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PubMed2004

C-H to N substitution dramatically alters the sequence-specific DNA alkylation, cytotoxicity, and expression of human cancer cell lines.

We designed and synthesized sequence-specific alkylating conjugates 1 and 2, which selectively alkylate matched sequences at nanomolar concentrations. Conjugates 1 and 2 differ only in that the C-H is substituted by an N in the second ring, which precisely recognizes and effectively alkylates DNA according to the recognition rule of Py-Im polyamides. We investigated sequence-specific DNA alkylation, cytotoxicity in 39 human cancer cell lines, and the effect on expression levels in cancer cell lines by Py-Im conjugates 1 and 2. The COMPARE analysis of the mean graphs showed that conjugates 1 and 2 did not correlate well with each other (r = 0.65) despite having a common DNA alkylating mechanism (purine N3 alkylation). Array-based gene expression analysis demonstrated that there are several oppositely regulated genes. The results suggest the intriguing possibility that DNA alkylating agents recognizing longer base-pair sequences may provide a promising approach for developing new types of antigene agents.

Alkylation↗
PubMed1998

DNA-Directed alkylating agents. 7. Synthesis, DNA interaction, and antitumor activity of bis(hydroxymethyl)- and bis(carbamate)-substituted pyrrolizines and imidazoles.

A series of bis(hydroxymethyl)-substituted imidazoles, thioimidazoles, and pyrrolizines and related bis(carbamates), linked to either 9-anilinoacridine (intercalating) or 4-(4-quinolinylamino)benzamide (minor groove binding) carriers, were synthesized and evaluated for sequence-specific DNA alkylation and cytotoxicity. The imidazole and thioimidazole analogues were prepared by initial synthesis of [(4-aminophenyl)alkyl]imidazole-, thioimidazole-, or pyrrolizine dicarboxylates, coupling of these with the desired carrier, and reduction to give the required bis(hydroxymethyl) alkylating moiety. The pyrrolizines were the most reactive alkylators, followed by the thioimidazoles, while the imidazoles were unreactive. The pyrrolizines and some of the thioimidazoles cross-linked DNA, as measured by agarose gel electrophoresis. Strand cleavage assays showed that none of the compounds reacted at purine N7 or N3 sites in the gpt region of the plasmid gpt2Eco, but the polymerase stop assay showed patterns of G-alkylation in C-rich regions. The corresponding thioimidazole bis(carbamates) were more selective than the bis(hydroxymethyl) pyrrolizines, with high-intensity bands at 5'-NCCN, 5'-NGCN and 5'-NCGN sequences in the PCR stopping assay ( indicates block sites). The data suggest that these targeted compounds, like the known thioimidazole bis(carbamate) carmethizole, alkylate exclusively at guanine residues via the 2-amino group, with little or no alkylation at N3 and N7 guanine or adenine sites. The cytotoxicities of the compounds correlated broadly with their reactivities, with the bis(hydroxymethyl)imidazoles being the least cytotoxic (IC50s >1 microM; P388 leukemia) and with the intercalator-linked analogues being more cytotoxic than the corresponding minor-groove-targeted ones. This was true also for the more reactive thioimidazole bis(carbamates) (IC50s 0.8 and 11 microM, respectively), but both were more active than the analogous "untargeted" carmethizole (IC50 20 microM). The bis(hydroxymethyl)pyrrolizine analogues were the most cytotoxic, with IC50s as low as 0.03 microM.

Alkylation↗
PubMedDate not supplied

Thioredoxin alkylation by a dihaloethane-glutathione conjugate.

Glutathione is a thiol-containing tripeptide which functions to protect cellular constituents from endogenous and xenobiotic electrophiles via conjugation and eventual excretion. In the case of compounds such as 1,2-dihaloethanes, however, conjugate formation results in bioactivation of the species rather than detoxification. The conjugate can then act as an alkylating agent toward cellular constituents including DNA, proteins, or lipids. Alkylation of protein thiols in cells exposed to dihaloethane may contribute substantially to the toxicity produced by these compounds. We examined the reactivity of the conjugate S-(2-chloroethyl)-glutathione (CEG) toward the model protein Escherichia coli thioredoxin. At physiological pH, treatment of thioredoxin by CEG resulted in the production of several bands visible on isoelectric focusing, which were determined by matrix-assisted laser desorption ionization (MALDI) mass spectrometry to be mono-, di-, tri-, and tetra-alkylated forms of thioredoxin. A concomitant loss of in vitro enzymatic activity was observed. These products were also observed when reaction was allowed to take place at pH 11.4. Treatment at pH 4.4 resulted in lesser alkylation of thioredoxin, with only the mono- and di-alkylated forms detected. Iodoacetic acid treatment of CEG-alkylated thioredoxin revealed that the iodoacetic acid-susceptible Cys32 was not carboxymethylated, suggesting that this is one of the sites alkylated by CEG.

Alkylating Agents↗
PubMed1999

Effect of cations on the formation of DNA alkylation products in DNA reacted with 1-(2-Chloroethyl)-1-nitrosourea.

The purpose of this study was to examine the influence of cations on the formation of the individual DNA alkylation products derived from 1-(2-chloroethyl)-1-nitrosourea (CNU). Reaction of calf-thymus DNA with [(3)H]CNU in 10 mM triethanolamine buffer produced 13 DNA adducts. Seven of these adducts were identified as N7-(2-hydroxyethyl)guanine, N7-(2-chloroethyl)guanine, 1, 2-(diguan-7-yl)ethane, N1-(2-hydroxyethyl)-2-deoxyguanosine, 1-(N1-2-deoxyguanosinyl)-2-(N3-2-deoxycytidyl)ethane, O(6)-(2-hydroxyethyl)-2-deoxyguanosine, and phosphotriesters. The ratios of the individual products indicated that the chloroethyl and hydroxyethyl adducts are derived from different alkylating intermediates. The influence of cations on the formation of these DNA alkylation products was investigated by the addition of either NaCl, MgCl(2), or spermine. The results demonstrated that (1) the levels of DNA alkylation were inversely proportional to ionic strength, (2) the extent of inhibition was dependent on the alkylation product, and (3) the order of relative effectiveness of inhibition of DNA alkylation by these cations was as follows: spermine > Mg > Na. These results support a model whereby reactions which proceed via an S(N)2 mechanism are more sensitive to the effects of ionic strength than reactions which proceed via an S(N)1 mechanism. In 9L cells treated with CNU, the same alkylation products were formed as in purified DNA; however, the product distribution was different. We interpret this to indicate that within cells, cations modify the reaction of intermediates derived from CNU with DNA.

Alkylating Agents↗
PubMed1989

Transcription-terminating lesions induced by bifunctional alkylating agents in vitro.

The present study was initiated to determine if DNA damage induced by the bifunctional anti-tumor alkylating agents melphalan, nitrogen mustard, a spontaneously activated derivative of cyclophosphamide or chlorambucil inhibits transcription in vitro, and to determine if the potential sites of transcription termination correlate with the sites of N7 guanine adducts predominantly formed by these agents. To assess drug effects on in vitro transcription, linearized plasmid DNA containing the 420-bp PstI fragment of exon two of the human c-myc oncogene was incubated with various concentrations of the drugs. After drug removal and further drug-free incubation, the sense strand of the c-myc insert was transcribed with either of two bacteriophage RNA polymerases in the presence of [32P]UTP. The labeled products of the reaction were electrophoresed next to the labeled products of RNA sequencing reactions, and the location of transcription termination along the DNA template was determined. The sites of transcription termination were then compared with the sites of drug-induced guanine N7 alkylation in the template, as determined by modified Maxam-Gilbert sequencing. At the drug exposures examined, all the drugs were shown to alkylate any guanine in the template. Transcription of this alkylated DNA, however, resulted in RNA molecules truncated not at every alkylated guanine, but at various discrete sites throughout the template. Transcription was terminated at every adenine pair examined in the melphalan-treated template, at selected guanine pairs in the nitrogen-mustard-treated template, and at selected adenine-guanine and guanine-adenine pairs in the chlorambucil-treated template. Transcription of cyclophosphamide-derivative-treated DNA was unaffected. These results suggest that only some bifunctional alkylating agents induce DNA damage capable of terminating transcription in vitro, and that these agents do so in a sequence-specific, drug-specific manner inconsistent with patterns of guanine N7 alkylation.

Alkylating Agents↗
PubMed1985

Different efficiencies of interaction between 3-aminobenzamide and various monofunctional alkylating agents in the induction of sister chromatid exchanges.

3-Aminobenzamide (3AB), an inhibitor of poly(ADP-ribose) polymerase, interacts synergistically with certain monofunctional alkylating agents to increase the frequency of sister chromatid exchanges (SCEs) in Chinese hamster ovary cells (CHO). Not all alkylating agents interact with 3AB to the same extent. Because 3AB has been reported to inhibit DNA strand break rejoining, experiments were carried out to determine if 3AB interacts preferentially with alkylation-induced DNA strand breaks or some other alkylated DNA lesion in the induction of SCEs. SCE frequency was determined in CHO cells that were pretreated with methyl methanesulfonate (MMS), ethyl methanesulfonate (EMS), ethylnitrosourea (ENU) or N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) for 2 h before being incubated with 3AB, and compared to the frequency of DNA strand breaks, as measured by alkaline elution, induced by these treatments. MNNG was by far the most potent DNA strand-breaking and SCE-inducing agent, followed in order of reactivity by MMS, EMS and ENU. The 3AB-enhanced SCE frequency was greatest, however, after MMS treatment. Thus the magnitude of the 3AB-mediated potentiation of SCE induction is not directly related to either the number of DNA strand breaks produced by the alkylating agent or the alkylation-induced SCE frequency. Furthermore, when MMS-treated cells were held in a non-cycling state for 2 days before release and treatment with 3AB, a procedure that results in a 60% reduction in MMS-induced SCE frequency, no synergism between MMS and 3AB in SCE induction was detected. These results suggest that 3AB does not interact with alkylation-induced DNA lesions in the induction of SCEs.

Alkylating Agents↗
PubMed1981

Analysis of alkylated sites at N-3 and N-7 positions of purines as an indicator for chemical carcinogens.

This study reports a rapid assay to distinguish depurination from other forms of alkaline-labile lesions induced in DNA by alkylating agents. Covalently closed circular duplex PM2 DNA was treated with various alkylating agents such as N-methyl-N-nitrosourea, dimethyl sulfate, methyl methanesulfonate, N-ethyl-N-nitrosourea, diethyl sulfate, and ethyl methanesulfonate at pH 6.5. Apurinic sites and subsequent strand breaks were introduced by the hydrolysis of the alkylated purines under nondenaturing conditions by heating alkylated DNA at 70 degrees for 1.5 hr with 0.05 M 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid:KOH (pH 7.4), 0.1 M KCl, 0.01 M MgCl2, 0.0005 M ethylenediaminetetraacetate, 0.05 M glycine, and 0.01 M putrescine. The number of strand breaks produced, representing the alkylated sites at N-3 and N-7 positions of purines, were quantitated by electrophoresis in 1% neutral agarose slab gels. These results were compared with previously reported carcinogenic and mutagenic effects of these compounds, and a correlation between the apurinic sites, the total alkylated sites, and the biological effect of the alkylating agent was determined.

Alkylating Agents↗
PubMed2001

Alkylation kinetics of proteins in preparation for two-dimensional maps: a matrix assisted laser desorption/ionization-mass spectrometry investigation.

All existing protocols for protein separation by two-dimensional (2-D) gel electrophoresis require the full reduction, denaturation, and alkylation as a precondition for an efficient and meaningful separation of such proteins. Existing literature provides a strong evidence to suggest that full reduction and denaturation can be achieved in a relatively short time; the same thing, however, can not be said for the alkylation process, which the present study shows that more than 6 h are required for a complete alkylation. We have used matrix assisted laser desorption/ionisation-time of flight-mass spectrometry (MALDI-TOF-MS) to monitor protein alkylation by iodoacetamide over the period 0-24 h at pH 9. The present, fast and specific MS method provided clear indication on the extent and speed of alkylation which reached approximately 70% in the first 2 min, yet the remaining 30% resisted complete alkylation up to 6 h. The use of sodium dodecyl sulfate (SDS) during the alkylation step resulted in a strong quenching of this reaction, whereas 2% 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate (CHAPS) exerted a much reduced inhibition. The implications of the present measurements on 2-D gel analysis in particular and proteomics in general are discussed.

Alkylation↗
PubMed2004

Cobalt-catalyzed cross-coupling reactions of alkyl halides with allylic and benzylic Grignard reagents and their application to tandem radical cyclization/cross-coupling reactions.

Details of cobalt-catalyzed cross-coupling reactions of alkyl halides with allylic Grignard reagents are disclosed. A combination of cobalt(II) chloride and 1,2-bis(diphenylphosphino)ethane (DPPE) or 1,3-bis(diphenylphosphino)propane (DPPP) is suitable as a precatalyst and allows secondary and tertiary alkyl halides--as well as primary ones--to be employed as coupling partners for allyl Grignard reagents. The reaction offers a facile synthesis of quaternary carbon centers, which has practically never been possible with palladium, nickel, and copper catalysts. Benzyl, methallyl, and crotyl Grignard reagents can all couple with alkyl halides. The benzylation definitely requires DPPE or DPPP as a ligand. The reaction mechanism should include the generation of an alkyl radical from the parent alkyl halide. The mechanism can be interpreted in terms of a tandem radical cyclization/cross-coupling reaction. In addition, serendipitous tandem radical cyclization/cyclopropanation/carbonyl allylation of 5-alkoxy-6-halo-4-oxa-1-hexene derivatives is also described. The intermediacy of a carbon-centered radical results in the loss of the original stereochemistry of the parent alkyl halides, creating the potential for asymmetric cross-coupling of racemic alkyl halides.

Alkylation↗
PubMed1990

Structure-activity studies on organoselenium alkylating agents.

A variety of organoselenium alkylating agents were synthesized, using 2-hydroxyethyl and 3-hydroxypropyl selenocyanate intermediates, and studied to determine their chemical reactivities with 4-(4-nitrobenzyl)pyridine (NBP) and cytotoxicities against CCRF-CEM, L1210/0, and L1210/L-PAM cells. The comparison between the 2-chloroethyl sulfides and selenides 1-4 revealed the markedly enhanced nucleophilicity of selenium (Se) over sulfur (S) by two or more orders of magnitude. This finding indicates that a major consideration in the design of antitumor alkylating organoselenides is the reactivity of selenium. A Taft plot of the experimental first-order rate constant, knbp, and sigma* in a series of 2-chloroethylseleno compounds gave a slope of -1.73 (rho*), with the exception of 2-chloroethyl 2-nitrophenyl selenide (10). The anomalous behavior of 10 is explained in terms of the ortho-nitro stabilization effect directly interacting with the selenium atom of ethyleneselenonium ion to form a 5-membered cyclic intermediate. In the same series, a 5000-fold difference in alkylating reactivity offered only a sixfold variation in cytotoxicity against CCRF-CEM cells. Increasing the alkylating chain length from ethlene to propylene units markedly reduced alkylating reactivities. In the CH3Se(CH2)n Cl series, 16 (n = 3) was 1.5 X 10(5) times slower than 2 (n = 2) in NBP alkylation, revealing that 3-chloro-n-propyl selenides are not chemically reactive enough to be biological alkylating agents despite the presence of the highly nucleophilic selenium atom. Replacement of chloride with mesylate in 3-substituted propyl selenides, such as 17 and 20, restored desirable reactivities and cytotoxicities.

Alkylating Agents↗
PubMed1985

Embryotoxicity induced by alkylating agents: left-sided preponderance of paw malformations induced by acetoxymethyl-methylnitrosamine in mice.

The alkylating agent acetoxymethyl-methylnitrosamine (DMN-OAc) triggers preferential left-sided paw defects in mice following IP administration on either day 11 or 12 of pregnancy. Predominantly, ectrodactyly and hypoplasia of the left paws were found. In an organ culture system, using limb buds of 11-day-old mouse embryos, differentiation is severely impaired following addition of 2 microM DMN-OAc to the culture medium. Left and right limbs are equally affected. In contrast, when DMN-OAc is administered in vivo to the dams with subsequent culturing of the limb buds, growth and differentiation of the left limb buds is more affected when compared to the right. Furthermore, DNA alkylation experiments were performed: in vitro, following addition of (14C)-DMN-OAc (2.3 microM) to the culture medium, the DNA alkylation rate of the limb bud DNA is determined. In vivo, following IP administration of 10 mg/kg DMN-OAc to the dams on day 11 of pregnancy, the extent of DNA alkylation of whole-embryo DNA is similar. However, the DNA alkylation rate of separately pooled left and right limb buds exhibits a two-fold difference according to the different teratogenic susceptibility. The results obtained with both in vivo and in vitro systems are consistent with the thesis that a certain amount of DNA alkylation in the tissue of the embryos is the initial step of alkylating agent-induced teratogenicity.

Abnormalities, Drug-Induced↗
PubMed1991

Fluosol-DA/carbogen with lonidamine or pentoxifylline as modulators of alkylating agents in the FSaIIC fibrosarcoma.

In an effort to increase the efficacy of several antineoplastic alkylating agents (CDDP, L-PAM, CTX, or BCNU), we examined the effect of the modulator Fluosol-DA/carbogen in combination with a second modulator, either lonidamine or pentoxifylline, on the survival of FSaIIC tumor cells and of bone marrow CFU-GM from tumor-bearing C3H mice. Fluosol-DA/carbogen increased the tumor-cell killing activity of each alkylating agent by about 10 times. In contrast, lonidamine alone did not significantly increase the cytocidal activity of any of the alkylating agents tested. However, in combination with Fluosol-DA/carbogen, the use of lonidamine produced about a 100-fold increase in the tumor cell kill achieved with CDDP as compared with CDDP alone. No increase in tumor cell kill over that produced with the single modulator Fluosol-DA/carbogen was seen following the addition of lonidamine to the combination treatment with L-PAM, CTX, or BCNU. Unfortunately, although neither lonidamine nor Fluosol-DA/carbogen alone significantly increased alkylator toxicity to bone marrow CFU-GM, the combination of modulators increased the toxicity of each alkylating agent to bone marrow by about 10 times. Pentoxifylline caused an increase in alkylator activity against the FSaIIC fibrosarcoma only when used with BCNU; this effect was further augmented by the addition of Fluosol-DA/carbogen. The combination of modulators pentoxifylline plus Fluosol-DA/carbogen was more effective than Fluosol-DA/carbogen alone only when the former was used with BCNU, whereas only minimal increases in tumor-cell killing activity were obtained with this modulator combination and CDDP, L-PAM, or CTX. Pentoxifylline increased the bone marrow CFU-GM toxicity of L-PAM by about 10 times. The bone marrow CFU-GM toxicity was further increased by Fluosol-DA/carbogen, as was the toxicity of each of the other alkylating agents. Lonidamine plus Fluosol-DA/carbogen may be useful in increasing the therapeutic efficacy of CDDP, and the combination of pentoxifylline plus Fluosol-DA/carbogen might improve the antitumor activity of BCNU.

Alkylating Agents↗
PubMed1992

Reductive metabolism of diaziquone (AZQ) in the S9 fraction of MCF-7 cells. II. Enhancement of the alkylating activity of AZQ by NAD(P)H: quinone-acceptor oxidoreductase (DT-diaphorase).

The alkylating activity of reduced diaziquone was studied by the nitrobenzylpyridine (NBP) assay and was compared to those of the parent compound and aziridine-containing N,N',N"-triethylenethiophosphoramide (Thio-TEPA). Diaziquone (AZQ) was reduced enzymatically by 2e- using S9 cell fraction from MCF-7 cells which is rich in NAA(P)H:quinone-acceptor oxidoreductase (DT-diaphorase) (QAO) activity. One electron enzymatic reduction was performed with NADPH-cytochrome c reductase. The alkylating activity of AZQ increased 3-fold when reduced by 2e-. This increase was inhibited by dicumarol, an inhibitor of QAO. In contrast, the alkylating activity of AZQ did not increase beyond that of the parent compound when reduced by 1e- using purified NADPH-cytochrome c reductase. Similar results were obtained when AZQ was reduced chemically with borohydride (2e-) and with NADPH (1e-). Anaerobic incubations of AZQ with the S9 fraction of MCF-7 cells (2e- reduction) resulted in an increase in NBP alkylation over its aerobic counterpart (1.8-fold) while maintaining the near 3-fold increase in alkylation over untreated AZQ. In contrast, AZQ incubations with NADPH-cytochrome c reductase (1e- reduction) under the same conditions did not result in an NBP alkylation increase over untreated AZQ. These results indicate that AZQ hydroquinone is most likely the responsible species for the observed alkylation of this antitumor agent to DNA and other nucleophiles. The results also suggest that NAD(P)H:quinone-acceptor oxidoreductase is a very important enzyme in the bioactivation of AZQ.

Alkylation↗
PubMed1991

Dealkylation rates of O6-alkyldeoxyguanosine, O4-alkylthymidine and related compounds in an alkyl-transfer system.

Bacterial O6-alkylguanine-DNA alkyltransferase (AGT) removes alkyl group from O6-alkylguanine and O4-alkylthymine residues in DNA, both of which are considered to be DNA damages most related to the induction of cancer and/or mutation. The repair process involves alkyl-transfer of an O-alkyl group to the active site of the enzyme, where an SH-group of cysteine residue plays the role of alkyl acceptor. In order to elucidate the chemical characteristics of substrates for this enzyme, dealkylation rates of O6-alkyldeoxyguanosine, O4-alkylthymidine and related compounds were measured using an alkyl-transfer system. Thiophenol-triethylamine system was employed as an alkyl acceptor and twenty-one O-alkyl compounds were tested. Dealkylation proceeded with pseudo first order kinetics. The half-life of O6-methyldeoxyguanosine (MedG) was 122 h and no remarkable dependence on N-9 substituents (H, CH3 and deoxyribose) was observed. A compound lacking 2-NH2 group underwent demethylation about three times faster than O6-methylguanines did, while, a compound lacking imidazole moiety underwent demethylation about 2.5 times more slowly. The half-life of O4-methylthymidine (MedT) was 38 h and no remarkable dependence on N-1 (H, CH3 and deoxyribose) and C-5 (H and CH3) substituents was observed. Deethylation proceeded much more slowly than demethylation. Substitution of selenophenol for thiophenol resulted in a 4.5 times faster MedG demethylation rate. Demethylation rates were moderately correlated with values for NMR chemical shift of CH3 group, an indicator of electron density, although the correlation curves of a series of MedG and MedT derivatives were quite different. This result suggests that some different rate-determining factors other than electron density are playing a role. These findings may be of help in resolving the details of the mechanisms of enzymic repair by bacterial and mammalian AGT.

Alkylation↗
PubMed1983

Carbinolamines and related structures--potential alkylating metabolites of clinically active anticancer drugs.

The precise biochemical mechanism by which a number of clinically-active anticancer compounds function remains unclear. Among these are procarbazine (NSC-77213), cyclophosphamide (NSC-26271), streptozotocin (NSC-85998), dacarbazine (NSC-45388), and hexamethylmelamine (NSC-13875). In all cases, there is an N-methyl or N-alkyl substituent which can be or has been shown to generate carbinolamine-like intermediates as a result of oxidative metabolism. Such intermediates can react with amines, imines, sulfhydryls and similar functional groups to form covalent linkages. Thus, carbinolamine metabolites of these clinically-active compounds are proposed as the active agents capable of altering covalently nucleic acids and proteins. It is this alkylating property that may be responsible for these compounds adversely effecting the mitosis of neoplastic cells. Thus, a unifying hypothesis is proposed whereby metabolic hydroxylation of various miscellaneous anticancer agents is the basis for biological activity. In essence, therefore, three broad classes of alkylating agents may be perceived: (1) the classical alkylators such as the nitrogen and sulfur mustards and the sulfonates, (2) bioreductive alkylating agents, and (3) biooxidative alkylating agents such as the carbinolamines. Though the chemical spectrum of each category may be highly diverse, nevertheless, all function as alkylating agents.

Alkylating Agents↗
PubMed1989

Monofunctional alkylating agent-induced S-phase-dependent DNA damage.

Alkylating agents are S-phase-dependent clastogenic agents: Chromosome aberrations are not observed unless the treated cells have first undergone a replicative DNA synthesis. While DNA gaps resulting from misreplication of the alkylated template are believed to underlie aberration formation, the specific alkylated DNA lesions that produce these DNA gaps are not known. To quantitate the DNA strand break induction that results from replication of an alkylated DNA template and attempt to identify those alkylated lesions which underlie DNA strand breakage. [14C]thymidine-labeled Chinese hamster ovary (CHO) cells were treated with either N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) or methyl methanesulfonate (MMS) in G1 and then allowed to progress through S phase in the presence of [3H]thymidine. When analyzed at the subsequent mitosis, DNA strand breaks were found in the nonalkylated ([3H]thymidine-labeled) DNA strand. This did not appear to be the consequence of any recombinational or endonuclease-mediated event and was more likely due to DNA gaps produced by incomplete replication off the alkylated template. A portion of these breaks probably result from a failure to replicate past 3-methyladenine. Differences between MNNG and MMS in the frequency of S-phase-dependent breaks they produce relative to the overall alkylation damage suggest that the O6-methylguanine lesion might also be involved in S-phase-dependent DNA strand breakage.

Alkylating Agents↗
PubMed1991

Modulation of resistance to alkylating agents in cancer cell by gossypol enantiomers.

Several cell lines resistant to alkylating agents possess increased activity of glutathione-S-transferase (GST) drug detoxifying enzymes. Inhibition of certain enzymes of the glutathione redox system may affect cellular sensitivity to alkylators. We report that the (-.)enantiomer of gossypol is a potent and selective inhibitor of GST alpha and GST pi isozymes, and that in combination with buthionine sulfoximine (BSO), causes the enhanced modulation of alkylator resistance in two drug resistant cell lines with increased GST activity. The use of (-)gossypol alone had no effect on the 2-5-fold resistance of MCF-7 Adr and Walker resistant cells to chlorambucil, melphalan and BCNU. Cellular depletion of glutathione with BSO resulted in a 2-4-fold modulation of cell sensitivity to these alkylators. However, the combination of (-)gossypol with BSO resulted in a markedly greater modulation of alkylator sensitivity than with either inhibitor alone. Therefore, the complementary inhibition of glutathione and GST by BSO and (-)gossypol, respectively, produced a synergistic modulation of alkylator cytotoxicity in these drug resistant cell lines. The favorable clinical pharmacokinetics of (-)gossypol suggest its further evaluation for use in combination with BSO and alkylating agents in clinical trials.

Alkylating Agents↗
PubMed2004

Novel semi-synthetic nocathiacin antibiotics: synthesis and antibacterial activity of bis- and mono-O-alkylated derivatives.

Several semi-synthetic bis- and mono-O-alkyl nocathiacin derivatives were synthesized and evaluated for antibacterial activity. Mono-O-alkyl N-hydroxyindole analogues 3a-l were prepared by regioselective alkylation. Bis-O-alkyl nocathiacins 4a-f were obtained by treatment with base and excess electrophile. A one-pot protection-alkylation-deprotection strategy was developed for the preparation of mono-O-alkyl hydroxypyridine analogues 5a,b. Most of the bis- and mono-O-alkyl nocathiacins maintained good in vitro activity but showed reduced in vivo efficacy when compared with the natural product. The excellent in vivo activity and improved water solubility of phosphate analogues 3m and 4g suggest their use as potential pro-drugs.

Alkylation↗