The instability of a polyisoprene (PI) thin film on a silicon substrate at room temperature in an aqueous environment was investigated by atomic force microscopy and optical microscopy. The instability mechanism changes from spinodal dewetting to hole nucleation with increasing film thickness, with the transitional thickness found to be around 46-50 nm. For PI films ≥50 nm, the dewetting was observed to proceed via successive stages of hole nucleation and growth, hole coalescence, cellular pattern formation and droplet formation. There is also a slowing down in the rate of the PI dewetting process and an increase in the pattern size as the film thickness is increased. In those films with observable holes, we also observed the coexistence of fine cellular cracking that is on a much smaller scale of hundreds of nanometres and extends only a few nanometres in depth from the film surface.