Identification and characterization of a novel human oxygenase responsible for the formation of 15(R)-HETE and 17(R)-HDHA in tumor cells and neutrophil granulocytes

• Applicant: Dr. Astrid Kahnt
• Subject Area: Pharmacy
• Term: since 2024
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 535161404

The lipid mediators 15-Hydroxyeicosatetraenoic acid (HETE) and 17-Hydroxydocosahexaenoic acid (HDHA) play a central role in a number of human physiological and pathophysiological processes, including inflammation initiation and resolution as well as cell proliferation and differentiation. These monohydroxylated arachidonic acid (ARA) and docosahexaenoic acid (DHA) derivatives are primarily produced by activated human leukocytes such as macrophages and granulocytes, but some solid malignant tumors are also known to produce significant amounts of these oxylipins. The formation of monohydroxylated oxylipins is an enzyme-driven process in which oxygen is stereospecifically incorporated into specific positions of the respective polyunsaturated fatty acid by various oxygenases. In humans, the formation of 15-HETE/17-HDHA is primarily carried out by two 15-lipoxygenases (15-LO-1 and 15-LO-2) which strictly incorporate the oxygen in S-configuration in position 15 of ARA / position 17 of DHA. In addition to the two 15-LOs, cyclooxygenase-1 (COX-1), aspirin-acetylated cyclooxygenase-2 (COX-2) and certain cytochrome P450 (CYP450) isozymes can also form small amounts of 15-HETE/17-HDHA. Here, however, the oxygen is predominantly R-configured in the molecule. There is evidence in literature that humans possess another, as yet unknown, oxygenase involved in the formation of 15-HETE and 17-HDHA. Indeed, our recent data show that several human cancer cell lines as well as neutrophilic granulocytes release significant amounts of 15-HETE and 17-HDHA even though they do not express any of the known 15-LOs. Closer examination revealed that the released 15-HETE/17-HDHA are largely R-configured and neither COX nor CYP450 isozymes are involved in their formation. Nonspecific oxidation could also be excluded. Therefore, our data clearly indicate the existence of a previously unknown enzyme that catalyses the formation of 15(R)-HETE from ARA or 17(R)-HDHA from DHA. Given the importance 15-HETE and 17-HDHA have in numerous human physiological and pathophysiological processes, this project aims to identify and systematically characterize the unknown oxygenase. Furthermore, the identified enzyme will be recombinantly expressed, purified and a suitable assay system will be established to screen for inhibitors with the help of a fatty acid mimetics library. In addition, expression of the enzyme in human leukocyte subsets and colon cancer cell lines will be studied, and the consequences of a knock-out of the new oxygenase on tumour cell and neutrophil function will be explored. These data will establish a novel pathway in the formation of bioactive lipids in humans and thus offers a new target for the treatment of human inflammatory diseases and cancer.

• DFG Programme: Research Grants
• Co-Investigators: Dr. Andreas Reuter; Dr. Dominique Thomas