Fig. 2: Generation of sAD iCOs and their pathological features.

a Generation of iCOs; 60 days-old iCOs were used for the experiments. b Experimental iCOs’ sets for this study; The iCOs were from both Pittsburgh compound B (PiB)-PET negative/positive participants’ iPSCs and ApoE3 (parental) /E4 (isogenic) iPSCs, which were generated by CRISPR-Cas9 gene-editing. c Quantification of AD hallmark proteins (Aβ1-42, Aβ1-40, total tau, p-tau) in the organoid conditioned media was performed between PiB⁻ iCOs and PiB⁺ iCOs (***p = 0.0003 for Aβ1-42, ****p < 0.0001 for Aβ1-40, *p = 0.0384 for Aβ1-42/1-40 ratio, **p = 0.0042 for p-tau, ***p = 0.0002 for t-tau, p = 0.1025 for p/t-tau ratio; two-sided p-values; unpaired t-test; Each 3–7 iCOs was used from n = 8 participants) or between non-E4 carriers and E4 carriers (**p = 0.0020 for Aβ1-42, **p = 0.0082 for Aβ1-40, *p = 0.0370 for Aβ1-42/1-40 ratio, *p = 0.0125 for p-tau, *p = 0.0173 for t-tau, p = 0.3296 for p/t-tau ratio; two-sided p-values; unpaired t-test; Each 3–7 iCOs was used from n = 4 participants). d Significant correlation was shown between the real human cerebral amyloid deposition (SUVR) and secreted AD hallmark proteins (*p = 0.0190 for Aβ1-42, ****p < 0.0001 for Aβ1-40, *p = 0.0183 for Aβ1-42/1-40 ratio, *p = 0.0407 for p-tau, **p = 0.0042 for t-tau, ^#p = 0.0550 for p/t-tau ratio; Pearson’s correlation; isotonic regression curve was shown). e Quantification of AD hallmark proteins (Aβ1-42, Aβ1-40, total tau, p-tau) in the organoid conditioned media was performed between E4^iso iCOs and E3^par iCOs (**p = 0.0013 for Aβ1-42, **p = 0.0034 for Aβ1-40, ***p = 0.0007 for Aβ1-42/1-40 ratio, **p = 0.0094 for p-tau, p = 0.1720 for t-tau, ***p = 0.0003 for p/t-tau ratio; two-sided p-values; unpaired t-test; Each 24 iCOs was used from E4^iso iCOs and E3^par iCOs). f, g Comparison of physiological responses of the iCOs (calcium oscillation analysis) was performed between PiB⁻ iCOs and PiB⁺ iCOs or E4^iso iCOs and E3^par iCOs. PiB⁺ iCOs and E4^iso iCOs had more number of peaks than PiB⁻ iCOs and E3^par iCOs (^*p = 0.0399 for PiB⁻ iCOs vs PiB⁺ iCOs, *p =  0.0254 for E4^iso iCOs vs E3^par iCOs; two-sided p-values; unpaired t-test). h Principal component analysis (PCA) plot showing transcriptomic expression patterns in RNA sequencing data. i Transcriptomic GO analyses between the PiB⁻ iCOs and PiB⁺ iCOs or E4^iso iCOs and E3^par iCOs were performed with the FDR-adjusted p-value < 0.05 (adjustments were made for multiple comparisons; FDR-corrected by Toppgene analysis). p-value criteria: *p < 0.05, **p < 0.01, ***p < 0.001, and ****p < 0.0001, two-sided p-values, unpaired t-test. PBMC peripheral blood mononuclear cells, SeV Sendai virus, EB embryoid body, Dor dorsomorphin, SB SB431542, NB neurobasal media, PiB Pittsburgh compound B, CM culture media, sAD sporadic AD, SUVR standardized uptake value ratio, Aβ beta-amyloid, p-tau phosphorylated tau, MF molecular function, BP biological process, CC cellular component.