bims-instec 2024-02-25 papers

bims-instec

Biomed News

on Intestinal stem cells and chemoresistance in colon cancer and intestinal regeneration

Issue of 2024‒02‒25
nineteen papers selected by
Maria-Virginia Giolito, Université Catholique de Louvain

The loss of antioxidant activities impairs intestinal epithelium homeostasis by altering lipid metabolism.
Chromatin Remodeling in Patient-Derived Colorectal Cancer Models.
A Proteomic Investigation to Discover Candidate Proteins Involved in Novel Mechanisms of 5-Fluorouracil Resistance in Colorectal Cancer.
Deciphering Endothelial and Mesenchymal Organ Specification in Vascularized Lung and Intestinal Organoids.
A growth factor-reduced culture system for colorectal cancer organoids.
Proteogenomic characterization of primary colorectal cancer and metastatic progression identifies proteome-based subtypes and signatures.
Nutrient metabolism in regulating intestinal stem cell homeostasis.
PPARG activation promotes the proliferation of colorectal cancer cell lines and enhances the antiproliferative effect of 5-fluorouracil.
Association of RNF43 Genetic Alterations With BRAFV600E and MSIhigh in Colorectal Cancer.
BECLIN1 is essential for intestinal homeostasis involving autophagy-independent mechanisms through its function in endocytic trafficking.
YAP mediates apoptosis through failed integrin adhesion reinforcement.
Circulating microRNA analysis in a prospective co-clinical trial identifies MIR652-3p as a response biomarker and driver of regorafenib resistance mechanisms in colorectal cancer.
SATB1 mediated tumor colonization and β-catenin nuclear localization are associated with colorectal cancer progression.
Aberrant PRDM2 methylation as an early event in serrated lesions destined to evolve into microsatellite-instable colorectal cancers.
Investigating the resistance mechanism of 5-fluorouracil in colorectal cancer based on surface markers of cancer stemness and cytokine level: A pre-clinical study.
Dual Inhibition of B7-H3 and EGFR Overcomes Acquired Chemoresistance in Colon Adenocarcinoma.
NOS2 Polymorphism in Aspect of Left and Right-Sided Colorectal Cancer.
Protocol for functional profiling of patient-derived organoids for precision oncology.
Crypt and Villus Enterochromaffin Cells are Distinct Stress Sensors in the Gut.

Exp Cell Res. 2024 Feb 18. pii: S0014-4827(24)00056-9. [Epub ahead of print] 113965

The loss of antioxidant activities impairs intestinal epithelium homeostasis by altering lipid metabolism.

Javier Ramos-León, Concepción Valencia, Mariana Gutiérrez-Mariscal, David-Alejandro Rivera-Miranda, Celina García-Meléndrez, Luis Covarrubias.

  Reactive oxygens species (ROS) are common byproducts of metabolic reactions and could be at the origin of many diseases of the elderly. Here we investigated the role of ROS in the renewal of the intestinal epithelium in mice lacking catalase (CAT) and/or nicotinamide nucleotide transhydrogenase (NNT) activities. Cat-/- mice have delayed intestinal epithelium renewal and were prone to develop necrotizing enterocolitis upon starvation. Interestingly, crypts lacking CAT showed fewer intestinal stem cells (ISC) and lower stem cell activity than wild-type. In contrast, crypts lacking NNT showed a similar number of ISCs as wild-type but increased stem cell activity, which was also impaired by the loss of CAT. No alteration in the number of Paneth cells (PCs) was observed in crypts of either Cat-/- or Nnt-/- mice, but they showed an evident decline in the amount of lysozyme. Cat deficiency caused fat accumulation in crypts, and a fall in the remarkable high amount of adipose triglyceride lipase (ATGL) in PCs. Notably, the low levels of ATGL in the intestine of Cat -/- mice increased after a treatment with the antioxidant N-acetyl cysteine. Supporting a role of ATGL in the regulation of ISC activity, its inhibition halt intestinal organoid development. These data suggest that the reduction of the intestine renewal capacity originates from fatty acid metabolic alterations caused by peroxisomal ROS.

Keywords:  Antioxidants; Paneth cells lipolysis; Reactive oxygen species; Stem cells; Tissue repair

DOI:  https://doi.org/10.1016/j.yexcr.2024.113965
Adv Sci (Weinh). 2024 Feb 21. e2303379

Chromatin Remodeling in Patient-Derived Colorectal Cancer Models.

Kun Xiang, Ergang Wang, John Mantyh, Gabrielle Rupprecht, Marcos Negrete, Golshid Sanati, Carolyn Hsu, Peggy Randon, Anders Dohlman, Kai Kretzschmar, Shree Bose, Nicholas Giroux, Shengli Ding, Lihua Wang, Jorge Prado Balcazar, Qiang Huang, Pasupathi Sundaramoorthy, Rui Xi, Shannon Jones McCall, Zhaohui Wang, Chongming Jiang, Yubin Kang, Scott Kopetz, Gregory E Crawford, Steven M Lipkin, Xiao-Fan Wang, Hans Clevers, David Hsu, Xiling Shen.

  Patient-Derived Organoids (PDO) and Xenografts (PDX) are the current gold standards for patient-derived models of cancer (PDMC). Nevertheless, how patient tumor cells evolve in these models and the impact on drug response remains unclear. Herein, the transcriptomic and chromatin accessibility landscapes of matched colorectal cancer (CRC) PDO, PDX, PDO-derived PDX (PDOX), and original patient tumors (PT) are compared. Two major remodeling axes are discovered. The first axis delineates PDMC from PT, and the second axis distinguishes PDX and PDO. PDOX are more similar to PDX than PDO, indicating the growth environment is a driving force for chromatin adaptation. Transcription factors (TF) that differentially bind to open chromatins between matched PDO and PDOX are identified. Among them, KLF14 and EGR2 footprints are enriched in PDOX relative to matched PDO, and silencing of KLF14 or EGR2 promoted tumor growth. Furthermore, EPHA4, a shared downstream target gene of KLF14 and EGR2, altered tumor sensitivity to MEK inhibitor treatment. Altogether, patient-derived CRC cells undergo both common and distinct chromatin remodeling in PDO and PDX/PDOX, driven largely by their respective microenvironments, which results in differences in growth and drug sensitivity and needs to be taken into consideration when interpreting their ability to predict clinical outcome.

Keywords:  ATAC-seq, Colorectal Cancer (CRC); Patient-Derived Models of Cancer (PDMC); Patient-Derived Organoids (PDO); Patient-Derived Xenografts (PDX)

DOI:  https://doi.org/10.1002/advs.202303379
Cells. 2024 Feb 14. pii: 342. [Epub ahead of print]13(4):

A Proteomic Investigation to Discover Candidate Proteins Involved in Novel Mechanisms of 5-Fluorouracil Resistance in Colorectal Cancer.

Mario Ortega Duran, Sadr Ul Shaheed, Christopher W Sutton, Steven D Shnyder.

  One of the main obstacles to therapeutic success in colorectal cancer (CRC) is the development of acquired resistance to treatment with drugs such as 5-fluorouracil (5-FU). Whilst some resistance mechanisms are well known, it is clear from the stasis in therapy success rate that much is still unknown. Here, a proteomics approach is taken towards identification of candidate proteins using 5-FU-resistant sublines of human CRC cell lines generated in house. Using a multiplexed stable isotope labelling with amino acids in cell culture (SILAC) strategy, 5-FU-resistant and equivalently passaged sensitive cell lines were compared to parent cell lines by growing in Heavy medium with 2D liquid chromatography and Orbitrap Fusion™ Tribrid™ Mass Spectrometry analysis. Among 3003 commonly quantified proteins, six (CD44, APP, NAGLU, CORO7, AGR2, PLSCR1) were found up-regulated, and six (VPS45, RBMS2, RIOK1, RAP1GDS1, POLR3D, CD55) down-regulated. A total of 11 of the 12 proteins have a known association with drug resistance mechanisms or role in CRC oncogenesis. Validation through immunodetection techniques confirmed high expression of CD44 and CD63, two known drug resistance mediators with elevated proteomics expression results. The information revealed by the sensitivity of this method warrants it as an important tool for elaborating the complexity of acquired drug resistance in CRC.

Keywords:  5-fluorouracil; colorectal cancer; drug resistance mechanisms; in vitro models; proteomics; stable isotope labelling with amino acids in cell culture (SILAC)

DOI:  https://doi.org/10.3390/cells13040342
bioRxiv. 2024 Feb 07. pii: 2024.02.06.577460. [Epub ahead of print]

Deciphering Endothelial and Mesenchymal Organ Specification in Vascularized Lung and Intestinal Organoids.

Yifei Miao, Cheng Tan, Nicole M Pek, Zhiyun Yu, Kentaro Iwasawa, Daniel O Kechele, Nambirajan Sundaram, Victor Pastrana-Gomez, Keishi Kishimoto, Min-Chi Yang, Cheng Jiang, Jason Tchieu, Jeffrey A Whitsett, Kyle W McCracken, Robbert J Rottier, Darrell N Kotton, Michael A Helmrath, James M Wells, Takanori Takebe, Aaron M Zorn, Ya-Wen Chen, Minzhe Guo, Mingxia Gu.

To investigate the co-development of vasculature, mesenchyme, and epithelium crucial for organogenesis and the acquisition of organ-specific characteristics, we constructed a human pluripotent stem cell-derived organoid system comprising lung or intestinal epithelium surrounded by organotypic mesenchyme and vasculature. We demonstrated the pivotal role of co-differentiating mesoderm and endoderm via precise BMP regulation in generating multilineage organoids and gut tube patterning. Single-cell RNA-seq analysis revealed organ specificity in endothelium and mesenchyme, and uncovered key ligands driving endothelial specification in the lung (e.g., WNT2B and Semaphorins) or intestine (e.g., GDF15). Upon transplantation under the kidney capsule in mice, these organoids further matured and developed perfusable human-specific sub-epithelial capillaries. Additionally, our model recapitulated the abnormal endothelial-epithelial crosstalk in patients with FOXF1 deletion or mutations. Multilineage organoids provide a unique platform to study developmental cues guiding endothelial and mesenchymal cell fate determination, and investigate intricate cell-cell communications in human organogenesis and disease.Highlights: BMP signaling fine-tunes the co-differentiation of mesoderm and endoderm.The cellular composition in multilineage organoids resembles that of human fetal organs.Mesenchyme and endothelium co-developed within the organoids adopt organ-specific characteristics.Multilineage organoids recapitulate abnormal endothelial-epithelial crosstalk in FOXF1-associated disorders.

DOI: https://doi.org/10.1101/2024.02.06.577460
Cancer Lett. 2024 Feb 19. pii: S0304-3835(24)00130-7. [Epub ahead of print] 216737

A growth factor-reduced culture system for colorectal cancer organoids.

Ronghui Tan, Ze Zhang, Peirong Ding, Yue Liu, Huidong Liu, Minyi Lu, Ye-Guang Chen.

  Although organoids derived from tumor tissues have been widely used in cancer research, it is a great challenge for cultured organoids to retain the characteristics of the original tumor tissues due to their heterogeneity. In this study, we explore organoid culture recipes to capture tumor features of colorectal cancers. We find that the activation of Wnt and EGF signaling and inhibition of BMP signaling are non-essential for the survival of most colorectal cancer organoids (CRCOs). We design a growth factor-reduced culture medium containing FGF10, A83-01 (TGF-β type I receptor inhibitor), SB202190 (p38 MAPK inhibitor), gastrin, and nicotinamide. Using this medium, we can maintain tumor features in long-term CRCO cultivation, as evidenced by histopathology, genetic stability, tumorigenicity, and response of clinical treatments. Our findings offer a reliable and economical strategy for CRCO culture, facilitating the utilization of organoids in colorectal cancer research and treatment.

Keywords:  Colorectal cancer; Culture system; Drug sensitivity; Organoids; Tumorigenesis

DOI:  https://doi.org/10.1016/j.canlet.2024.216737
Cell Rep. 2024 Feb 19. pii: S2211-1247(24)00138-4. [Epub ahead of print]43(2): 113810

Proteogenomic characterization of primary colorectal cancer and metastatic progression identifies proteome-based subtypes and signatures.

Atsushi Tanaka, Makiko Ogawa, Yihua Zhou, Kei Namba, Ronald C Hendrickson, Matthew M Miele, Zhuoning Li, David S Klimstra, Patrick G Buckley, Jeffrey Gulcher, Julia Y Wang, Michael H A Roehrl.

  Metastatic progression of colorectal adenocarcinoma (CRC) remains poorly understood and poses significant challenges for treatment. To overcome these challenges, we performed multiomics analyses of primary CRC and liver metastases. Genomic alterations, such as structural variants or copy number alterations, were enriched in oncogenes and tumor suppressor genes and increased in metastases. Unsupervised mass spectrometry-based proteomics of 135 primary and 123 metastatic CRCs uncovered distinct proteomic subtypes, three each for primary and metastatic CRCs, respectively. Integrated analyses revealed that hypoxia, stemness, and immune signatures characterize these 6 subtypes. Hypoxic CRC harbors high epithelial-to-mesenchymal transition features and metabolic adaptation. CRC with a stemness signature shows high oncogenic pathway activation and alternative telomere lengthening (ALT) phenotype, especially in metastatic lesions. Tumor microenvironment analysis shows immune evasion via modulation of major histocompatibility complex (MHC) class I/II and antigen processing pathways. This study characterizes both primary and metastatic CRCs and provides a large proteogenomics dataset of metastatic progression.

Keywords:  CP: Cancer; biomarkers; colorectal cancer; hypoxia; mass spectrometry; metastasis; molecular signature; proteomics; stemness; subtyping; tumor immune microenvironment

DOI:  https://doi.org/10.1016/j.celrep.2024.113810
Cell Prolif. 2024 Feb 22. e13602

Nutrient metabolism in regulating intestinal stem cell homeostasis.

Ruicheng Shi, Bo Wang.

Intestinal stem cells (ISCs) are known for their remarkable proliferative capacity, making them one of the most active cell populations in the body. However, a high turnover rate of intestinal epithelium raises the likelihood of dysregulated homeostasis, which is known to cause various diseases, including cancer. Maintaining precise control over the homeostasis of ISCs is crucial to preserve the intestinal epithelium's integrity during homeostasis or stressed conditions. Recent research has indicated that nutrients and metabolic pathways can extensively modulate the fate of ISCs. This review will explore recent findings concerning the influence of various nutrients, including lipids, carbohydrates, and vitamin D, on the delicate balance between ISC proliferation and differentiation.

DOI: https://doi.org/10.1111/cpr.13602
BMC Cancer. 2024 Feb 20. 24(1): 234

PPARG activation promotes the proliferation of colorectal cancer cell lines and enhances the antiproliferative effect of 5-fluorouracil.

Leah Schöckel, Christine Woischke, Sai Agash Surendran, Marlies Michl, Tobias Schiergens, Andreas Hölscher, Florian Glass, Peter Kreissl, Frederick Klauschen, Michael Günther, Steffen Ormanns, Jens Neumann.

  BACKGROUND: Peroxisome proliferator-activated receptor gamma (PPARG) is a member of the nuclear receptor family. It is involved in the regulation of adipogenesis, lipid metabolism, insulin sensitivity, vascular homeostasis and inflammation. In addition, PPARG agonists, known as thiazolidinediones, are well established in the treatment of type 2 diabetes mellitus. PPARGs role in cancer is a matter of debate, as pro- and anti-tumour properties have been described in various tumour entities. Currently, the specific role of PPARG in patients with colorectal cancer (CRC) is not fully understood.MATERIAL AND METHODS: The prognostic impact of PPARG expression was investigated by immunohistochemistry in a case-control study using a matched pair selection of CRC tumours (n = 246) with either distant metastases to the liver (n = 82), lung (n = 82) or without distant metastases (n = 82). Its effect on proliferation as well as the sensitivity to the chemotherapeutic drug 5-fluorouracil (5-FU) was examined after activation, inhibition, and transient gene knockdown of PPARG in the CRC cell lines SW403 and HT29.
RESULTS: High PPARG expression was significantly associated with pulmonary metastasis (p = 0.019). Patients without distant metastases had a significantly longer overall survival with low PPARG expression in their tumours compared to patients with high PPARG expression (p = 0.045). In the pulmonary metastasis cohort instead, a trend towards longer survival was observed for patients with high PPARG expression in their tumour (p = 0.059). Activation of PPARG by pioglitazone and rosiglitazone resulted in a significant dose-dependent increase in proliferation of CRC cell lines. Inhibition of PPARG by its specific inhibitor GW9662 and siRNA-mediated knockdown of PPARG significantly decreased proliferation. Activating PPARG significantly increased the CRC cell lines sensitivity to 5-FU while its inhibition decreased it.
CONCLUSION: The prognostic effect of PPARG expression depends on the metastasis localization in advanced CRC patients. Activation of PPARG increased malignancy associated traits such as proliferation in CRC cell lines but also increases sensitivity towards the chemotherapeutic agent 5-FU. Based on this finding, a combination therapy of PPARG agonists and 5-FU-based chemotherapy constitutes a promising strategy which should be further investigated.

Keywords:  5-fluorouracil; Colorectal cancer; metastasis; Diabetes mellitus; PPARG

DOI:  https://doi.org/10.1186/s12885-024-11985-5
JCO Precis Oncol. 2024 Feb;8 e2300411

Association of RNF43 Genetic Alterations With BRAFV600E and MSIhigh in Colorectal Cancer.

Arndt Vogel, Karthikeyan Murugesan, Gajanan Kendre, Julia C F Quintanilha, Jeffrey S Ross, Tilman Brummer, Anna Saborowski.

PURPOSE: Recent studies have provided evidence for a predictive value of RNF43 genetic alterations (GAs) as biomarkers for targeted therapies in microsatellite-stable (MSS) colorectal cancer (CRC). These data have the potential to prioritize treatment strategies in patients with BRAFV600E-mutant CRC and help to identify a subgroup that is more likely to derive benefit versus those patients for whom alternative treatment approaches are needed. We were therefore interested in defining the precise frequency of BRAFV600E and RNF43 GAs and their respective overlap in a large cohort of patients with CRC.METHODS: To address this question, we performed a retrospective analysis that included 52,969 patients diagnosed with CRC from the FoundationCORE database.
RESULTS: We observed a striking association of RNF43 GAs with MSI and tumor mutational burden status and BRAFV600E mutations. Overall, 23% of MSS patients with confirmed BRAFV600E mutation harbor an RNF43 GA-which accounts for 1.1% of all patients with CRC and for 15.7% of all CRC BRAFV600E cases.
CONCLUSION: Ongoing phase III clinical trials, such as BREAKWATER, should aim to incorporate broader genetic profiling to further validate the superior sensitivity of patients with RNF43-mutant, MSS BRAFV600E CRC to anti-EGFR-/BRAFi-based therapies.

DOI: https://doi.org/10.1200/PO.23.00411
Commun Biol. 2024 Feb 20. 7(1): 209

BECLIN1 is essential for intestinal homeostasis involving autophagy-independent mechanisms through its function in endocytic trafficking.

Sharon Tran, Juliani Juliani, Tiffany J Harris, Marco Evangelista, Julian Ratcliffe, Sarah L Ellis, David Baloyan, Camilla M Reehorst, Rebecca Nightingale, Ian Y Luk, Laura J Jenkins, Sonia Ghilas, Marina H Yakou, Chantelle Inguanti, Chad Johnson, Michael Buchert, James C Lee, Peter De Cruz, Kinga Duszyc, Paul A Gleeson, Benjamin T Kile, Lisa A Mielke, Alpha S Yap, John M Mariadason, W Douglas Fairlie, Erinna F Lee.

Autophagy-related genes have been closely associated with intestinal homeostasis. BECLIN1 is a component of Class III phosphatidylinositol 3-kinase complexes that orchestrate autophagy initiation and endocytic trafficking. Here we show intestinal epithelium-specific BECLIN1 deletion in adult mice leads to rapid fatal enteritis with compromised gut barrier integrity, highlighting its intrinsic critical role in gut maintenance. BECLIN1-deficient intestinal epithelial cells exhibit extensive apoptosis, impaired autophagy, and stressed endoplasmic reticulum and mitochondria. Remaining absorptive enterocytes and secretory cells display morphological abnormalities. Deletion of the autophagy regulator, ATG7, fails to elicit similar effects, suggesting additional novel autophagy-independent functions of BECLIN1 distinct from ATG7. Indeed, organoids derived from BECLIN1 KO mice show E-CADHERIN mislocalisation associated with abnormalities in the endocytic trafficking pathway. This provides a mechanism linking endocytic trafficking mediated by BECLIN1 and loss of intestinal barrier integrity. Our findings establish an indispensable role of BECLIN1 in maintaining mammalian intestinal homeostasis and uncover its involvement in endocytic trafficking in this process. Hence, this study has important implications for our understanding of intestinal pathophysiology.

DOI: https://doi.org/10.1038/s42003-024-05890-7
Cell Rep. 2024 Feb 21. pii: S2211-1247(24)00139-6. [Epub ahead of print]43(3): 113811

YAP mediates apoptosis through failed integrin adhesion reinforcement.

Lidan Shi, Elisabeth Nadjar-Boger, Hamidreza Jafarinia, Aurélie Carlier, Haguy Wolfenson.

  Extracellular matrix (ECM) rigidity is a major effector of cell fate decisions. Whereas cell proliferation on stiff matrices, wherein Yes-associated protein (YAP) plays a pivotal role, is well documented, activation of apoptosis in response to soft matrices is poorly understood. Here, we show that YAP drives the apoptotic decision as well. We find that in cells on soft matrices, YAP is recruited to small adhesions, phosphorylated at the Y357 residue, and translocated into the nucleus, ultimately leading to apoptosis. In contrast, Y357 phosphorylation levels are dramatically low in large adhesions on stiff matrices. Furthermore, mild attenuation of actomyosin contractility allows adhesion growth on soft matrices, leading to reduced Y357 phosphorylation levels and resulting in cell growth. These findings indicate that failed adhesion reinforcement drives rigidity-dependent apoptosis through YAP and that this decision is not determined solely by ECM rigidity but rather by the balance between cellular forces and ECM rigidity.

Keywords:  CP: Cell biology; Src; YAP; actomyosin; anchorage-independence; apoptosis; c-Abl; mechanosensing; pYAP-Y357; rigidity sensing

DOI:  https://doi.org/10.1016/j.celrep.2024.113811
Clin Cancer Res. 2024 Feb 20.

Circulating microRNA analysis in a prospective co-clinical trial identifies MIR652-3p as a response biomarker and driver of regorafenib resistance mechanisms in colorectal cancer.

Somaieh Hedayat, Luciano Cascione, David Cunningham, Marta Schirripa, Andrea Lampis, Jens C Hahne, Nina Tunariu, Sung Pil Hong, Silvia Marchetti, Khurum Khan, Elisa Fontana, Valentina Angerilli, Mia Delrieux, Daniel Nava Rodrigues, Letizia Procaccio, Sheela Rao, David Watkins, Naureen Starling, Ian Chau, Chiara Braconi, Nicos Fotiadis, Ruwaida Begum, Naomi Guppy, Louise Howell, Melanie Valenti, Scott Cribbes, Bernadett Kolozsvari, Vladimir Kirkin, Sara Lonardi, Michele Ghidini, Rodolfo Passalacqua, Raghad Elghadi, Luca Magnani, David J Pinato, Federica Di Maggio, Filippo Ghelardi, Elisa Sottotetti, Guglielmo Vetere, Paolo Ciraci, Georgios Vlachogiannis, Filippo Pietrantonio, Chiara Cremolini, Alessio Cortellini, Fotios Loupakis, Matteo Fassan, Nicola Valeri.

BACKGROUND: The multi-kinase inhibitor regorafenib has demonstrated efficacy in chemo-refractory metastatic colorectal cancer (mCRC) patients. However, lack of predictive biomarkers and concerns over significant toxicities hamper the use of regorafenib in clinical practice.METHODS: Serial liquid biopsies were obtained at baseline and monthly until disease progression in chemo-refractory mCRC patients treated with regorafenib in a phase II clinical trial (PROSPECT-R n=40; NCT03010722) and in a multicentric validation cohort (n=241). Tissue biopsies collected at baseline, after 2 months and at progression in the PROSPECT-R trial were used to establish Patient-Derived Organoids (PDOs) and for molecular analyses. MicroRNA profiling was performed on baseline bloods using the NanoString nCounter platform and results were validated by digital droplet PCR and/or In Situ Hybridization in paired liquid and tissue biopsies. PDOs co-cultures and PDO-xenotransplants were generated for functional analyses.
RESULTS: Large-scale microRNA expression analysis in longitudinal matched liquid and tissue biopsies from the PROSPECT-R trial identified MIR652-3p as a biomarker of clinical benefit to regorafenib. These findings were confirmed in an independent validation cohort and in a "control" group of 100 patients treated with lonsurf. Using ex vivo co-culture assays paired with single-cell RNA-sequencing of PDO established pre- and post-treatment, we modelled regorafenib response observed in vivo and in patients, and showed that MIR652-3p controls resistance to regorafenib by impairing regorafenib-induced lethal autophagy and by orchestrating the switch from neo-angiogenesis to vessel co-option.
CONCLUSIONS: Our results identify MIR652-3p as potential biomarker and as a driver of cell and non-cell autonomous mechanisms of resistance to regorafenib.

DOI: https://doi.org/10.1158/1078-0432.CCR-23-2748
Cancer Biol Ther. 2024 Dec 31. 25(1): 2320307

SATB1 mediated tumor colonization and β-catenin nuclear localization are associated with colorectal cancer progression.

Luan Sun, Feng Wang, Xufei Wang, Feiying Zhang, Sujuan Ma, Jinghuan Lv.

  Colorectal cancer (CRC) is a malignancy with high incidence and poor prognosis. It is urgent to identify valuable biomarkers for early diagnosis and potent therapeutic targets. It has been reported that SATB1 is associated with the malignant progression in CRC. To explore the role of SATB1 in CRC progression and the underlying mechanism, we evaluated the expression of SATB1 in the paired CRC tissues with immunohistochemistry. The results showed that the expression of SATB1 in lymph node metastasis was higher than that in primary lesion, and that in distant organ metastasis was higher than that in primary lesion. The retrospective analysis showed that patients with high expression of SATB1 had a significantly worse prognosis than those with negative and moderate expression. In vitro experiments that employing SATB1 over-expressing and depleted CRC cell lines confirmed that SATB1 contributes to cell proliferation and colonization, while inhibiting cell motility. Furthermore, the tissue immunofluorescence assay, Co-IP and Western blot were conducted to reveal that SATB1 induced translocation of β-catenin and formed a protein complex with it in the nuclei. In conclusion, SATB1 mediated tumor colonization and β-catenin nuclear localization are associated with the malignant progression and poor prognosis of CRC.

Keywords:  Colorectal cancer progression; SATB1; tumor colonization; β-catenin

DOI:  https://doi.org/10.1080/15384047.2024.2320307
J Pathol Clin Res. 2024 Mar;10(2): e348

Aberrant PRDM2 methylation as an early event in serrated lesions destined to evolve into microsatellite-instable colorectal cancers.

David Efwm van Toledo, Arne Gc Bleijenberg, Andrea Venema, Mireille J de Wit, Susanne van Eeden, Gerrit A Meijer, Beatrice Carvalho, Evelien Dekker, Peter Henneman, Joep Eg IJspeert, Carel Jm van Noesel.

  Up to 30% of colorectal cancers (CRCs) develop from sessile serrated lesions (SSLs). Within the serrated neoplasia pathway, at least two principally distinct oncogenetic routes exist generating microsatellite-stable and microsatellite-instable CRCs, respectively. Aberrant DNA methylation (DNAm) is found early in the serrated pathway and might play a role in both oncogenetic routes. We studied a cohort of 23 SSLs with a small focus (<10 mm) of dysplasia or cancer, 10 of which were MLH1 deficient and 13 MLH1 proficient. By comparing, for each SSL, the methylation status of (1) the region of dysplasia or cancer (SSL-D), (2) the nondysplastic SSL (SSL), and (3) adjacent normal mucosa, differentially methylated probes (DMPs) and regions (DMRs) were assessed both genome-wide as well as in a tumor-suppressor gene-focused approach. By comparing DNAm of MLH1-deficient SSL-Ds with their corresponding SSLs, we identified five DMRs, including those annotating for PRDM2 and, not unexpectedly, MLH1. PRDM2 gene promotor methylation was associated with MLH1 expression status, as it was largely hypermethylated in MLH1-deficient SSL-Ds and hypomethylated in MLH1-proficient SSL-Ds. Significantly increased DNAm levels of PRDM2 and MLH1, in particular at 'critical' MLH1 probe sites, were to some extent already visible in SSLs as compared to normal mucosa (p = 0.02, p = 0.01, p < 0.0001, respectively). No DMRs, nor DMPs, were identified for SSLs destined to evolve into MLH1-proficient SSL-Ds. Our data indicate that, within both arms of the serrated CRC pathway, the majority of the epigenetic alterations are introduced early during SSL formation. Promoter hypermethylation of PRDM2 and MLH1 on the other hand specifically initiates in SSLs destined to transform into MLH1-deficient CRCs suggesting that the fate of SSLs may not necessarily result from a stochastic process but possibly is already imprinted and predisposed.

Keywords:  colorectal cancer; methylation; serrated neoplasia pathway; sessile serrated lesions

DOI:  https://doi.org/10.1002/cjp2.348
J Cancer Res Ther. 2023 Jan 01. 19(Suppl 2): S560-S568

Investigating the resistance mechanism of 5-fluorouracil in colorectal cancer based on surface markers of cancer stemness and cytokine level: A pre-clinical study.

Samir G Patel, Alkeshkumar Patel, Nupur Patel, Birva Raiya, Hemangini Vora, Neeraj Jain.

BACKGROUND: Colorectal cancer (CRC) is the deadliest malignancy in the world. The first-line chemotherapy used for CRC is 5-fluorouracil (5-FU). 5-FU completely eradicates rapidly proliferating and terminally differentiated tumor cells but fails to target cancer stem cells (CSCs). As a result, the tumor may shrink temporarily, but remnant CSC multiplies and forms a tumor again more aggressively. The recurrence and resistance lead to metastasis.METHODOLOGY: CRC was induced in 12 Sprague-Dawley (RPCP/IAEC/2019-20/R2) rats by 1,2 dimethyl hydrazine. Later, animals were treated with 5-FU for 7 weeks at a 10 mg/kg dose by the subcutaneous route. At the end of treatment, half population was sacrificed (6), whereas the remaining half (6) was left without treatment of 5-FU for 5 weeks and then sacrificed. Parameters such as body weight, complete blood count (CBC), immune cell subset (CD4, CD8, and NK cells), colon length to weight index, interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-α) level, occult blood in stool, tumor multiplicity, and liver metastasis were estimated. In addition, the dissected colon was fixed in formalin and sent to the histology lab for hematoxylin-eosin staining and immunohistochemistry at both intervals.
RESULTS: All blood and tissue-based markers have shown significant differences (p < 0.05) between the animals sacrificed at the end of the 27th week and the end of the 32nd week for 5-FU treatment.
CONCLUSION: It can be concluded that 5-FU up-regulates inflammatory cytokines and cell surface markers of CSC that promote CRC stemness via the Wnt/β-catenin pathway. Also, involvement of Nf-κB, fibronectin, MMP-9, and RANKL leads to tumorigenesis, disease aggressiveness, metastasis, and resistance.

DOI: https://doi.org/10.4103/jcrt.jcrt_1299_22
J Cancer. 2024 ;15(6): 1750-1761

Dual Inhibition of B7-H3 and EGFR Overcomes Acquired Chemoresistance in Colon Adenocarcinoma.

Liang-Chi Chen, Pei-Chen Yang, Chia-Yi Chen, Shu-Fen Chiang, Tsung-Wei Chen, William Tzu-Liang Chen, Tao-Wei Ke, Ji-An Liang, An-Cheng Shiau, K S Clifford Chao, Kevin Chih-Yang Huang.

  Despite advances in therapeutic strategies for colorectal cancer (CRC), CRC has a high disease incidence with significant morbidity and mortality worldwide. Notably, immunotherapy has shown limited efficacy in treating metastatic CRC, underscoring the need for alternative immunotherapeutic targets for the management of metastatic colorectal cancer (mCRC). In the present study, we evaluated the levels of the immune checkpoint proteins PD-L1, PD-L2 and B7-H3 in a large cohort retrospective study. We found that tumor B7-H3 (52.7%) was highly expressed in primary tumors compared to that in PD-L1 (33.6%) or PD-L2 (34.0%). Elevated B7-H3 expression was associated with advanced stage and the risk of distant metastasis and correlated with poor disease-free survival (DFS), suggesting that tumor B7-H3 was an independent prognostic factor associated with worse DFS in colon adenocarcinoma patients (COAD), especially high-risk COAD patients who received adjuvant chemotherapy. Furthermore, we found that B7-H3 significantly promoted cell proliferation and tumor growth in CRC. B7-H3 may stabilize EGFR to activate its downstream pathway for cancer cell proliferation and resistance to oxaliplatin (OXP). Dual targeting of B7-H3 and EGFR markedly rescued the susceptibility to chemotherapy in colorectal cancer cells in vitro and in vivo. Overall, these results showed that B7-H3 exhibited a high prevalence in COAD patients and was significantly associated with worse prognosis in COAD patients. Dual targeting of B7-H3 and EGFR signaling might be a potential therapeutic strategy for high-risk COAD patients.

Keywords:  B7-H3; Colorectal cancer; Intratumoral infiltrating lymphocytes; PD-L1

DOI:  https://doi.org/10.7150/jca.91089
J Clin Med. 2024 Feb 06. pii: 937. [Epub ahead of print]13(4):

NOS2 Polymorphism in Aspect of Left and Right-Sided Colorectal Cancer.

Justyna Klusek, Piotr Lewitowicz, Ruslan Oblap, Ewa Orlewska, Bartosz Witczak, Michał Tomasz Marzec, Monika Kozłowska-Geller, Łukasz Nawacki, Monika Wawszczak-Kasza, Kamila Kocańda, Artur Jóźwik, Stanisław Głuszek.

  Background: The NOS2 gene polymorphism rs2297518 is associated with an increased level of NO, which could contribute to colorectal cancer (CRC) development. We hypothesized that the potential influence of the NOS2 gene polymorphism on cancer development may vary between right-sided and left-sided colon cancers, and rectal cancers. The aim of this study was to determine the rs2297518 polymorphism influence on colorectal cancer development with regard to tumor localization. Methods: This case-control study included 199 patients with CRC and 120 controls. The qPCR endpoint genotyping was conducted using the TaqMan® genotyping assay. Results: This study revealed significant differences in tumor characteristic and in the minor alelle A frequency in the NOS2 genotype between colorectal cancers with different localizations. The mucinous adenocarcinoma was diagnosed significantly more often in right-sided cancers than in left-sided (30.6% vs. 10.9%, p = 0.009) and rectal cancers (30.6% vs. 7.1%, p = 0.0003). The minor allele A of the NOS2 genotype was observed more frequently in right-sided cancers than in left-sided cancers (44.9% vs. 23.1%, p = 0.0137) and more frequently in rectal cancers than in left-sided cancers (40.0% vs. 23.1%, p = 0.0285). Conclusions: In conclusion, the results support the hypothesis that the SNP rs2297518 of the NOS2 gene influences colorectal cancer development with regard to tumor localization.

Keywords:  NOS2; colorectal cancer; gene polymorphism right-sided CRC; left-sided CRC

DOI:  https://doi.org/10.3390/jcm13040937
STAR Protoc. 2024 Feb 15. pii: S2666-1667(24)00052-2. [Epub ahead of print]5(1): 102887

Protocol for functional profiling of patient-derived organoids for precision oncology.

Niloofar Nemati, Nina Boeck, Giorgia Lamberti, Rebecca Lisandrelli, Zlatko Trajanoski.

  Functional precision oncology-a strategy based on perturbing primary tumor cells from cancer patients-could provide a road forward for personalized treatment. Here, we present a comprehensive protocol covering generation and culture of patient-derived colorectal organoids, isolation and expansion of tumor-infiltrating lymphocytes (TILs), and isolation and culture of peripheral blood mononuclear cells (PBMCs). With this protocol, samples fulfilling the demands for performing multi-omics analysis, e.g., RNA sequencing (RNA-seq), whole-exome sequencing (WES), single-cell RNA sequencing (scRNA-seq), and (phospho-)proteomics, can be generated. For complete details on the use and execution of this protocol, please refer to Plattner et al. (2023).1.

Keywords:  Cancer; Organoids; RNAseq; Sequencing; Systems biology

DOI:  https://doi.org/10.1016/j.xpro.2024.102887
bioRxiv. 2024 Feb 08. pii: 2024.02.06.579180. [Epub ahead of print]

Crypt and Villus Enterochromaffin Cells are Distinct Stress Sensors in the Gut.

Kouki K Touhara, Nathan D Rossen, Fei Deng, Tifany Chu, Onur Cil, Yulong Li, David Julius.

The crypt-villus structure of the small intestine serves as an essential protective barrier, with its integrity monitored by the gut's sensory system. Enterochromaffin (EC) cells, a subtype of sensory epithelial cells that release serotonin (5-HT), surveil the mucosal environment and signal both within and outside the gut. However, it remains unclear whether EC cells in intestinal crypts and villi respond to different stimuli and elicit distinct responses. In this study, we introduce a new reporter mouse model to observe the release and propagation of serotonin in live intestines. Using this system, we show that crypt EC cells exhibit two modes of serotonin release: transient receptor potential A1 (TRPA1)-dependent tonic serotonin release that controls basal ionic secretion, and irritant-evoked serotonin release that activates gut sensory neurons. Furthermore, we find that a thick protective mucus layer prevents TRPA1 receptors on crypt EC cells from responding to luminal irritants such as reactive electrophiles; if this mucus layer is compromised, then crypt EC cells become susceptible to activation by luminal irritants. On the other hand, villus EC cells detect oxidative stress through TRPM2 channels and co-release serotonin and ATP. Our work highlights the physiological importance of intestinal architecture and differential TRP channel expression in sensing noxious stimuli that elicit nausea and/or pain sensations in the gut.

DOI: https://doi.org/10.1101/2024.02.06.579180