Synthesis and characterization of the biological activity of the cisplatin analogs, cis-PtCl(2)(dexrazoxane) and cis-PtCl(2)(levrazoxane), of the topoisomerase II inhibitors dexrazoxane (ICRF-187) and levrazoxane (ICRF-186)

The individual stereoisomers cis-PtCl(2)(dexrazoxane) and cis-PtCl(2)(levrazoxane) were synthesized and their structures were determined by X-ray crystallography. Dexrazoxane and levrazoxane inhibit cell growth because they are strong catalytic inhibitors of DNA topoisomerase II, whereas cisplatin acts through the formation of DNA cross-links. It was hypothesized that platinum(II) complexes of dexrazoxane and levrazoxane would retain both activities and yield drugs with a dual mode of action. Both cis-PtCl(2)(dexrazoxane) and cis-PtCl(2)(levrazoxane) inhibited Chinese hamster ovary cell growth, but more weakly than dexrazoxane and levrazoxane did. Based on their weak topoisomerase II inhibitory activity, it was concluded that these compounds did not inhibit cell growth by targeting topoisomerase II. A comparison of the conformation of cis-PtCl(2)(dexrazoxane) to that of dexrazoxane bound to the dimer interface of topoisomerase II showed that the highly constrained cis-PtCl(2)(dexrazoxane) was in a highly unfavorable conformation for binding. Neither of the platinum complexes were able to cross-link DNA. Thus the cell growth inhibitory activity of these complexes was also not likely due to any cisplatin-type cross-linking activity.     

The design of peptidyldiazomethane inhibitors to distinguish between the cysteine proteinases calpain II, cathepsin L and cathepsin B.

A series of peptidyldiazomethanes was synthesized and tested as inactivators of the cysteine proteinases calpain II, cathepsin L and cathepsin B. Inactivators that react rapidly and that show a degree of selectivity between the enzymes were identified. Z-Tyr(I)-Ala-CHN2 (where Z represents benzyloxycarbonyl) reacts rapidly with cathepsin L and more slowly with cathepsin B, but does not inhibit calpain II. Z-Leu-Leu-Tyr-CHN2 reacts rapidly with cathepsin L and calpain II but very slowly with cathepsin B. Boc-Val-Lys(epsilon-Z)Leu-Tyr-CHN2 (where Boc represents t-butyloxycarbonyl) reacts more rapidly with calpain II than with cathepsin L or cathepsin B. The discriminating inhibitory effects of these compounds make them potentially useful for investigation of enzyme functions in vivo. The data presented also provide insights into the subsite specificity of calpain.     

Dicarboxylate platinum(II) complexes as inhibitors of plasma membrane H(+)-ATPase in the yeast Saccharomyces cerevisiae

A series of cytotoxic neutral dicarboxylatoplatinum(II) complexes containing D(+), L(-) or DL-malate dianion and ethylenediamine or 1-ethylimidazole as ligands were examined using ATPase activity assays and the proton extrusion test. ATPase activity assays in vitro on plasma membrane H+-ATPase and on mitochondrial ATPase were carried out. The concentrations of compounds inhibiting enzyme activity to 50 per cent (J50) was determined. The new platinum complexes showed a stronger level of inhibition of both ATPases than the reference carboplatin; this inhibitory activity is related to a stereoisomeric form of anionic platinum ligands. ATPase inhibition in vivo was tested by glucose-stimulated proton extrusion and the influence of platinum compounds on this process in yeast cells was determined. Significant differences in activity levels were observed between those complexes with 1-ethylimidazole and those with ethylenediamine.     

Terpyridine-platinum(II) complexes are effective inhibitors of mammalian topoisomerases and human thioredoxin reductase 1

Terpyridine-platinum(II) (TP-Pt(II)) complexes are known to possess DNA-intercalating activity and have been regarded as potential antitumor agents. However, their cytotoxic mechanism remains unclear. To investigate the possible mechanism, a series of TP-Pt(II) compounds were prepared and their biological activities assessed. The DNA binding activities of the aromatic thiolato[TP-Pt(II)] complexes were stronger than the aliphatic 2-hydroxylethanethiolato(2,2′:6′,2′′-terpyridine)platinum(II) [TP(HET)]. TP-Pt(II) complexes inhibited topoisomerase IIα or topoisomerase I activity at IC₅₀ values of about 5 μM and 10-20 μM, respectively, whereas the human thioredoxin reductase 1 (hTrxR1) activity was inhibited with IC₅₀ values in the range of 58-78 nM. At the cellular level, they possessed cytotoxicity with IC₅₀ values between 7 and 19 μM against HeLa cells. Additionally, using X-ray crystallography and matrix-assisted laser desorption/ionization (MALDI) mass spectrometry, we elucidated that the TP-Pt(II) complexes inhibited hTrxR1 activity by blocking its C-terminal active-site selenocysteine. Therefore, TP-Pt(II) complexes possess inhibitory activities against multiple biological targets, and they may be further studied as anticancer agents.     

Combinatorial discovery of new asymmetric cis platinum anticancer complexes is made possible with solid-phase synthetic methods

To efficiently access asymmetric cis platinum (II) complexes for biological evaluation, a new solid-phase synthesis was designed. This synthesis was used for the preparation of a small library of platinum compounds. Several compounds from this library revealed promising activity during a cytotoxicity screen. Two active compounds were, therefore, synthesised on a larger scale and tested more extensively against a larger panel of cell-lines, confirming their high potential as antitumour compounds. The work presented illustrates how a combination of a new methodology and established techniques can speed up the search for platinum complexes with improved cytotoxic profiles compared to cisplatin.     

Inhibition of mitogen-activated protein kinase (MAPK) and cyclin-dependent kinase 2 (Cdk2) by platinum(II) phenanthroline complexes

Inhibition of protein kinases in the fight against disease remains a constant challenge for medicinal chemists, who have screened multitudes of predominantly planar organic scaffolds, natural and synthetic, to identify potent—albeit not always selective—kinase inhibitors. Herein, in an effort to investigate the potential biological utility of metal-based compounds as inhibitors against the cancer-relevant targets mitogen-activated protein kinase and cyclin-dependent kinase 2, we explore various parameters in planar platinum(II) complexes with substituted phenanthroline ligands and aliphatic diamine chelate co-ligands, to identify combinations that yield promising inhibitory activity. The individual ligands’ steric requirements as well as their pattern of hydrogen bond donors/acceptors appear to alter inhibitory potency when modulated.     

Platinum complexes of diaminocarboxylic acids and their ethyl ester derivatives: the effect of the chelate ring size on antitumor activity and interactions with GMP and DNA

A number of new Pt(II) complexes is described having the general formula PtCl(2)(LL), where LL is a chelating diamine ligand. Ligands LL were chosen as D,L-2,3-diaminopropionic acid and its ethyl ester, and D,L-2,4-diaminobutyric acid and its ethyl ester. The compounds were characterized using analytical and spectroscopic methods. The influence of the size of the chelate ring and its functionalization on the biological properties was studied. It was demonstrated by circular dichroism (CD) that the effects on the secondary structure of DNA induced by the four complexes are different. The interaction takes place at the N7 position of the purine bases, as shown by NMR studies. The platinum complexes of 2,3-diaminopropionic acid and 2,4-diaminobutyric acid are able to form intrastrand adducts with DNA and to distort the double helix by changing the base stacking. The ethyl ester derivatives uncoil the DNA from the B form to the C form. The interactions with 5'-GMP and DNA were compared with their antitumor activity. The platinum complexes of diaminocarboxylic acids exhibit cytotoxic activity in the A431, HeLa, and HL-60 cell lines in a dose- and time-dependent manner.     

In vivo delayed rejection of tumors and inhibition of delayed - type hypersensitivity by HT-29 human colonic adenocarcinoma cell line

Summary Cis-diamminedichloroplatinum (II) (cis-DDP) and its structural analogue cis-diamminediaquoplatinum(II) nitrate (cis-aq) were complexed via an intermediate dextran carrier to antibodies specifically reactive with B lymphoma cells (38C-13). The potential use of these drugs in site-directed immunotargeting was evaluated. The two platinum(II) compounds were previously shown to form pharmacologically active complexes with carboxymethyl dextran (CM-dex). For the purpose of preparing drug-antibody complexes, CM-dex was first conjugated to idiotypic antibodies that recognize a specific membrane IgM on the B lymphoma cells. The conjugates were prepared by a modified water-soluble carbodiimide method in which N-hydroxysuccinimide was used to enhance the coupling reaction. The conjugation was followed by separation of the CM-dex-IgG conjugates from unconjugated CM-dex or IgG. The platinum(II) compounds were then complexed to the CM-dex-IgG resulting in complexes carrying up to 50 mole drug/mole IgG. Both cis-DDP and cis-aq complexes of CM-dex-antibody conjugates maintained most of the original cell-binding activity of the antibodies. An in vitro assay was used to demonstrate selective binding to tumor cells in which the target cells were treated with specific immune complexes and washed before culture. In this assay the specific complexes showed preferential cytotoxicity for the B lymphoma cells in comparison to the free drugs, drug CM-dex, or nonspecific immune complexes.     

Anti-proliferative and metastatic protease inhibitory activities of protoberberines: An in silico and in vitro approaches

In metastasis, tumour cells interact with numerous factors and one of them is extracellular matrix (ECM). Earlier studies have emphasised the on role of matrix metalloproteases (MMPs) and cathepsin B in ECM degradation and cancer metastasis. In silico docking approaches are a boon in exploring the physicochemical characteristics like angiogenesis, growth and repair of cancerous cells. The binding affinities of berberrubine, jatrorrhizine and thalifendine with caspase 3, MMP-9, cathepsin B and telomeric DNA were performed using hex 6v and iGEMDOCK v2.1 software tools. The present investigation on berberrubine, jatrorrhizine and thalifendine revealed the formation of potentially stable complexes with caspase 3, cathepsin B and telomeric DNA similar to doxorubicin, a well-known anticancer drug. Further, in vitro approaches were made to study anti-proliferative activity against colon, lymphoma and ovarian cancer cell lines and enzyme inhibition activity against MMP-9 and cathepsin B. The results obtained reveal that these protoberberines alkaloids have potential inhibitory effect on cancer cell proliferation as well as metastatic proteases.     

On the stability and biological behavior of cyclometallated Pt(IV) complexes with halido and aryl ligands in the axial positions

A series of cyclometallated platinum(IV) compounds (3a, 3a′ and 3b′) with a meridional [C,N,N′] terdentate ligand, featuring an halido and an aryl group in the axial positions has been evaluated for electrochemical reduction and preliminary biological behavior against a panel of human adenocarcinoma (A-549 lung, HCT-116 colon, and MCF-7 breast) cell lines and the normal bronquial epithelial BEAS-2B cells. Cathodic reduction potentials (shifting from −1.463 to −1.570 V) reveal that the platinum(IV) compounds under study would be highly reluctant to be reduced in a biological environment. Actually ascorbic acid was not able to reduce complex 3a′, the most prone to be reduced according its reduction potential, over a period of one week. These results suggest an intrinsic activity for the investigated platinum(IV) complexes (3a, 3a′ and 3b′), which exhibit a remarkable cytotoxicity effectiveness (with IC₅₀ values in the low micromolar range), even greater than that of cisplatin. The IC₅₀ for A-549 lung cells and clog P values were found to follow the same trend: 3b′ > 3a′ > 3a. However, no correlation was observed between reduction potential and in vitro activity. As a representative example, cyclometallated platinum(IV) compound 3a′, exercise its antiproliferative activity directly over non-microcytic A-549 lung cancer cells through a mixture of cell cycle arrest (13% arrest at G1 phase and 46% arrest at G2 phase) and apoptosis induction (increase of early apoptosis by 30 times with regard to control). To gain further insights into the mode of action of the investigated platinum(IV) complexes, drug uptake, cathepsin B inhibition and ROS generation were also evaluated. Interestingly an increased ROS generation could be related with the antiproliferative activity of the cyclometallated platinum(IV) series under study in the cisplatin-resistant A-549 lung and HCT-116 cancer cell lines.     

Antitumor steroidal-cis-platinum(II)-o-catecholato conjugates: preliminary evaluation on breast cancer MCF-7 cells

Six new steroidal-cis-platinum(II)-o-catecholato complexes (5–9 and 15) were prepared by treatment of either [4-(2-aminoethyl)1,2-benzenediolato(2-)-O, O′]-bis(triphenylphosphine)platinum(II) or [3,4-dihydroxybenzenepropionic acid (2-)-O³, O⁴]-bis-(triphenylphosphine)platinum(II) with appropriate functionalized steroids. The biological effect of the air-stable conjugates on a human breast tumor cell line, MCF-7, was compared with that of cis-dichloro-diaminoplatinum(II) (cis-DDP). The activity of the new compounds proved to be of the same order of magnitude as cis-DDP.     

Effect of biologically active compounds of divalent platinum on the properties of the liquid crystal "microphase" of DNA

The optical properties of the "microphases" modeling the state of the DNA molecule in the cell and formed of both the low molecular DNA and the DNA complexes with cis- and trans-isomers of dichlorodiamine platininum (II) were studied. It was shown that the intensive band characteristic of the circular dichroism spectrum of the initial DNA "microphase" was decreasing with binding of DNA to cis-Pt (II) or trans-Pt (II). The effect of cis-Pt (II) on the "microphase" optical properties was more significant than that of trans-Pt (II). The effect correlated with the biological activity of the cis- and trans-compounds of platinum. Possible causes of the decrease in the optical activity of the DNA "microphase" are discussed.     

Inhibition of cathepsin L-induced degradation of epidermal growth factor receptors by c-Ha-ras gene products

The inhibitory activities of c-Ha-ras gene products (p21s) toward several cysteine proteinases have been investigated. The activity of cathepsin L was inhibited by p21s most effectively while those of cathepsin B and papain were slightly inhibited by p21s. p21s did not show any inhibitory activity toward cathepsin H. In order to connect the protease-inhibitor activity of p21s with cell growth, the degradation of epidermal growth factor receptors (EGF-receptors) was investigated. EGF-receptors were preferentially cleaved by cathepsin L but not by cathepsin B or H. The cleavage of EGF-receptors by cathepsin L was inhibited by p21s dose-dependently. These results raise the possibility that p21s can suppress the degradation of growth-related proteins such as EGF-receptors and thereby affect cell growth.     

Aminomalonato(1,2-diaminocyclohexane)platinum(II): A competitive antitumor compound within a new class of neutral,chemically stable,water soluble,functionalized platinum(II) complexes

Antitumor, neutral, chemically stable, water soluble and functionalized aminomalonato-platinum(II) complexes have been prepared and their mode of coordination characterized by elemental analysis and infrared spectra. Among this new class of compounds, aminomalonato(1,2-diaminocyclohexane)platinum(II) has been selected for ¹³C NMR measurements and for initial evaluation against L1210 and B 16 melanoma. The preliminary biological results reveal the high anti- neoplastic potential of this compound.     

Platinum complexes with anticancer potential and their evaluation by a colorimetric lambda prophage induction assay

A simple biochemical phage induction assay (BIA) showed significant activity with 90% of the antitumor platinum compounds tested and lack of activity for all Pd(II) compounds and Pt(II) cationic complexes, compounds that are expected to be inactive. Structure-activity relationships for a large number of chemicals can be studied simultaneously by this simple, rapid, inexpensive and quantitative biochemical assay. Fifty-three platinum complexes were tested, including a number of ethylenediamines synthesized for this work. The magnitude of inducing activity varied over a 25-fold range; differences among analogs reflected structural differences in a chemically consistent manner. Seven platinum complexes showed greater activity than that of cis-diamminedichloroplatinum(II) (cisplatin, cis-DDP), while other compounds appeared to be substantially less toxic. The assay was predictive for most compounds with very high or very low activity in vivo against L1210. For compounds with intermediate levels of activity, no correlation between inducing and antitumor activity was observed.     

Inhibitory effect of di- and tripeptidyl aldehydes on calpains and cathepsins

Eight different di- and tripeptidyl aldehyde derivatives, each having at its C-terminus an aldehyde analog of L-norleucine, L-methionine, or L-phenylalanine with a preceding L-leucine residue, were synthesized and tested for their inhibitory effects on several serine and cysteine endopeptidases. These compounds showed almost no inhibition of trypsin, and only weak inhibition of alpha-chymotrypsin and cathepsin H, while they exhibited marked inhibition of cathepsin B less than calpain II congruent to calpain I less than cathepsin L, being stronger in this order. The mode of inhibition of these cysteine proteinases was competitive for the peptide substrate used and inhibitor constants (Ki) were calculated from the Dixon plot. The best inhibitors found were: 4-phenyl-butyryl-Leu-Met-H for calpain I (Ki, 36 nM) and calpain II (Ki, 50 nM); acetyl-Leu-Leu-nLeu-H for cathepsin L (Ki, 0.5 nM); acetyl-Leu-Leu-Met-H for cathepsin B (Ki, 100 nM).     

Biological activity of palladium(II) and platinum(II) complexes of the acetone Schiff bases of S-methyl- and S-benzyldithiocarbazate and the X-ray crystal structure of the [Pd(asme)2] (asme=anionic form of the acetone Schiff base of S-methyldithiocarbazate) complex

Palladium(II) and platinum(II) complexes of general empirical formula, [M(NS)(2)] (NS=uninegatively charged acetone Schiff bases of S-methyl- and S-benzyldithiocarbazate; M=Pt(II) and Pd(II)) have been prepared and characterized by a variety of physicochemical techniques. Based on conductance, IR and electronic spectral evidence, a square-planar structure is assigned to these complexes. The crystal and molecular structure of the [Pd(asme)(2)] complex (asme=anionic form of the acetone Schiff base of S-methyldithiocarbazate) has been determined by X-ray diffraction. The complex has a distorted cis-square planar structure with the ligands coordinated to the palladium(II) ions as uninegatively charged bidentate NS chelating agents via the azomethine nitrogen and the mercaptide sulfur atoms. The distortion from a regular square-planar geometry is attributed to the restricted bite angles of the ligands. Antimicrobial tests indicate that the Schiff bases exhibit strong activities against the pathogenic bacteria, Bacillus subtilis (mutant defective DNA repair), methicillin-resistant Staphylococcus aureus, B. subtilis (wild type) and Pseudomonas aeruginosa and the fungi, Candida albicans (CA), Candida lypotica (2075), Saccharomyces cerevisiae (20341) and Aspergillus ochraceous (398)-the activities exhibited by these compounds being greater than that of the standard antibacterial and antifungal drugs, streptomycin and nystatin, respectively. The palladium(II) and platinum(II) complexes are inactive against most of these organisms but, the microbe, Pseudomonas aeruginosa shows strong sensitivity to the platinum(II) complexes. Screening of the compounds for their cytotoxicities against T-lymphoblastic leukemia cancer cells has shown that the acetone Schiff base of S-methyldithiocarbazate (Hasme) exhibits a very weak activity, whereas the S-benzyl derivative (Hasbz) is inactive. However, the palladium(II) complexes exhibit strong cytotoxicities against this cancer; their activities being more than that of the standard anticancer drug, tamoxifen. The [Pt(asme)(2)] complex exhibits a very weak cytotoxicity, whereas [Pt(asbz)(2)] is inactive against leukemic cells.     

Differential anti-proliferative properties of novel hydroxydicarboxylatoplatinum(II) complexes with high or low reactivity with thiols

We have examined the anti-proliferative effect of 13 recently synthesised platinum dicarboxylate complexes, very similar in their chemical, structural and kinetic properties to carboplatin. We used the L5178Y model: two murine lymphoma sublines, which differ in nucleotide excision repair ability and hence, in sensitivity to those platinum complexes that react with DNA. The anti-proliferative effect of the examined compounds mainly depends on the kind of amine ligand. Complexes with the primary amine (ethylenediamine) are more effective than complexes containing the tertiary amine (1-alkylimidazole). The ethylenediaminemalatoplatinum(II) complexes show a differential in vitro anti-proliferative activity in the L5178Y model; hence, it may be expected that they inflict DNA lesions that are repaired by the nucleotide excision system. The cytotoxicity of these complexes is directly correlated with reactivity with glutathione (GSH). The 1-alkylimidazole complexes are of low toxicity and moderate to low reactivity with GSH; in contrast to the ethylenediaminemalatoplatinum(II) complexes, their cytotoxicity is inversely correlated with reactivity with GSH. Two of the 1-alkylimidazole complexes, bis(1-ethylimidazole)(L-malato)platinum(II) and bis(1-propylimidazole (L-malato)platinum(II), show a considerable ability to arrest cells in G2 phase. We expect that the properties of these two groups of platinum complexes may be exploited in combined platinum complex treatment and irradiation.     

Water-soluble platinum(II) complexes of diamine chelating ligands bearing amino-acid type substituents: the effect of the linked amino acid and the diamine chelate ring size on antitumor activity, and interactions with 5'-GMP and DNA

Six new Pt(II) complexes are described having the general formula PtCl(2)(LL), in which LL is a chelating diamine ligand bearing an amino acid as substituent. The amino acids chosen are l-alanine and its methyl ester, and l-phenylalanine. The compounds have been characterized using analytical and spectroscopic methods. The influence on the biological properties of the size of the chelate ring and the structure of the amino acid substituent has been studied. The effect of the presence of a carboxylic or carboxylate group on the amino acid C-terminus has also been determined. It is demonstrated by circular dichroism (CD) that the effect on the secondary structure of DNA induced by the six complexes differ from each other. In all cases, the interaction takes place at the N7 position of the purine bases, as shown by NMR monitoring. The general behavior of these platinum complexes, with one exception, is to uncoil the DNA from the B form to the C form. The interactions with 5'-GMP and DNA have been compared with their expected antitumour activity. The complexes with l-alanine and l-phenylalanine exhibit cytotoxic activity in HeLa and HL-60 cell lines, in a dose- and time-dependent manner. No cytotoxic activity of the methyl ester derivatives have been determined because of their low solubility in aqueous solution.     

Interactions of the anticancer antibiotic altromycin B with copper(II), palladium(II) and platinum(II) ions and in vitro activity of the formed complexes

Interaction of the anticancer antibiotic altromycin B with Cu(II), Pd(II) and Pt(II) ions was studied using 1H-NMR, EPR, electronic absorption and circular dichroism spectroscopy. The results derived from NMR studies where that the Pt(II) and Pd(II) ions interact with the nitrogen atom of the dimethylamino group of the C(10)-disaccharide, while the C(2)-epoxide group does not participate and remains intact. Cu(II) ions interact in a different way with altromycin B as was concluded by EPR and circular dichroism spectra. Altromycin B coordinates to the Cu(II) ions via the oxygen atoms of the C(11) phenolic and the C(12) carbonyl group while the nitrogen atom does not participate in the complexation. The presence of these metal ions improves the stability of altromycin B in solution. These complexes were studied in vitro against K562 leukemia sensitive and doxorubicin-resistant cells and GLC4 lung tumor cells, sensitive and doxorubicin-resistant. The activity of the complexes compared to the free drug is improved against resistant cells and is affected moderately against sensitive cells. Finally, 20% of platinum added as altromycin B metal complex entered GLC4 cells.