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Carbamoylphosphonates (CPOs) have recently been identified as extracellular in vivo active inhibitors of the cancer and metastasis-promoting extracellular enzymes, carbonic anhydrases (CAs) IX and XII, and matrix metalloproteinase-2 (MMP-2). This article describes the stereoselective synthesis and the evaluation of the biological properties of a pair of enantiomeric aminoalkylcarbamoylphosphonates, which have been enantioselectively synthesized from l-serine, using functional group-transformations. The structures of the enantiomeric products have been determined by X-ray crystallography. The enantiomeric purities of compounds have been confirmed by chiral shift reagent NMR experiments and by circular dichroism. In vitro evaluation of the CPOs synthesized revealed that they possess micromolar IC50 inhibitory action against CAIX and CAXII. The two enantiomers as well as the racemic or optically inactive ones do not differ by any significant extent from previously reported CPOs’ CA inhibitory values. On the other hand, MMPs inhibitory activities are rather weak; only MMP-2 showed a notable IC50 value.

Seven 4-phenoxybenzenesulfonamidopolymethylene carbamoylphosphonates (CPOs) bearing two to eight methylene units in the polymethylene chain were synthesized and evaluated as matrix metalloproteinase (MMP) inhibitors. The five lowest homologues [(CH₂)₂-₆] are selective MMP-2 inhibitors, whereas the two with the longest linkers [(CH₂)₇,₈] lack inhibitory activity. The most potent homologues are those with (CH₂)₅,₆; these two were evaluated for antimetastatic activity in a murine melanoma model and showed good potency both by oral and intraperitoneal administration without any toxic--including musculoskeletal--side effects. In contrast to the previously reported cis-ACCP, which was shown to inhibit MMP-2 for ∼30 min, the new compounds inhibit MMP activity for the duration of measurement, lasting several hours. Pharmacokinetic evaluation revealed, on the one hand, low oral bioavailability; on the other hand, a relatively large calculated volume of distribution, consistent with the observed reversible absorption of CPO 5 to hydroxyapatite, as a model for bone.

Low molecular weight compounds were isolated by high-performance liquid chromatography from the maggot or haemolymph extracts of Lucilia sericata (Meigen) (Diptera: Calliphoridae). Using gas chromatography-mass spectrometry analysis, three compounds were obtained: p-hydroxybenzoic acid (molecular weight 138 Da), p-hydroxyphenylacetic acid (molecular weight 152 Da) and octahydro-dipyrrolo[1,2-a;1',2'-d] pyrazine-5,10-dione (molecular weight 194 Da), also known as the cyclic dimer of proline (or proline diketopiperazine or cyclo[Pro,Pro]). All three molecules revealed antibacterial activity when tested against Micrococcus luteus and/or Pseudomonas aeruginosa, and the effect was even more pronounced when these molecules were tested in combination and caused lysis of these bacteria.

Bisacylphosphonates have previously been shown to possess in vitro activity with regard to hydroxyapatite (HAP) formation and dissolution, and in vivo on tissue calcification and bone resorption. This work was aimed at elucidating the role of the keto groups in the biological activity of these compounds. For this purpose, we have synthesized a series of long-chain bisphosphonates possessing different functional groups adjacent to the phosphonic functions and examined them in some in vitro and in vivo models. The functional groups introduced into the newly synthesized bisphosphonates (hydroxyl, amide, and difluoromethylene) were chosen as to represent certain isolated aspects of the keto groups in the original compounds. None of the functional groups introduced into the long chain bisphosphonates bestowed higher activity on the molecules than the carbonyl groups. The unique effect of the α-keto groups in rendering long-chain bisphosphonates active is presumably attained through a combination of chelating ability with sufficiently low pKa values to enable the bisacylphosphonate molecules to be fully ionized at physiological pH. © 2000 John Wiley & Sons, Inc. Heteroatom Chem 11:470–479, 2000

Solid-state 31P and 13C magic angle spinning (MAS) NMR spectra were used to characterize pure bisphosphonates and also bisphosphonates adsorbed on hydroxyapatite. Four geminal bisphosphonates, including the clinically used compounds ethane-1-hydroxy-1,1-diphosphonic acid, 3-amino-1-hydroxypropane-1,1-diphosphonic acid and 4-amino-1-hydroxybutane-1,1-diphosphonic acid, five α,ω-bisphosphonates and phosphonoacetic acid were investigated. NMR spectra of pure and adsorbed bisphosphonates differ in the observed linewidths and in the isotropic chemical shifts. The broad lines reflect mainly the poor crystallinity of the adsorbed compounds. Shifts of δiso in both directions do not reveal a correlation with the molecular structures. The molar ratio of phosphonates adsorbed on hydroxyapatite determined by 31P spectra without cross-polarization (CP) is approximately two times larger for geminal bisphosphonates than for α,ω-bisphosphonates and phosphonoacetic acid. 13C CP/MAS spectra of pure and adsorbed bisphosphonates recorded in two cases for identification of adsorbed compounds give additional information about the state of adsorbed compounds. Disodium α,ω-dihydroxypolymethylene-α,ω-bisphosphonates in the solid state show characteristic 13C chemical shifts which are indicative of either odd or even numbers of CH2 groups. Copyright © 2000 John Wiley & Sons, Ltd.

Bisphosphonates are clinically used mainly to reduce bone resorption. We studied the transplacental effects of two bisphosphonates on the fetal skeleton in rats. Pregnant rats were treated during days 11-20 of pregnancy with daily subcutaneous injections of 0.1 mg/kg of alendronate or a newly synthesized bisphosphonate, VS-b6. This period of pregnancy was chosen because the active development of bones from mesenchyme through cartilaginous models occurs during that time. Histological examination of midlongitudinal sections of the 21-day-old fetuses showed an increase in the amount of diaphyseal bone trabeculae with slight shortening of the diaphysis in the experimental fetuses, in comparison to controls. Computerized histomorphometric studies similarly showed an increase in the amount of diaphyseal bone trabeculae with a concomitant decrease in bone marrow volume, but no change in cartilage volume. In addition, chemical analysis of the fetal bones showed an increase in calcium content in the treated fetuses. 14C-alendronate was shown to pass through the rat placenta and accumulate in the fetuses, most probably in their bones. This is presumed because bisphosphonates are known to accumulate in bone, being stored there for long periods of time. It is important, in light of our results, to give careful consideration to the treatment of women with bisphosphonates at childbearing age, whenever this is needed.

We describe results of our initial experiments regarding the use of various protecting groups for the preparation of some N-protected aminoacylphosphonates and amino hydroxyimino phosphonates derived from them. The protecting groups that have been examined are benzyloxycarbonyl (Cbz), [(9-fluorenylmethyl)oxy]carbonyl (Fmoc), and phthaloyl (Pht)

Modified amino acids and peptides are in the focus of medicinal chemistry as potential mimics of natural compounds that may act as inhibitors and modulators of natural processes. Along with many other types of analogs, considerable effort was invested into the synthesis of phospha amino acids. Presently phospha analogs for all amino acids and for many peptides are known and some of these are of considerable practical importance.

The reaction of benzoylphosphonic acid (3) with thionyl chloride leads to the formation of benzoylphosphonic dichloride (5) and P,P′-dibenzoylpyrophosphonic dichloride (6). These products were identified by 31P nmr spectra and through their products of alcoholysis with 2-propanol.

Arbuzov reactions of diethyl 2,2,2-trihaloethyl phosphites (3) with benzoyl chloride afforded ethyl 2,2,2-trihaloethyl benzoylphosphonates (4). The reactions of 4 with NH2OH.HCl led to the formation of methyl benzoate and ethyl methyl H-phosphonate as a result of alcoholysis of 4, followed by alkoxy group exchange. Methanol solutions of benzoylphosphonates 4 were found by 31P NMR spectroscopy to contain considerable proportions of hemiacetals, 7, which undergo base-catalyzed CP bond cleavage. The formation of hemiacetals from benzoylphosphonates 4 is suppressed in 2-propanol, and in this solvent the corresponding oximes 2 could be obtained in good yields. Reactions of methyl benzoylphosphonochloridate (10) with 2,2,2-trihaloethanols, in CH2Cl2, gave methyl 2,2,2-trihaloethyl benzoylphosphonates (11) which could be converted directly to oximes 12 by NH2OH.HCl in a one-pot procedure. In contrast to the previously studied dimethyl (E)-α-hydroxyiminobenzylphosphonate, which underwent thermal Beckmann rearrangement to afford N-benzoylphosphoramidates, both (E) and (Z)-trihaloethyl esters 2 and 12 underwent fragmentation to benzonitrile and to the corresponding dialkyl hydrogen phosphate, reflecting the increased electrophilicity of the phosphorus in these compounds. Demethylation of methyl esters 12 was effected smoothly by iodide or bromide ions to yield benzoylphosphonate salts 15, which in turn were converted to oxime salts 14 by treatment with hydroxylamine. In contrast, attempted deethylation of ethylesters 2 in refluxing acetonitrile led to benzonitrile and pyrophosphate type product as indicated by 31P spectroscopic examination of the reaction mixture. Oxime salts 14 behaved similarly when heated. Acidification of lithium 2,2,2-trifluoroethyl α-hydroxyiminobenzylphosphonate (14a) gave the corresponding hydrogen trifluoroethyl phosphonate (19a). The fragmentation of 19a in 0.6 N ethanolic hydrogen chloride to ethyl trifluoroethyl hydrogen phosphate and benzonitrile at room temperature had a T1/2 value of approx 18 hours, which is greater by a factor of 2 than that of the corresponding methyl ester. When the fragmentation of 19 was carried out in solvent mixtures of either water with methanol or 2-propanol, or methanol with tbutanol, the composition of the solvents was reflected in the products, indicating a dissociative type mechanism, involving metaphosphate as reactive intermediates.

Methyl benzoylphosphonochloridate 3 reacted with 2-cyanoethanol or p-nitrophenethyl alcohol to yield methyl 2-cyanoethyl benzoylphosphonate 4a or methyl p-nitrophenethyl benzoylphosphonate 4b. Ketones 4a and 4b were converted into the corresponding oximes, 5a and 5b as a mixture of E and Z isomers, by hydroxylamine. Oximes 5a and 5b were demethylated by treatment with sodium iodide or lithium bromide to give the corresponding monester monosalts 6a and 6b. Reaction of lithium 2-cyanoethyl or p-nitrophenethyl benzoylphosphonate with methoxylamine afforded the respective oxime methyl ether salts 8a and 8b as predominantly E isomers. Base treatment of 6a and 6b caused their dealkylation to the anion of α-hydroxyiminobenzylphosphonic acid 9 which did not decompose further. In contrast, 8a and 8b in aqueous base gave the dianion of α-methoxyaminobenzylphosphonic acid 10, which slowly fragmented to benzonitrile and phosporic acid. The fragmentation is interpreted in terms of a dissociative mechanism involving the formation of metaphosphate anion.

Arbuzov reactions of diethyl 2,2,2-trihaloethyl phosphites (3) with benzoyl chloride afforded ethyl 2,2,2-trihaloethyl benzoylphosphonates (4). The reactions of 4 with NH2OH.HCl led to the formation of methyl benzoate and ethyl methyl H-phosphonate as a result of alcoholysis of 4, followed by alkoxy group exchange. Methanol solutions of benzoylphosphonates 4 were found by 31P NMR spectroscopy to contain considerable proportions of hemiacetals, 7, which undergo base-catalyzed CP bond cleavage. The formation of hemiacetals from benzoylphosphonates 4 is suppressed in 2-propanol, and in this solvent the corresponding oximes 2 could be obtained in good yields. Reactions of methyl benzoylphosphonochloridate (10) with 2,2,2-trihaloethanols, in CH2Cl2, gave methyl 2,2,2-trihaloethyl benzoylphosphonates (11) which could be converted directly to oximes 12 by NH2OH.HCl in a one-pot procedure. In contrast to the previously studied dimethyl (E)-α-hydroxyiminobenzylphosphonate, which underwent thermal Beckmann rearrangement to afford N-benzoylphosphoramidates, both (E) and (Z)-trihaloethyl esters 2 and 12 underwent fragmentation to benzonitrile and to the corresponding dialkyl hydrogen phosphate, reflecting the increased electrophilicity of the phosphorus in these compounds. Demethylation of methyl esters 12 was effected smoothly by iodide or bromide ions to yield benzoylphosphonate salts 15, which in turn were converted to oxime salts 14 by treatment with hydroxylamine. In contrast, attempted deethylation of ethylesters 2 in refluxing acetonitrile led to benzonitrile and pyrophosphate type product as indicated by 31P spectroscopic examination of the reaction mixture. Oxime salts 14 behaved similarly when heated. Acidification of lithium 2,2,2-trifluoroethyl α-hydroxyiminobenzylphosphonate (14a) gave the corresponding hydrogen trifluoroethyl phosphonate (19a). The fragmentation of 19a in 0.6 N ethanolic hydrogen chloride to ethyl trifluoroethyl hydrogen phosphate and benzonitrile at room temperature had a T1/2 value of approx 18 hours, which is greater by a factor of 2 than that of the corresponding methyl ester. When the fragmentation of 19 was carried out in solvent mixtures of either water with methanol or 2-propanol, or methanol with tbutanol, the composition of the solvents was reflected in the products, indicating a dissociative type mechanism, involving metaphosphate as reactive intermediates.

This article reviews the unique reactivities of � -hydroxyiminoalkylphosphonates, which result from the proximity and the interaction between the hydroxyimino and the phosphonic groups. The reaction course of � -hydroxy- iminoalkylphosphonates depends markedly on the type of groups linked to the phosphorus and on the stereochemistry of the oxime function. (Z)-� -Hydroxyiminoalkylphosphonates and phosphinates undergo, upon heating, fragmentation to a nitrile and a phosphate or phosphonate, while (E)-� -hydroxyiminoalkylphosphonates and -phosphinates undergo Beckmann rearrangement to N-acylphosphoramidates or N-acylphosphonamidates, respectively. In contrast to the diesters, � -hydroxyiminoalkylphosphonic acids (and monoesters) are unstable and fragment to metaphosphates, which can perform in situ phosphorylation of appropriate groups. Kinetic and mechanistic studies of the fragmentations of � - hydroxyiminobenzylphosphonates to metaphosphoric acid and esters are reviewed. The fragmentation of � -hydroxyimino- benzylphosphonic acid monoesters to metaphosphate esters requires acid catalysis and slows down at about pH 3. In contrast, � -hydroxyiminobenzylphosphonic acid undergoes fragmentation in aqueous solutions at a wide range of pH, from strongly acidic up to about pH 9. The � -hydroxyiminophosphonate fragmentation methodology was applied to phosphorylation of silica gel, which can potentially be useful as chromatographic stationary phase. A variety of stable precursors of � - hydroxyiminophosphonates that can be induced to fragment thermally, photochemically or by base catalysis is described. (E)-� -Hydroxyiminobenzylphosphonamidates of various types have been found to undergo Beckmann rearrangement to yield N-acylphosphordiamidates upon heating in nonpolar solvents such as toluene, or fragmentation to metaphosphonamidate by heating in polar solvents. The fragmentation of (E)-� -hydroxyiminobenzyl- phosphonates to metaphosphates is considered a special case of the Beckmann fragmentation.

Methyl trifluoropyruvate (1) and tris(trimethylsilyl) phosphite (3) reacted to give methyl 2-[bis(trimethylsiloxy)phosphoryl]- 3,3,3-trifluoro-2-(trimethylsiloxy) propionate (4). Partial hydrolysis furnished propionate 6, the molecular structure of which was obtained in the solid state. Attempted trimethylsilylation of the methylcarboxylato group in 4 using iodotrimethylsilane caused the formation of bis(trimethylsily1) [(2,2- difluoro- 1 -trimethylsiloxy)ethenyl]phosphonate (8). For comparison, methyl pyruvate (2) and 3 gave methyl 2-[bis(trimethylsiloxy)phosphoryl]-2-(trimethylsiloxy)propionate (5).

1,1,3,3,3-Pentafluoro-2-pentafluorophenyl-1,2-epoxypropane 1 reacted with trimethylphosphite giving two diastereomers, (Z)- and (E)-3,6-bis(trifluoromethyl)-3,6-bis(pentafluorophenyl)-1,4-dioxan-2,5-dione 2a, b in a 1:1 ratio, cyclodimerisation product of the intermediately generated α-lactone 4. Compounds 2a, b were hydrolysed to furnish 3,3,3-trifluoro-2-hydroxy-2-(2,3,4,5,6-pentafluorophenyl)propionic acid 5.

The thermodynamics, pH dependency and solvent effects of the fragmentation reaction of a series of α-oxyiminobenzylphosphonate monomethyl esters [(E)-1a–f] were examined in water and other hydroxylic solvents by UV and by 31P NMR spectroscopy at pH 0–3.1. The fragmentation of compounds (E)-1a–f was found to be a first-order reaction in substrate over the acidity range studied, while the dependence on the acidity is more complex, with rate constants k1 and k2. The ρ values corresponding to the first and second order rate constants were −1.12 and −0.835, respectively, indicating that the reaction is facilitated by electron-donating substituents, which probably enhance the protonation of the oxime OH group. Activation parameters for k1 and k2 reactions were also calculated. The near- zero values of the entropies of activation obtained are consistent with a dissociative transition state with almost no bonding to a nucleophilic solvent. Monitoring the fragmentation reaction of (E)-1a in several binary alcohol–water mixtures at different acidities showed that the reaction rate is enhanced by the alcohol’s acidity and not hampered by the steric requirements of the alcohol molecule. This rules out in our opinion, the likelihood for nucleophilic solvent assistance in the rate-determining step. On the other hand, product studies show that both the nucleophilicity and the steric requirements of the alcohol are of importance in determining the product formed in the fragmentation of (E)-1a. The highest selectivity (S) value was found for MeOH, while S values of

Proton magnetic resonance spectra of trans and cis-2,3-diphenylaziridine (1 and 2) and their N-ethyl derivatives 3 and 4 were measured in carbon tetrachloride, chloroform, and benzene-d6 at low temperatures (1 and 3) and in dry conditions (1 and 2). On the basis of these results it was concluded that an N-ethyl group exerts a shielding influence on a cis ring proton and a deshielding influence on a trans ring proton. From results obtained by measuring the 1H n.m.r. spectra of 1–4 in deuterochlorofom-trifluoroacetic acid it was derived that the lone pair of the aziridine nitrogen exerts a shielding influence on cis related ring hydrogens. In most N-alkylaziridines the effect of the N-alkyl group predominates.

Reactions of aryl cyclopropyl ketones with formamide, in the presence of magnesium chloride, give predominantly the corresponding 1-formyl-2-arylpyrrolidines (II).

The synthesis of some p-substituted phenylmethylcyclopropylcarbinols of the structure (I) is herein reported. Dehydration of the cyclopropylcarbinols by means of acetic anhydride yielded mixtures of α-cyclopropylstyrenes (V) and 4-arylpent-3-enyl acetates (VI) the ratio of which depended on both the polar characteristics of the substituent X and on reaction conditions. The relative order of the effect of substitution in the phenyl group on V/VI ratio is Cl > H > CH3 > OCH3. The data presented being interpreted in terms of stability of the cyclopropylbenzylium ions (III). The more stable carbonium ions (III or IV) would favor the formation of rearranged products (VI) whereas the less stable would favor the production of V.

1, 3, 3, 4, 4-Pentaphenylazetidinone-2 (I) is converted by sulfuric acid into 1, 3, 3, 4-tetraphenylhydrocarbostyril. Upon treatment with lithium aluminum hydride (I) undergoes a fragmentation reaction from which benzophenone anil is isolated.

Condensation of 3-ketobisnorcholenal (1) with malononitrile could be directed selectively to the aldehyde group to afford dicyanomethylene ketone 3. Ketalization of 3 with ethylene glycol followed by alkaline hydrolysis gave the aldehydo dioxolane 5, which served as starting material for the syntheses of 1,3-propanediol derivatives 7 and 10 and their respective dicarbamates 9 and 11.

A new synthetic approach to vinylphosphonates is presented. The reaction of diethyl benzoylphosphonate (1) with diethyl methoxycarbonyl-methyl-phosphonate (2) gives diethyl 2-methoxycarbonyl-l-styrylphosphonate (3) as a mixture of geometrical isomers (Z-3) and (E-3) in the ratio of 83:17. In contrast, reaction of 1 with the heterocyclic phosphonate reagent, 2-methoxycarbonyl-methyl-2-oxo-4,5-dimethyl [1,3,2] dioxaphospholane (4) leads to a mixture of the isomers (Z-3) and (E-3) in the ratio of 20:80.

The reaction of 2-benzoyl-2-oxo-4,4,5,5–tetramethyl [1,3,2] dioxaphospholane (3) with hydroxylamine yields the corresponding E oxime (E-4) and benzonitrile (5), which arises from fragmentation of the Z oxime via an intramolecular attack of the N-OH on the phosphorus.

(E)-α-Hydroxyiminobenzylphosphonic acid decomposes in aqueous solution to benzonitrile and phosphoric acid in a pH dependent manner, which is consistent with a dissociative mechanism involving, in the first step, the formation of monomeric metaphosphate.

Reaction of N-methyl-C-phenyl nitrone with sodio triethyl phosphonoacetate leads to trans-I-methyl-2-ethoxycarbonyl-3-phenylaziridine and to 1-benzyl-2-ethoxycarbonylaziridine.

Reaction of 5,5-dimethyl-Δ1-pyrroline N-oxide (2) with 2-cyanomethyl-4,5-dimethyl-2-oxo-1,3,2-dioxaphospholan (1b) gives exo-6-cyano-2,2-dimethyl-1-azabicyclo[3.1.0]hexane (3), but reaction of (2) with 2-cyanomethyl-5,5-dimethyl-2-oxo-1,3,2-dioxaphosphorinan (1c) gives 5,5-dimethyl-2-cyanomethylenepyrrolidine (4).

Acylphosphonic acids (5) and methyl hydrogen acylphosphonates (3) were synthesized by di- and mono-demethylation of dimethyl acylphosphonates (1). Spectroscopic data (i.r., 31P and 1H n.mr.) are reported for these types of compounds for the first time. Examination of their hydrolytic stability under acidic and basic conditions revealed that except for methyl hydrogen acylphosphonates (3) that are unstable under highly alkaline pH conditions, the C–P bond in these types of compounds is stable in most cases. Nucleophilic reagents, e.g. amines, borohydride, or hydroxylamine react with the carbonyl group of ionized acylphosphonates with the preservation of the C–P bond, to yield α-imino-, α-hydroxy-, or α-oxyimino-alkylphosphononate anions, respectively. Semi-empirical quantum mechanical (MNDO/H) calculations were performed on benzoylphosphonic acid (5c) and on the esters and anions derived from it, as representatives of their classes, in order to assess bond lengths and preferred conformations, and to estimate charges on the carbonyl and phosphoryl groups. Calculations show that for both neutral and ionized (anions) compounds free rotation around the C–P bond is expected due to the low energy barriers.

Der Aldehyd (I) reagiert mit Acetylaceton (II) zum Kondensationsprodukt (III), dessen Saurehydrolyse die Verbindung (IV) ergibt.

Although dialkyl acylphosphonates (1) have been known for over four decades,1 their dealkylated derivatives, namely acylphosphonic acids (2) and alkyl hydrogen acylphosphonates (3), have only been isolated as salts. Characterization of compounds of type 2 and 3 have not been previously reported, neither have their chemical and physical properties been described. In this paper we wish to describe some new chemical properties of these types of compounds.