MOLECULAR AND CELLULAR TARGETS INVOLVED IN THE ACTIVITY OF DRUGS AND BIOLOGICALLY-ACTIVE PRODUCTS
Research team:
Prof. Francesco Dall'Acqua (Tel. 049/8275708; This e-mail address is being protected from spambots. You need JavaScript enabled to view it )
Prof.ssa Daniela Vedaldi (Tel. 049/8275707; This e-mail address is being protected from spambots. You need JavaScript enabled to view it )
Dott.ssa Alessia Salvador
General remarks.
It is well known that activity of drugs involves in general interactions with specific targets, and these interactions are responsible of the pharmacological effects.
A general strategy which is developed in the research group consists in recognizing the targets involved both at cellular and at molecular level and evidencing the physical chemical and biological modifications which occur in the interactions between the targets and the biological active molecules.
In general the research group deals with :
Evaluation of drugs for thalassemia treatment:
The research for the treatment of such haematological diseases as beta-thalassemia and sickle cell anaemia is now based on the pharmacological regulation of gamma-globin gene expression, with consequent haemoglobin synthesis. This makes patients more independent from transfusion therapy.
Among the recently studied gamma-globin inducers, linear and angular psoralens present excellent ability to induce gamma-globin biosynthesis, through cellular differentiation, in different experimental models. A model in which various derivatives will be evaluated is the erythroblastoid line, K562. Different compounds will be studied for their erythrodifferentiating properties and for their haemoglobin production. The more interesting molecules in this model will be selected and their activity will be checked using another experimental model such as human stem cells by healthy volunteers or even by thalassemic patients.
Evaluation of new potential antitumour compounds:
The research in tumour therapy recently focuses on the following objectives: the project and study of compounds with selectivity towards a specific target in tumour cells to damage them. The research we carry out consists in the evaluation of different compounds, usually heterocyclic ones, obtained by molecular project and subsequent synthesis. First of all, their antiproliferative activity is checked in tumour cell lines. We evaluate their capability to induce apoptosis (normally in Jurkat cells), and the mechanism of cell death (caspase activation, mitochondria interference, etc.).
The assessment of an eventual cell cycle block is carried out by flow cytometry; thus we can obtain precise indications; for instance, if we observe a M (mitosis) block, we hypothesize a possible involvement of microtubules. Moreover, we are able to study ROS formation and topoisomerase I and II interactions. Thanks to these tests, their mechanism of action will be elucidated, and they will be eventually proposed as leads for further research developments.
Evaluation of compounds with potential photonuclease activity and antitopoisomerase activity.
We study the DNA binding properties of new compounds, synthesised with aim of obtaining DNA binders, and their capability to induce DNA single strand breaks after UVA light activation. The plan is to verify their eventual photonuclease activity and then to use specific enzymes which cleave DNA in particular tracts. The DNA binding mode by these new derivatives will be evaluated in detail using different spectroscopic techniques. Moreover, plasmid DNA photocleavage experiments will be carried out and the photocleavage kinetics and the precise sites will be investigated through sequencing gels.
It is well known that molecules which undergo intercalation inside duplex DNA, as quinolizinium derivatives , can interact effectively with Topoisomerases, which are fundamental enzymes for DNA replications. They initially induce DNA strand breaks following by reorganization and rejoining of the break chains of DNA . Some quinolozinium derivatives form a tertiary complex between DNA and Topoisomerase, inhibiting the repair of DNA and consequently inducing cell death, and therefore exhibit potential antitumor activity
Main techniques and methodologies:
The research is carried out both in model substrates (cell membranes, proteins, DNA and oligonucleotides) and cell cultures (tumor and not tumor) with the use of UV-Vis spectrometry, fluorimetry, circular dichroism, flow linear dichroism, chromatographic techniques (HPLC), electrophoresis (SDS-PAGE and agarose), flow cytometry, ELISA (Enzyme Linked Immunosorbent assay), Western blot.