Eight human telomerase inhibitors (5a-5h) having the core of N-acyl-4,5-dihydropyrazole with anticancer effects were identified in this study. Biological results revealed that a few compounds had potent anticancer activities against three common tumor cell lines (SGC-7901, HepG2 and MGC-803). Among them, compound 5c, with a molecular weight of only 272.2 Da, had antiproliferative activities against SGC-7901 and MGC-803 with EC50 values of 2.06 ± 0.17 and 2.89 ± 0.62 μM, respectively, better than 5-Fluorouracil. Compound 5c inhibited the enzyme of telomerase with an IC50 value of 1.86 ± 0.51 μM, surpassing the control compound, ethidium bromide. Modeling study showed that this compound can reside in the binding pocket of the telomerase/TNA:DNA hairpin complex. When the moiety of N-acyl was changed to N-sulfonyl, the gotten compounds (8a-8i) had deteriorative activities against both these three cancer cell lines and the enzyme of telomerase.