Urological malignancies: translational cancer research
Pagano Lab’s research work is focused on urological malignancies, with particular attention to prostate and bladder cancer. Cancer is the prototype signal transduction disease where aberrant signaling leads to the re-programming of normal cells turning them into malignant tumor cells. We are interested in finding novel and feasible points of intervention within the intricate signaling network that governs cell death in cancer cells. The main goal of the lab is a fast transfer of the basic research information into the clinical practice. The research design is based on both an in vitro model based on cancer cells and an ex vivo approach where surgically obtained tissue blocks from patients are cultured and treated in the lab.
Novel strategies for drug-treatment of prostate cancer
Figure 1. Light microscopy images of the prostate cancer cells PC-3, treated with Serenoa repens extract. (A) Control cells (time 0). (B) Cells after 4 h of treatment, showing extensive cytosolic vacuolization and 40% increase in size; (C) massive apoptosis of the cells after 24 h of treatment.
We are focusing on new drugs to be used in prostate cancer cure or prevention. We are currently working on an natural extract from palm tree (Serenoa repens extract), which, in the field of men’s health, is the most common dietary supplement used for treatment of benign prostate hyperplasia (BPH). This drug, a natural oil without significant side-effects, shows the potentiality to be used for prostate cancer prophylaxis and long-term therapy. In fact, when used on prostate cancer cell lines, this extract is able to induce massive apoptosis though the mitochondrial pathway. A remarkable finding was the selectivity of the effect for prostate cancer cell lines, which probably reflects their unique metabolic features. Also cells negative for the androgen receptor were responsive, suggesting that the extract could be useful for the killing of tumor cells resistant to antiandrogen therapy. Given the high tolerability and the clinical absence of side-effects, it is possible to hypothesize a future use of Serenoa repens extract for long-term therapy or for prostate cancer prophylaxis in high risk pathway.
Prostate carcinogenesis in Benign Prostate Hyperplasia (BPH)
It is critical to use in translational prostate research an experimental model which can faithfully represent the real prostatic tissue. In our lab, we are working on an ex vivo model based on tridimensional prostate tissue blocks obtained from surgery. The tissue is sampled in the surgical room and quickly transferred to the lab.
Carcinogenesis in the prostate seems to derive from an abnormal proliferation of the cells with intermediate differentiation. These cells increase in BPH. We hypothesisze that these cells could be involved in prostate carcinogenesis in BPH. Moreover, we have recently observed that drugs able to induce apoptosis in BPH, such as Serenoa repens extract, block the proliferation of the intermediate cells in the blocks ex vivo. This information could have important implications for prostate cancer prevention in BPH patients.
Ex vivo assessment of drug induced apoptosis in prostate cancer
We have previously shown that detectable mitochondrial changes can be used as a marker of apoptosis after drug treatment of several cancer cell lines in vitro. Moving to the clinical setting, a translational project has been started, looking at apoptosis induction in human cancer tissue ex vivo. For this purpose, short term cultures of prostate cancer blocks are being used. Using novel apoptosis-inducing drugs, we have been able to detect an apoptotic response in the blocks by light microscopy, immunohistochemistry or electron microscopy. The future aim of this project is to identify a pathway from bedside to bench and viceversa, where patients’ samples are tested in the lab ex vivo and the results from the lab are transferred to clinical practice. This approach could be helpful in selecting new and promising individualized treatment strategies for prostate cancer patients in the near future.
Investigating drug resistance in bladder cancer: a functional genomic approach
Figure 2. DNA microarray data of genes that potentially suppress Gemcitabine mediated cell death. The more resistant the cells are to Gemcitabine, the higher is the expression of these genes.
The most practical and unbiased way for identifying differences between cancer and normal tissue is the use of DNA microarrays for the measurement of gene expression changes. The size of a stamp, DNA microarrays can quantify the expression of every known gene (more than 40000) in a given biological sample. Hence, the gene expression in cancer tissue can be thoroughly compared to normal tissue in order to find out which genes are aberrantly expressed in cancer. For this reason, in recent years DNA microarrays have been employed very successfully for the discovery of novel tumor markers, i.e. genes whose expression can distinguish between sub-types of cancer that respond differently to treatment but that could not be identified by traditional diagnostics.
DNA microarrays were also thought to be a good tool for the detection of genes whose stronger expression in cancer contributes to cancer formation. The products of these genes (the proteins) could serve as “points of attack” at which novel therapies could be directed. However, finding these genes turned out to be harder than thought because most of the expression changes in cancer are not a cause of cancer formation but a consequence. Attacking these genes/proteins would therefore not help in fighting this disease.
We use DNA microarrays in a functional genomics strategy to identify genes that cause inhibition of cell death in cancer. By first assigning a function to the genes we can be much more confident that the genes we find actually contribute to cancer formation when they are overexpressed in tumor tissue.
Currently, we are investigating which mechanisms bladder cancer cells utilize in order to avoid cell death by the chemotherapeutic drug Gemcitabine. This drug is being used for treatment of a wide variety of cancer types; among them is also metastatic bladder cancer. As with any cancer drug, resistance to its action from the outset or the recurrence of a resistant tumor after seemingly successful therapy constitutes a great problem and prevents a better treatment. We compare the gene expression of cancer cells with different levels of resistance towards Gemcitabine and identify the genes whose expression most closely correlates with the degree of resistance. Genes that are stronger expressed in the more resistant cells are potential modulators of resistance. Subsequently, we focus further studies on the genes coding for proteins that may be inhibited by small molecules or antibodies and we attempt to elucidate how these genes influence cell death decisions at the molecular level.