Background: Pyrrolo[2,3-d]pyrimidines have been recently reported to have anticancer activities
through inhibition of different targets such as, Epidermal Growth Factor Receptor (EGFR) tyrosine kinase,
Janus Kinase (JAK), mitotic checkpoint protein kinase (Mps1), carbonic anhydrase, MDM-2. On the other hand,
aryl urea moieties which are found in some tyrosine kinase inhibitors such as Sorafenib and Linifanib have
aroused recent attention as responsible for anticancer activities. The aims of this paper are to synthesize pyrrolo[
2,3-d]pyrimidine derivatives containing urea moiety and evaluate their anti-cancer activity against human
lung cancer cell line (A549), prostate cancer cell line (PC3), human colon cancer cell line (SW480) and human
breast cancer cell line (MCF-7).
Methods: A series of new pyrrolo[2,3-d]pyrimidines containing urea moieties have been synthesized as
Scheme 1. In vitro cytotoxicity of target compounds were evaluated against, SW480, PC3, A549 and MCF-7
human cancer cell lines using a MTT assay. In order to evaluate the mechanism of cytotoxic activity of
compounds 9e, 10a and 10b, having the best cytotoxic activity, Annexin V binding assay, cell cycle analysis
and western blot analysis were performed.
Results: Among the target compounds, 10a (IC50 = 0.19 µM) was found to be the most potent derivative against
PC3 cells. Compound 10b and 9e showed the strong cytotoxic activity against MCF-7 and A549 cells with IC50
value of 1.66 µM and 4.55 µM, respectively. Flow cytometry data suggest that the cytotoxic activity of the
compounds on cancer cells might be mediated by apoptosis revealing a significant increase in the percentage of
late apoptotic cells and causing a cell cycle arrest at different stages. Western blot analysis of apoptosis marker
demonstrated that these compounds induce apoptosis through the intrinsic pathway.
Conclusion: Compound 9e displayed the strongest cytotoxicity against A549 cancer cell line, and induced late
apoptosis in A549, as confirmed by cell cycle arrest in G0/G1 phase. In addition, compound 9e reduced expression
of the anti-apoptotic protein Bcl-2 and enhanced expression of the pro-apoptotic protein Bax, besides increased
caspase-9 and caspase-3, as well as cleavage of PARP levels. These results suggest that compound 9e
showed a cytotoxic effect in A549 cells through activation of the mitochondrial apoptotic pathway. Further
studies will be undertaken in our laboratory to improve cytotoxic activity of compound 9e and to identify the
biological targets of 9e which are responsible for anticancer activity.