Inhibitor Of S-adenosylmethionine Decarboxylase Research Articles

CGP 48664, a potent and specific S-adenosylmethionine decarboxylase inhibitor: effects on regulation and stability of the enzyme.

Mammalian S-adenosylmethionine decarboxylase (AdoMetDC) catalyses a regulatory important step in the biosynthesis of polyamines and is a potential target for therapeutic agents against various parasitic diseases and proliferative disorders. In the present study we examined the effects of a newly synthesized AdoMetDC inhibitor. 4-amidinoindan-1-one 2'-amidinohydrazone (CGP 48664), on polyamine metabolism in the mouse leukaemia cell line L1210. Treatment of the cells with 2 microM CGP 48664 led to a depletion of cellular spermidine and spermine. The putrescine content, in contrast, was markedly increased. Cells seeded in the presence of the inhibitor showed a significant decrease in growth rate, which was fully reversed by the addition of 2 microM spermidine or 1 microM spermine. The syntheses of ornithine decarboxylase and AdoMetDC were greatly increased in cells treated with CGP 48664. These increases were not correlated with similar changes in the mRNA levels, indicating the involvement of a translational mechanism. CGP 48664 was demonstrated to be a very poor competitor of spermidine uptake in the L1210 cells. L1210 cells deficient in polyamine transport were as sensitive to the antiproliferative effect of the inhibitor as were the parental cells, indicating that CGP 48664 did not enter the cells by the polyamine transport system. In addition to inhibiting AdoMetDC, CGP 48664 stabilized the enzyme against degradation. In the present study we also demonstrated that aminoguanidine (AMG), which is frequently used in cellular systems to inhibit any action of serum polyamine oxidase, apparently inhibits AdoMetDC by an irreversible mechanism that markedly stabilizes the enzyme against proteolytic degradation. CGP 48664 and the parental compound methylglyoxal bis(guanylhydrazone), which is also a potent inhibitor of AdoMetDC, contain one or two AMG-like moieties; the importance of these residues in the inhibition of AdoMetDC is discussed.

Structure-activity relations of S-adenosylmethionine decarboxylase inhibitors on the growth of MCF-7 breast cancer cells.

SAMDC is a key enzyme in the biosynthesis of spermidine and spermine, 2 polyamines that are essential for cell proliferation. Inhibition of polyamine biosynthesis is often targeted as a therapeutic strategy to suppress cancer cell growth as these cells contain elevated levels of polyamines. We examined the effect of a new group of SAMDC inhibitors, CGP33829, CGP35753, CGP36958, CGP39937, and CGP48664, (obtained from Ciba-Geigy, Basel, Switzerland), and their parent compound, MGBG, on the proliferation of MCF-7 breast cancer cells. MGBG had minimal effects on the proliferation of MCF-7 cells up to 6 microM concentration. In contrast, CGP48664 and CGP39937, containing 2 aromatic rings that delocalize the pi electron system of the backbone of MGBG, were potent inhibitors with 50% growth inhibition at 0.5 microM concentration. Other CGP compounds were less effective in inhibiting cell growth. The ability of CGP48664 to inhibit MCF-7 cell proliferation was related to its ability to inhibit SAMDC and to consequently deplete spermidine and spermine levels in the cell. Exogenous spermidine and spermine could reverse the growth inhibitory effects of this compound. CGP compounds also increased the activity of ODC, another enzyme involved in polyamine biosynthesis. Northern blot analysis of mRNA from MCF-7 cells progressing in cell cycle after G1 synchronization did not show an increase in ODC mRNA level by CGP48664. These data demonstrate structure-activity relationships of a series of MGBG derivatives on cell growth, enzyme activities, and polyamine biosynthesis in a hormone-responsive breast cancer cell line and suggest potential application of SAMDC inhibitors as therapeutic agents.

CHARACTERIZATION OF ALPHA-DIFLUOROMETHYLORNITHINE RESISTANT LEISHMANIA DONOVANIAND ITS SUSCEPTIBILITY TO OTHER INHIBITORS OF THE POLYAMINE BIOSYNTHETIC PATHWAY

The promastigote form of Leishmania donovaniis sensitive to growth inhibition by DL- α-difluoromethylornithine, an irreversible inhibitor of ornithine decarboxylase, exhibiting an IC 50of 125 μM. Exposure of a wild type cell population to gradually increasing concentrations of DL- α-difluoromethylornithine resulted in the selection of a strain of Leishmania, which was capable of proliferating in 5mM DFMO with an IC 50value of 10mM. This resistant strain showed increased ornithine decarboxylase activity and higher levels of putrescine. It was sensitive to growth inhibition by bis (benzyl) analogs of spermine and a potent S-adenosylmethionine decarboxylase inhibitor, CGP 40215A. These results indicate the potential value of other inhibitors of the polyamine biosynthetic pathway as therapeutic agents in conditions where ODC inhibitors are ineffective.

In vitro trypanocidal activities of new S-adenosylmethionine decarboxylase inhibitors

A series of novel aromatic derivatives based on the structure of methylglyoxal bis(guanylhydrazone) (MGBG) was examined for in vitro antitrypanosomal activities and cytotoxicities for human cells. One-third of the compounds tested showed trypanocidal activity at concentrations below 0.5 microM after an incubation period of 72 h. Structure-activity analysis revealed that bicyclic compounds with homocyclic rings and unmodified termini were the most active compounds. Results obtained in three laboratories employing different methods and trypanosome populations consistently ranked compound CGP 40215A highest. This compound had a 50% inhibitory concentration of 0.0045 microM for Trypanosoma brucei rhodesiense, was also active against other trypanosome species, including a multidrug-resistant Trypanosoma brucei brucei, and was significantly less toxic than other compounds tested for a human adenocarcinoma cell line, with a 50% inhibitory concentration of 1.14 mM. The effect of CGP 40215A was time and dose dependent, and low concentrations of the compound required exposure times of > 2 days to exert trypanocidal activity. Compounds were inactive against Leishmania donovani and Trypanosoma cruzi amastigotes in murine macrophages in vitro.

In vivo trypanocidal activities of new S-adenosylmethionine decarboxylase inhibitors.

A series of novel aromatic derivatives based on the structure of methylglyoxal bis(guanylhydrazone) (MGBG) was examined for trypanocidal activities in human and veterinary trypanosomes of African origin. One agent, CGP 40215A, a bicyclic analog of MGBG which also resembles the diamidines diminazene (Berenil) and pentamidine, was curative of infections by 19 isolates of Trypanosoma brucei subspecies as well as a Trypanosoma congolense isolate. Several of these isolates were resistant to standard trypanocides. Curative doses were < or = 25 mg/kg of body weight/day for 3 days in these acute laboratory model infections. In addition, CGP 40215A also cured a model central nervous system infection in combination with the ornithine decarboxylase inhibitor DL-alpha-difluoromethylornithine (DFMO; Ornidyl, eflornithine). Curative combinations were 14 days of oral 2% DFMO (approximately 5 g/kg/day) plus 5, 10, or 25 mg/kg/day for 3 or 7 days given by intraperitoneal injection or with a miniosmotic pump. Combinations were most effective if CGP 40215A was given in the second half or at the end of the DFMO regimen. MGBG has modest activity as an inhibitor of trypanosome S-adenosylmethionine decarboxylase (50% inhibitory concentration [IC50]. 130 microM), while CGP 40215A was a more active inhibitor (IC50, 20 microM). Preincubation of trypanosomes with CGP 40215A for 1 h caused a reduction in spermidine content (36%) and an increase in putrescine content (20%), indicating that one possible mechanism of its action may be inhibition of polyamine biosynthesis.

Regulation by Spermine of Native Inward Rectifier K+ Channels in RBL-1 Cells

Polyamines have been shown to participate in the rectification of cloned inwardly rectifying potassium channels, a class of potassium channel proteins that conducts inward current more readily than outward current. Here, basophil leukemia cells were used to determine the effects of polyamines on a native, inwardly rectifying potassium current. Rat basophil leukemia cells were cultured in the presence of two different polyamine biosynthesis inhibitors, and both the electrophysiological properties and the polyamine levels were monitored. Treatment with alpha-difluoromethylornithine, a specific ornithine decarboxylase inhibitor, resulted in no significant change of electrophysiological properties. In contrast, treatment with 5'-[(Z)-4-amino-2-butenyl]- methyl-amino-5'-deoxyadenosine (MDL73811), an inhibitor of S-adenosylmethionine decarboxylase, resulted in increased outward currents through inwardly rectifying potassium channels while intracellular putrescine was markedly increased and spermidine and spermine levels were decreased. Fluctuations of intracellular polyamine concentrations as imposed by MDL73811 were directly translated in an altered cell excitability. Based on these results we conclude that the rectification properties of native inwardly rectifying potassium channels are largely controlled by intracellular spermine.

Inhibition of Polyamine Synthesis Alters Hair Follicle Function and Fiber Composition

The activities of ornithine decarboxylase and S-adenosylmethionine decarboxylase, two of the enzymes involved in the synthesis of the polyamines, were found to be high in follicle-rich homogenates of sheep skin, and to be responsive to the nutrition of the animal. Systemic provision of the inhibitor of ornithine decarboxylase, alpha difluoromethylornithine, markedly altered the length, diameter, and composition of the fiber, the last being accompanied by an increase in the proportion of the fiber occupied by paracortical cells and an increase in the level of mRNA encoding a cysteine-rich family of keratin proteins. The growth of wool follicles cultured in media containing alpha-difluoromethylornithine was not inhibited, even at high concentrations. In contrast, low concentrations of methylglyoxal (bis)guanylhydrazone, the inhibitor of S-adenosylmethionine decarboxylase, completely inhibited fiber growth in culture follicles. Addition of spermidine to the media overcame this inhibition but spermine had no effect. Further evidence that spermine is not required for normal follicle function was provided by incubating follicles with the specific inhibitor of spermine synthase, n-butyl-1,3-diaminopropane. This inhibitor, even at high concentrations, had no effect on fiber growth in vitro. Spermidine partially overcame the growth depression that occurred in follicles cultured in methionine-deficient media, suggesting that part of the requirement for methionine is for spermidine synthesis in the follicle. These investigations provide strong evidence that the polyamines in general , and spermidine in particular, play a major role in hair growth.

Use of 4-fluoro- l-ornithine to monitor metabolic flux through the polyamine biosynthetic pathway

The mechanistic effectiveness of various polyamine analogs and enzyme inhibitors is typically determined by their ability to deplete intracellular polyamine pools. In this study, we describe an assay that may prove useful in augmenting this relatively static assessment of drug action. The assay relies upon the substitution of 4-fluoro- l-ornithine (Fl-Orn) for ornithine as a polyamine precursor to provide a means to measure metabolic flux through polyamine pools. At concentrations up to 500 μM, the analog did not inhibit the growth of L1210 murine leukemia cells during incubations of up to 72 hr. Using HPLC, the analog was processed metabolically over time to what was deduced to be 2-fluoroputrescine, 6-fluorospermidine and 6-fluorospermine. The relative proportion of fluorinated polyamine analog to the natural polyamine increased with time and Fl-Orn concentration. The sum of the two was found to be nearly identical to the respective polyamine pool of control cells exposed instead to 500 μM ornithine. This indicates that Fl-Orn was recognized and utilized as a precursor at a rate very similar to that of ornithine itself. Using L1210 cells at different stages of cell growth, it was determined that the metabolic flux through the pools, as indicated by the rate of appearance of individual fluorinated polyamine species, reflected the proliferation status of the cells—non-growing cells failed to incorporate the analog. Likewise, in cell types with varying polyamine pool profiles, such as polyamine enzyme overproduces or those with constitutively different spermidine of spermine ratios, the incorporation of the fluorinated analogs into pools was found to be proportional to the size to the natural polyamine pool. In cells treated with inhibitors of S-adenosylmethionine decarboxylase, Fl-Orn incorporation indicated a total blockade of polyamine synthesis at that enzyme site. Overall, the Fl-Orn assay has demonstrated that polyamine pool profiles generally reflect the rate of flux through the pathway in proliferating cells, suggesting that most intracellular polyamines are freely exchangeable with those undergoing metabolic flux.

Biochemical and growth-modulatory effects of the new S-adenosylmethionine decarboxylase inhibitor CGP 48664 in malignant and immortalized normal human breast epithelial cells in culture.

CGP 48664 [4-aminoindanon-1-(2'-amidino)hydrazone dihydrochloride monohydrate] is a newly introduced inhibitor of S-adenosylmethionine decarboxylase (SAMDC) with increased selectivity of action and reduced toxicity. We analyzed the biochemical and antiproliferative effects of this compound in a panel of hormone-dependent (3 clones of MCF-7, T47D) and -independent (MDA-MB-231, BT-20) human breast cancer cell lines in culture. For comparison, we also tested its effects in the spontaneously immortalized human breast epithelial cell line MCF-10A. All cell lines were highly sensitive to the growth-inhibitor effect of CGP 48664 with an IC50 between 0.1 and 0.5 microM. A dose-dependent bell-shaped increase in SAMDC was observed in normal and malignant breast cells resulting from enzyme stabilization by the inhibitor as supported by Western blot analysis. While ornithine decarboxylase (ODC) activity consistently increased, the effect of CGP 48664 on spermidine/spermine N'acetyltransferase (SSAT) was variable in the breast cancer cell lines. In contrast, the inhibitor consistently reduced SSAT activity level in the MCF-10A cell line and its derivative partially transformed by a mutated ras oncogene. As expected cellular putrescine levels were markedly increased by CGP 48664 administration, whereas spermidine and spermine contents were reduced. However, the degree of reduction was usually only moderate. Furthermore, exogenous polyamine administration was relatively ineffective in rescuing the antiproliferative effect of CGP 48664 in MCF-7 cells, while exerting a more complete rescue in the MDA-MB-231 cell line. We conclude that CGP 48664 exerts a potent growth-inhibitory effect on mammary cells in culture. However, its action may not always be entirely mediated through the polyamine pathway.

Treatment of nude mice with 4-amidinoindan-1-one2???-amidinohydrazone, a new S-adenosylmethionine decarboxylase inhibitor, delays growth and inhibits metastasis of human melanoma cells

CGP 48664A (4-amidinoindan-1-one2'-amidinohydrazone) is a novel inhibitor of S-adenosyl-methionine decarboxylase (SAMDC), a key enzyme in the biosynthesis of polyamines, which are themselves essential for proliferation of mammalian cells. Seven different human melanoma cell lines were treated in vitro with CGP 48664A. High, intermediate and low levels of cytostasis were induced in four, one and two melanoma lines, respectively. This cytostasis was reversed by the addition of exogenous spermidine or spermine to the culture medium. The heterogeneous low metastatic (CGP 48664A-resistant) A375P cells and highly metastatic (CGP 48664A-sensitive) A375SM cells were implanted into the subcutis or injected intravenously into nude mice. Systemic daily administration of CGP 48664A significantly reduced the size of cutaneous lesions and the number of lung metastases in mice implanted with A375SM cells. No beneficial effects were found in mice injected with A375P cells. Drug activity was dose dependent, and maximal effects were observed when treatment began in mice with small tumour burdens. The data suggest that CGP 48664A is effective against melanoma metastasis in nude mice and that its activity should be tested in combination with other cytoreductive agents.

S-(5'-deoxy-5'-adenosyl)-1-aminoxy-4-(methylsulfonio)-2-cyclopentene (AdoMao): an irreversible inhibitor of S-adenosylmethionine decarboxylase with potent in vitro antitrypanosomal activity.

The S-adenosylmethionine (AdoMet) analogue S-(5'-deoxy-5'-adenosyl)-1-aminoxy-4-(methylsulfonio)-2-cycl opentene (AdoMao) was synthesized in two of its four possible diastereomeric forms using a facile chemoenzymatic route. The trans-1R,4R- and trans-1S,4S-diastereomers of AdoMao, as well as the corresponding diastereomers of the unmethylated precursor molecule nor-AdoMao, were then evaluated as inhibitors of S-adenosylmethionine decarboxylase (AdoMet-DC) from both bacterial and human sources. All four of the analogues acted as time-dependent, irreversible inhibitors of AdoMet-DC from Escherichia coli, exhibiting remarkably constant Ki values ranging between 20.6 and 23.7 microM. These analogues also inhibited the human form of AdoMet-DC, although this form of the enzyme was able to discriminate between AdoMao (Ki values of 21.2 microM for the trans-1R,4R form and 19.6 microM for the trans-1S,4S form) and nor AdoMao (Ki values of 95.2 microM for the trans-1R,4R form and 30.9 microM for the trans-1S,4S form). The trans diastereomers of AdoMao and nor-AdoMao were next evaluated for their ability to inhibit trypanosomal growth in vitro against cultured Trypanosoma brucei brucei bloodforms. All four of these analogues were effective growth inhibitors, with IC50 values ranging between 0.9 and 10.1 microM. The two most effective analogues, trans-1S,4S-AdoMao (IC50 0.9 microM) and trans-1S,4S-AdoMao (IC50 3.0 microM) were also effective against two clinical isolates of the pathogenic organism Trypanosoma brucei rhodesiense, KETRI 243 and KETRI 269. The most promising analogue in all respects was trans-1S,4S-AdoMao, which was subsequently found to have minimal effects on cell growth, AdoMet-DC activity, and intracellular polyamine levels in the sensitive human promyelocytic leukemia cell line HL60. Thus, the S-adenosylmethionine analogue trans-1S,4S-AdoMao acts as an effective inhibitor of AdoMet-DC and appears to serve as a parasite-specific trypanocidal agent in vitro.

In vivo growth inhibition of L1210 leukemia by 4-amidinoindan-1-one 2'-amidinohydrazone (CGP 48664A), a new inhibitor of S-adenosylmethionine decarboxylase.

We studied the in vivo growth-inhibitory effect of the new S-adenosylmethionine decarboxylase inhibitor 4-amidinoindan-1-one 2'-amidinohydrazone (CGP 48664A). L1210-bearing DBA-2 mice were treated with increasing CGP 48664A doses from 1 day after i.p. L1210 cell inoculation. Treatment was continued for 4 days, after which all mice were killed. CGP 48664A caused dose-related exponential decreases of L1210 cell numbers and spermidine and spermine contents. Putrescine contents increased exponentially. Polyamine changes in spleen and liver were less profound. L1210 growth inhibition was not accompanied by changes in cell cycle phase distribution. It is concluded that CGP 48664A is an effective inhibitor of S-adenosylmethionine decarboxylase but that CGP 48664A-induced changes in intracellular polyamine compositions are not necessarily the cause of growth inhibition.

Polyamines as targets for therapeutic intervention.

Polyamines are ubiquitous cell components essential for normal growth. Compounds interfering with polyamine biosynthesis or function have considerable potential for use as therapeutic agents. Inhibitors of ornithine decarboxylase have been shown to be valuable for the treatment of diseases caused by parasitic protozoa, most notably African sleeping sickness. They may also be useful chemopreventive and antineoplastic agents. Inhibitors of S-adenosylmethionine decarboxylase also have potential as treatments of these diseases. Protocols minimizing uptake of exogenous polyamines via the polyamine-transport system will probably be needed for the full potential of the inhibitors to be realized. Polyamine analogues, notably those with ethyl or benzyl groups on the terminal nitrogen atoms, have potent antiproliferative activity and are promising agents for the treatment of cancer. These analogues are transported by the polyamine-transport system, and their therapeutic effects are less likely to be blocked by the availability of the exogenous polyamines.

SYSTEMIC ADMINISTRATION OF 4-AMIDINOINDANON-1-(2'-AMIDINO)-HYDRAZONE, A NEW INHIBITOR OF S-ADENOSYLMETHIONINE DECARBOXYLASE, PRODUCES CYTOSTASIS OF HUMAN PROSTATE-CANCER IN ATHYMIC NUDE-MICE

CGP 48664A, a new S-adenosylmethionine decarboxylase inhibitor, blocks the production of spermidine and spermine, two polyamines that play critical roles in cellular proliferation. Under in vitro conditions, CGP 48664A produced cytostasis of the human prostate cancer cell lines LNCaP, LNCaP-LN3, PC-3M, and PC-3M-MM2 in a dose-dependent manner. This cytostasis was reversed by the addition of exogenous polyamines to the culture medium. LNCaP-LN3 cells or PC-3M-MM2 cells were implanted into the prostate of nude mice. Daily administration of CGP 48664A significantly inhibited tumor size and serum levels of prostate-specific antigen in mice implanted with LNCaP-LN3 cells. The therapeutic effect was related to the time the treatment was initiated, the volume of disease, and the length of treatment. CGP 48664A was not effective against the fast-growing PC-3M-MM2 tumor. These data suggest that to broaden its effectiveness, CGP 48664A should be combined with other cytoreductive agents.

α-cyano-substituted analogoues of decarboxylated S-adenosylmethionine as enzyme activated, irreversible inhibitors of S-adenosylmethionine decarboxylase

The S-adenosylmethionine analogues 2a and 2b were synthesized and evaluated as enzyme activated, irreversible inhibitors of the pyruvoyl enzyme S-adenosylmethionine decarboxylase. Each compound inactivated both the Escherichia coli and human forms of the enzyme in a time dependent manner. The synthesis of these compounds and the associated enzyme kinetic studies are described.

Preparation of the pure diastereomeric forms of S- (5′-deoxy-5′-adenosyl)-1-ammonio-4-methylsulfonio-2- cyclopentene and their evaluation as irreversible inhibitors of S-adenosylmethionine decarboxylase from Escherichia coli

The conformationally restricted S-adenosylmethionine analogue AdoMac ( S-(5′-deoxy-5′-adenosyl)-1-ammonio-4-methylsulfonio-2-cyclopentene has been shown to act as an enzyme activated, irreversible inhibitor of the Escherichia coli form of the enzyme S-adenosylmethionine decarboxylase. Inactivation of the enzyme is presumably initiated by formation of an imine linkage between the inhibitor and the terminal pyruvate of the enzyme, followed by base-catalyzed elimination of methylthioadenosine and generation of a latent electrophile. Removal of the driving force for the elimination of methylthioadenosine resulted in a reversibly binding inhibitor. Thus, the thioether analogue corresponding to AdoMac, and the corresponding dihydro derivative (H 2-AdoMac), reversibly inhibit the enzyme. AdoMac was resolved into its four pure diastereomeric forms, and each diastereomer was evaluated as an irreversible inhibitor of the enzyme. The K I values for the individual diastereomers range between 3.83 and 39.6 μM, with the cis-1 S,4 R diastereomer being the most potent inhibitor. However, the k inact values for the four diastereomers are not significantly different, suggesting that the binding of each diastereomer to the enzyme is configuration-dependent, while the subsequent inactivation likely proceeds through a single intermediate which is formed from each of the four diastereomers. Since each pure diastereomer represents a distinct conformational mimic exhibiting restricted sidechain rotation, the data suggests that these and related analogues may be useful as conformational probes for the catalytic site of AdoMet-DC.

Effects of polyamine biosynthesis inhibitors on the progesterone-initiated increase in intracellular free Ca2+ and acrosome reactions in human sperm.

This laboratory has previously reported that progesterone can initiate a rapid transient increase in the concentration of intracellular free Ca2+ ([Ca2+]i) and an increase in a Ca(2+)-requiring exocytotic event, the acrosome reaction (AR) in human sperm. Rapid increases in Ca2+ fluxes of some mammalian cells caused by another steroid, testosterone, require polyamine biosynthesis. Herein, we tested two polyamine biosynthesis suicide inhibitors for their effects on the progesterone-initiated increase in [Ca2+]i and AR in capacitated human sperm in vitro: DL-alpha-(difluoromethyl)ornithine hydrochloride (DFMO), an inhibitor of putrescine synthesis by ornithine decarboxylase and (5'[[(Z)-4-amino-2-butenyl]methylamino]-5'-deoxyadenosine (MDL 73811), an inhibitor of S-adenosylmethionine decarboxylase (required for spermidine and spermine synthesis). Sperm were capacitated in vitro and preincubated 10 min with 4.9 mM DFMO or 9.8 microM MDL 73811 with or without various polyamines (245 microM). Progesterone (3.09 microM final concentration) or progesterone solvent (ethanol, 0.1% final concentration) was then added, sperm fixed 1 min after additions and AR assayed by indirect immunofluorescence or with fluorescein-labeled Con A lectin. DFMO strongly inhibited the AR, but putrescine (product of ornithine decarboxylase and precursor of spermidine and spermine) reversed that inhibition. Preincubation for 25 min with DMFO + spermidine also reversed DFMO inhibition. MDL 73811 inhibited the progesterone-initiated AR, and a 10 min preincubation with spermidine, but not putrescine or spermine, reversed that inhibition. Preincubations with putrescine alone or with spermidine alone followed by addition of the progesterone solvent did not initiate the AR, and such preincubations followed by progesterone addition did not increase the AR more than progesterone alone.(ABSTRACT TRUNCATED AT 250 WORDS)

Restricted rotation analogs of S-adenosylmethionine: synthesis, evaluation as inhibitors of S-adenosylmethionine decarboxylase, and potential use as selective antitrypanosomal agents

A series of restricted rotation analogs of S-adenosylmethionine were synthesized and evaluated as inhibitors of S-adenosylmethionine decarboxylase from Escherichia coli. These analogs were also evaluated for inhibitory activity against cultured mammalian cells, and against the parasite Trypanosoma brucei brucei. All analogs tested appeared to selectively inhibit trypanosomal growth. In particular, compound 5 (AdoMac) inhibits the growth of Trypanosoma brucei brucei with an IC 50 of 5.2μM.

S-adenosylmethionine decarboxylase inhibitors: new aryl and heteroaryl analogues of methylglyoxal bis(guanylhydrazone).

A series of 3-acylbenzamidine (amidino)hydrazones 7a-h, the corresponding (hetero)aromatic congeners 7i-p, and 3,3'-bis-amidino-biaryls 25a-e were synthesized. The hydrazones 7a-p were prepared by conversion of the corresponding acyl nitriles 1a,c-d,i,n-p to the imido esters 3a,c-d,i and the amidines 5a,c-d,h-i, followed by a reaction with aminoguanidine, or vice versa. Similarly, the biaryl 3,3'-dinitriles 23a-e were converted, via the imino esters 24a-c or the imino thioesters 27d-e, to the diamidines 25a-e. These new products are conformationally constrained analogues of methylglyoxal bis(guanylhydrazone) (MGBG). They are up to 100 times more potent as inhibitors of rat liver S-adenosylmethionine decarboxylase (SMDC) and generally less potent inhibitors of rat small intestine diamine oxidase (DAO) than MGBG. Some of these SAMDC inhibitors, e.g., compounds 7a, 7e, 7i, 25a, and 25d, have shown antiproliferative effects against T24 human bladder carcinoma cells. These products, whose structure-activity relationships are discussed, are of interest as potential anticancer agents and drugs for the treatment of protozoal and Pneumocystis carinii infections.

Changes in Polyamine Metabolism during Somatic Embryogenesis in Picea abies

Summary The concentrations of free, conjugated and bound polyamines (putrescine and spermidine) and the activity of a key enzyme, [S-adenosylmethionine decarboxylase (SAMdc)], which participates in the biosynthesis of spermidine, were studied in embryogenic and non-embryogenic callus cultures of Picea abies (L.) Karst. When the calli were transferred on ABA (10 gM) containing maturation media, where proembryos in embryogenic callus develop to mature embryos, both tissues contained high levels of free and conjugated putrescine. After 5 days the putrescine levels decreased in both tissues and did not rise thereafter. The total spermidine level (free, conjugated and bound) was nearly four-fold higher in the embryogenic callus in comparison to the non-embryogenic tissue during the development of globular embryos in the embryogenic callus. In our cultures a positive correlation was found between spermidine concentration and SAMdc-activity. If methylglyoxal bis(guanylhydrazone) (0.1 mM), an inhibitor of SAMdc, was added to the nutrient medium of embryogenic cultures, the levels of total spermidine decreased to nearly half, although further development of somatic embryos was not inhibited.