Hypoxia-inducible factor (HIF) is a central regulator involved in detection and adaption to cellular oxygen stress through regulation of the hypoxic transcriptional program in angiogenesis, erythropoiesis, and metabolism. The HIF pathway is involved in many diseases. On one hand, overexpression of the HIF pathway is associated with solid tumors such as renal cell carcinoma (RCC). On the other hand, suppression of the HIF pathway is correlated with inflammatory, anemia, and other hypoxic-ischemic diseases. Therefore, modulation of the HIF pathway has been perceived as a promising strategy for treating HIF-related diseases. Recent advances in understanding of the biochemistry underlying the HIF pathway have stimulated small-molecule drugs development, and therapeutically, manipulation of the HIF-mediated response has been shown to have considerable medicinal potential. This review will summarize and provide insight into recent advances in research that have expanded our knowledge of the HIF pathway, including its structural basis and biology, small-molecule modulators of the pathway, including inhibitors and activators, and the potential therapeutic applications of these modulators.