Tumor molecular biology is an increasingly important consideration when choosing therapy for patients with advanced non-small cell lung cancer (NSCLC). A number of potential biological markers are under active investigation in the hope that it will be possible to identify markers that assist in patient selection for specific therapies. Distinguishing prognostic from predictive makers is crucial to the development of customized drug therapy. Some markers, such as mutations in the tyrosine kinase domain of the epidermal growth factor receptor (EGFR), are prognostic; patients with EGFR-mutant NSCLC have prolonged survival compared with those with wild-type disease, regardless of the treatment received. Although EGFR mutations are predictive of response to EGFR tyrosine kinase inhibitor (TKI) therapy, they do not appear to be predictive of a differential effect on survival. Other EGFR markers, such as protein expression or gene amplification, may be better predictors of a survival benefit from EGFR TKI. HER2 expression status and K-ras mutations provide additional information that may be useful in evaluating a patient for EGFR TKI therapy. Biological markers for chemosensitivity and resistance are also emerging. Patients with an elevated DNA repair capacity, evidenced by increased tumor expression of excision repair cross-complementing 1 or ribonucleotide reductase subunit M1 messenger RNA, may benefit less from cisplatin and gemcitabine, respectively, than from other agents. Increased levels of class III beta-tubulin are associated with taxane-resistance, and K-ras mutations have been associated with a lack of survival benefit from adjuvant chemotherapy in early stage NSCLC. It is likely that in the future, clinicians will evaluate a panel of biological markers in order to customize therapy for individual patients with NSCLC.