As the cure rates for haematological malignancies have improved, the exploration of the balance between efficacy and side effects has become a major research target. The antifolate methotrexate is widely used in the treatment of acute lymphoblastic leukaemia, non-Hodgkin lymphoma, and osteosarcoma. Even when given identical methotrexate doses, patients vary significantly in their response and pattern of toxicities. This diversity can, to some extent, be linked to sequence variations in genes involved in drug absorption, metabolism, excretion, cellular transport, and effector targets or target pathways. In the coming years pharmacogenomics is expected to change our approaches to individualised therapy with methotrexate. However, genetic polymorphisms affect the pharmacokinetics and dynamics of all the drugs a patient receive as well as the normal tissues tolerance to a given drug exposure. Thus, although high-throughput techniques will allow mapping of tens of thousands of genetic polymorphisms in one run, it will be a major challenge to dissect out which of these have the strongest impact on efficacy and toxicity and hence should be the targets for intervention. This paper discusses the pharmacology of methotrexate and reviews studies on haematological malignancies that have attempted to predict the risk of toxicity by specific clinical or genetic features.