Lipooxygenase Inhibitors as Potential Anti-Cancer Drugs Ted Holman, Chemistry & Biochemistry Lipoxygenases are enzymes implicated in a broad range of human cancers, as well as cardiac and inflammatory diseases. Ted Holman's laboratory examines the enzymatic mechanism and biological function of lipoxygenase in the hopes of developing novel inhibitors. In collaboration with UCSC Professor Phil Crews, his laboratory has identified potent lipoxygenase inhibitors and are currently characterizing their structure/function reactivity. The results of this work will shed light on their potential as anti-cancer agents. (Keywords: proteomics, cancer, enzymes, bioinorganic chemistry, lipoxygenase, heme binding)

Potential disease targets for human lipoxygenases inhibitors

Proposed mechanism of the soy bean lipoxygenase (SLO)

Potential sites for the allosteric cavities of SLO

Lipoxgenaseinhibitors found in the marine natural products (MNP) library of UCSC Professor Phil Crews

Lipoxygenases have also been implicated in a wide range of human diseases. There are three major human lipoxygenases (HLO): 5-, 12-, and 15-HLO. Their primary enzymatic difference is their positional specificity on arachidonic acid (AA). These lipoxygenase products are the precursors of hormones, such as leukotrienes and lipoxins, which have been implicated as critical signaling molecules in a variety of inflammatory diseases and cancers. 5-HLO products cause bronchial constriction, while 12-HLO plays a major role in psoriasis, and 15-HLO oxidizes low-density lipoproteins that are thought to initiate the primary stage of atherosclerosis. Lipoxygenase isozymes are also involved in uncontrolled cell growth and/or regulation. 15-HLO has been shown to be a key biological agent in colorectal cancers, while 12-HLO is involved in pancreatic, breast and prostate cancers. 5-HLO is up-regulated in prostate cancer and its inhibition abolishes all cell proliferation, inducing apoptosis.

Examining the Mechanisms of Lipoxygenase Inhibition

 The broad implication of lipoxygenase in human disease has elicited great interest in this protein as a potential therapeutic target and has provided the motivation for the Holman laboratory's investigation of lipoxygenase activity and inhibition.

 In order to develop lipoxygenase inhibitors, one must first understand the protein's enzymatic function. Holman and his colleagues have used a variety of spectroscopic and kinetic methods to study three specific lipoxygenase enzymes: soybean 15-LO (SLO), human 12-LO (12-HLO) and human 15-LO (15-HLO). Because it is more robust than the human variants, the laboratory uses the soybean enzyme as a model system to develop a detailed mechanism for lipoxygenase as a class of enzymes. They have compared the mechanism of SLO with the human enzymes and determined that these mechanisms are remarkably similar, despite a 100-fold difference in rate.

 Holman and his colleagues have used this mechanistic information to investigate inhibition of lipoxygenase and determined that both SLO and HLO have allosteric inhibitory sites. Holman believes that the allosteric site may be critical to the regulation of lipoxygenase activity by either suppressing or activating the enzyme, and that the site might serve as a potential target for a novel class of lipoxygenase inhibitors. With this in mind, the laboratory is using stopped-flow kinetics to establish the binding constraints of this site. Such information would contribute significantly to rational design of an allosteric inhibitor.

Characterization of Novel Lipoxygenase Inhibitors

 Another research project in the laboratory involves identification and characterization of novel inhibitors to lipoxygenase. The research group has discovered over 20 unique lipoxygenase inhibitors by screening the marine natural products (MNP) library of their collaborator, UCSC Professor Phil Crews. The Crews MNP library is one of the worlds largest and offers a unique source of novel chemical structures unavailable through standard synthetic or combinatorial methods.

 Holman and his colleagues perform biochemical and spectroscopic methods to determine how these lipoxygenase inhibitors bind and function. They are also testing their effects on the overall biochemistry of cancer cell with proteomics (the induction and/or repression of all the proteins in a particular cell line). For example, in one set of experiments, they plan to challenge prostrate cancer cells with the addition of different lipoxygenase inhibitors, measuring their effectiveness in killing cancer cells and determining what biochemical pathways are affected. Classification of each inhibitor by its potency, mode of action, and selectivity is expected to contribute to the design the next generation of lipoxygenase pharmaceuticals.