Fig. 5. Cryptic HDGFL2 is elevated in CSF of sporadic ALS and of C9orf72 mutation carriers, including in the presymptomatic stage.

a, Cryptic HDGFL2 CSF:diluent MSD signal ratios measured in CSF of healthy controls (n = 16), migraine controls (n = 25), NPH controls (n = 25), presymptomatic (n = 81) and symptomatic (n = 76) C9orf72 mutation carriers, and sporadic ALS (n = 44). Values and statistics are shown in Extended Data Table 1. Ratios >1 indicate elevated signal. Data presented as mean ± s.d. Data points represent individuals with longitudinal CSF measurements averaged as applicable. Mann–Whitney U-test with Holm–Bonferroni correction, *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001. b, Cryptic HDGFL2 concentrations are negatively correlated with disease duration in NINDS symptomatic C9orf72 mutation carriers (Spearman, two-tailed, r = −0.45, P = 0.040), suggesting that cryptic HDGFL2 levels tend to be higher earlier in disease. c,d, CSF NfL (c) and pNfH (d) concentrations are positively correlated with disease duration during the first 25 months of symptomatic disease (Pearson, two-tailed, r = 0.58, P = 0.012 (c); r = 0.60, P = 0.006 (d)) and negatively correlated with disease duration after 25 months (Pearson, two-tailed, r = −0.59, P = 0.0003 (c); r = −0.66, P = 1.7 × 10⁻⁵ (d). e–g, Change in CSF cryptic HDGFL2 (e), NfL (f) and pNfH (g) levels (ng ml⁻¹) in presymptomatic C9orf72 mutation carriers across age (left) and in symptomatic C9orf72 mutation carriers across disease progression (right). Each line represents one individual. h, Proposed ALS staging model (adapted from Benatar et al.²⁴) based on the dynamic of NF subunit and cryptic HDGFL2 accumulation in CSF of C9orf72 mutation carriers. While CSF NF levels rise during the prodromal phase and continue increasing during the first few years of symptomatic disease, CSF cryptic HDGFL2 may peak before symptom onset and decrease during symptomatic disease progression. Due to these temporal differences, we propose a staging scale whereby the ratio of CSF NfL:cryptic HDGFL2 or pNfH:cryptic HDGFL2 concentrations would be <1 during the presymptomatic stage of ALS–FTD and increase to >1 during symptomatic disease. i, Among presymptomatic C9orf72 mutation carriers with detectable cryptic HDGFL2 levels and available NF measurements (n = 8), eight of eight (100%) individuals had NfL:cryptic HDGFL2 and pNfH:cryptic HDGFL2 ratios <1. Of the symptomatic C9orf72 mutation carriers with detectable cryptic HDGFL2 levels and available NfL (n = 13) or pNfH (n = 14) measurements, nine of 13 (69.2%) and ten of 14 (71.4%) individuals had NfL:cryptic HDGFL2 and pNfH:cryptic HDGFL2 ratios >1, respectively. Data presented as mean ± s.d.