Branka Zorc

Antiradical, chelating and antioxidant activities of hydroxamic acids and hydroxyureas.

Reactive oxygen species, along with reactive nitrogen species, may play an important role in the pathogenesis and progress of many diseases, including cancer, diabetes and sickle cell disease. It has been postulated that hydroxyurea, one of the main treatments in sickle cell disease, achieves its activity partly also through its antioxidant properties. A series of hydroxyurea derivatives of L- and D-amino acid amides and cycloalkyl-N-aryl-hydroxamic acids was synthesized and investigated for their radical scavenging activity, chelating properties and antioxidant activity. All the compounds showed exceptional antiradical activities. For example, free radical scavenging activities of investigated hydroxyureas were higher than the activity of standard antioxidant, butylated hydroxyanisole (BHA). Moreover, most of the investigated hydroxamic acids were stronger Fe2+ ion chelators than quercetin. In addition, the investigated compounds, especially hydroxamic acids, were proven to be excellent antioxidants. They were as effective as BHA in inhibiting β-carotene-linoleic acid coupled oxidation. It is reasonable to assume that the antioxidant activity of the investigated compounds could contribute to their previously proven biological properties as cytostatic and antiviral agents.

Hydantoin derivatives of L- and D-amino acids: synthesis and evaluation of their antiviral and antitumoral activity.

3,5-Disubstituted hydantoin (1,3-imidazolidinedione) derivatives 5a-h were prepared by base induced cyclization of the corresponding N-(1-benzotriazolecarbonyl)-L- and D-amino acid amides 4a-h. Compounds 5a-h were evaluated for their cytostatic and antiviral activities. Among all the compounds evaluated, 3-benzhydryl-5-isopropyl hydantoin (5a) showed a weak but selective inhibitory effect against vaccinia virus (EC(50) = 16 microg/mL; selectivity index: 25). 3-Cyclohexyl-5-phenyl hydantoin (5g) showed inhibitory activity against cervical carcinoma (HeLa, IC(50) = 5.4 microM) and breast carcinoma (MCF-7, IC(50) = 2 microM), but also cytotoxic effects towards human normal fibroblasts (WI 38). On the contrary, the 3-benzhydryl-5-phenyl substituted hydantoin derivative 5h showed moderate inhibitory activity towards HeLa, MCF-7, pancreatic carcinoma (MiaPaCa-2), lung carcinoma (H 460) and colon carcinoma (SW 620) (IC(50) = 20-23 microM), but no effect on WI 38.

The novel phosphoramidate derivatives of NSAID 3-hydroxypropylamides: synthesis, cytostatic and antiviral activity evaluations.

The target phosphoramidates 5a-e were prepared in one step from 3-hydroxypropyl derivatives 3a-e of nonsteroidal anti-inflammatory drugs (fenoprofen, ketoprofen, ibuprofen, indomethacin, diclofenac). The products 3a-e and 5a-e were evaluated for their cytostatic and antiviral activity against malignant tumour cell lines and normal human fibroblasts (WI 38). All phosphoramidate derivatives 5a-e possess significantly greater inhibitory activities than the corresponding 3-hydroxypropyl derivatives 3a-e, whereby compound 5a showed the most potent inhibitory activities against cervical, pancreatic and colon carcinoma cell lines (IC(50)=5-7 microM).

Urea and carbamate derivatives of primaquine: synthesis, cytostatic and antioxidant activities.

The novel urea primaquine derivatives 3 were prepared by aminolysis of primaquine benzotriazolide 2 with several hydroxyamines and ethylendiamine, while carbamates 4 were synthesized from the same precursor 2 and alcohols. All compounds are fully chemically characterized and evaluated for their cytostatic and antioxidant activities. The most prominent antiproliferative activity was obtained by compounds 3c, 3d, 3g, and 5b (IC(50)=9-40 microM). 1-(5-Hydroxypentyl)-3-[4-(6-methoxy-quinolin-8-ylamino)-pentyl]urea (3c) showed extreme selectivity toward SW 620 colon cancer cells (IC(50)=0.2 microM) and a bit less toward lung cancer cells H 460. Hydroxyurea 3h showed the highest interaction with DPPH. Primaquine twin drug 3g showed very significant inhibition on LOX soybean (IC(50)=62 microM). Almost all the tested derivatives highly inhibited lipid peroxidation, significantly stronger than primaquine phosphate.

Macromolecular prodrugs. IX. Synthesis of polymer-fenoprofen conjugates.

Synthesis of several polymer-fenoprofen conjugates is described. Fenoprofen was first chemically modified into benzotriazolide 2 and amino acid amide derivatives: glycine fenoprofenamide (3a) and beta-alanine fenoprofenamide (3b) and their benzotriazolides 6a and 6b. Compounds 2 and 6 readily reacted with polyhydroxy aspartamide-type polymers, i.e. poly[alpha,beta-(N-2-hydroxyethyl-DL-aspartamide)] (PHEA) and poly[alpha,beta-(N-3-hydroxypropyl-DL-aspartamide)] (PHPA) forming conjugates 5, 8a,b and 9a,b, respectively. Conjugate 11 was obtained by partial aminolysis of poly-DL-(2,5-dioxo-1,3-pyrrolidinediyl) (PSI) with 2-aminoethyl fenoprofenamide (3c), followed by total aminolysis with 2-hydroxyethylamine. The synthesised polymer-drug conjugates differed in type of covalent bounding, type and/or length of spacer and drug-loading.

Gemfibrozil encapsulation and release from microspheres and macromolecular conjugates.

The purpose of this study was to evaluate and compare the ability of the macromolecular conjugates and microspheres to modify the release rate of gemfibrozil (Gem). Gem was covalently linked to two similar polymers: poly[alpha,beta-(N-2-hydroxyethyl-DL-aspartamide)] (PHEA) and poly[alpha,beta-(N-3-hydroxypropyl-DL-aspartamide)] (PHPA) by an ester linkage. The polymer-drug conjugates obtained (PHEA-G(1-3) and PHPA-G) differ in weight-average molecular weight, length of spacer and Gem content. Microspheres, composed of chitosans of different molecular weight alone or as a mixture with (2-hydroxypropyl)methylcellulose (HPMC), PHEA or PHPA and with different theoretical polymer/drug ratio (2:1 and 3:1, w/w) were prepared by spray drying. The microparticulate systems were morphologically characterised by scanning electron microscopy, particle size analysis and Gem content was determined. In vitro dissolution tests were performed to evaluate the feasibility of conjugates and microspheres in modulating Gem release. The results obtained show that microspheres are always suitable to modulate Gem release and that the best conditions are achieved by microspheres composed of the low molecular weight chitosan (CL) combined with PHPA or HPMC with either 2:1 or 3:1 (w/w) polymer/drug ratio. The PHEA-G conjugates exhibited rapid Gem release within less than 2 h, while the PHPA-G conjugate showed sustained Gem release profiles over a 10-h period.

Macromolecular prodrugs. II: Esters of L-dopa and α-methyldopa

l-Dopa and α-methyldopa were attached by an ester linkage to α,β-poly(N-hydroxyethyl)-dl-aspartamide (PHEA), a hydrophilic polymer, previously proposed as a drug carrier. Ester bonding was achieved by means of 1-benzotriazolycarbonyl (Btc) group as both an N-protecting and C-activating group in the starting amino acids. In the same way several simple esters of l-dopa and α-methyldopa were prepared. Release of active substances based on hydrolysis of PHEA adducts was studied in vitro, and the following (pseudo) first order release rate constants for l-dopa and α-methyldopa were obtained, 1.06 × 10−3 and 6.91 × 10−4 min−1, respectively. In addition, characterization of the PHEA-l-dopa adduct was carried out in vivo using an intracerebral microdialysis technique in order to evaluate the prolonged release eventually achieved.

The Novel Ketoprofen Amides - Synthesis and Biological Evaluation as Antioxidants, Lipoxygenase Inhibitors and Cytostatic Agents

The novel amides of ketoprofen and its reduced derivatives (5a–f, 4a–n, 6a–g) with aromatic and cycloalkyl amines or hydroxylamines were prepared and screened for their reducing and cytostatic activity as well as for their ability to inhibit soybean lipoxygenase and lipid peroxidation. 1,1‐Diphenyl‐picrylhydrazyl test for reducing ability revealed that ketoprofen amides were more potent antioxidants than the amides of the reduced ketoprofen derivatives. The most active compound was benzhydryl ketoprofen amide 5f. Lipoxygenase inhibition of the tested compounds varied from strong to very weak. The most potent compound was benzhydryl derivative 6f (IC50 = 20.5 μm). Aromatic and cycloalkyl amides 4 and 5 were more potent lipoxygenase inhibitors than derivatives with carboxylic group. Aromatic amides of series 4 and 5 showed excellent lipid peroxidation inhibition (92.2–99.9%). On the other hand, the most pronounced cytostatic activity was exerted by O‐benzyl derivative 4i, although in general all tested reduced and non‐reduced lipophilic derivatives showed similar activity.

Antioxidant activity of NSAID hydroxamic acids

Antioxidant activity of NSAID hydroxamic acids In the present study, seven hydroxamic acid derivatives of nonsteroidal anti-inflammatory drugs (NSAIDs) (ibuprofen, fenoprofen, ketoprofen, indomethacin and diclofenac) were found to possess significant antioxidant, radical scavenging and metal chelating activities. The most active antioxidant and radical scavenger was N-methylhydroxamic acid of diclofenac (ANT = 88.0% and EC50 = 60.1 μg mL-1). The activity of the standard substance, butylated hydroxyanisole, in the two assays was ANT = 86.9% and EC50 = 18.8 μg mL-1, respectively. Ibuproxam was the strongest iron chelator among investigated hydroxamic acids (EC50 = 255.6 μg mL-1), yet significantly weaker than the standard substance, EDTA (EC50 = 29.1 μg mL-1). It seems that different mechanism is involved in metal chelating activity than in antioxidant and radical scavenging activity. Antioxidant and radical scavenging activities may be connected with conjugation of the nitrogen lone electron pair with the carbonyl group. On the other hand, more hydrophilic substances tend to be better iron chelators. Antioksidativno djelovanje NSAID hidroksamskih kiselina U radu je opisano antioksidativno djelovanje te sposobnost hvatanja slobodnih radikala i stvaranja kelata sedam hidroksamskih kiselina, derivata nesteroidnih protuupalnih lijekova ibuprofena, fenoprofena, ketoprofena, indometacina i diklofenaka. Najjače antioksidativno djelovanje i najjaču sposobnost hvatanja slobodnih radikala imala je N-metilhidroksamska kiselina diklofenaka (ANT = 88,0% i EC50 = 60,1 μg mL-1). Vrijednosti za standardnu supstanciju, butilirani hidroksianisol, bile su: ANT = 86,9% i EC50 = 18,8 μg mL-1. Derivat ibuprofena bio je najjači kelator među ispitivanim hidroksamskim kiselinama (EC50 = 255,6 μg mL-1), ali značajno slabiji od standardne supstancije, EDTA (EC50 = 29,1 μg mL-1). Pretpostavlja se da su različiti mehanizmi uključeni u keliranje metala i antioksidativno djelovanje, odnosno hvatanje slobodnih radikala. Antioksidativno djelovanje i sposobnost hvatanja slobodnih radikala moglo bi biti povezano s konjugacijom slobodnog para elektrona na dušiku s karbonilnom skupinom. S druge strane, hidrofilnije supstancije pokazale su se kao jači kelatori iona željeza.

Fenoprofen and ketoprofen amides as potential antitumor agents.

Following numerous experimental observations that various non‐steroidal anti‐inflammatory drugs have antitumor potentials, a series of fenoprofenamides (1a–g) and ketoprofenamides (2a–c) was tested on proliferation of different human tumor cell lines and normal human fibroblasts in vitro. Fenoprofen and ketoprofen showed modest antiproliferative activity, whereas the growth inhibitory activity of the tested amides clearly demonstrates that the substituents linked by an amide bond are essential for the significantly stronger cytostatic activity, probably because of a greater lipophilicity and/or better cell uptake. Additionally, it was shown that the most active derivatives (1d and 2a) induced cell cycle arrest at the G1 phase, as well as apoptosis.