In neuromuscular illnesses, the intricacy of RNA metabolism has become essential, particularly with regard to spinal muscular atrophy (SMA). In fact, RNA metabolism involves survival motor neuron protein (SMN) in small nuclear ribonucleoprotein synthesis, alternative splicing, trafficking of RNA-binding proteins, and translation of mRNAs in neurites. Biologically active SMN is restored by Nusinersen, an antisense oligonucleotide that modifies the SMN2 gene’s pre-mRNA splicing. In this work we aimed to evaluate the transcriptomic signature in cerebrospinal fluid (CSF) of three adult SMA patients before and after Nusinersen treatment at different time points. We collected RNA from CSF of adult SMA patients (n=3) at baseline (T0), after 6 months of Nusinersen therapy, at fifth injection (T4) and proportionally at T6, T8, T10, T13 and T16 injection. Differentially expressed (DE) cell-free RNAs (cfRNAs) were examined by RNA sequencing, with all the injection’ time points being compared to T0. We showed a maximum peak of DE genes (protein coding) between T4 and T6, which progressively decreases reaching an expression plateau at later time points. Enriched DE pathways are related to inflammation (HSP90AB), ribosomal physiology, and actin or myosin filaments. The specific molecular response was transferable on a clinical level, since the trend in gene deregulation reflects the modulation of the clinical phenotype of patients who, after an initial improvement tend to stabilize their pathological condition. This study provides preliminary insights into modulation of gene expression dependent on Nusinersen treatment and lays the foundation for prognostic biomarkers discovery.