Drosophila models of metastasis

Epithelial–Mesenchymal Transition in Tumor Metastasis: A Method to the Madness

Cancer is one of the leading causes of death. A primary tumor forms when cells start to proliferate in an uncontrolled way and stop reacting to restraining signals. Tumors that reach a critical volume induce angiogenesis, a vascular remodeling process that provides nutrients and oxygen to the degenerated cell mass. Upon further tumor progression to a malignant cancer, cells acquire the ability to invade the surrounding tissue. In order to do so, formerly epithelial tumor cells undergo an epithelial to mesenchymal transition (EMT). This process of cell-cell detachment, breaching through the basement membrane and gaining migratory capabilities is the first step of the metastatic cascade. Metastasis is a process that allows tumor cells to leave the primary lesion and disseminate via the vascular system to secondary sites. Metastases, as the fatal feature of cancer, lead in most of the cases to patients’ death. Furthermore, metastasis, as the end stage of malignant disease, until now is incurable. This is because metastases are spread systemically throughout the body. Moreover, they even can establish from a disseminated tumor cell months after the primary tumor has already been surgically removed. Another barely controllable feature of cancer is tumor relapse. Even after a thorough surgery with the aim to completely remove the primary tumor and eventual draining lymph nodes combined with chemotherapy, patients relapse. Resistance to chemotherapy and establishment of metastases has both been accounted for to the abundance of cancer stem cells (CSC) within the tumor mass. These CSCs are endowed with the ability to evade chemotherapeutic drugs, they are mesenchymal in nature, migratory and invasive, and, most importantly, they are able to establish cancer and metastasis de novo. The work of my thesis has been dedicated to investigate the process of metastasis and the function of cancer initiation. Tumor initiation has recently been associated with EMT. To understand the functional circumstances how EMT cells gain the ability to form tumors, I used cellular murine breast cancer models. These model systems allowed me to study cells’ behavior before and after EMT in vitro and in vivo. In vitro experiments validated the observation of others that EMT cells indeed resemble cancer stem cells by being able to form hollow spheres and being susceptible to the CSC-specific drug salinomycin. In vivo studies revealed that EMT cells initiate tumors with a much earlier onset and with a higher efficiency when limited amounts of cells were injected orthotopically into mice compared to their epithelial counterpart. Moreover, EMT cell-generated cancers are highly vascularized already in the early phase of tumor establishment. Knockdown studies of the main pro-angiogenic factor VEGF-A revealed that it is required for early tumor onset. Thus, tumor angiogenesis is not only an effect of early and fast EMT-tumor progression. Further supporting this notion are limiting dilution experiments, which suggest that tumor initiation in EMT cells is a multifactorial event. This concept was validated by the observation that 10 EMT cells could initiate a tumor, whereas VEGF‑A knockdown cells could not. Hence, EMT-induced tumor initiation is achieved by the ability of promoting angiogenesis. EMT can be induced by the cytokine TGFβ. Normally, cells that experience TGFβ signaling become quiescent or die due to induction of apoptosis. Cancer cells can overcome these effects and react to TGFβ by undergoing EMT. As part of my thesis, I could show that the transcription factor Dlx2 is an important switch that allows cells to react to TGFβ by undergoing EMT without facing apoptosis induction. Dlx2 exerts its anti-apoptotic, pro-survival function by directly reducing TGFβRI expression and inducing the expression of the epidermal growth factor receptor ligand betacellulin. Another feature of EMT is the gain of cell motility. Cell migration is extremely important for cancer cells in order to leave their primary site and to disseminate. I have found ephrinB2 expression upregulated during EMT. EphrinB2 is a member of the Eph-ephrin signaling network that is known to be crucial for cell-cell communication. Thereby, cells that do not belong to the same entity repulse each other, restricting intermingling of tissue. Furthermore, ephrinB2 is required for neuronal axon guidance and angiogenesis. In my thesis, I showed that EMT cells need ephrinB2 to efficiently migrate. As an explanation for this phenotype I described that a knockdown for ephrinB2 led to an over-stabilization of focal adhesions. Cells normally use focal adhesions to hold on to an extracellular matrix (ECM) surface. Over-stabilization of focal adhesions attaches cells too firmly to the ECM and, hence, cells cannot retract their rear-end anymore which decreases cell motility. In summary, I succeeded in gathering further insights into tumor initiation, EMT, and with this, the metastatic process.

The epithelial to mesenchymal transition (EMT) is a developmental program in which epithelial cells downregulate their cell-cell junctions, acquire spindle cell morphology and exhibit cellular motility. In breast cancer, EMT facilitates invasion of surrounding tissues and correlates closely with cancer metastasis and relapse. Previous studies in our lab showed that the candidate tumor suppressor hCLCA2 is expressed in differentiated, growth-arrested mammary epithelial cells but is downregulated during tumor progression and EMT. We further showed that hCLCA2 is a p53-inducible growth inhibitor whose loss indicates an increased risk of metastasis. Subsequent studies have revealed that another member of the CLCA gene family, hCLCA4, is also expressed in mammary epithelial levels, albeit at lower levels than hCLCA2. We find that hCLCA4 is also induced by stress and is similarly downregulated in breast cancer cell lines. Like hCLCA2, hCLCA4 is downregulated when EMT is induced by TGF beta, ectopic expression of mesenchymal transcription factors, or dilution of immortalized cells. Furthermore, knockdown of hCLCA4 in HMLE by shRNAs caused EMT. RNA and protein profiling for EMT markers showed that knockdown cells have lower expression epithelial marker E-cadherin and higher expression level of mesenchymal markers vimentin and fibronectin. Moreover, we found that hCLCA4 is co-regulated with hCLCA2 in HMLE cells. Knockdown of hCLCA2 led to down-regulation of hCLCA4 and vice versa. These findings suggest that hCLCA4 is, like hCLCA2, a marker of epithelial differentiation whose loss contributes to EMT and possibly metastasis. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2420. doi:1538-7445.AM2012-2420

<div>Abstract<p>The epithelial–mesenchymal transition (EMT) has been postulated as a mechanism by which cancer cells acquire the invasive and stem-like traits necessary for distant metastasis. However, direct <i>in vivo</i> evidence for the role of EMT in the formation of cancer stem–like cells (CSC) and the metastatic cascade remains lacking. Here we report the first isolation and characterization of mesenchymal-like and EMT tumor cells, which harbor both epithelial and mesenchymal characteristics, in an autochthonous murine model of prostate cancer. By crossing the established <i>Pb-Cre</i><sup>+/−</sup><i>;Pten<sup>L/L</sup>;Kras<sup>G12D</sup></i><sup>/+</sup> prostate cancer model with a <i>vimentin-GFP</i> reporter strain, generating <i>CPKV</i> mice, we were able to isolate epithelial, EMT, and mesenchymal-like cancer cells based on expression of vimentin and EpCAM. <i>CPKV</i> mice (but not mice with <i>Pten</i> deletion alone) exhibited expansion of cells with EMT (EpCAM<sup>+</sup>/Vim-GFP<sup>+</sup>) and mesenchymal-like (EpCAM<sup>−</sup>/Vim-GFP<sup>+</sup>) characteristics at the primary tumor site and in circulation. These EMT and mesenchymal-like tumor cells displayed enhanced stemness and invasive character compared with epithelial tumor cells. Moreover, they displayed an enriched tumor-initiating capacity and could regenerate epithelial glandular structures <i>in vivo</i>, indicative of epithelia–mesenchyme plasticity. Interestingly, while mesenchymal-like tumor cells could persist in circulation and survive in the lung following intravenous injection, only epithelial and EMT tumor cells could form macrometastases. Our work extends the evidence that mesenchymal and epithelial states in cancer cells contribute differentially to their capacities for tumor initiation and metastatic seeding, respectively, and that EMT tumor cells exist with plasticity that can contribute to multiple stages of the metastatic cascade. <i>Cancer Res; 75(13); 2749–59. ©2015 AACR</i>.</p></div>

<div>Abstract<p><b>Purpose:</b> MicroRNAs in the delta-like 1 homolog–deiodinase, iodothyronine 3 (DLK1-DIO3) cluster have been shown to be critical for embryonic development and epithelial to mesenchymal transition (EMT). DLK1-DIO3 cluster miRNAs are elevated in the serum of patients with metastatic cancer. However, the biologic functions of these miRNAs in the EMT and metastasis of cancer cells are poorly understood. We previously demonstrated the oncogenic and metastatic role of miR-409-3p/5p, a member of this cluster, in prostate cancer. In this study, we defined the role of miR-154* and miR-379, two key members of this cluster, in prostate cancer progression and bone metastasis in both cell line models and clinical specimens.</p><p><b>Experimental Design:</b> Genetic manipulation of miR-154* and miR-379 was performed to determine their role in tumor growth, EMT, and bone metastasis in mouse models. We determined the expression of miR-154* in prostate cancer clinical samples and bone metastasis samples using <i>in situ</i> hybridization and quantum dot labeling.</p><p><b>Results:</b> Elevated expression of miR-154* and miR-379 was observed in bone metastatic prostate cancer cell lines and tissues, and miR-379 expression correlated with progression-free survival of patients with prostate cancer. Intracardiac inoculation (to mimic systemic dissemination) of miR-154* inhibitor-treated bone metastatic ARCaP<sub>M</sub> prostate cancer cells in mice led to decreased bone metastasis and increased survival.</p><p><b>Conclusion:</b> miR-154* and miR-379 play important roles in prostate cancer biology by facilitating tumor growth, EMT, and bone metastasis. This finding has particular translational importance because miRNAs in the DLK1-DIO3 cluster can be attractive biomarkers and possible therapeutic targets to treat bone metastatic prostate cancer. <i>Clin Cancer Res; 20(24); 6559–69. ©2014 AACR</i>.</p></div>

Pancreatic ductal adenocarcinoma (PDAC) has an extremely poor prognosis, due in part to early invasion and metastasis, which in turn involves epithelial–mesenchymal transition (EMT) of the cancer cells. Prompted by the discovery that two PDAC cell lines of the quasi-mesenchymal subtype (PANC-1, MIA PaCa-2) exhibit neuroendocrine differentiation (NED), we asked whether NED is associated with EMT. Using real-time PCR and immunoblotting, we initially verified endogenous expressions of various NED markers, i.e., chromogranin A (CHGA), synaptophysin (SYP), somatostatin receptor 2 (SSTR2), and SSTR5 in PANC-1 and MIA PaCa-2 cells. By means of immunohistochemistry, the expressions of CHGA, SYP, SSTR2, and the EMT markers cytokeratin 7 (CK7) and vimentin could be allocated to the neoplastic ductal epithelial cells of pancreatic ducts in surgically resected tissues from patients with PDAC. In HPDE6c7 normal pancreatic duct epithelial cells and in epithelial subtype BxPC-3 PDAC cells, the expression of CHGA, SYP, and neuron-specific enolase 2 (NSE) was either undetectable or much lower than in PANC-1 and MIA PaCa-2 cells. Parental cultures of PANC-1 cells exhibit EM plasticity (EMP) and harbor clonal subpopulations with both M- and E-phenotypes. Of note, M-type clones were found to display more pronounced NED than E-type clones. Inducing EMT in parental cultures of PANC-1 cells by treatment with transforming growth factor-β1 (TGF-β1) repressed epithelial genes and co-induced mesenchymal and NED genes, except for SSTR5. Surprisingly, treatment with bone morphogenetic protein (BMP)-7 differentially affected gene expressions in PANC-1, MIA PaCa-2, BxPC-3, and HPDE cells. It synergized with TGF-β1 in the induction of vimentin, SNAIL, SSTR2, and NSE but antagonized it in the regulation of CHGA and SSTR5. Phospho-immunoblotting in M- and E-type PANC-1 clones revealed that both TGF-β1 and, surprisingly, also BMP-7 activated SMAD2 and SMAD3 and that in M- but not E-type clones BMP-7 was able to dramatically enhance the activation of SMAD3. From these data, we conclude that in EMT of PDAC cells mesenchymal and NED markers are co-regulated, and that mesenchymal–epithelial transition (MET) is associated with a loss of both the mesenchymal and NED phenotypes. Analyzing NED in another tumor type, small cell carcinoma of the ovary hypercalcemic type (SCCOHT), revealed that two model cell lines of this disease (SCCOHT-1, BIN-67) do express CDH1, SNAI1, VIM, CHGA, SYP, ENO2, and SSTR2, but that in contrast to BMP-7, none of these genes was transcriptionally regulated by TGF-β1. Likewise, in BIN-67 cells, BMP-7 was able to reduce proliferation, while in SCCOHT-1 cells this occurred only upon combined treatment with TGF-β and BMP-7. We conclude that in PDAC-derived tumor cells, NED is closely linked to EMT and TGF-β signaling, which may have implications for the therapeutic use of TGF-β inhibitors in PDAC management.

The epithelial-mesenchymal transition (EMT) is a process through which epithelial cells trans-differentiate and acquire an aggressive mesenchymal phenotype. In tumor cells, EMT is a vital step of tumor progression and metastasis. Amid the increasing interest in tumor EMT, only a few studies focused on the soluble mediators secreted by tumor cells passing through this phenotypic switch. In this review, we focus on the essential role of interleukin-8 (IL-8) signaling for the acquisition and maintenance of tumor EMT via direct and indirect mechanisms. Besides the autocrine loop between IL-8 and tumor cells that have gone through EMT, IL-8 could potentiate adjacent epithelial tumor cells into a mesenchymal phenotype via a paracrine mode. Moreover, understanding the role of IL-8 in EMT will provide insight into the pathogenesis of tumor progression and may facilitate the development of an effective strategy for the prevention and treatment of metastatic cancer.

Recent studies have highlighted tumor microenvironment as important in carcinogenesis and progression. Especially in prostate cancer (PC), it has been suggested that tumor microenvironment plays an important role in progression, acquisition of androgen independence, and distant metastasis. However, little is known about the genetic basis of human PC microenvironment. The prognosis of PC is related to invasion and metastasis. Epithelial-to-mesenchymal transition (EMT) has gained considerable attention as a conceptual paradigm to explain invasive and metastatic behavior during cancer progression. EMT is a normal physiologic process by which cells of epithelial origin convert into cells with mesenchymal characteristics. It has been proposed that EMT is induced by cancer cells during metastatic dissemination from a primary organ to secondary sites, but how EMT occurs during PC invasion and metastasis remains uncertain. To clarify the mechanism of PC progression and metastasis, we investigated cross-interaction of PC epithelial cells and stromal cells by analyzing genome-wide gene expression profiles. We hypothesized that PC cells could affect on stromal cells to change their characteristics into so-called cancer-associated fibroblasts (CAFs), which might contribute to cancer invasion and metastasis. Likewise, CAFs could affect on surrounding normal epithelial cells to change their characteristics into PC cells. We first analyzed three of genome-wide gene expression profiles; normal human prostate stromal cells (PrSC) co-cultured with human prostate cancer cells (LNCaP), LNCaP co-cultured with PrSC, and human prostate epithelial cells (PrEC) co-cultured with cancer-associated PrSC which is PrSC previously co-cultured with LNCaP. Before microarray analysis, we confirmed the overexpression of alpha-smooth muscle actin in PrSC co-cultured with LNCaP, which means the transition from normal stromal cells to CAFs. We then analyzed networks among PC cells, stromal cells, and surrounding normal epithelial cells by analyzing gene expression profiles and validated our data using real-time PCR, immunohistochemistry, and bioinformatics. These data provide clues to the molecular mechanisms underlying PC invasion and metastasis, and suggest candidate genes the products of which might serve as molecular targets for the treatment of PC. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4282. doi:1538-7445.AM2012-4282

Distant metastasis of tumor cell is a series of continuous, selectable cascades of events regulated by multiple factors and genes. Epithelial-mesenchymal transition (EMT) is a critical step during cancer metastasis. However, the mechanism of EMT in tumor is not yet fully elucidated. MicroRNAs (miRNAs) are a class of non-coding small endogenous RNAs that negatively regulate EMT-related genes at the post-transcriptional level and play critical roles in cancer metastasis. This review mainly focuses on the topics that include the relationship of EMT and tumor metastasis, transcription factors involved in EMT, and the effect of miRNAs on tumor metastasis by targeting the EMT-related transcription factors.

The acquisition of highly malignant traits by carcinoma cells is often associated with the activation of a transdifferentiation program termed the epithelial-mesenchymal transition (EMT). The EMT program is deployed in a variety of steps of embryonic morphogenesis, enabling the interconversion of epithelial and mesenchymal cell types. When both normal and neoplastic epithelial cells undergo an EMT, they acquire addition changes in addition to expressing characteristic mesenchymal cell markers. In particular, they acquire motility, an increased resistance to apoptosis and, in the case of carcinoma cells, invasiveness. The motility and invasiveness components associated with the EMT would appear to empower many types of cancer cells to metastasize. An interesting question is how the resulting disseminated cancer cells are able to spawn a macroscopic tumor mass at distant anatomical sites, since this clonal expansion would appear to require an ability to self-renew. Recently, we have examined the cellular products of an EMT. Contrary to our expectations, the resulting cells acquired many of the attributes of stem cells, including the expression of stem cell-associated cell-surface markers. Conversely, mammary epithelial cells that naturally express these markers also express greatly elevated levels of at least four pleiotropically acting transcription factors that have been found able to induce an EMT. More recent studies have confirmed that the product of an EMT in mouse mammary epithelial cells is a cell that has a greatly increased ability to generate an entire mammary ductal epithelial tree. Moreover, we find that cells with the attributes of stem cells exist naturally as minor subpopulations in cultured human mammary epithelial cells. These cells can differentiate spontaneously into non-stem cells and, conversely, the non-stem cells appear capable of dedifferentiating spontaneously into cells with progenitor or stem-cell phenotypes. Hence, the study of certain aspects of mammary epithelial cell biology, including the biology of mammary carcinoma stem cells, may be feasible with cultured cells. Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr A1-1.

Transitions between epithelial and mesenchymal states in development and disease

Objective To investigate the expressions of Yes-associated protein-1 (YAP1) in gallbladder mucosal epithelium of normal persons, in patients with simple/calculous cholecystitis, and in patients with gallbladder carcinoma; and to study the mechanism of YAP1 in gallbladder carcinoma development. Methods Immunohistochemistry was used to detect the expression and distribution of YAP1 protein in 50 persons with normal gallbladder, 101 patients with simple cholecystitis/calculous cholecystitis and 100 patients with gallbladder carcinoma. RT-PCR and western-blot were used to detect the mRNA and protein levels of YAP1 in normal and malignant gallbladder mucosal epithelium cells. siRNA was used to shut down the expression of YAP1 in SGC996 cells. MTT was used to test cell vitality. Flow cytometry was used to measure cell cycle. Results Immunohistochemistry revealed the expression rates of YAP1 in the gallbladder carcinoma group, the cholecystitis/gallstone group and the control group to be 87.0% (87/100), 56.4% (57/101) and 5.0% (1/20), respectively (P<0.01). The YAP1 protein levels were higher in gallbladder carcinoma tissues and cells when compared to normal tissues and cells. RT-PCR showed the mRNA levels of gallbladder carcinoma cells to be 12.5±1.2 times of normal gallbladder mucosal epithelial cells (P<0.05). After using siRNA to shut down the YAP1 expression, EMT associated proteins were down-regulated, cell vitality was decreased, and cell cycle was arrested in the S-phase. Conclusions YAP1 is closely related to cell proliferation and metastasis of gallbladder carcinoma. It may promote tumor progression through epithelial-mesenchymal transition. Key words: Primary gallbladder carcinoma; Yes-associated protein 1; Epithelial-mesenchymal transition; Tumor progress

BackgroundThe airway epithelial cell plays a central role in coordinating the pulmonary response to injury and inflammation. Here, transforming growth factor-β (TGFβ) activates gene expression programs to induce stem cell-like properties, inhibit expression of differentiated epithelial adhesion proteins and express mesenchymal contractile proteins. This process is known as epithelial mesenchymal transition (EMT); although much is known about the role of EMT in cellular metastasis in an oncogene-transformed cell, less is known about Type II EMT, that occurring in normal epithelial cells. In this study, we applied next generation sequencing (RNA-Seq) in primary human airway epithelial cells to understand the gene program controlling Type II EMT and how cytokine-induced inflammation modifies it.ResultsGeneralized linear modeling was performed on a two-factor RNA-Seq experiment of 6 treatments of telomerase immortalized human small airway epithelial cells (3 replicates). Using a stringent cut-off, we identified 3,478 differentially expressed genes (DEGs) in response to EMT. Unbiased transcription factor enrichment analysis identified three clusters of EMT regulators, one including SMADs/TP63 and another NF-κB/RelA. Surprisingly, we also observed 527 of the EMT DEGs were also regulated by the TNF-NF-κB/RelA pathway. This Type II EMT program was compared to Type III EMT in TGFβ stimulated A549 alveolar lung cancer cells, revealing significant functional differences. Moreover, we observe that Type II EMT modifies the outcome of the TNF program, reducing IFN signaling and enhancing integrin signaling. We confirmed experimentally that TGFβ-induced the NF-κB/RelA pathway by observing a 2-fold change in NF-κB/RelA nuclear translocation. A small molecule IKK inhibitor blocked TGFβ-induced core transcription factor (SNAIL1, ZEB1 and Twist1) and mesenchymal gene (FN1 and VIM) expression.ConclusionsThese data indicate that NF-κB/RelA controls a SMAD-independent gene network whose regulation is required for initiation of Type II EMT. Type II EMT dramatically affects the induction and kinetics of TNF-dependent gene networks.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-015-1707-x) contains supplementary material, which is available to authorized users.

In normal endometrium, stromal factors regulate the growth of epithelial cells. However, epithelial cells in endometriotic lesions display increased proliferation and decreased apoptosis. This work tested the hypothesis that in endometriosis stromal cells lose the ability to regulate survival signaling and cell growth in epithelial cells. Primary normal, endometriotic eutopic and ectopic epithelial cells were cultured in the presence of medium conditioned by normal, eutopic and ectopic endometriotic endometrial stromal cells. Endometriotic epithelial cells showed higher Survivin expression than normal epithelial cells. Conditioned medium (CM) from normal or eutopic endometriotic stromal cells significantly inhibited the Survivin expression and AKt phosphorylation in normal or eutopic endometriotic epithelial cells. However, CM from ectopic endometriotic stromal cells did not have an inhibitory effect on normal or ectopic endometriotic epithelial cells. Inhibition of AKt phosphorylation and Survivin expression in normal or eutopic endometriotic epithelial cells in the presence of stromal factors from normal or eutopic endometriotic stromal cells was enhanced by progesterone, whereas progesterone had little effect in the presence of stromal factors from ectopic endometriotic stromal cells. The inability of ectopic endometriotic stromal cells to regulated PI3K/AKt/Survivin signaling and mediate the progesterone response in endometriotic epithelial cells may facilitate epithelial cell proliferation in endometriosis and promote the survival of endometriotic lesions.

BACKGROUNDThere is growing evidence proving that many human carcinomas, including colon cancer, can overexpress immunoglobulin (Ig); the non B cancer cell-derived Ig usually displayed unique V(D)J rearrangement pattern that are distinct from B cell-derived Ig. Especially, the cancer-derived Ig plays important roles in cancer initiation, progression, and metastasis. However, it still remains unclear if the colon cancer-derived Ig can display unique V(D)J pattern and sequencing, which can be used as novel target for colon cancer therapy.AIMTo investigate the Ig repertoire features expressed in human colon cancer cells.METHODSSeven cancerous tissue samples of colon adenocarcinoma and corresponding noncancerous tissue samples were sorted by fluorescence-activated cell sorting using epithelial cell adhesion molecule as a marker for epithelial cells. Ig repertoire sequencing was used to analyze the expression profiles of all 5 classes of Ig heavy chains (IgH) and the Ig repertoire in colon cancer cells and corresponding normal epithelial cells.RESULTSWe found that all 5 IgH classes can be expressed in both colon cancer cells and normal epithelial cells. Surprisingly, unlike the normal colonic epithelial cells that expressed 5 Ig classes, our results suggested that cancer cells most prominently express IgG. Next, we found that the usage of Ig in cancer cells caused the expression of some unique Ig repertoires compared to normal cells. Some VH segments, such as VH3-7, have been used in cancer cells, and VH3-74 was frequently present in normal epithelial cells. Moreover, compared to the normal cell-derived Ig, most cancer cell-derived Ig showed unique VHDJH patterns. Importantly, even if the same VHDJH pattern was seen in cancer cells and normal cells, cancer cell-derived IgH always displayed distinct hypermutation hot points.CONCLUSIONWe found that colon cancer cells could frequently express IgG and unique IgH repertoires, which may be involved in carcinogenesis of colon cancer. The unique IgH repertoire has the potential to be used as a novel target in immune therapy for colon cancer.