Mao-Chin Liu

Synthesis of several pyrimidine l-nucleoside analogues as potential antiviral agents

Abstract β- l -5-Iodo-2′-deoxyuridine (β- l -IUdR, 7) and 1-[(β- l - arabinofuranosyl )-E-5-(2- bromovinyl)]uracil (β- l -BV-ara-U, 10) have been synthesized via a multi-step synthesis from l -arabinose. 2′,3′-Dideoxy-β- l -5-azacytidine (18), 2′,3′-dideoxy-β- l -2-thiocytidine (20) and their respective α-anomers, compounds 19 and 21, also were synthesized by direct coupling of 1-O- acetyl -5-O-(tert- butyldimethylsilyl )-2,3-dideoxy- l - ribofuranose (13) with the corresponding silylated bases, in the presence of EtAlCl2 in CH2Cl2, followed by separation of the α- and β-isomers and deblocking of the 5′-protecting groups. In addition, 2′,3′-dideoxy-β- l -5-fluorocytidine (34), a potent anti-HIV and anti-HBV agent, was synthesized by an alternative methodology from 2′,3′-dideoxy-β- l -5-fluorouridine (31) via a 4-triazolylpyrimidinone intermediate. These l -nucleoside analogues were tested in vitro against HIV, HBV, HSV-1, and HSV-2. Among these compounds, 2′,3′-dideoxy-β- l -5-azacytidine (18) was found to show significant activity against HBV in vitro at approximately the same level as 2′,3′-dideoxy-β- d -cytidine (ddC), which is a known potent anti-HBV agent.

1 Chemical and Biological Properties of Cytotoxic α-(N)-Heterocyclic Carboxaldehyde Thiosemicarbazones

Publisher Summary This chapter discusses chemical and biological properties of cytotoxic α-(N)-heterocyclic carboxaldehyde. Ribonucleoside diphosphate reductase is a critical enzyme in the de novo synthesis of the deoxyribonucleotide precursors of DNA and, as such, is essential for cellular replication. Thus, its presence and activity is closely correlated with cellular growth rates. It seems reasonable that a strong inhibitor of ribonucleoside diphosphate reductase would be a useful weapon in the therapeutic armamentarium against cancer. Several different classes of agents are relatively specific inhibitors of ribonucleoside diphosphate reductase. These have included α-(N)-heterocyclic carboxaldehyde thiosemicarbazones (HCTs), hydroxyurea, N-hydroxy-N´-aminoguanidine derivatives and polyhydroxybenzohydroxamates. The HCTs, as a class, are among the most potent known inhibitors of ribonucleoside diphosphate reductase, being 80-5000 times more effective, depending upon the HCT, than hydroxyurea, a clinically useful anticancer agent. The primary metabolic lesion created by the HCTs is interference with the biosynthesis of DNA, an action resulting from the potent inhibition of ribonucleotide reductase activity.

Overexpression of the multidrug resistance genes mdr1, mdr3, and mrp in L1210 leukemia cells resistant to inhibitors of ribonucleotide reductase

L1210 MQ-580 is a murine leukemia cell line resistant to the cytotoxic activity of the alpha-(N)-heterocyclic carboxaldehyde thiosemicarbazone class of inhibitors of ribonucleotide reductase. The line is cross-resistant to etoposide, daunomycin, and vinblastine. L1210 MQ-580 cells expressed 8-fold resistance to 3-aminopyridine-2-carboxaldehyde thiosemicarbazone (3-AP), a relatively newly developed inhibitor of ribonucleotide reductase. The accumulation of [14C]3-AP by L1210 MQ-580 cells was 5- to 6-fold less than by parental L1210 cells. An increased rate of efflux of 3-AP was responsible for the lower steady-state concentration of 3-AP in resistant cells. In reverse transcription-polymerase chain reaction assays, L1210 MQ-580 cells were found to overexpress the multidrug resistance genes mdr1, mdr3, and mrp, but not the mdr2 gene, compared with parental L1210 cells. Measurement of the steady-state concentration of doxorubicin, a potential substrate for both the mdr and mrp gene products, demonstrated that L1210 MQ-580 cells accumulated 4-fold less anthracycline than parental cells. These findings indicate that drug efflux is a major determinant of the pattern of cross-resistance of L1210 MQ-580 cells. To extrapolate these observations to the human homologues of the mdr1, mdr3, and mrp murine genes, the effects of 3-AP were measured in L1210/VMDRC0.06 and NIH3T3 36-8-32 cells transfected with human MDR1 and MRP cDNAs, respectively. The transfectants were 2- to 3-fold resistant to the cytotoxic effects of 3-AP and accumulated less [14C]3-AP than their parental mock-transfected counterparts. Moreover, the cytotoxic activity of 3-AP was significantly greater in two double mrp gene knockout cell lines than in parental W 9.5 embryonic stem cells. Thus, the results suggest that 3-AP is a substrate for both the P-glycoprotein and MRP and that baseline MRP expression has the capacity to exert a protective role against the toxicity of this agent.

Synthesis and biological evaluation of l- and d-configuration 1,3-dioxolane 5-azacytosine and 6-azathymine nucleosides

Novel L- and D-configuration dioxolane 5-azacytosine and 6-azathymine nucleosides have been synthesized and evaluated for biological activity. (-)-(2S,4S)-1-[2-(Hydroxymethyl)-1,3-dioxolan-4-yl]-5-azacytosine (6) showed significant activity against HBV, whereas the D-configuration analogue (14) has been found to exhibit potent anti-HIV activity.

4-nitrobenzyloxycarbonyl derivatives of O(6)-benzylguanine as hypoxia-activated prodrug inhibitors of O(6)-alkylguanine-DNA alkyltransferase (AGT), which produces resistance to agents targeting the O-6 position of DNA guanine.

A series of 4-nitrobenzyloxycarbonyl prodrug derivatives of O(6)-benzylguanine (O(6)-BG), conceived as prodrugs of O(6)-BG, an inhibitor of the resistance protein O(6)-alkylguanine-DNA alkyltransferase (AGT), were synthesized and evaluated for their ability to undergo bioreductive activation by reductase enzymes under oxygen deficiency. Three agents of this class, 4-nitrobenzyl (6-(benzyloxy)-9H-purin-2-yl)carbamate (1) and its monomethyl (2) and gem-dimethyl analogues (3), were tested for activation by reductase enzyme systems under oxygen deficient conditions. Compound 3, the most water-soluble of these agents, gave the highest yield of O(6)-BG following reduction of the nitro group trigger. Compound 3 was also evaluated for its ability to sensitize 1,2-bis(methylsulfonyl)-1-(2-chloroethyl)-2-[(methylamino)carbonyl]hydrazine (laromustine)-resistant DU145 human prostate carcinoma cells, which express high levels of AGT, to the cytotoxic effects of this agent under normoxic and oxygen deficient conditions. While 3 had little or no effect on laromustine cytotoxicity under aerobic conditions, significant enhancement occurred under oxygen deficiency, providing evidence for the preferential release of the AGT inhibitor O(6)-BG under hypoxia.

SYNTHESIS OF HALOGEN-SUBSTITUTED 3-DEAZAADENOSINE AND 3-DEAZAGUANOSINE ANALOGUES AS POTENTIAL ANTITUMOR/ANTIVIRAL AGENTS

Various 2-halogen-substituted analogues (38, 39, 43 and 44), 3-halogen- substituted analogues (51 and 52), and 2′, 3′-dihalogen-substituted analogues (57–60) of 3-deazaadenosine and 3-halogen-substituted analogues (61 and 62) of 3-deazaguanosine have been synthesized as potential anticancer and/or antiviral agents. Among these compounds, 3-deaza-3-bromoguanosine (62) showed significant cytotoxicity against L1210, P388, CCRF-CEM and B16F10 cell lines in vitro, producing IC50 values of 3, 7, 9 and 7,μM, respectively. Several 3-deazaadenosine analogues (38, 51, 57 and 59) showed moderate to weak activity against hepatitis B virus. †Deceased.

A stereospecific synthesis of 2′,3′-dideoxy-β-l-cytidine (β-l-ddC), a potent inhibitor against human hepatitis B virus (HBV) and human immunodeficiency virus (HIV)

Abstract 2′,3′-Dideoxy-β- l -cytidine (β- l -ddC), a potent inhibitor against human hepatitis B virus (HBV) and human immunodeficiency virus (HIV), has been stereospecifically synthesized from l -arabinose in 9 steps.

SYNTHESIS OF A SERIES OF PURINE 2',3'-DIDEOXY-L-NUCLEOSIDE ANALOGUES AS POTENTIAL ANTIVIRAL AGENTS

Abstract Various 2′,3′-dideoxy-L-nucleoside analogues, 6-amino-9-(2,3-dideoxy-β-L-ribofuranosyl) purine (19), 2-chloro-6-amino-9-(2,3-dideoxy-β-L-ribofuranosyl)-purine (20), 2-chloro-6-amino-9-(2,3-dideoxy-4-thio-β-L-ribofuranosyl) purine (21), 2,6-diamino-9-(2,3-dideoxy-β-L-ribofuranosyl) purine (26), 2,6-diamino-9-(2,3-dideoxy-β-thio-β-L-ribofuranosyl)-purine (27), 2-amino-6-chloro-9-(2,3-dideoxy-β-L-ribofuranosyl) purine (28), 6-chloro-9-(2,3-dideoxy-4-thio-β-L-ribofuranosyl) purine (29), and 6-amino-9-(2,3-dideoxy-4-thio-β-L-ribofuran-osyl) purine (30) have been synthesized by coupling of the sodium salt of 2-amino-6-chloropurine (1), 6-chloropurine (2), and 2,6-dichloropurine (3) with 1-O-acetyl-5-O-(tert-butyldimethylsilyl)-2,3-dideoxy-L-ribofuranose (4) or 1-O-acetyl-5-O-(tert-butyldimethylsilyl)-2,3-dideoxy-4-thio-L-ribofuranose (5) in anhydrous MeCN in the presence of either EtAlCl2 or Et2AlCl followed by separation of the α/β-anomers and deprotection of the blocking groups. However, the synthesi...

Synthesis and biological evaluation of 2- and 7-substituted 9-deazaadenosine analogues.

A series of 2-halogen and 7-alkyl substituted analogues of 9-deazaadenosine and 2′-deoxy-9-deazaadenosine was synthesized by new efficient methodology involving transformation of corresponding 9-deazaguanosine and 2′-deoxyguanosine, which in turn were synthesized by direct C-glycosylation of 1-benzyl-9-deazaguanine with 1-O -acetyl-2,3,5-tri-O -benzoyl-d-ribofuranose and methyl 2-deoxy-3,5-di-O -(p -toluoyl)-d-ribofuranoside, respectively. Deoxychlorination of C6 and diazotization/chloro- or fluoro-dediazoniation of the sugar-protected 9-deazaguanosine, followed by selective ammonolysis at C6 and deprotection of the sugar moiety, gave 2-chloro- and 2-fluoro-9-deazaadenosine (6 and 9). Substitution of the 7-position of the dihalogen-intermediate with alkyl groups, followed by ammonolysis and deprotection, provided 2-chloro-7-alkyl-9-deazaadenosines (13a–e) and 2-fluoro-7-benzyl-9-deazaadenosine (13f). Catalytic hydrogenation of 13a–e gave 7-alkyl-9-deazaadenosines 14a–e. Similarly, 2-chloro-2′-deoxy-9-deazaadenosine (21), 2-chloro-2′-deoxy-7-methyl-9-deazaadenosine (25), 2′-deoxy-9-deazaadenosine (22), and 2′-deoxy-7-methyl-9-deazaadenosine (26) were prepared from sugar-protected 2′-deoxy-9-deazaguanosine. Among these compounds, 7-benzyl-9-deazaadenosine (14b) showed the most potent cytotoxic activity, with IC50 values of 0.07, 0.1, 0.2 and 1.5 µM, while both 7-methyl-9-deazaadenosine (14a) and 2-fluoro-9-deazaadenosine (9) also demonstrated significant cytotoxic activity with IC50 values of 0.4, 0.7, 0.3, and 1.5 µM, and 1.5, 0.9, 0.3, and 5 µM against L1210 leukemia, P388 leukemia, CCRF-CEM lymphoblastic leukemia, and B16F10 melanoma cells, respectively.

Structure and Function Converge To Identify a Hydrogen Bond in a Group I Ribozyme Active Site

The determination of how enzymes achieve their catalytic power requires an understanding of how structural motifs are used to position functional groups of enzymes and substrates within active sites. The recent explosion of RNA crystal structures provides an extraordinary opportunity to delve deeply into the relationship between ribozyme structure and function. The Tetrahymena group I ribozyme provides an attractive system for such studies because of the wealth of structural information, with ten crystal structures of group I introns solved in the past five years,[1–5] and extensive functional information[6] that enables incisive analysis of the energetics of catalysis.