J Cancer Prev 2022; 27(1): 1-6
Published online March 30, 2022
© Korean Society of Cancer Prevention
Mizuho Nakayama1,2 , Dong Wang1,2 , Sau Yee Kok1,3 , Hiroko Oshima1,2 , Masanobu Oshima1,2
1Division of Genetics, Cancer Research Institute, Kanazawa University, 2WPI Nano-Life Science Institute (Nano-LSI), Kanazawa University, Kanazawa, Japan, 3Cancer Immunology and Immunotherapy Unit, Cancer Research Malaysia, Selangor, Malaysia
Correspondence to :
Masanobu Oshima, E-mail: email@example.com, https://orcid.org/0000-0002-3304-0004
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License, which permits unrestricted noncommercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
Comprehensive genome analyses have identified frequently mutated genes in human colorectal cancers (CRC). These include APC, KRAS, SMAD4, TP53, and FBXW7. The biological functions of the respective gene products in cell proliferation and homeostasis have been intensively examined by in vitro experiments. However, how each gene mutation or combinations of specific mutations drive malignant progression of CRC in vivo has not been fully understood. Based on the genomic information, we generated mouse models that carry multiple mutations of CRC driver genes in various combinations, and we performed comprehensive histological analyses to link genetic alteration(s) and tumor phenotypes, including liver metastasis. In this review article, we summarize the phenotypes of the respective genetic models carrying major driver mutations and discuss a possible mechanism of mutations underlying malignant progression.
Keywords: Transgenic mice, Organoid, Colorectal cancer, Mutations, Elastic modulus
It has been proposed that the accumulation of genetic alterations causes the development and malignant progression of colorectal cancer (CRC) as a well-known concept of multistep tumorigenesis . According to this concept,
Wnt signaling plays an important role in the regulation of stemness of tissue stem cells . Wnt signaling via Frizzled receptors induces dissociation of the destruction complex, which consists of APC, AXIN and β-catenin, resulting in stabilization and activation of β-catenin, and the induction of the expression of Wnt-target. Thus, mutations in
Oncogenic Wnt activation is also regulated at the receptor level. RNF43 is a ubiquitin ligase that targets the Frizzled receptor. Thus,
TGF-β binds the TGF-β type II receptor (TGFβRII), leading to the association and phosphorylation of TGFβRI, which further phosphorylates Smad2 or Smad3. Phosphorylated Smad2/3 forms a complex with Smad4, resulting in the expression of TGF-β target genes. TGF-β signaling induces the differentiation of intestinal epithelial cells, and thus plays a role as a tumor suppressor. When the mouse intestinal mucosa is damaged by irradiation, the remaining stem cells actively proliferate to regenerate. Notably, in
A gene expression analysis of established organoid cells indicated that the expression of mutant p53 R270H resulted in the significant upregulation of innate immunity and inflammatory pathway, including TNF, NFκB, TLR4, IRF7, and IFNG . These results suggest that GOF p53 mutation in cancer cells contributes to the generation of an inflammatory microenvironment, which may support tumor cell survival and proliferation (Fig. 1).
In the in vitro study, we found that the disruption of
We further generated mice carrying four or five driver mutations in their intestinal tumor cells, and established intestinal tumor-derived organoids from all mice . To examine the metastatic ability of the tumor cells, we transplanted established organoids to the mouse spleen and examined liver metastasis via the portal vein. Notably, tumor organoid cells that carried triple mutations,
Importantly, when AKTP cells with
Histological analyses indicate that hepatic stellate cells (HSCs) are activated and proliferate inside or outside of the liver sinusoid, where metastatic AKTP cells are arrested, which leads to the generation of a fibrotic niche that surrounds CRC cells . It has been established that TGF-β signaling is responsible for HSC activation and liver fibrosis. Notably, the disruption of host TGF-β signaling by
Genetic alterations induce distinct gene expression profiles, leading to the acquisition of malignant phenotypes, such as metastatic ability. Mouse and organoid model studies have successfully linked genotypes and phenotypes. To further understand how genetic alterations cause the acquisition of malignant phenotypes, such as increased migration and invasiveness, we performed a nano-scale topography and mechanical property analysis using scanning ion conductance microscopy (SICM). An SICM analysis can create live images that simultaneously show topography and stiffness by contact-free scanning of the cell surface with a nanopipette that measures the ion migration through the pipette tip . Notably, we observed a distinct topographic feature—a microridge structure—on the surface of AKT-common and AKF organoid cells, which actively fluctuate (Fig. 3) . Moreover, cell surface stiffness was found to be significantly low (softer) in AKT-common highly metastatic tumor organoid cells. Therefore, SICM analyses indicate that fluctuating structures and softer surface properties are characteristic to metastatic cells (Fig. 3). Such information about the physical properties of CRC cells will be helpful for understanding the relationships among genetic alteration, expression profiles, and malignant phenotypes. We also believe that SICM can be used for evaluating the metastatic ability of CRC cells.
Comprehensive genome analyses revealed frequently mutated genes in CRC as possible driver genes. Based on the genomic information, mouse genetics and organoid transplantation experiments demonstrated the role of each driver mutation and specific combinations of mutations in the promotion of malignant progression, including metastasis. Mouse and organoid model studies also showed the tumor-induced generation of microenvironment that promotes the survival and proliferation of cancer cells. These results will be helpful to understand the biological mechanism of malignant progression of CRC and the future development of clinical strategies for the treatment of malignant CRC.
We thank Manami Watanabe, Ayako Tsuda, and Yoshie Jomen for technical support in our mouse model and organoid studies.
This work was supported by Grants-in-Aid for Scientific Research (A) (18H04030) from the Ministry of Education, Cultures, Sports, Science and Technology of Japan; and AMED (21ck0106541h0002, 21gm4010012h0001) from the Japan Agency for Medical Research and Development, Japan.
No potential conflicts of interest were disclosed.
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