Synthesis of the Multidrug Reversal Agent Ko143 and Its Parent Natural Product Fumitremorgin C

Ko143 is a tetracyclic, synthetic analog of the fungal metabolite fumitremorgin C. Ko143 is a potent and specific inhibitor of the membrane-bound efflux transporter ABCG2, and it reverses ABCG2-mediated drug resistance in cancer cells. Here, we describe an improved synthesis of Ko143 that relies on the highly selective, substrate-controlled reduction of an imine that is formed in a Bischler–Napieralski reaction with the amide derived from 6-methoxy-l -tryptophan methyl ester and isovaleric acid as a key step. We have also developed a new route to 6-methoxy-l -tryptophan methyl ester from Cbz-l -aspartic acid methyl ester, m-anisidine and differently substituted benzaldehydes. With p-nitrobenzaldehyde as one of the starting materials, this route gave access to 6-methoxy-l -tryptophan methyl ester in five steps and 20 % overall yield; however, it is less efficient than a previously reported synthesis of 6-methoxy-l -tryptophan methyl ester from 6-methoxy indole.     

6-aza-5,8,10-trideaza analogues of tetrahydrofolic acid and tetrahydroaminopterin: Synthesis and biological studies

6-Aza-5,8,10-trideaza-5,6,7,8-tetrahydrofolic acid ( 3 ) and 6-aza-5,8,10-trideaza-5,6,7,8-tetrahydroamino-pterin ( 4 ) were synthesized from 6-aza-5,8,10-trideaza-5,6,7,8-tetrahydropteroic acid ( 5 ) and 4-amino-6-aza-5,8,10-trideaza-4-deoxy-5,6,7,8-tetrahydropteroic acid ( 6 ), respectively, by mixed carboxylic-carbonic anhydride condensation with dimethyl L-glutamate followed by ester hydrolysis. The pteroic acid analogues 5 and 6 were prepared in several steps from 1-benzyl-3-carbethoxypiperidin-4-one via 2-amino-6-benzyl-5,6,7,8-te-trahydropyrido[4,3-d]pyrimidin-4(3H)-one ( 7 ). Compound 3 did not inhibit the growth of L1210 mouse leukemia cells in culture, and was not an inhibitor of dihydrofolate reductase (DHFR) or thymidylate synthase (TS). It was a very poor substrate for mouse liver folylpolyglutamate synthetase (FPGS). The 2,4-diamino analogue 4 was only a marginal substrate for FPGS, yet showed activity comparable to methotrexate as a DHFR inhibitor and as an inhibitor of tumor cell growth. The cytotoxicity of 4 is noteworthy because this compound is, to our knowledge, the first example of a classical antifolate which forms polyglutamates poorly even though it contains an intact p-aminobenzoyl-L-glutamic acid side-chain. The inability of 3 and 4 to form polyglutamates indicates that a basic nitrogen at position 6 is highly unfavorable for binding to FPGS.     

Quinazoline Antifolate Thymidylate Synthase Inhibitors: Replacement of Glutamic Acid by Aminophosphonic Acids

The synthesis of six analogues of the potent thymidylate synthase (TS) inhibitor N -[4-[ N -[(3,4-dihydro-2-methyl-4-oxo-6-quinazolinoyl)-methyl]- N -prop-2-ynylamino]benzoyl]- L -glutamic acid 2 is described in which the glutamic acid residue has been replaced by DL -aminophosphonic acids. New antifolates were tested as inhibitors of TS isolated from mouse L1210 leukemic cells as well as inhibitors of growth mouse leukemic L5178Y cells. In general these modifications result in compounds that are considerably less potent than 2 as TS inhibitors with K i 's 0.17-1.10 w M. Very poor solubility in water limited their proper assay of growth cells inhibition.     

Methotrexate analogs. 7. Synthesis of two higher homologs and a positional isomer of methotrexate diethyl ester as potential antitumor agents

Analogs of methotrexate diethyl ester ( 1 ) were prepared, in which the distances separating the ester functions from each other and from the carboxamide function of the p-aminobenzoate moiety were varied via the use of methylene groups as “spacers”. The diethyl esters 3 and 4 , with D,L-α-aminoadipate and D,L-α-aminopimelate side chains in place of L-glutamate, displayed approximately the same order of activity as compound 1 against bacterial and mammalian cells in culture, and were inhibitors of the enzyme dihydrofolate reductase. When given intraperitoneally to L1210 1eukemic mice at a dose of 120 mg./kg. q3d 1,4,7, compound 4 produced a 67% increase in survival and no evidence of toxicity, whereas methotrexate diethyl ester ( 1 ) gave a 44% increase in survival at a dose of 45 mg./kg. q3d 1,4,7 but was toxic at higher doses. The positional isomer 2 was inactive.     

Cytotoxicity of cobalt complexes of furan oximes in murine and human tissue‐cultured cell lines

The copper complexes of 2‐furaldehyde and furan oximes have previously demonstrated potent cytotoxicity, L1210 DNA synthesis inhibition, DNA topoisomerase II inhibition and DNA fragmentation. Currently a series of cobalt metal complexes of 2‐furaldehyde oximes were compared with copper complexes of furan oximes to determine whether the type of metal is important to the cytotoxicity and mode of action of the complexes. The cobalt complexes of furan oximes, like the copper complexes, were shown to be cytotoxic to suspended tumor cell lines, e.g. leukemias, lymphomas, acute monocytic leukemia and HeLa‐S³ uterine carcinoma. The cobalt complexes did not demonstrate dramatic cytotoxicity against the growth of tumors derived from solid human tumor lines. The cobalt complexes preferentially inhibited L1210 DNA synthesis, followed by inhibition of RNA and protein synthesis from 25 to 100 µM over 60 min. These agents, like the copper complexes of 2‐furaldehyde and furan oximes, were inhibitors of DNA polymerase α activity and de novo purine synthesis with marginal inhibition of ribonucleoside reductase and dihydrofolate reductase activities with DNA fragmentation. Unlike the copper complexes, the cobalt complexes did not inhibit L1210 DNA topoisomerase II activity but did reduce thymidylate synthetase activity. Thus, varying the type of metal within the complexes of 2‐furaldehyde and furan oximes produces differences in both cytotoxicity and mode of action. Copyright © 1999 John Wiley & Sons, Ltd.     

Synthesis and biological evaluation of -n[4-(2-trans-[([2,6-diamino-4(3H)-oxopyrimidin-5-yl]methyl)thio]cyclobutyl)benzoyl] -l-glutamic acid a novel 5-thiapyrimidinone inhibitor of dihydrofolate reductase

The synthesis and biological evaluation of N-[4-(2-trans-[([2,6-diamino-4(3H)-oxopyrimidin-5-yl]methyl)thio]cyclobutyl)benzoyl]-L-glutamic acid (1) is reported. Compound 1 is a potent dihydrofolate reductase (DHFR) inhibitor (K_j = 12 nM) with excellent in vitro cell culture growth inhibition (L1210, IC₅₀ = 29 nM). Protection experiments showed that the cell growth inhibitory activity was due to DHFR inhibition. The key step in the synthesis was the coupling of a cyclobutylmethylthiol with the 5-bromo-2,6-diamino-4-oxopyrimidine ⁸.     

Exclusive Free Radical Mechanisms of Cellular Photosensitization

In order to determine the specific effects of radical-induced reactions in the absence of complicating excited-state pathways, four different thiohydroxamic esters and their parent molecule, N-hydroxypyridine-2(1H)-thione, have been studied in murine L1210 leukemia cells for their ability to produce photobiological damage. Irradiation (X_exc= 355 nm) of cells in the presence of thiopyridone esters, specific photolytic precursors of sulfur-, carbon- and oxygen-centered radicals, caused toxicity that was unambiguously demonstrated to result from radical photosensitization mechanisms. Cellular morphological changes were observed following irradiation but apoptosis was not found to take place. A good correlation was evident between lipid peroxidation, measured by the thiobarbituric acid method, and phototoxicity, assessed by the trypan blue exclusion assay, indicating that the ester derivatives exert their effects mainly in plasma and/ or subcellular membranes. Irradiation performed under deaerated conditions also induced significant phototoxicity but the effects of deaeration were dependent on the ester used and are discussed in terms of the nature of the primary radical species generated in each case. Irradiation of L1210 cells in the presence of N-hydroxypyridine-2(1H)-thione, a nonspecific, photochemical source of hydroxyl radical, was also found to trigger phototoxicity and lipid peroxidation. However in this case, photodamage cannot yet be definitely attributed to a radical or type II mechanism although the apparent oxygen independence of phototoxicity would indicate that type II contribution is not significant.     

The carbocyclic analog of cytidine,synthesis and antineoplastic activity

The carbocyclic analog (VI) of cytidine was prepared from the carbocyclic analog (I) of uridine. The intermediate stages were a tetrabenzoyl derivative of I, the tribenzoyl derivative of the uridine analog, and the tribenzoyl 4-chloropyrimidinone obtained from the latter derivative. The cytidine analog (VI) is active against KB cells in culture and against L1210 leukemia in mice. In the initial tests against L1210 leukemia, the highest dose (200 mg./kg./day, q.d. 1-9), of three active doses, increased lifespan by 82% and showed no evidence of toxicity.     

Synthesis of novel β-propanamides to inhibit cholesteryl ester transfer protein(CETP)

A novel series of β-propanamide derivatives as inhibitors of cholesteryl ester transfer protein(CETP)were synthesized.Previously,H3(IC_(50) 2 μmol/L) was observed to inhibit CETP moderately(Xie et ah,2016).Structural modifications based on H3 led to discovery of the successful CETP inhibitor,known as 1-methyl-4-arylpyrazole.Using a similar approach,compound Q08 was identified as a highly potent CETP inhibitor with an IC_(50) of 490 nmol/L in vitro.     

Folic acid analogs. Modifications in the benzene-ring region. 5. 2′,6′-diazafolic acid

The synthesis of 2′,6′-diazafolic acid was accomplished by the condensation of 2-acetylamino-4(3H)pteridinone-6-earboxaldehyde (XIV) with diethyl N-[(5-amino-2-pyrimidinyl)carbonyl]-L-glutamate (XIII) followed by reduction of the anil double bond and alkaline hydrolylic cleavage of the N²-acetyl and ethyl ester protecting groups. Intermediate XIII was prepared by starling with 5-nitro-2-styrylpyrimidine (VI) and proceeding via 5-arnino-2-styrylpyrimidine (IX). The henzyloxycarbonyl derivative of IX was prepared and oxidized to the corresponding 5-benzyloxycarbonylaminopyrimidine-2-carboxylic acid (XI). The coupling of XI with diethyl L-glutamate followed by hydrogenolysis of the henzyloxycarbonyl function afforded the desired intermediate XIII. 2′,6′-Diazafolic acid was a potent inhibitor of Streptococcus faecium and displayed marginal activity against leukemia 1,1210 in mice.     

Folate antagonists. 17. Synthesis and biological properties of a 2,4-diamino-6-thioquinazoline analog of aminopterin

A multistep route for the synthesis of N-[4-[(2,4-diamino-6-quinazolinyl)thio]benzoyl]-L-glutamic acid (2) from 4-mercaptobenzoic acid and 5-chloro-2-nitrobenzonitrile is described. Although this aminopterin analog lacked significant antimalarial activity, it was a potent inhibitor of dihydrofolate reductase from Trypanosoma cruzi. The pteroic ester analog 11 , however, was active against Plasmodium berghei infections in mice at high doses.     

Synthesis and preliminary biological evaluation of 5-fluoro-5,8-dideazaisoaminopterin

The new folate antagonist, 5-fluoro-5,8-dideazaisoaminopterin was synthesized in four steps beginning with 2,4-diamino-5-fluoroquinazoline. It was found to be a potent inhibitor of human dihydrofolate reductase. Against L1210 leukemia in mice, 5-fluoro-5,8-dideazaisoaminopterin was equiactive with methotrexate at approximately one half of the total dose employed.     

Synthesis and biological evaluation of 9‐thia‐5,10‐dideazafolic acid

The folate analogue, 9‐thia‐5,10‐dideazafolic acid ( 3b ), was obtained in an efficient two‐step procedure in an overall yield of 60%. The previously unknown intermediate dimethyl‐thiocarbamic acid S‐(2‐amino‐3,4‐dihydo‐4‐oxo‐pyrido[2,3‐d]pyrimidin‐6‐yl) ester ( 5 ) was prepared via the condensation of 2,6‐diamino‐3H‐pyrimidin‐4‐one and S‐(2‐malonaldehyde)‐1,1,3,3‐tetramethylthiouronium bromide ( 4 ). Compound 5 , in a one pot procedure, was deprotected using sodium hydroxide and then coupled to diethyl N‐[(4‐chloromethyl)benzoyl]‐L‐glutamate, followed by saponification of the ethyl esters to give the 9‐thia‐5,10‐dideazafolic acid ( 3b ). Compound 3b was a potent inhibitor of human 5‐aminoimidazole‐4‐carboxamide ribonucleotide transformylase (K_i of 8 ± 5 μM) and showed no inhibition of human glycinamide ribonu‐cleotide transformylase at concentrations as high as 50 μM. Compound 3b was screened by the National Cancer Institute Developmental Therapeutics Program against 60 human tumors and was found to be active against a leukemia RPMI‐8226 cell line where the LC₅₀ was 1 μM.     

Synthesis and resolution of diethyl (1S,2S)-1-amino-2-vinylcyclopropane-1-phosphonate for HCV NS3 protease inhibitors

We herein describe an efficient synthesis of optically active diethyl 1-amino-2-vinylcyclopropane-1-phosphonate (analogous to 1-amino-2-vinylcyclopropane-1-carboxylate). The racemic phosphonate diethyl ester was obtained from an imine derived from aminomethylphosphonate diester and trans-1,4-dibromo-2-butene. Crystallizations of the dibenzoyl-l-tartaric acid salt allowed for separation of enantiomers. The enantiomerically pure material was used to synthesize an extremely potent tripeptide phosphonate inhibitor of HCV NS3 protease. X-ray crystal structure of the inhibitor bound to the HCV NS3 protease confirmed the absolute stereochemistry of the title compound.     

Colchicine Models: Synthesis and Binding to Tubulin of Tertamethoxybiphenyls

Syntheis of tetramethoxybiphenyl 21 was accomplished from 4-phenylcylohexane-1,3-dione 13 by aromatization to biphenyl 19 and reductive removal of the phenolic OH group as phenyltetrazolyl ether. Tertramethoxybiphenyls 34 and 40 were obtained from 4-phenylcyclohexenone 26 via ester 27 . The tetramethoxybiphenyls 21, 34 , and 40 , and analogs 28, 29 , and 31 were evaluated for antitubulin activity and as antimitotic agents with L1210 murine leukemia cells. Compounds 31 and 34 had significant effects on the in-vitro polymerization of tubulin. Compound 31 was the most cytotoxic of the six new biphenyls studied (IC₅₀ for cell growth, 0.6M ) and caused the accumulation of cells in metaphase arrest.     

Stereocontrolled Total Synthesis of (+)‐trans‐Dihydronarciclasine

A highly stereoselective and efficient total synthesis of trans‐dihydronarciclasine from a readily available chiral starting material was developed. The synthesis defines two of the five stereogenic centers of the natural product by an amino acid ester–enolate Claisen rearrangement. The other three stereogenic centers are created in a highly stereocontrolled fashion via a six‐ring vinylogous ester intermediate, which is generated from the γ,δ‐unsaturated ester functional group of the Claisen rearrangement product in an efficient three‐step sequence. This concise total synthesis exemplifies the use of a highly regioselective Friedel–Crafts‐type cyclization to form the B ring via an isocyanate intermediate derived from an N‐Boc group, which is superior to the conventional method using an imino triflate intermediate. This same N‐Boc group is employed to give high selectivity in the Claisen rearrangement earlier in the sequence.     

2-Hydroxy-4-methoxy-trans-cinnamic acid as a precursor of herniarin inArtemisia dracunculus

The isolation of (E)-2-hydroxy-4-methoxycinnamic acid (1) fromArtemisia dracunculus L. supports strongly the assumption that this compound is an intermediate in the biosynthesis of 7-methoxycoumarin (herniarin,3). The structure of the UV-unstable compound1 was derived from¹H-NMR data and by comparison of the stable dimethylated derivative with synthetic (E)-2,4-dimethoxycinnamic acid methyl ester (2).Dedicated to Prof. Dr.Kurt L. Komarek on the occasion of his 60th birthday.     

Syntheses and biological evaluation of vinblastine congeners

Sixty-two congeners of vinblastine (VLB), primarily with modifications of the piperidine ring in the carbomethoxycleavamine moiety of the binary alkaloid, were synthesized and evaluated for cytotoxicity against murine L1210 leukemia and RCC-2 rat colon cancer cells, and for their ability to inhibit polymerization of microtubular protein at < 10(-6) M, and for induction of spiralization of microtubular protein, and for microtubular disassembly at 10(-4) M concentrations. An ID50 range of >10(7) M concentrations was found for L1210 inhibition by these compounds, with the most active 1000x as potent as vinblastine.     

Synthesis of D- and L-5-Oxaproline and of a New Captopril Analogue

The 1,3-dipolar cycloaddition of the C-t-butyloxycarbonyl-N -mannosyl-nitrone 5 , formed in situ from the partially protected D -mannose-oxime 3 and the glyoxylate 4 , to ethylene gave preferentially the (3S)-N-glycosyl-isoxazolidine 6 which was transformed into the 3-isoxazolidine-carboxylate (L -5-oxaproline ester) 12 and into some derivatives thereof. The (S)-configuration of 12 was proved by chemical correlation with a derivative of L -asparagine. The D -5-oxaproline ester was obtained from the corresponding N-ribosyl-nitrone 24 . Two protected dipeptides containing either C-terminal- ( 28 ) or N-terminal-5-oxaproline (= Opro) ( 30 ) were synthesized. Starting from 12 , the analogue 1 of captopril® ( 2 ) was prepared and its activity as an inhibitor of the angiotensin-converting-enzyme (ACE) was examined.     

trans‐3‐Ethyl‐cis‐2,6,6‐trimethyl‐4‐oxocyclohexanecarboxylic acid: an intermediate in the synthesis of a highly potent estrogen

The title compound, C₁₂H₂₀O₃, (IV), the ethyl ester of which is an intermediate in the synthesis of a compound reported to be highly estrogenic, has been prepared. After the initial steps reported for the synthesis of this ester intermediate were followed, it was converted into the crystalline acid, (IV), for X‐ray analysis. It was verified that (IV) was racemic when prepared. X‐ray analysis showed that anti‐hydrogenation of the double bond had occurred in the synthesis, making the orientation of the carboxyl group cis to the 2‐methyl group and trans to the 3‐ethyl group. NMR spectroscopy showed that the stereochemistry of (IV) was identical with that of its ester precursor. While the earlier report did not note the stereochemistry of this ester, it pointed out that the estrogenic product derived from it possessed the opposite carboxyl‐2‐methyl orientation, i.e.trans, although no X‐ray analysis was performed. In the light of these results and the importance of correlating biological activity with compound structure, the unequivocal characterization of the highly estrogenic compound is warranted.