Activity-dependent neuroprotective protein deficiency models synaptic and developmental phenotypes of autism-like syndrome
Gal Hacohen-Kleiman, Shlomo Sragovich, Gidon Karmon, Andy Y. L. Gao, Iris Grigg, Metsada Pasmanik-Chor, Albert Le, Vlasta Korenková, R. Anne McKinney, Illana Gozes
Gal Hacohen-Kleiman, Shlomo Sragovich, Gidon Karmon, Andy Y. L. Gao, Iris Grigg, Metsada Pasmanik-Chor, Albert Le, Vlasta Korenková, R. Anne McKinney, Illana Gozes
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Research Article

Activity-dependent neuroprotective protein deficiency models synaptic and developmental phenotypes of autism-like syndrome

Abstract

Previous findings showed that in mice, complete knockout of activity-dependent neuroprotective protein (ADNP) abolishes brain formation, while haploinsufficiency (Adnp+/–) causes cognitive impairments. We hypothesized that mutations in ADNP lead to a developmental/autistic syndrome in children. Indeed, recent phenotypic characterization of children harboring ADNP mutations (ADNP syndrome children) revealed global developmental delays and intellectual disabilities, including speech and motor dysfunctions. Mechanistically, ADNP includes a SIP motif embedded in the ADNP-derived snippet drug candidate NAP (NAPVSIPQ, also known as CP201), which binds to microtubule end–binding protein 3, essential for dendritic spine formation. Here, we established a unique neuronal membrane–tagged, GFP-expressing Adnp+/– mouse line allowing in vivo synaptic pathology quantification. We discovered that Adnp deficiency reduced dendritic spine density and altered synaptic gene expression, both of which were partly ameliorated by NAP treatment. Adnp+/–mice further exhibited global developmental delays, vocalization impediments, gait and motor dysfunctions, and social and object memory impairments, all of which were partially reversed by daily NAP administration (systemic/nasal). In conclusion, we have connected ADNP-related synaptic pathology to developmental and behavioral outcomes, establishing NAP in vivo target engagement and identifying potential biomarkers. Together, these studies pave a path toward the clinical development of NAP (CP201) for the treatment of ADNP syndrome.

Authors

Gal Hacohen-Kleiman, Shlomo Sragovich, Gidon Karmon, Andy Y. L. Gao, Iris Grigg, Metsada Pasmanik-Chor, Albert Le, Vlasta Korenková, R. Anne McKinney, Illana Gozes

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Figure 6

Adnp+/– pups exhibit significantly delayed growth as well as significantly impaired gait at 18 to 40 days of age, affected by NAP treatment, in a sex-dependent manner.

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Adnp+/– pups exhibit significantly delayed growth as well as significan...
(A and B) A 2-way, repeated-measures ANOVA with Bonferroni’s means separation test revealed significant differences in length between Adnp+/+ and Adnp+/– littermate mice (males: Adnp+/+ n = 18, Adnp+/– n = 12, Adnp+/– NAP, n = 12; females: Adnp+/+ n = 20, Adnp+/– n = 9, Adnp+/– NAP, n = 23). For males, main effects for group [F(1,28) = 24.025, P < 0.001], day [F(21,573) = 2122.663, P < 0.001], and interaction [F(21,573) = 3.703, P < 0.001] were found, with significant differences between Adnp+/+ and Adnp+/– mice (*P < 0.05 and **P < 0.01). For females, main effects for group [F(1,27) = 16.178, P < 0.001], day [F(21,554) = 1487.989, P < 0.001], and interaction [F(21,554) = 5.267, P < 0.001] were found, with significant differences between Adnp+/+ and Adnp+/– mice (*P < 0.05 and ***P < 0.001). NAP treatment did not affect length acquisition in male or female Adnp+/– pups. (C–E) For gait analysis, a 2-way ANOVA with Tukey’s post hoc test was performed. An unpaired Student’s t test was also used to determine sex differences (males: Adnp+/+ n = 104, Adnp+/– n = 72, Adnp+/– NAP, n = 64; females: Adnp+/+ n = 96, Adnp+/– n = 44, Adnp+/– NAP, n = 116 paw replicates per experimental group). For standing (seconds), main genotype [F(1,316) = 23.683, P < 0.001] and interaction [F(1,316) = 18.030, P <0.001] effects were found in males and main interaction effect in females (F(1,352) = 4.894, P < 0.05). For step cycles (seconds), main genotype [F(1,316) = 32.116, P < 0.001] and interaction [F(1,316) = 18.086, P < 0.001] effects were found in males. In females, a main interaction effect was found [F(1,352) = 4.974, P = 0.026]. For stride length (cm), main genotype [F(1,316) = 38.359, P <0.001] and treatment [F(1,316) = 6.152, P = 0.014] effects were found in males. In females, main genotype [F(1,352) = 21.286, P < 0.001], treatment [F(1,352) = 5.371, P = 0.021], and interaction [F(1,352) = 7.117, P = 0.008] effects were found. For standing, step cycle, and stride length parameters, significant differences between Adnp+/+ and Adnp+/– mice (***P < 0.001) and NAP- versus vehicle-treated Adnp+/– mice (***P < 0.001) were found. Sex differences were observed for all 3 gait parameters (**P < 0.01 and ***P < 0.001, Student’s t test). (A and B) Adnp+/+ data are reshown in Supplemental Figure 11, A and B; (C–E) Adnp+/+ data are also shown in Supplemental Figure 13, A–C.