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. 2021 Feb 10;3(2):257-266.
doi: 10.1016/j.xkme.2020.12.007. eCollection 2021 Mar-Apr.

Type IV Collagen Variants in CKD: Performance of Computational Predictions for Identifying Pathogenic Variants

Cole Shulman  1   2 , Emerald Liang  1   2 , Misato Kamura  3 , Khalil Udwan  1   2 , Tony Yao  1   2 , Daniel Cattran  1   2   4   5 , Heather Reich  1   2   4   5 , Michelle Hladunewich  1   2   4   5 , York Pei  1   2   4   5 , Judy Savige  6 , Andrew D Paterson  7   8 , Mary Ann Suico  3 , Hirofumi Kai  3 , Moumita Barua  1   2   4   5
Affiliations

Type IV Collagen Variants in CKD: Performance of Computational Predictions for Identifying Pathogenic Variants

Cole Shulman et al. Kidney Med. .

Abstract

Rationale & objective: Pathogenic variants in type IV collagen have been reported to account for a significant proportion of chronic kidney disease. Accordingly, genetic testing is increasingly used to diagnose kidney diseases, but testing also may reveal rare missense variants that are of uncertain clinical significance. To aid in interpretation, computational prediction (called in silico) programs may be used to predict whether a variant is clinically important. We evaluate the performance of in silico programs for COL4A3/A4/A5 variants.

Study design setting & participants: Rare missense variants in COL4A3/A4/A5 were identified in disease cohorts, including a local focal segmental glomerulosclerosis (FSGS) cohort and publicly available disease databases, in which they are categorized as pathogenic or benign based on clinical criteria.

Tests compared & outcomes: All rare missense variants identified in the 4 disease cohorts were subjected to in silico predictions using 12 different programs. Comparisons between the predictions were compared with: (1) variant classification (pathogenic or benign) in the cohorts and (2) functional characterization in a randomly selected smaller number (17) of pathogenic or uncertain significance variants obtained from the local FSGS cohort.

Results: In silico predictions correctly classified 75% to 97% of pathogenic and 57% to 100% of benign COL4A3/A4/A5 variants in public disease databases. The congruency of in silico predictions was similar for variants categorized as pathogenic and benign, with the exception of benign COL4A5 variants, in which disease effects were overestimated. By contrast, in silico predictions and functional characterization classified all 9 pathogenic COL4A3/A4/A5 variants correctly that were obtained from a local FSGS cohort. However, these programs also overestimated the effects of genomic variants of uncertain significance when compared with functional characterization. Each of the 12 in silico programs used yielded similar results.

Limitations: Overestimation of in silico program sensitivity given that they may have been used in the categorization of variants labeled as pathogenic in disease repositories.

Conclusions: Our results suggest that in silico predictions are sensitive but not specific to assign COL4A3/A4/A5 variant pathogenicity, with misclassification of benign variants and variants of uncertain significance. Thus, we do not recommend in silico programs but instead recommend pursuing more objective levels of evidence suggested by medical genetics guidelines.

Keywords: ARUP; Alport syndrome; ClinVar; FSGS; LOVD; computational predictions; genomics; gnomAD; in silico predictions; type IV collagen variants.

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Figures

Figure 1
Figure 1
Number of rare missense variants predicted to be pathogenic in: (A) the focal segmental glomerulosclerosis (FSGS) study cohort and (B) Genome Aggregation Database (gnomAD). For rare missense COL4A3, COL4A4, and COL4A5 variants in our FSGS cohort, 43% (6/14), 40% (4/10), and 33% (2/6) were predicted to be deleterious by at least 10 of 12 programs, respectively. For rare missense COL4A3, COL4A4, and COL4A5 variants identified in gnomAD, 35% (301/851), 32% (306/949), and 41% (197/483) were predicted to be deleterious by at least 10 of 12 programs, respectively.
Figure 2
Figure 2
Comparison of COL4A3, COL4A4, and COL4A5 in silico predictions with disease database categorization. For ARUP COL4A5 pathogenic variants, there was 97% (317/327) concordance with in silico predictions. For ClinVar COL4A3/A4/A5 pathogenic variants, there was 75% (12/16), 100% (9/9), and 89% (258/287) concordance with in silico predictions, respectively. For LOVD COL4A3/A4/A5 pathogenic variants, there was 82% (28/34), 86% (42/49), and 94% (650/699) concordance. Congruency of in silico predictions was similar for variants categorized as benign, with the exception of COL4A5 variants documented in ARUP and ClinVar, in which the effects were overestimated by in silico programs, though there were fewer variants to interrogate. In ARUP, 57% (4/7) of COL4A5 variants were classified correctly by in silico predictions. In ClinVar, 100% (6/6), 100% (9/9), and 71% (5/7) of COL4A3/A4/A5 variants, respectively, were correctly assigned. Finally, for LOVD, 100% (2/2), 85% (23/37), and 100% (6/6) of COL4A3/A4/A5 variants were correctly classified.
Figure 3
Figure 3
Spearman correlation coefficient heatmap comparing results of various prediction models. Most programs had similar prediction scores when comparing with each other except for FATHMM and M-CAP. Dark blue signifies a strong direct correlation while dark red signifies a strong indirect correlation. Squares that are lighter in color signify a weak correlation between the results of the 2 prediction models. Figure created using the corrplot package available in Rstudio.
Figure 4
Figure 4
Functional characterization of COL4A3 and COL4A5 using the split-luciferase assay. Scatterplots of the intracellular/secreted relative light unit (RLU) ratio from human embryonic kidney 293 (HEK293T) cells expressing (A, B) mutant α3 chain or (C, D) mutant α5 chain compared to wild type (WT) using N-terminal and C-terminal split-luciferase tagged constructs. Pathogenic α3 and α5 chain mutants showed clearer secretory defect with N-terminal tagged constructs. Solid line: Y = X, dotted line: Y = X + 50, Y = X – 50. Square, WT; red circles are pathogenic variants and grey circles are variants of uncertain significance. Any data point under the −50 line was considered as a significant secretory defect. Experiments were performed in triplicate. Data presented are representative of 2 independent experiments.
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References

    1. Klarenbach S.W., Tonelli M., Chui B., Manns B.J. Economic evaluation of dialysis therapies. Nat Rev Nephrol. 2014;10(11):644–652. - PubMed
    1. Canadian Institute for Health Information . Canadian Institute for Health Information; Ottawa, ON: 2015. National Health Expenditure Trends, 1975 to 2015.
    1. Yao T., Udwan K., John R. Integration of genetic testing and pathology for the diagnosis of adults with FSGS. Clin J Am Soc Nephrol. 2019;14:213–223. - PMC - PubMed
    1. Groopman E.E., Marasa M., Cameron-Christie S. Diagnostic utility of exome sequencing for kidney disease. N Engl J Med. 2019;380(2):142–151. - PMC - PubMed
    1. Kashtan C.E. Alport syndrome. An inherited disorder of renal, ocular, and cochlear basement membranes. Medicine (Baltimore) 1999;78(5):338–360. - PubMed

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