John C. DiCesare

Non-enzymatic hydrolysis of creatine ethyl ester.

The rate of the non-enzymatic hydrolysis of creatine ethyl ester (CEE) was studied at 37 degrees C over the pH range of 1.6-7.0 using (1)H NMR. The ester can be present in solution in three forms: the unprotonated form (CEE), the monoprotonated form (HCEE(+)), and the diprotonated form (H(2)CEE(2+)). The values of pK(a1) and pK(a2) of H(2)CEE(2+) were found to be 2.30 and 5.25, respectively. The rate law is found to be Rate=-dCCEE/dt=k++[H2CEE2+][OH-]+k+[HCEE+][OH-]+k0[CEE][OH-] where the rate constants k(++), k(+), and k(0) are (3.9+/-0.2)x10(6)L mol(-1)s(-1), (3.3+/-0.5)x10(4)L mol(-1)s(-1), and (4.9+/-0.3)x10(4)L mol(-1)s(-1), respectively. Calculations performed at the density functional theory level support the hypothesis that the similarity in the values of k(+) and k(0) results from intramolecular hydrogen bonding that plays a crucial role. This study indicates that the half-life of CEE in blood is on the order of one minute, suggesting that CEE may hydrolyze too quickly to reach muscle cells in its ester form.

Parallel Synthesis of An Oligomeric Imidazole-4,5-dicarboxamide Library

A library of oligomeric compounds was synthesized based on the imidazole-4,5-dicarboxylic acid scaffold along with amino acid esters and chiral diamines derived from amino acids. The final compounds incorporate nonpolar amino acids (Leu, Phe, Trp), polar amino acids (Ser, Asp, Arg), and neutral amino acids (Gly, Ala), and were designed to be useful in screening for inhibitors of protein-protein interactions. Many of the protected and deprotected oligomers show evidence of conformational isomers persistent at room temperature in aqueous solution. A total of 317 final oligomers, out of 441 targeted compounds, were obtained in high analytical purity and of sufficient quantity to submit them for high-throughput screening as part of the NIH Roadmap.

Induction of Cell Death by a Novel Naphthoquinone Containing a Modified Anthracycline Ring System

The novel naphthoquinone adduct 12,13‐Dihydro‐N‐methyl‐6,11,13‐trioxo‐5H‐benzo[4,5]cyclohepta[1,2‐b]naphthalen‐5,12‐imine (hereafter called TU100) was synthesized as a potential chemotherapeutic agent. TU100 arrests tissue culture cells in S and G2/M phases of the cell cycle, followed by rapid induction of apoptosis. Evaluation by the Developmental Therapeutics Program at the National Cancer Institute revealed TU100 differentially inhibits growth of tissue‐specific human cancer cell lines and has in vivo efficacy in a hollow fiber assay. These data were evaluated against previously analyzed compounds using the COMPARE algorithm and predicted that TU100 has a unique mechanism of action. Further analysis revealed TU100 does not intercalate into DNA despite structural similarity to anthracyclines. Cells treated with the drug do exhibit DNA damage, however, as indicated by phosphorylation of histone H2A.X. This damage and effects on cell viability are likely mediated in part by TU100‐induced reactive oxygen species. Based on these results, TU100 shows promise as a chemotherapeutic drug owing to its unique structure, cellular targets, and efficacy against selected panels of tissue‐specific cancer cell lines.

Progress in Developing Nerve Agent Sensors Using Combinatorial Techniques

Development of a sensor capable of selective detection of specific nerve agents is imperative in today’s atmosphere of terrorism. The sensor needs to be inexpensive, portable, reliable, absent of false positives and available to all military and first responders. By utilizing the techniques of molecular imprinting, combinatorial chemistry, silica sol-gel synthesis and lanthanide luminescence, a sensor for the detection of the hydrolysis product of the nerve agent soman is being developed. There are many parameters that require investigation in order for the sensor to become a reality. These parameters include 1) the selection of a chelate that can bind to the lanthanide and anchor the nerve agent simulant during the formation of the molecularly imprinted polymer, 2) the determination of the environment best suited for this complex formation, 3) the formation, as well as modification of the silica sol-gel for molecular imprinting to take place, and 4) the proper quantity and ratios of monomers used to create the three dimensional imprint. Key to the success of optimizing these parameters is the development of a combinatorial assay that allows for the synthesis and testing of tens of thousands of combinations of parameters. Work on the development of the combinatorial assay has lead to a method of preparing thin film polymers capable of analyzing the presence of nerve agent simulants. Current work is underway to validate the combinatorial assay and to synthesize and evaluate a library of sensor materials selective for nerve agents.

Parallel Synthesis of a Library of Symmetrically- and Dissymmetrically-disubstituted Imidazole-4,5-dicarboxamides Bearing Amino Acid Esters

The imidazole-4,5-dicarboxylic acid scaffold is readily derivatized with amino acid esters to afford symmetrically- and dissymmetrically-disubstituted imidazole-4,5-dicarboxamides with intramolecularly hydrogen bonded conformations that predispose the presentation of amino acid pharmacophores. In this work, a total of 45 imidazole-4,5-dicarboxamides bearing amino acid esters were prepared by parallel synthesis. The library members were purified by column chromatography on silica gel and the purified compounds characterized by LC-MS with LC detection at 214 nm. A selection of the final compounds was also analyzed by 1H-NMR spectroscopy. The analytically pure final products have been submitted to the Molecular Library Small Molecule Repository (MLSMR) for screening in the Molecular Library Screening Center Network (MLSCN) as part of the NIH Roadmap.

Parallel Synthesis of an Imidazole-4,5-dicarboxamide Library Bearing Amino Acid Esters and Alkanamines

The imidazole-4,5-dicarboxylic acid scaffold is readily derivatized with amino acid esters and alkanamines to afford compounds with intramolecularly hydrogen bonded conformations that mimic substituted purines and therefore are hypothesized to be potential inhibitors of kinases through competitive binding to the ATP site. In this work, a total of 126 dissymmetrically disubstituted imidazole-4,5-dicarboxamides with amino acid ester and alkanamide substituents were prepared by parallel synthesis. The library members were purified by column chromatography on silica gel and the purified compounds characterized by LC-MS with LC detection at 214 nm. A selection of the final compounds was also analyzed by 1H-NMR spectroscopy. The analytically pure final products have been submitted to the Molecular Library Small Molecule Repository (MLSMR) for screening in the Molecular Library Screening Center Network (MLSCN) as part of the NIH Roadmap.

Modification of the Titanium(IV) Isopropoxide Reductive Amination Reaction: Application to Solid Phase Synthesis

Abstract Secondary amines are synthesized by a modification of the titanium(IV) isopropoxide reductive amination reaction. The modified procedure was utilized for both solution and solid phase synthesis with excellent purity and yields.

The Synthesis of 1-(4-Triethoxysilyl)phenyl)-4,4,4-trifluoro-1,3-butanedione, a Novel Trialkoxysilane Monomer for the Preparation of Functionalized Sol-gel Matrix Materials

The title compound, 1-(4-triethoxysilyl)phenyl)-4,4,4-trifluoro-1,3-butanedione, was synthesized in a three-step sequence starting from 2-(4-bromophenyl)propene. Containing both a trialkoxysilyl and a substituted 1,3-butanedione functional grouping within its structure, this new silane is a viable starting material for the preparation of functionalized sol-gel materials.

LARGE SCALE REDUCTIVE CLEAVAGE OF DIBENZOTHIOPHENE

A modified procedure for the large scale reductive cleavage of dibenzothiophene to prepare 2-phenylthiophenol without the formation of the over reduction products, hydrogen sulfide and biphenyl, has been developed. 2-Phenylthiophenol is an important intermediate in the hydrodesulfurization of dibenzothiophene, a common component of crude oil (1). Knowledge of the precise thermochemical properties of 2-phenylthiophenol is needed to develop improvements in refinery design and other petroleum processing technology. A large, high purity (>99.9%) sample of 2phenylthiophenol was needed for thermodynamic studies (2). 2-Phenylthiophenol can be synthesized directly from dibenzothiophene by reductive cleavage using lithium or lithium biphenyl as the reducing agent followed by addition of aqueous acid to quench the resulting dianion. The yields of these reactions were modest, 40 60%, for the purified product (1a, 3). Our attempts to synthesize 2-phenylthiophenol by the lithium biphenyl method resulted in similar yields. The formation of hydrogen sulfide upon quenching with aqueous acidic solution was an indication of over reduction to form biphenyl. Hydrogen sulfide is an OSHA regulated chemical with exposure limits in the 10-50 ppm range (4). A procedure that eliminated the formation of hydrogen sulfide would be beneficial, especially for larger scale preparations. The formation of biphenyl and hydrogen sulfide could occur either in the initial addition of reagents or as a competitive process during the course of the reaction. In order to elucidate when the formation was occurring, lithium naphthaiide was utilized as the reducing agent (5). The change in reducing agent was needed to determine when biphenyl was being produced, since the end product of lithium biphenyl is the same as the over reduction product. The progress of the reaction was monitored by quenching aliquots with aqueous acid and analyzing the ratio of dibenzothiophene, naphthalene, 2-phenylthiophenol and biphenyl formed in the reaction. It was found that biphenyl was being formed early in the course of the reaction. The Vol. 6, No. I, 2000 Large scale reductive cleavage of dibenzothiophene literature procedures call for adding dibenzothiophene to a 0 °C solution of reducing agent. This procedure allows for excess reducing agent to be present during the addition of the dibenzothiophene, a situation facilitating over reduction especially during large scale reactions where addition times are greatly increased. Attempts to reverse the addition of reagents by adding the reducing agent to the cooled solution of dibenzothiophene by cannula resulted in some initial formation of the biphenyl dianion presumably from localized high concentration gradients of lithium naphthalide. It was discovered that cooling the reaction below -20 °C effectively stopped all reactions from taking place. Therefore, cooling a solution of lithium naphthalide to below -20 °C and maintaining the temperature while adding the dibenzothiophene via an addition funnel afforded a large scale preparation of 2-phenylthiophenol without the formation of biphenyl or hydrogen sulfide. After complete addition of dibenzothiophene, GC analysis showed that only starting material was present in the solution. The solution was allowed to warm to 0 °C and stirred until the reaction was complete. The solution was diluted with ethyl acetate and extracted with 1N NaOH, the aqueous solution was neutralized with 3N HCl and extracted with ethyl acetate. The final product was recrystallized from 2-propanol. Isolated and purified yields as high as 92% on the one mole scale have been achieved. Acknowledgement We thank the U.S. Department of Energy for funds through subcontracts DE-RP22-92PC11008 and G4S60381 at the National Institute for Petroleum and Energy Research, Bartlesville, OK. References (1) a) P.R. Stafford, T.B. Rauchfuss, A.K. Verma and S.R. Wilson, J. Organomet. Chem., 526. 203-214 (1996). b) C. Bianchini, J.A. Casares, A. Meli, V. Sernau, F. Vizza and R.A. Sancez-Delagdo, Polyhedron, 18, 3099-3114 (1997). c) J.J. Garcia, A. Arevalo, V. Montiel, F. Del Rio, B. Quiroz, H. Adams and P.M. Maitlis, Organometallics, 16, 3216-3220 (1997); (2) The thermodynamic properties will be reported by personnel of the National Institute for Petroleum and Energy Research, Bartlesville, OK 74005. (3) a) D.D. Ridley, and M.A. Smal, Aust. J. Chem., 36, 795-802 (1983). b) J.J. Eisch, J. Org. Chem., 28, 707-710 (1963) c) H. Gilman and J.J. Dietrich, J. Org. Chem., 22, 851-853 (1957). (4) Occupational Safety & Health Administration: Standard Number: 1910.1000 TABLE Z-2 (5) J. March, Advanced Organic Chemistry, 4 ed, John Wiley, New York, 1992, pp 729. Received on December 13, 1999