Synergistic enhancement of carcinogenicity by tobacco smoke constituents
There is considerable evidence indicating that the weakly acidic phenolic fraction of tobacco smoke condensate (TSC), elicits strong tumor-promoting activity in polynuclear aromatic hydrocarbons (PAHs)-initiated animals. Pure phenols by themselves are weak tumor-promoters at very high dose and can not account for the tumor-promoting activity of the whole phenolic fraction. The mechanism(s) underlying the tumor-promoting activity of the phenolic fraction is not known. We observed that TSC phenolic fraction at non-cytotoxic concentrations attenuates BPDE-induced (i) p53 accumulation and p21 expression in human lung small airway epithelial (SAE) cells and (ii) activation of ERKs and NF-κB in mouse epidermal JB6 (P+) cells. Our ongoing effort in this respect is to understand whether TSC phenolic fraction inhibits p53 function by abrogating (i) DNA binding (in vitro and in vivo) and transcriptional activities of p53, (ii) p21 response, cell-cycle arrest, expression of G1 cyclins, activation of cdks, and phosphorylation of Rb protein, (iii) p53 stability, p53 phosphorylation/acetylation at serine and lysine residues respectively, p53- Mdm2 interaction, and PI3-K/Akt-mediated phosphorylation of Mdm2 which regulates p53-Mdm2 interaction (iv) the activation of DNA damage-induced kinases and their ability to phosphorylate p53 and (v) p53 transcription and NF-kappaB activation. The data from these studies will help in assessing the health risk presented by tobacco smoke constituents.
Mechanism of carcinogenesis of polynuclear aromatic hydrocarbons and their aza- and thia-analogues
Polynuclear aromatic hydrocarbons and their heterocyclic analogs are ubiquitous environmental contaminants which are introduced in our environment through incomplete combustion of organic matters such as fossil fuels, tobacco, etc. It has been demonstrated that some of these compounds are potent carcinogens in laboratory animals, and may be responsible for causing majority of human cancers. There is now considerable evidence indicating that these environmental contaminants are metabolically activated to reactive metabolites which binds to DNA thereby inducing carcinogenesis. We are currently investigating various major biochemical pathways that may be involved in the bioactivation of these polynuclear aromatic compounds and their heterocyclic analogs. Our research is currently focused on:
- Synthesis of the potential metabolites of PAHs and their heterocyclic analogs.
- vivo and in vitro metabolism of PAHs and their heterocyclic analogs, and the characterization of the metabolites produced using modern instrumental techniques (HPLC, GC, GC-MS etc).
- In vitro and in vivo bioassay of carcinogens and their various metabolites. These studies involve (i) cell transformation activity of parent compounds and their metabolites using various bacterial strains, and animal or human cells, and (ii) tumorigenic activity in animal models (mouse skin etc).
- Characterization and evaluation of the potential role of DNA adducts produced by environmental carcinogens and their metabolites using various in vitro and in vivo models in order to understand the structural requirements and the function of these DNA adducts in transforming normal cell to cancerous cells.
- Characterization of specific enzymes, especially cytochrome P450s (in animals and humans) involved in the metabolic activation of chemical carcinogens. Cytochrome P450 is an important class of oxidative enzyme which play important role in the metabolic activation of environmental carcinogens. Cytochrome P450s exit in multiple forms and each form (isozyme) may differ significantly from others in its substrate specificity, regioselectivity, and stereoselctivity in the metabolism of carcinogens to their carcinogenic metabolites. Thus the characterization of specific P450s that are involved mainly in the metabolic activation of environmental carcinogens is of significant interest to cancer researchers.
- Potential role of non-carcinogenic metabolites on inhibitory and synergistic effect on the carcinogenicity of carcinogenic metabolites of carcinogens. This study is initiated to understand why certain environmental carcinogens (such as benzo[a]pyrene, dibenz[a,l]pyrene) exhibit high carcinogenic activity compared to their widely accepted ultimate carcinogens, bay-region diol epoxides.
The collective information produced from these and other mechanistic studies will help in assessing the health risk presented by various classes of environmental carcinogens, and in developing various strategies in the prevention of cancer and related diseases caused by these ubiquitous environmental carcinogens/toxicants.
Environmental metal pollutants, co-carcinogenesis, p53 signaling events and apoptosis
Cadmium is an environmental polutant and is one of the major metal constituents of tobacco smoke. There is considerable evidence indicating that cadmium elicits synergistic enhancement of cell transformation when combined with benzo[a]pyrene (BP) or other PAHs. Smokers are particularly at a high risk of exposure to mixture of PAHs and cadmium. The mechanism of the synergistic interaction of heavy metals particularly cadmium with PAHs is not established. We observed that cadmium at non-cytotoxic concentrations attenuates PAH-induced (i) p53 accumulation, (ii) NF-κB activation and (iii) DNA fragmentation (indicative of apoptosis) in different cell lines. We also observed that BPDE-induced NF-κB activation and DNA fragmentation are inhibited by inhibitors of p53 and NF-κB respectively. In order to examine whether the synergistic activity of cadmium is due to its interference with BP-induced signaling events which mediate p53 stability, p53 activation and p53 function toward apoptosis, we are investigating the effect of cadmium on (i) NF-κB-mediated p53 transcription, (ii) activation of signaling pathways which regulate NF-κB activation, (iii) the regulation of NF-κB subunits which determines its apoptotic or anti-apoptotic function and (iv) the expression of anti- and pro-apoptotic proteins. The data obtained from these studies will help in assessing the associated health risk presented by tobacco smoke constituents and will be useful for therapeutic strategies in the prevention of cancer.
DNA damage by poly-nuclear aromatic hydrocarbons present in tobacco smoke and the mechanism of tumor promotion by phorbol ester
Polynuclear aromatic hydrocarbon (PAH) are ubiquitous environmental pollutants. There is considerable evidence showing that the prototype PAH bnzo[a] pyrene (BP) induces p53 in human and mouse cells. DNA damage caused by benzo[a]pyrene (BP) or other PAHs induce p53 protein as a protective measure to eliminate the possibility of mutagenic fixation of the DNA damage. 12-O-tetradecanoylphorbol-13-acetate (TPA) inhibits p53 response induced by BP and other DNA-damaging agents and may cause tumor promotion. The molecular mechanism of attenuation of BP-induced p53 response by TPA is not known. We are investigating the interference of TPA with (i) the upstream regulation of p53 and (ii) the downstream signaling pathways which are involved in p53 function. In this regard the role of MAP kinase and NF-kappaB signaling pathways in p53 stabilization and function has been focused.Developing mechanism based chemopreventive agents
These studies include (i) developing mechanism-based suicide substartes or inhibitors of cytochrome P450s involved in the metabolic activation of P450s, (ii) developing various synthetic and naturally occurring compounds that potentially interfere with various signaling processes involved in cancer induction and its metastatis.
