Preparation of Potent Leptin Receptor Antagonists and Their Therapeutic Use in Mouse Models of Uremic Cachexia and Kidney Fibrosis.

Leptin antagonists (L39A/D40A/F41A mutants) of mouse, human, rat and ovine leptins were developed in our laboratory by rational mutagenesis, expressed in Escherichia coli, refolded and purified to homogeneity. Pegylation of these antagonists resulted in long-acting reagents suitable for in-vivo studies. Further selection of high-affinity leptin antagonists was achieved by random mutagenesis of the whole open reading frame followed by yeast-surface display; an additional mutation (D23L) increased their affinity toward leptin receptor 60-fold. This superactive pegylated mouse leptin antagonist (PLA) exhibited a strong orexigenic effect, leading, in 10-14 days, to a 40% increase in body weight resulting mainly from obesity; this was reversed once PLA treatment was ceased. Cachexia is common in patients with Chronic Kidney Disease (CKD). Our studies suggested that leptin mediates cachexia by decreasing food intake while increasing energy consumption in CKD mice. We showed that PLA ameliorates CKD-associated cachexia in mice. Leptin may also contribute to the development of muscle and renal fibrosis in CKD, serious complications associated with increased morbidity and mortality. Transforming growth factor (TGF)-beta signaling may be the most potent mediator of fibrogenesis in multiple organs, and leptin is a co-activator of TGF-beta. Muscle fibrosis was evident in our CKD mice and PLA treatment significantly reduced the mRNA levels of TGF-beta 1 and its downstream targets in their muscle and renal tissues. PLA may offer a novel therapeutic strategy for CKD-associated cachexia, muscle and renal fibrosis to improve CKD patients' survival and quality of life.