Reports by the International Agency for Research on Cancer have classified secondhand tobacco smoke as an established human carcinogen. There are over 4000 chemical compounds in secondhand smoke; 200 of which are known to be poisonous, and upwards of 60 have been identified as carcinogens. Exposure to secondhand smoke or even just a past smoking history can also increase the likelihood of bladder cancer in offspring. One of the most commonly used antineoplastic agents for the treatment of advanced bladder cancer is cisplatin, but the development of resistance to cisplatin during treatment is common and constitutes a major obstacle to the cure of sensitive tumors. Although many studies have been conducted on the molecular mechanism of drug resistance, little is known about the treatment of these drug-resistant tumors, which still remains a significant problem. A number of reports have suggested that cancer patients who smoke while receiving treatment have poorer outcomes compared with their nonsmoking counterparts, possibly because of lower rates of response. Retrospective series of patients with renal, bladder, and especially glottic cancers also indicate a link between smoking during treatment and decreased efficacy of cancer therapies. However, there are no direct data showing that cigarette smoke could actually induce resistance to chemotherapeutic agents, such as cisplatin. Deletion of chromosome 9p frequently occurs in bladder tumors. Depending on the respective investigation, frequencies between 30 and 70% have been published. Studies by Blaveri et al., indicate that in primary bladder cancer, loss of clones across the entire chromosome 9 occurs with an average frequency of 47% for 9p and 46% for 9q. Chromosome 9 carries important genes involved in adenine metabolism, namely AK1, AK2 and AK3. All three AKs are nuclear-encoded proteins and synthesized in the cytoplasm. AK1 remains ASP1517 HIF inhibitor located mainly in the cytosol of different tissues. Mature AK2 and AK3 are imported into mitochondria, where AK2 resides in the intermembrane space whereas AK3 is located exclusively in the mitochondrial matrix. Cigarette smoke is also known to induce mitochondrial damage as well as dysfunction which may in turn increase cisplatin resistance in bladder cancer cells. In this study we examined the relationship between tobacco exposure and cisplatin resistance in relation to mitochondria function and specifically to a mitochondria-resident protein AK3. We showed here that cigarette smoke condensate vapor can induce cisplatin resistance. We also found that the expression of AK3 is affected by CSC vapor and that restoration of this gene sensitizes the cells to cisplatin. Secondhand smoke exposure or exposure to environmental smoke has been assessed as a risk of bladder cancer. Our approach here is based on recent studies on normal oral keratinocytes indicating that exposure of cells to vapor component of cigarette smoke extract is as effective as direct treatment of cigarette smoke extract. These studies also indicated that chronic exposure to cigarette smoke provide a better model in vivo than acute exposure to cigarette smoke. Studies with environmental cigarette smoke or secondhand smoke in mice have demonstrated a variety of early alterations, including cytogenetic damage in bone marrow and peripheral blood, formation of lipid peroxidation products in lung, increase of bulky DNA adducts and oxidatively generated DNA damage. Our observation in this model that AK3 protein expression is decreased by CSC vapor builds on an old observation that cigarette smoke poisons lung cilia through a direct effect on adenylate kinases. Also increase of AK3 mRNA level and AK3 enzyme activity were previously observed in rat skeletal muscle. In addition, AK3 protein was found to increase 10-fold during neural differentiation of P19 embryonal carcinoma cells. The induction of AK3 mRNA was also shown in response to hypoxia in HeLa cells depending on the presence of hypoxiainducible factor-1. Here we show a direct effect on cancer cells and provide novel evidence that decreased expression of AK3 in the presence of CSC vapor is accompanied with decreased sensitivity of bladder cells to cisplatin and restoration of AK3 sensitizes cells to cisplatin. By linking AK3, our data support the notion that mitochondria plays an important role in cigarette smoke induced cisplatin resistance. Studies indicate that components present in cigarette smoke extract are able to pass through the membranes of mitochondria. Further it has been proven that highly reactive components like polycyclic aromatic hydrocarbons, aldehydes, phenols, heavy metals, and amines are lipophilic candidates that easily enter the cell and disturb mitochondria. AK3 is present in the mitochondrial matrix and probably functions in transferring the high-energy phosphate to AMP from GTP that is synthesized by the TCA cycle. Decrease of cellular ATP and rapid depolarization of mitochondrial membrane potential and induction of apoptosis by cigarette smoke have been well established in various forms of cancer. The role ofmitochondria in cisplatin resistance is further supported by recent data which showed that cisplatin may directly interact with mitochondria. Cisplatin introduces DNA damage by forming inter- and intra-strand nuclear DNA crosslinks. However, only a low percentage of intracellular platinum is bound to nuclear DNA, while a great majority of the intracellular drug interacts with nucleophilic sites on other molecules, including mitochondrial DNA. Mitochondrial DNA-cisplatin adducts may be more common than cisplatin adducts with nuclear DNA in the same cell line treated with the same concentration of cisplatin. Furthermore, Yang et al showed that glutathione counteracted the cytotoxicity of cisplatin by preventing ROS production rather than inhibiting formation of platinum/DNA adducts. Mitochondria are the predominant source of ROS produced in most apoptotic systems and mitochondrial homeostasis is critical in regulating apoptosis. In a normal system, apoptosis at the mitochondrial level is initiated by depolarization of the mitochondrial membrane, followed by release of cytochrome c.