleotides in anti-HIV therapy.

2023

49) M. Materna, J. Stawinski & M. Sobkowski
"A new approach to phosphorylation of nucleosides using oxyonium phosphobetaines as intermediates"
Biotechnologia, 50, 3836-3844, 2020.
Open Access
Abstract: Oxyonium phosphobetaines are recently discovered molecules with a unique ^-O–P–O–N⁺ bond system, which makes them useful and versatile intermediates for the synthesis of phosphates and their derivatives. In this paper, the preliminary data on the application of these compounds in nucleoside phosphorylation were presented.