Although NGLY1 deficiency has been discovered as a result of mutations in the NGLY1 gene, cellular and molecular mechanisms underlying the neurological abnormalities due to NGLY1 malfunction in the brain remain mostly unknown. Using human cerebral organoid (CO) models and systems biology techniques, we uncovered NGLY1 deficiencyinduced alterations at the early stage of cerebral development. Despite the similar vitality and cellular pluripotency of NGLY1-functional and -deficient WA09 hESCs, COs developed from the NGLY1-deficient hESCs had the defective formation of SATB2+ upper-layer neurons and attenuation of STAT3 and HES1 signaling critical for sustaining radial glia. The NGLY1-deficient CO cells, compared with the NGLY1-functional ones, also presented higher vulnerability to multiple stressors. Bulk and single-cell analysis of transcriptomes revealed that NGLY1-deficient COs showed a propensity for premature neuronal differentiation, accompanied by significant downregulation of secretory and transcription factors, including TTR, IGFBP2, and ID4. Supplementing recombinant TTR to NGLY1-deficient CO cells reduced their susceptibility to proteasome inhibition. Ectopic expression of NGLY1 led to IGFBP2 and ID4 upregulation in CO cells developed from NGLY-deficient patientderived induced pluripotent stem cells (iPSCs). Moreover, ID4 expression, STAT3 signaling, and proliferation were enhanced by treatment of recombinant IGFBP2 in CO cells developed from the NGLY1-deficient WA09 hESCs and patient-derived iPSCs. Our findings indicate that NGLY1 could be critical for regulating various stress responses and maintaining neural stem cells (NSCs) in the developing cerebrum. In patients, NGLY1 deficiencyassociated neurological abnormalities, including microcephaly, may be a consequence of aberrations in NSC signaling sustained by NGLY1.