Fluid pressurization in rock fractures under extreme conditions, such as reservoir impoundment and hydraulic fracturing, may accelerate the instability of fractured rock masses. Fluid pressure can reduce the effective normal stress on rock fractures and cause the degradation of the shear strength, resulting in destructive geohazards. Recent evidences show that the factor of safety (FOS) defined by the traditional methods may not predict the stability of rock masses appropriately. In our study, laboratory experiments, numerical simulations, and theoretical derivation are conducted to investigate the effect of fluid pressure distribution on the FOS assessment. We find that the Mohr-Coulomb failure criterion is inapplicable to predicting the instability of low-permeability fractures under significant fluid pressure gradients, and the FOS evaluated based on the failure criterion can be much higher than 1. The more considerable the fluid pressure gradient, the higher the FOS at fracture failure. Thus, a significant fluid pressure gradient may cause the premature failure of a rock fracture. Our analysis also indicates that the upper bound of the FOS at fracture failure under a non-linear fluid pressure gradient depends on the initial FOS of the fracture without fluid pressurization. The findings help advance the understanding of rock mass instability due to fluid pressurization and the safety control of rock engineering projects under extreme conditions.