Activation and detoxification of lung cancer carcinogens
Initial Award Abstract
Cigarette smoking is the greatest risk factor for lung cancer, and inhalation of tobacco smoke exposes the airways to at least 4000 chemicals, more than 60 of which are considered carcinogenic (cancer-causing). One of the classes of compounds contained in tobacco smoke, the polycyclic aromatic hydrocarbons (PAHs), has been shown to have cancer-causing effects in animals and is believed to be one of the major contributors to smoking-related lung cancer. After PAHs are inhaled, they are actively metabolized by specialized enzymes, the cytochromes P450 (CYPs), into more reactive (activated) forms. A model substrate of the class of PAHs is benzo(a)pyrene (BaP), which has been extensively studied and shown to undergo metabolism and activation to forms that are known to be mutagenic. This means that in its activated form, BaP metabolites bind irreversibly to DNA, creating mutations that can affect cell behavior and lead to a cancerous episode. While the process of bioactivation clearly underlies the initiation of a toxic response, a separate set of specialized enzymes, the UDP-glucuronosyltransferases (UGTs), are able to modify and inactivate toxic chemicals such as activated BaP, making them less toxic and more easily excreted from the body.
A tightly controlled relationship between activation and detoxification by the UGTs is essential for normal cellular function. It is now being understood that human populations carry selective inactivating allelic polymorphisms in some of the UGT genes, and these individuals are at heightened risk to the dangers of selective cancers of the GI and upper respiratory tracts. These findings indicate that substantial interindividual variability in UGT activities exist among the normal population and that UGT-deficient individuals are at risk from the reactive intermediate-mediated effects of environmental toxicants such as those present in tobacco smoke. We are proposing in this application to generate strains of mice that carry those human UGT genes with the intention to explore their impact in protecting the whole body against tobacco born mutagens.
In order to achieve the goals set out in this application, it is first necessary to prove that the human UGT genes can be genetically manipulated so that incorporation of the DNA into the germ line of mice can be achieved. We have recently accomplished this goal by first isolating a portion of mouse chromosome 1 that spans over 250 thousand bases of DNA. This fragment of DNA was incorporated into fertilized female mouse blastocyst and following introduction into female mice produced offspring that now express the human UGT genes. These new transgenic UGT (Tg-UGT1) mouse strains have been shown to regulate these genes in a pattern similar to what has been observed in human tissues, demonstrating that regulatory forces present in the whole animal can mimic the actions as observed in humans. Most important, we have now developed a living animal model to examine the contribution of human glucuronidation and its ability to protect the animal against selective environmental toxicants that are ubiquitous in tobacco products.
Having fully “humanized” mice with the human UGTs, it will now be possible to examine the contribution of these proteins toward the initiation of a carcinogenic episode. Having available animals that contain no functional UGTs and those that are now fully humanized with these genes, we can examine the specific physiological and biochemical events that underlie the initiation of lung cancer. While the biochemical and molecular events that underlie the etiology of a carcinogenic event are immensely complex, we will be able for the first time to evaluate through genetics the importance of the human UGTs toward the onset lung cancer. We anticipate the development of these unique animal models will provide us the very unique opportunity to evaluate what we believe is an important genetic component to tobacco induced diseases. |