CDKL5 deficiency disorder (CDD) is an X-linked neurodevelopmental disorder caused by mutations in the CDKL5 gene. CDD is characterized by early-onset epilepsy, global developmental delay, intellectual disability, autistic features, visual impairment, and motor impairment. The CDKL5 gene is located on the X-chromosome and is highly expressed in the forebrain. Despite some molecular and cellular functions of CDKL5 being revealed, gaps and controversies still exist between different studies. The molecular actions of CDKL5 in human-derived neurons and the effects of CDKL5 disruption remain unclear. To fully understand the biological functions of CDKL5 and identify more reliable therapeutics, it is crucial to identify its bona fide substrates and dissect its function in clinically relevant cell types and models.The emergence of induced pluripotent stem cells (iPSCs) and the advancement of three-dimensional brain organoids have provided powerful tools to unravel human neurodevelopmental processes. This study aims to utilize cerebral organoids generated from patient-derived iPSCs as a model system to characterize early neurodevelopmental deficits in CDD. By employing single-cell RNA sequencing and immunostaining techniques, we have observed impaired neurogenesis and altered neuronal fate in CDD. Ongoing work involves using biochemical methods to dissect the downstream pathways involved. The findings from this study will enhance our understanding of CDKL5 function and propose a novel molecular mechanism for CDD pathology. Moreover, this study not only provides a robust model for studying X-linked neurodevelopmental disorders but also opens up new avenues for therapeutic development targeting neurodevelopmental disorders (NDDs).