The Effect And Mechanism Of New MTOR Inhibitor Y31 Retarding The Progression Of Rat Polycystic Kidney Disease

Background and objectiveAs a monogenic diseases, Autosomal dominant polycystic kidney disease(ADPKD) is the most common inherited kidney disease known. It has a very high incidence which is 1:400 to 1:1000. It thought currently Cyst initiation and expansion arise from abnormal cell proliferation, fluid secretion and extracellular, matrix defects and results in kidney enlargement and interstitial fibrosis after the gene mutation. The cysts constantly enlarge while compress the normal nephron structure, eventually leading to irreversible damage of the kidney structure and function. As report, by the age of 60 years, half of all ADPKD patients will go to End-stage renal disease(ESRD) and require dialysis or transplantation.The formation and growth of renal cysts in ADPKD patients are due to mutations in either the PKD1( 85%) or the PKD2( 15%) gene mutation, which encode polycystin-1(PC1) or polycystin-2(PC2) respectively. PC1 and PC2 constitute complexes by the interaction of the C-terminus and then act as cation channels to transfer cell signaling.The pathogenesis of ADPKD is not yet clear but lots of studies have investigated its pathogenesis including m TOR signal transduction pathway upregulated, vasopressin V2 receptor induced c AMP production,Cilia machinery- calcium influx disorders and excessive activation of the complement pathway and so on.At present, for the treatment of ADPKD drug research is mainly directed against the above mechanism, but there is still not a drug with definite therapeutic effect and less side effects. Therefore continuing to explore and develop an effective drug for ADPKD treatment is still a urgent medical problem.So we use the drug Y31, a new m TOR inhibitor to study ites role and possible mechanism. Our early study has shown that the pharmacokinetics and water-soluble of Y31 are better than traditional m TOR inhibitor. So Y31 might be new effective drugs for treatment of ADPKD.It is clear that m TOR can regulate the phosphorylation of stat1/3 and our previous study has proved that stat1/3 can activate CFB. So we will observed if m TOR inhibitor would influence the activation of the complement system in this study.As the m TOR signaling pathway networks are so complicated even involving the negative feedback path way that monotherapy could not the best choice. Maybe, combination therapy is the better approach. As it has been reported, Metformin inhibit renal cystogenesis by activating AMPK and suppressing m TOR and CFTR in Pkd1 mice. Besides, it also can suppress AKT, a key protein in m TOR negative feedback path way via activatintg phosphatase and tensin homolog deleted on chromosome ten(PTEN). Because of these reasons, we choose Metformin to combine with m TOR inhibitor to discuss the synergistic effects and possible mechanism.At last, we ues Han:SPRD rat model and Human cyst lining cells to study the possible mechanisms of Y31 in treating ADPKD. Also, this kind combination therapy is the fist attempt in the study of treatment in ADPKD. We are aiming at exploring new therapeutic drugs and treatment modalities for ADPKD through this study.Methods We use Han:SPRD rat model and Human cyst lining cells WT9-12 cell in this study. Evaluate the therapeutic effect of the drug through gavage administration and detecting renal function(Scr and BUN). It is through these to explore the mechanism of the drug by that Using western blotting method to detect the change of p-P70S6 K,which are the key protein in m TOR signal transduction pathway and the change of p-AKT expression, which are the key protein in m TOR negative feedback path way in Han:SPRD kidney specimens and WT9-12 cells; using western blotting method to detect variance of CFB and C3 degradation products expression in Han:SPRD kidney specimens and WT9-12 cells; Using tissue biopsy staining and MTT methods to observe the effects of drug on the growth of rat kidney cyst and detect the cell proliferation of WT9-12 cells administered by different drug concentrations of Y31 at different time points.ConclusionOur experimental results indicate that Y31 is an effective m TOR inhibitor. It can significantly improve the SCR and BUN, also it reduce two kidney/body weight ratio, cyst index and the Proliferation rate by indicating m TOR signal pathway. Moreover, we find high dosage Y31 also can inhibit the expresion of p-AKT expression, which are the key protein in m TOR negative feedback path way in Han:SPRD rat and WT9-12 cells. In the detection of the complement system component, both Rapamycin and Y31 can inhibit Complement system activity by reducing the expresion of CFB and the C3 degradation products including. This finding is the first in m TOR inhibitor treament of ADPKD. So we hope this study will give us a new understanding of the m TOR inhibitor in ADPKD treatment, especially the new m TOR inhibitor Y31.