Cancer is a major cause of deaths across the globe due to chemoresistance and lack of selective chemotherapy. Pyrido[2,3-d]pyrimidine is an emerging scaffold in medicinal chemistry having a broad spectrum of activities, including antitumor, antibacterial, CNS depressive, anticonvulsant, and antipyretic activities. In this study, we have covered different cancer targets, including tyrosine kinase, extracellular regulated protein kinases – ABL kinase, phosphatidylinositol-3 kinase, mammalian target of rapamycin, p38 mitogen-activated protein kinases, BCR-ABL, dihydrofolate reductase, cyclin-dependent kinase, phosphodiesterase, KRAS and fibroblast growth factor receptors, their signaling pathways, mechanism of action and structure–activity relationship of pyrido[2,3-d]pyrimidine derivatives as inhibitors of the above-mentioned targets. This review will represent the complete medicinal and pharmacological profile of pyrido[2,3-d]pyrimidines as anticancer agents, and will help scientists to design new selective, effective and safe anticancer agents.