A Targeted, Low-Throughput Compound Screen in a Drosophila Model of Neurofibromatosis Type 1 Identifies Simvastatin and BMS-204352 as Potential Therapies for Autism Spectrum Disorder (ASD)

Knock-down of Nf1 in the embryo is sufficient to induce synaptic transmission deficits in the third instar larval stage. Full genotypes for those abbreviated in the figure (i.e., GFP^RNAi and Nf1^RNAi) are explained in the legend for Figure 1. A–F, Raising Nf1^RNAi larvae at 30°C, sufficient to ensure constitutive knock-down of Nf1, mimics the Nf1^−/− larval phenotype (Dyson et al., 2022), in that it does not affect EJP amplitude (p = 0.5334) but increases mEJP frequency and amplitude, and reduces quantal content. G–L, Knock-down of Nf1 within the embryonic period has a similar effect on synaptic transmission as constitutive knock-down on all parameters examined, including no significant impact on (G) EJP amplitude (p = 0.8967). M–R, In contrast, knock-down of Nf1 only during larval stages has no effect on (M) EJP amplitude (p = 0.3010), (N) mEJP frequency (p = 0.3230), (O) mEJP amplitude (p = 0.3694), or (P) quantal content (p = 0.9998). Data in each panel were analyzed via unpaired, two-tailed Student’s t test. For each experiment, n = 13. Data are presented as mean ± SEM.

A targeted pharmacological screen identifies simvastatin and BMS-204352 to improve tactile hypersensitivity in Nf1^P1 larvae. Twenty compounds were screened for their ability to reduce the number of Nf1^P1 larvae responding to a mechanical stimulus, with three proving toxic at the concentration (50 μm) tested. Only administration of simvastatin (50% of vehicle-treated Nf1^P1) and BMS-204352 (61.1% of vehicle-treated Nf1^P1) resulted in a significant decrease in the number of responding larvae. Data are presented as a percentage of the number of Nf1^P1 larvae raised on an equivalent concentration of DMSO (vehicle) that responded to the stimulus (dotted line = 100%). Statistical comparisons were conducted on raw data using Fisher’s exact test between compound-treated and vehicle-treated Nf1^P1 larvae.

Simvastatin and BMS-204352 consistently improve, but do not fully rescue, tactile hypersensitivity in Nf1^P1 larvae, while having no effect on overall activity. A, Across four independent trials, 50 μm Simvastatin significantly reduces the mean percentage responding larvae, as does (B) 50 μm BMS-204352. C, The presence of simvastatin solely during the embryo has no impact on the percentage of Nf1^P1 larvae responding to stimulation (p = 0.3026). D, Conversely, exposing only larval progeny to simvastatin results in a significant reduction in the mean percentage of responding Nf1^P1 larvae. E, The presence of BMS-204352 solely during the embryo results in a significant reduction in the percentage of Nf1^P1 larvae responding to stimulation. F, The percentage of responding Nf1^P1 larvae is also significantly reduced when only larval progeny are exposed to BMS-204352. G, K33 and Nf1^P1 larvae do not significantly differ in their total distance traveled over a 3-min period at room temperature, nor is this impacted by treatment with 50 μm simvastatin (p = 0.9518). H, 50 μm BMS-204352 treatment also does not alter distance traveled in Nf1^P1 larvae, which again show no significant difference in crawling behavior compared with K33 controls (p = 0.1881). In panels A–F, each data point represents the percentage responding larvae from a single trial, with n = 20 per trial, such that n = 80 larvae overall. Data in these panels were analyzed via unpaired, two-tailed Student’s t test. Comparisons were made between individual trials, such that n = 4 per group. Data in panels G, H were analyzed via one-way ANOVA followed by Tukey’s post hoc test. All data are presented as mean ± SEM.