A Novel Leflunomide Analog, UTL-5b (GBL-5b), Suppresses JAK3, MAP3K2, and LITAF Genes

Abstract

UTL-5b (GBL-5b) is a novel analog of leflunomide with anti-inflammatory and antiarthritic effects. It has been shown to lower serum tumor necrosis factor-alpha (TNF-α) level induced by lipopolysaccharide (LPS) in an animal model. In this study, the effect of UTL-5b on nitric oxide (NO) and dihydroorotate dehydrogenase (DHODH) was investigated. Our in vitro studies showed that (1) UTL-5b is a stronger inhibitor of NO production as compared to leflunomide and its active metabolite, teriflunomide, and (2) Unlike leflunomide, a potent inhibitor of DHODH, UTL-5b does not inhibit DHODH activity. These findings show that UTL-5b acts in a manner different from that of leflunomide. To further investigate the mode of action of UTL-5b, an ex vivo gene array study was performed. C57BL/6 mice were injected subcutaneously with of UTL-5b 24 hr before injection of E. coli LPS. Mice were sacrificed 90 min later and the whole spleen mRNA was isolated for gene microarray analysis. The results showed that UTL-5b significantly suppressed three genes that are relevant to the TNF- pathway: Janus kinase 3 (JAK3), mitogen-activated protein kinase kinase kinase 2 (MAP3K2) and lipopolysaccharide-induced TNF- factor (LITAF). In summary, our results showed that UTL-5b has a stronger inhibitory effect on NO production than leflunomide; yet, unlike leflunomide, UTL-5b does not inhibit DHODH in vitro. In addition, gene array analysis showed that the biological effects of UTL-5b are attributed at least in part to the suppression of JAK3, MAP3K2 and LITAF gene expression.