bims-tucedo Biomed News
on Tumor cell dormancy
Issue of 2020‒09‒27
eighteen papers selected by
Isabel Puig Borreil
Vall d’Hebron Institute of Oncology

  1. Cancer Res. 2020 Sep 23. pii: canres.0360.2020. [Epub ahead of print]
      Histone methyltransferase NSD3 is frequently dysregulated in human cancers, yet the epigenetic role of NSD3 during cancer development remains elusive. Here we report that NSD3-induced methylation of H3K36 is crucial for breast tumor initiation and metastasis. In breast cancer patients, elevated expression of NSD3 was associated with recurrence, distant metastasis, and poor survival. In vivo, NSD3 promoted malignant transformation of mammary epithelial cells, a function comparable to that of HRAS. Furthermore, NSD3 expanded breast cancer-initiating cells and promoted epithelial-mesenchymal transition to trigger tumor invasion and metastasis. Mechanistically, the long isoform (full-length transcript) of NSD3, but not its shorter isoform lacking a catalytic domain, cooperated with EZH2 and RNA polymerase II to stimulate H3K36me2/3-dependent transactivation of genes associated with NOTCH receptor cleavage, leading to nuclear accumulation of NICD and NICD-mediated transcriptional repression of E-cadherin. Furthermore, mice harboring primary and metastatic breast tumors with overexpressed NSD3 showed sensitivity to NOTCH inhibition. Together, our findings uncover the critical epigenetic role of NSD3 in the modulation of NOTCH-dependent breast tumor progression, providing a rationale for targeting the NSD3-NOTCH signaling regulatory axis in aggressive breast cancer.
  2. Oncogene. 2020 Sep 22.
      Metastasis is the cause for 90% of cancer-related mortalities. Identification of genetic drivers promoting dissemination of tumor cells may provide opportunities for novel therapeutic strategies. We previously reported an in vivo gain-of-function screen that identified ~30 genes with a functional role in metastasis promotion and characterized detailed mechanistic functions of two hits. In this study, we characterized the contribution of one of the identified genes, MBIP (MAP3K12 binding inhibitory protein), towards driving tumor invasion and metastasis. We demonstrate that expression of MBIP significantly enhances the cellular proliferation, migration and invasion of NSCLC cells in vitro and metastasis in vivo. We functionally characterized that MBIP mediates activation of the JNK pathway and induces expression of matrix metalloproteinases (MMPs), which are necessary for the invasive and metastatic phenotype. Our findings establish a novel mechanistic role of MBIP as a driver of NSCLC progression and metastasis.
  3. Nat Immunol. 2020 Sep 21.
      Acquisition of a lipid-laden phenotype by immune cells has been defined in infectious diseases and atherosclerosis but remains largely uncharacterized in cancer. Here, in breast cancer models, we found that neutrophils are induced to accumulate neutral lipids upon interaction with resident mesenchymal cells in the premetastatic lung. Lung mesenchymal cells elicit this process through repressing the adipose triglyceride lipase (ATGL) activity in neutrophils in prostaglandin E2-dependent and -independent manners. In vivo, neutrophil-specific deletion of genes encoding ATGL or ATGL inhibitory factors altered neutrophil lipid profiles and breast tumor lung metastasis in mice. Mechanistically, lipids stored in lung neutrophils are transported to metastatic tumor cells through a macropinocytosis-lysosome pathway, endowing tumor cells with augmented survival and proliferative capacities. Pharmacological inhibition of macropinocytosis significantly reduced metastatic colonization by breast tumor cells in vivo. Collectively, our work reveals that neutrophils serve as an energy reservoir to fuel breast cancer lung metastasis.
  4. Cancers (Basel). 2020 Sep 19. pii: E2677. [Epub ahead of print]12(9):
      Metastasis-related complications account for the overwhelming majority of breast cancer mortalities. Triple negative breast cancer (TNBC), the most aggressive breast cancer subtype, has a high propensity to metastasize to distant organs, leading to poor patient survival. The forkhead transcription factor, FOXM1, is especially upregulated and overexpressed in TNBC and is known to regulate multiple signaling pathways that control many key cancer properties, including proliferation, invasiveness, stem cell renewal, and therapy resistance, making FOXM1 a critical therapeutic target for TNBC. In this study, we test the effectiveness of a novel class of 1,1-diarylethylene FOXM1 inhibitory compounds in suppressing TNBC cell migration, invasion, and metastasis using in vitro cell culture and in vivo tumor models. We show that these compounds inhibit the motility and invasiveness of TNBC MDA-MB-231 and DT28 cells, along with reducing the expression of important epithelial to mesenchymal transition (EMT) associated genes. Further, orthotopic tumor studies in NOD-SCID-gamma (NSG) mice demonstrate that these compounds reduce FOXM1 expression and suppress TNBC tumor growth as well as distant metastasis. Gene expression and protein analyses confirm the decreased levels of EMT factors and FOXM1-regulated target genes in tumors and metastatic lesions in the inhibitor-treated animals. The findings suggest that these FOXM1 suppressive compounds may have therapeutic potential in treating triple negative breast cancer, with the aim of reducing tumor progression and metastatic outgrowth.
    Keywords:  EMT; FOXM1; invasion; metastasis; migration; triple negative breast cancer
  5. Nat Commun. 2020 09 24. 11(1): 4840
      Immunotherapies revolutionized cancer treatment by harnessing the immune system to target cancer cells. However, most patients are resistant to immunotherapies and the mechanisms underlying this resistant is still poorly understood. Here, we report that overexpression of BMP7, a member of the TGFB superfamily, represents a mechanism for resistance to anti-PD1 therapy in preclinical models and in patients with disease progression while on immunotherapies. BMP7 secreted by tumor cells acts on macrophages and CD4+ T cells in the tumor microenvironment, inhibiting MAPK14 expression and impairing pro-inflammatory responses. Knockdown of BMP7 or its neutralization via follistatin in combination with anti-PD1 re-sensitizes resistant tumors to immunotherapies. Thus, we identify the BMP7 signaling pathway as a potential immunotherapeutic target in cancer.
  6. Cancer Res. 2020 Sep 25. pii: canres.1162.2020. [Epub ahead of print]
      Lung squamous carcinoma (LUSC) is a highly metastatic disease with a poor prognosis. Using an integrated screening approach, we found that miR-671-5p reduces LUSC metastasis by inhibiting a circular RNA (circRNA), CDR1as. Although the putative function of circRNA is through miRNA sponging, we found that miR-671-5p more potently silenced an axis of CDR1as and its antisense transcript, cerebellar degeneration related protein 1 (CDR1). Silencing of CDR1as or CDR1 significantly inhibited LUSC metastases and CDR1 was sufficient to promote migration and metastases. CDR1, which directly interacted with adaptor protein 1 (AP1) complex subunits and COPI proteins, no longer promoted migration upon blockade of Golgi trafficking. Therapeutic inhibition of the CDR1as/CDR1 axis with miR-671-5p mimics reduced metastasis in vivo. This report demonstrates a novel role for CDR1 in promoting metastasis and Golgi trafficking. These findings reveal a miRNA/circRNA axis that regulates LUSC metastases through a previously unstudied protein, CDR1.
  7. Cancer Cell. 2020 Sep 14. pii: S1535-6108(20)30427-X. [Epub ahead of print]
      PIK3CA, encoding the PI3Kα isoform, is the most frequently mutated oncogene in estrogen receptor (ER)-positive breast cancer. Isoform-selective PI3K inhibitors are used clinically but intrinsic and acquired resistance limits their utility. Improved selection of patients that will benefit from these drugs requires predictive biomarkers. We show here that persistent FOXM1 expression following drug treatment is a biomarker of resistance to PI3Kα inhibition in ER+ breast cancer. FOXM1 drives expression of lactate dehydrogenase (LDH) but not hexokinase 2 (HK-II). The downstream metabolic changes can therefore be detected using MRI of LDH-catalyzed hyperpolarized 13C label exchange between pyruvate and lactate but not by positron emission tomography measurements of HK-II-mediated trapping of the glucose analog 2-deoxy-2-[18F]fluorodeoxyglucose. Rapid assessment of treatment response in breast cancer using this imaging method could help identify patients that benefit from PI3Kα inhibition and design drug combinations to counteract the emergence of resistance.
    Keywords:  FDG-PET; FOXM1; MRI; PI3K alpha inhibition; biomarker; breast cancer; hexokinase 2; hyperpolarized [1-(13)C]pyruvate; lactate dehydrogenase; treatment response
  8. Mol Ther. 2020 Sep 05. pii: S1525-0016(20)30428-7. [Epub ahead of print]
      Bladder cancer patients with lymph node (LN) metastasis have an extremely poor prognosis and no effective treatment. The alternative splicing of precursor (pre-)mRNA participates in the progression of various tumors. However, the precise mechanisms of splicing factors and cancer-related variants in LN metastasis of bladder cancer remain largely unknown. The present study identified a splicing factor, non-POU domain-containing octamer-binding protein (NONO), that was significantly downregulated in bladder cancer tissues and correlated with LN metastasis status, tumor stage, and prognosis. Functionally, NONO markedly inhibited bladder cancer cell migration and invasion in vitro and LN metastasis in vivo. Mechanistically, NONO regulated the exon skipping of SETMAR by binding to its motif, mainly through the RRM2 domain. NONO directly interacted with splicing factor proline/glutamine rich (SFPQ) to regulate the splicing of SETMAR, and it induced metastasis suppression of bladder cancer cells. SETMAR-L overexpression significantly reversed the metastasis of NONO-knockdown bladder cancer cells, both in vitro and in vivo. The further analysis revealed that NONO-mediated SETMAR-L can induce H3K27me3 at the promotor of metastatic oncogenes and inhibit their transcription, ultimately resulting in metastasis suppression. Therefore, the present findings uncover the molecular mechanism of lymphatic metastasis in bladder cancer, which may provide novel clinical markers and therapeutic strategies for LN-metastatic bladder cancer.
    Keywords:  NONO; SETMAR; alternative splicing; bladder cancer; lymphatic metastasis
  9. Nat Commun. 2020 09 23. 11(1): 4807
      Non-invasive assessment of the risk of lymph node metastasis (LNM) in patients with papillary thyroid carcinoma (PTC) is of great value for the treatment option selection. The purpose of this paper is to develop a transfer learning radiomics (TLR) model for preoperative prediction of LNM in PTC patients in a multicenter, cross-machine, multi-operator scenario. Here we report the TLR model produces a stable LNM prediction. In the experiments of cross-validation and independent testing of the main cohort according to diagnostic time, machine, and operator, the TLR achieves an average area under the curve (AUC) of 0.90. In the other two independent cohorts, TLR also achieves 0.93 AUC, and this performance is statistically better than the other three methods according to Delong test. Decision curve analysis also proves that the TLR model brings more benefit to PTC patients than other methods.
  10. Mol Cancer Res. 2020 Sep 24. pii: molcanres.0353.2020. [Epub ahead of print]
      Rapid tumor growth, widespread brain-invasion, and therapeutic resistance critically contribute to glioblastoma (GBM) recurrence and dismal patient outcomes. Although GBM stem cells (GSCs) are shown to play key roles in these processes, the molecular pathways governing the GSC phenotype (GBM-stemness) remain poorly defined. Here, we show that epigenetic silencing of miR-146a significantly correlated with worse patient outcome and importantly, miR-146a level was significantly lower in recurrent tumors compared with primary ones. Further, miR-146a overexpression significantly inhibited the proliferation and invasion of GBM patient-derived primary cells and increased their response to temozolomide (TMZ), both in vitro and in vivo. Mechanistically, miR-146a directly silenced POU3F2 and SMARCA5, two transcription factors that mutually regulated each other, significantly compromising GBM-stemness and increasing TMZ response. Collectively, our data show that miR-146a-POU3F2/SMARCA5 pathway plays a critical role in suppressing GBM-stemness and increasing TMZ-response, suggesting that POU3F2 and SMARCA5 may serve as novel therapeutic targets in GBM. Implications: MicroRNA-146a predicts favorable prognosis and the miR-146a-POU3F2/SMARCA5 pathway is important for the suppression of stemness in GBM.
  11. Cancers (Basel). 2020 Sep 22. pii: E2719. [Epub ahead of print]12(9):
      Cancer cells facilitate growth and metastasis by using multiple signals from the cancer-associated microenvironment. However, it remains poorly understood whether prostate cancer (PCa) cells may recruit and utilize bone marrow cells for their growth and survival. Furthermore, the regulatory mechanisms underlying interactions between PCa cells and bone marrow cells are obscure. In this study, we isolated bone marrow cells that mainly constituted populations that were positive for CD11b and Gr1 antigens from xenograft PC-3 tumor tissues from athymic nu/nu mice. We found that the tumor-infiltrated cells alone were unable to form tumor spheroids, even with increased amounts and time. By contrast, the tumor-infiltrated cells together with PCa cells formed large numbers of tumor spheroids compared with PCa cells alone. We further utilized xenograft athymic nu/nu mice bearing bone metastatic lesions. We demonstrated that PCa cells were unable to survive and give rise to colony-forming units (CFUs) in media that were used for hematopoietic cell colony-formation unit (CFU) assays. By contrast, PC-3M cells survived when bone marrow cells were present and gave rise to CFUs. Our results showed that PCa cells required bone marrow cells to support their growth and survival and establish bone metastasis in the host environment. We showed that PCa cells that were treated with either siRNA for PIP5K1α or its specific inhibitor, ISA-2011B, were unable to survive and produce tumor spheroids, together with bone marrow cells. Given that the elevated expression of PIP5K1α was specific for PCa cells and was associated with the induced expression of VEGF receptor 2 in PCa cells, our findings suggest that cancer cells may utilize PIP5K1α-mediated receptor signaling to recruit growth factors and ligands from the bone marrow-derived cells. Taken together, our study suggests a new mechanism that enables PCa cells to gain proliferative and invasive advantages within their associated host microenvironment. Therapeutic interventions using PIP5K1α inhibitors may not only inhibit tumor invasion and metastasis but also enhance the host immune system.
    Keywords:  PIP5K1α; bone marrow cells; prostate cancer metastasis; therapeutic interventions
  12. Cancer Discov. 2020 Sep 24. pii: CD-20-0487. [Epub ahead of print]
      Gastrointestinal stromal tumors (GIST) harboring activating mutations of PDGFRA respond to imatinib with the notable exception of the most common mutation, D842V. Avapritinib is a novel, potent KIT/PDGFRA inhibitor with substantial clinical activity in patients with the D842V genotype. To date, only a minority of PDGFRA-mutant patients treated with avapritinib have developed secondary resistance. Tumor and plasma biopsies in 6 of 7 patients with PDGFRA primary mutations who progressed on avapritinib or imatinib had secondary resistance mutations within PDGFRA exons 13, 14 and 15 that interfere with avapritinib binding. Secondary PDGFRA mutations causing V658A, N659K, Y676C, and G680R substitutions were found in 2 or more patients each, representing recurrent mechanisms of PDGFRA GIST drug resistance. Notably, most PDGFRA-mutant GIST refractory to avapritinib remain dependent on the PDGFRA oncogenic signal. Inhibitors that target PDGFRA protein stability or inhibition of PDGFRA-dependent signaling pathways may overcome avapritinib resistance.
  13. Clin Cancer Res. 2020 Sep 23. pii: clincanres.2269.2020. [Epub ahead of print]
      PURPOSE: Cyclin-dependent kinase 4/6 (CDK4/6) inhibitors have improved progression free survival for metastatic, estrogen receptor positive (ER+) breast cancers, but their role in the non-metastatic setting remains unclear. We sought to understand the effects of CDK4/6 inhibition (CDK4/6i) and radiation (RT) in multiple preclinical breast cancer models.METHODS: Transcriptomic and proteomic analyses were used to identify significantly altered pathways after CDK4/6i. Clonogenic assays were used to quantify the RT enhancement ratio (rER). DNA damage was quantified using γH2AX staining and the neutral comet assay. DNA repair was assessed using RAD51 foci formation and non-homologous end joining (NHEJ) reporter assays. Orthotopic xenografts were used to assess the efficacy of combination therapy.
    RESULTS: Palbociclib significantly radiosensitized multiple ER+ cell lines at low nanomolar, sub IC50 concentrations (rER: 1.21 - 1.52) and led to a decrease in the surviving fraction of cells at 2 Gy (p < 0.001). Similar results were observed in ribociclib- (rER: 1.08 - 1.68) and abemaciclib-treated (rER: 1.19 - 2.05) cells. Combination treatment decreased RAD51 foci formation (p < 0.001), leading to a suppression of HR activity, but did not affect NHEJ efficiency (p > 0.05). Immortalized breast epithelial cells and cells with acquired resistance to CDK4/6i did not demonstrate radiosensitization (rER: 0.94 - 1.11) or changes in RAD51 foci. In xenograft models, concurrent palbociclib and RT led to a significant decrease in tumor growth.
    CONCLUSIONS: These studies provide preclinical rationale to test CDK4/6i + RT in women with locally-advanced ER+ breast cancer at high risk for locoregional recurrence.
  14. Nat Commun. 2020 Sep 25. 11(1): 4875
      Single-cell whole-exome sequencing (scWES) is a powerful approach for deciphering intratumor heterogeneity and identifying cancer drivers. So far, however, simultaneous analysis of single nucleotide variants (SNVs) and copy number variations (CNVs) of a single cell has been challenging. By analyzing SNVs and CNVs simultaneously in bulk and single cells of premalignant tissues and tumors from mouse and human BRCA1-associated breast cancers, we discover an evolution process through which the tumors initiate from cells with SNVs affecting driver genes in the premalignant stage and malignantly progress later via CNVs acquired in chromosome regions with cancer driver genes. These events occur randomly and hit many putative cancer drivers besides p53 to generate unique genetic and pathological features for each tumor. Upon this, we finally identify a tumor metastasis suppressor Plekha5, whose deficiency promotes cancer metastasis to the liver and/or lung.
  15. Clin Cancer Res. 2020 Sep 22. pii: clincanres.2793.2020. [Epub ahead of print]
      PURPOSE: Predicting prognosis in HR+/HER2-negative MBC might be clinically useful; however, no validated prognostic biomarkers exist in this setting to date.EXPERIMENTAL DESIGN: In phase III, EGF30008 trial, 484 patients with HER2-negative MBC who received letrozole and placebo or lapatinib were selected. PAM50 data, ECOG PS, visceral disease, number of metastasis, biopsy type, and age were evaluated. A progression-free survival (PFS) Cox model was evaluated. The final model (PAM50MET) with a prespecified cutoff was validated in patients (n=261) with HR+/HER2-negative aBC from BOLERO-2 (phase III trial that evaluated exemestane and placebo or everolimus).
    RESULTS: In EGF30008, prognostic models with PAM50 plus clinical variables yielded higher C-index values versus models with only PAM50 or clinical variables. The PAM50MET model combined 21 variables: 2 PAM50 subtypes, basal signature, 14 genes, and 4 clinical variables. In EGF30008, the optimized cutoff was associated with PFS (hazard ratio [HR]=0.37; 95% confidence interval [CI] 0.29-0.47; P<0.0001) and overall survival (OS; HR=0.37; 0.27-0.51; P<0.0001). The median (months) (95% CI) PFS and OS were 22.24 (19.0-24.9) and not reached in PAM50MET-low versus 9.13 (8.15-11.0) and 33.0 (28.0-40.0) in PAM50MET-high groups, respectively. In BOLERO-2, the PAM50MET-low was associated with better PFS (HR=0.72; 95% CI 0.53-0.96; P=0.028) and OS (HR=0.51; 95% CI 0.35-0.69; P<0.0001). The median (months) (95% CI) PFS and OS were 6.93 (5.57-11.0) and 36.9 (33.4-NA) in PAM50MET-low versus 5.23 (4.2-6.8) and 23.5 (20.2-28.3) in PAM50MET-high groups, respectively.
    CONCLUSION: PAM50MET is prognostic in HR+/HER2-negative MBC, and further evaluation might help identify candidates for ET only or novel therapies.
  16. N Engl J Med. 2020 Sep 20.
      BACKGROUND: We previously reported that olaparib led to significantly longer imaging-based progression-free survival than the physician's choice of enzalutamide or abiraterone among men with metastatic castration-resistant prostate cancer who had qualifying alterations in homologous recombination repair genes and whose disease had progressed during previous treatment with a next-generation hormonal agent. The results of the final analysis of overall survival have not yet been reported.METHODS: In an open-label, phase 3 trial, we randomly assigned patients in a 2:1 ratio to receive olaparib (256 patients) or the physician's choice of enzalutamide or abiraterone plus prednisone as the control therapy (131 patients). Cohort A included 245 patients with at least one alteration in BRCA1, BRCA2, or ATM, and cohort B included 142 patients with at least one alteration in any of the other 12 prespecified genes. Crossover to olaparib was allowed after imaging-based disease progression for patients who met certain criteria. Overall survival in cohort A, a key secondary end point, was analyzed with the use of an alpha-controlled, stratified log-rank test at a data maturity of approximately 60%. The primary and other key secondary end points were reported previously.
    RESULTS: The median duration of overall survival in cohort A was 19.1 months with olaparib and 14.7 months with control therapy (hazard ratio for death, 0.69; 95% confidence interval [CI], 0.50 to 0.97; P = 0.02). In cohort B, the median duration of overall survival was 14.1 months with olaparib and 11.5 months with control therapy. In the overall population (cohorts A and B), the corresponding durations were 17.3 months and 14.0 months. Overall, 86 of 131 patients (66%) in the control group crossed over to receive olaparib (56 of 83 patients [67%] in cohort A). A sensitivity analysis that adjusted for crossover to olaparib showed hazard ratios for death of 0.42 (95% CI, 0.19 to 0.91) in cohort A, 0.83 (95% CI, 0.11 to 5.98) in cohort B, and 0.55 (95% CI, 0.29 to 1.06) in the overall population.
    CONCLUSIONS: Among men with metastatic castration-resistant prostate cancer who had tumors with at least one alteration in BRCA1, BRCA2, or ATM and whose disease had progressed during previous treatment with a next-generation hormonal agent, those who were initially assigned to receive olaparib had a significantly longer duration of overall survival than those who were assigned to receive enzalutamide or abiraterone plus prednisone as the control therapy, despite substantial crossover from control therapy to olaparib. (Funded by AstraZeneca and Merck Sharp & Dohme; PROfound number, NCT02987543.).
  17. Cancer Metab. 2020 ;8 19
      Pancreatic ductal adenocarcinoma (PDAC) is one of the most malignant forms of cancer. Lack of effective treatment options and drug resistance contributes to the low survival among PDAC patients. In this study, we investigated the metabolic alterations in pancreatic cancer cells that do not respond to the EGFR inhibitor erlotinib. We selected erlotinib-resistant pancreatic cancer cells from MiaPaCa2 and AsPC1 cell lines. Metabolic profiling of erlotinib-resistant cells revealed a significant downregulation of glycolytic activity and reduced level of glycolytic metabolites compared to the sensitive cells. The resistant cells displayed elevated expression of the pentose phosphate pathway (PPP) enzymes involved in ROS regulation and nucleotide biosynthesis. The enhanced PPP elevated cellular NADPH/NADP+ ratio and protected the cells from reactive oxygen species (ROS)-induced damage. Inhibition of PPP using 6-aminonicotinamide (6AN) elevated ROS levels, induced G1 cell cycle arrest, and sensitized resistant cells to erlotinib. Genetic studies identified elevated PPP enzyme glucose-6-phosphate dehydrogenase (G6PD) as an important contributor to erlotinib resistance. Mechanistically, our data showed that upregulation of inhibitor of differentiation (ID1) regulates G6PD expression in resistant cells thus contributing to altered metabolic phenotype and reduced response to erlotinib. Together, our results highlight an underlying role of tumor metabolism in PDAC drug response and identify G6PD as a target to overcome drug resistance.
    Keywords:  Erlotinib resistance; Metabolic reprogramming; Pancreatic cancer
  18. Cancers (Basel). 2020 Sep 22. pii: E2710. [Epub ahead of print]12(9):
      Colorectal cancer is a heterogeneous disease caused by both genetic and epigenetics factors. Analysing DNA methylation changes occurring during colorectal cancer progression and metastasis formation is crucial for the identification of novel epigenetic markers of patient prognosis. Genome-wide methylation sequencing of paired samples of colon (normal adjacent, primary tumour and lymph node metastasis) showed global hypomethylation and CpG island (CGI) hypermethylation of primary tumours compared to normal. In metastasis we observed high global and non-CGI regions methylation, but lower CGI methylation, compared to primary tumours. Gene ontology analysis showed shared biological processes between hypermethylated CGIs in metastasis and primary tumours. After complementary analysis with The Cancer Genome Atlas (TCGA) cohort, FIGN, HTRA3, BDNF, HCN4 and STAC2 genes were found associated with poor survival. We mapped the methylation landscape of colon normal tissues, primary tumours and lymph node metastasis, being capable of identified methylation changes throughout the genome. Furthermore, we found five genes with potential for methylation biomarkers of poor prognosis in colorectal cancer patients.
    Keywords:  DNA methylation; colorectal cancer; genome-wide analysis; lymph node; metastasis