Splice modulation of COL4A5 reinstates collagen IV assembly in an organoid model of Alport syndrome
Hassan Saei, Bruno Estebe, Nicolas Goudin, Mahsa Esmailpour, Julie Haure, Olivier Gribouval, Christelle Arrondel, Vincent Moriniere, Pinyuan Tian, Rachel Lennon, Corinne Antignac, Geraldine Mollet, Guillaume Dorval
Hassan Saei, Bruno Estebe, Nicolas Goudin, Mahsa Esmailpour, Julie Haure, Olivier Gribouval, Christelle Arrondel, Vincent Moriniere, Pinyuan Tian, Rachel Lennon, Corinne Antignac, Geraldine Mollet, Guillaume Dorval
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Research Article Genetics Nephrology

Splice modulation of COL4A5 reinstates collagen IV assembly in an organoid model of Alport syndrome

Abstract

Kidney organoids are an emerging tool for disease modeling, especially genetic diseases. Among these diseases, X-linked Alport syndrome (XLAS) is a hematuric nephropathy affecting the glomerular basement membrane (GBM) secondary to pathogenic variations in the COL4A5 gene encoding the α5 subunit of type IV collagen [α5(IV)]. In patients carrying pathogenic variations affecting splicing, the use of antisense oligonucleotides (ASOs) offers immense therapeutic hope. In this study, we develop a framework combining the use of patient-derived cells and kidney organoids to provide evidence of the therapeutic efficacy of ASOs in XLAS patients. Using multiomics analysis, we describe the development of GBM in WT and mutated human kidney organoids. We show that GBM maturation is a dynamic process, which requires long organoid culture. Then, using semi-automated quantification of α5(IV) at basement membranes in organoids carrying the splicing variants identified in patients, we demonstrate the efficacy of ASO treatment for α5(IV) restoration. These data contribute to our understanding of the development of GBM in kidney organoids and pave the way for a therapeutic screening platform for patients.

Authors

Hassan Saei, Bruno Estebe, Nicolas Goudin, Mahsa Esmailpour, Julie Haure, Olivier Gribouval, Christelle Arrondel, Vincent Moriniere, Pinyuan Tian, Rachel Lennon, Corinne Antignac, Geraldine Mollet, Guillaume Dorval

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Figure 5

Analysis of BM defects in XLAS.

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Analysis of BM defects in XLAS. (A) Immunofluorescence staining for coll...
(A) Immunofluorescence staining for collagen α3(IV), α4(IV), α5(IV), and α6(IV) chains in organoids (day 38), showing the absence of type IV collagen networks (both α5α5α6 and α3α4α5) in the severe XLAS model and substantial decrease in the moderate XLAS model. White arrows point to the GBM. Scale bars: 100 μm. (B–D) Quantification of collagen α5(IV) mean fluorescence intensities within GBM (B), PEC-BM (C), and TBM (D) in the severe, moderate, and control organoids (n = 60 objects in each group; each object is either tubule or glomerulus) (values below each condition indicate the mean intensity, 1-way ANOVA with Tukey’s multiple-comparison test). No collagen α5(IV) secretion was observed in the XLAS severe models, and significant reduction in collagen α5(IV) deposition was observed in all BMs in the moderate model.