Cancer is a leading cause of human death worldwide. One of the greatest challenges in cancer therapy is the discovery and design of novel products with potential anti-tumor activities. In this study, a new protocol involves three-component condensation of the 3-amino-1,2,4-triazole as a 1,3-binucleophile, versatile aldehydes and N-methyl-1-(methylthio)-2-nitroethenamine as an enamine analogous in the presence of trichloroacetic acid as a Brønsted-Lowry acidic promoter leads to new functionalized N-alkyl-6-nitro-3,5-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-7-amine in moderate to good yields. The presence of five nitrogen heteroatoms in the product structure has gathered immense attention among chemists and biologists due to their biological values. Therefore, we evaluated the anti-tumor activity of our synthetic compounds on different cancer cells including human malignant melanoma cells (A375), prostate cancer cells (PC3 cells, LNCaP cells) and normal cells HDF (human dermal fibroblast). Notably, we found that compound 4b that contains a nitro group has the best anti-tumor activity on three different cancer cells. By using DAPI staining, we showed cancer cells death. Apoptosis induction was shown using quantitative real time PCR (qRT-PCR) by evaluating of Bax and Bcl2 mRNA levels. Finally, we demonstrated that 4b has epithelial-to-mesenchymal transition (EMT) inhibition effect on cancer cells (by induction of E-cadherin and reduction of vimentin mRNA expression levels as two potential EMT markers). So, 4b could be an anti-cancer promising drug. Although, in vivo experiments will be required to evaluate possible side effects.

癌症是全球人类死亡的主要原因。癌症治疗中最大的挑战之一是发现和设计具有潜在抗肿瘤活性的新产品。在这项研究中，一个新的协议涉及3-氨基-1,2,4-三唑作为1,3-双亲核试剂，多用途醛和N-甲基-1-（甲硫基）-2-硝基乙胺的三组分缩合。苯胺类似物在三氯乙酸的存在下作为布朗斯台德-劳里酸性助催化剂，可产生新的功能化N-烷基-6-硝基-3,5-二氢-[1,2,4]三唑[1,5- a] pyrimidin-7-amine，产量中等至良好。产品结构中五个氮杂原子的存在，由于它们的生物学价值而引起了化学家和生物学家的极大关注。因此，我们评估了合成化合物对不同癌细胞（包括人类恶性黑色素瘤细胞（A375），前列腺癌细胞（PC3细胞，LNCaP细胞）和正常细胞HDF（人类皮肤成纤维细胞）的抗肿瘤活性。值得注意的是，我们发现含有硝基的化合物4b对三种不同的癌细胞具有最佳的抗肿瘤活性。通过使用DAPI染色，我们显示了癌细胞的死亡。通过评估Bax和Bcl2 mRNA水平，使用定量实时PCR（qRT-PCR）显示了凋亡诱导。最后，我们证明了4b对癌细胞具有上皮到间质转化（EMT）抑制作用（通过诱导E-钙粘蛋白和降低波形蛋白mRNA表达水平作为两个潜在的EMT标记）。因此，4b可能是一种抗癌的有前途的药物。虽然，将需要体内实验来评估可能的副作用。