The title compound 2-[2-(2-fluorobenzylidene)hydrazinyl]-4-(1-methyl-1H-indol-3-yl) thieno[3,2-d]pyrimidine (C22H16FN5S) was prepared and its structure was confirmed by IR, H-1 NMR, MS, elemental analyses and X-ray diffraction. The crystal of the title compound belongs to the monoclinic system, space group P2(1)/c with a = 14.4546(17) angstrom, b = 17.0895(19) angstrom, c = 17.9621(15) angstrom, alpha = 90 degrees, beta = 122.717(6), gamma = 90 degrees, V = 3733.1(7) angstrom(3), Z = 4, and R = 0.0412 for 4816 observed reflections with I > 2 sigma(I). In addition, the compound possesses distinct effective inhibition on the proliferation of HT-29, A549 and MKN45 cell lines.