Septin 9 (SEPT9), a GTPase, known as the fourth cytoskeleton, is widely expressed in various cells and tissues. The functions of SEPT9 are partly similar to other cytoskeletons as a structure protein. Further, SEPT9 can interact with other cytoskeletons, participating in actin dynamics and microtubule regulation. SEPT9 is associated with various diseases, such as cancers. Thus, it could be a potential drug target. However, there are no small molecule SEPT9 inhibitors and the only reported septin inhibitor, forchlorfenuron, has no effects on SEPT9 inhibition from our study results. Therefore, the derivatives of forchlorfenuron were synthesized, and their activities were evaluated by a direct SEPT9 inhibition screening platform, followed by localized surface plasmon resonance (LSPR) and cell-based assays. The screening results conveyed that 6b, 8a, and 8b are SEPT9 inhibitors with IC