bims-instec 2024-12-01 papers

bims-instec

Biomed News

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

Issue of 2024–12–01
twenty-one papers selected by
Maria-Virginia Giolito, Université Catholique de Louvain

FZD5 controls intestinal crypt homeostasis and colonic Wnt surrogate agonist response.
PIEZO-dependent mechanosensing is essential for intestinal stem cell fate decision and maintenance.
IFN-γ-dependent regulation of intestinal epithelial homeostasis by NKT cells.
UGT8 mediated sulfatide synthesis modulates BAX localization and dictates apoptosis sensitivity of colorectal cancer.
Frizzled5 controls murine intestinal epithelial cell plasticity through organization of chromatin accessibility.
Spatially organized tumor-stroma boundary determines the efficacy of immunotherapy in colorectal cancer patients.
Preparing Chamber Slides With Pressed Collagen for Live Imaging Monolayers of Primary Human Intestinal Stem Cells.
Genetic variation at 11q23.1 confers colorectal cancer risk by dysregulation of colonic tuft cell transcriptional activator POU2AF2.
Antibody-Drug Conjugates Targeting the EGFR Ligand Epiregulin Elicit Robust Anti-Tumor Activity in Colorectal Cancer.
Early onset metastatic colorectal cancer patients as a distinctive clinical and molecular phenomenon.
ACSL4 and polyunsaturated lipids support metastatic extravasation and colonization.
TPM4 overexpression drives colon epithelial cell tumorigenesis by suppressing differentiation and promoting proliferation.
Hedgehog Signalling Pathway and Its Role in Shaping the Architecture of Intestinal Epithelium.
AXIN2 is a non-redundant regulator of AXIN1 stability and β-catenin in colorectal cancer cells.
Oxaliplatin and 5-fluorouracil promote epithelial-mesenchymal transition via activation of KRAS/ERK/NF-κB pathway in KRAS-mutated colon cancer cells.
Loss of peroxiredoxin 6 (PRDX6) alters lipid composition and distribution resulting in increased sensitivity to ferroptosis.
Basal cell of origin resolves neuroendocrine-tuft lineage plasticity in cancer.
Targeting extracellular matrix interaction in gastrointestinal cancer: Immune modulation, metabolic reprogramming, and therapeutic strategies.
CPT1A mediates radiation sensitivity in colorectal cancer.
The single-cell spatial landscape of stage III colorectal cancers.
Classification of colorectal cancer patients based on serum micronutrients: An exploratory investigation.

Dev Cell. 2024 Nov 19. pii: S1534-5807(24)00662-2. [Epub ahead of print]

FZD5 controls intestinal crypt homeostasis and colonic Wnt surrogate agonist response.

Qinghui Mu, Andrew Ha, Antonio J M Santos, Yuan-Hung Lo, Vincent van Unen, Yi Miao, Madeline Tomaske, Veronica K Guzman, Samira Alwahabi, Jenny J Yuan, Lu Deng, Linheng Li, K Christopher Garcia, Calvin J Kuo.

  The rapidly regenerating intestinal epithelium requires crypt intestinal stem cells (ISCs). Wnt/β-catenin signaling maintains crypt homeostasis and Lgr5+ ISCs, and WNT ligands bind Frizzled receptors (FZD1-10). Identifying specific FZD(s) essential for intestinal homeostasis has been elusive; however, bioengineered antagonists blocking Wnt binding to FZD5 and FZD8 deplete the gut epithelium in vivo, highlighting potential roles. Here, an epithelial-specific Fzd5 knockout (KO) elicited lethal pan-intestinal crypt and villus loss, whereas an Lgr5+ ISC-specific Fzd5 KO depleted Lgr5+ ISCs via premature differentiation and repressed Wnt target genes. Fzd5-null phenotypes were rescued by constitutive β-catenin activation in vivo and in both mouse and human enteroids. KO of Fzd5, not Fzd8, in enteroids ablated responsiveness to dual-specificity FZD5/FZD8-selective Wnt surrogate agonists, which ameliorated DSS-induced colitis in wild-type and Fzd8 KO mice. Overall, FZD5 is a dominant and essential regulator of crypt homeostasis, Lgr5+ ISCs, and intestinal response to Wnt surrogate agonists, with implications for therapeutic mucosal repair.

Keywords:  Fzd5; Lgr5; Wnt signaling; colitis; intestinal stem cells

DOI:  https://doi.org/10.1016/j.devcel.2024.10.022
Science. 2024 Nov 29. 386(6725): eadj7615

PIEZO-dependent mechanosensing is essential for intestinal stem cell fate decision and maintenance.

Meryem B Baghdadi, Ronja M Houtekamer, Louisiane Perrin, Abilasha Rao-Bhatia, Myles Whelen, Linda Decker, Martin Bergert, Carlos Pérez-Gonzàlez, Réda Bouras, Giacomo Gropplero, Adrian K H Loe, Amin Afkhami-Poostchi, Xin Chen, Xi Huang, Stephanie Descroix, Jeffrey L Wrana, Alba Diz-Muñoz, Martijn Gloerich, Arshad Ayyaz, Danijela Matic Vignjevic, Tae-Hee Kim.

Stem cells perceive and respond to biochemical and physical signals to maintain homeostasis. Yet, it remains unclear how stem cells sense mechanical signals from their niche in vivo. In this work, we investigated the roles of PIEZO mechanosensitive channels in the intestinal stem cell (ISC) niche. We used mouse genetics and single-cell RNA sequencing analysis to assess the requirement for PIEZO channels in ISC maintenance. In vivo measurement of basement membrane stiffness showed that ISCs reside in a more rigid microenvironment at the bottom of the crypt. Three-dimensional and two-dimensional organoid systems combined with bioengineered substrates and a stretching device revealed that PIEZO channels sense extracellular mechanical stimuli to modulate ISC function. This study delineates the mechanistic cascade of PIEZO activation that coordinates ISC fate decision and maintenance.

DOI: https://doi.org/10.1126/science.adj7615
Cell Rep. 2024 Nov 23. pii: S2211-1247(24)01299-3. [Epub ahead of print]43(12): 114948

IFN-γ-dependent regulation of intestinal epithelial homeostasis by NKT cells.

Marta Lebrusant-Fernandez, Tom Ap Rees, Rebeca Jimeno, Nikolaos Angelis, Joseph C Ng, Franca Fraternali, Vivian S W Li, Patricia Barral.

  Intestinal homeostasis is maintained through the combined functions of epithelial and immune cells that collaborate to preserve the integrity of the intestinal barrier. However, the mechanisms by which immune cell populations regulate intestinal epithelial cell (IEC) homeostasis remain unclear. Here, we use a multi-omics approach to study the immune-epithelial crosstalk and identify CD1d-restricted natural killer T (NKT) cells as key regulators of IEC biology. We find that NKT cells are abundant in the proximal small intestine and show hallmarks of activation at steady state. Subsequently, NKT cells regulate the survival and the transcriptional and cellular composition landscapes of IECs in intestinal organoids, through interferon-γ (IFN-γ) and interleukin-4 secretion. In vivo, lack of NKT cells results in an increase in IEC turnover, while NKT cell activation leads to IFN-γ-dependent epithelial apoptosis. Our findings propose NKT cells as potent producers of cytokines that contribute to the regulation of IEC homeostasis.

Keywords:  CP: Immunology; IFN-γ; NKT cell; intestinal epithelial cell; intestinal organoids

DOI:  https://doi.org/10.1016/j.celrep.2024.114948
Cell Death Differ. 2024 Nov 23.

UGT8 mediated sulfatide synthesis modulates BAX localization and dictates apoptosis sensitivity of colorectal cancer.

Le Zhang, Prashanthi Ramesh, Lidia Atencia Taboada, Rebecca Roessler, Dick W Zijlmans, Michiel Vermeulen, Daisy I Picavet-Havik, Nicole N van der Wel, Frédéric M Vaz, Jan Paul Medema.

Elevated de novo lipid synthesis is a remarkable adaptation of cancer cells that can be exploited for therapy. However, the role of altered lipid metabolism in the regulation of apoptosis is still poorly understood. Using thermal proteome profiling, we identified Manidipine-2HCl, targeting UGT8, a key enzyme in the synthesis of sulfatides. In agreement, lipidomic analysis indicated that sulfatides are strongly reduced in colorectal cancer cells upon treatment with Manidipine-2HCl. Intriguingly, this reduction led to severe mitochondrial swelling and a strong synergism with BH3 mimetics targeting BCL-XL, leading to the activation of mitochondria-dependent apoptosis. Mechanistically, Manidipine-2HCl enhanced mitochondrial BAX localization in a sulfatide-dependent fashion, facilitating its activation by BH3 mimetics. In conclusion, our data indicates that UGT8 mediated synthesis of sulfatides controls mitochondrial homeostasis and BAX localization, dictating apoptosis sensitivity of colorectal cancer cells.

DOI: https://doi.org/10.1038/s41418-024-01418-y
Dev Cell. 2024 Nov 15. pii: S1534-5807(24)00661-0. [Epub ahead of print]

Frizzled5 controls murine intestinal epithelial cell plasticity through organization of chromatin accessibility.

Lu Deng, Xi C He, Shiyuan Chen, Ning Zhang, Fengyan Deng, Allison Scott, Yanfeng He, Dai Tsuchiya, Sarah E Smith, Michael Epp, Seth Malloy, Fang Liu, Mark Hembree, Qinghui Mu, Jeffrey S Haug, Ermanno Malagola, Huzaifa Hassan, Kaitlyn Petentler, Rhonda Egidy, Lucinda Maddera, Jonathon Russell, Yan Wang, Hua Li, Chongbei Zhao, Anoja Perera, Timothy C Wang, Calvin J Kuo, Linheng Li.

  The homeostasis of the intestinal epithelium relies on intricate yet insufficiently understood mechanisms of intestinal epithelial plasticity. Here, we elucidate the pivotal role of Frizzled5 (Fzd5), a Wnt pathway receptor, as a determinant of murine intestinal epithelial cell fate. Deletion of Fzd5 in Lgr5+ intestinal stem cells (ISCs) impairs their self-renewal, whereas its deletion in Krt19+ cells disrupts lineage generation, without affecting crypt integrity in either case. However, a broader deletion of Fzd5 across the epithelium leads to substantial crypt deterioration. Integrated analysis of single-cell RNA sequencing (scRNA-seq) and single-cell ATAC-seq (scATAC-seq) identifies that Fzd5 governs chromatin accessibility, orchestrating the regulation of stem- and lineage-related gene expression mainly in ISCs and progenitor cells. In summary, our findings provide insights into the regulatory role of Fzd5 in governing intestinal epithelial plasticity.

Keywords:  Fzd5; Wnt pathway; chromatin accessibility; intestinal stem cell; plasticity; single-cell transcriptome and epigenome

DOI:  https://doi.org/10.1016/j.devcel.2024.10.021
Nat Commun. 2024 Nov 26. 15(1): 10259

Spatially organized tumor-stroma boundary determines the efficacy of immunotherapy in colorectal cancer patients.

Yu Feng, Wenjuan Ma, Yupeng Zang, Yanying Guo, Young Li, Yixuan Zhang, Xuan Dong, Yi Liu, Xiaojuan Zhan, Zhizhong Pan, Mei Luo, Miaoqing Wu, Ao Chen, Da Kang, Gong Chen, Longqi Liu, Jingying Zhou, Rongxin Zhang.

Colorectal cancer (CRC) patients with mismatch repair (MMR)-deficient (dMMR) but not MMR-proficient (pMMR) tend to benefit from immune checkpoint blockade (ICB) therapy. To profile the tumor microenvironments (TME) underlying these varied therapeutic responses, we integrate spatial enhanced resolution omics-sequencing (Stereo-seq), single-cell RNA sequencing, and multiplexed imaging analysis to create high-definition spatial maps of tumors from treatment-naïve and ICB-treated CRC patients. Our results identify the spatial organization and immune status of the tumor-stroma boundary as a distinctive feature of dMMR and pMMR CRCs, which associates with ICB response. The physical interactions and abundance of LAMP3+DCs and CXCL13+T cells may shape the ICB-responsive tumor-stroma boundary, whereas CXCL14+cancer-associated fibroblasts tend to remodel extracellular matrix to form a structural barrier in non-responders. Our work therefore points out the importance of the molecular and cellular spatial structures of tumors in ICB response, raising the possibility of reprogramming tumor-stroma boundary for sensitizing immunotherapies in the majority of CRCs.

DOI: https://doi.org/10.1038/s41467-024-54710-3
Bio Protoc. 2024 Nov 20. 14(22): e5116

Preparing Chamber Slides With Pressed Collagen for Live Imaging Monolayers of Primary Human Intestinal Stem Cells.

Joseph Burclaff, Scott T Magness.

  Primary human intestinal stem cells (ISCs) can be cultured and passaged indefinitely as two-dimensional monolayers grown on soft collagen. Culturing ISCs as monolayers enables easy access to the luminal side for chemical treatments and provides a simpler topology for high-resolution imaging compared to cells cultured as three-dimensional organoids. However, the soft collagen required to support primary ISC growth can pose a challenge for live imaging with an inverted microscope, as the collagen creates a steep meniscus when poured into wells. This may lead to uneven growth toward the center of the well, with cells at the edges often extending beyond the working distance of confocal microscopes. We have engineered a 3D-printed collagen mold that enables the preparation of chamber slides with flat, smooth, and reproducible thin collagen layers. These layers are adequate to support ISC growth while being thin enough to optimize live imaging with an inverted microscope. We present methods for constructing the collagen press, preparing chamber slides with pressed collagen, and plating primary human ISCs for growth and analysis. Key features • This protocol describes how to construct and use collagen presses for chamber slides, as demonstrated in Cotton et al. [1]. • The soft collagen and culture media presented are optimized for primary human intestinal stem cells. • The full protocol, including 3D printing, preparing collagen-coated chamber slides, and plating cells can be completed in under one week. • This protocol requires access to a 3D printer.

Keywords:  Chamber slide; Collagen press; Live imaging; Microphysiological platform; Primary human intestinal stem cells

DOI:  https://doi.org/10.21769/BioProtoc.5116
Gut. 2024 Nov 28. pii: gutjnl-2024-332121. [Epub ahead of print]

Genetic variation at 11q23.1 confers colorectal cancer risk by dysregulation of colonic tuft cell transcriptional activator POU2AF2.

Vidya Rajasekaran, Bradley T Harris, Ruby T Osborn, Claire Smillie, Kevin Donnelly, Marion Bacou, Edward Esiri-Bloom, Li-Yin Ooi, Morven Allan, Marion Walker, Stuart Reid, Alison Meynert, Graeme Grimes, James P Blackmur, Peter G Vaughan-Shaw, Philip J Law, Ceres Fernández-Rozadilla, Ian Tomlinson, Richard S Houlston, Kevin B Myant, Farhat Vn Din, Maria Timofeeva, Malcolm G Dunlop, Susan M Farrington.

   BACKGROUND: Common genetic variation at 11q23.1 is associated with colorectal cancer (CRC) risk, exerting local expression quantitative trait locus (cis-eQTL) effects on POU2AF2, COLCA1 and POU2AF3 genes. However, complex linkage disequilibrium and correlated expression has hindered elucidation of the mechanisms by which genetic variants impart underlying CRC risk.
OBJECTIVE: Undertake an interdisciplinary approach to understand how variation at 11q23.1 locus imparts CRC risk.
DESIGN: We employ analysis of RNA sequencing, single-cell RNA sequencing, chromatin immunoprecipitation sequencing and single-cell ATAC sequencing data to identify, prioritise and characterise the genes that contribute to CRC risk. We further validate these findings using mouse models and demonstrate parallel effects in human colonic mucosa.
RESULTS: We establish rs3087967 as a prime eQTL variant at 11q23.1, colocalising with CRC risk. Furthermore, rs3087967 influences expression of 21 distant genes, thereby acting as a trans-eQTL hub for a gene-set highly enriched for tuft cell markers. Epigenomic analysis implicates POU2AF2 as controlling the tuft cell-specific trans-genes, through POU2F3-correlated genomic regulation. Immunofluorescence confirms rs3087967 risk genotype (T) to be associated with a tuft cell deficit in the human colon. CRISPR-mediated deletion of the 11q23.1 risk locus genes in the mouse germline exacerbated the ApcMin/+ mouse phenotype on abrogation of Pou2af2 expression specifically.
CONCLUSION: We demonstrate that genotype at rs3087967 controls a portfolio of genes through misregulation of POU2AF2. POU2AF2 is the primary transcriptional activator of tuft cells with a tumour suppressive role in mouse models. We therefore implicate tuft cells as having a key tumour-protective role in the large bowel epithelium.

Keywords:  cancer; cancer susceptibility; colorectal cancer; colorectal cancer genes

DOI:  https://doi.org/10.1136/gutjnl-2024-332121
bioRxiv. 2024 Nov 21. pii: 2024.02.20.581056. [Epub ahead of print]

Antibody-Drug Conjugates Targeting the EGFR Ligand Epiregulin Elicit Robust Anti-Tumor Activity in Colorectal Cancer.

Joan Jacob, Yasuaki Anami, Peyton High, Zhengdong Liang, Shraddha Subramanian, Sukhen C Ghosh, Solmaz AghaAmiri, Cara Guernsey-Biddle, Ha Tran, Julie H Rowe, Ali Azhdarinia, Kyoji Tsuchikama, Kendra S Carmon.

As colorectal cancer (CRC) remains a leading cause of cancer-related death, identifying therapeutic targets and approaches is essential to improve patient outcomes. The EGFR ligand epiregulin (EREG) is highly expressed in RAS wildtype and mutant CRC with minimal expression in normal tissues, making it an attractive target for antibody-drug conjugate (ADC) development. In this study, we produced and purified an EREG monoclonal antibody (mAb), H231, that had high specificity and affinity for human and mouse EREG. H231 also internalized to lysosomes, which is important for ADC payload release. ImmunoPET and ex vivo biodistribution studies showed significant tumor uptake of 89 Zr-labeled H231 with minimal uptake in normal tissues. H231 was conjugated to either cleavable dipeptide or tripeptide chemical linkers attached to the DNA-alkylating payload duocarmycin DM, and cytotoxicity of EREG ADCs was assessed in a panel of CRC cell lines. EREG ADCs incorporating tripeptide linkers demonstrated the highest potency in EREG-expressing CRC cells irrespective of RAS mutations. Preclinical safety and efficacy studies showed EREG ADCs were well-tolerated, neutralized EGFR pathway activity, caused significant tumor growth inhibition or regression, and increased survival in CRC cell line and patient-derived xenograft models. These data suggest EREG is a promising target for the development of ADCs for treating CRC and other cancer types that express high levels of EREG. While the efficacy of clinically approved anti-EGFR mAbs are largely limited by RAS mutational status, EREG ADCs may show promise for both RAS mutant and wildtype patients, thus improving existing treatment options. Significance: EREG-targeting antibody-drug conjugates demonstrate acceptable safety and robust therapeutic efficacy in RAS mutant and wildtype colorectal cancer, suggesting their potential as an alternative to EGFR-targeted therapy to benefit a broader patient population.

DOI: https://doi.org/10.1101/2024.02.20.581056
Br J Cancer. 2024 Nov 27.

Early onset metastatic colorectal cancer patients as a distinctive clinical and molecular phenomenon.

Andrea Pretta, Pina Ziranu, Eleonora Perissinotto, Filippo Ghelardi, Federica Marmorino, Riccardo Giampieri, Mariangela Puci, Maria Caterina De Grandis, Eleonora Lai, Vincenzo Nasca, Paolo Ciraci, Marco Puzzoni, Krisida Cerma, Carolina Sciortino, Ada Taravella, Gianluca Pretta, Lorenzo Giuliani, Camilla Damonte, Valeria Pusceddu, Giovanni Sotgiu, Rossana Berardi, Sara Lonardi, Francesca Bergamo, Filippo Pietrantonio, Chiara Cremolini, Mario Scartozzi.

BACKGROUND: Despite a reduction of both incidence and mortality from CRC, recent studies have shown an increase in the incidence of early-onset CRC (EO-CRC). Data on this setting are limited. The aim of our study was to evaluate the clinical and molecular profiles of metastatic EO-CRC patients in order to identify differences compared to a late-onset CRC (LO-CRC) control group.
METHODS: We retrospectively collected data from 1272 metastatic colorectal cancers from 5 different Italian Institutions. The main objective was to the evaluate clinical outcome for EO-CRC patients in comparison to patients included in the control group.
RESULTS: In the overall population, mOS was 34,7 in EO-CRC pts vs 43,0 months (mo) (p < 0,0001). In the RAS/BRAF mutated subgroup mOS in EO-CRC pts was 30,3 vs 34,0 mo (p = 0,0156). In RAS/BRAF wild-type EO-CRC mOS was 43,0 vs 50,0 mo (p = 0,0290). mPFS was 11,0 in EO-CRC pts vs 14,0 mo (p < 0,0001).
CONCLUSION: Findings indicate a general worse prognosis for patients with early-onset colorectal cancer compared to late-onset patients. Interestingly this seems to occur regardless of the molecular status. These observations might have a considerable impact on clinical practice and research.

DOI: https://doi.org/10.1038/s41416-024-02902-5
Cell. 2024 Nov 19. pii: S0092-8674(24)01270-4. [Epub ahead of print]

ACSL4 and polyunsaturated lipids support metastatic extravasation and colonization.

Yuqi Wang, Mangze Hu, Jian Cao, Fengxiang Wang, Jingrong Regina Han, Tianshu William Wu, Luxiao Li, Jinshi Yu, Yujing Fan, Guanglei Xie, Heyuan Lian, Yueying Cao, Nathchar Naowarojna, Xi Wang, Yilong Zou.

  Metastatic dissemination to distant organs demands that cancer cells possess high morphological and metabolic adaptability. However, contributions of the cellular lipidome to metastasis remain elusive. Here, we uncover a correlation between metastasis potential and ferroptosis susceptibility in multiple cancers. Metastases-derived cancer cells exhibited higher ferroptosis sensitivity and polyunsaturated fatty acyl (PUFA)-lipid contents than primary-tumor-derived cells from ovarian cancer patients. Metabolism-focused CRISPR screens in a mouse model for ovarian cancer distant metastasis established via two rounds of in vivo selection revealed the PUFA-lipid biosynthesis enzyme acyl-coenzyme A (CoA) synthetase long-chain family member 4 (ACSL4) as a pro-hematogenous metastasis factor. ACSL4 promotes metastatic extravasation by enhancing membrane fluidity and cellular invasiveness. While promoting metastasis, the high PUFA-lipid state creates dependencies on abhydrolase-domain-containing 6, acylglycerol lipase (ABHD6), enoyl-CoA delta isomerase 1 (ECI1), and enoyl-CoA hydratase 1 (ECH1)-rate-limiting enzymes preparing unsaturated fatty acids (UFAs) for β-oxidation. ACSL4/ECH1 co-inhibition achieved potent suppression of metastasis. Our work establishes the dual functions of PUFA-lipids in tumor progression and metastasis that may be exploitable for therapeutic development.

Keywords:  ACSL4; ECH1; cancer metastasis; extravasation; ferroptosis susceptibility; in vivo CRISPR screens; lipid metabolism; metastatic colonization; ovarian cancer; polyunsaturated lipids

DOI:  https://doi.org/10.1016/j.cell.2024.10.047
Neoplasia. 2024 Nov 27. pii: S1476-5586(24)00134-9. [Epub ahead of print]59 101093

TPM4 overexpression drives colon epithelial cell tumorigenesis by suppressing differentiation and promoting proliferation.

Rajsumeet S Macwan, Giulio Ferrero, Barbara Pardini, Alessio Naccarati, Piotr B Kozlowski, Michael J Papetti.

   OBJECTIVE: The high morbidity and mortality associated with colorectal cancer (CRC) and the recent increases in early-onset CRC obviate the need for novel methods to detect and treat this disease, particularly at early stages. We hypothesize that aberrant expression of genes involved in the crypt-luminal migration of colon epithelial cells, a process necessary for their growth arrest and maturation, may disrupt differentiation and transition cells from a normal to tumorigenic state.
METHODS: We searched for contractility- and motility-related genes that are dysregulated in human CRC relative to normal colon. RNA expression of one such gene, tropomyosin 4 (TPM4), was measured by qRT-PCR and RNA-seq in human colorectal tissues at various stages of tumorigenesis: CRC, adenoma, and at-risk (grossly normal mucosa from a patient with Familial Adenomatous Polyposis, or FAP), relative to controls. Effects of aberrant TPM4 expression on colon epithelial cell proliferation and maturation were determined by overexpression using stable transfection in spontaneously differentiating Caco2 cells or silencing using siRNA in proliferating cells.
RESULTS: TPM4 is overexpressed at various stages of tumorigenesis, including CRC, adenoma, and grossly normal FAP colon tissue, as well as in proliferating versus differentiating Caco2 cells. TPM4.2 overexpression in differentiating Caco2 cells markedly inhibits certain aspects of maturation, notably sucrase isomaltase and glutathione-S-transferase alpha1 expression, and causes morphological and cell junction abnormalities. Conversely, siRNA-mediated suppression of TPM4.2 inhibits Caco2 proliferation.
CONCLUSIONS: TPM4 overexpression attenuates colon epithelial cell differentiation and promotes proliferation. Therefore, TPM4 expression may be a biomarker to enhance strategies for CRC diagnosis and treatment.

Keywords:  Adenoma; Differentiation; Familial adenomatous polyposis; Human colorectal cancer; Proliferation; Tropomyosin, Cell:cell junctions

DOI:  https://doi.org/10.1016/j.neo.2024.101093
Int J Mol Sci. 2024 Nov 08. pii: 12007. [Epub ahead of print]25(22):

Hedgehog Signalling Pathway and Its Role in Shaping the Architecture of Intestinal Epithelium.

Adrianna Konopka, Kamil Gawin, Marcin Barszcz.

  The hedgehog (Hh) signalling pathway plays a key role in both embryonic and postnatal development of the intestine and is responsible for gut homeostasis. It regulates stem cell renewal, formation of the villous-crypt axis, differentiation of goblet and Paneth cells, the cell cycle, apoptosis, development of gut innervation, and lipid metabolism. Ligands of the Hh pathway, i.e., Indian hedgehog (Ihh) and Sonic hedgehog (Shh), are expressed by superficial enterocytes but act in the mesenchyme, where they are bound by a Patched receptor localised on myofibroblasts and smooth muscle cells. This activates a cascade leading to the transcription of target genes, including those encoding G1/S-specific cyclin-D2 and -E1, B-cell lymphoma 2, fibroblast growth factor 4, and bone morphogenetic protein 4. The Hh pathway is tightly connected to Wnt signalling. Ihh is the major ligand in the Hh pathway. Its activation inhibits proliferation, while its blocking induces hyperproliferation and triggers a wound-healing response. Thus, Ihh is a negative feedback regulator of cell proliferation. There are data indicating that diet composition may affect the expression of the Hh pathway genes and proteins, which in turn, induces changes in mucosal architecture. This was shown for fat, vitamin A, haem, berberine, and ovotransferrin. The Hh signalling is also affected by the intestinal microbiota, which affects the intestinal barrier integrity. This review highlights the critical importance of the Hh pathway in shaping the intestinal mucosa and summarises the results obtained so far in research on the effect of dietary constituents on the activity of this pathway.

Keywords:  Indian hedgehog; Sonic hedgehog; crypt; intestine; mesenchyme; morphogen; nutrition; proliferation; villous

DOI:  https://doi.org/10.3390/ijms252212007
FEBS J. 2024 Nov 25.

AXIN2 is a non-redundant regulator of AXIN1 stability and β-catenin in colorectal cancer cells.

Lin Liu, John Silke.

  AXIN proteins are major components of the β-catenin destruction complex or degradasome, which limits β-catenin nuclear translocation and Wnt signalling activation at steady state. Schmidt et al. performed quantitative analysis of cellular AXIN protein levels in human colorectal cancer cells and revealed that AXIN2 plays a non-redundant role in regulating the total AXIN pool and Wnt/β-catenin signalling activity. Tankyrase (TNKS) inhibitors failed to inhibit Wnt/β-catenin signalling in AXIN2 knockout cells, suggesting that AXIN2 is essential for TNKS inhibitors to function. Mechanistically, the authors show that AXIN2 recruits TNKS to AXIN1 and promotes TNKS-mediated degradation of AXIN1. These findings may have important implications for anti-cancer therapy by TNKS small molecule inhibitors.

Keywords:  AXIN1; AXIN2; Wnt/β‐catenin signalling; colorectal cancer; tankyrase; tankyrase inhibitor

DOI:  https://doi.org/10.1111/febs.17336
Mol Cell Biochem. 2024 Nov 25.

Oxaliplatin and 5-fluorouracil promote epithelial-mesenchymal transition via activation of KRAS/ERK/NF-κB pathway in KRAS-mutated colon cancer cells.

Tadafumi Hoshida, Masanobu Tsubaki, Tomoya Takeda, Ryota Asano, Ik-Hyun Choi, Koudai Takimoto, Ayano Inukai, Motohiro Imano, Kazufumi Tanabe, Noriaki Nagai, Shozo Nishida.

  Oxaliplatin (L-OHP) and 5-fluorouracil (5-FU) are used to treat colon cancer; however, resistance contributes to poor prognosis. Epithelial-mesenchymal transition (EMT) has been induced in tumor tissues after administration of anticancer drugs and may be involved in drug resistance. We investigated the mechanism of EMT induction in colon cancer cells treated with 5-FU and L-OHP. We found that L-OHP and 5-FU at clinical steady-state concentrations induced EMT in LoVo and DLD-1 cells (KRAS G13D-mutated), but not in HT-29 and Caco-2 cells (KRAS wild type). L-OHP and 5-FU elevated vimentin, N-cadherin, Twist, Slug, and Snail and decreased E-cadherin expressions. Moreover, 5-FU- and L-OHP -induced EMT cells showed increased cell migration and decreased sensitivity to 5-FU and L-OHP. L-OHP and 5-FU treatment promoted KRAS, ERK1/2, and NF-κB activation. Combined administration with KRAS siRNA, MEK1/2 inhibitor trametinib, and NF-κB inhibitor dimethyl fumarate (DMF), suppressed L-OHP- and 5-FU-induced EMT. These results suggest that KRAS/ERK/NF-κB pathway activation is important for EMT induction by L-OHP and 5-FU treatment. Thus, MEK1/2 and NF-κB inhibitors may facilitate the resistance acquisition to L-OHP and 5-FU therapy in KRAS G13D-mutated colon cancer.

Keywords:  5-fluorouracil; Colon cancer; Epithelial-mesenchymal transition; KRAS; Oxaliplatin

DOI:  https://doi.org/10.1007/s11010-024-05157-z
Biochem J. 2024 Nov 27. pii: BCJ20240445. [Epub ahead of print]

Loss of peroxiredoxin 6 (PRDX6) alters lipid composition and distribution resulting in increased sensitivity to ferroptosis.

Daniel J Lagal, Ángel Ortiz-Alcántara, José R Pedrajas, Brian McDonagh, J Antonio Bárcena, Raquel Requejo-Aguilar, C Alicia Padilla.

  Peroxiredoxin 6 (PRDX6) is a multifunctional enzyme involved in phospholipid peroxide repair and metabolism. In this study we investigated the global lipid composition of a human hepatocarcinoma cell line SNU475 lacking PRDX6 and lipid related cellular processes. There was a general decrease in multiple lipids species upon loss of PRDX6, in particular sphingomyelins and acylcarnitines, consistent with previously observed alterations in cell signaling pathways and mitochondrial dysfunction. Deprivation of docosahexaenoic acid and related species was also evident. However, a few striking exceptions are worth highlighting: 1) Three specific arachidonic acid (AA) containing phophatidylcholines (PC) increased significantly. The increase of sn1-stearic/sn2-PUFA containing PC and sn2-AA containing plasmenyls are indicative of a preference of PRDX6 iPLA2 activity for these AA storage glycerophospholipids. 2) Several polyunsaturated fatty acids (PUFA) and PUFA containing triacylglycerols accumulated together with increased formation of lipid droplets, an indication of altered FA flux and PUFA sequestration in PRDX6 knockout cells. Loss of PRDX6 resulted in increased sensitivity to erastin-induced ferroptosis, independent of selenium and GPX4, as a consequence of increased levels of lipid hydroperoxides, that reverted to normal levels upon rescue with PRDX6. <SPAN style="font-weight: 400;">The results presented demonstrate that all three enzymatic activities of PRDX6 contribute to the role of this multifunctional enzyme in diverse cellular processes, including membrane phospholipid remodeling and glycerophospholipid functional diversity, resulting in altered lipid peroxides and modulation of AA disposition and traffic. These contributions highlight the complexity of the changes that loss of PRDX6 exerts on cell functionality.</SPAN>.

Keywords:  Lipid peroxidation; ferroptosis; lipid droplet; lipid raft; lysophospholipids; phospholipase A2

DOI:  https://doi.org/10.1042/BCJ20240445
bioRxiv. 2024 Nov 15. pii: 2024.11.13.623500. [Epub ahead of print]

Basal cell of origin resolves neuroendocrine-tuft lineage plasticity in cancer.

Abbie S Ireland, Sarah B Hawgood, Daniel A Xie, Margaret W Barbier, Scarlett Lucas-Randolph, Darren R Tyson, Lisa Y Zuo, Benjamin L Witt, Ramaswamy Govindan, Afshin Dowlati, Justin C Moser, Sonam Puri, Charles M Rudin, Joseph M Chan, Andrew Elliott, Trudy G Oliver.

Neuroendocrine and tuft cells are rare, chemosensory epithelial lineages defined by expression of ASCL1 and POU2F3 transcription factors, respectively 1,2 . Neuroendocrine cancers, including small cell lung cancer (SCLC), frequently display tuft-like subsets, a feature linked to poor patient outcomes 3-13 . The mechanisms driving neuroendocrine-tuft tumour heterogeneity, and the origins of tuft-like cancers are unknown. Using multiple genetically-engineered animal models of SCLC, we demonstrate that a basal cell of origin (but not the accepted neuroendocrine origin) generates neuroendocrine-tuft-like tumours that highly recapitulate human SCLC. Single-cell clonal analyses of basal-derived SCLC further uncovers unexpected transcriptional states and lineage trajectories underlying neuroendocrine-tuft plasticity. Uniquely in basal cells, introduction of genetic alterations enriched in human tuft-like SCLC, including high MYC, PTEN loss, and ASCL1 suppression, cooperate to promote tuft-like tumours. Transcriptomics of 944 human SCLCs reveal a basal-like subset and a tuft-ionocyte-like state that altogether demonstrate remarkable conservation between cancer states and normal basal cell injury response mechanisms 14-18 . Together, these data suggest that the basal cell is a plausible origin for SCLC and other neuroendocrine-tuft cancers that can explain neuroendocrine-tuft heterogeneity-offering new insights for targeting lineage plasticity.

DOI: https://doi.org/10.1101/2024.11.13.623500
Biochim Biophys Acta Rev Cancer. 2024 Nov 25. pii: S0304-419X(24)00156-2. [Epub ahead of print] 189225

Targeting extracellular matrix interaction in gastrointestinal cancer: Immune modulation, metabolic reprogramming, and therapeutic strategies.

Jiyifan Li, Wenxin Zhang, Lu Chen, Xinhai Wang, Jiafeng Liu, Yuxin Huang, Huijie Qi, Li Chen, Tianxiao Wang, Qunyi Li.

  The extracellular matrix (ECM) is a major constituent of the tumor microenvironment, acting as a mediator that supports the progression of gastrointestinal (GI) cancers, particularly in mesenchymal subtypes. Beyond providing structural support, the ECM actively shapes the tumor microenvironment (TME) through complex biochemical and biomechanical remodeling. Dysregulation of ECM composition and signaling is closely linked to increased cancer aggressiveness, poor prognosis, and resistance to therapy. ECM components, such as collagen, fibronectin, laminin, and periostin, influence tumor growth, metastasis, immune modulation, and metabolic reprogramming by interacting with tumor cells, immune cells, and cancer-associated fibroblasts. In this review, we highlight the heterogeneous nature of the ECM and the dualistic roles of its components across GI cancers, with a focus on their contributions to immune evasion and metabolic remodeling via intercellular interactions. Additionally, we explore therapeutic strategies targeting ECM remodeling and ECM-centered interactions, emphasizing their potential in enhancing existing anti-tumor therapies.

Keywords:  Extracellular matrix; Gastrointestinal cancer; Immunotherapy; Tumor metabolism; Tumor microenvironment; cancer-associated fibroblast

DOI:  https://doi.org/10.1016/j.bbcan.2024.189225
Elife. 2024 Nov 28. pii: RP97827. [Epub ahead of print]13

CPT1A mediates radiation sensitivity in colorectal cancer.

Zhenhui Chen, Lu Yu, Zhihao Zheng, Xusheng Wang, Qiqing Guo, Yuchuan Chen, Yaowei Zhang, Yuqin Zhang, Jianbiao Xiao, Keli Chen, Hongying Fan, Yi Ding.

  The prevalence and mortality rates of colorectal cancer (CRC) are increasing worldwide. Radiation resistance hinders radiotherapy, a standard treatment for advanced CRC, leading to local recurrence and metastasis. Elucidating the molecular mechanisms underlying radioresistance in CRC is critical to enhance therapeutic efficacy and patient outcomes. Bioinformatic analysis and tumour tissue examination were conducted to investigate the CPT1A mRNA and protein levels in CRC and their correlation with radiotherapy efficacy. Furthermore, lentiviral overexpression and CRISPR/Cas9 lentiviral vectors, along with in vitro and in vivo radiation experiments, were used to explore the effect of CPT1A on radiosensitivity. Additionally, transcriptomic sequencing, molecular biology experiments, and bioinformatic analyses were employed to elucidate the molecular mechanisms by which CPT1A regulates radiosensitivity. CPT1A was significantly downregulated in CRC and negatively correlated with responsiveness to neoadjuvant radiotherapy. Functional studies suggested that CPT1A mediates radiosensitivity, influencing reactive oxygen species (ROS) scavenging and DNA damage response. Transcriptomic and molecular analyses highlighted the involvement of the peroxisomal pathway. Mechanistic exploration revealed that CPT1A downregulates the FOXM1-SOD1/SOD2/CAT axis, moderating cellular ROS levels after irradiation and enhancing radiosensitivity. CPT1A downregulation contributes to radioresistance in CRC by augmenting the FOXM1-mediated antioxidant response. Thus, CPT1A is a potential biomarker of radiosensitivity and a novel target for overcoming radioresistance, offering a future direction to enhance CRC radiotherapy.

Keywords:  CPT1A; FOXM1; cancer biology; colorectal cancer cells; human; mouse; radiosensitivity; reactive oxygen species

DOI:  https://doi.org/10.7554/eLife.97827
bioRxiv. 2024 Nov 11. pii: 2024.11.07.622577. [Epub ahead of print]

The single-cell spatial landscape of stage III colorectal cancers.

Andrew Su, HoJoon Lee, Minh Tran, Richard D Cruz, Anuja Sathe, Xiangqi Bai, Ignacio Wichmann, Lance Pflieger, Bryce Moulton, Tyler Barker, Derrick Haslem, David Jones, Lincoln Nadauld, Quan Nguyen, Hanlee P Ji, Terence Rhodes.

We conducted a spatial analysis using imaging mass cytometry applied to stage III colorectal adenocarcinomas. This study used multiplexed markers to distinguish individual cells and their spatial organization from 52 colorectal cancers. We determined the landscape features of cellular spatial features in the CRC tumor microenvironment. This spatial single-cell analysis identified 10 unique cell phenotypes in the tumor microenvironment that included stromal and immune cells with a subset which had a proliferative phenotype. These special features included spatial neighborhood interactions between single cells as well as different tissue niches, especially the tumor infiltrating lymphocyte regions. We applied a robust statistical analysis to identify significant correlations of cell features with phenotypes such as microsatellite instability or recurrence. We determined that microsatellite stable (MSS) colorectal cancers had an increased risk of recurrence if they had the following features: 1) a low level of stromal tumor-infiltrating lymphocytes, and 2) low interactions between CD4+ T cells and stromal cells. Our results point to the utility of spatial single-cell interaction analysis in defining novel features of the tumor immune microenvironments and providing useful clinical cell-related spatial biomarkers.

DOI: https://doi.org/10.1101/2024.11.07.622577
J Trace Elem Med Biol. 2024 Nov 15. pii: S0946-672X(24)00184-6. [Epub ahead of print]86 127564

Classification of colorectal cancer patients based on serum micronutrients: An exploratory investigation.

Krishnamurthy Hari, Rosanna Squitti, Jophi Bosco, Vasanth Jayaraman, Karthik Krishna, Amit Pal, Anastasia De Luca, Laura Di Veroli, Gioia Mastromoro, Gianluca Rizzo, Vincenzo Tondolo, Mauro Rongioletti.

   BACKGROUND: Colorectal cancer (CRC) is a growing global health challenge with a multifactorial etiology encompassing genetic susceptibility, nutrition, and inflammation in the bowel.
OBJECTIVE: To examine micronutrient status in CRC patients undergoing CRC resection.
DESIGN: We performed a case-control study including 13 consecutive CRC patients and 10 healthy controls (CTRL) comparing the serum levels of 29 micronutrients, namely Copper, Zinc, Selenium, Chromium, Manganese, Carnitine, Choline, Inositol, Methylmalonic acid (MMA), Vitamin (Vit) B1, Vit B2, Vit B3, Vit B5, Vit B6, Vit C, Vit A, Vit D3, Vit E, Vit K1, Vit K2 and the amino acids Serine, Valine, Leucine, Isoleucine, Asparagine, Glutamine, Arginine, Citrulline and Cysteine.
RESULTS: After considering the effect of age and sex, copper, arginine, and cysteine were increased, while zinc, selenium, chromium, Vit B1, Vit K1, and Vit A were decreased in CRC patients in comparison with CTRL. Zinc levels perfectly predicted the diagnosis of CRC, and were associated with lymph nodes (pN), of the pTNM staging. Copper levels in serum were strongly associated with the pathological pTNM staging of CRC.
CONCLUSION: Though this is a preliminary study that needs confirmation with a larger longitudinal cohort, our results show that serum micronutrients are linked to tumor growth, likely caused by increased demand from tumor tissue associated with an aberrant cell proliferation and changes in the antioxidant function.

Keywords:  Colorectal cancer; Cysteine; Micronutrients copper; Selenium; Vitamin B1; Zinc; pTNM

DOI:  https://doi.org/10.1016/j.jtemb.2024.127564