Loss of angiogenin function is related to earlier ALS onset and a paradoxical increase in ALS duration

Abstract

0.5-1% of ALS (Amyotrophic Lateral Sclerosis) and Parkinson's disease (PD) are associated with mutations in the angiogenin (ANG). These mutations are thought to cause disease through a loss of ANG function, but this hypothesis has not been evaluated statistically. In addition, the potential for ANG to promote disease has not been considered. With the goal of better defining the etiology of ANG-ALS, we assembled all clinical onset and disease duration data and determined if these were correlated with biochemical properties of ANG variants. Loss of ANG stability and ribonuclease activity were found to correlate with early ALS onset, confirming an aspect of the prevailing model of ANG-ALS. Conversely, loss of ANG stability and ribonuclease activity correlated with longer survival following diagnosis, which is inconsistent with the prevailing model. These results indicate that functional ANG appears to decrease the risk of developing ALS but exacerbate ALS once in progress. These findings are rationalized in terms of studies demonstrating that distinct mechanisms contribute to ALS onset and progression and propose that ANG replacement or stabilization would benefit pre-symptomatic ANG-ALS patients. However, this study challenges the prevailing hypothesis that augmenting ANG will benefit symptomatic ANG-ALS patients. Instead, our results suggest that silencing of ANG activity may be beneficial for symptomatic ALS patients. This study will serve as a call-to-arms for neurologists to consistently publish ALS and PD patient's clinical data-if all ANG-ALS patients' data were available our findings could be tested with considerable statistical power.

Figure 1

Lack of correlation between ANG stability and ANG ribonuclease activity and ANG aggregation propensity. Spearman’s Coefficient and Kendall Tau’s coefficient were used for analyzing correlation. A significance level of 0.05 was used. (a) Scatter plot demonstrating no significant correlation between ANG stability and ANG ribonuclease activity (b) Scatterplot demonstrating no significant correlation between ANG ribonuclease activity and Aggregation propensity (c) Scatterplot showing no significant correlation between ANG stability and aggregation propensity.

Figure 2

Destabilization of ANG variants correlates with faster ALS onset. Spearman’s Coefficient (p value 0.010) and Kendall Tau’s coefficient (p value 0.014) were used for analyzing correlation. Kaplan-Meier survival analysis was performed and the statistical significance of differences in survival between the categories was evaluated using Log-rank (p value 0.054), Breslow (p value 0.015), and Tarone-ware (p value 0.027) tests. A significance level of 0.05 was used. (a) Scatter plot demonstrating correlation between thermal destabilization and ALS onset. Sizes of points are proportional to number of cases. (b) Kaplan-Meier curves illustrating significant differences in ALS onset between patients with ANG variants with ∆∆G less than or equal to −1 kcal/mol and variants with ∆∆G greater than −1 kcal/mol.

Figure 3

Loss of ribonuclease activity of ANG variants does not correlate with ALS onset. Spearman’s Coefficient (p value 0.340) and Kendall Tau’s coefficient (p value 0.294) were used for analyzing correlation. Kaplan-Meier survival analysis was performed and the statistical significance of differences in survival between the categories was evaluated using Log-rank (p value 0.235), Breslow (p value 0.119), and Tarone-ware (p value 0.173) tests. A significance level of 0.05 was used. (a) Scatter plot demonstrating no significant correlation between ribonuclease activity and ALS onset. Sizes of points are proportional to number of cases. (b) Kaplan-Meier curves illustrating no significant differences in ALS onset between patients with ANG variants with %WT ribonuclease activity less than or equal to 10% and variants with %WT ribonuclease activity greater than −10%.

Figure 4

Destabilization of ANG variants correlates with longer ALS duration. Spearman’s Coefficient (p value 0.016) and Kendall Tau’s coefficient (p value 0.021) were used for analyzing correlation. Kaplan-Meier survival analysis was performed and the statistical significance of differences in survival between the categories was evaluated using Log-rank (p value 0.072), Breslow (p value 0.086), and Tarone-ware (p value 0.077) tests. A significance level of 0.05 was used. (a) Scatter plot representing correlation between thermal destabilization and disease duration. Sizes of points are proportional to number of cases. (b) Kaplan-Meier curves illustrating differences in ALS onset between patients with ANG variants with ∆∆G less than or equal to −1 kcal/mol and variants with ∆∆G greater than −1 kcal/mol.

Figure 5

Loss of ANG ribonuclease activity correlates with longer ALS duration. Spearman’s Coefficient (p value 0.002) and Kendall Tau’s coefficient (p value 0.005) were used for analyzing correlation. Kaplan-Meier survival analysis is performed and the statistical significance of differences in survival between the categories was evaluated using Log-rank (p value 0.002), Breslow (p value 0.01), and Tarone-ware (p value 0.005) tests. (a) Scatter plot demonstrating significant correlation between ribonuclease activity and ALS onset. Sizes of points are proportional to number of cases. (b) Kaplan-Meier curves illustrating significant differences in disease duration between patients with ANG variants with %WT ribonuclease activity less than or equal to 10% and variants with %WT ribonuclease activity greater than −10%.

Figure 6

Intraperitoneal treatment of tgSOD1^G93A-ALS mice with 10 μg recombinant huANG post disease onset did not prolong survival. (a) Kaplan-Meier curves illustrated no significant differences in survival between tgSOD1^G93A-ALS mice treated with vehicle and tgSOD1G93A-ALS mice treated with 10 μg recombinant hANG (i.p., 3 times/week). Kaplan-Meier survival analysis was performed and the statistical significance of differences in survival between the categories was evaluated using Log-rank (p value 0.426), Breslow (p value 0.702), and Tarone-ware (p value 0.55) tests. A significance level of 0.05 was used.