Projects

Alkylpyridinium polymers (polyAPS) are natural products isolated from the Mediterranean species of marine sponge Reniera sarai. At first, these compounds were recognized as acetylcholinesterase (AChE) inhibitors, but preliminary experiments with their synthetic analogues have revealed they are much stronger antagonists of ?7 nicotinic acetylcholine receptors (nAChRs). Natural polyAPS are cytotoxic for a non-small cell lung cancer (NSCLC) adenocarcinoma cell line (A549), but not for normal lung fibroblasts that do not express ?7 or other nAChR subtypes, which are less susceptible to polyAPS. Preliminary experiments also revealed the cytotoxic effects of polyAPS on a lung adenocarcinoma primary cell line as well as their ability to limit the growth of solid tumors in mice. However, serious drawbacks of the polyAPS compounds include their large, heterogeneous molecular size, their propensity to form even larger supramolecular aggregates in aqueous solutions and their non-specific cytolytic activity. Therefore a therapeutic use of these natural products seems unlikely. Recently, we prepared a series of smaller synthetic alkylpyridinium oligomers with similar structural and functional properties. Each has a defined molecular weight and is not prone to aggregate in aqueous solutions. Initial experiments have shown that at least one analogue (8Br-1) is very potent ?7 nAChR antagonist at pM concentrations. This concentration is several orders of magnitude lower than is required for AChE inhibition. This finding is important since AChE inhibition might enhance cancer cell proliferation, which would oppose the effects of nAChR antagonists that reduce proliferation of NSCLC, SCLC and MPM (mezothelioma) lung cancer cell lines. The ?7 nAChR also is important for mediating cognitive functions in the central nervous system, protected by the blood brain barrier. In nonneural cells expressing nAChRs, binding of the agonist nicotine triggers antiapoptotic and proliferative signaling pathways which prevent cell death and enhance tumor growth. By using nAChR antagonists, we would like to achieve the opposite effect: to activate proapoptotic pathways that lead to cell death and destruction of cancer cells and inhibit their proliferation. We believe that the smaller, synthetic alkypyridinium analogs, due to their high affinity for ?7 nAChRs, may become useful drugs in treating one of the most common types of lung cancer connected with smoking. These compounds may also serve as tools to elucidate certain signaling pathways and the roles of proapoptotic proteins involved in mitochondrial damage and activation of caspases. Our in vitro studies will provide a solid foundation for our subsequent tests of these compounds in animal models of these cancers.

We expect that the results of our project proposal will reveal the possibility to implement the nAChR antagonists as a new group of chemotherapeutics for treatment of those tumor types which express specific nAChR. To implement such a therapy it is of outmost importance to understand cell apoptotic signaling pathways. Our compounds should be therefore also useful as molecular tools to elucidate and to better understand the complex mechanisms of signaling pathways. Worldwide, these processes are widely studied, however the role of nAChR and their antagonists which can trigger apoptosis is rather new. Furthermore, the number of antagonists of nAChR, which could be used without significant toxic side effects is rather limited. The revelation that nAChR are over expressed in certain cancer cells, and that certain subtypes of nAChR prevail on their membranes, opens new possibilities in therapeutic strategies. In addition, in cancer cells most elements of cholinergic loop are up regulated, acetylcholine binds to nAChR and triggers the antiapoptotic pathways. In this regard the application of specific antagonists is even more interesting. In our case such approach seems particularly important since antiapoptotic pathways in cancer cells of majority of lung tumors are also triggered by binding of nAChR agonist nicotine. Nicotine is the major alkaloid in tobacco smoke inhaled by smokers. The prevention of endogenic acetylcholine as well as exogenic nicotine binding to nAChR by the use of nAChR antagonists could be a successful approach to limit or eliminate the tumor promoting effects of nicotine in lung cancer cancerogenesis.