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Heterogeneous epigenetic changes affect DNA repair system, leading to heterogeneous tumor development patterns. See also Supplementary Fig. 5. See also Supplementary Data 4. a GSEA analysis using normalized promoter methylation score of PDOs. The KEGG signaling pathway pathways are categorized and indicated with representative colors. Tumor-related pathways (n = 14) were significantly dysregulated compared to the normal mucosa. b Differentially methylated genes separating C and F-TO from the rest (standard deviation < 0.1). Hypermethylated promoter regions (n = 63) and hypomethylated promoter sites (n = 54) were identified (red, hypermethylated; blue, hypomethylated). c The sigmoid curves of PDOs treated with two different types of methyltransferase inhibitors. Each PDO is marked with representative colors. The error bars represent the standard deviations (n = 3).
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Multifocal colorectal cancer (CRC) comprises both clonally independent primary tumors caused by inherited predisposition and clonally related tumors mainly due to intraluminal spreading along an intact basement membrane. The distinction between these multifocal CRCs is essential because therapeutic strategies vary according to the clonal associatio...
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... used the normalized value of transcripts (n = 35,993) in the original tumors (AC-FC), which are log2 transformed values of tumor TPM/normal mucosa TPM. With respect to transcriptomic patterns of synchronous tumors, 85.12% of transcriptomes accounted for scattered patterns of principle component 1 (PC1) and principle component (PC2) after removing the lower 10% of variables based on variance ( Supplementary Fig. 4a). This suggested that each multifocal tumor distinctively regulates gene expressions in synchronous intestinal cancer. ...
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... further investigated that the derived models also retained expression heterogeneity identified in synchronous tumors. To discover the overlaps of transcriptomes that affect intertumoral heterogeneity between tumors, we collected the top 10% of genes that contribute to PC1 of the original tumors and their derivatives including PDOs, PDOCs, and PDCs, respectively ( Supplementary Fig. 4a, b, Supplementary Data 3b). Although transcriptomes of cancer cell lines and organoids are likely affected by varying composition of culture media and cell density 24 , contributing components (genes) that accounted for heterogeneous patterns of the original tumor were intersected more than 57% of their derivatives. ...
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... clearly separated PDOs from tissue samples as expected based on the results of the prior hierarchical clustering analysis. In parallel Supplementary Fig. 4 and Supplementary Data 3. a Gene Set Enrichment Assay (GSEA) analysis using normalized transcripts of all genes (n = 35,993) of tumor tissues and corresponding derivatives compared to the normal tissue. The KEGG signaling pathway pathways are categorized and indicated with representative colors. ...
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... compared to normal mucosa, which revealed that 14 tumor-related pathways were significantly dysregulated. Although pathway enrichment scores across six different PDOs were mostly analogous to each other, we demonstrated that aberrations with cell proliferation, communication, and genetic process pathways accounted for the onset of this SIC case ( Fig. 4a; Supplementary Data 4a). "Focal_Adhesion" pathway exhibited an organ-specific enrichment score, in which it was specifically depleted in PDOs derived from the small bowel (E-TO and F-TO) compared to the rest (Supplementary Data ...
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... pathway enrichment scores across six different PDOs were mostly analogous to each other, we demonstrated that aberrations with cell proliferation, communication, and genetic process pathways accounted for the onset of this SIC case ( Fig. 4a; Supplementary Data 4a). "Focal_Adhesion" pathway exhibited an organ-specific enrichment score, in which it was specifically depleted in PDOs derived from the small bowel (E-TO and F-TO) compared to the rest (Supplementary Data 4a). ...
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... selected the specific regions in which the standard deviation (SD) of methylation scores (m) between C and F organoids was <0.1. This enables us to assign the 63 promoter sites where m of C and F-TO ≥ 0 as CF-specific hypermethylated regions and the 54 promoter sites where m of C and F-TO < 0 as CF-specific hypo-methylated regions. The heatmap in Fig. 4b summarized this result using the color variance ranging from −1 (hypo) to 1 (hyper). Genes that encode DNA damage repair proteins such as MLH1 and MGMT were hyper-methylated in C and F-TO compared to the rest as indicated in Fig. 4b. This might account for the relatively high mutational burden in C and F-TO (Fig. 2c). Besides, the ...
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... regions and the 54 promoter sites where m of C and F-TO < 0 as CF-specific hypo-methylated regions. The heatmap in Fig. 4b summarized this result using the color variance ranging from −1 (hypo) to 1 (hyper). Genes that encode DNA damage repair proteins such as MLH1 and MGMT were hyper-methylated in C and F-TO compared to the rest as indicated in Fig. 4b. This might account for the relatively high mutational burden in C and F-TO (Fig. 2c). Besides, the promoter region of multiple genes involved in the regulation of cell cycle, proliferation, and migration such as membrane receptor in TGF-β signaling (ACVR2A), FGF signaling (FGFR2), and WNT signaling (CTNNB1) pathways was specifically ...
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... cell cycle, proliferation, and migration such as membrane receptor in TGF-β signaling (ACVR2A), FGF signaling (FGFR2), and WNT signaling (CTNNB1) pathways was specifically hyper-methylated in C and F-TO. In contrast, the rest of PDOs were hyper-methylated in regions encoding chromatin regulator (ZMYM2) and cytoplasmic ribosomal protein (RPL22) ( Fig. 4b; Supplementary Data 4b). Such heterogeneous methylation profiles suggested that post-translational regulation was considerably involved in further tumor progression and diverse clonal expansions of ...
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... transcriptomic, and epi-genomic patterns. Both azacitidine and decitabine inhibit cell growth by inhibiting methylatransferase, yet azacitidine is capable of interfering into RNA to a larger extent than into DNA whereas decitabine is only incorporated into DNA. Considering the better responses of azacitidine than decitabine in organoid C-and F-TO (Fig. 4c, Supplementary Data 4c), we deduced that specific RNA hypermethylation in C and F organoids accounts for the particularly effective response of azacitidine. Overall, these data validate that considerable epi-genomic heterogeneity exists within SICs, which reflects different regulations of multiple pathways including cell growth, signaling transduction, DNA ...
Citations
... It was noted that organoid TILs had a functional reaction to PD-1 immune checkpoint suppression, as seen in the patients [99,100]. In a high-throughput study, patient-derived organoids (PDO), cell lines (PDC), and organoid-derived cell lines (PDOC) produced from each of the six regions of a single synchronous intestinal tumor (SIC) were subjected to genomic, transcriptomic, and epigenetic sequencing and drug screening utilizing a panel of 25 chosen drugs [101]. Short tandem repeat (STR) profiling confirmed the absence of cross-contamination, indicating that all derivatives had matched loci. ...
... Based on mutational analysis of PDOs, PDCs, and PDOCs, all derivatives preserved >75% of genetic variations in each associated tumor. PDOs and PDCs were found to replicate the pathway enrichment of respective tissues [101]. Notably, it was reported that PDOs preserved the patterns of multi-site transcriptomic heterogeneity as well as traits of clonal association found in synchronous tumors [101]. ...
... PDOs and PDCs were found to replicate the pathway enrichment of respective tissues [101]. Notably, it was reported that PDOs preserved the patterns of multi-site transcriptomic heterogeneity as well as traits of clonal association found in synchronous tumors [101]. ...
Organoid models have recently been utilized to study 3D human-derived tissue systems to uncover tissue architecture and adult stem cell biology. Patient-derived organoids unambiguously provide the most suitable in vitro system to study disease biology with the actual genetic background. With the advent of much improved and innovative approaches, patient-derived organoids can potentially be used in regenerative medicine. Various human tissues were explored to develop organoids due to their multifold advantage over the conventional in vitro cell line culture approach and in vivo models. Gastrointestinal (GI) tissues have been widely studied to establish organoids and organ-on-chip for screening drugs, nutraceuticals, and other small molecules having therapeutic potential. The function of channel proteins, transporters, and transmembrane proteins was also explained. The successful application of genome editing in organoids using the CRISPR-Cas approach has been reported recently. GI diseases such as Celiac disease (CeD), Inflammatory bowel disease (IBD), and common GI cancers have been investigated using several patient-derived organoid models. Recent advancements on organoid bio-banking and 3D bio-printing contributed significantly in personalized disease management and therapeutics. This article reviews the available literature on investigations and translational applications of patient-derived GI organoid models, notably on elucidating gut-microbial interaction and epigenetic modifications.
Graphical Abstract
... Organoid also allows more precise profiling of CRC thus driving drug screening, more effectively cater to subpopulation of patients. Yan et al. [146] revealed key mutational and transcriptomic alterations in microsatellite stable sporadic early-onset CRC (EOCRC). CRC PDOs also have been utilized as a genetic model for investigation of the function of driver genes. ...
Colorectal cancer (CRC) is a heterogeneous disease. Conventional two-dimensional (2D) culture employing cell lines was developed to study the molecular properties of CRC in vitro. Although these cell lines which are isolated from the tumor niche in which cancer develop, the translation to human model such as studying drug response is often hindered by the inability of cell lines to recapture original tumor features and the lack of heterogeneous clinical tumors represented by this 2D model, differed from in vivo condition. These limitations which may be overcome by utilizing three-dimensional (3D) culture consisting of spheroids and organoids. Over the past decade, great advancements have been made in optimizing culture method to establish spheroids and organoids of solid tumors including of CRC for multiple purposes including drug screening and establishing personalized medicine. These structures have been proven to be versatile and robust models to study CRC progression and deciphering its heterogeneity. This review will describe on advances in 3D culture technology and the application as well as the challenges of CRC-derived spheroids and organoids as a mode to screen for anticancer drugs.
The establishment of self-organizing 'mini-gut' organoid models has brought about a significant breakthrough in biomedical research. Patient-derived tumor organoids have emerged as valuable tools for preclinical studies, offering the retention of genetic and phenotypic characteristics of the original tumor. These organoids have applications in various research areas, including in vitro modelling, drug discovery and personalized medicine. The present review provided an overview of intestinal organoids, focusing on their unique characteristics and current understanding. The progress made in colorectal cancer (CRC) organoid models was then delved into, discussing their role in drug development and personalized medicine. For instance, it has been indicated that patient-derived tumor organoids are able to predict response to irinotecan-based neoadjuvant chemoradiotherapy. Furthermore, the limitations and challenges associated with current CRC organoid models were addressed, along with proposed strategies for enhancing their utility in future basic and translational research.
