There is clear epidemiological evidence that plasma levels of high-density lipoprotein (HDL) - cholesterol are inverse and independent predictors of cardiovascular disease risk, fuelling interest in novel therapies capable of raising HDL-cholesterol. However, the relevance of HDL-cholesterol as a surrogate marker for HDL-related risk has been questioned. Latest failures of HDL-cholesterol raising drugs and a recent study that showed no causal association between risk for myocardial infarction and genetically raised plasma HDL-cholesterol indicate that steady-state HDL-cholesterol concentrations may provide limited information regarding the potential antiatherogenic functions of HDL. There is accumulating evidence that HDL composition determines its functional properties, rather than the levels of circulating HDL-cholesterol. Therefore, assessing HDL composition and function might provide more relevant information than steady-state HDL-cholesterol levels. Recent mass spectrometric analyses revealed that protein composition of HDL is complex, expanding our understanding of the functions and structures of lipoproteins. Significant alterations were identified in the composition and function of circulating HDL of patients with high cardiovascular risk, as well as in HDL isolated from atherosclerotic tissue. These novel insights may help to develop therapies that target the functionality of HDL and further enable the identification of patients at increased cardiovascular risk.