bims-instec 2021-12-26 papers

bims-instec

Biomed News

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

Issue of 2021–12–26
ten papers selected by
Maria-Virginia Giolito, IRFAC/UMR-S1113 INSERM

MLH1 mediates cytoprotective nucleophagy to resist 5-Fluorouracil-induced cell death in colorectal carcinoma.
PHLDB2 mediates cetuximab resistance via interacting with EGFR in latent metastasis of colorectal cancer.
Desmoplakin maintains transcellular keratin scaffolding and protects from intestinal injury.
Targeted deletion of keratin 8 in intestinal epithelial cells disrupts tissue integrity and predisposes to tumorigenesis in the colon.
Reuterin in the healthy gut microbiome suppresses colorectal cancer growth through altering redox balance.
Colorectal cancer-associated fibroblasts promote metastasis by up-regulating LRG1 through stromal IL-6/STAT3 signaling.
The Colorectal Cancer Tumor Microenvironment and Its Impact on Liver and Lung Metastasis.
Therapeutic Targets of KRAS in Colorectal Cancer.
Spatial discordances between mRNAs and proteins in the intestinal epithelium.
Mesalazine initiates an anti-oncogenic β-catenin / MUCDHL negative feed-back loop in colon cancer cells by cell-specific mechanisms.

Neoplasia. 2021 Dec 21. pii: S1476-5586(21)00107-X. [Epub ahead of print]24(2): 76-85

MLH1 mediates cytoprotective nucleophagy to resist 5-Fluorouracil-induced cell death in colorectal carcinoma.

Shaista Manzoor, Maha Saber-Ayad, Azzam A Maghazachi, Qutayba Hamid, Jibran Sualeh Muhammad.

  Colorectal Cancer (CRC) with Microsatellite instability (MSI) and mutLhomolog-1 (MLH1) gene deficiency are less aggressive than MLH1 proficient cancers. MLH1 is involved in several cellular processes, but its connection with the autophagy-dependent cellular response towards anticancer drugs remains unclear. In this study, we aimed to investigate the interaction between MLH1 and the autophagy marker LC3, which facilitated nucleophagy induction, and its potential role in determining sensitivity to 5-Fluorouracil (5-FU) induced cell death. To examine the role of MLH1 in DNA-damage-induced nucleophagy in CRC cells, we utilized a panel of MLH1 deficient and MLH1 proficient CRC cell lines. We included a parental HCT116 cell line (MLH1-/-) and its isogenic cell line HCT116 MLH1+/- in which a single allele of the MLH1 gene was introduced using CRISPR-Cas9 gene editing. We observed that MLH1 proficient cells were less sensitive to the 5-FU-induced cytotoxic effect. The 5-FU induced DNA damage led to LC3 up-regulation, which was dependent on MLH1 overexpression. Moreover, immunofluorescence and immunoprecipitation data showed LC3 and MLH1 were colocalized in CRC cells. Consequently, MLH1 dependent 5-FU-induced DNA damage contributed to the formation of micronuclei. These micronuclei colocalize with autolysosome, indicating a cytoprotective role of MLH1 dependent nucleophagy. Interestingly, siRNA knockdown of MLH1 in HCT116 MLH1+/- prevented LC3 upregulation and micronuclei formation. These novel data are the first to show an essential role of MLH1 in mediating the chemoresistance and survival of cancer cells by increasing the LC3 expression and inducing nucleophagy in 5-FU treated CRC cells.

Keywords:  Autophagy; CRISPR-Cas9; Chemoresistance; Colorectal cancer; LC3; Lamin; Microsatellite instability; Mismatch Repair; Nucleophagy; SIRT1

DOI:  https://doi.org/10.1016/j.neo.2021.12.003
Cell Mol Gastroenterol Hepatol. 2021 Dec 21. pii: S2352-345X(21)00261-7. [Epub ahead of print]

PHLDB2 mediates cetuximab resistance via interacting with EGFR in latent metastasis of colorectal cancer.

Maochao Luo, Zhao Huang, Xingyue Yang, Yan Chen, Jingwen Jiang, Lu Zhang, Li Zhou, Siyuan Qin, Ping Jin, Shuyue Fu, Liyuan Peng, Bowen Li, Yongting Fang, Wenchen Pu, Yanqiu Gong, Yu Liu, Zhixiang Ren, Qiu-Luo Liu, Cun Wang, Fangqiong Xiao, Du He, Hongying Zhang, Changlong Li, Heng Xu, Lunzhi Dai, Yong Peng, Zong-Gung Zhou, Canhua Huang, Hai-Ning Chen.

   BACKGROUND & AIMS: Latent metastasis of colorectal cancer (CRC) frequently develops months or years after primary surgery followed by adjuvant therapies and may progress rapidly even with targeted therapy administered, but the underlying mechanism remains unclear. Here, we aim to explore the molecular basis for the aggressive behavior of latent metastasis in CRC.
METHODS: Transcriptional profiling and pathway enrichment analysis of paired primary and metastatic tumor samples were performed. The underlying mechanisms of PHLDB2 in CRC were investigated by RNA immunoprecipitation assay, immunohistochemistry, mass spectrometry analysis and Duolink in situ proximity ligation assay. The efficacy of targeting PHLDB2 in cetuximab treatment was elucidated in CRC cell lines and mouse models.
RESULTS: Based on the transcriptional profile of paired primary and metastatic tumor samples, we identified PHLDB2 as a potential regulator in latent liver metastasis. A detailed mechanistic study revealed that chemotherapeutic agent-induced oxidative stress promotes METTL14-mediated m6A modification of PHLDB2 mRNA, facilitating its protein expression. Upregulated PHLDB2 stabilizes EGFR and promotes its nuclear translocation, which in turn results in EGFR signaling activation and consequent cetuximab resistance. Moreover, Arg1163 (R1163) of PHLDB2 is crucial for interaction with EGFR, and the R1163A mutation abrogates its regulatory function in EGFR signaling.
CONCLUSIONS: PHLDB2 plays a crucial role in cetuximab resistance and is proposed to be a potential target for the treatment of CRC.

Keywords:  Cetuximab resistance; Colorectal cancer; EGFR; Latent metastasis; PHLDB2

DOI:  https://doi.org/10.1016/j.jcmgh.2021.12.011
Cell Mol Gastroenterol Hepatol. 2021 Dec 17. pii: S2352-345X(21)00259-9. [Epub ahead of print]

Desmoplakin maintains transcellular keratin scaffolding and protects from intestinal injury.

Annika Gross, Biaohuan Zhou, Lisa Bewersdorf, Nicole Schwarz, Gabriel M Schacht, Peter Boor, Konrad Hoeft, Bernd Hoffmann, Elaine Fuchs, Rafael Kramann, Rudolf Merkel, Rudolf E Leube, Pavel Strnad.

   BACKGROUND AND AIM: Desmosomes are intercellular junctions connecting keratin intermediate filaments of neighboring cells. The cadherins desmoglein 2 (Dsg2) and desmocollin 2 mediate cell-cell adhesion, whereas desmoplakin (Dsp) provides the attachment of desmosomes to keratins. While the importance of the desmosome-keratin-network is well established in mechanically challenged tissues, we aimed to assess the currently understudied function of desmosomal proteins in intestinal epithelia.
METHODS: We analyzed the intestine-specific villin-Cre DSP (DSPΔIEC) and the combined, intestine-specific DSG2/DSPΔIEC (ΔDsg2/Dsp) knockout mice. Cross-breeding with keratin 8 (K8)-YFP knock-in mice and generation of organoids was performed to visualize the keratin network. A Dsp-deficient colorectal carcinoma HT29-derived cell line was generated and the role of Dsp in adhesion and mechanical stress was studied in dispase assays, after exposure to uniaxial cell stretching and during scratch assay.
RESULTS: The intestine of DSPΔIEC mice was histopathologically inconspicuous. Intestinal epithelial cells, however, displayed an accelerated migration along the crypt and an enhanced shedding into the lumen. Increased intestinal permeability and altered levels of desmosomal proteins were detected. An inconspicuous phenotype was also seen in ΔDsg2/Dsp mice. After dextran sodium sulfate treatment, DSPΔIEC mice developed a more pronounced colitis. A retracted keratin network was seen in the intestinal epithelium of DSPΔIEC/K8-YFP mice and organoids derived from these mice presented a collapsed keratin network. The level, phosphorylation status, and solubility of keratins were not affected. Dsp-deficient HT29 cells had an impaired cell adhesion and suffered from increased cellular damage after stretch.
CONCLUSIONS: Our results demonstrate that Dsp is required for proper keratin network architecture in intestinal epithelia, mechanical resilience and adhesion thereby protecting from injury.

Keywords:  apical junctional complex; cell adhesion; desmosome; intestinal epithelial barrier; keratin

DOI:  https://doi.org/10.1016/j.jcmgh.2021.12.009
Cell Mol Life Sci. 2021 Dec 24. 79(1): 10

Targeted deletion of keratin 8 in intestinal epithelial cells disrupts tissue integrity and predisposes to tumorigenesis in the colon.

Carl-Gustaf A Stenvall, Mina Tayyab, Tove J Grönroos, Maria A Ilomäki, Keijo Viiri, Karen M Ridge, Lauri Polari, Diana M Toivola.

  Keratin 8 (K8) is the main intestinal epithelial intermediate filament protein with proposed roles for colonic epithelial cell integrity. Here, we used mice lacking K8 in intestinal epithelial cells (floxed K8 and Villin-Cre1000 and Villin-CreERt2) to investigate the cell-specific roles of intestinal epithelial K8 for colonocyte function and pathologies. Intestinal epithelial K8 deletion decreased K8 partner proteins, K18-K20, 75-95%, and the remaining keratin filaments were located at the colonocyte apical regions with type II K7, which decreased 30%. 2-Deoxy-2-[18F]-fluoroglucose positron emission tomography in vivo imaging identified a metabolic phenotype in the lower gut of the conditional K8 knockouts. These mice developed intestinal barrier leakiness, mild diarrhea, and epithelial damage, especially in the proximal colon. Mice exhibited shifted differentiation from enterocytes to goblet cells, displayed longer crypts and an increased number of Ki67 + transit-amplifying cells in the colon. Significant proproliferative and regenerative signaling occurred in the IL-22, STAT3, and pRb pathways, with minor effects on inflammatory parameters, which, however, increased in aging mice. Importantly, colonocyte K8 deletion induced a dramatically increased sensitivity to azoxymethane-induced tumorigenesis. In conclusion, intestinal epithelial K8 plays a significant role in colonocyte epithelial integrity maintenance, proliferation regulation and tumor suppression.

Keywords:  Barrier; Colon cancer; Goblet cell; Notch; Proliferation; Tumorigenesis; Villin-Cre

DOI:  https://doi.org/10.1007/s00018-021-04081-5
Cancer Cell. 2021 Dec 14. pii: S1535-6108(21)00613-9. [Epub ahead of print]

Reuterin in the healthy gut microbiome suppresses colorectal cancer growth through altering redox balance.

Hannah N Bell, Ryan J Rebernick, Joshua Goyert, Rashi Singhal, Miljan Kuljanin, Samuel A Kerk, Wesley Huang, Nupur K Das, Anthony Andren, Sumeet Solanki, Shannon L Miller, Peter K Todd, Eric R Fearon, Costas A Lyssiotis, Steven P Gygi, Joseph D Mancias, Yatrik M Shah.

  Microbial dysbiosis is a colorectal cancer (CRC) hallmark and contributes to inflammation, tumor growth, and therapy response. Gut microbes signal via metabolites, but how the metabolites impact CRC is largely unknown. We interrogated fecal metabolites associated with mouse models of colon tumorigenesis with varying mutational load. We find that microbial metabolites from healthy mice or humans are growth-repressive, and this response is attenuated in mice and patients with CRC. Microbial profiling reveals that Lactobacillus reuteri and its metabolite, reuterin, are downregulated in mouse and human CRC. Reuterin alters redox balance, and reduces proliferation and survival in colon cancer cells. Reuterin induces selective protein oxidation and inhibits ribosomal biogenesis and protein translation. Exogenous Lactobacillus reuteri restricts colon tumor growth, increases tumor reactive oxygen species, and decreases protein translation in vivo. Our findings indicate that a healthy microbiome and specifically, Lactobacillus reuteri, is protective against CRC through microbial metabolite exchange.

Keywords:  Lactobacillus reuteri; Microbiome; Reuterin; colorectal cancer; metabolites; protein oxidation

DOI:  https://doi.org/10.1016/j.ccell.2021.12.001
Cell Death Dis. 2021 Dec 20. 13(1): 16

Colorectal cancer-associated fibroblasts promote metastasis by up-regulating LRG1 through stromal IL-6/STAT3 signaling.

Beiping Zhong, Bing Cheng, Xiaoming Huang, Qian Xiao, Zhitong Niu, Yu-Feng Chen, Qiang Yu, Wenyu Wang, Xiao-Jian Wu.

Cancer-associated fibroblasts (CAFs) have been shown to play a strong role in colorectal cancer metastasis, yet the underlying mechanism remains to be fully elucidated. Using CRC clinical samples together with ex vivo CAFs-CRC co-culture models, we found that CAFs induce expression of Leucine Rich Alpha-2-Glycoprotein 1(LRG1) in CRC, where it shows markedly higher expression in metastatic CRC tissues compared to primary tumors. We further show that CAFs-induced LRG1 promotes CRC migration and invasion that is concomitant with EMT (epithelial-mesenchymal transition) induction. In addition, this signaling axis has also been confirmed in the liver metastatic mouse model which displayed CAFs-induced LRG1 substantially accelerates metastasis. Mechanistically, we demonstrate that CAFs-secreted IL-6 (interleukin-6) is responsible for LRG1 up-regulation in CRC, which occurs through a direct transactivation by STAT3 following JAK2 activation. In clinical CRC tumor samples, LRG1 expression was positively correlated with CAFs-specific marker, α-SMA, and a higher LRG1 expression predicted poor clinical outcomes especially distant metastasis free survival, supporting the role of LRG1 in CRC progression. Collectively, this study provided a novel insight into CAFs-mediated metastasis in CRC and indicated that therapeutic targeting of CAFs-mediated IL-6-STAT3-LRG1 axis might be a potential strategy to mitigate metastasis in CRC.

DOI: https://doi.org/10.1038/s41419-021-04461-6
Cancers (Basel). 2021 Dec 09. pii: 6206. [Epub ahead of print]13(24):

The Colorectal Cancer Tumor Microenvironment and Its Impact on Liver and Lung Metastasis.

Raghav Chandra, John D Karalis, Charles Liu, Gilbert Z Murimwa, Josiah Voth Park, Christopher A Heid, Scott I Reznik, Emina Huang, John D Minna, Rolf A Brekken.

  Colorectal cancer (CRC) is the third most common malignancy and the second most common cause of cancer-related mortality worldwide. A total of 20% of CRC patients present with distant metastases, most frequently to the liver and lung. In the primary tumor, as well as at each metastatic site, the cellular components of the tumor microenvironment (TME) contribute to tumor engraftment and metastasis. These include immune cells (macrophages, neutrophils, T lymphocytes, and dendritic cells) and stromal cells (cancer-associated fibroblasts and endothelial cells). In this review, we highlight how the TME influences tumor progression and invasion at the primary site and its function in fostering metastatic niches in the liver and lungs. We also discuss emerging clinical strategies to target the CRC TME.

Keywords:  colorectal cancer; colorectal liver metastasis; colorectal pulmonary metastasis; immuno-oncology; novel anticancer therapy; tumor microenvironment

DOI:  https://doi.org/10.3390/cancers13246206
Cancers (Basel). 2021 Dec 11. pii: 6233. [Epub ahead of print]13(24):

Therapeutic Targets of KRAS in Colorectal Cancer.

Shafia Rahman, Shimon Garrel, Michael Gerber, Radhashree Maitra, Sanjay Goel.

  Patients with metastatic colorectal cancer have a 5-year overall survival of less than 10%. Approximately 45% of patients with metastatic colorectal cancer harbor KRAS mutations. These mutations not only carry a predictive role for the absence of response to anti-EGFR therapy, but also have a negative prognostic impact on the overall survival. There is a growing unmet need for a personalized therapy approach for patients with KRAS-mutant colorectal cancer. In this article, we focus on the therapeutic strategies targeting KRAS- mutant CRC, while reviewing and elaborating on the discovery and physiology of KRAS.

Keywords:  KRAS mutation; colorectal cancer; targeted therapy

DOI:  https://doi.org/10.3390/cancers13246233
Nat Metab. 2021 Dec;3(12): 1680-1693

Spatial discordances between mRNAs and proteins in the intestinal epithelium.

Yotam Harnik, Lisa Buchauer, Shani Ben-Moshe, Inna Averbukh, Yishai Levin, Alon Savidor, Raya Eilam, Andreas E Moor, Shalev Itzkovitz.

The use of transcriptomes as reliable proxies for cellular proteomes is controversial. In the small intestine, enterocytes operate for 4 days as they migrate along villi, which are highly graded microenvironments. Spatial transcriptomics have demonstrated profound zonation in enterocyte gene expression, but how this variability translates to protein content is unclear. Here we show that enterocyte proteins and messenger RNAs along the villus axis are zonated, yet often spatially discordant. Using spatial sorting with zonated surface markers, together with a Bayesian approach to infer protein translation and degradation rates from the combined spatial profiles, we find that, while many genes exhibit proteins zonated toward the villus tip, mRNA is zonated toward the villus bottom. Finally, we demonstrate that space-independent protein synthesis delays can explain many of the mRNA-protein discordances. Our work provides a proteomic spatial blueprint of the intestinal epithelium, highlighting the importance of protein measurements for inferring cell states in tissues that operate outside of steady state.

DOI: https://doi.org/10.1038/s42255-021-00504-6
Biomed Pharmacother. 2021 Dec 17. pii: S0753-3322(21)01330-5. [Epub ahead of print]146 112543

Mesalazine initiates an anti-oncogenic β-catenin / MUCDHL negative feed-back loop in colon cancer cells by cell-specific mechanisms.

Emilie Bersuder, Chloe Terciolo, Mathilde Lechevrel, Elisabeth Martin, Celine Quesnelle, Jean-Noel Freund, Jean-Marie Reimund, Isabelle Gross.

  Chronic inflammation associated with intestinal architecture and barrier disruption puts patients with inflammatory bowel disease (IBD) at increased risk of developing colorectal cancer (CRC). Widely used to reduce flares of intestinal inflammation, 5-aminosalicylic acid derivatives (5-ASAs) such as mesalazine appear to also exert more direct mucosal healing and chemopreventive activities against CRC. The mechanisms underlying these activities are poorly understood and may involve the up-regulation of the cadherin-related gene MUCDHL (CDHR5). This atypical cadherin is emerging as a new actor of intestinal homeostasis and opposes colon tumorigenesis. Here, we showed that mesalazine increase mRNA levels of MUCDHL and of other genes involved in the intestinal barrier function in most intestinal cell lines. In addition, using gain / loss of function experiments (agonists, plasmid or siRNAs transfections), luciferase reporter genes and chromatin immunoprecipitation, we thoroughly investigated the molecular mechanisms triggered by mesalazine that lead to the up-regulation of MUCDHL expression. We found that basal transcription of MUCDHL in different CRC cell lines is regulated positively by CDX2 and negatively by β-catenin through a negative feed-back loop. However, mesalazine-stimulation of MUCDHL transcription is controlled by cell-specific mechanisms, involving either enhanced activation of CDX2 and PPAR-γ or repression of the β-catenin inhibitory effect. This work highlights the importance of the cellular and molecular context in the activity of mesalazine and suggests that its efficacy against CRC depends on the genetic alterations of transformed cells.

Keywords:  5-aminosalicylic acid; Adhesion; CDHR5; Colorectal neoplasm; Ulcerative Colitis; WNT pathway

DOI:  https://doi.org/10.1016/j.biopha.2021.112543