bims-instec 2022-09-11 papers

bims-instec

Biomed News

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

Issue of 2022–09–11
twenty-one papers selected by
Maria-Virginia Giolito, IRFAC/UMR-S1113 INSERM

Intestinal epithelial organoids: regeneration and maintenance of the intestinal epithelium.
Modeling Colorectal Cancer Progression Reveals Niche-Dependent Clonal Selection.
Patient-Derived Organoids from Colorectal Cancer with Paired Liver Metastasis Reveal Tumor Heterogeneity and Predict Response to Chemotherapy.
Long-term resistance to 5-fluorouracil promotes epithelial-mesenchymal transition, apoptosis evasion, autophagy, and reduced proliferation rate in colon cancer cells.
From stem to stern: LGR5+ as a marker for intestinal crypts.
In vivo tumor immune microenvironment phenotypes correlate with inflammation and vasculature to predict immunotherapy response.
Modelling Biochemical Gradients in Vitro to Control Cell Compartmentalization in a Microengineered 3D Model of the Intestinal Epithelium.
Small leucine zipper protein functions as a modulator for metabolic reprogramming of colorectal cancer cells by inducing nutrient stress-mediated autophagy.
Towards a precision immune checkpoint blockade immunotherapy in patients with colorectal cancer: Strategies and perspectives.
TIMP1 Promotes Cell Proliferation and Invasion Capability of Right-sided Colon Cancers via the FAK/Akt Signaling Pathway.
The role of heat shock proteins in metastatic colorectal cancer: A review.
Novel antigen presenting cell imparts Treg-dependent tolerance to gut microbiota.
Myeloid-derived suppressor cells in colorectal cancer: prognostic biomarkers and therapeutic targets.
The cAMP/PKA/CREB and TGF-β/SMAD4 pathways regulate stemness and metastatic potential in colorectal cancer cells.
Disulfiram increases the efficacy of 5-fluorouracil in organotypic cultures of colorectal carcinoma.
Comprehensive analysis of the prognostic and immunotherapeutic implications of STAT family members in human colorectal cancer.
Insights into Early Onset Colorectal Cancer through Analysis of Normal Colon Organoids of Familial Adenomatous Polyposis Patients.
Checkpoint molecules on infiltrating immune cells in colorectal tumor microenvironment.
Traject3d allows label-free identification of distinct co-occurring phenotypes within 3D culture by live imaging.
Regorafenib in combination with immune checkpoint inhibitors for mismatch repair proficient (pMMR)/microsatellite stable (MSS) colorectal cancer.
Acquired Genomic Alterations on First-Line Chemotherapy With Cetuximab in Advanced Colorectal Cancer: Circulating Tumor DNA Analysis of the CALGB/SWOG-80405 Trial (Alliance).

Curr Opin Genet Dev. 2022 Sep 01. pii: S0959-437X(22)00086-7. [Epub ahead of print]76 101977

Intestinal epithelial organoids: regeneration and maintenance of the intestinal epithelium.

Mayumi Oda, Yoshiko Hatano, Toshiro Sato.

Vital functions of the intestines: digestion, absorption, and surface barrier are performed by the intestinal epithelium, which consists of various differentiated cells and intestinal stem cells. Recent technological advances in sequencing technology, including single-cell transcriptomics and epigenetic analysis, have facilitated the genetic characterization of diverse intestinal epithelial cell types and surrounding mesenchymal niche environments. Organoids have allowed biological analysis of the human intestinal epithelium in coordination with genome engineering, genetic lineage tracing, and transplantation into orthotopic tissue. Together, these technologies have prompted the development of organoid-based regenerative therapies for intestinal diseases, including short-bowel syndrome. This article provides an overview of the current understanding of intestinal epithelial self-renewal during homeostasis and regeneration and provides a perspective for future organoid medicine.

DOI: https://doi.org/10.1016/j.gde.2022.101977
Cancers (Basel). 2022 Aug 31. pii: 4260. [Epub ahead of print]14(17):

Modeling Colorectal Cancer Progression Reveals Niche-Dependent Clonal Selection.

Nuria Vaquero-Siguero, Nikolai Schleussner, Julia Volk, Manuel Mastel, Jasmin Meier, Rene Jackstadt.

  Colorectal cancer (CRC) is among the deadliest cancers worldwide, with metastasis being the main cause of patient mortality. During CRC progression the complex tumor ecosystem changes in its composition at virtually every stage. However, clonal dynamics and associated niche-dependencies at these stages are unknown. Hence, it is of importance to utilize models that faithfully recapitulate human CRC to define its clonal dynamics. We used an optical barcoding approach in mouse-derived organoids (MDOs) that revealed niche-dependent clonal selection. Our findings highlight that clonal selection is controlled by a site-specific niche, which critically contributes to cancer heterogeneity and has implications for therapeutic intervention.

Keywords:  clonal selection; colorectal cancer; metastasis; mouse models; mouse-derived organoids; niche; orthotopic transplantation; tumor heterogeneity

DOI:  https://doi.org/10.3390/cancers14174260
Adv Sci (Weinh). 2022 Sep 04. e2204097

Patient-Derived Organoids from Colorectal Cancer with Paired Liver Metastasis Reveal Tumor Heterogeneity and Predict Response to Chemotherapy.

Shaobo Mo, Peiyuan Tang, Wenqin Luo, Long Zhang, Yaqi Li, Xiang Hu, Xiaoji Ma, Yikuan Chen, Yichao Bao, Xingfeng He, Guoxiang Fu, Xiaoya Xu, Xinxin Rao, Xiaomeng Li, Ruoyu Guan, Shengzhi Chen, Yun Deng, Tao Lv, Peiyuan Mu, Qiang Zheng, Simin Wang, Fangqi Liu, Yiwei Li, Weiqi Sheng, Dan Huang, Chen Hu, Jianjun Gao, Zhen Zhang, Sanjun Cai, Hans Clevers, Junjie Peng, Guoqiang Hua.

  There is no effective method to predict chemotherapy response and postoperative prognosis of colorectal cancer liver metastasis (CRLM) patients. Patient-derived organoid (PDO) has become an important preclinical model. Herein, a living biobank with 50 CRLM organoids derived from primary tumors and paired liver metastatic lesions is successfully constructed. CRLM PDOs from the multiomics levels (histopathology, genome, transcriptome and single-cell sequencing) are comprehensively analyzed and confirmed that this organoid platform for CRLM could capture intra- and interpatient heterogeneity. The chemosensitivity data in vitro reveal the potential value of clinical application for PDOs to predict chemotherapy response (FOLFOX or FOLFIRI) and clinical prognosis of CRLM patients. Taken together, CRLM PDOs can be utilized to deliver a potential application for personalized medicine.

Keywords:  chemotherapy response; colorectal cancer liver metastasis; patient-derived organoid; prognosis prediction; tumor heterogeneity

DOI:  https://doi.org/10.1002/advs.202204097
Eur J Pharmacol. 2022 Sep 03. pii: S0014-2999(22)00514-3. [Epub ahead of print] 175253

Long-term resistance to 5-fluorouracil promotes epithelial-mesenchymal transition, apoptosis evasion, autophagy, and reduced proliferation rate in colon cancer cells.

Annie Cristhine Moraes Sousa-Squiavinato, Diego Alfonso Arregui Ramos, Mônica Wanger Silveira, Josiane Weber Tessmann, Julio Cesar Madureira de-Freitas-Junior, Jose Andrés Morgado-Díaz.

  The drug, 5-fluorouracil (5FU) is a standard first-line treatment for colorectal cancer (CRC) patients. However, drug resistance acquisition remains an important challenge for effective clinical outcomes. Here, we established a long-term drug-resistant CRC model and explored the cellular events underlying 5FU resistance. We showed that 5FU-treated cells (HCT-116 5FUR) using a prolonged treatment protocol were significantly more resistant than parental cells. Likewise, cell viability and IC50 values were also observed to increase in HCT-116 5FUR cells when treated with increasing doses of oxaliplatin, indicating a cross-resistance mechanism to other cytotoxic agents. Moreover, HCT-116 5FUR cells exhibited metabolic and molecular changes, as evidenced by increased thymidylate synthase levels and upregulated mRNA levels of ABCB1. HCT-116 5FUR cells were able to overcome S phase arrest and evade apoptosis, as well as activate autophagy, as indicated by increased LC3B levels. Cells treated with low and high doses displayed epithelial-mesenchymal transition (EMT) features, as observed by decreased E-cadherin and claudin-3 levels, increased vimentin protein levels, and increased SLUG, ZEB2 and TWIST1 mRNA levels. Furthermore, HCT-116 5FUR cells displayed enhanced migration and invasion capabilities. Interestingly, we found that the 5FU drug-resistance gene signature is positively associated with the mesenchymal signature in CRC samples, and that ABCB1 and ZEB2 co-expressed at high levels could predict poor outcomes in CRC patients. Overall, the 5FU long-term drug-resistance model established here induced various cellular events, and highlighted the importance of further efforts to identify promising targets involved in more than one cellular event to successfully overcome drug-resistance.

Keywords:  5-Fluorouracil; Apoptosis; Autophagy; Colorectal cancer; Drug-resistance; Epithelial–mesenchymal transition

DOI:  https://doi.org/10.1016/j.ejphar.2022.175253
Nat Rev Gastroenterol Hepatol. 2022 Sep 06.

From stem to stern: LGR5+ as a marker for intestinal crypts.

Ghada Mohamed.

DOI: https://doi.org/10.1038/s41575-022-00683-x
Nat Commun. 2022 Sep 09. 13(1): 5312

In vivo tumor immune microenvironment phenotypes correlate with inflammation and vasculature to predict immunotherapy response.

Aditi Sahu, Kivanc Kose, Lukas Kraehenbuehl, Candice Byers, Aliya Holland, Teguru Tembo, Anthony Santella, Anabel Alfonso, Madison Li, Miguel Cordova, Melissa Gill, Christi Fox, Salvador Gonzalez, Piyush Kumar, Amber Weiching Wang, Nicholas Kurtansky, Pratik Chandrani, Shen Yin, Paras Mehta, Cristian Navarrete-Dechent, Gary Peterson, Kimeil King, Stephen Dusza, Ning Yang, Shuaitong Liu, William Phillips, Pascale Guitera, Anthony Rossi, Allan Halpern, Liang Deng, Melissa Pulitzer, Ashfaq Marghoob, Chih-Shan Jason Chen, Taha Merghoub, Milind Rajadhyaksha.

Response to immunotherapies can be variable and unpredictable. Pathology-based phenotyping of tumors into 'hot' and 'cold' is static, relying solely on T-cell infiltration in single-time single-site biopsies, resulting in suboptimal treatment response prediction. Dynamic vascular events (tumor angiogenesis, leukocyte trafficking) within tumor immune microenvironment (TiME) also influence anti-tumor immunity and treatment response. Here, we report dynamic cellular-level TiME phenotyping in vivo that combines inflammation profiles with vascular features through non-invasive reflectance confocal microscopic imaging. In skin cancer patients, we demonstrate three main TiME phenotypes that correlate with gene and protein expression, and response to toll-like receptor agonist immune-therapy. Notably, phenotypes with high inflammation associate with immunostimulatory signatures and those with high vasculature with angiogenic and endothelial anergy signatures. Moreover, phenotypes with high inflammation and low vasculature demonstrate the best treatment response. This non-invasive in vivo phenotyping approach integrating dynamic vasculature with inflammation serves as a reliable predictor of response to topical immune-therapy in patients.

DOI: https://doi.org/10.1038/s41467-022-32738-7
Adv Healthc Mater. 2022 Sep 07. e2201172

Modelling Biochemical Gradients in Vitro to Control Cell Compartmentalization in a Microengineered 3D Model of the Intestinal Epithelium.

Gizem Altay, Aina Abad-Lázaro, Emilio J Gualda, Jordi Folch, Claudia Insa, Sébastien Tosi, Xavier Hernando-Momblona, Eduard Batlle, Pablo Loza-Álvarez, Vanesa Fernández-Majada, Elena Martinez.

  Gradients of signaling pathways within the intestinal stem cell (ISC) niche are instrumental for cellular compartmentalization and tissue function, yet how are they sensed by the epithelium is still not fully understood. Here we present a new in vitro model of the small intestine based on primary epithelial cells (i), apically accessible (ii), with native tissue mechanical properties and controlled mesh size (iii), 3D villus-like architecture (iv), and precisely controlled biomolecular gradients of the ISC niche (v). Biochemical gradients are formed through hydrogel-based scaffolds by free diffusion from a source to a sink chamber. To confirm the establishment of spatiotemporally controlled gradients, we employ light-sheet fluorescence microscopy and in-silico modelling. The ISC niche biochemical gradients coming from the stroma and applied along the villus axis lead to the in vivo-like compartmentalization of the proliferative and differentiated cells, while changing the composition and concentration of the biochemical factors affects the cellular organization along the villus axis. This novel 3D in vitro intestinal model derived from organoids recapitulates both the villus-like architecture and the gradients of ISC biochemical factors, thus opening the possibility to study in vitro the nature of such gradients and the resulting cellular response. This article is protected by copyright. All rights reserved.

Keywords:  3D architecture; biomolecular gradients; engineering organoids; in-silico modelling; intestinal stem cell niche; light sheet fluorescence microscopy; photolithography

DOI:  https://doi.org/10.1002/adhm.202201172
Cell Mol Life Sci. 2022 Sep 04. 79(9): 505

Small leucine zipper protein functions as a modulator for metabolic reprogramming of colorectal cancer cells by inducing nutrient stress-mediated autophagy.

Suhyun Kim, Minseok Oh, Minsoo Kang, Jesang Ko.

  In multiple cancers, autophagy promotes tumor development by recycling intracellular components into metabolic pathways. Autophagy-induced metabolic reprogramming and plasticity lead to cancer cell survival and resistance to anticancer therapy. We investigated the role of small leucine zipper protein (sLZIP) in autophagy and cell survival under nutrient-deficient conditions in colorectal cancer (CRC). sLZIP was induced by nutrient stress and increased the transcription of microtubule-associated protein 1A/1B-light chain 3 (LC3), by directly binding to its promoter. Under nutrient stress conditions, sLZIP activated autophagy and promoted the survival of CRC cells. sLZIP induced metabolic reprogramming of CRC cells, to activate glutaminolysis and the tricarboxylic acid cycle. sLZIP also enhanced the autophagic degradation of Keap1 and the nuclear accumulation of Nrf2, leading to NQO1 expression, for maintenance of redox homeostasis. sLZIP-knockout CRC cells exhibited impaired autophagy induction in the glycolytic inhibition state. Xenograft mice lacking sLZIP showed decreased tumor growth, by rendering CRC cells sensitive to glycolysis inhibition. The expression of sLZIP and LC3B was highly elevated in tumors of CRC patients compared to that in normal tissues, and correlated with the progression of CRC. These findings suggest that sLZIP drives autophagy and metabolic reprogramming to promote colorectal tumorigenesis.

Keywords:  Autophagy; Colorectal cancer; Metabolic reprogramming; Transcriptional regulation

DOI:  https://doi.org/10.1007/s00018-022-04535-4
Biomed Pharmacother. 2022 May;pii: S0753-3322(22)00312-2. [Epub ahead of print]149 112923

Towards a precision immune checkpoint blockade immunotherapy in patients with colorectal cancer: Strategies and perspectives.

Guanglin Cui.

  To date, immune checkpoint blockade (ICB) immunotherapy has become one of promise strategies in the management of patients with unresectable or metastatic colorectal cancer (CRC). However, clinical observations showed that not all the patients responded equally to ICBs, certain group of CRC patients with microsatellite-instability-low (MSI-L) phenotype was not sensitive to ICB immunotherapy. In addition, some primary responders might lose their sensitivity and become resistant to ICBs overtime. To obtain a better response rate and therapeutic efficacy, considerable attempts have been made toward to a precision medicine algorithm. Studies showed that multiple strategies based on the patient's individual condition might improve the response and therapeutic efficacy to ICBs. Therefore, we focused on and discussed precision strategies and perspectives e.g., how to early define candidates who will benefit from ICB immunotherapy prior treatment, overcome the primary and acquired resistance and improve the therapeutic response to ICBs in CRC patients with different microsatellite-instability statuses within the context of precision medicine algorithm in this review.

Keywords:  Blockade; Immune checkpoint; Precise medicine, colorectal cancer

DOI:  https://doi.org/10.1016/j.biopha.2022.112923
Cancer Sci. 2022 Sep 08.

TIMP1 Promotes Cell Proliferation and Invasion Capability of Right-sided Colon Cancers via the FAK/Akt Signaling Pathway.

Beibei Ma, Hiroyuki Ueda, Koichi Okamoto, Masahiro Bando, Shota Fujimoto, Yasuyuki Okada, Tomoyuki Kawaguchi, Hironori Wada, Hiroshi Miyamoto, Mitsuo Shimada, Yasushi Sato, Tetsuji Takayama.

  Although right-sided colorectal cancer (CRC) shows a worse prognosis than left-sided CRC, underlying mechanism remains unclear. We established patient-derived organoids (PDOs) from both-sided CRCs and directly compared cell proliferation and invasion capability between them. We then analyzed the expression of numerous genes in signal transduction pathways to clarify the mechanism of the differential prognosis. Cell proliferation activity and invasion capability in right-sided cancer PDOs were significantly higher than in left-sided cancer PDOs and normal PDOs, as revealed by CellTiter-Glo and transwell assays, respectively. We then used quantitative RT-PCR to compare 184 genes in 30 pathways among right-sided and left-sided cancer and normal PDOs, and found that the TIMP1 mRNA level was highest in right-sided PDOs. TIMP1 protein levels were upregulated in right-sided PDOs compared with normal PDOs but was downregulated in left-sided PDOs. TIMP1 knockdown with shRNA significantly decreased cell proliferation activity and invasion capability in right-sided PDOs but not in left-sided PDOs. Moreover, TIMP1 knockdown significantly decreased pFAK and pAkt expression levels in right-sided PDOs but not in left-sided PDOs. A TCGA database analysis revealed that TIMP1 expression in right-sided CRCs was significantly higher than in left-sided CRCs. Kaplan-Meier survival analysis showed significantly shorter overall survival in high-TIMP1 patients versus low-TIMP1 patients with right-sided CRCs but not left-sided CRCs. Our data suggest that TIMP1 is overexpressed in right-sided CRCs and promotes cell proliferation and invasion capability through the TIMP1/FAK/Akt pathway, leading to a poor prognosis. The TIMP1/FAK/Akt pathway can be a target for therapeutic agents in right-sided CRCs.

Keywords:  Colorectal cancer; Organoids; Prognosis; TIMP1; Tumor location

DOI:  https://doi.org/10.1111/cas.15567
J Cell Biochem. 2022 Sep 05.

The role of heat shock proteins in metastatic colorectal cancer: A review.

Hossein Javid, Pedram Hashemian, Shaghayegh Yazdani, Alireza Sharbaf Mashhad, Mehdi Karimi-Shahri.

  Heat shock proteins (HSPs) are a large molecular chaperone family classified by their molecular weights, including HSP27, HSP40, HSP60, HSP70, HSP90, and HSP110. HSPs are likely to have antiapoptotic properties and participate actively in various processes such as tumor cell proliferation, invasion, metastases, and death. In this review, we discuss comprehensively the functions of HSPs associated with the progression of colorectal cancer (CRC) and metastasis and resistance to cancer therapy. Taken together, HSPs have numerous clinical applications as biomarkers for cancer diagnosis and prognosis and potential therapeutic targets for CRC and its related metastases.

Keywords:  colorectal cancer (CRC); heat shock proteins (HSPs); metastatic colorectal cancer

DOI:  https://doi.org/10.1002/jcb.30326
Nature. 2022 Sep 07.

Novel antigen presenting cell imparts Treg-dependent tolerance to gut microbiota.

Blossom Akagbosu, Zakieh Tayyebi, Gayathri Shibu, Yoselin A Paucar Iza, Deeksha Deep, Yollanda Franco Parisotto, Logan Fisher, H Amalia Pasolli, Valentin Thevin, Rasa Elmentaite, Maximilian Knott, Saskia Hemmers, Lorenz Jahn, Christin Friedrich, Jacob Verter, Zhong-Min Wang, Marcel van den Brink, Georg Gasteiger, Thomas G P Grünewald, Julien C Marie, Christina Leslie, Alexander Y Rudensky, Chrysothemis C Brown.

Establishing and maintaining tolerance to self- or innocuous foreign antigens is vital for preservation of organismal health. Within the thymus, medullary thymic epithelial cells (mTECs) expressing AutoImmune Regulator, Aire, play a critical role in self-tolerance through deletion of autoreactive T cells and promotion of thymic regulatory T (Treg) cell development1-4. Within weeks of birth, a separate wave of Treg cell differentiation occurs in the periphery, upon exposure to dietary and commensal microbiota derived antigens5-8, yet the cell types responsible for the generation of peripheral Treg (pTreg) cells are not known. Here we identified a new class of RORγt+ antigen-presenting cells (APC), dubbed Thetis cells (TCs), with transcriptional features of both mTECs and dendritic cells (DCs), comprising 4 major sub-groups (TC I-IV). We uncovered a developmental wave of TCs within intestinal lymph nodes during a critical early life window, coincident with the wave of pTreg cell differentiation. While TC I and III expressed the signature mTEC nuclear factor Aire, TC IV lacked Aire expression and were enriched for molecules required for pTreg generation, including the TGF-β activating integrin αvβ8. Loss of either MHCII or Itgb8 expression by TCs led to a profound impairment in intestinal pTreg differentiation, with ensuing colitis. In contrast, MHCII expression by RORγt+ group 3 innate lymphoid cells (ILC3) and classical DCs was neither sufficient nor required for pTreg generation, further implicating TC IV as the tolerogenic RORγt+ APC with an essential early life function. Our studies reveal parallel pathways for establishment of tolerance to self and foreign antigen within the thymus and periphery, respectively, marked by involvement of shared cellular and transcriptional programs.

DOI: https://doi.org/10.1038/s41586-022-05309-5
Explor Target Antitumor Ther. 2022 ;3(4): 497-510

Myeloid-derived suppressor cells in colorectal cancer: prognostic biomarkers and therapeutic targets.

Mohammad A Al-Mterin, Eyad Elkord.

  Myeloid-derived suppressor cells (MDSCs) are a group of immature myeloid cells, which are expanded in most cancer patients. MDSCs suppress host immune responses, leading to cancer growth and progression. Several studies demonstrated that there was a relationship between levels of MDSCs and tumorigenesis in colorectal cancer (CRC) patients. MDSCs are now being investigated for their role as possible therapeutic targets in cancer treatment. This review summarizes available studies that investigated MDSC expansion in CRC patients, as well as their role in CRC tumorigenesis, prognosis, and targeting. Based on the available studies, there is a possible relationship between high levels of MDSCs and CRC progression. Additionally, targeting MDSCs in CRC patients selectively represents a significant challenge for the development of targeted treatments. Targeting of MDSCs could be exploited in different ways including MDSC depletion, inhibition of MDSC function and recruitment, and enhancing MDSC differentiation. Overall, MDSCs could be exploited as prognostic biomarkers and potential therapeutic targets in CRC.

Keywords:  Myeloid-derived suppressor cells; colorectal cancer; prognostic biomarkers; targeting

DOI:  https://doi.org/10.37349/etat.2022.00097
Cancer Res. 2022 Sep 06. pii: CAN-22-1369. [Epub ahead of print]

The cAMP/PKA/CREB and TGF-β/SMAD4 pathways regulate stemness and metastatic potential in colorectal cancer cells.

Teruaki Fujishita, Yasushi Kojima, Rie Kajino-Sakamoto, Emi Mishiro-Sato, Yasuhiro Shimizu, Waki Hosoda, Rui Yamaguchi, Makoto Mark Taketo, Masahiro Aoki.

Metastasis is responsible for the majority of deaths of cancer patients. However, mechanisms governing metastasis in colorectal cancer (CRC) remain largely unknown. Here we investigated how CRC cells acquire metastatic potential using a novel mouse model of CRC that spontaneously develops liver metastasis, generated by introducing sporadic mutations of Ctnnb1, Kras, Trp53, and Smad4 (CKPS) genes. Proteomic analyses revealed elevated expression of CRC stem cell markers ALCAM (CD166) and PROM1 (CD133) in CRC cells from the metastatic model compared with those from a non-metastatic model. Spleen-to-liver metastasis assays using CRC cells derived from the CKPS model (CKPS cells) demonstrated the functional importance of ALCAM and PROM1 in initiating metastasis. Genetic and pharmacological analyses using CKPS cells in 2D and spheroid culture revealed that expression of ALCAM and PROM1 is regulated positively and negatively by the cAMP/PKA/CREB and TGF-β/SMAD4 pathways, respectively. Consistently, phospho-CREB was expressed in both primary and metastatic lesions of CKPS mice and CRC patients, and knockout of CREB in CKPS cells reduced their spheroid-forming and metastasis-initiating abilities. Treatment with a CREB inhibitor potentiated the effect of irinotecan in suppressing liver metastasis by CKPS cells. These results reveal the essential roles of ALCAM and PROM1, as well as their upstream regulators, the cAMP/PKA/CREB and TGF-β/SMAD4 pathways, in maintaining the stemness and metastatic potential of CRC cells and indicate that CREB inhibition may be a potential therapeutic strategy against metastatic CRC.

DOI: https://doi.org/10.1158/0008-5472.CAN-22-1369
Biomed Pharmacother. 2022 Sep;pii: S0753-3322(22)00854-X. [Epub ahead of print]153 113465

Disulfiram increases the efficacy of 5-fluorouracil in organotypic cultures of colorectal carcinoma.

Michal Hendrych, Kamila Říhová, Barbora Adamová, Vojtěch Hradil, Marek Stiborek, Petr Vlček, Markéta Hermanová, Jana Vašíčková, Petr Beneš, Jan Šmarda, Viktor Kanický, Jan Preisler, Jarmila Navrátilová.

  Drug efficacy determined in preclinical research is difficult to transfer to clinical practice. This is mainly due to the use of oversimplified models omitting the effect of the tumor microenvironment and the presence of various cell types participating in the formation of tumors in vivo. In this study, we used robust three-dimensional models including spheroids grown from colon cancer cell lines and organotypic cultures prepared from the colorectal carcinoma tissue to test novel therapeutic strategies. We developed a multi-modal approach combining brightfield and fluorescence microscopy for evaluating drug effects on organotypic cultures. Combined treatment with 5-fluorouracil and disulfiram/copper efficiently eliminated cancer cells in these 3D models. Moreover, disulfiram/copper down-regulated the expression of markers associated with 5-fluorouracil resistance, such as thymidylate synthase and CD133/CD44. Thus, we propose combined therapy of 5-fluorouracil and disulfiram/copper for further testing as a treatment for colorectal carcinoma. In addition, we show that organotypic cultures are suitable models for anti-cancer drug testing.

Keywords:  5-fluorouracil; Colorectal carcinoma; Disulfiram/copper; Organotypic culture; Spheroid; Synergism

DOI:  https://doi.org/10.1016/j.biopha.2022.113465
Front Genet. 2022 ;13 951252

Comprehensive analysis of the prognostic and immunotherapeutic implications of STAT family members in human colorectal cancer.

Dingchang Li, Yanan Jiao, Wenxing Gao, Shidong Hu, Dingling Li, Wen Zhao, Peng Chen, Lujia Jin, Yingjie Zhao, Zhaofu Ma, Xiansheng Wu, Yang Yan, Wen Sun, Xiaohui Du, Guanglong Dong.

  Background: Colorectal cancer (CRC) is the third most prevalent cancer worldwide and the second leading cause of cancer mortality. Signal transducer and activator of transcription (STAT) proteins are a group of transcription factors implicated in cell signal transduction and gene transcription in several cancer types. However, the level of expression, genetic alterations, and biological function of different STATs, as well as their prognostic and immunotherapeutic value in CRC remain unclear. Methods: The mRNA and protein expression levels, genetic alterations, prognostic value, gene-gene and protein-protein interaction networks, and biological function of STATs in CRC were studied using the GEPIA, HPA, cBioPortal, PrognoScan, Kaplan-Meier plotter, GeneMANIA, STRING, and Metascape databases. The expression of STATs in CRC was confirmed using immunohistochemistry (IHC). Finally, the relationship between STAT expression and immune infiltration as well as immunotherapy-associated indicators was also investigated. Results: The expression levels of STAT2/5A/5B are downregulated in CRC, and the STAT1/3/4/5B expressions were significantly associated with the tumor stage of patients with CRC. The abnormal expression of STAT2/4/5B in patients with CRC is related to the prognosis of patients with CRC. The STATs and their neighboring proteins are primarily associated with lymphocyte activation, cytokine-mediated signaling pathways, positive regulation of immune response, regulation of cytokine production, and growth hormone receptor signaling pathways in cancer. The expression of STATs was significantly associated with immune infiltration and immunotherapy response-associated indicators. Conclusion: This study may help further understand the molecular mechanism of CRC and provide new prognostic biomarkers and immunotherapy targets in patients with CRC.

Keywords:  STAT transcription factors; bioinformatics analysis; colorectal cancer; immune infiltration; prognostic value; tumor immunotherapy

DOI:  https://doi.org/10.3389/fgene.2022.951252
Cancers (Basel). 2022 Aug 26. pii: 4138. [Epub ahead of print]14(17):

Insights into Early Onset Colorectal Cancer through Analysis of Normal Colon Organoids of Familial Adenomatous Polyposis Patients.

Matthew A Devall, Stephen Eaton, Mourad W Ali, Steven M Powell, Li Li, Graham Casey.

  Early onset colorectal cancer (EOCRC) rates have increased in recent decades. While lowering the recommended age for routine colonoscopies to 45 may reduce this burden, such measures do not address those who develop CRC before that age. Additional measures are needed to identify individuals at-risk for CRC. To better define transcriptomic events that precede the development of CRC, we performed RNA-sequencing analysis in colon organoids derived from seven healthy and six familial adenomatous polyposis (FAP) patients. This led to the identification of 2635 significant differentially expressed genes (FDR < 0.05). Through secondary analysis of publicly available datasets, we found that these genes were enriched for significant genes also present in FAP CRC and non-hereditary CRC datasets, including a subset that were unique to EOCRC. By exposing FAP colon organoids to a three-day ethanol treatment, we found that two EOCRC-relevant genes were also targets of CRC related lifestyle factors. Our data provides unique insight into the potential, early mechanisms of CRC development in colon epithelial cells, which may provide biomarkers for patient monitoring. We also show how modifiable lifestyle factors may further alter genes relevant to EOCRC, adding weight to the hypothesis that such factors represent an important contributor to increased EOCRC incidence.

Keywords:  RNA-seq; colon organoid; early onset colorectal cancer; familial adenomatous polyposis

DOI:  https://doi.org/10.3390/cancers14174138
Front Med (Lausanne). 2022 ;9 955599

Checkpoint molecules on infiltrating immune cells in colorectal tumor microenvironment.

Iman M Talaat, Noha M Elemam, Shroque Zaher, Maha Saber-Ayad.

  Colorectal cancer (CRC) is one of the most prevalent cancer types worldwide, with a high mortality rate due to metastasis. The tumor microenvironment (TME) contains multiple interactions between the tumor and the host, thus determining CRC initiation and progression. Various immune cells exist within the TME, such as tumor-infiltrating lymphocytes (TILs), tumor-associated macrophages (TAMs), and tumor-associated neutrophils (TANs). The immunotherapy approach provides novel opportunities to treat solid tumors, especially toward immune checkpoints. Despite the advances in the immunotherapy of CRC, there are still obstacles to successful treatment. In this review, we highlighted the role of these immune cells in CRC, with a particular emphasis on immune checkpoint molecules involved in CRC pathogenesis.

Keywords:  colorectal cancer; immune checkpoint; lymphocytes; macrophages; neutrophils; tumor microenvironment

DOI:  https://doi.org/10.3389/fmed.2022.955599
Nat Commun. 2022 Sep 09. 13(1): 5317

Traject3d allows label-free identification of distinct co-occurring phenotypes within 3D culture by live imaging.

Eva C Freckmann, Emma Sandilands, Erin Cumming, Matthew Neilson, Alvaro Román-Fernández, Konstantina Nikolatou, Marisa Nacke, Tamsin R M Lannagan, Ann Hedley, David Strachan, Mark Salji, Jennifer P Morton, Lynn McGarry, Hing Y Leung, Owen J Sansom, Crispin J Miller, David M Bryant.

Single cell profiling by genetic, proteomic and imaging methods has expanded the ability to identify programmes regulating distinct cell states. The 3-dimensional (3D) culture of cells or tissue fragments provides a system to study how such states contribute to multicellular morphogenesis. Whether cells plated into 3D cultures give rise to a singular phenotype or whether multiple biologically distinct phenotypes arise in parallel is largely unknown due to a lack of tools to detect such heterogeneity. Here we develop Traject3d (Trajectory identification in 3D), a method for identifying heterogeneous states in 3D culture and how these give rise to distinct phenotypes over time, from label-free multi-day time-lapse imaging. We use this to characterise the temporal landscape of morphological states of cancer cell lines, varying in metastatic potential and drug resistance, and use this information to identify drug combinations that inhibit such heterogeneity. Traject3d is therefore an important companion to other single-cell technologies by facilitating real-time identification via live imaging of how distinct states can lead to alternate phenotypes that occur in parallel in 3D culture.

DOI: https://doi.org/10.1038/s41467-022-32958-x
Cancer Treat Rev. 2022 Aug 27. pii: S0305-7372(22)00129-3. [Epub ahead of print]110 102460

Regorafenib in combination with immune checkpoint inhibitors for mismatch repair proficient (pMMR)/microsatellite stable (MSS) colorectal cancer.

Tugba Akin Telli, Giacomo Bregni, Michele Vanhooren, Rita Saude Conde, Alain Hendlisz, Francesco Sclafani.

  Immune checkpoint inhibitors (ICIs) have marked a new era of cancer treatment, showing remarkable efficacy in a wide range of solid malignancies. In colorectal cancer (CRC), however, the therapeutic potential of ICIs is limited to the small group (≈5%) of patients with mismatch repair deficient (dMMR)/high microsatellite instable (MSI-H) tumours, which are characterised by high mutational/neo-antigen burden, and an inflammatory tumour microenvironment with abundant tumour-infiltrating lymphocytes. Over the last few years, research has focused on immuno-modulatory strategies that could overcome the inherent resistance to ICIs that is observed in the vast group (≈95%) of patients with mismatch repair proficient (pMMR)/microsatellite stable (MSS) tumours. Among these, the combination of ICIs with multi-kinase inhibitors has gained traction in preclinical studies and clinical trials. Thanks to their multiple targets and mechanisms of action, generally involving key cancer pathways such as oncogenesis, angiogenesis, metastasis, and tumour immunity, these agents can exert synergistic effects with ICIs, eventually turning inherently cold cancers into hot tumours, that can be efficiently recognised and targeted by an activated immune system. Regorafenib is routinely used for chemorefractory CRC with limited efficacy. Preliminary evidence, however, suggests that this multi-kinase inhibitor could be an optimal combination partner for ICIs. In this review article, we explain the biological rationale underlying the synergism between regorafenib and ICIs, discuss the available clinical data in CRC, and take a glance into future perspectives by presenting ongoing trials and possible research developments in this setting.

Keywords:  Colorectal cancer; Immune checkpoint inhibitors; Microsatellite stable; Mismatch repair proficient; Regorafenib

DOI:  https://doi.org/10.1016/j.ctrv.2022.102460
J Clin Oncol. 2022 Sep 06. JCO2200365

Acquired Genomic Alterations on First-Line Chemotherapy With Cetuximab in Advanced Colorectal Cancer: Circulating Tumor DNA Analysis of the CALGB/SWOG-80405 Trial (Alliance).

Kanwal Raghav, Fang-Shu Ou, Alan P Venook, Federico Innocenti, Ryan Sun, Heinz-Josef Lenz, Scott Kopetz.

Clinical trials frequently include multiple end points that mature at different times. The initial report, typically based on the primary end point, may be published when key planned co-primary or secondary analyses are not yet available. Clinical Trial Updates provide an opportunity to disseminate additional results from studies, published in JCO or elsewhere, for which the primary end point has already been reported.Acquired genomic alterations (Acq-GAs), specifically RAS, BRAF, and EGFR-ectodomain mutations and ERBB2 and MET amplifications, are recognized as major mechanisms of resistance to later-line anti-EGFR-antibody therapy in metastatic colorectal cancer (mCRC). However, data regarding emergence of these Acq-GAs under the selective pressure of first-line anti-EGFR-chemotherapy are lacking. We performed next-generation sequencing (Guardant360) on circulating tumor DNA obtained from paired plasma samples (pretreatment and postprogression) from the CALGB/SWOG-80405 trial, which randomly assigned patients with mCRC between first-line chemotherapy with cetuximab (anti-EGFR-chemotherapy) or bevacizumab (anti-VEGF-chemotherapy). The primary objective was to determine the prevalence of Acq-GAs on anti-EGFR-chemotherapy and compare this to the prevalence with anti-VEGF-chemotherapy on trial and pooled estimates (N = 292) seen with later-line anti-EGFR-antibody therapy as reported in the literature. Among the 61 patients on anti-EGFR-chemotherapy, only four (6.6%) developed ≥ 1 Acq-GAs of interest compared with 10.1% (7) on anti-VEGF-chemotherapy (odds ratio, 0.62; 95% CI, 0.20 to 2.11) and 62.0% on anti-EGFR-antibody therapy in later lines (odds ratio, 0.09; 95% CI, 0.03 to 0.23). Acq-GAs, classically associated with anti-EGFR-antibody resistance in later lines (RAS, BRAF, and EGFR-ectodomain mutations; ERBB2 and MET amplifications), were rare with up-front use of anti-EGFR-chemotherapy indicating divergent resistance mechanisms. These findings have critical translational relevance to timing and value of circulating tumor DNA-guided anti-EGFR rechallenge in patients with mCRC, especially those treated with anti-EGFR therapy upfront.

DOI: https://doi.org/10.1200/JCO.22.00365