Chronic lymphocytic leukemia (CLL) is the most common lymphoid neoplasia in Western societies and is currently incurable. Multiple treatment options are practiced, but the available small molecule drugs suffer from dose-limiting toxicity and undesirable side effects. The need for new, less toxic treatments is a pressing concern. Here, we demonstrate that (-)-agelastatin A (1a), a pyrrole-imidazole alkaloid obtained from a marine sponge, exhibits potent in vitro activity against primary cell lines of CLL and disclose the synthesis of several analogues that are equipotent or exceed the potency of the natural product. The novel synthetic analogue, 13-debromo-13-trifluoromethyl agelastatin A (1j), showed higher activity than the natural product when tested against the same cell lines and is the most potent agelastatin derivative reported to date. A detailed in vitro structure-activity relationship of 1a in CLL compared to that of 22 synthetic analogues is described along with preliminary in vivo pharmacokinetic and metabolism studies on the most potent compounds.