Tubulin proteins, as components of the microtubule cytoskeleton perform critical cellular functions throughout all phases of the cell cycle. Altered tubulin isotype composition of microtubules is emerging as a feature of aggressive and treatment refractory cancers. Emerging evidence highlighting a role for tubulin isotypes in differentially influencing microtubule behaviour and broader functional networks within cells is illuminating a complex role for tubulin isotypes regulating cancer biology and chemotherapy resistance. This review focuses on the role of different tubulin isotypes in microtubule dynamics as well as in oncogenic changes that provide a survival or proliferative advantage to cancer cells within the tumour microenvironment and during metastatic processes. Consideration of the role of tubulin isotypes beyond their structural function will be essential to improving the current clinical use of tubulin-targeted chemotherapy agents and informing the development of more effective cancer therapies.