Protein disulfide isomerase (PDI) family is a group of multifunctional endoplasmic reticulum (ER) enzymes mediating the formation of disulfide bonds, catalyzing the cysteine-based redox reactions and assisting the quality control of the client proteins. Recent structural and functional studies have demonstrated that PDI members not only play an essential role in the proteostasis in the ER but also exert diverse effects in numerous human diseases including cancer and neurodegenerative diseases. Increasing evidence suggests that PDI is actively involved in the proliferation, survival, and metastasis of several types of cancer cells. Although the molecular mechanism by which PDI contributes to tumorigenesis and metastasis remains to be understood, PDI is now emerging as a new therapeutic target for cancer treatment. In fact, several attempts have been made to develop PDI inhibitors as anti-cancer drugs. In this review, we discuss the properties and diverse functions of human PDI proteins and focus on recent findings on their roles in disease states including cancer and neurodegeneration.