A series of 2,5- and 3,5-diphenylpyridine derivatives was synthetised in high yields. A versatile chemical strategy allows the design of diphenylpyridines differently substituted with cationic or neutral side chains. The interaction of the molecules with DNA was investigated by biophysical and biochemical methods and an AT-binder (20) was characterised. A few cytotoxic molecules were identified but their antiproliferative activity does not correlate with DNA binding. Two compounds 18 and 22 showed significant antiproliferative activity and provide a novel route to potential anticancer agents.