Fazlul Huq

Studies on the interaction between Cd2+ ions and DNA

Cadmium is a potent carcinogen in rodents and has recently been accepted by the International Agency for Research on Cancer as a category 1 (human) carcinogen, but the molecular mechanism of its action remains largely unclear. It has however been suggested that cadmium-induced carcinogenesis may involve either direct or indirect interaction of Cd(2+) with DNA. Cd(2+) is believed to bind covalently with N7 centres of adenine and guanine. At low concentrations (< or =50 mM), Cd(2+) is found to react with plasmid DNA to produce a mixture of Form I and Form II bands whereas at higher concentrations (> or =100 mM), Cd(2+) causes extensive damage to DNA at a pH 5.8 solution of cadmium nitrate. Within the range 0-100 mM (when pH is adjusted to 7.4 by adding NaOH) an increase in concentration of Cd(2+) is found to cause a decrease in the gel mobility rate of plasmid and an increase in the intensity of the Form II band. When plasmid DNA is digested with BamH1, only the Form III band is observed both in the presence and absence of Cd(2+). However, the mobility of the band is found to decrease with the increase in the concentration of Cd(2+). When the enzyme Ssp1 which cuts plasmid DNA at the AT sites is used instead of BamH1, two bands are observed in the presence of cadmium as against one band in the absence of cadmium. These results suggest that Cd(2+) binds covalently with DNA (possibly at G, A and T centres) and can form intrastrand bifunctional AT adducts but not the GG adducts. It may also be that neither GG nor AT adducts are formed and yet Ssp1 digestion is prevented because of a structural modification introduced in adenine by its interaction with Cd(2+). In the presence of antioxidants such as cysteine, glutathione and ascorbate (especially cysteine and ascorbate), DNA damage is found to be greater than expected for the combined effects of the antioxidant and Cd(2+). The increased DNA damage is believed to be due to the formation of reactive oxygen species (ROS).

Studies on the interaction between Cd2+ ions and nucleobases and nucleotides

Cadmium is a potent carcinogen in rodents and has recently been accepted by the International Agency for Research on Cancer as a category 1 (human) carcinogen, but the molecular mechanism of its action remains largely unclear. It has however been suggested that cadmium-induced carcinogenesis may involve either direct or indirect interaction of Cd(2+) with DNA. In this study it is found that when Cd(2+) is allowed to interact with adenine and guanine, there is a marked change in the high performance liquid chromatography (HPLC) retention time for adenine but not for guanine. Since Cd(2+) is believed to bind covalently to adenine and guanine, the changes in retention time but absence of any cadmium in the peak fraction point to the following: (i) lability of cadmium-nucleobase adducts, and (ii) introduction of some kind of chemical modification in adenine but not in guanine as a result of covalent binding. This result is different from that for Ni(2+) in which case a change in retention time was observed for guanine but not for adenine.

Studies on the interaction between Ag(+) and DNA.

The interaction between silver ion and DNA has been followed by submarine gel electrophoresis. When pBR322 plasmid DNA was allowed to interact with silver(I) acetate, it was found to contain Form I and Form II bands whose intensity remained unchanged as the concentration of Ag(+) was increased from 0 to 50 mM. However, the mobility of the bands decreased as the concentration of Ag(+) was increased, indicating the occurrence of increased covalent binding of the metal ion with DNA. When 1:1 mixtures of silver(I) acetate and ascorbate were allowed to interact with plasmid and genomic DNAs, it was found that the mixtures were much more damaging to plasmid as well as genomic DNAs than silver(I) acetate or ascorbate alone. In the case of pBR322 plasmid DNA, the mixture at 12.5 mM concentration was found to be more damaging than the mixtures at both higher and lower concentrations. The increased DNA damage is believed to be due to free radicals produced from the oxidation of ascorbate by molecular oxygen where the metal ion was playing a catalytic role.

Synthesis, characterisation, activities, cell uptake and DNA binding of a trinuclear complex: [{trans-PtCl(NH3)}2μ-{trans-Pd(NH3)(2-hydroxypyridine)-(H2N(CH2)6NH2)2]Cl4

The trinuclear complex: [[trans-PtCl(NH3)](2)mu-[trans-Pd(NH(3))(2-hydroxypyridine)-(H(2)N(CH(2))(6)NH(2))(2)]Cl(4) (code named CH25) has been synthesized and characterized. The activity of the compound against human ovarian cancer cell lines: A2780, A2780 cisR and A2780 ZD0473R, cell up take, level of binding with DNA and nature of its interaction with pBR322 plasmid DNA have been determined. The compound is found to exhibit significant anticancer activity against the cell lines-about 45 times as active as cisplatin against A2780 cell line, about 76 times as active as cisplatin against A2780(cisR) cell line and about seven times as active as cisplatin against A2780cell line. The higher activity of CH25 suggests that the compound is able to overcome multiple mechanisms of resistance operating in A2780(cisR) and A2780(ZD0473R) cell lines. The compound is believed to form a range of interstrand GG adducts with duplex DNA that induces global changes in the DNA conformation, unlike cisplatin and ZD0473 [also known as AMD473 and JM473: cis-(2-methylpyridine)(ammine)dichloroplatinum(II)] that form mainly intrastrand adducts that induces a local kink in a DNA strand. The increasing prevention of BamH1 digestion of form I and form II pBR322 plasmid DNA with the increase in concentration of the compound is believed to be due to interstrand binding that brings about global changes in DNA conformation.

IN VITRO ASSESSMENT OF INDIVIDUAL AND INTERACTIVE EFFECTS OF AROMATIC HYDROCARBONS ON EMBRYONIC DEVELOPMENT OF THE RAT

There have been reports of disruption of embryonic development following exposure of pregnant women to aromatic hydrocarbons. In the present study, the embryotoxicity of toluene, xylene, benzene, styrene, and its metabolite, styrene oxide, was evaluated using the in vitro culture of postimplantation rat embryos. Possible interactions between toluene, xylene, and benzene were also studied using mixtures of these solvents. The results of the study showed that toluene, xylene, benzene, and styrene all have a concentration-dependent embryotoxic effect on the developing rat embryo in vitro. Styrene was embryotoxic at a lower concentration (1.00 mumol/mL) than benzene (1.56 mumol/mL), toluene (2.25 mumol/mL), or xylene (1.89 mumol/mL). The metabolite of styrene, styrene oxide, was embryotoxic at a concentration (0.038 mumol/mL). more than 20 times less than the parent compound. There was no evidence of a synergistic interaction between toluene, xylene, and benzene in causing embryotoxicity; the solvents interacted in an additive manner. The embryos were exposed to the solvents for 40 h of the organogenic period. When the levels of solvents found to be embryotoxic in the present study are compared to blood levels in the human following industrial exposure or solvent abuse, it appears unlikely that the threshold blood levels for embryotoxicity would be exceeded in the workplace. However, the possibility that exposure to solvents earlier or later or throughout the entire organogenic period might result in a different conclusion cannot be excluded.

Studies on the synthesis, characterization, binding with DNA and activities of two cis-planaramineplatinum(II) complexes of the form:cis-PtL(NH3)Cl2 where L = 3-hydroxypyridine and 2,3-diaminopyridine

Background Cis-planaramineplatinum(II) complexes like their trans isomers are often found to be active against cancer cell lines. The present study deals with the synthesis, characterization and determination of activity of new cis-planaramineplatinum(II) complexes. Results Two cis-planaramineplatinum(II) complexes: cis-(3-hydroxypyridine)(ammine)dichloroplatinum(II) (code named AH3) and cis-(2,3-diaminopyridine)(ammine)dichloroplatinum(II) (code named AH7) have been prepared and characterised based on elemental analyses, IR, Raman, mass and 1H NMR spectral measurements. The interactions of the compounds with pBR322 plasmid DNA have been investigated and their activity against ovarian cancer cell lines: A2780, A2780cisR and A2780ZD047Rhave been determined. Like cisplatin, AH3 and AH7 are believed to form mainly monofunctional N7(G) and bifunctional intrastrand N7(G)N7(G) adducts with DNA, causing a local distortion of a DNA strand. As a result, gel mobility of the DNA changes. Both AH3 and AH7 are found to be less active than cisplatin against the three cell lines with AH3 being the more active compound of the two. The higher activity of AH3 is in line with its lower molar conductivity value corresponding to a lower degree of dissociation. Conclusion The differences in activity of AH3, AH7 and cisplatin against the cell lines illustrate structure-activity relationship.

Synthesis and Activity of a Trinuclear Platinum Complex: [{trans‐PtCl(NH3)2}2μ‐{trans‐Pt(3‐hydroxypyridine)2(H2N(CH2)6NH2)2}]Cl4 in Ovarian Cancer Cell Lines

This paper describes the synthesis, characterisation, and cytotoxicity of a novel trinuclear platinum complex code named TH1. In addition to its activity against human ovarian cancer cell lines: A2780, A2780cisR, and A2780ZD0473R, cell uptake, DNA‐binding, and the nature of the compound interaction with pBR322 plasmid DNA have been determined. TH1 is found to be significantly more cytotoxic than cisplatin ‐ two times more active than cisplatin against the parent cell line A2780, thirteen times more active against the cisplatin‐resistant cell line A2780cisR and 11.5 times more active against the cell line A2780ZD0473R. Whereas the resistance factors for cisplatin as applied to the cell lines A2780 and A2780cisR, and A2780 and A2780ZD0473R are 12.9 and 3.0 respectively, the corresponding values for TH1 are 1.98 and 0.5. The results suggest that TH1 has been able to significantly overcome resistance in A2780cisR and A2780ZD0473R cell lines. Whereas cisplatin binds with DNA forming mainly intrastrand GG adduct that causes local bending of a DNA strand, TH1 should bind with DNA forming mainly interstrand GG adducts that would cause more of a global change in DNA conformation. Provided it has favourable toxicity profile, TH1 has the potential to be developed into a highly active anticancer drug with a wider spectrum of activity than cisplatin.

In Vitro Assessment of the Effect of Halogenated Hydrocarbons: Chloroform, Dichloromethane, and Dibromoethane on Embryonic Development of the Rat

Halogenated hydrocarbons are widely used in industry, the laboratory, and in the home. In the present study three of these solvents--chloroform, dichloromethane, and dibromoethane--were examined for embryotoxic/teratogenic potential using rat embryo culture. The results showed that each of the solvents had a concentration-dependent embryotoxic effect on the developing rat embryo in vitro. The effect and no-effect concentrations (expressed in mumol/ml culture medium), respectively, for each of the halogenated hydrocarbons tested were: dibromoethane--0.33, < 0.18; chloroform--2.06, 1.05; dichloromethane--6.54, 3.46. The levels of chloroform and dichloromethane found to be embryotoxic in the present study were compared to reported blood levels attained following controlled human exposure. In the industrial situation, if the current exposure levels are adhered to, chloroform and dichloromethane appear to have little potential for reproductive toxicity in the human. Fatal or near fatal solvent levels would be required in the mother for the embryotoxic level to be reached. For dibromoethane, there are no reports following controlled human exposure presumably due to its carcinogenicity. In an attempt to elucidate the mechanism of embryotoxicity, histological studies were performed after exposure of rat embryos to an embryotoxic level of each of the halogenated hydrocarbons studied, for increasing time periods up to the standard 40-hour culture. Marked cell death in the neuroepithelium of the developing neural tube was a prominent feature in all embryos exposed to an embryotoxic level of these solvents for periods of 16 hours of longer.

Synthesis and cytotoxicity of three trans-palladium complexes containing planaramine ligands in human ovarian tumor models.

The present study deals with the synthesis, characterization, and activity against human ovarian cancer cell lines A2780, A2780cisR, A2780ZD0473R, and SKOV‐3 of three trans‐planaraminepalladium(II) complexes of the form trans‐PdL2Cl2, coded as EH1, EH3, and EH4, for which L=2‐methylpyridine, imidazole, and 1,2‐α‐imidazopyridine, respectively. The cellular accumulation of palladium, palladium–DNA binding levels, and the nature of interactions of the compounds with salmon sperm and pBR322 plasmid DNA were also determined. All three compounds were found to be less active than cisplatin, but unlike cisplatin they were found to be equally or more active against the resistant cell lines A2780cisR and A2780ZD0473R than against the parent cell line A2780. Among the three palladium complexes, EH4 (which has the bulkiest carrier ligand) was found to be most active, in line with the highest cellular accumulation of palladium and highest level of palladium‐DNA binding resulting from the compound. EH4 was also found to cause the greatest conformational change to pBR322 plasmid DNA. The results of this study illustrate structure–activity relationships; in particular, they support the idea that the decreased reactivity of trans‐palladium complexes through the introduction of bulky ligands can make them more active against tumors.

Studies on the Nature and Strength of Pt . . .H(-N) Interactions. The Crystal Structures of Chloro[N-(2-aminoethyl)-N-(2-ammonioethyl)ethane-1,2-diamine]platinum(II) Chloride and Dichloro[4,7-diaza-1-azoniacyclononane]platinum(II) Tetrachlor

Two complexes, namely, chloro[N-(2-aminoethyl)-N-(2-ammonioethyl)ethane-1,2-diamine]platinum(II) chloride {[PtCl(tren+H)]Cl2} and dichloro[4,7-diaza-1-azoniacyclononane]platinum(II) tetrachloroplatinate(II)–water (1/2) {[PtCl2(tacn+H)]2[PtCl4]·2H2O}, have been prepared and structurally characterized by single-crystal X-ray diffractometry as part of a study of the nature and strength of Pt···H(–N) interactions. Crystals of [PtCl(tren+H)]Cl2 are monoclinic, space group P21/c, a 8.293(2), b 14.396(6), c 11.305(3) A, β 107.34(2)o, Z 4, and the structure has been refined to a residual of 0.042 based on 1631 reflections. Crystals of [PtCl2(tacn+H)]2[PtCl4]·2H2O are monoclinic, space group P21/a, a 12.834(4), b 8.206(4), c 13.116(8) A, β 93.01(4)˚, Z 2, and the structure has been refined to a residual of 0.035 based on 1974 reflections. In [PtCl(tren+H)]2+, the protonated amine forms hydrogen bonds with chloride anions and no close contacts with the metal ion. In [PtCl2(tacn+H)]+, a short intramolecular contact is observed between the metal and the protonated amine and the results of molecular mechanics modelling are consistent with there being a Pt···H hydrogen bond. Molecular mechanics modelling of [PtCl(tren+H)]2+ and [PtCl2(dien+H)]+ shows that the protonated amines could readily form close contacts with the metal. It is concluded that there is evidence for the formation of Pt···H(–N) hydrogen bonds but these bonds are very weak, being similar or lower in energy than Cl···H(–NPt) hydrogen bonds.