Dendrimer, a new class of hyper-branched polymer with predetermined molecular weight, is being received much attention in nano biomedical applications such as anticancer drug delivery, gene therapy, disease diagnosis and etc. In this study, polyamidoamine (PAMAM)-based dendrimer generation 3.0 (G 3.0) was synthesized and subsequently pegylated. Obtained results showed that pegylation degree of the dendrimer was around 31% for its external amine groups. TEM image of the pegylated dendrimer exhibited spherical shape and nano sizes ranging from 30 to 40 nm. The fluorouracil (5-FU)-loaded pegylated dendrimer showed a slow release profile of the drug. In vitro study, at the primary screening concentration of 100 microg/mL, the PAMAM dendrimer presented higher toxicity in MCF-7 cells as compared to its pegylated counterpart. Meanwhile, the (5-FU)-loaded pegylated dendrimer exhibited the antiproliferative activity against the cell line with the IC50 of 9.92 +/- 0.19 microg/mL. In vivo tumor xenograft study, we succeeded in generating MCF-7 cells-derived cancer tumors on mice that was well-confirmed by using flow cytometer assay. The 5-FU encapsulated pegylated dendrimer exhibited a significant decrement in volume of the tumors which was generated by MCF-7 cancer cells.