Carcinogenesis In Mice Research Articles

Inhibition of phospholipase D1 reduces pancreatic carcinogenesis in mice partly through a FAK-dependent mechanism.

Phospholipase D (PLD) plays a critical role in cancer progression. However, its role in pancreatic cancer remains unclear. Thus, we evaluated the role of PLD1, one of two classical isoforms of PLD, in pancreatic carcinogenesis in vivo. The role of PLD1 in tumor growth was evaluated by subcutaneously transplanting human MIA PaCa-2 cells expressing endogenous PLD1 levels (Ctr KD cells) or cells in which PLD1 was knocked down (Pld1 KD cells) into immunodeficient mice. Twenty days post-implantation, tumors that arose from Pld1-KD cells were significantly smaller, compared to controls (Ctr KD). Then, we assessed the role of PLD1 in the tumor microenvironment, by subcutaneously implanting mouse LSL-KrasG12D/+;Trp53R172H/+;Pdx-1-Cre (KPC) cells into wild-type (WT) or PLD1 knockout (Pld1-/-) mice. Compared to WT, tumor growth was attenuated in Pld1-/- mice by 39%, whereas treatment of Pld1-/- mice with gemcitabine reduced tumor growth by 79%. When PLD1 was ablated in LSL-KrasG12D;Ptf1Cre/+ (KC) mice, no reduction in acinar cell loss was observed, compared to KC mice. Finally, treatment of KC mice with a small molecule inhibitor of PLD1 and PLD2 (FIPI) significantly reduced acinar cell loss and cell proliferation, compared to vehicle-treated mice. Mechanistically, the effect of PLD on tumor growth is mediated, partly, by the FAK pathway. In conclusion, while PLD1 is a critical regulator of pancreatic xenograft and allograft growth, playing an important role at the tumor and at the microenvironment levels, inhibition of PLD1 and PLD2 are necessary to reduce pancreatic carcinogenesis in KC mice, and might represent a novel therapeutic target.

Anti-VEGFR2 neutralising antibody slows the progression of multistep oral carcinogenesis.

Angiogenesis plays an important role in cancer growth and metastasis, and it is considered a therapeutic target to control tumour growth following anti-angiogenic therapy. However, it is still unclear when tissues initiate angiogenesis during malignant transformation from premalignant condition and whether this premalignant condition could be a therapeutic target of anti-angiogenic therapy. In this study, we aimed to analyse the onset of angiogenesis by evaluating morphological and functional alterations of microvessels during oral multistep carcinogenesis using a 4-nitroquinoline 1-oxide (4NQO)-induced oral carcinogenesis mouse model. In the study, we initially confirmed that with the use of 4NQO, oral lesions develop in a stepwise manner from normal mucosa through oral epithelial dysplasia (OED) to oral squamous cell carcinoma (OSCC). Evaluation of CD31-immunostained specimens revealed that microvessel density (MVD) increases in a stepwise manner from OEDs. Histological and functional analyses revealed the structural abnormalities and leakage of blood vessels had already taken place in OED. Then we evaluated the expression profiles of Hif1a and Vegfa along with hypoxic status and found that OED exhibited increased Vegfa expression under hypoxic conditions. Finally, we tested the possibility of OEDs as a target of anti-angiogenic therapy and found that anti-VEGFR2 neutralising antibody in OED slowed the disease progression from OED to OSCC. These data indicate that an angiogenic switch occurs at the premalignant stage and morphological, and functional alterations of microvessels already exist in OED. These findings also elucidate the tumour microenvironment, which gradually develops along with carcinogenic processes, and highlight usefulness of the 4NQO-induced carcinogenesis model in the study of epithelial and stromal components, which will support epithelial carcinogenesis. © 2024 The Author(s). The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Antioxidant and Chemopreventive Potential of Uraria picta Root Extracts against DMBA/croton Oil Induced Skin Carcinogenesis in Mice

Antioxidant and Chemopreventive Potential of Uraria picta Root Extracts against DMBA/croton Oil Induced Skin Carcinogenesis in Mice

P2RY6 deletion promotes UVB-induced skin carcinogenesis by activating the PI3K/AKT signal pathway.

Our previous research has demonstrated that P2RY6 functions as an oncogene in DMBA/TPA-induced two-stage chemical skin carcinogenesis in mice. However, considering that human skin cancer is predominantly attributed to UV radiation from sunlight, additional investigations are needed to elucidate the role of P2RY6 in UVB-induced skin carcinogenesis. Surprisingly, we found that P2ry6-deficient mice exhibited marked promotion to UVB-induced skin papilloma formation compared with wild-type mice, suggesting its tumor-suppressive role in UVB-induced skin cancer. Additionally, a P2ry6 gene knockout promoted skin hyperplasia induced by short-term UVB irradiation, while UDP, the ligand of P2RY6, could inhibit the short-term UVB-induced increase of epiderma thickness in mouse skin. Furthermore, UVB irradiation could significantly upregulate P2RY6 expression in human and mouse skin cells. These results indicated that P2RY6 may play a crucial protective role in resisting the UVB-induced formation of skin tumors. At the molecular level, the loss of the P2RY6 gene inhibits the ubiquitination modification and expression of XPC after UVB irradiation in skin keratinocytes, resulting in the accumulation of CPDs (cyclobutane pyrimidine dimers). We have also demonstrated that P2RY6 deletion activates the PI3K/AKT signaling pathway both invitro and invivo. The CPD accumulation and acute inflammatory response enhanced by the loss of the P2RY6 gene can be reversed by an AKT inhibitor. These findings suggest that P2RY6 may act as a tumor suppressor in UVB-induced skin cancer by regulating the PI3K/AKT signaling pathway.

Effects of Vitamin D on tumor cell proliferation and migration, tumor initiation and anti-tumor immune response in head and neck squamous cell carcinomas

BackgroundHead and neck squamous cell carcinomas (HNSCCs) are among the six most common cancers, with a constantly poor prognosis. Vitamin D has been found to have antineoplastic and immunomodulatory properties in various cancers. This study investigated the impact of Vitamin D on the initiation and progression as well as antitumor immune response in HNSCCs, both in vitro and in vivo. MethodsAn immunocompetent, orthotopic oral carcinogenesis mouse model was used to examine the influence of Vitamin D3 substitution on HNSCC initiation and progression in vivo. Tumor immune infiltration was analyzed by immunohistochemistry targeting CD3, CD8, NKR-P1C, FOXP3, and CD163. Two HPV- and two HPV+ HNSCC cell lines were treated with 1,25-dihydroxyvitamin D3 to analyze effects on tumor cell proliferation, migration and transcriptomic changes using RNA-sequencing, differential gene expression and gene set enrichment analysis. ResultsVitamin D3 treatment led to a significant suppression of HNSCC initiation and progression, while also stimulating tumor immune infiltration with CD3+, CD8+ and NKR-P1C+ cells and lowering levels of M2 macrophages and Treg cells in vivo. In vitro experiments showed an inhibition of HNSCC cell proliferation and migration in HPV+ and HPV- cell lines. RNA-sequencing showed significant regulations in IL6 JAK STAT3, hypoxia signaling and immunomodulatory pathways upon Vitamin D3 treatment. ConclusionThe findings of our study highlight the promising potential of Vitamin D in the therapeutic repertoire for HNSCC patients given its immune modulating, anti-proliferative and anti-migratory properties. Clinical transferability of those in vitro and in vivo effects should be further validated in clinical trials.

MAD2L1 supports MYC-driven liver carcinogenesis in mice and predicts poor prognosis in human hepatocarcinoma.

Mitotic arrest deficient 2 like 1 (MAD2L1) is a component of the mitotic spindle assembly checkpoint implicated in cancer cell proliferation and tumorigenesis. The functional role of MAD2L1 in hepatocellular carcinoma (HCC) has not been adequately investigated, especially in vivo. In the current manuscript, we sought to address the function of MAD2L1 in hepatocarcinogenesis. We found that MAD2L1 expression is upregulated in human HCCs, where its expression is associated with higher aggressive tumor grade, elevated proliferative activity, and poor prognosis. In human HCC cell lines, MAD2L1 knockdown led to decreased cell growth. Moreover, RNA-seq results demonstrated that MAD2L1 silencing induces the expression of genes associated with cell cycle, DNA replication, and various cancer-related pathways, supporting the critical role of MAD2L1 during HCC growth and differentiation. In a c-MYC-induced mouse HCC model, we revealed an increased expression of Mad2l1. Furthermore, Mad2l1 CRIPSR-mediated silencing prevented c-MYC-driven mouse liver development. Altogether, our study suggests that MAD2L1 plays a crucial role in hepatocarcinogenesis, and that its suppression could be a promising therapeutic strategy for treating human HCC.

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.

Gastrodin Attenuates Colitis and Prevents Tumorigenesis in Mice by Interrupting TLR4/MD2/NF-κB Signaling Transduction.

Chronic inflammation is one of the causative factors for tumorigenesis. Gastrodin is a main active ingredient isolated from Gastrodia elata Blume, a famous medicinal herb with a long edible history. This study aimed to explore the effects of gastrodin on colitis-associated carcinogenesis (CRC) in mice and to elucidate its potential molecular mechanisms. Balb/c mice were induced with azoxymethane (AOM) and dextran sulfate sodium (DSS) for 12 weeks. Gastrodin (50 mg/kg) was administered via oral gavage three times per week until the end of the experiment. Disease indexes, including body weight, bloody diarrhea, colon length, histopathological score, and tumor size, were measured. Tumor cell proliferation was evaluated by BrdU incorporation assay and tumor cell cytotoxicity was assessed by cell counting kit (CCK-8). The expression levels of toll-like receptor 4 (TLR4)/nuclear factor kappa-B (NF-κB) signaling molecules, NF-κB luciferase, and pro-inflammatory cytokines were determined by real-time fluorescence quantitative polymerase chain reaction (RT-qPCR), immunoblotting, immunohistochemistry (IHC), enzyme-linked immunosorbent assay (ELISA), or reporter gene assays. The binding affinity between gastrodin and myeloid differentiation protein-2 (MD2) was analyzed by molecular docking and cellular thermal shift assay (CETSA). Gastrodin administration was demonstrated to mitigate various CRC-related symptoms in mice, including weight loss, diarrhea, and tissue abnormalities. Notably, gastrodin suppressed tumor cell growth during colitis- associated tumorigenesis, resulting in fewer and smaller adenomas in the colon. Unlike irinotecan, a broadspectrum antitumor drug, gastrodin did not exhibit apparent cytotoxicity in various colorectal adenocarcinoma cell lines. Additionally, gastrodin downregulated TLR4/NF-κB signaling molecules and pro-inflammatory mediators in mice and macrophages. Molecular docking and CETSA experiments suggested that gastrodin binds to the MD2 protein, potentially interfering with the recognition of lipopolysaccharide (LPS) by TLR4, leading to NF-κB pathway inhibition. This study provides evidence for the first time that gastrodin attenuated colitis and prevented colitisrelated carcinogenesis in mice, at least partially, by diminishing tumor-promoting cytokines through the interruption of TLR4/MD2/NF-κB signaling transduction.

The Impact of Fat Quantity and Composition in Pancreatic Carcinogenesis in Mice: Mitigating Obesity-Induced Tumor Progression With Dietary Changes

The Impact of Fat Quantity and Composition in Pancreatic Carcinogenesis in Mice: Mitigating Obesity-Induced Tumor Progression With Dietary Changes

Integrin-Linked Kinase in the Development of Gastric Tumors Induced by Helicobacter pylori: Regulation and Prevention Potential.

Integrin-linked kinase (ILK) is crucial in solid tumors by regulating the Hippo-Yes-associated protein 1 (YAP) pathway. This study aimed to uncover how Helicobacter pylori influences ILK levels and its role in regulating YAP during H. pylori-induced gastric cancer. GES-1 cells with stable Ilk knockdown and overexpression and a mouse carcinogenesis model for H. pylori infection were constructed. And ILK, the phosphorylated mammalian STE20-like protein kinase 1 (MST1), large tumor suppressor 1 (LATS1; S909, T1079), and YAP (S109, S127) were detected in cells, and mice by western blotting, as well as fluorescence intensity of YAP were assayed by immunofluorescence. YAP downstream genes Igfbp4 and Ctgf, the pathological changes and tumor necrosis factor alpha (TNF-α), interleukin-6 (IL-6), interleukin-1beta (IL-1β), and nitric oxide (NO) levels in mice gastric tissues were detected by real-time PCR, H&E, and ELISA assays. In this study, stable Ilk knockdown cells exhibited significantly higher phosphorylated levels of MST1, LATS1, and YAP, as well as increased YAP in the nuclei of GES-1 cells. Conversely, cells with Ilk overexpression showed opposite results. H. pylori infection led to decreased ILK levels in gastric epithelial cells but increased ILK levels in gastric cancer cell lines (MGC803, SGC7901) and gastric cancer tissues in mice. Treatment with the ILK inhibitor OST-T315 elevated the phosphorylated MST, LATS1, and YAP levels, and inhibited the mRNA levels of Igfbp4 and Ctgf at 44, 48 week-aged mice. OST-T315 also reduced the release of TNF-α, IL-6, IL-1β, and NO, as well as the progression of gastric cancer caused by H. pylori and N-Nitroso-N-methylurea (NMU) treatment. Upon initiation of gastric tumorigenesis signals, H. pylori increases ILK levels and suppresses Hippo signaling, thereby promoting YAP activation and gastric cancer progression. ILK can serve as a potential prevention target to impede H. pylori-induced gastric cancer.

A high-fat diet induces changes in mesenteric adipose tissue accelerating early-stage pancreatic carcinogenesis in mice

Increased adiposity is a significant risk factor for pancreatic cancer development. Multiple preclinical studies have documented that high-fat, high calorie diets, rich in omega-6 fatty acids (FA) accelerate pancreatic cancer development. However, the effect of a high-fat, low sucrose diet (HFD), on pancreatic carcinogenesis remains unclear. We evaluated the impact of a HFD on early-stage pancreatic carcinogenesis in the clinically relevant KrasLSL-G12D/+; Ptf1aCre/+ (KC) genetically engineered mouse model, and characterized the role of the mesenteric adipose tissue (MAT). Cohorts of male and female KC mice were randomly assigned to a control diet (CD) or a HFD, matched for FA composition (9:1 of omega-6 FA: omega-3 FA), and fed their diets for 8 weeks. After 8 weeks on a HFD, KC mice had significantly higher body weight, fat mass, and serum leptin compared to CD-fed KC mice. Furthermore, a HFD accelerated pancreatic acinar-to-ductal metaplasia (ADM) and proliferation, associated with increased activation of ERK and STAT3, and macrophage infiltration in the pancreas, compared to CD-fed KC mice. Metabolomics analysis of the MAT revealed sex differences between diet groups. In females, a HFD altered metabolites related to FA (α-linolenic acid and linoleic acid) and amino acid metabolism (alanine, aspartate, glutamate). In males, a HFD significantly affected pathways related to alanine, aspartate, glutamate, linoleic acid, and the citric acid cycle. A HFD accelerates early pancreatic ADM through multifaceted mechanisms, including effects at the tumor and surrounding MAT. The sex-dependent changes in MAT metabolites could explain some of the sex differences in HFD-induced pancreatic ADM.

Comprehensive Proteogenomic Profiling Reveals the Molecular Characteristics of Colorectal Cancer at Distinct Stages of Progression.

Colorectal cancer is the second most common malignant tumor worldwide. Analysis of the changes that occur during colorectal cancer progression could provide insights into the molecular mechanisms driving colorectal cancer development and identify improved treatment strategies. In this study, we performed an integrated multiomic analysis of 435 trace tumor samples from 148 patients with colorectal cancer, covering nontumor, intraepithelial neoplasia (IEN), infiltration, and advanced stage colorectal cancer phases. Proteogenomic analyses demonstrated that KRAS and BRAF mutations were mutually exclusive and elevated oxidative phosphorylation in the IEN phase. Chr17q loss and chr20q gain were also mutually exclusive, which occurred predominantly in the IEN and infiltration phases, respectively, and impacted the cell cycle. Mutations in TP53 were frequent in the advanced stage colorectal cancer phase and associated with the tumor microenvironment, including increased extracellular matrix rigidity and stromal infiltration. Analysis of the profiles of colorectal cancer based on consensus molecular subtype and colorectal cancer intrinsic subtype classifications revealed the progression paths of each subtype and indicated that microsatellite instability was associated with specific subtype classifications. Additional comparison of molecular characteristics of colorectal cancer based on location showed that ANKRD22 amplification by chr10q23.31 gain enhanced glycolysis in the right-sided colorectal cancer. The AOM/DSS-induced colorectal cancer carcinogenesis mouse model indicated that DDX5 deletion due to chr17q loss promoted colorectal cancer development, consistent with the findings from the patient samples. Collectively, this study provides an informative resource for understanding the driving events of different stages of colorectal cancer and identifying the potential therapeutic targets. Significance: Characterization of the proteogenomic landscape of colorectal cancer during progression provides a multiomic map detailing the alterations in each stage of carcinogenesis and suggesting potential diagnostic and therapeutic approaches for patients.

Evaluation of antitumor activity of saphia alkali K90 on dmba-induced carcinogenesis in swiss mice

The present study aimed to initially evaluate the anti-tumor effect of orally administered Saphia Alkali K90 in 7,12-dimethylbenz(a)anthracene (DMBA)-induced carcinogenesis in Swiss mice. Mice exposed to DMBA (total dose of 6 mg, p.o.) were treated with SAK90 at 15 ml/kg BW and 45 ml/kg BW daily. The vehicle was given to both normal and diseased controls. The change in body weight, mortality rate, tumor incidence, tumor burden, tumor yield, histopathology, and hematological and biochemical parameters were evaluated after 16 weeks of treatment. 23,26% of DMBA-exposed mice presented neoplasms. SAK90 tended to reduce death, tumor incidence, and tumor yield compared to the DMBA group. The formulation exhibited protective effects against DMBA-induced hepatotoxicity through a trend in drop transaminase levels and increased albumin concentration concurrently compared to the DMBA group. This study suggested that SAK90 has potential anticancer activity.

Pantoprazole suppresses carcinogenesis and growth of hepatocellular carcinoma by inhibiting glycolysis and Na+/H+ exchange.

Hepatocellular carcinoma (HCC) is one of the deadliest cancers. The prevention and therapy for this deadly disease remain a global medical challenge. In this study, we investigated the effect of pantoprazole (PPZ) on the carcinogenesis and growth of HCC. Both diethylnitrosamine (DEN) plus CCl4-induced and DEN plus high fat diet (HFD)-induced HCC models in mice were established. Cytokines and cell proliferation-associated gene in the liver tissues of mice and HCC cells were analyzed. Cellular glycolysis and Na+/H+ exchange activity were measured. The preventive administration of pantoprazole (PPZ) at a clinically relevant low dose markedly suppressed HCC carcinogenesis in both DEN plus CCl4-induced and HFD-induced murine HCC models, whereas the therapeutic administration of PPZ at the dose suppressed the growth of HCC. In the liver tissues of PPZ-treated mice, inflammatory cytokines, IL1, CXCL1, CXCL5, CXCL9, CXCL10, CCL2, CCL5, CCL6, CCL7, CCL20, and CCL22, were reduced. The administration of CXCL1, CXCL5, CCL2, or CCL20 all reversed PPZ-suppressed DEN plus CCL4-induced HCC carcinogenesis in mice. PPZ inhibited the expressions of CCNA2, CCNB2, CCNE2, CDC25C, CDCA5, CDK1, CDK2, TOP2A, TTK, AURKA, and BIRC5 in HCC cells. Further results showed that PPZ reduced the production of these inflammatory cytokines and the expression of these cell proliferation-associated genes through the inhibition of glycolysis and Na+/H+ exchange. In conclusion, PPZ suppresses the carcinogenesis and growth of HCC, which is related to inhibiting the production of inflammatory cytokines and the expression of cell proliferation-associated genes in the liver through the inhibition of glycolysis and Na+/H+ exchange.

5-Methylcoumarin-4β-glucoside mitigated colon tumor progression in mice with AOM/DSS-induced colon carcinogenesis

ContextThe in vitro cytotoxicity profile of 5-Methylcoumarin-4β-glucoside (5-MC4βG) has been demonstrated to inhibit the proliferation of HT-29 adenocarcinoma cells. Aim: The need for newer and affordable chemotherapeutic agents is critical. Main methodsWe investigated the effect of 5-MC4βG on an azoxymethane /dextran sodium sulfate-induced colon carcinogenesis model in six groups of laboratory BALB/c mice for six weeks. While the first and second groups of mice served as vehicle and disease controls respectively, the third, fourth, and fifth groups were administered oral graded doses (25, 50, and 100 mg/kg) of 5-MC4βG. The sixth group was treated with 5-Flourouracil (15 mg/kg). Key findingsThe 100 mg/kg dose of 5-MC4βG upregulated APC mRNA expression by two-fold and downregulated β-catenin mRNA expression by two-fold compared to their respective disease controls. Furthermore, tumorigenic gene transcripts (MDM2, BCL2, CDK1, and cyclin D1) were downregulated by one-fold (except cyclin D1 which was downregulated by two-fold), whereas pro-apoptotic genes (p53, Bax, and Casp3) were upregulated by two-fold following treatment with 100 mg/kg dose of 5-MC4βG. At the metabolic level, the bioactive compound lowered the expression of classical tumor markers; tissue polypeptide antigen, tumor-associated glycoprotein 72, and carcinoembryonic antigen by at least half. Histologically, 5-MC4βG intervention revealed a reduction in neoplastic cells associated with cellular necrosis. Significance5-MC4βG reduced colon carcinogenesis in mice. Thus, this compound may be a promising candidate for colorectal cancer chemotherapy. Further development of 5-MC4βG will hopefully lead to the development of a potential anti-colon cancer drug candidate.

Abstract LB377: The combination of epigallocatechin gallate (EGCG) and resveratrol prevents 4-NQO-induced oral carcinogenesis in mice

Abstract Head and neck cancer (HNC), commonly known as oral cancer, is a devastating disease and the 6th most common cancer worldwide. Most HNC patients are diagnosed with advanced-stage disease for which the 5-year survival is below 50%, stressing the need for chemoprevention. Recently, we have reported that the combination of resveratrol and EGCG induces synergistic apoptosis and inhibits xenografted HNC growth by inhibiting the AKT-mTOR pathway. This study investigated the chemopreventive efficacy of resveratrol, EGCG, and their combination using the 4NQO-induced oral carcinogenesis model. C57BL/6 mice were exposed to 4-NQO (50 μg/ml) via drinking water for 10 weeks, followed by treatment with vehicle (50% sweetened condensed milk), resveratrol (30 mg/kg), EGCG (30 mg/kg) and their combination for 8 weeks, 5 days/week. The mice were sacrificed on week 24, and the number of visible and microscopic lesions was counted. Resveratrol alone and in combination with EGCG significantly inhibited the number of visible lesions. In contrast, the number of microscopic lesions and lesion area was significantly inhibited only in the combination group. The expression of Ki-67 was also significantly inhibited in resveratrol and combination groups when compared with untreated control. Furthermore, RNASeq and qPCR analysis using an HNC cell line identified GDF15, ATF3, p21, p27, and Bim as significantly upregulated genes, with GDF15 being the most upregulated one. Expression of GDF15 and ATF3 proteins was confirmed by Western blotting. Taken together, our data strongly demonstrate the chemopreventive potential of the combination of EGCG and resveratrol and pave the way for further clinical developments. (Supported by NIH Grants R15DE032063 and P20GM103434) Citation Format: Adeoluwa A. Adeluola, Lukmon Raji, Saroj Sigdel, ASM Anisuzzaman, A.R.M. Ruhul Amin. The combination of epigallocatechin gallate (EGCG) and resveratrol prevents 4-NQO-induced oral carcinogenesis in mice [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr LB377.

Vanillic acid protects mortality and toxicity induced by N-ethyl-N-nitrosourea in mice; in vivo model of chronic lymphocytic leukemia

Alkylating agents such as N-Ethyl-N-Nitrosourea (ENU) are ubiquitous within living cells and in the environment. This study designed to evaluate the chemopreventive activity of vanillic acid on ENU-induced toxicity and carcinogenesis in mice as an animal model of chronic lymphocytic leukemia (CLL). The female, Swiss albino mice were divided into three groups each with 7 mice, group I received normal saline, group II, mice received ENU at a dose of 80 mg/kg body weight i.p. to induce CLL on the 31th day of the study, and group III, the mice pretreated with vanillic acid at a dose of 20 mg/kg body weight/day, i.p. up to 30 days and received ENU. The animals were monitored for weight changes and mortality during 120 days, and then were sacrificed for isolation of lymphocytes, as target cells in CLL. Cellular parameters like reactive oxygen species (ROS) formation, malondialdehyde (MDA) production, depletion of glutathione (GSH), mitochondrial membrane potential (MMP) and lysosomal membrane integrity were studied. We found that pretreatment with vanillic acid significantly increased the survival of mice up to 57%, delay in death time (30%) and prevented weight changes after exposure to ENU. In addition, it was found that vanillic acid protected ROS formation, lipid peroxidation mitochondrial dysfunction, and lysosomal membrane destabilization in isolated lymphocytes. These data suggest that vanillic acid exhibited significant protection against ENU-induced toxicity and carcinogenicity, which might be related to the protection of the mitochondria and lysosomes and the reduction of ROS formation and oxidative stress.

Assessment of the mode of action of perchloroethylene-induced mouse liver tumors.

Perchloroethylene (PCE) is used as a solvent and chemical intermediate. Following chronic inhalation exposure, PCE selectively induced liver tumors in mice. Understanding the mode of action (MOA) for PCE carcinogenesis in mice is important in defining its possible human cancer risk. The proposed MOA is based on the extensive examination of the peer-reviewed studies that have assessed the mouse liver effects of PCE and its major oxidative metabolite trichloroacetic acid (TCA). Similar to PCE, TCA has also been demonstrated to liver tumors selectively in mice following chronic exposure. The Key Events (KE) of the proposed PCE MOA involve oxidative metabolism of PCE to TCA [KE 1]; activation of the peroxisome proliferator-activated receptor alpha (PPARα) [KE 2]; alteration in hepatic gene expression including cell growth pathways [KE 3]; increase in cell proliferation [KE 4]; selective clonal expansion of hepatic preneoplastic foci [KE 5]; and formation of hepatic neoplasms [KE 6]. The scientific evidence supporting the PPARα MOA for PCE is strong and satisfies the requirements for a MOA analysis. The PPARα liver tumor MOA in rodents has been demonstrated not to occur in humans; thus, human liver cancer risk to PCE is not likely.

Effect of inactivating heterozygous mutations in DNA repair genes on experimental lung carcinogenesis in mice

Background. Inactivating mutations in Chek2 and Gprc5a genes are known to be associated with cancer development. Experimental carcinogenesis studies in genetically modified mice generate new data on their influence on pathology development.Aim. In the present study in a model of lung carcinogenesis, survival parameters as well as tumor multiplicity and size in mice with Chek2 and Gprc5a heterozygous inactivating mutations were evaluated.Material and methods. F2 hybrid mice from crosses between CBAB6F1 males heterozygous for the studied mutations and wild-type BALB / c females were used: Chek2dAA-carriers (76 males and 64 females) and Gprc5ainsA-carriers (60 males and 42 females). Starting at four months of age, mice received urethane (ethyl carbamate) intraperitoneally at a dose of 600 mg / kg weekly for 6 weeks. After genotyping by allele-specific PCR, animals were allocated to groups. Carcinogenesis parameters were evaluated 40 weeks after the beginning of the experiment.Results. The proportion of mice with mutations surviving to the age of three months roughly followed the Mendelian distribution (35 / 41 males and 33 / 31 females) for the offspring of males heterozygous for Chek2dAA and was significantly lower in the case of Gprc5ainsA (20 / 40 males and 17 / 25 females, p = 0.043). The death of Gprc5ainsA carriers during the experiment was also higher than in the control group (p = 0.0506 in females). Synchronous lung and thymus neoplasms were found in 2 out of 4 Gprc5ainsA females that died before the end of the experiment, which were not found in other groups. At the end of the experiment, no significant differences in tumor multiplicity, mean linear size, and volume were found between the groups of mice with and without mutations.Conclusion. It was found that heterozygous inactivating mutation Chek2dAA does not affect early age development and does not modify the parameters of induced lung carcinogenesis in mice. Heterozygous carriage of Gprc5ainsA mutation in mice increases the risk of early death and sensitivity to the toxic and carcinogenic effects of urethane.

Abstract 3038: PPARδ promotes gastric adenocarcinoma carcinogenesis by enhancing ZBP1-induced necroptosis and inflammation and fostering gastric tumor immunosuppressive microenvironment

Abstract Overexpression of peroxisome proliferator-activated receptor delta (PPARδ) in villin-positive gastric progenitor cells (VGPCs) has been demonstrated to spontaneously induce gastric adenocarcinoma (GAC) in mice. PPARδ is upregulated in human GAC tissues, and its expression level is positively associated with human GAC grades and stages. However, the potential for targeting this tumorigenic effect of PPARδ, along with the underlying molecular mechanisms, remains elusive. In this study, we investigated the impact of PPARδ genetic deletion/loss of function in VGPCs on Helicobacter felis (H. felis)-induced GAC carcinogenesis by orally exposing the mice carrying wild-type (WT) PPARδ or genetically deleted PPARδ in VGPCs to H. felis infection. We also examined the effect of PPARδ inhibition by its specific antagonist, GSK3787, on PPARδ-induced GAC carcinogenesis in mice. Furthermore, we conducted RNA-seq for transcriptomic profiling analysis and validated the results through quantitative reverse transcription-polymerase chain reaction (qRT-PCR) on the gastric tissues of those mice and the mice with WT PPARδ or PPARδ overexpression in VGPCs at ages 10, 25 and 55 weeks. We found that 1) PPARδ genetic deletion in VGPCs significantly inhibited H. felis-induced gastric chronic inflammation and GAC carcinogenesis; GSK3787 significantly suppressed PPARδ-induced GAC carcinogenesis in mice; 2) PPARδ overexpression in VGPCs significantly enhanced while PPARδ genetic deletion in VGPCs significantly inhibited ZBP1-induced necroptosis and the related inflammatory signaling pathways such as interferon-gamma signaling, IL6-JAK-STAT3 signaling, and IL2-STAT5 signaling evidenced by gene set enrichment analysis of RNA-seq data, which were further validated by qRT-PCR; and 3) PPARδ overexpression in VGPCs markedly increased the expression of CCL20 in gastric epithelial cells of the mice before (age 10 weeks) and after GAC initiation and progression (age 25 weeks-early stage GAC and age 55 weeks-late stage GAC), and gastric infiltrations of CCR6+ myeloid-derived suppressor cells and CCR6+ tumor-associated macrophages, thereby creating a gastric tumor immunosuppressive microenvironment. In contrast, PPARδ genetic deletion in VGPCs or the administration of GSK3787 diet significantly reversed this PPARδ-induced immune suppression in mice. In conclusion, our findings highlight the pivotal role of PPARδ in VGPCs as a key host factor in GAC carcinogenesis, especially in individuals exposed to chronic H. pylori infection that could lead to an upregulation of PPARδ expression. This suggests that targeting PPARδ signaling could be a novel therapeutic approach for GAC prevention and therapy. Citation Format: Yi Liu, Daoyan Wei, Lily Wang, James C. Yao, Imad Shureiqi, Xiangsheng Zuo. PPARδ promotes gastric adenocarcinoma carcinogenesis by enhancing ZBP1-induced necroptosis and inflammation and fostering gastric tumor immunosuppressive microenvironment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3038.