Chronic lymphocytic leukemia (CLL) is a common B-cell malignancy characterized by a highly variable clinical course. The behavior of the disease is believed to be influenced by microenvironmental signals that regulate the proliferation and survival of the malignant B-cells. Signals transduced through Toll-like-receptor-9 (TLR9) may play a particularly important role, as they could drive the expansion of a subset of cells that express B-cell receptors reactive with DNA or DNA-containing complexes. Interestingly, leukemic cells from patients with aggressive disease respond more effectively to TLR9 stimulation than their less aggressive counterparts, suggesting that the capacity to respond to TLR9 signals can define distinct prognostic subsets in CLL. The exact mechanism(s) accounting for the variability in the response to TLR9 engagement are still unclear, although important differences have been observed between prognostic groups in terms of downstream signaling events and gene- and miRNA-expression profiles. Understanding the mechanism(s) that underlie the different TLR9 responses should provide further insight in the pathophysiology of CLL and may lead to the identification of novel targets for therapeutic intervention.