Different Stages Of Colorectal Cancer Research Articles

Unravelling the molecular mechanistic pathway underlying the anticancer effects of kaempferol in colorectal cancer: a reverse pharmacology network approach.

Colorectal cancer (CRC) is the third most diagnosed and highly fatal malignancy, presenting serious health concerns worldwide. The search for an effective cure for CRC is challenging and poses a serious concern. Kaempferol is a potent anti-cancerous bioactive compound often suggested for treating various cancers, including CRC. However, its underlying molecular mechanism against CRC remains unclear. The present study delves into kaempferol's molecular pathways and underlying molecular mechanisms against CRC targets. The target protein-coding genes for kaempferol were retrieved, and the CRC-associated genes were curated. Twelve common targets with a disease specificity index of > 0.6 were validated for their protein expression at different stages of CRC. Over-expressed USP1, SETD7, POLH, TDP1 and RACGAP1 were selected for further studies. The binding affinities of kaempferol to the corresponding proteins were evaluated using molecular docking and Molecular Dynamics (MD) simulations. SETD7 exhibited the highest binding affinity with the lowest binding energy (-8.06kcal/mol). Additionally, the MD simulation, and MM-PBSA conferred SETD7-kaempferol complex had the least root-mean-square deviation with lower interaction energy and higher conformational stability. The protein-protein interaction of SETD7 constructed revealed direct interactors, namely, DNMT1, FOXO1, FOXO3, FOXO4, H3-3B, H3-4, H3C12, H3C13, SETD7, SIRT1 and TP53, have a potential role in cancer progression through FOXO signalling. In summary, our study revealed kaempferol's multi-target and synergistic effect on multiple CRC targets and its underlying mechanisms. Finally, the study recommends in-vitro and in-vivo trials for validation of anti-cancerous drugs for CRC.

Early administration of Wumei Wan inhibit myeloid-derived suppressor cells via PI3K/Akt pathway and amino acids metabolism to prevent colitis-associated colorectal cancer

Ethnopharmacological relevanceWumei Wan (WMW), a traditional Chinese medicine prescription, has been proved to be effective in treating Colitis-associated colorectal cancer (CAC), but it has not been proven to be effective in different stages of CAC. Aim of the studyThe purpose of our study is to investigate the therapeutic effect and mechanism of WMW on the progression of CAC. Materials and methodsAzioximethane (AOM) and dextran sulfate sodium (DSS) were used to treat mice for the purpose of establishing CAC models. WMW was administered in different stages of CAC. The presentative chemical components in WMW were confirmed by LC-MS/MS under the optimized conditions. The detection of inflammatory cytokines in the serum and colon of mice were estimated by qRT-PCR and ELISA. The changes of T cells and myeloid-derived suppressor cells (MDSCs) in each group were detected by flow cytometry. The metabolic components in serum of mice were detected by UPLC-MS/MS. Expression of genes and proteins were detected by eukaryotic transcriptomics and Western blot to explore the key pathway of WMW in preventing CAC. ResultsWMW had significant effect on inhibiting inflammatory responses and tumors during the early development stage of CAC when compared to other times. WMW increased the length of mice's colons, reduced the level of IL-1β, IL-6, TNF-α in colon tissues, and effectively alleviated colonic inflammation, and improved the pathological damage of colon tissues. WMW could significantly reduce the infiltration of MDSCs in the spleen, increase CD4+ T cells and CD8+ T cells in the spleen of CAC mice, and effectively reform the immune microenvironment in CAC mice.Transcriptomics analysis revealed that 2204 genes had different patterns of overlap in the colon tissues of mice between control group, AOM + DSS group, and early administration of WMW group. And KEGG enrichment analysis showed that PI3K/Akt signaling pathway, ECM-receptor interaction, IL-17 signaling pathway, MAPK signaling pathway, pancreatic secretion, thermogenesis, and Rap1 signaling pathway were all involved.The serum metabolomics results of WMW showed that the metabolic compositions of the control group, AOM + DSS group and the early stage of WMW were different, and 42 differential metabolites with the opposite trends of changes were screened. The metabolic pathways mainly included pyrimidine metabolism, glycine, serine and threonine metabolism, tryptophan metabolism, and purine metabolism. And amino acids and related metabolites may play an important role in WMW prevention of CAC. ConclusionWMW can effectively prevent the occurrence and development of CAC, especially in the initial stage. WMW can reduce the immune infiltration of MDSCs in the early stage. Early intervention of WMW can improve the metabolic disorder caused by AOM + DSS, especially correct the amino acid metabolism. PI3K/Akt signaling pathway was inhabited in early administration of WMW, which can regulate the amplification and function of MDSCs.

Abstract 2299: More information from limited DNA: simultaneous measurement of genetics, 5hmC and 5mC in cell-free DNA

Abstract Liquid biopsy for profiling of cell free DNA (cfDNA) in blood holds huge promise to transform how we experience and manage cancer by early detection and identification of residual disease and subtype. While early work in liquid biopsy focused on the identification of actionable somatic variations at specific loci, the past decade has seen an expansion into non-genetic features, notably methylation. 5-methylcytosine (5-mC) profiles of cancer are differential from non-cancer at many more loci and so provide a stronger signal. Moreover, recent research has suggested that 5-hydroxymethylcytosine (5-hmC) profiles in cfDNA can be a marker for early cancer. However, a standard blood draw yields an average of only 10ng of cfDNA, presenting the dilemma of how to use limited sample to obtain maximum information. Existing methods for measuring methylation using next-generation sequencing cannot distinguish between 5-mC and 5-hmC and are limited in their ability to detect mutations, with approaches that measure both 5-mC and 5-hmC requiring two separate workflows. Hence, to measure complete genetics, 5-mC and 5-hmC using existing technologies requires three separate workflows, each of which require separate DNA input. We will present a technology which sequences at single base resolution the complete genetic sequence of input DNA fragments integrated with the modification status (unmodified, 5-mC or 5-hmC) for each CpG from low nanogram input quantities of DNA. A hairpin construct is used to create a copy of the original DNA strand. Enzymatic conversion followed by next-generation sequencing enables coupled decoding of bases across the original and copy strand, uniquely reporting one of A, C, G, T, 5-mC, or 5-hmC for each position in the input DNA fragment. Using this technology, we generated whole genome 6-letter data (measuring A, C, G, T, 5-mC, and 5-hmC at single-base resolution across the genome) on cell-free DNA extracted from plasma of healthy volunteers and patients with colorectal cancer at various stages of progression. We demonstrate how the technology can be used to compare 5-mC and 5-hmC methylomic profiles, genomics, and fragmentomics, across different stages of colorectal cancer. In particular, we show how disease progression can be marked by changes in 5-mC and 5-hmC across several loci. We propose that the ability to measure 5-mC and 5-hmC at high accuracy and single base resolution, alongside genomic and fragmentomic profiles, from a limited quantity of DNA will enable greater insight into the ctDNA in plasma and help advance the field of liquid biopsy towards fulfilling its promise. Citation Format: Tom J. Charlesworth, Fabio Puddu, Robert Crawford, Annelie Johansson, Ermira Lleshi, Aurelie Modat, Jamie Scotcher, Michael Wilson, Nicholas Harding, Jean Teyssandier, Jens Fullgrabe, Walraj Gosal, Paidi Creed. More information from limited DNA: simultaneous measurement of genetics, 5hmC and 5mC in cell-free DNA [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 2299.

Progressive Changes of Colitis Into Colorectal Cancer Associated With Changes in Neutrophil Lymphocyte Ratio, Serum, Micrornas and Infiltrate Neutrophils

Introduction: Understand the progression of colorectal cancer from the beginning until the advance stages is difficult and challenging. However, this could be overcome with a good animal model. Methods: In this study, a modified approach had been used to develop colorectal cancer model. The model was developed and monitored from colitis formation until the late stage of colorectal cancer. The changes of neutrophil lymphocyte ratio (NLR), serum microRNAs and infiltrate neutrophil in different stages of colorectal cancer were assessed in this study. Results: Results showed that the progression of the disease is correlated with NLR as early as the formation of colitis (r=0.121, p<0.026). Meanwhile, the size of the tumor at each stage is also associated with the NLR value (r=0.185, p<0.0012). In the serum microRNAs study, it was found microRNAs expression in blood serum change in different stages of colorectal cancer. In the early stage of colitis formation, miR223 (> 3 fold expression, p < 0.0025) were abundantly found in the blood serum. Meanwhile in others stage mild (miRNA345 > 2.5 fold, p<0.0011), moderate (miRNA347 & miR512 > 3 fold, p<0.002) and severe (miR31 & miR145 > 2 fold, p<0.0001) microRNAs were also found expressed differently. The quantities of infiltrate neutrophil were varied in different stages of the disease. Conclusion: This study provides an insight into the immunity and molecular level of colorectal cancer and it allows a progressive monitoring on the changes in the molecular, cellular and histological level.

Evaluation of BMP-2 as a differentiating and radiosensitizing agent for colorectal cancer stem cells.

Despite effective clinical responses, a large proportion of patients undergo resistance to radiotherapy. The low response rate to current treatments in different stages of colorectal cancer depends on the prominent role of stem cells in cancer. In the present study, the role of BMP-2 as an ionizing radiation-sensitive factor in colorectal cancer cells was investigated. A sphere formation assay was used for the enrichment of HCT-116 cancer stem cells (CSCs). The effects of combination therapy (BMP-2+ radiation) on DNA damage response (DDR), proliferation, and apoptosis were evaluated in HCT-116 and CSCs. Gene expressions of CSCs and epithelial-mesenchymal transition (EMT) markers were also evaluated. We found that the sphere formation assay showed a significant increase in the percentage of CSCs. Moreover, expression of CSCs markers, EMT-related genes, and DNA repair proteins significantly decreased in HCT-116 cells compared to the CSCs group after radiation. In addition, BMP-2 promoted the radiosensitivity of HCT-116 cells by decreasing the survival rate of the treated cells at 2, 4, and 6 Gy compared to the control group in HCT-116 cells. Our findings indicated that BMP-2 could affect numerous signaling pathways involved in radioresistance. Therefore, BMP-2 ‎ can be considered an appealing therapeutic target for the treatment of radioresistant human colorectal cancer.

YTE-17 inhibits colonic carcinogenesis by resetting antitumor immune response via Wnt5a/JNK mediated metabolic signaling

The density and composition of lymphocytes infiltrating colon tumors serve as predictive factors for the clinical outcome of colon cancer. Our previous studies highlighted the potent anti-cancer properties of the principal compounds found in Garcinia yunnanensis (YTE-17), attributing these effects to the regulation of multiple signaling pathways. However, knowledge regarding the mechanism and effect of YTE-17 in the prevention of colorectal cancer is limited. In this study, we conducted isobaric tags for relative and absolute quantification (iTRAQ) analysis on intestinal epithelial cells (IECs) exposed YTE-17, both in vitro and invivo, revealing a significant inhibition of the Wnt family member 5a (Wnt5a)/c-Jun N-terminal kinase (JNK) signaling pathway. Subsequently, we elucidated the influence and mechanism of YTE-17 on the tumor microenvironment (TME), specifically focusing on macrophage-mediated T helper 17 (Th17) cell induction in a colitis-associated cancer (CAC) model with Wnt5a deletion. Additionally, we performed the single-cell RNA sequencing (scRNA-seq) on the colonic tissue from the Wnt5a-deleted CAC model to characterize the composition, lineage, and functional status of immune mesenchymal cells during different stages of colorectal cancer (CRC) progression. Remarkably, our findings demonstrate a significant reduction in M2 macrophage polarization and Th17 cell phenotype upon treatment with YTE-17, leading to the restoration of regulatory T (Treg)/Th17 cell balance in azoxymethane (AOM)/dextran sodium sulfate (DSS) model. Furthermore, we also confirmed that YTE-17 effectively inhibited the glycolysis of Th17 cells in both direct and indirect co-culture systems with M2 macrophages. Notably, our study shed light on potential mechanisms linking the non-canonical Wnt5a/JNK signaling pathway and well-established canonical β-catenin oncogenic pathway in vivo. Specifically, we proposed that Wnt5a/JNK signaling activity in IECs promotes the development of cancer stem cells with β-catenin activity within the TME, involving macrophages and T cells. In summary, our study undergoes the potential of YTE-17 as a preventive strategy against CRC development by addressing the imbalance with the immune microenvironment, thereby mitigating the risk of malignancies.

Features of the immunophenotype of cancer stem cells in different stages of colorectal cancer

ABSTRACT. This study examines the immunophenotype characteristics of the stem cell population in colon adenocarcinoma at various stages of the disease and evaluates their impact on the prevalence of the oncological process. The research involved analyzing surgical specimens from 67 patients with histologically confirmed colon adenocarcinoma using microscopic and immunohistochemical techniques. The patients were divided into two groups based on the extent of the neoplastic process. The first group consisted of 32 patients with localized colorectal cancer (T34aN0M0), while the second group included 35 patients with colorectal cancer with synchronous monoorgan liver metastases (T34aN02M1a (hep)).The study assessed the immunophenotype of the tumor and investigated its correlation with the disease stage. The analysis revealed no significant correlation between the proliferative activity index of the immunohistochemical marker Ki67 and the stage of the disease (p = 0.108). However, when comparing the immunophenotype of stem tumor cells in the group of patients with metastatic colorectal cancer, statistically significantly higher levels of expression were observed for the following immunohistochemical markers: ALDH1+ (p = 0.028), ALDH1+Ki67+ (p = 0.0002), and CXCR4+ (p = 0.005). These results indicate a strong association between immunohistochemical markers characterizing the stem cell population and the prevalence of the tumor. Therefore, markers such as ALDH1+, ALDH1+Ki67+, and CXCR4+ can provide valuable insights into the molecular and biological properties of stem tumor cells, aiding in the clinical assessment of the potential aggressiveness of colon adenocarcinoma. The study also found that the median stem cell proliferation index, which considers the proportion of proliferating cells only within the stem cell population and not the entire cell pool, in the group of patients with localized colorectal cancer was 16%. Notably, none of the patients in stage II of the disease had a stem cell proliferation level below 50%, and the stem cell proliferation index ranged from 0% to 50%. Notably, none of the patients in stage II of the disease had a stem cell proliferation level below 50%, and the stem cell proliferation index varied from 0% to 50%. In contrast, the median index of stem cell proliferation in the group of patients with metastatic colorectal cancer was 45.5%, with values ranging from 7% to 100%. The stem cell proliferation index in this group was significantly higher (p = 0.00001) than that in the group with localized tumor processes. Considering the small population of stem tumor cells, determining the index of stem cell proliferation is advisable for a more accurate assessment of their proliferative activity.

Combined detection of SDC2/ADHFE1/PPP2R5C methylation in stool DNA for colorectal cancer screening.

Colorectal cancer (CRC) is a disease of global concern, and its increasing incidence suggests the need for early and accurate diagnosis. The aim of this study was to investigate the value of combined detection of SDC2, ADHFE1 and PPP2R5C gene methylation in stool samples for early CRC screening. Stool samples from patients with CRC (n = 105), advanced adenoma (AA) (n = 54), non-advanced adenoma (NA) (n = 57), hyperplastic or other polyps (HOP) (n = 47) or no evidence of disease (NED) (n = 100) were collected from September 2021 to September 2022. The methylation levels of SDC2, ADHFE1 and PPP2R5C were quantified by quantitative methylation-specific polymerase chain reaction (qMSP), and faecal immunochemical testing (FIT) was performed. The diagnostic value was assessed using reporter operating characteristic (ROC) curve analysis. The sensitivity of combined detection of SDC2/ADHFE1/PPP2R5C methylation in predicting CRC (0-IV) was 84.8%, the specificity was 98.0%, and the AUC was 0.930 (95% CI 0.889-0.970). Compared to FIT and serum tumour biomarkers, it showed better diagnostic performance for different stages of CRC. The results of this study verified that the methylation levels of SDC2, ADHFE1 and PPP2R5C in stool DNA were significantly increased in CRC patients. Combined detection of SDC2/ADHFE1/PPP2R5C methylation is a potential non-invasive diagnostic method for CRC and precancerous lesion screening. Chinese Clinical Trials Registry, ChiCTR2100046662, registered on 26 May 2021, prospective registration.

Abdominal fat and muscle distributions in different stages of colorectal cancer

BackgroundThe purpose of this study is to explore the difference of abdominal fat and muscle composition, especially subcutaneous and visceral adipose tissue, in different stages of colorectal cancer (CRC).Materials and methodsPatients were divided into 4 groups: healthy controls (patients without colorectal polyp), polyp group (patients with colorectal polyp), cancer group (CRC patients without cachexia), and cachexia group (CRC patients with cachexia). Skeletal muscle (SM), subcutaneous adipose tissue (SAT), visceral adipose tissue (VAT), and intermuscular adipose tissue (IMAT) were assessed at the third lumbar level on computed tomography images obtained within 30 days before colonoscopy or surgery. One-way ANOVA and linear regression were used to analyze the difference of abdominal fat and muscle composition in different stages of CRC.ResultsA total of 1513 patients were divided into healthy controls, polyp group, cancer group, and cachexia group, respectively. In the development of CRC from normal mucosa to polyp and cancer, the VAT area of the polyp group was significantly higher than that of the healthy controls both in male (156.32 ± 69.71 cm2 vs. 141.97 ± 79.40 cm2, P = 0.014) and female patients (108.69 ± 53.95 cm2 vs. 96.28 ± 46.70 cm2, P = 0.044). However, no significant differences were observed of SAT area between polyp group and healthy controls in both sexes. SAT area decreased significantly in the male cancer group compared with the polyp group (111.16 ± 46.98 cm2 vs. 126.40 ± 43.52 cm2, P = 0.001), while no such change was observed in female patients. When compared with healthy controls, the SM, IMAT, SAT, and VAT areas of cachexia group was significantly decreased by 9.25 cm2 (95% CI: 5.39–13.11 cm2, P < 0.001), 1.93 cm2 (95% CI: 0.54–3.32 cm2, P = 0.001), 28.84 cm2 (95% CI: 17.84–39.83 cm2, P < 0.001), and 31.31 cm2 (95% CI: 18.12–44.51 cm2, P < 0.001) after adjusting for age and gender.ConclusionAbdominal fat and muscle composition, especially SAT and VAT, was differently distributed in different stages of CRC. It is necessary to pay attention to the different roles of subcutaneous and visceral adipose tissue in the development of CRC.

Increased ENT2 expression and its association with altered purine metabolism in cell lines derived from different stages of colorectal cancer.

Colorectal cancer (CRC) is one of the most prevalent malignant cancer types worldwide. Although the purine metabolism pathway is vital for cancer cell survival, little is known about the role of equilibrative nucleoside transporter 2 (ENT2) in CRC development and its association with purine metabolites. The aim of the present study was to evaluate the levels of hypoxanthine phosphoribosyl transferase (HPRT), hypoxanthine and uric acid (UA), as well as xanthine oxidase (XO) activity, and investigate their association with ENT2 expression levels in a normal human colon cell line and CRC cell lines derived from different stages of CRC. These analyses were performed using the normal colon CCD-841CoN cell line and a panel of human CRC cell lines comprising SW480, HCT15 and HCT116, which represent Dukes' B, C and D stages, respectively. Reverse transcription-quantitative PCR was performed to determine the level of ENT2 mRNA expression. In cells of all CRC stages, the levels of HPRT and hypoxanthine were significantly higher (P<0.05), while XO activity and UA levels were significantly decreased (P<0.05), compared with those in the CCD-841CoN cell line. ENT2 expression was found to be elevated in cells derived from all stages of CRC. The Dukes' D stage cell line had higher levels of HPRT and hypoxanthine, although its ENT2 level was not significantly lower than that of the Dukes' B and C stage cell lines. Increased levels of HPRT and hypoxanthine in various stages of CRC may indicate an increase in the activity of the salvage pathway. The increased expression of ENT2 implies the importance of the ENT2 protein in facilitating hypoxanthine transport, which is required for enhanced DNA synthesis via hypoxanthine recycling. In conclusion, ENT2 may have potential as a target in the development of CRC therapeutics.

Exploring Core Genes by Comparative Transcriptomics Analysis for Early Diagnosis, Prognosis, and Therapies of Colorectal Cancer.

Colorectal cancer (CRC) is one of the most common cancers with a high mortality rate. Early diagnosis and therapies for CRC may reduce the mortality rate. However, so far, no researchers have yet investigated core genes (CGs) rigorously for early diagnosis, prognosis, and therapies of CRC. Therefore, an attempt was made in this study to explore CRC-related CGs for early diagnosis, prognosis, and therapies. At first, we identified 252 common differentially expressed genes (cDEGs) between CRC and control samples based on three gene-expression datasets. Then, we identified ten cDEGs (AURKA, TOP2A, CDK1, PTTG1, CDKN3, CDC20, MAD2L1, CKS2, MELK, and TPX2) as the CGs, highlighting their mechanisms in CRC progression. The enrichment analysis of CGs with GO terms and KEGG pathways revealed some crucial biological processes, molecular functions, and signaling pathways that are associated with CRC progression. The survival probability curves and box-plot analyses with the expressions of CGs in different stages of CRC indicated their strong prognostic performance from the earlier stage of the disease. Then, we detected CGs-guided seven candidate drugs (Manzamine A, Cardidigin, Staurosporine, Sitosterol, Benzo[a]pyrene, Nocardiopsis sp., and Riccardin D) by molecular docking. Finally, the binding stability of four top-ranked complexes (TPX2 vs. Manzamine A, CDC20 vs. Cardidigin, MELK vs. Staurosporine, and CDK1 vs. Riccardin D) was investigated by using 100 ns molecular dynamics simulation studies, and their stable performance was observed. Therefore, the output of this study may play a vital role in developing a proper treatment plan at the earlier stages of CRC.

Novel DNA methylation biomarkers in stool and blood for early detection of colorectal cancer and precancerous lesions

BackgroundEarly detection and prevention of precancerous lesions can significantly reduce the morbidity and mortality of colorectal cancer (CRC). Here, we developed new candidate CpG site biomarkers for CRC and evaluated the diagnostic value of their expression in blood and stool samples of CRC and precancerous lesions.MethodsWe analyzed 76 pairs of CRC and adjacent normal tissue samples, 348 stool samples, and 136 blood samples. Candidate biomarkers for CRC were screened using a bioinformatics database and identified using a quantitative methylation-specific PCR method. The methylation levels of the candidate biomarkers were validated using blood and stool samples. The divided stool samples were used to construct and validate a combined diagnostic model and to analyze the independent or combined diagnostic value of candidate biomarkers in stool samples of CRC and precancerous lesions.ResultsTwo candidate CpG site biomarkers for CRC, cg13096260 and cg12993163, were identified. Although both biomarkers demonstrated diagnostic performance to a certain extent when using blood samples, they showed better diagnostic value for different stages of CRC and AA with stool samples.Conclusionscg13096260 and cg12993163 detection in stool samples could be a promising approach for screening and early diagnosis of CRC and precancerous lesions.

Isolation and proteomic profiling of urinary exosomes from patients with colorectal cancer

Exosomes in the body fluid are effective cell-derived membranous structures transferring various molecules to mediate intercellular communication. The expression of protein in the urinary exosomes from the colorectal cancer (CRC) patients could reflect the characteristics of tumorigenesis. The urinary exosomes with globular membrane structure, the size of 30 ~ 100 nm and positive expression of CD9, CD63 and CD81 were successfully isolated from 9 CRC patients and 3 heathy adults using the density gradient ultracentrifugation. Proteome profiles revealed by label-free liquid chromatography-tandem mass spectrometry (LC–MS/MS) indicated that several proteins were differentially expressed among different stages of CRC. Compared with normal controls, 67 proteins in CRC urinary exosomes were upregulated and 74 proteins were downregulated. The bioinformatics analysis revealed the decreased proteins were related to ESCRT III complex disassembly. The CHMP family was further determined to be the hub of interaction network of proteins enriched in ESCRT signaling. The significant decrease of CHMP4A, CHMP4B, CHMP2A, CHMP2B and CHMP1B were respectively found in the total CRC group and distant metastasis group compared with NC group. Moreover, the CEACAM family also showed significant aberrant changes in the urinary exosomes of CRC patients. The CEACAM7 and CEACAM1 were increased in the CRC patients compared with healthy individuals (P < 0.05). Significant changes of proteomic profile could be found in the urinary exosomes in the CRC patients. The differential expressed urinary exosomes derived proteins showed potential usage in diagnosis and prognosis of CRC.

Use of glycoproteome profiles to detect advanced adenomas and colorectal cancer.

69 Background: Colorectal cancer (CRC) remains a leading cancer despite current screening modalities. Precancerous lesions, or Advanced Adenomas (AA), commonly precede invasive cancer development by years. Newer technologies use circulating tumor DNA and/or proteins for CRC detection but have not been able to effectively detect AA. Aberrant protein glycosylation is associated with (pre-)malignant lesions. To detect glycoproteome profiles associated with the occurrence of AA, we studied serum glycoproteins in AA/CRC. Methods: A novel platform combining liquid-chromatography/mass-spectrometry (LC-MS) and artificial-intelligence (AI)-powered data processing allowing high resolution, high throughput glycoproteomic profiling was used to identify glycoprotein biomarkers in peripheral blood. Samples were sourced from biorepositories and included patients diagnosed with CRC, AA, ulcerative colitis (UC) and controls. The samples were split into a training (50%) and a hold-out testing set (50%) for the development of a machine learning (ML)-based multivariable predictive model. Statistical analysis was performed on normalized data to identify biomarkers differentiating AAs and different stages of CRC from controls. Results: We studied 563 patient samples: 196 controls (mean age 51.7; 52% female); 32 AA (mean age 68.6; 53% female); 247 CRC (mean age 65.6; 50% female) and 88 UC (mean age 44.1; 47% female). There were 250 differentially abundant (FDR &lt; 0.05) glycopeptides/peptides when comparing CRC and AA samples with healthy and UC controls. A subset was assessed, generating a six (6) biomarker ML classification model. This model was applied to the hold-out test and achieved an overall sensitivity of 91.4% and specificity of 91.8% for predicting AA/CRC versus healthy/UC with an area under the receiver operating characteristic of 0.962. AA and CRC separately were predicted with a sensitivity of 84.4% and 92.8%, respectively, relative to healthy/UC with sensitivities for CRC stage 1/2 and stage 3/4 being 91.2% and 93.2%, respectively). Conclusions: Glycoproteomic serum profiles accurately detect precancerous AA in addition to CRC and offer a new approach to effective CRC screening. We will have completed an interim analysis of a large prospective observational study at the time of the meeting. Clinical trial information: NCT05445570 . [Table: see text]

Metabolomics analysis of amino acid and fatty acids in colorectal cancer patients based on tandem mass spectrometry.

Metabolic differences between colorectal cancer (CRC) and NI (NI) play an important role in early diagnoses and in-time treatments. We investigated the metabolic alterations between CRC patients and NI, and identified some potential biomarkers, and these biomarkers might be used as indicators for diagnosis of CRC. In this study, there were 79 NI, 50 CRC I patients, 52 CRC II patients, 56 CRC III patients, and 52 CRC IV patients. MS-MS was used to measure the metabolic alterations. Univariate and multivariate data analysis and metabolic pathway analysis were applied to analyze metabolic data and determine differential metabolites. These indicators revealed that amino acid and fatty acids could separate these groups. Several metabolites indicated an excellent variables capability in the separation of CRC patients and NI. Ornithine, arginine, octadecanoyl carnitine, palmitoyl carnitine, adipoyl carnitine, and butyryl carnitine/propanoyl carnitine were selected to distinguish the CRC patients and NI. And methionine and propanoyl carnitine, were directly linked to different stages of CRC. Receiver operating characteristics curves and variables importance in projection both represented an excellent performance of these metabolites. In conclusion, we assessed the difference between CRC patients and NI, which supports guidelines for an early diagnosis and effective treatment.

Myeloid-derived suppressor-like cells - does their frequency change in patients with different stages of CRC?

Background/Aim. Colorectal cancer (CRC) is one of the most common cancers in the population, often leading to lethal outcomes. Myeloid-derived suppressor cells (MDSCs) belong to a heterogeneous group of immature cells thought to have an immunosuppressive effect that may aid in tumor development and spreading. The aim of this study was to analyze the frequency and significance of MDSC-like cells at different stages in patients with CRC. Methods. Peripheral blood (PB) samples of 83 patients at different stages of the disease and 12 healthy subjects (control group) were analyzed. MDSC-like cells were identified and enumerated in the PB samples of the participants based on the immuno-phenotypic characteristics of the cells. Results. A statistically significant increase in the absolute and relative number of polymorphonuclear (PMN) MDSC (PMN-MDSC)-like cells was observed in the PB of all the patients with CRC, com-pared to the healthy control group (p &lt; 0.0001). No significant increase was observed in monocytic MDSC (M-MDSC)-like cells when they were analyzed without CRC stage stratification (p &gt; 0.05). When the relative and absolute numbers of PMN-MDSC-like cells were analyzed in relation to the stages of CRC disease (TNM classification), a statistically significant difference was observed between the control group and patients in stages III and IV of the dis-ease (p = 0.0005 vs. p = 0.0003 and p &lt; 0.0001 vs. p &lt; 0.0001, respectively). There was, as well, a significant difference when the numbers of PMN-MDSC-like cells in patients in stages I and II were compared to numbers in patients in stage IV of the CRC (p = 0.0161 vs. p &lt; 0.0001 and p = 0.0065 vs. p &lt; 0.0001, respectively). A statistically significant difference in the relative and absolute number of M-MDSC-like cells was observed only between patients in stages II and IV of the disease (p = 0.0014 and p = 0.0002, respectively). The highest number of MDSC-like cells was observed in stage IV of the disease according to the TNM classification. A positive correlation between the presence of these cells and the number of organs affected by metastatic changes was observed (p &lt; 0.0001 for the relative and absolute number of PMN-MDSC-like cells and p = 0.003 and p = 0.0004 for the relative and absolute number of M-MDSC-like cells). Conclusion. CRC patients had a statistically significant increase in PMN-MDSC-like cells compared to healthy controls. The increase in absolute and relative numbers of these cells mostly follows the growth and progression of CRC, while a statistically significant difference in the number of M-MDSC-like cells is observed only between stages II and IV of the disease. The absolute and relative numbers of both subtypes of MDSC-like cells significantly correlate with the number of organs affected by CRC metastases.

Rewiring glucose metabolism improves 5-FU efficacy in p53-deficient/KRASG12D glycolytic colorectal tumors

Despite the fact that 5-fluorouracil (5-FU) is the backbone for chemotherapy in colorectal cancer (CRC), the response rates in patients is limited to 50%. The mechanisms underlying 5-FU toxicity are debated, limiting the development of strategies to improve its efficacy. How fundamental aspects of cancer, such as driver mutations and phenotypic heterogeneity, relate to the 5-FU response remains obscure. This largely relies on the limited number of studies performed in pre-clinical models able to recapitulate the key features of CRC. Here, we analyzed the 5-FU response in patient-derived organoids that reproduce the different stages of CRC. We find that 5-FU induces pyrimidine imbalance, which leads to DNA damage and cell death in the actively proliferating cancer cells deficient in p53. Importantly, p53-deficiency leads to cell death due to impaired cell cycle arrest. Moreover, we find that targeting the Warburg effect in KRASG12D glycolytic tumor organoids enhances 5-FU toxicity by further altering the nucleotide pool and, importantly, without affecting non-transformed WT cells. Thus, p53 emerges as an important factor in determining the 5-FU response, and targeting cancer metabolism in combination with replication stress-inducing chemotherapies emerges as a promising strategy for CRC treatment.

Regulation of transforming growth factor-β signaling as a therapeutic approach to treating colorectal cancer.

According to data from 2020, Slovakia has long been among the top five countries with the highest incidence rate of colorectal cancer (CRC) worldwide, and the rate is continuing to rise every year. In approximately 80% of CRC cases, allelic loss (loss of heterozygosity, LOH) occurs in the long arm of chromosome 18q. The most important genes that can be silenced by 18q LOH or mutations are small mothers against decapentaplegic homolog (SMAD) 2 and SMAD4, which are intracellular mediators of transforming growth factor (TGF)-β superfamily signals. TGF-β plays an important role in the pro-oncogenic processes, including such properties as invasion, epithelial-mesenchymal transition (commonly known as EMT), promotion of angiogenesis, and immunomodulatory effects. Several recent studies have reported that activation of TGF-β signaling is related to drug resistance in CRC. Because the mechanisms of drug resistance are different between patients in different stages of CRC, personalized treatment is more effective. Therefore, knowledge of the activation and inhibition of factors that affect the TGF-β signaling pathway is very important.

Abstract 3062: Modifiable factors and gut microbiome alteration among colorectal cancer patients

Abstract Background: Previous study have indicated an association of microbial dysbiosis with stages of colorectal cancer (CRC). Meanwhile, higher microbial stochasticity has been introduced as the Anna Karenina principle (AKP) effect, which originates from the opening line of Leo Tolstoy and is translated in the microbiome as “all healthy microbiomes are similar; each dysbiotic microbiome is dysbiotic in its own way”. Here, we aim to elucidate the effect of modifiable factors on microbiome stochasticity related to dysbiosis in CRC. Methods: Fecal samples from 331 CRC patients, were collected prior to their surgery at Seoul National University Hospital. Bacteria that were enriched in early- and late-stage CRC groups were identified by linear discrimination analysis effect size (LEfSE) method, which was then used to calculate microbial dysbiosis index (MDI). The association of dietary choices with microbiome variability was addressed using the LEfSE analysis. The difference of intra-sample similarity index among lifestyle factors and metabolic diseases was tested to examine the AKP effect. A tree-based analysis of food choices was constructed. We then applied Procrustes framework to analyze the shape of dietary diversity and microbiome. Results: The LEfSE identified Leuconostoc, Oxalobacter, Acidaminococcus, and Methanobrevibacter were enriched in early-stage CRC, whereas Bacteroides, Fusobacterium, Lachnoanaerobaculum, Clostridium, and Granulicatella were enriched in late-stage CRC patients. The MDI derived from these bacteria had 64.2% predictive ability for CRC stages, with sensitivity and specificity of 61% and 69%, respectively. The AKP effect was found for history of smoking, alcohol consumption, and diabetes, whereas obesity and hypertension showed anti-AKP effect. For dietary choices, the hierarchical tree of foods and nutrients did not shape the microbial between-sample diversity. Additionally, high MDI was related with a high intake of pizza/hamburger, poultry, and light-vegetable combination. Conclusion: Our study contributed to current evidence of the microbiome structure and dysbiosis at different stages of CRC. Furthermore, history of smoking, alcohol consumption, and diabetes showed AKP effect in CRC patients. Non-significant association between dietary choices and microbiome diversity supported a small variation of microbiome profiles explained by dietary intake at the population level. Citation Format: Tung Hoang, Min Jung Kim, Ji Won Park, Seung-Yong Jeong, Jeeyoo Lee, Aesun Shin. Modifiable factors and gut microbiome alteration among colorectal cancer patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3062.

Lipidomic profiling of exosomes from colorectal cancer cells and patients reveals potential biomarkers.

Strong evidence suggests that differences in the molecular composition of lipids in exosomes depend on the cell type and has an influence on cancer initiation and progression. Here, we analyzed by liquid chromatography–mass spectrometry (LC‐MS) the lipidomic signature of exosomes derived from the human cell lines normal colon mucosa (NCM460D), and colorectal cancer (CRC) nonmetastatic (HCT116) and metastatic (SW620), and exosomes isolated from the plasma of nonmetastatic and metastatic CRC patients and healthy donors. Analysis of this exhaustive lipid study highlighted changes in some molecular species that were found in the cell lines and confirmed in the patients. For example, exosomes from primary cancer patients and nonmetastatic cells compared with healthy donors and control cells displayed a common marked increase in phosphatidylcholine (PC) 34 : 1, phosphatidylethanolamine (PE) 36 : 2, sphingomyelin (SM) d18 : 1/16 : 0, hexosylceramide (HexCer) d18 : 1/24 : 0 and HexCer d18 : 1/24 : 1. Interestingly, these same lipids species were decreased in the metastatic cell line and patients. Further, levels of PE 34 : 2, PE 36 : 2, and phosphorylated PE p16 : 0/20 : 4 were also significantly decreased in metastatic conditions when compared to the nonmetastatic counterparts. The only molecule species found markedly increased in metastatic conditions (in both patients and cells) when compared to controls was ceramide (Cer) d18 : 1/24 : 1. These decreases in lipid species in the extracellular vesicles might reflect function‐associated changes in the metastatic cell membrane. Although these potential biomarkers need to be validated in a larger cohort, they provide new insight toward the use of clusters of lipid biomarkers rather than a single molecule for the diagnosis of different stages of CRC.