Ability of luteinizing hormone releasing hormone-Pseudomonas aeruginosa exotoxin 40 binding to LHRH receptor on human liver cancer cells

AIM: TO explore the ability of recombinant toxin luteinizinghormone releasing hormone-Pseudomonas aeruginosaexotoxin 40 (LHRH-PE40) and LHRH binding to LHRH receptor(LHRHR) on the membrane surface of human liver cancerHEPG cells.METHODS: LHRH was labeled by using 12sI with enzymaticreaction. The affinity and receptor volume of LHP, H-PE40and LHRH binding to LHRHR on the membrane surface ofhuman liver cancer cells were measured with radioligandreceptor assay.RESULTS: The specific activity of LHRH labeled with 12sIwas 2.7x 104 kBq/l^L, and its radiochemical purity reachedto 99.2-99.7%. The binding of 12sI to LHRH was maximalfor 240 min in the warm cultivation, and this binding wasstabilized. The inhibiting rates of ~2SI-LHRH and LHRH onthe proliferation of human liver cancer HEPG cells werenot significantly different. On the basis of the saturationcurve of ~2SI-LHRH binding to the membrane LHRHR of HEPGcells, ~2SI-LHRH of lx10s cpm was selected for radioligandreceptor assay. The affinity constants (Kd) of LHRH-PE40and LHRH binding to the membrane LHRHR of HEPG cellswere 0.43~0.12 nmol/L and 4.86~ 1.47 nmol/L, respectively,and their receptor volumes were 0.37~0.15 l^mol/g and0.42=I=0.13 l^mol/g, respectively. The binding of LHRH-PE40to the membrane protein of normal liver cells was notobserved.CONCLUSION: The recombinant toxin LHRH-PE40 binding tothe membrane surface of LHRHR of human liver cancer HEPGcells was very strong, while the specific binding of it to normalliver cells was not observed. The results provide an importantexperimental basis for the clinical application of LHRH-PE.