N-terminal anthranoyl-phenylalanine derivatives as CCK1 receptor antagonists: the final approach.

Starting from our lead compound, VL-0395, an anthranilic acid based CCK1 receptor antagonist, and following the well established "step by step" lead investigation strategy, we describe the final step of the anthranilic acid N-terminal optimization. Improvements for both affinity and selectivity towards CCK1 receptors have been accomplished through introduction of the fluoro substituent at C-5 and C-7 position of the indole ring together with the appropriate configuration of the aminoacidic chiral center.

Synthesis of N-terminal substituted anthranilic acid dimer derivatives for evaluation on CCK receptors.

A series of new N-substituted anthranilic acid dimer derivatives having a C-terminal Phe residue was synthesized and evaluated for their affinity for CCK receptors. These compounds resulted from a blended approach based firstly on the use of an alternative substructure embedded within asperlicin and secondly on the derivatization of this template with substituents chosen considering the C-terminal primary structure of the endogenous ligand. Although these compounds exhibited a regnylogical-type organization similar to that of CCK-4, they are characterized by about 1000-fold greater affinity for CCK-A receptor than the C-terminal tetrapeptide.

Synthesis of new anthranilic acid dimer derivatives and their evaluation on CCK receptors.

We have described previously an innovative bond disconnection strategy of asperlicin, a naturally occurring CCK receptor antagonist, leading to anthranilic acid dimer and tryptophan synthons. We have also demonstrated that when the tryptophan residue is connected to the C- or N-terminal sides of the anthranilic acid dimer, compounds with similar micromolar CCK-A receptor affinities are obtained. In order to investigate the binding effects of different N-terminal substitution, in this paper we describe a new series of anthranilic acid dimer derivatives, characterized by the presence of the tryptophan residue in the C-terminus of the dimer. Among the compounds synthesized, the N-1H-indol-3-propionyl derivative exhibited an improved, at the micromolar range, affinity for the CCK-A receptor in comparison to that of either, the N-unsubstituted derivative and asperlicin. The lead compound emerging from this key step of our investigation represents the new starting point for the development of a new class of CCK-A receptor ligands.

C-terminal anthranoyl-anthranilic acid derivatives and their evaluation on CCK receptors.

A series of C-terminal anthranoyl-anthranilic acid derivatives arising from a strict bond disconnection approach of asperlicin were synthesized and examined for their CCK receptor affinities. These compounds represent the second step of our investigation directed toward the search for alternative substructures of asperlicin as a starting point for the development of a new class of CCK ligands. The obtained micromolar affinities for CCK-A rather than CCK-B receptor confirm that the anthranilic acid dimer represents a useful template for the development of selective CCK-A receptor ligands.

Improved activity in acidic media of immobilized lysozyme.

Polyethylene glycols (PEGs) of various chain length were used to crosslink lysozyme onto an insoluble support such as oxirane. A very high degree of modification and no inactivation of lysozyme were obtained with PEG 20000, but enzymatic activity increased up to 20 times at pH 3.0, at which point the activity of the native enzyme was lower when using Leuconostok oenus as a macromolecular substrate.

Quinazolinone derivatives: synthesis and binding evaluation on cholecystokinin receptors.

A series of 2-methyl-3-amino-4(H)-quinazolinone and of 2-phenyl-3-amino-4(H)-quinazolinone derivatives were synthesized and examined for their CCK receptor affinities. These compounds displayed micromolar affinities for CCK-B rather than CCK-A receptor and the obtained results confirm that the 4(3H)-quinazolinone nucleous represent a useful template for the development of selective CCK-B receptor ligands.

Quinolone derivatives: synthesis and binding evaluation on cholecystokinin receptors.

A series of 1,2-dihydro-4-phenylquinolin-2-one-3-carboxylic acid and of 3-amino-4-phenylcarbostyril derivatives were synthesized and examined for their CCK receptor affinities. These compounds displayed micromolar affinities for CCK-A rather than CCK-B receptor and the results have been discussed on the basis of a molecular modelling study.

Synthesis and antitumor activity of hydrosoluble analogs of p-(3,3-dimethyl-1-triazeno) benzoic acid potassium salt.

The hydrosoluble triazene derivatives of phenylacetic, phenylbutyric and cinnamic acid have been synthesized and their logP and pKa values were simultaneously determined according to a multiparametric fitting of potentiometric data. The antitumor activity caused by the synthesized compounds in mice bearing either Lewis lung carcinoma or TLX5 lymphoma was evaluated and discussed in comparison with the parent compound (p-(3,3-dimethyl-1-triazeno)benzoic acid potassium salt (DM-COOK, CAS 70055-49-1). The tested compounds were at least as active as DM-COOK, the cinnamic and the phenylacetic derivatives being the more active compounds in mice bearing TLX5 lymphoma and Lewis lung carcinoma, respectively.

Antineoplastic action of p-(3-methyl-1-triazeno)benzoic acid potassium salt, a monomethyl derivative of the antimetastatic compound DM-COOK.

The antitumor and antimetastatic effects of p-(3-methyl-1-triazeno)benzoic acid potassium salt (MM-COOK) as compared with those of the parent 3,3-dimethyl derivative (DM-COOK) were examined using Lewis lung carcinoma, MCa mammary carcinoma of the CBA mouse and TLX5 lymphoma. Similarly to DM-COOK, MM-COOK reduces metastasis formation and significantly prolongs the survival of mice bearing the Lewis lung carcinoma when given at a daily dose corresponding to one-half that of DM-COOK. Unlike DM-COOK, MM-COOK exhibits significant cytotoxicity to metastatic foci and pronounced inhibition of primary tumor development. MM-COOK also causes cytotoxic effects on TLX5 lymphoma cell growing in the peritoneal cavity, even when used at low doses. The antimetastatic effects observed in mice bearing MCa mammary carcinoma are unrelated to the inhibition of primary tumor growth and are more likely due to the selection of clones endowed with lower metastatic ability. It appears that MM-COOK exhibits the same antineoplastic activity as DM-COOK, but the former does so at a lower daily dose and produces interesting cytotoxic effects other than those reflecting its antimetastatic properties. It thus seems to be a valid alternative to DM-COOK, in view of the possible introduction of newer aryltriazenes into clinical use.

Kinetic investigation of the aqueous stability and antitumor activity of a hydrosoluble diaryltriazene, AVIS, related to the antimetastatic agent DM-COOK.

The aim of this study is to investigate the hydrolysis of 1,3-di(p-carboxyphenyl)triazene dipotassium salt, AVIS (1), over a pH range of 2.60-8.50. This compound decomposes into p-aminobenzoic acid and the corresponding diazonium cation with no formation of alkylcarbo cations; the same compounds are formed from hydrolyses of DM-COOK (2), an antimetastatic agent, and of its possible demethylated metabolite, MM-COOK (3), a chemical xenogenization inducer. In these latter cases, however, a methylcarbo cation is formed. The pH dependence of the pseudo-first-order rate constants is intermediate between 2 and 3. Preliminary data on its toxicity and antitumor activity on both Lewis lung carcinoma and TLX5 lymphoma seem to indicate the essential role of alkylcarbo cation in mediating the antitumor action of aryldimethyltriazenes.

Kinetic study employing UV derivative spectroscopy of DM-COOK, antitumor and antimetastatic agent.

The hydrolysis of p-(3,3-dimethyl-l-triazeno)benzoic acid potassium salt (1;DM-COOK) a highly active antimetastatic and anti-disseminative agent, has been studied in buffered aqueous solution over a pH range of 2.8-8.8 degrees C. The pH dependence of the pseudo first-order rate constants showed two different routes. Under physiological conditions the hydrolysis reactions are carried out by acid catalysis. A procedure based on fourth-order derivative UV spectroscopy (D4) has been developed for the calculation of the kinetic constants at pH greater than or equal to 4.00 and no spectral interferences resulted from decomposition products. The application of derivative UV spectroscopy proved to be suitable fpr rapid, sensitive and reproducible studies of hydrolysis of this class of compounds.

Synthesis and antitumor activity of aminoacid derivatives of the antimetastatic agent DM-COOK.

Esters and amides of N-(3,3-dimethyl-1-triazeno)benzoylamino acids have been synthesized as inhibitors of cell surface neutral proteases. Preliminary data on activity against P-388 lymphocytic leukemia are also reported.