We report the design, synthesis and self‐assembly of a series of tandem triblock shape amphiphile regioisomers composed of one rod‐like fluorinated polyhedral oligomeric silsesquioxane (FPOSS) cage tethered with two spherical tert‐butyl‐POSS (tBPOSS) cages in para‐, meta‐, and ortho‐configurations. Precision syntheses are achieved through sequential “click” reactions and the compounds are thoroughly characterized by combined techniques of NMR, FT‐IR and MALDI‐TOF MS spectroscopy. Only the ortho‐isomer forms an ordered lamellae structure induced by the crystallization of perfluoroalkyl‐chains and the other two remain amorphous due to frustrated packing by unfavorable regio‐configuration. The distinct assembly behaviors of these three regioisomers reflect the delicate influence of a minute structural difference on precision macromolecular assembly.