Efficacy of Hydroxy-L-proline (HYP) analogs in the treatment of primary hyperoxaluria in Drosophila Melanogaster

Huan Yang,Musa Male,Juncheng Hu,Shan Jin,Yang Li,Ning Wang,Hua Xu,Zhiqiang Chen,Zhangqun Ye,Chenming Zhao

doi:10.1186/s12882-018-0980-8

Huan Yang, Musa Male + Show 8 more

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https://doi.org/10.1186/s12882-018-0980-8

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Abstract

BackgroundSubstrate reduction therapy with analogs reduces the accumulation of substrates by inhibiting the metabolic pathways involved in their biosynthesis, providing new treatment options for patients with primary hyperoxalurias (PHs) that often progress to end-stage renal disease (ESRD). This research aims to evaluate the inhibition efficacy of Hydroxy-L-proline (HYP) analogs against calcium oxalate (CaOx) crystal formation in the Drosophila Melanogaster (D. Melanogaster) by comparing them with Pyridoxine (Vitamin B6).MethodsThree stocks of Drosophila Melanogaster (W118, CG3926 RNAi, and Act5C-GAL4/CyO) were utilized. Two stocks (CG3926 RNAi and Act5C-GAL4 /CyO) were crossed to generate the Act5C > dAGXT RNAi recombinant line (F1 generation) of D. Melanogaster which was used to compare the efficacy of Hydroxy-L-proline (HYP) analogs inhibiting CaOx crystal formation with Vitamin B6 as the traditional therapy for primary hyperoxaluria.ResultsNephrolithiasis model was successfully constructed by downregulating the function of the dAGXT gene in D. Melanogaster (P-Value = 0.0045). Furthermore, the efficacy of Hydroxy-L-proline (HYP) analogs against CaOx crystal formation was demonstrated in vivo using D. Melanogaster model; the results showed that these L-Proline analogs were better in inhibiting stone formation at very low concentrations than Vitamin B6 (IC50 = 0.6 and 1.8% for standard and dietary salt growth medium respectively) compared to N-acetyl-L-Hydroxyproline (IC50 = 0.1% for both standard and dietary salt growth medium) and Baclofen (IC50 = 0.06 and 0.1% for standard and dietary salt growth medium respectively). Analysis of variance (ANOVA) also showed that Hydroxy-L-proline (HYP) analogs were better alternatives for CaOx inhibition at very low concentration especially when both genetics and environmental factors are intertwined (p < 0.0008) for the dietary salt growth medium and (P < 0.063) for standard growth medium.ConclusionAddition of Hydroxy-L-Proline analogs to growth medium resulted in the reduction of CaOx crystals formation. These analogs show promise as potential inhibitors for oxalate reduction in Primary Hyperoxaluria.

Highlights

Substrate reduction therapy with analogs reduces the accumulation of substrates by inhibiting the metabolic pathways involved in their biosynthesis, providing new treatment options for patients with primary hyperoxalurias (PHs) that often progress to end-stage renal disease (ESRD)
Melanogaster of W1118 (#3605, Bloomington, USA), Actin-GAL4/CyO (#4414, Bloomington, USA) from Bloomington Drosophila Stock Center [https:// bdsc.indiana.edu/] and CG3926 RNAi (#TH02225.N, Beijing) from Tsinghua Fly Center [http://fly.redbux.cn/ rnai.php?lang=en] were used in these experiments. These flies were either fed with standard growth medium in this study to evaluate the relationship of dAGXT gene and calcium oxalate (CaOx) crystal formation or with dietary salt growth medium containing 0.05% Sodium oxalate [http://en.reagent.com.cn/ enshowproduct.jsp?id=10020118] which was used to investigate the exacerbation effects of extrinsic factors on the genetics as well as the efficacy of the inhibitors
We demonstrated the Drosophila model for urolithiasis utilizing knocking-down of the dAGXT gene implicated in the formation of stone effectuating with adult D

Summary

Introduction

Substrate reduction therapy with analogs reduces the accumulation of substrates by inhibiting the metabolic pathways involved in their biosynthesis, providing new treatment options for patients with primary hyperoxalurias (PHs) that often progress to end-stage renal disease (ESRD). This research aims to evaluate the inhibition efficacy of Hydroxy-L-proline (HYP) analogs against calcium oxalate (CaOx) crystal formation in the Drosophila Melanogaster Methods: Three stocks of Drosophila Melanogaster (W118, CG3926 RNAi, and Act5C-GAL4/CyO) were utilized. Melanogaster which was used to compare the efficacy of Hydroxy-L-proline (HYP) analogs inhibiting CaOx crystal formation with Vitamin B6 as the traditional therapy for primary hyperoxaluria. Calcium oxalate (CaOx) is the primary component of nephrolithiasis which accounts for about 80% of all stones [1,2,3,4,5]. Many factors can induce nephrolithiasis, and these encompass acidic urinary pH, dehydration, hypercalciuria, particular medications, hyperoxaluria and hereditary disorders. Among these factors, hyperoxaluria is the gravest. Hyperoxaluria is either acquired or inherited [6, 7]

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