• protein kinases
• pyrimido-naphtyridines
• pyrimido-quinazolines
• VEGFR-2
• VEGFR-2 inhibitors

Protein kinases (PKs) represent a wide family of homologous enzymes, both transmembranous and cytoplasmic, that catalyze the transfer of the γ-phosphate group from ATP to a hydroxy group of selected target protein substrates. This covalent post-translational modification is a pivotal component of normal cellular communication and maintenance of homeostasis.1
PKs can be divided in two principal classes: Tyrosine kinases (TKs) and Serin-Threonin kinases (STKs). The first one catalyze the transfer of the phosphate group to the phenolic group of a tyrosine, STKs phosphorylate the alcoholic group of a serine or a threonine. Phosphorylation/de-phosphorylation are the more common post-translational modifications and the more potent mechanism for signalling transduction and enzymatic regulation. Kinases are key regulators of almost all cellular processes, they control the activity, localization and overall function of many proteins, they are essential in signal transduction and coordination of complex biological functions such as growth, differentiation, metabolism and apoptosis, in response to external and internal stimuli. Recent advances have confirmed the role of tyrosine kinases in the pathophysiology of cancer. Though their activity is tightly regulated in normal cells, they may acquire transforming functions due to mutation(s), overexpression and autocrine paracrine stimulation, leading to malignancy. For this reason kinase proteins represent a valid target in the therapy against cancer.
The molecular basis of tumour angiogenesis has been extensively studied and the Vascular Endothelial Growth Factor (VEGF) pathway has emerged as one of the most important positive modulators of this process. The various members of the VEFG family have overlapping abilities to interact with a set of tyrosine kinase receptors: VEGFR-1 and VEGFR-2 (KDR), largely expressed in endothelial cells and primarily involved in angiogenesis, and VEGFR-3, located in lymphatic vessels, where it seems to be critical in regulating lymphoangiogenesis. The demonstration that expression of VEGF and of its receptors correlates with the degree of vascularization of many experimental and clinical tumours led to the rational design and development of agents targeting this pathway. The new agents range from anti-VEGF monoclonal antibodies, such as bevacizumab, to small-molecule ATP-competitive VEGFR inhibitors, including compounds from distinct heterocyclic classes.2,3
Many derivatives characterized by a variously substituted pyrimidine ring, either isolated or fused to form different heterocyclic structures, have been reported as interesting VEGFR inhibitors. 4
Inside the class of fused pyrimidines, those bearing the aniline substituted pyrimidine core showed to possess excellent antiangiogenic effects, and they have widely been used as a basis for the design of new VEGFR-2 competitive inhibitors.
The research program performed in the laboratory where I worked for my thesis has always been devoted to the preparation and evaluation of new antiproliferative agents. On the basis of previous results, a serie of benzothiopyrane- and pyridothiopyrane-fused pyrimidines, characterized by an aniline group in the 2-position, has been recently synthesized.5
Actually it was hypothesized that these new products might share the pharmacophoric requirements suitable for anchoring to the ATP binding site, in particular the aniline NH group and the pyrimidine N1 and N3 atoms. Moreover it seemed plausible to investigate if the thiopyrane sulfur atom might be directly implicated in additional H-bonding interactions with critical residues at the catalytic cleft, as it has been reported for the previously described thiophene-containing scaffold.
In this regard, my thesis work concerned the preparation of novel heteropolycyclic compounds, characterized by an aniline (variously substituted) inserted in an appropriate position of the scaffold.
In particular I synthesized a series of 2-anilino substituted pyrimido-naphtyridines and 2-anilino substituted pyrimidoquinazolines.
The ability of the novel derivatives to inhibit the kinase activity of VEGFR-2, as well as their cytotoxic effect, will be determined by the research group of the University of Padua.
In particular, the antiproliferative activity of new thiopyrano-fused pyrimidines will be performed on HeLa (cervix adenocarcinoma), A-431 (epidermoid carcinoma) and MSTO-211H (biphasic mesothelioma) cell lines and the results will be expressed as IC50 values, i.e. the concentration of compound able to produce 50% cell death with respect to the control culture. With the aim to investigate the mechanism of action responsible for the antiproliferative effect, the ability of new derivatives to inhibit the KDR tyrosine kinase activity will be determined by a biochemical assay performed with a recombinant human kinase insert domain receptor.

References:
[1] Manash, K P.; Mukhopadhyay, A. Tyrosine kinase-role and significance in cancer. Int. J. Med. Sci. 2004, 1, 101-115
[2] Irby, R.B.; Jiang, Y. Tyrosine Kinase Update: Role and Response in Cancer Therapy, Curr. Cancer Ther. Rev., 2011, 7, 155-166.
[3] Natoli, C.; Perrucci, B.; Perrotti, F.; Falchi, L.; Iacobelli, S. on behalf of Consorzio Interuniversitario Nazionale per la Bio-Oncologia (CINBO), Tyrosine Kinase Inhibitors Current Cancer Drug Targets, 2010, 10, 462-483.
[4] Musumeci, F.; Radi, M.; Brullo, C.; Schenone, S. Vascular Endothelial Growth Factor (VEGF) Receptors: Drugs and New Inhibitors, J. Med. Chem. 2012, 55, 10797−10822.
[5] Dalla Via, L.; Gia, O.; Marciani Magno, S.; Marini, A.M.; Salerno, S.; Da Settimo, F.; Fornaciari, G.; Simorini, F.; La Motta, C.; Taliani, S.; “Synthesis, Antiproliferative Activity And VEGFR-2 Inhibition of New Anilino-Substituted Thiopyrano Fused Pyrimidines”, XX National meeting on Medicinal Chemistry, September 12-16 2010 Abano Terme-Padova.