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R L Kisliuk

Publications and source records attributed to R L Kisliuk.

At least 55 records · Page 3Linked to original sources
PubMed1986

Synthesis and biological activity of resolved C-10 diastereomers of 10-methyl- and 10-ethyl-10-deazaminopterin.

Synthesis and evaluation of the antitumor drugs 10-methyl- and 10-ethyl-10-deazaminopterin (15a,b) were previously reported for the diastereomeric mixtures, lacking resolution at the C-10 position. In order to assess biological properties of the individual diastereomers, the C-10 isomers of 4-amino-4-deoxy-10-methyl- and 10-ethyl-10-deazapteroic acids (13a,b) were prepared by total synthesis. Coupling with L-glutamate afforded the appropriate diastereomers of the title compounds. Biochemical, transport, and cell growth inhibitory properties in L1210 cells and folate-dependent bacteria were measured. Differences were generally less than 2-fold between diastereomeric pairs, but a factor of 3 was noted for d,L-15b vs. l,L-15b in inhibition of DHFR from L1210 cells and in cytotoxicity toward L1210 cells. An in vivo comparison of the isomers of 15b with racemic compound against L1210 in mice did not show a significant efficacy difference (ILS) among the compounds. However, d,L-15b showed an acute toxicity about 2.5 times that of l,L-15b.

Aminopterin↗
PubMed1986

Syntheses and antifolate activity of 5-methyl-5-deaza analogues of aminopterin, methotrexate, folic acid, and N10-methylfolic acid.

Evidence indicating that modifications at the 5- and 10-positions of classical folic acid antimetabolites lead to compounds with favorable differential membrane transport in tumor vs. normal proliferative tissue prompted an investigation of 5-alkyl-5-deaza analogues. 2-Amino-4-methyl-3,5-pyridinedicarbonitrile, prepared by hydrogenolysis of its known 6-chloro precursor, was treated with guanidine to give 2,4-diamino-5-methylpyrido[2,3-d]pyrimidine-6-carbonitrile which was converted via the corresponding aldehyde and hydroxymethyl compound to 6-(bromomethyl)-2,4-diamino-5-methylpyrido[2,3-d]pyrimidine. Reductive condensation of the nitrile 8 with diethyl N-(4-amino-benzoyl)-L-glutamate followed by ester hydrolysis gave 5-methyl-5-deazaaminopterin. Treatment of 12 with formaldehyde and Na(CN)BH3 afforded 5-methyl-5-deazamethotrexate, which was also prepared from 15 and dimethyl N-[(4-methylamino)benzoyl]-L-glutamate followed by ester hydrolysis. 5-Methyl-10-ethyl-5-deazaaminopterin was similarly prepared from 15. Biological evaluation of the 5-methyl-5-deaza analogues together with previously reported 5-deazaaminopterin and 5-deazamethotrexate for inhibition of dihydrofolate reductase (DHFR) isolated from L1210 cells and for their effect on cell growth inhibition, transport characteristics, and net accumulation of polyglutamate forms in L1210 cells revealed the analogues to have essentially the same properties as the appropriate parent compound, aminopterin or methotrexate (MTX), except that 20 and 21 were approximately 10 times more growth inhibitory than MTX. In in vivo tests against P388/0 and P388/MTX leukemia in mice, the analogues showed activity comparable to that of MTX, with the more potent 20 producing the same response in the P388/0 test as MTX but at one-fourth the dose; none showed activity against P388/MTX. Hydrolytic deamination of 12 and 20 produced 5-methyl-5-deazafolic acid and 5,10-dimethyl-5-deazafolic acid, respectively. In bacterial studies on the 2-amino-4-oxo analogues, 5-deazafolic acid proved to be a potent inhibitor of Lactobacillus casei DHFR and also the growth of both L. casei ATCC 7469 and Streptococcus faecium ATCC 8043. Its 5-methyl congener 22 is also inhibitory toward L. casei, but its IC50 for growth inhibition is much lower than its IC50 values for inhibition of DHFR or thymidylate synthase from L. casei, suggesting an alternate site of action.

Aminopterin↗
PubMed1986

Folate analogues. 25. Synthesis and biological evaluation of N10-propargylfolic acid and its reduced derivatives.

N10-Propargylfolic acid (2), which is the closest pteridine analogue of the thymidylate synthase inhibitor N10-propargyl-5,8-dideazafolic acid (PDDF), was synthesized starting from diethyl [p-(N-propargylamino)benzoyl]-L-glutamate (5) and N-(3-bromo-2-oxopropyl)phthalimide (8). The 7,8-dihydro derivative of propargylfolic acid served as a synthetic substrate of Lactobacillus casei dihydrofolate reductase. Propargylfolic acid and its reduced derivatives were weak inhibitors of L. casei thymidylate synthase compared to PDDF. All derivatives of propargylfolate were active against the growth of Streptococcus faecium, but with the exception of 7,8-dihydropropargylfolic acid, all were inactive against L. casei. Although less potent than PDDF, marked inhibition of thymidylate synthase by 2 was observed in permeabilized L1210 cells.

Animals↗
PubMed1986

Folate analogues. 26. Syntheses and antifolate activity of 10-substituted derivatives of 5,8-dideazafolic acid and of the poly-gamma-glutamyl metabolites of N10-propargyl-5,8-dideazafolic acid (PDDF).

The poly-gamma-glutamyl derivatives of n10-propargyl-5,8-dideazafolic acid (PDDF) with a chain length of up to five glutamate residues were synthesized from N10-propargyl-5,8-dideazapteroic acid by the solid-phase procedure. These compounds were evaluated for their antifolate activity using folate-requiring microorganisms and intact and permeabilized L1210 cells and as inhibitors of dihydrofolate reductase and thymidylate synthase derived from L. casei. The polyglutamylated derivatives of PDDF (1) were more active than the parent compound in inhibiting the growth of L. casei, thymidylate synthesis in permeabilized L1210 cells, and L. casei thymidylate synthase. Two analogues of 5,8-dideazafolic acid (2 and 3), one with a 2-butyne and another with a cyclopropylmethyl substituent at N10, were also synthesized and evaluated for their antifolate activities using the above-mentioned test systems. They were considerably less active than PDDF or its polyglutamylated derivatives. N10-Propargyl-5,8-dideazapteroyl tri-, tetra-, and pentaglutamates were equipotent with 5-fluorodeoxyuridylate as inhibitors of thymidylate synthesis in permeabilized L1210 cells. The polyglutamyl metabolites of PDDF were shown to be the most potent antifolate inhibitors of L. casei and L1210 thymidylate synthases yet described.

Animals↗
PubMed1986

Synthesis and antifolate properties of 5,10-methylenetetrahydro-8,10-dideazaminopterin.

The synthesis of the 5,10-methylene analogue of 5,6,7,8-tetrahydro-8,10-dideazaminopterin, a potential dual inhibitor of dihydrofolate reductase (DHFR) and thymidylate synthase (TS) enzymes, is described. The dimethyl ester of 10-carboxy-4-amino-4-deoxy-8,10-dideazapteroic acid was converted to the tetrahydro derivative by hydrogenation. Thermally induced cyclization of the 10-carbomethoxy and the 5-NH groups afforded the 5,10-carbonyl analogue. Reduction of the lactam with borane readily yielded the key 5,10-methylene-4-amino-4-deoxy-8,10-dideazatetrahydropteroic acid methyl ester. Saponification of the benzoate ester and coupling with L-glutamate concluded the synthesis. The title compound was a modest inhibitor of growth in folate-dependent bacteria. Streptococcus faecium and Lactobacillus casei, but inhibition of DHFR or TS derived from L. casei was poor. The compound was also a weak inhibitor of DHFR derived from L1210 murine leukemia and was a weak inhibitor of L1210 growth in culture.

Aminopterin↗
PubMed1986

Interaction of polyglutamyl derivatives of methotrexate, 10-deazaaminopterin, and dihydrofolate with dihydrofolate reductase.

Polyglutamyl derivatives of methotrexate (MTX) and 10-deazaaminopterin (10-DAM) containing a total of one through six glutamate residues (Glu residues) were tested as inhibitors of dihydrofolate reductase (DHFR) derived from sheep, chicken, and beef liver. The ability of dihydropteroylpentaglutamate to antagonize the inhibitory activity of these analogues was also studied. The most striking effects were seen with sheep liver DHFR, where polyglutamylation of MTX causes stepwise decreases in the concentration required for 50% inhibition (IC50) with each additional Glu residue until MTX with a total of six Glu residues has an IC50 value 1/3 that of MTX. With 10-DAM the pattern is more complex. The IC50 values increase with addition of Glu residues until a maximum is reached with 10-DAM having a total of three Glu residues which has a value twice that of 10-DAM. 10-DAM with a total of four Glu residues and 10-DAM with a total of five Glu residues have progressively lower IC50 values, the latter being equipotent with 10-DAM. With dihydropteroylpentaglutamate as substrate instead of dihydrofolate, the IC50 values are increased 2- to 5-fold for both MTX and 10-DAM derivatives. The results obtained with chicken liver and beef liver DHFR are generally similar to those described for the sheep liver enzyme, but the effects of polyglutamylation are less pronounced. The addition of 0.2 M KCl to the assay system reduces the differences in inhibitory potency of the polyglutamyl derivatives with all three enzymes tested. We conclude that polyglutamylation can alter the interaction of folate analogues and dihydrofolate with DHFR.

Aminopterin↗
PubMed1984

Synthesis and antifolate properties of 10-alkyl-8,10-dideazaminopterins.

The synthesis of 10-alkyl analogues of the potent antitumor agent 8,10-dideazaminopterin is described. Alkylation of appropriate alpha-alkyl homoterephthalate esters with 2,4-diamino-6-(bromomethyl)-8-deazapteridine afforded 10-alkyl-10-carboxy-4-amino-4-deoxy-8,10-dideazapteroic acid diesters. Ester cleavage and decarboxylation at C-10 were accomplished by heating with sodium cyanide in Me2SO at 170-180 degrees C to afford the 2,4-diamino-10-alkyl-8,10-dideazapteroic acids. The acids were coupled with diethyl glutamate, followed by saponification, to give the 10-alkyl-8,10-dideazaminopterins. The compounds were potent inhibitors of growth in folate-dependent bacteria, Streptococcus faecium and Lactobacillus casei. The 10-methyl and 10-ethyl analogues gave the highest percent increases in life span for mice infected with L1210 leukemia with ILS values of +203 and +235%, respectively.

Aminopterin↗
PubMed1983

The interaction of pteroylpolyglutamates with calf thymus thymidylate synthase.

Calf thymus thymidylate synthase was purified to homogeneity using a pteroyltetraglutamyllysine Sepharose affinity column. Inhibition of the purified enzyme by folate or methotrexate was enhanced by addition of gamma-Glu residues. Tetrahydrofolate, tetrahydropteroyltriglutamate and tetrahydropteroylheptaglutamate, all having the natural configuration at C-6, all showed similar Km values near 15 microM. Thus, although calf thymus thymidylate synthase showed a higher affinity for pteroylpolyglutamates than pteroylmonoglutamate, this was not reflected in lowered Km values with the corresponding tetrahydropteroylpolyglutamate substrates.

Animals↗
PubMed1983

Folate analogues. 20. Synthesis and antifolate activity of 1',2',3',4',5',6'-hexahydrohomofolic acid.

The synthesis of 1',2',3',4',5',6'-hexahydrohomofolic acid (3), a close analogue of homofolic acid (2), has been carried out by replacement of the benzene ring of 2 with a cyclohexane ring. The synthetic methods employed here were based on the Boon-Leigh strategy to obtain products with unambiguous structures. Based on a number of chemical and spectral observations, a tentative cis stereochemistry was assigned to the 1,4-substituents of the cyclohexane ring of both the homopteroate analogue 13 and the target compound 3. We investigated hexahydrohomopteroic acid (13), hexahydrohomofolic acid (3), and their 7,8-dihydro and d,l-5,6,7,8-tetrahydro derivatives for antifolate activity employing several biological test systems. The dihydro and tetrahydro derivatives of both 13 and 3 were active against Streptococcus faecium, whereas they were inactive against Lactobacillus casei. These compounds were neither substrates nor inhibitors of L. casei dihydrofolate reductase or thymidylate synthase.

Biological Assay↗
PubMed1983

Folate analogues. 21. Synthesis and antifolate and antitumor activities of N10-(cyanomethyl)-5,8-dideazafolic acid.

A close analogue of the antileukemic agent 5,8-dideaza-N10 propargylfolic acid (2) was synthesized by replacing the propargyl moiety of 2 with a cyanomethyl group. This compound, N10-(cyanomethyl)-5,8-dideazafolic acid (3), was evaluated for its antifolate and antitumor activities in several biological test systems. Alkylation of diethyl N-(4-aminobenzoyl)-L-glutamate with bromoacetonitrile gave diethyl N-[4-[(cyanomethyl)amino]benzoyl]-L-glutamate (7). Reaction of 7 with 2 amino-6-(bromomethyl)-4-hydroxyquinazoline (9) in dimethylacetamide gave the corresponding diethyl ester 11, which was hydrolyzed to the target compound 3. The known antileukemic agent 2 was also synthesized for comparative studies by employing a modified procedure, which resulted in a better yield of this product. Both compounds 2 and 3 were evaluated for their antifolate activities by using two folate-requiring microorganisms, Streptococcus faecium and Lactobacillus casei. They were further evaluated as inhibitors of thymidylate synthase and dihydrofolate reductase derived from the above organisms, as well as for their antitumor activity by using selected tumor cells in culture. Compound 2 was found to be as equally potent as methotrexate (MTX) against S. faecium, and it was an excellent inhibitor of L. casei thymidylate synthase. The cyanomethyl analogue 3 was less active than 2 in all the test systems, except the inhibition of dihydrofolate reductase.

Antineoplastic Agents↗
PubMed1983

Folate analogues. 22. Synthesis and biological evaluation of two analogues of dihydrofolic acid possessing a 7,8-dihydro-8-oxapterin ring system.

Two analogues of dihydrofolic acid possessing a 7,8-dihydro-8-oxapterin ring system have been synthesized and evaluated for their antifolate activities. These compounds, N-[(2-amino-4-hydroxy-7,8-dihydro-8-oxa-6-pteridinyl)benzoyl]-L-glutamic acid (3) and N-[[(2-amino-4-hydroxy-7,8-dihydro-8-oxa-6-pteridinyl) methyl]benzoyl]-L-glutamic acid (4), were synthesized by reacting the appropriately substituted alpha-halo ketones with 2,5-diamino-4,6-dihydroxypyrimidine (2). Elaboration of p-carbomethoxybenzaldehyde (5) to p-carbomethoxyphenacyl bromide (7) was accomplished by its oxidation with Jones reagent and the successive treatment of the oxidation product with SOCl2, CH2N2, and HBr. Commercially available p-vinylbenzoic acid (11) was converted to its glutamate conjugate 12 and was further converted to the bromo ketone, diethyl N-[p-(1-bromo-2-oxopropyl)benzoyl]-L-glutamate (17), by a series of reactions involving epoxidation, oxirane ring opening with HBr, Jones oxidation, Zn/HOAc reduction, and successive treatment of the reduction product 16 with SOCl2, CH2N2, and HBr. These bromo ketones, 7 and 17, upon reaction with pyrimidine 2, gave the diethyl esters of the target compounds, which were hydrolyzed to 3 and 4 with NaOH. Compound 4 underwent an interesting acid-catalyzed isomerization where the double bond of 4 was shifted from the 5,6-position to the 6,9-position to give the isomer 19. Both compounds 3 and 4 were inactive against Lactobacillus casei (ATCC 7469) and did not serve as synthetic substrates of L. casei dihydrofolate reductase. Compound 4 showed activity against Streptococcus faecium (ATCC 8043), but 3 was inactive against this organism.

Enterococcus faecalis↗
PubMed1983

Association of RNA with thymidylate synthase from methotrexate-resistant Streptococcus faecium.

Thymidylate synthase (5,10-methylenetetrahydrofolate:dUMP C-methyltransferase, EC 2.1.1.45) from methotrexate-resistant Streptococcus faecium has a UV absorbance peak at 259 nm and stains with acridine orange because of the presence of RNA on the protein. Material having an absorbance peak at 254 nm, obtained from the enzyme by phenol extraction, is degraded by treatment with pancreatic RNase, T1 RNase, and alkali but is stable to DNase. Dowex-1 chromatography of the pure enzyme yields two polynucleotide fragments in addition to the apoenzyme. As estimated from their absorbance, these fragments contain 4 and 11 mononucleotide residues per mole of enzyme, respectively. In crude extracts, thymidylate synthase is associated with rapidly sedimenting material that is sensitive to RNase. Treatment of crude extracts with RNase, as is done routinely during thymidylate synthase purification, most likely results in the formation of the small polynucleotides found on the enzyme. The RNA is not required for enzyme activity.

Drug Resistance, Microbial↗
PubMed1982

Further studies stereospecificity at carbon 6 for membrane transport of tetrahydrofolates. Diastereoisomers of 5-methyltetrahydrofolates as competitive inhibitors of transport of methotrexate in L1210 cells.

The unnatural d diastereoisomer at carbon 6 of 5-methyltetrahydrofolate was only slightly less effective than the natural 1 diastereoisomer as a competitive inhibitor of the carrier-mediated membrane transport of [3H]methotrexate into L1210 murine leukemia cells. The apparent Ki for a mixture containing equal amounts of both natural and unnatural diastereoisomers was not significantly different from that found for the unnatural form. These results show that the reduced folate carrier system in these cells has a strong affinity for the unnatural stereoisomer, a finding in contrast to that obtained with the corresponding diastereoisomer of 5-formyltetrahydrofolate.

Animals↗
PubMed1982

10-Propargylaminopterin and alkyl homologues of methotrexate as inhibitors of folate metabolism.

Reported antifolate activity against leukemia L1210 by N-[14-[[(2-amino-4-hydroxy-6-quinazolinyl)methyl]-propargylamino]benzoyl]]-L-glu tamic acid through potent inhibition of thymidylate synthase (EC 2.1.1.45) prompted us to include the propargyl group in a study of the effect on folate metabolism and membrane transport of replacing the 10-methyl group of methotrexate with other groups. Along with the propyl (8a) and octyl (8b) homologues of methotrexate, the propargyl compound 8c was prepared for evaluation. Syntheses of 8a,b were achieved by a standard multistep sequence involving preparation of the side-chain precursors via tosylated intermediates and then their alkylation with 6-(bromomethyl)-2,4-pteridinediamine hydrobromide. The side-chain precursor to 8c was prepared by direct alkylation of diethyl N-(4-aminobenzoyl)-L-glutamate with propargyl bromide and was separated from unchanged amine and dipropargyl coproduct by a combination of methods, including dry-column chromatography and recrystallization. Subsequent steps leading to 8c were like those used to prepare 8a,b. Biological evaluations of the three compounds consisted of studies of their effects on enzyme inhibition [(dihydrofolate reductase (EC 1.5.1.3) and thymidylate synthase)], L1210 cell growth inhibition, cellular membrane transport with various murine cell types (L210, S180, Ehrlich, and epithelial), in vivo (mice) activity vs. L1210 leukemia and S180 ascites, and plasma clearance in mice. The in vivo results vs. S180 ascites offered evidence that 8c might have a better therapeutic index against this tumor than methotrexate, but no other result from either of these compounds suggested significant superiority over methotrexate.

Aminopterin↗
PubMed1982

Synthesis and antitumor activity of 10-alkyl-10-deazaminopterins. A convenient synthesis of 10-deazaminopterin.

Requirements for large-scale synthesis of the potent antitumor drug 10-deazaminopterin have led to development of a facile synthesis of this compound and its 10-alkyl analogues. The lithium diisopropyl amide generated dianions of appropriate p-alkylbenzoic acids were alkylated with 3-methoxyallyl chloride. The resulting 4-(p-carboxyphenyl)-1-methoxy-1-butenes were brominated at pH 7-8 to afford the 2-bromo-4-(p-carboxyphenyl)butyraldehydes. Condensation with 2,4,5,6-tetraminopyrimidine and subsequent in situ oxidation of the resulting dihydropteridines yielded crystalline 10-alkyl-10-deaza-4-amino-4-deoxypteroic acids. The pteroic acids were coupled with diethyl glutamate via the mixed anhydride method, followed by saponification at room temperature, to give the target 10-deazaminopterins. The 10-alkyl compounds were approximately equipotent to 10-deazaminopterin as growth inhibitors of folate-dependent bacteria. Their abilities to inhibit Lactobacillus casei and L1210 derived dihydrofolate reductases were also similar. Transport properties in vitro were suggestive of an improved therapeutic index for the 10-alkyl analogues. Against L1210 in mice, the percent increase in life span at the LD10 dosage was +151% (methotrexate), +178% (10-deazaminopterin), +235% (10-methyl analogue), and +211% (10-ethyl analogue). 10,10-Dimethyl-10-deazaminopterin was less effective at an equimolar dosage, but the ILS at the maximum dose tested (72 mg/kg) was +135%. It was far less toxic than the other analogues possibly because of enhanced clearance.

Aminopterin↗
PubMed1981

Pteroylpolyglutamates.

Reduced derivatives of the vitamin pteroylglutamic acid (folic acid) are essential coenzymes for the biosynthesis of purine nucleotides, methionine, thymidylate and for many other enzyme catalyzed reactions involving the transfer, oxidation and reduction of single carbon units. Pteroylglutamic acid is found in tissues in the form of poly-gamma-glutamyl derivatives of varying chain length. The present review covers the detection, distribution, synthesis, degradation, coenzyme function and inhibitory activities of pteroyl-gamma-glutamates. The biosynthesis and inhibitory activities of poly-gamma-glutamyl derivatives of methotrexate, an analog of pteroylglutamic acid having antitumor activity, are also considered. An hypothesis on the coenzymatic role of pteroylpoly-gamma-glutamates in the coordination of sequential enzymatic steps in the metabolism of single carbon units is presented.

Amide Synthases↗
PubMed1981

Polyglutamyl derivatives of tetrahydrofolate as substrates for Lactobacillus casei thymidylate synthase.

Tetrahydropteroylpolyglutamates containing up to seven Glu residues were tested as substrates for Lactobacillus casei thymidylate synthase. The Km values decreased from 24 microM for the monoglutamate to 1.8 microM for the triglutamate. Addition of residues 4, 5, 6, and 7 did not decrease the Km further. When monoglutamate and polyglutamate substrates were simultaneously incubated with the enzyme, the rate observed was characteristic of the polyglutamate even when the monoglutamate concentration was 44 times that of the polyglutamate. Iodoacetamide treatment inhibited the enzyme to the same extent with monoglutamate and polyglutamate substrates. Addition of 0.3 M NaCl doubled the rate obtained with the polyglutamate substrate whereas the rate with the monoglutamate was inhibited 25%. MgCl2 stimulated the reaction only 10% with the polyglutamate substrate compared with 80% stimulation obtained with the monoglutamate. Inhibition by fluorodeoxyuridylate was similar with both mono- and polyglutamate substrates; however, with the phosphonate derivative of fluorodeoxyuridine, the polyglutamate substrate enhanced inhibition 5- to 8-fold.

Deoxyuridine↗