For most of the 25,000 human genes, the removal of introns by pre-messenger RNA splicing represents an essential step towards the production of functional messenger RNAs. Moreover, a majority of pre-messenger RNAs is alternatively spliced to yield different messenger RNAs. Cancer cells often display aberrant profiles of alternative splicing producing isoforms that can stimulate cell proliferation and migration or improve resistance to apoptosis. While mutations at splice sites or in splicing control elements have been identified, changes are also caused by alterations in the expression of proteins that affect splice site selection. Current challenges consist in documenting the functional diversity generated by alternative splicing and its contribution to different types of cancers. The development of innovative approaches aimed at reprogramming alternative splicing offers promising perspectives in cancer therapy.