Aromatic Diamidines Research Articles

Inhibition of the haemolytic complement activity by derivatives of benzamidine

The influence of a series of benzamidine derivatives on the haemolytic complement activity is studied. Complement-mediated haemolysis is inhibited by the derivatives tested to a different extent. Aromatic diamidines are found to be relatively potent inhibitors. Hydrophobic substituents of benzamidine are believed to enhance the inhibitory activity. Relations to the inhibitory action of benzamidine derivatives on serine proteinases of the blood are discussed.

Studies on the structure—activity relationship among aliphatic and aromatic bisguanylhydrazones and some related compounds

A total of 18 compounds consisting of 7 aliphatic and 7 aromatic bis(guanylhydrazones), p-quinone-bis(guanylhydrazone), one monoguanylhydrazone, one diamidine and one diguanidine were studied spectrophotometrically to determine their ability to interact with native calf-thymus DNA and the possible correlation of binding with biological activity. In each case, the ability of a compound to bind to DNA correlated with its ability to inhibit the activity of DNA-dependent DNA polymerase (EC 2.7.7.7) extracted from mouse leukemia L1210 cells. For example, all the aromatic bis-guanylhydrazones and diamidine (hydroxystilbamidine), which were good inhibitors of the enzyme activity, showed a biphasic interaction with DNA. All the aliphatic compounds displayed no detectable interaction with DNA in the Tris buffer used, and were also poor inhibitors of the polymerase activity. Interaction of decamethylene diguanide (Synthalin) with DNA could not be determined because the compound does not absorb light in the UV-VIS region. However, in similarity with other aliphatic compounds, this agent was a poor inhibitor of DNA polymerase reaction. The p-quinone-bis(guanylhydrazone) and p-phenylbenzaldehyde-monoguanylhydrazone showed only a monophasic interaction with DNA and caused an intermediate inhibition of the enzyme activity. When tested for possible anti-leukemic activity against i.p. L1210 leukemia in syngeneic DBA/2J mice, all the aromatic bisguanylhydrazones as well as hydroxystilbamidine caused prolongation of survival of tumor-bearing mice. Among the aliphatic bisguanylhydrazones, all of which showed no binding to DNA and caused at the most only a very slight inhibition of DNA polymerase, only methylglyoxal-bis(guanylhydrazone) (CH 3-G) had antileukemic activity. Synthalin also inhibited leukemic growth. Evidences presented indicate that the mechanisms of action of aliphatic and aromatic bisguanylhydrazones may be quite different. Furthermore, the ability to bind to DNA may be a useful criterion to predict the antileukemic activity of aromatic guanylhydrazones and possibly other aromatic bis-cationic compounds, but not that of aliphatic congeners.

ChemInform Abstract: NOVEL BIS(BENZAMIDINO) COMPOUNDS WITH AN AROMATIC CENTRAL LINK. INHIBITORS OF THROMBIN, PANCREATIC KALLIKREIN, TRYPSIN, AND COMPLEMENT

AbstractUnter Verwendung bekannter Verfahren werden verschiedene Bis‐benzamidine mit aromatischen Bindegliedern hergestellt.

Pentamidine transport in Trypanosoma brucei—Kinetics and specificity

Pentamidine, an aromatic diamidine used in the treatment and control of African trypanosomiases, is rapidly absorbed by Trypanosoma brucei and is lethal to the parasite both in vitro and in vivo. The pentamidine transport system in T. brucei exhibits saturation kinetics and shows specificity for the aromatic amidine moiety of this compound. The inhibition of uptake by salicyl hydroxamic acid (SHAM) and iodoacetate (IAA), and the retention of intracellular substrate against large chemical gradients indicate a coupling of energy to this system. The system shows high affinity for pentamidine with an average K m value of 2.68 μM. Structural specificity in relationship to varying drug affinities for this transport site and the role it might play in drug resistance are discussed.

New aromatic diamidines with central alpha-oxyalkane or alpha, omega-dioxyalkane chains. Structure-activity relationships for the inhibition of trypsin, pancreatic kallikrein, and thrombin and for the inhibition of the overall coagulation process.

A series of omega-amidinophenylalkyl amidinophenyl ethers was synthesized and examined for inhibitory activity against trypsin, pancreatic kallikrein, and thrombin. Modifications of the compounds included lengthening of the alkane chain, variation in the position of the amidino groups, and substitution of halogen on the benzene rings. The compounds act as competitive reversible inhibitors, and many of them possess considerable potency. An outstanding trypsin inhibitor was found in 4-amidinophenylethyl 4 amidino-2-bromophenyl ether (compound 7) with a Ki value of 7.3 x 10(-8) M (pH 8.1, 37 degrees). A number of aromatic diamidines with a central dioxyalkane chain were similarly studied. Here, 1-(4-amidino-2-iodophenoxy)-5-(3-amidinophenoxy)pentane (compound 32) was a highly effective inhibitor of bovine thrombin (Ki = 1.1 x 10(-6) M), of human thrombin, and of the overall clotting process of human plasma.

Interaction Between Aromatic Diamidines and Nucleic Acids: Possible Implications for Chemotherapy

Interaction between aromatic diamidines (pentamidine, propamidine, and stilbamidine) and nucleic acids were studied to elucidate the mechanism underlying renal toxicity included by pentamidine in patients. Pentamidine, propamidine, and stilbamidine precipitated with RNA and DNA of all species and types. Furthermore, the diamidines interacted and precipitated with all nucleoside triphosphates and nucleoside diphosphates, but not with nucleoside monophosphates, nucleosides, or free bases. Nucleoside diphosphate is the minimum unit necessary for interaction with the diamidines. This interaction between diamidines and nucleotides or nucleic acids may be implicated in the etiology of renal damage and skin reactions associated with these drugs.

The Inhibition of Urokinase by Aromatic Diamidines

1. Structure-activity relationships have been established for the inhibition of urokinase by aromatic diamidines. In an assay system employing purified urokinase and human plasminogen the most potent inhibitor was found in 4',4''-diamidino-2-hydroxy-1,4-diphenoxybutane which proved 5600 times more active on a molar bases than epsilon-aminocaproic acid (E-ACA). 2. 4',4''-diamidino-2-hydroxy-1,4-diphenoxybutane behaved as a competitive inhibitor of the urokinase catalyzed hydrolysis of N-alpha-acetyl-L-lysine methyl ester. At pH 7.85 and 37 degrees C the K-1 value was determined as 3.18 times 10-6 M which compares with a value of 6.79 times 10-5 M for p-aminobenzamidine and 3.57 times 10-2 M for E-ACA. 3. In two fibrinolytic tests including urokinase as activator the superiority of diamidines over E-ACA was less marked than in the pure plasminogen activation system. This was due to the presence of certain plasma proteins in the fibrinolysis assays which augmented the inhibitory strength of E-ACA. The order of effectiveness of diamidines in the lysis tests was also different from the one in the activation test. In a human fibrin clot lysis test the most active inhibitor was 3',3''-diamidino-2-hydroxy-1,4-diphenoxybutane which was 1700 times more effective on a molar basis than E-ACA. In a human plasma clot lysis test the strongest inhibitor, 2-hydroxy-stilbamidine, was 70 times more powerful than E-ACA.

Structure-Activity Relationships for the Inhibition of Plasmin and Plasminogen Activation by Aromatic Diamidines and a Study of the Effect of Plasma Proteins on the Inhibition Process

Summary1. Structure-activity relationships for the inhibition of human plasmin were established for a large series of aromatic diamidines. The compounds are reversible competitive inhibitors and block the amidase and fibrinolytic activities of the enzym. The results confirm pentamidine (4,4’-diamidino α ω-diphenoxy-pentane) as the leading inhibitor (Ki = 3.3 (μM) and show distinct differences in the inhibitory spectrum of diamidines against plasmin as compared with trypsin, pancreatic kallikrein and thrombin.2. Diamidines are potent inhibitors of the SK-dependent activation of human plasminogen and of the activation of bovine plasminogen by the SK-human plasmin activator complex. Pentamidine is again the most powerful inhibitor of these systems.3. In fibrinolytic assays of plasmin and in plasminogen activation tests the relative strength of diamidines as compared with E-ACA is greatly influenced by the test conditions. The decisive factor is the presence in the incubation mixtures of lesser or greater amounts of plasma or serum proteins which bring about a fall in the absolute strength of diamidines and an increase in the absolute strength of E-ACA. In the fibrinolytic assay of plasmin, this modifying effect of added serum is based on a time- dependent interaction with the enzyme, thereby presumably altering its susceptibility to inhibition.

Effects of Protease Inhibitors on Protein Breakdown in Escherichia coli

Abstract In an attempt to learn more about the proteolytic enzymes responsible for protein turnover in cells, the effects of various protease inhibitors on protein catabolism in Escherichia coli were examined. Diisopropyl fluorophosphate, phenylmethane sulfonyl fluoride, and toluenesulfonyl fluoride, all of which inhibit serine proteases, were found to decrease protein breakdown in E. coli, starved for a carbon source. These compounds partially inhibited protein breakdown at doses which did not inhibit growth on minimal medium. The effects of the sulfonyl fluorides were reversed by washing the cells. Protein breakdown in starving cells could also be inhibited reversibly by the aromatic diamidines, dibromopropamidine and pentamidine, which are potent trypsin inhibitors. The effects of pentamidine and p-toluenesulfonyl fluoride did not require concomitant protein synthesis. Tosyl lysine chloromethyl ketone, an inhibitor of trypsin, also inhibited protein breakdown in starving cells. A number of other protease inhibitors were tested and found either to have no effect on intact cells or to prevent normal cell growth. These findings suggest that at least one of the enzymes responsible for the increased protein breakdown in starving cells is a serine protease, possibly similar to trypsin. In cells deprived of a carbon or nitrogen source, synthesis of new enzymes should be dependent upon the breakdown of previously existent cell proteins. Treatment with the sulfonyl fluorides or aromatic diamidines inhibited amino acid incorporation into proteins under such conditions, but did not do so in growing cells. Similarly these compounds reversibly inhibited the induction of β-galactosidase and tryptophanase in cells deprived of a carbon or nitrogen source, but did not do so in cells growing on glycerol. The sulfonyl fluorides, the aromatic diamidines, and tosyl lysine chloromethyl ketone did not reduce the relatively low amount of protein breakdown found in growing cells. Growing cells degrade selectively or aberrant proteins, and the rate of protein breakdown in such cells increased markedly when they were grown on amino acid analogs, canavanine or threo-α-amino-β-chlorobutyric acid. This accelerated degradation also was not inhibited by the sulfonyl fluorides or the diamidines. These results thus suggest that there may exist at least two proteolytic systems in E. coli: one operating in all cells, which can degrade abnormal proteins, and one found in starving cells which is sensitive to sulfonyl fluorides and aromatic diamidines.

Drug effects on the fine structure of trypanosoma rhodesiense: Diamidines

Three aromatic diamidine trypanocides, Berenil, hydroxystilbamidine and pentamidine, have been found to show similar and specific lesions in the fine structure of Trypanosoma rhodesiense taken from infected mice 5 to 6 hours after treatment with a curative dose of the drug. The primary effect of all 3 compounds was to disintegrate selectively the kinetoplast DNA core into round dense granules and micro-fibrils; the normal core is a uniform fibrillar lamellar structure. There was no effect on the nucleus. Possible causes for this selectivity are discussed in relation to the properties of kinetoplast DNA and the behaviour of trypanocides with an affinity for kinetoplast localization. Later effects of pentamidine include aggregation and loss of ribosomes, nucleolar dispersion, and formation of cytoplasmic membrane whorls intimately related to bodies resembling myelin figures. The latter may have been formed as a result of lysosomal enyme action, as large numbers of “empty” lysosomes were present. Lysosomal function is also discussed in relation to drug uptake and interaction, with some speculation on the rôle of lysosomes in drug-induced intracellular lesions in the trypanosome.

Inhibition of Coagulation and Fibrinolysis by Aromatic Amidino Compounds

Summary1. Aromatic diamidines which are potent inhibitors of trypsin possess a marked inhibitory effect on the clotting activity of human thrombin and on the prothrombin time and partial thromboplastin time of human plasma. They also block the contact activation phase of the coagulation process. The strongest inhibitor among the compounds tested was M & B 4596 which was followed in second place by pentamidine.2. Pentamidine was 10 times more active than ε-ACA in impeding streptokinase-induced lysis of human plasma clots. It was 100-200 times stronger than ε-ACA in inhibiting the activation of bovine plasminogen by activators formed from the interaction between streptokinase and either human plasmin(ogen) or human plasma.3. The prothrombin time and partial thromboplastin time of canine plasma were less susceptible to inhibition by pentamidine than the same tests on human plasma. Clot lysis in the canine system was inhibited by pentamidine to a similar degree as in the human system. After intravenous injection of pentamidine in the dog there occurred the expected prolongation of the partial thromboplastin time and of the clot lysis time.

Inhibitory effect of aromatic diamidines on trypsin and enterokinase.

Aromatische Diamidinoverbindungen sind hochwirksame Hemmstoffe des Trypsins. 2,2′-Dibromopropamidin, die starkste der hier untersuchten Verbindungen, besitzt gegenuber der BANA-Hydrolyse eine Inhibitions-Konstante von 7.6×10−7 M. Amicarbalid und M & B 4596 sind gute Hemmstoffe der Enterokinase, doch erreichen sie die Wirkung derp-Amidinophenylbrenztraubensaure nicht.

Therapy with amphotericin B in North American blastomycosis.

Excerpt The therapy of North American blastomycosis has not been completely satisfactory in the past. Although the aromatic diamidines, particularly stilbamidine and the less toxic 2-hydroxystilbam...

The toxicity of 2-hydroxystilbamidine: Probable fatal toxic reaction during treatment of blastomycosis.

Excerpt Following the original report of Schoenbach et al.1of the successful therapy of a case of systemic North American blastomycosis treated with the aromatic diamidine, stilbamidine, numerous p...

The treatment of North American blastomycosis with amphotericin B.

The many reports in the medical literature on the treatment of North American blastomycosis with the aromatic diamidines attest to the beneficial effect obtained by the use of these compounds in this specific fungus infection. 1-13 There is increasing evidence, however, that stilbamidine and 2-hydroxystilbamidine are not effective in curing all instances of the systemic type of North American blastomycosis. 9,14,15 This systemic form of the disease, which was usually fatal 16 prior to the use of the aromatic diamidines, has been beneficially influenced by this therapy, but often not cured. The relapses which have occurred following the proper administration of the stilbenes would appear to have developed in that group of patients exhibiting the immunological status comprising a negative blastomycin skin test and a persistently positive complement-fixation test, a type of blastomycosis which Smith 17 has categorized as having the poorest prognosis. Relapse following stilbamidine

Combined therapy in histoplasmosis and coccidioidomycosis; methyltestosterone and meth-dia-mer-sulfonamides.

THERE IS no reported instance of treatment of systemic fungous disease in man with testosterone compounds. In 1953 Lamb and his associates 1 reported the cure of a mycetoma caused by an atypical Nocardia under treatment with large doses of pregnenolone acetate. Trial of this steroid in disseminated coccidioidomycosis has been unsuccessful. * The use of aromatic diamidines structurally related to synthetic estrogens is being thoroughly investigated by Curtis and his co-workers. 2 The present paper presents in vitro and in vivo laboratory studies of testosterone compounds as fungistatic agents and a report of the clinical trial of methyltestosterone and meth-dia-mer-sulfonamides (triple sulfonamides) in three cases of the deep mycoses. LABORATORY ASSAY OF ANTIMYCOTIC EFFECT OF TESTOSTERONE COMPOUNDS AND SULFONAMIDES In Vitro Assay of Testosterone, Methyltestosterone, and Testosterone Propionate .—In 1949 Reiss 3 demonstrated that testosterone, methyltestosterone, and other steroids and steroid-like compounds possess moderate fungistatic activity in vitro against Histoplasma capsulatum and other pathogenic fungi. Using a pour

A new selective medium for Haemophilus pertussis, containing a diamidine, sodium fluoride and penicillin.

1. A highly selective medium forH. pertussishas been made by incorporating 4:4'-diamidinodiphenylamine dihydrochloride (M & B 938), penicillin and sodium fluoride in a partially defined base.2. M & B 938 has an interesting selectivity which is much influenced by its environment, especially by the ionic composition of the medium. No pattern has emerged from a study of the effects of eighty salts and many other substances on its activity; but the gain in selectivity forH. pertussisproduced by cysteine and magnesium malonate has been usefully exploited.3. In a strict comparison with Bordet-Gengou the new medium yielded 28% more positive cultures from per-nasal swabs and 75% more from post-nasal swabs. With the new medium the incidence of serious rival growth from any part of the nasopharynx has been about 4% and, in contrast with Bordet-Gengou, postnasal swabs have yielded as many positive cultures as per-nasal swabs.4. The new medium is simple to make, stable, reliable, economical of material and time and suitable for routine use with per-nasal, post-nasal or supralaryngeal swabs, sputum or vomit.5. The new medium growsH. parapertussispoorly; but a highly selective medium forH. parapertussisis easily made by adding streptomycin and penicillin to Bordet-Gengou.6. Whatever the medium, the nature and number of swabs are important factors affecting the isolation ofH. pertussis:with the new medium positive diagnoses from per- and post-nasal swabs taken at the same time were about 40% more than from either swab alone.7. A statistical analysis strongly supports the opinion of Bradford & Slavin (1940) that the relatively poor results with post-nasal swabs on Bordet-Gengou medium are due to the overgrowth ofH. pertussisby rivals rather than to the absence ofH. pertussisfrom the pharynx. It also indicates that the expectation of isolatingH. pertussison Bordet–Gengou varies directly with the numbers present and inversely with the numbers of rivals, especially of diphtheroids and streptococci.8. Addition of 32μg. stilbamidine di-isethionate and 0·25 unit penicillin/ml. to Bordet–Gengou makes an effective selective medium for oral streptococci related to, or of, groups K and L, but not forH. pertussis.I wish to thank my clinical colleagues, especially Dr N. O. Richards and Dr A. Bogdan, for kindly sending swabs from their patients, Professor R. J. V. Pulvertaft for his continuous support, Dr J. A. Fraser Roberts for advice and help with the statistics, Dr A. J. Ewins and Dr H. J. Barber, May and Baker Ltd., for gifts of aromatic diamidines, Dr H. King for gifts of alkyl diamidines and Mr M. Breach, F.I.M.L.T., and Mr A. Drake, F.I.M.L.T., for their help with the preparation of sera, media and routine cultures. I am indebted also to the Central Research Fund of London University and to the Governors of Westminster Hospital for grants for the purchase of materials.

Treatment of systemic blastomycosis with 2-hydroxystilbamidine.

PRIOR to the introduction of stilbamidine in the treatment of systemic blastomycotic infections, by Schoenbach and co-workers,* there had been no generally effective therapy for this disease, although administration of iodides in conjunction with desensitization has been reported to have been successfully used in the treatment of some cases.3Elson's observation that certain diamidines exerted a fungistatic effect upon Blastomyces dermatitidis4led to the clinical trial of stilbamidine in patients who had systemic blastomycotic infections. The therapeutic results obtained in the treatment of this disease were excellent.† Subsequently, Snapper and co-workers treated patients with extensive blastomycotic infections with 2-hydroxystilbamidine with good results.‡ Stilbamidine is one of the aromatic diamidines which has been used effectively in the treatment of kala-azar. However, because of undesirable sequelae developing in patients treated with this drug, it has been replaced to a great extent by propamidine, a less effective agent in the

Sporotrichosis successfully treated with stilbamidine.

SPOROTRICHOSIS has been treated successfully with the iodides for many years. Nevertheless, there are occasional patients with this disease who are resistant to iodide therapy. The report of Elson1in 1945 showing that Propamidine, one of the aromatic diamidines, exhibits in vitro antifungal activity opened up an entirely new group of chemotherapeutic agents in the treatment of the deep mycotic infections. One of the members of this diamidine group, stilbamidine (4,4′-stilbenedicarboxyamidine), is now being used extensively in the treatment of North American blastomycosis* and has also been successfully used in actinomycosis.5Since sporotrichosis has apparently never been treated with stilbamidine, it was decided to determine the in vivo effect of this fungicidal agent. REPORT OF A CASE W. E., an 18-year-old white male farmer, was referred to the dermatology department of the University Hospital on June 26, 1953, because of an ulcer of the distal end of the

Antifungal activity of some aromatic diamidines.

Abstract Selective in vitro antifungal activity of four aromatic diamidines against B. dermatitidis , B. brasiliensis , C. neoformans , H. capsulatum and C. albicans has been demonstrated. The addition of serum to the medium did not interfere with the activity of the compounds but, rather, enhanced the activity of phenamidine against C. neoformans.