Colon Tumorigenesis In Mice Research Articles

Neohesperidin protects against colitis-associated colorectal cancer in mice via suppression of the NF-κB/p65 and MAPK pathways.

Patients with inflammatory bowel disease (IBD) are at increased risk of developing colitis-associated colorectal cancer (CAC). Neohesperidin (NHP), a flavanone glycoside derived from citrus fruits, has been reported to have anti-inflammatory, antioxidant, and anticancer potential. However, the function of NHP on tumorigenesis has not been well understood. To investigate the potential chemopreventive effects of NHP on CAC development, an in vivo azoxymethane (AOM)/dextran sulfate sodium (DSS)-induced mouse model was used and NHP was administered by daily gavage for 10 weeks throughout the model period. In this study, we found that NHP effectively ameliorated AOM/DSS-induced pathological symptoms of colitis and thus inhibited colon tumorigenesis in mice. NHP treatment attenuated tumor proliferation, induced apoptosis, and inhibited angiogenesis during CAC development. In addition, NHP inhibited macrophage infiltration and reduced the expression of proinflammatory cytokines such as TNF-α, IL-1β, IL-6, and COX-2 at both mRNA and protein levels, and the higher the concentration of NHP, the better the inhibition. It is worth noting that the positive therapeutic agent mesalazine (100 mg/kg) had a therapeutic effect comparable to that of a low concentration of NHP (50 mg/kg), but less effective than the same concentration of NHP (100 mg/kg). In addition, NHP may exert anti-inflammatory and anticancer effects by inhibiting the NF-κB/p65 and ERK/p38 MAPK pathways. Our findings highlight the potential of NHP as a potential therapeutic candidate for IBD and CAC.

PH-sensitive nanoformulation of acetyl-11-keto-beta-boswellic acid (AKBA) as a potential antiproliferative agent in colon adenocarcinoma (in vitro and in vivo)

BackgroundDeveloping a drug delivery system that can transport a higher concentration to the target cells can improve therapeutic efficacy. This study aimed to develop a novel delivery system for acetyl-11-keto-beta-boswellic Acid (AKBA) using chitosan-sodium alginate–calcium chloride (CS-SA-CaCl2) nanoparticles. The objectives were to evaluate the antiproliferative activity of these nanoparticles against colorectal cancer (CRC) cells and to improve the bioavailability and therapeutic efficacy of AKBA.ResultsWith an extraction efficiency of 12.64%, AKBA was successfully extracted from the gum resin of B. serrata. The nanoparticle delivery system exhibited superior cytotoxicity against HT29 cells compared to free AKBA, AKBA extract (BA-Ex), and 5-FU. Furthermore, the nanoformulation (nano-BA-Ex) induced apoptosis in HT29 cells more effectively than the other treatments. In vivo results showed that nanoformulation inhibited chemically induced colon tumorigenesis in mice and significantly reduced the number of aberrant crypt foci (ACFs).ConclusionsThe developed CS-SA-CaCl2 nanoparticles loaded with AKBA extract exhibit potential as a potent drug delivery mechanism for the colorectal cancer model. Nano-BA-Ex is a promising strategy for enhancing the solubility, bioavailability, and therapeutic efficacy of BA derivatives. With its multiple effects on cancer cells and controlled drug release through nanocapsules, nano-BA-Ex stands out as a compelling candidate for further preclinical and clinical evaluation in CRC therapy.

Vitamin D3 Upregulated Protein 1 Deficiency Promotes Azoxymethane/Dextran Sulfate Sodium-Induced Colorectal Carcinogenesis in Mice.

VDUP1 acts as a tumor suppressor gene in various cancers. VDUP1 is expressed at low levels in sporadic and ulcerative-colitis-associated colorectal cancer. However, the effects of VDUP1 deficiency on CAC remain unclear. In this study, we found that VDUP1 deficiency promoted CAC development in mice. Wild-type (WT) and VDUP1 KO mice were used to investigate the role of VDUP1 in the development of azoxymethane (AOM)- and dextran sulfate sodium (DSS)-induced CAC. VDUP1 levels significantly decreased in the colonic tumor and adjacent nontumoral tissues of WT mice after AOM/DSS treatment. Moreover, AOM/DSS-treated VDUP1 KO mice exhibited a worse survival rate, disease activity index, and tumor burden than WT mice. VDUP1 deficiency significantly induced cell proliferation and anti-apoptosis in tumor tissues of VDUP1 KO mice compared to WT littermates. Additionally, mRNA levels of interleukin-6 and tumor necrosis factor-alpha and active forms of signal transducer and activator of transcription 3 and nuclear factor-kappa B p65 were significantly increased in the tumor tissues of VDUP1 KO mice. Overall, this study demonstrated that the loss of VDUP1 promoted AOM/DSS-induced colon tumorigenesis in mice, highlighting the potential of VDUP1-targeting strategies for colon cancer prevention and treatment.

Partially Hydrolyzed Guar Gum Increases Susceptibility to Colitis and Colon Tumorigenesis in Mice

Partially Hydrolyzed Guar Gum Increases Susceptibility to Colitis and Colon Tumorigenesis in Mice

N-glycosylation Regulates Intrinsic IFN-γ Resistance in Colorectal Cancer: Implications for Immunotherapy

Background & AimsAdvanced colorectal carcinoma (CRC) is characterized by a high frequency of primary immune evasion and refractoriness to immunotherapy. Given the importance of interferon (IFN)-γ in CRC immunosurveillance, we investigated whether and how acquired IFN-γ resistance in tumor cells would promote tumor growth, and whether IFN-γ sensitivity could be restored. MethodsSpontaneous and colitis-associated CRC development was induced in mice with a specific IFN-γ pathway inhibition in intestinal epithelial cells. The influence of IFN-γ pathway gene status and expression on survival was assessed in patients with CRC. The mechanisms underlying IFN-γ resistance were investigated in CRC cell lines. ResultsThe conditional knockout of the IFN-γ receptor in intestinal epithelial cells enhanced spontaneous and colitis-associated colon tumorigenesis in mice, and the loss of IFN-γ receptor α (IFNγRα) expression by tumor cells predicted poor prognosis in patients with CRC. IFNγRα expression was repressed in human CRC cells through changes in N-glycosylation, which decreased protein stability via proteasome-dependent degradation, inhibiting IFNγR-signaling. Downregulation of the bisecting N-acetylglucosaminyltransferase III (MGAT3) expression was associated with IFN-γ resistance in all IFN-γ–resistant cells, and highly correlated with low IFNγRα expression in CRC tissues. Both ectopic and pharmacological reconstitution of MGAT3 expression with all-trans retinoic acid increased bisecting N-glycosylation, as well as IFNγRα protein stability and signaling. ConclusionsTogether, our results demonstrated that tumor-associated changes in N-glycosylation destabilize IFNγRα, causing IFN-γ resistance in CRC. IFN-γ sensitivity could be reestablished through the increase in MGAT3 expression, notably via all-trans retinoic acid treatment, providing new prospects for the treatment of immune-resistant CRC.

Commensal microbiota from patients with inflammatory bowel disease produce genotoxic metabolites.

Microbiota-derived metabolites that elicit DNA damage can contribute to colorectal cancer (CRC). However, the full spectrum of genotoxic chemicals produced by indigenous gut microbes remains to be defined. We established a pipeline to systematically evaluate the genotoxicity of an extensive collection of gut commensals from inflammatory bowel disease patients. We identified isolates from divergent phylogenies whose metabolites caused DNA damage and discovered a distinctive family of genotoxins-termed the indolimines-produced by the CRC-associated species Morganella morganii. A non-indolimine-producing M. morganii mutant lacked genotoxicity and failed to exacerbate colon tumorigenesis in mice. These studies reveal the existence of a previously unexplored universe of genotoxic small molecules from the microbiome that may affect host biology in homeostasis and disease.

Comparative Effects of Traditional Versus Genetically Modified Soybean Oils on Colon Tumorigenesis in Mice.

Soybean oil, which has high abundance of linoleic acid (LA, 18:2ω-6), is the most commonly consumed edible oil. Recent studies support that a high dietary intake of LA is linked with increased risks of developing colonic inflammation and colon cancer. Here we studied the effects of the genetically modified Plenish® soybean oil, which has low abundance of LA as well as α-linolenic acid (ALA, 18:3ω-3), on development of azoxymethane (AOM)/dextran sulfate sodium (DSS)-induced colon tumorigenesis in mice. Compared with a diet rich in traditional soybean oil, administration of a diet enriched with the Plenish oil has little impact on AOM/DSS-induced colon tumorigenesis, colonic infiltration of immune cells, expressions of inflammatory genes, and tumor markers. These results suggest that the traditional and the Plenish soybean oils have similar effects on development of AOM/DSS-induced colon cancer in mice.

Fecal Microbiota Transfer to Determine the Contribution of the Gut Microbiome in a Mouse Model of Western Diet-Enhanced Colorectal Cancer

ObjectivesThe role of the gut microbiome in the etiology of colorectal cancer (CRC) and the therapeutic potential for modulation of gut microbiota is under intense investigation. Previously, we showed that consumption of the total Western diet (TWD) in mice increased gut inflammation, promoted colon tumorigenesis, and altered the microbiome composition compared to mice fed a healthy diet, AIN93G (AIN). However, it is unclear whether the gut microbiome contributes directly to colitis-associated CRC in this model. The objective of this study was to determine whether fecal microbiota transfer (FMT) from host mice fed either AIN or TWD basal diets would alter colitis symptoms or colitis-associated CRC in recipient mice, which were fed either AIN or TWD directly.MethodsThe resident microbiome of recipient mice was depleted using broad spectrum antibiotics and then replenished by FMT using fecal samples collected during a prior experiment from mice fed either AIN diet (resulting in modest colitis and low tumor burden) or TWD diet (extreme colitis and high tumor burden) in a time-matched fashion by week. Recipient mice were fed either AIN or TWD basal diets, such that four experiment groups were generated: 1) AIN-fed recipients with FMT from AIN-fed host, 2) AIN-fed recipients with FMT from TWD-fed host, 3) TWD-fed recipients with FMT from AIN-fed host, 2) TWD-fed recipients with FMT from TWD-fed host. In recipients, the standard azoxymethane + dextran sodium sulfate model of colitis-associated CRC was used.ResultsPreliminary results indicate that time-matched FMT from mice fed TWD did not significantly enhance symptoms of colitis, colon epithelial inflammation, mucosal injury, or colon tumor burden in recipient mice fed AIN diet. Conversely, FMT from AIN-fed mice did not impart a protective effect for recipient mice fed the TWD.ConclusionsFMT from mice fed either basal diet with differing colitis or tumor outcomes did not shift colitis symptoms of colon tumorigenesis in recipient mice, regardless of the basal diet they consumed. These observations suggest that the gut microbiome may not contribute directly to the development of disease in this animal model. Gut microbiome 16s rRNA sequencing and analyses are pending.Funding SourcesFunding support USDA NIFA grant no. 2018–67,017-27,516.

Polysaccharides from Lachnum sp. Inhibited colitis-associated colon tumorigenesis in mice by modulating fecal microbiota and metabolites

It is still uncertain whether the consumption of Lachnum sp. polysaccharides (LEP) alleviates colorectal cancer (CRC) through the gut microbiota. In this study, our efforts are focused on the influence of LEP on CRC, intestinal barrier and inflammation, and fecal microbiota and the metabolites, in azoxymethane (AOM)/dextran sodium sulfate (DSS)-induced CRC mice. Results showed that LEP inhibited CRC mouse colon shortening and weight loss, decreased tumor incidence, restored intestinal barrier integrity, and reduced excessive inflammation. LEP consumption significantly altered microbiota overall structure and community, with reduced pernicious bacteria (such as Parabacteroides, Escherichia_Shigella, Desulfovibrio and Helicobacter), and increased beneficial bacterium (such as Alistipes, Alloprevotella and Ruminiclostridium). Fecal-metabolome profile indicated that a total of 43 metabolites were clearly changed, with 10 down-regulated and 33 up-regulated metabolites. In addition, short-chain fatty acids (SCFAs), including acetic acid, propionic acid and n-butyric acid, were significantly increased after LEP administration. Moreover, a strong correlation between the fluctuant gut microbiota and metabolites was found. These findings provided not only deeper insights into the responsibility of LEP for CRC alleviation, and but also the potential of LEP as a promising candidate for CRC prevention and treatment.

Bacterial Genotoxin Accelerates Transient Infection-Driven Murine Colon Tumorigenesis.

Chronic and low-grade inflammation associated with persistent bacterial infections has been linked to colon tumor development; however, the impact of transient and self-limited infections in bacterially driven colon tumorigenesis has remained enigmatic. Here we report that UshA is a novel genotoxin in attaching/effacing (A/E) pathogens, which include the human pathogens enteropathogenic Escherichia coli, enterohemorrhagic E. coli, and their murine equivalent Citrobacter rodentium (CR). UshA harbors direct DNA digestion activity with a catalytic histidine-aspartic acid dyad. Injected via the type III secretion system (T3SS) into host cells, UshA triggers DNA damage and initiates tumorigenic transformation during infections in vitro and in vivo. Moreover, UshA plays an indispensable role in CR infection-accelerated colon tumorigenesis in genetically susceptible Apc MinΔ716/+ mice. Collectively, our results reveal that UshA, functioning as a bacterial T3SS-dependent genotoxin, plays a critical role in prompting transient and noninvasive bacterial infection-accelerated colon tumorigenesis in mice. SIGNIFICANCE: We identified UshA, a novel T3SS-dependent genotoxin in A/E pathogens that possesses direct DNA digestion activity and confers bacterially accelerated colon tumorigenesis in mice. Our results demonstrate that acute and noninvasive infection with A/E pathogens harbors a far-reaching impact on the development of colon cancer.This article is highlighted in the In This Issue feature, p. 1.

Loss of aryl hydrocarbon receptor suppresses the response of colonic epithelial cells to IL22 signaling by upregulating SOCS3.

IL22 signaling plays an important role in maintaining gastrointestinal epithelial barrier function, cell proliferation, and protection of intestinal stem cells from genotoxicants. Emerging studies indicate that the aryl hydrocarbon receptor (AhR), a ligand-activated transcription factor, promotes production of IL22 in gut immune cells. However, it remains to be determined if AhR signaling can also affect the responsiveness of colonic epithelial cells to IL22. Here, we show that IL22 treatment induces the phosphorylation of STAT3, inhibits colonic organoid growth, and promotes colonic cell proliferation in vivo. Notably, intestinal cell-specific AhR knockout (KO) reduces responsiveness to IL22 and compromises DNA damage response after exposure to carcinogen, in part due to the enhancement of suppressor of cytokine signaling 3 (SOCS3) expression. Deletion of SOCS3 increases levels of pSTAT3 in AhR KO organoids, and phenocopies the effects of IL22 treatment on wild-type (WT) organoid growth. In addition, pSTAT3 levels are inversely associated with increased azoxymethane/dextran sulfate sodium (AOM/DSS)-induced colon tumorigenesis in AhR KO mice. These findings indicate that AhR function is required for optimal IL22 signaling in colonic epithelial cells and provide rationale for targeting AhR as a means of reducing colon cancer risk.NEW & NOTEWORTHY AhR is a key transcription factor controlling expression of IL22 in gut immune cells. In this study, we show for the first time that AhR signaling also regulates IL22 response in colonic epithelial cells by modulating SOCS3 expression.

Suppression of Colon Tumorigenesis in Mutant Apc Mice by a Novel PDE10 Inhibitor that Reduces Oncogenic β-Catenin.

Previous studies have reported that phosphodiesterase 10A (PDE10) is overexpressed in colon epithelium during early stages of colon tumorigenesis and essential for colon cancer cell growth. Here we describe a novel non-COX inhibitory derivative of the anti-inflammatory drug, sulindac, with selective PDE10 inhibitory activity, ADT 061. ADT 061 potently inhibited the growth of colon cancer cells expressing high levels of PDE10, but not normal colonocytes that do not express PDE10. The concentration range by which ADT 061 inhibited colon cancer cell growth was identical to concentrations that inhibit recombinant PDE10. ADT 061 inhibited PDE10 by a competitive mechanism and did not affect the activity of other PDE isozymes at concentrations that inhibit colon cancer cell growth. Treatment of colon cancer cells with ADT 061 activated cGMP/PKG signaling, induced phosphorylation of oncogenic β-catenin, inhibited Wnt-induced nuclear translocation of β-catenin, and suppressed TCF/LEF transcription at concentrations that inhibit cancer cell growth. Oral administration of ADT 061 resulted in high concentrations in the colon mucosa and significantly suppressed the formation of colon adenomas in the Apc+/min-FCCC mouse model of colorectal cancer without discernable toxicity. These results support the development of ADT 061 for the treatment or prevention of adenomas in individuals at risk of developing colorectal cancer. PREVENTION RELEVANCE: PDE10 is overexpressed in colon tumors whereby inhibition activates cGMP/PKG signaling and suppresses Wnt/β-catenin transcription to selectively induce apoptosis of colon cancer cells. ADT 061 is a novel PDE10 inhibitor that shows promising cancer chemopreventive activity and tolerance in a mouse model of colon cancer.

Loss of Aryl Hydrocarbon Receptor Promotes Colon Tumorigenesis in ApcS580/+; KrasG12D/+ Mice.

The mutational genetic landscape of colorectal cancer has been extensively characterized; however, the ability of "cooperation response genes" to modulate the function of cancer "driver" genes remains largely unknown. In this study, we investigate the role of aryl hydrocarbon receptor (AhR), a ligand-activated transcription factor, in modulating oncogenic cues in the colon. We show that intestinal epithelial cell-targeted AhR knockout (KO) promotes the expansion and clonogenic capacity of colonic stem/progenitor cells harboring ApcS580/+; KrasG12D/+ mutations by upregulating Wnt signaling. The loss of AhR in the gut epithelium increased cell proliferation, reduced mouse survival rate, and promoted cecum and colon tumorigenesis in mice. Mechanistically, the antagonism of Wnt signaling induced by Lgr5 haploinsufficiency attenuated the effects of AhR KO on cecum and colon tumorigenesis. IMPLICATIONS: Our findings reveal that AhR signaling plays a protective role in genetically induced colon tumorigenesis at least by suppressing Wnt signaling and provides rationale for the AhR as a therapeutic target for cancer prevention and treatment.

Chemoprevention of Colon Cancer by Dietary Intake of Sulforaphane

BackgroundSulforaphane (SFN), a phytochemical compound, which belongs to isothiocyanates family found in abundance in broccoli sprouts, potently induces a variety of antioxidant enzymes via NF‐E2‐related factor 2‐Kelch‐like ECH‐associated protein 1‐mediated pathway, thereby protects cells from injury induced by various kinds of oxidative stresses. We have previously shown that SFN protects gastric mucosa from oxidative injury induced by H. pyloriinfection. SFN also down‐regulates histone deacetylase (HDAC) activity, thereby induces apoptosis and inhibits proliferation of tumor cells in variety of tissues. On the other hand, colon cancer has been increasing in Japan. Since numerous epidemiological studies have shown that colon cancer is inversely associated with intake of anti‐oxidant vegetables, we examined if daily intake of SFN prevents colon tumorigenesis in mice, and in human subjects.Methods1. Effects of SFN on Colonic tumorigenesis in Mice Treated with Chemical Carcinogen: Effects of SFN on colonic tumorigenesis were examined in the ICR male mice, pretreated with a chemical carcinogen, azoxymethane (AOM) (15 mg/kg). The mice treated with AOM for 3 or 6 times were fed for 8 or 24 weeks with or without sulforaphane glucosinolates (SGS: 2,200 ppm/kg/day), which is a precursor of SFN. Effects of SGS treatment on formation of the microscopic aberrant crypt foci (ACF), and the macroscopic tumors in colonic mucosa were evaluated. 2. Effects of SFN on formation of Colonic ACF in patients with colonic adenoma: Effects of intake of raw broccoli sprouts (BS), 50 g/day, which contains 220 mg SGS every other day, for 6 months on changes in the number of ACF in rectal mucosa was examined by colonoscopy in patients with colonic adenoma. 3. Effects of SFN on intestinal microbiota in human subjects: Effects of dietary intake of raw BS, 20 g/day, which contains 88 mg SGS every other day, for 2 weeks on intestinal microbiota in healthy volunteers was assessed by measuring composition of stool bacteria, using a method of terminal restriction fragment length polymorphism flora analysis. In human studies, alfalfa sprouts, which contains no SFN was used as a placebo control.Results1. Daily administration of SGS suppressed formation of microscopic ACF and macroscopic colonic tumors in the AOM‐pretreated mice in vivo. 2. Intake of BS for 6 months tended to decrease the number of colonic ACF in patients with colonic adenoma. 3. Intake of BS for 2 weeks increased percentages of Bifidobacterium and Clostridium XVIa, which has been shown to enhance protection of colonic mucosa by increasing butyrate production in colonic lumen.Conclusions1. Daily intake of SFN affords chemoprotection against colonic tumors in the mice treated with a chemical carcinogen. 2. The present study further suggests that, in addition to previously reported mechanisms, changes in the intestinal microbiota by SFN intake may also play a role in chemoprevention against colon cancer. Further studies are required to prove this possibility.Support or Funding InformationThis study was supported in part by Murakami Farm Inc., Japan.

Expression of Free Fatty Acid Receptor 2 by Dendritic Cells Prevents Their Expression of Interleukin 27 and Is Required for Maintenance of Mucosal Barrier and Immune Response Against Colorectal Tumors in Mice

Intestinal microbes and their metabolites affect the development of colorectal cancer (CRC). Short-chain fatty acids are metabolites generated by intestinal microbes from dietary fiber. We investigated the mechanisms by which free fatty acid receptor 2 (FFAR2), a receptor for short-chain fatty acids that can affect the composition of the intestinal microbiome, contributes to the pathogenesis of CRC. We performed studies with ApcMin/+ mice, ApcMin/+Ffar2-/- mice, mice with conditional disruption of Ffar2 in dendritic cells (DCs) (Ffar2fl/flCD11c-Cre mice), ApcMin/+Ffar2fl/flCD11c-Cre mice, and Ffar2fl/fl mice (controls); some mice were given dextran sodium sulfate to induce colitis, with or without a FFAR2 agonist or an antibody against interleukin 27 (IL27). Colon and tumor tissues were analyzed by histology, quantitative polymerase chain reaction, and 16S ribosomal RNA gene sequencing; lamina propria and mesenteric lymph node tissues were analyzed by RNA sequencing and flow cytometry. Intestinal permeability was measured after gavage with fluorescently labeled dextran. We collected data on colorectal tumors from The Cancer Genome Atlas. ApcMin/+Ffar2-/- mice developed significantly more spontaneous colon tumors than ApcMin/+ mice and had increased gut permeability before tumor development, associated with reduced expression of E-cadherin. Colon tumors from ApcMin/+Ffar2-/- mice had a higher number of bacteria than tumors from ApcMin/+ mice, as well as higher frequencies of CD39+CD8+ T cells and exhausted or dying T cells. DCs from ApcMin/+Ffar2-/- mice had an altered state of activation, increased death, and higher production of IL27. Administration of an antibody against IL27 reduced the numbers of colon tumors in ApcMin/+ mice with colitis. Frequencies of CD39+CD8+ T cells and IL27+ DCs were increased in colon lamina propria from Ffar2fl/flCD11c-Cre mice with colitis compared with control mice or mice without colitis. ApcMin/+Ffar2fl/flCD11c-Cre mice developed even more tumors than ApcMin/+Ffar2fl/fl mice, and their tumors had even higher numbers of IL27+ DCs. ApcMin/+ mice with colitis given the FFAR2 agonist developed fewer colon tumors, with fewer IL27+ DCs, than mice not given the agonist. DCs incubated with the FFAR2 agonist no longer had gene expression patterns associated with activation or IL27 production. Loss of FFAR2 promotes colon tumorigenesis in mice by reducing gut barrier integrity, increasing tumor bacterial load, promoting exhaustion of CD8+ T cells, and overactivating DCs, leading to their death. Antibodies against IL27 and an FFAR2 agonist reduce tumorigenesis in mice and might be developed for the treatment of CRC.

Triclocarban exposure exaggerates colitis and colon tumorigenesis: roles of gut microbiota involved

ABSTRACT Triclocarban (TCC) is a widely used antimicrobial ingredient in consumer products and is a ubiquitous contaminant in the environment. In 2016, the FDA removed TCC from over-the-counter handwashing products, but this compound is still approved for use in many other personal care products. A better understanding of its impact on human health could lead to significant impact for public health and regulatory policies. Here we show that exposure to low-dose TCC exaggerated the severity of colitis and exacerbated the development of colitis-associated colon tumorigenesis, via gut microbiota-dependent mechanisms. Exposure to TCC increased dextran sodium sulfate (DSS)- and interleukin 10 (IL-10) knockout-induced colitis, and exaggerated azoxymethane (AOM)/DSS-induced colon tumorigenesis in mice. Regarding the mechanisms, TCC exposure reduced the diversity and altered the composition of gut microbiota and failed to promote DSS-induced colitis in mice lacking the microbiota, supporting that the presence of the microbiota is critical for the pro-colitis effects of TCC. Together, these results support TCC could be a novel risk factor for colitis and colitis-associated colon cancer, and further regulatory policies on this compound could be needed.

Thermally Processed Oil Exaggerates Colonic Inflammation and Colitis-Associated Colon Tumorigenesis in Mice.

Frying in vegetable oil is a popular cooking and food processing method worldwide; as a result, the oils used for frying are widely consumed by the general public and it is of practical importance to better understand their health impacts. To date, the effects of frying oil consumption on human health are inconclusive, making it difficult to establish dietary recommendations or guidelines. Here we show that dietary administration of frying oil, which was prepared under the conditions of good commercial practice, exaggerated dextran sodium sulfate (DSS)-induced colitis and azoxymethane (AOM)/DSS-induced colon tumorigenesis in mice. In addition, dietary administration of frying oil impaired intestinal barrier function, enhanced translocation of lipopolysaccharide (LPS) and bacteria from the gut into the systemic circulation, and increased tissue inflammation. Finally, to explore the potential compounds involved in the actions of the frying oil, we isolated polar compounds from the frying oil and found that administration of the polar compounds exacerbated DSS-induced colitis in mice. Together, our results showed that dietary administration of frying oil exaggerated development of inflammatory bowel disease (IBD) and IBD-associated colon tumorigenesis in mice, and these effects could be mediated by the polar compounds in the frying oil.

Trp53 null and R270H mutant alleles have comparable effects in regulating invasion, metastasis, and gene expression in mouse colon tumorigenesis

Somatic APC (adenomatous polyposis coli), TP53, KRAS mutations are present in roughly 80%, 60%, and 40%, respectively, of human colorectal cancers (CRCs). Most TP53 mutant alleles in CRCs encode missense mutant proteins with loss-of-function (LOF) of p53's transcriptional activity and dominant negative (DN) effects on wild-type p53 function. Missense mutant p53 proteins have been reported to exert gain-of-function (GOF) effects in cancer. We compared the phenotypic effects of the common human cancer-associated TP53 R273H missense mutation to p53 null status in a genetically engineered mouse CRC model. Inactivation of one allele of Apc together with activation of a Kras mutant allele in mouse colon epithelium instigated development of serrated and hyperplastic epithelium and adenomas (AK mice). Addition of a Trp53R270H or Trp53null mutant allele to the model (AKP mice) led to markedly shortened survival and increased tumor burden relative to that of AK mice, including adenocarcinomas in AKP mice. Comparable life span and tumor burden were seen in AKP mice carrying Trp53R270H or Trp53null alleles, along with similar frequencies of spontaneous metastasis to lymph nodes, lung, and liver. The fraction of adenocarcinomas with submucosa or deeper invasion was higher in AKP270/fl mice than in AKPfl/fl mice, but the incidence of adenocarcinomas per mouse did not differ significantly between AKPfl/fl and AKP270/fl mice. In line with their comparable biological behaviors, mouse primary tumors and tumor-derived organoids with the Trp53R270H or Trp53null alleles had highly similar gene expression profiles. Human CRCs with TP53 R273 missense mutant or null alleles also had essentially homogeneous gene expression patterns. Our findings indicate the R270H/R273H p53 mutant protein does not manifest definite GOF biological effects in mouse and human CRCs, suggesting possible GOF effects of mutant p53 in cancer phenotypes are likely allele-specific and/or context-dependent.

Chemoprevention against colon cancer by dietary intake of sulforaphane

Background: Sulforaphane (SFN), a phytochemical compound, which belongs to isothiocyanates family found in abundance in broccoli sprouts, potently induces a variety of antioxidant enzymes via nrf2-keap1 mediated pathway, thereby protects cells from injury induced by various kinds of oxidative stresses. We have previously shown that SFN protects gastric mucosa from oxidative injury induced by H. pylori infection. SFN also down-regulates histone deacetylase (HDAC) activity, thereby induces apoptosis and inhibits proliferation of tumor cells in variety of tissues. On the other hand, colon cancer has been increasing in Japan. Since numerous epidemiological studies have shown that colon cancer is inversely associated with intake of anti-oxidant vegetables, we examined if dietary SGS prevents colon tumorigenesis in mice, and in human subjects.Methods: 1. Effects of SFN on Colonic tumorigenesis in Mice Treated with Chemical Carcinogen: Effects of SFN on colonic tumorigenesis were examined in the ICR male mice, pretreated with a chemical carcinogen, azoxymethane (AOM) (15 mg/kg). The AOM-treated mice were fed for 8 weeks with or without sulforaphane glucosinolates (SGS: 2,200 ppm/kg/day), which is a precursor of SFN. Effects of SGS treatment on formation of the microscopic aberrant crypt foci (ACF), and the macroscopic tumors in colonic mucosa were evaluated. 2. Effects of SFN on formation of Colonic Aberrant Crypt Foci (ACF) in patients with colonic adenoma: Effects of intake of raw broccoli sprouts (BS), 50 g/day, which contains 220 mg SGS every other day, for 6 months on changes in the number of ACF in rectal mucosa was examined by colonoscopy in patients with colonic adenoma. 3. Effects of SFN on intestinal microbiota in human subjects: Effects of dietary intake of raw broccoli sprouts (BS), 20 g/day, which contains 88 mg SGS every other day, for 2 weeks on intestinal microbiota in healthy volunteers was assessed by measuring composition of stool bacteria, using T-RFLP method. In human studies, alfalfa sprouts, which contains no SFN was used as a placebo control. Results: 1. Daily administration of SGS suppressed formation of microscopic ACF and macroscopic colonic tumors in the AOM-pretreated mice in vivo. 2. Intake of BS for 6 months tended to decrease the number of colonic ACF in patients with colonic adenoma. 3. Intake of BS for 2 weeks increased percentages of Bifidobacterium and Clostridium XVIa, which has been shown to enhance protection of colonic mucosa by increasing butyrate production in colonic lumen.Conclusions: 1. SFN affords chemoprotection against colonic cancer, presumably by up-regulating nrf2-dependent antioxidant enzymes in normal colonic epithelial cells, and by inducing apoptosis of colonic tumor cells through inhibition of HDAC activity. The present study further suggests that changes in intestinal microbiota by SFN, may play a role in chemoprevention against colon cancer.Keywords: sulforaphane, colon cancer, chemoprevention, aberrant crypt foci, intestinal microbiota

Abstract 2601: HMGA1 is induced by procarcinogenic bacteria within the microbiome where it drives expansion in the colon stem cell pool and tumorigenesis

Abstract Cancer cells undergo chromatin remodeling and epigenetic reprogramming to hijack stem cell transcriptional networks and drive tumorigenesis, although the molecular underpinnings remain unclear. High Mobility Group A (HMGA1) chromatin remodeling proteins are architectural transcription factors that bind AT-rich sequences where they “open” chromatin and recruit transcriptional complexes to modulate gene expression. The HMGA1 gene is highly expressed during embryogenesis, but silenced postnatally in differentiated tissues. HMGA1 is re-expressed in aggressive cancers where high levels portend adverse clinical outcomes. In colorectal cancer, HMGA1 is among the genes most overexpressed compared to normal colon epithelium. We previously found that HMGA1 induces genes involved in EMT and drives tumor progression in colon cancer models. Hmga1 transgenic mice develop aberrant proliferation and polyposis in the small intestine and colon. In intestinal stem cells, Hmga1 amplifies Wnt signals to enhance self-renewal. Surprisingly, Hmga1 also induces Sox-9 to drive Paneth cell differentiation and “build” a stem cell niche. To determine how HMGA1 functions in the colon, we examined mouse models with varied levels of Hmga1. Hmga1 overexpression in colon stem cells leads to expansion of the colon stem cell pool. Both goblet cells and colonic mucous are increased. In DSS models of inflammatory bowel disease, Hmga1 enhances repair and tissue regeneration, while Hmga1 overexpression promotes aberrant proliferation and polyposis with advancing age. To determine how HMGA1 is induced during carcinogenesis, we examined mice harboring a heterozygous Apc mutation (Min+/-) following colonization with the human symbiotic bacteria, enterotoxigenic Bacterioides fragilis (ETBF). ETBF triggers increased Wnt signaling by inducing E-cadherin cleavage with release of membrane bound β-catenin, leading to enhanced distal colon tumorigenesis in mice. Further, ETBF colonization in humans is linked to colon cancer. ETBF colonization in mice induces Hmga1 and increases stem cell self-renewal in preliminary studies. Inflammatory cytokine genes are also induced. Bacteroides fragilis toxin (BFT) induces Hmga1 in wildtype and Min+/- mouse colon organoids. HMGA1 also increases modestly in human HT29/C1 cells after exposure to BFT. Studies are underway to identify transcriptional networks and epigenetic alterations governed by HMGA1 in this setting. This work not only provides new insights into the role of HMGA1 in colon epithelial homeostasis by maintaining both the stem cell pool and promoting regeneration following injury, but also suggests that HMGA1 can be aberrantly induced by signals from the microbiome to promote tumorigenesis. Our results also highlight the interactions between the microbiome and HMGA1 as a potential therapeutic opportunity in colon carcinogenesis. Citation Format: Linda M S Resar, Lingling Xian, Jawara Allen, Shaoguang Wu, Lionel Chia, Cynthia Sears. HMGA1 is induced by procarcinogenic bacteria within the microbiome where it drives expansion in the colon stem cell pool and tumorigenesis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2601.