The Experimental Study On The Effect Of Epidermal Growth Factor Receptor Tyrosine Kinase Inhibitor On Polycystic Kidney Disease

Autosomal dominant polycystic kidney disease is one of the most common human fatal hereditary renal diseases with a morbidity of 1 %o~2 %o. The disease is characteristic by innumerable fluid-filled cysts in both kidneys and other organs, eventually resulting in renal failure. There is no sensitive method to monitor the progression of the disease or effective medicine to treat it. It is reported that $6 billion was used annually for treating and nursing ADPKD patients in America. Therefore, research and development of drugs for ADPKD is of great significance and value to science, society and economy.Further understanding of the pathogenesis of ADPKD suggests new directions for treatment. Blocking the crucial pathological process of the disease becomes a viable treatment for ADPKD. As is the case with cancer, various growth factors are involved in ADPKD, Enlightened by anticancer therapies, researchers wondered wether anticancer drugs could be used for the treatment of PKD. In 2000, this hypothesis was proven by experiments in which the epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI) was shown to have a therapeutic effect on mice with autosomal recessive polycystic kidney disease (ARPKD).In order to conform the effect of EGFR tyrosine kinase inhibitor on ADPKD, our research group began the study in 2000, trying to develop the first drug for ADPKD. The study consists of four parts. 1. Role of EGF/EGFR on autosomal dominant polycystic kidney diseaseTo investigate efficiency and safety of EGFR-TKI for ADPKD, firstly we planned to use human cyst-lining epithelia cell, Han:SPRD rat of the ADPKD model and ADPKD patients to testify the role of EGF/EGFR in the development of the disease. By in situ hybridization and immunohistochemistry, we detected distribution of EGF and EGFR in the kidneys of ADPKD rats and patients. The EGFR and phosphorylated EGFR in cells and kidneys were determined by Western blot. The quantitative information of EGF in the culture medium of cyst-lining epithelia cells and bodyfluids of ADPKD rats and patients were assayed by ELISA. Cellular proliferation stimulated by rhEGF and cyst fluids was tested by MTT, and relative signal transduction mechanisms were analyzed. The studies at cell and tissue levels revealed quality and quantity abnormalities of EGF and EGFR. As is the case with normal condition, EGF was located on the apical membrane of distal tubules of the kidneys. Although EGF was down regulated in the kidneys (PO.05), it can be secreted by cystic cells. As a result, there was EGF with mitotic concentration (103.56 pg/ml) in the cyst fluids from ADPKD patients; EGFR in ADPKD rats and patients was obviously overexpressed (/><0.05), making the cells more sensitive to rhEGF. In addition, EGFR was also mis-located on the apical (luminal) surface of cyst-lining epithelia. When the luminal-located EGFR bound to the ligands, the tyrosine kinase was activated by an autocrine-paracrine stimulation. After being self-phosphorylated, EGFR transferred the signals to the nucleus, the nuclear factor c-fos, c-jun were expressed, resulting in the uncontrollable proliferation of cells and growth of cysts (PO.001). Our study demonstrates the important role for EGF/ EGFR in ADPKD. Therefore, the abnormal EGFR could be an effective target of therapy.2. Molecular docking and mechanisms of EGFR-TKIBased on the pathogenetic role of abnormal EGFR, we planned to use small molecule TKI treating PKD. Asa novel anticancer drug, TKIs have become a big family with various structures and mechanisms. By a computer-aided drug design approach, we docked some TKIs reported in the literature into EGFR tyrosine kinase. AG was compared with experimental data KD. To explore the inhibitory mechanism, we investigated the interaction between inhibitors and EGFR by LIGPLOT. The results showed that AG was highly correlate with KD (r2 =0.925), which proved the reliability of docking model. The study on the structure activity relationship of TKI gave a clue to the further structure modification. With...