Metastasis Of Colorectal Carcinoma Research Articles

P4-13 cMET promotes metastasis and epithelial-mesenchymal transition in colorectal carcinoma via repressing RKIP

Colorectal cancer (CRC) ranks among the most frequent malignancies globally and is associated with high metastasis incidence. Today, it is becoming more prevalent among Asian populations, especially in the Chinese. There is increasing evidence that c-mesenchymal-epithelial transition factor (cMET) plays an important role in malignant progression, and its overexpression is closely linked with poor survival and enhanced resistance to drug therapy, hence abnormal cMET is viewed as a signature event in CRC. As a metastasis suppressor, loss of raf kinase inhibitory protein (RKIP) has been reported in many of cancer types. Both of them may serve to regulate CRC initiation and progression. It is therefore necessary to address the interaction between cMET/ERK signal axis and RKIP. In this study, expression of cMET and RKIP in CRC tissues and cell lines were determined, and their crosstalk and potential biological effects were explored using. Immunofluorescence staining and Chromatin immunoprecipitation. Our results showed cMET was inversely correlated with RKIP. Both cMET upregulation and RKIP downregulation implied poor clinical outcomes. Moreover, the MAPK/ERK signal pathway was involved in the regulation of cMET and RKIP. Overexpression of cMET promoted epithelial mesenchymal transition (EMT), invasion, migration and chemoresistance of tumor cells, whereas the impact could be efficiently impeded by increased RKIP. Notably, shRNA-mediated cMET knockdown dramatically suppressed cell proliferation, although no influence of RKIP on cell growth was observed in CRC. Altogether, cMET overexpression may contribute to tumor progression by inhibiting anti-oncogene RKIP, which provides preclinical justification for targeting RKIP to treat cMET-induced metastasis in CRC.

Metabolic Reprogramming of Colorectal Cancer Cells and the Microenvironment: Implication for Therapy.

Colorectal carcinoma (CRC) is one of the most frequently diagnosed carcinomas and one of the leading causes of cancer-related death worldwide. Metabolic reprogramming, a hallmark of cancer, is closely related to the initiation and progression of carcinomas, including CRC. Accumulating evidence shows that activation of oncogenic pathways and loss of tumor suppressor genes regulate the metabolic reprogramming that is mainly involved in glycolysis, glutaminolysis, one-carbon metabolism and lipid metabolism. The abnormal metabolic program provides tumor cells with abundant energy, nutrients and redox requirements to support their malignant growth and metastasis, which is accompanied by impaired metabolic flexibility in the tumor microenvironment (TME) and dysbiosis of the gut microbiota. The metabolic crosstalk between the tumor cells, the components of the TME and the intestinal microbiota further facilitates CRC cell proliferation, invasion and metastasis and leads to therapy resistance. Hence, to target the dysregulated tumor metabolism, the TME and the gut microbiota, novel preventive and therapeutic applications are required. In this review, the dysregulation of metabolic programs, molecular pathways, the TME and the intestinal microbiota in CRC is addressed. Possible therapeutic strategies, including metabolic inhibition and immune therapy in CRC, as well as modulation of the aberrant intestinal microbiota, are discussed.

BMAL1 Knockdown Leans Epithelial-Mesenchymal Balance toward Epithelial Properties and Decreases the Chemoresistance of Colon Carcinoma Cells.

The circadian clock coordinates biological and physiological functions to day/night cycles. The perturbation of the circadian clock increases cancer risk and affects cancer progression. Here, we studied how BMAL1 knockdown (BMAL1-KD) by shRNA affects the epithelial–mesenchymal transition (EMT), a critical early event in the invasion and metastasis of colorectal carcinoma (CRC). In corresponding to a gene set enrichment analysis, which showed a significant enrichment of EMT and invasive signatures in BMAL1_high CRC patients as compared to BMAL1_low CRC patients, our results revealed that BMAL1 is implicated in keeping the epithelial–mesenchymal equilibrium of CRC cells and influences their capacity of adhesion, migration, invasion, and chemoresistance. Firstly, BMAL1-KD increased the expression of epithelial markers (E-cadherin, CK-20, and EpCAM) but decreased the expression of Twist and mesenchymal markers (N-cadherin and vimentin) in CRC cell lines. Finally, the molecular alterations after BMAL1-KD promoted mesenchymal-to-epithelial transition-like changes mostly appeared in two primary CRC cell lines (i.e., HCT116 and SW480) compared to the metastatic cell line SW620. As a consequence, migration/invasion and drug resistance capacities decreased in HCT116 and SW480 BMAL1-KD cells. Together, BMAL1-KD alerts the delicate equilibrium between epithelial and mesenchymal properties of CRC cell lines, which revealed the crucial role of BMAL1 in EMT-related CRC metastasis and chemoresistance.

Epithelial-Mesenchymal Transition in Colorectal Carcinoma: Comparison Between Primary Tumor, Lymph Node and Liver Metastases.

There is emerging evidence suggesting that epithelial-mesenchymal transition (EMT) and mesenchymal-epithelial transition (MET) play an important role in colorectal carcinoma (CRC), but their exact role remains controversial. Our aim was to analyze the miR-200 family as EMT markers and their target genes expression at invasive tumor front and in nodal and liver metastases. Sixty-three formalin-fixed paraffin-embedded tissue samples from 19 patients with CRC were included. Using a micropuncture technique, tissue was obtained from central part and invasive front of the primary tumor, and nodal and liver metastases. Expression of the miR-200 family and their target genes CDKN1B, ONECUT2, PTPN13, RND3, SOX2, TGFB2 and ZEB2 was analyzed using real-time PCR. We found miR-200 family down-regulation at invasive front compared to central part, and up-regulation of miRNA-200a/b/c and miR-429 in metastases compared to invasive front. At invasive front, TGFB2 was the only gene with inverse expression to the miR-200 family, whereas in metastases inverse expression was found for ONECUT2 and SOX2. CDKN1B, PTPN13 and ZEB2 were down-regulated at invasive front and up-regulated in metastases. Our results suggest the involvement of partial EMT at invasive tumor front, and partial MET in metastases in CRC, based on miR-200 family and its target genes expression.

Abelson interactor 1 splice isoform-L plays an anti-oncogenic role in colorectal carcinoma through interactions with WAVE2 and full-length Abelson interactor 1.

BACKGROUNDExpression of the full-length isoform of Abelson interactor 1 (ABI1), ABI1-p65, is increased in colorectal carcinoma (CRC) and is thought to be involved in one or more steps leading to tumor progression or metastasis. The ABI1 splice isoform-L (ABI1-SiL) has conserved WAVE2-binding and SH3 domains, lacks the homeo-domain homologous region, and is missing the majority of PxxP- and Pro-rich domains found in full-length ABI1-p65. Thus, ABI1-SiL domain structure suggests that the protein may regulate CRC cell morphology, adhesion, migration, and metastasis via interactions with the WAVE2 complex pathway.AIMTo investigate the potential role and underlying mechanisms associated with ABI1-SiL-mediated regulation of CRC.METHODSABI1-SiL mRNA expression in CC tissue and cell lines was measured using both qualitative reverse transcriptase-polymerase chain reaction (RT-PCR) and real-time quantitative RT-PCR. A stably ABI1-SiL overexpressing SW480 cell model was constructed using Lipofectamine 2000, and cells selected with G418. Image J software, CCK8, and transwell assays were used to investigate SW480 cell surface area, proliferation, migration, and invasion. Immunoprecipitation, Western blot, and co-localization assays were performed to explore intermolecular interactions between ABI1-SiL, WAVE2, and ABI1-p65 proteins.RESULTSABI1-SiL was expressed in normal colon tissue and was significantly decreased in CRC cell lines and tissues. Overexpression of ABI1-SiL in SW480 cells significantly increased the cell surface area and inhibited the adhesive and migration properties of the cells, but did not alter their invasive capacity. Similar to ABI1-p65, ABI1-SiL still binds WAVE2, and the ABI1-p65 isoform in SW480 cells. Furthermore, co-localization assays confirmed these intermolecular interactions.CONCLUSIONThese results support a model in which ABI1-SiL plays an anti-oncogenic role by competitively binding to WAVE2 and directly interacting with phosphorylated and non-phosphorylated ABI1-p65, functioning as a dominant-negative form of ABI1-p65.

A study of the correlation between M2 macrophages and lymph node metastasis of colorectal carcinoma

BackgroundLymph node metastasis is a major prognostic sign of colorectal carcinoma and an important indicator for individualized treatment. M2 macrophages play a key role in carcinogenesis and tumor development by enhancing invasiveness and promoting lymph node metastasis. The purpose of this study was to investigate the effect of CD163-positive M2 macrophages on lymph node metastasis in colorectal carcinoma.MethodsPostoperative lymph node tissues were obtained from 120 patients with colorectal carcinoma who underwent radical surgery in the First Affiliated Hospital of Jinzhou Medical University between December 2019 and May 2020. We detected the expression of the CD163 protein in lymph nodes using immunohistochemistry. Furthermore, the relationships between M2 macrophages identified by expression of CD163 and lymph node metastasis were analyzed using the independent sample t-test and Chi-square test.ResultsM2 macrophages were increased in metastatic lymph nodes and non-metastatic lymph nodes adjacent to the cancer. The M2 macrophage count was higher in patients with macro-metastases than in patients with micro-metastases.ConclusionsThe presence of M2 macrophages represents an important indicator for lymph node metastasis in colorectal carcinoma and may be a potential marker for its prediction. Thus, M2 macrophage localization might offer a new target for the comprehensive treatment of colorectal carcinoma.

Membrane-cytoplasm translocation of annexin A4 is involved in the metastasis of colorectal carcinoma.

Annexin A4 (ANXA4) is a Ca2+- and phospholipid-binding protein that belongs to the annexin family, which is involved in the development of multiple tumour types via NF-κB signalling. In this study, we verified the high expression and membrane-cytoplasm translocation of ANXA4 in colorectal carcinoma (CRC). Calcium/calmodulin-dependent protein kinase II gamma (CAMK2γ) was found to be important for high ANXA4 expression in CRC, whereas carbonic anhydrase (CA1) promoted ANXA4 aggregation in the cell membrane. An increased Ca2+ concentration attenuated the small ubiquitin-like modifier (SUMO) modification of cytoplasmic ANXA4 and ANXA4 stabilization, and relatively high expression of ANXA4 promoted CRC tumorigenesis and epithelial-mesenchymal transition (EMT).

ENC1 Facilitates Colorectal Carcinoma Tumorigenesis and Metastasis via JAK2/STAT5/AKT Axis-Mediated Epithelial Mesenchymal Transition and Stemness.

Ectodermal neural cortex 1 (ENC1) is an actin-binding protein and has been known to be upregulated in several cancers, but the molecular mechanisms through which it contributes to the pathology of CRC have largely been elusive. We utilized data mining and validated the aberrant expression of ENC1, following which phenotypic traits of malignancy were assessed in vitro. Ruxolitinib was used as a surrogate to compare the effects of ENC1 expression and silencing on the JAK-STAT-AKT pathway. In vivo models were employed to confirm the in vitro observations. Computation analysis, strengthened by in situ and in vitro data, confirmed the overexpression of ENC1 in CRC and predicted a poor prognosis, while enhanced cell proliferation, invasion, migration, EMT, and stemness were associated with ENC1 overexpression. Silencing of ENC1 downregulated the phenotypes. Additionally, silencing of ENC1 significantly reduced the activation of JAK2 and consequent activation of STAT5 and AKT comparable to ruxolitinib inhibition of JAK2. Silencing of ENC1 resulted in lesser tumor volumes and fewer numbers of tumors, in vivo. These data suggest that ENC1 induces CRC through the JAK2-STAT5-AKT axis. ENC1 is a suitable diagnostic marker for CRC detection, and ENC1 targeting therapies may suppress CRC progression.

Unusual case of colorectal carcinoma with sarcoid-like mediastinal adenopathy and near-fatal DPD deficiency

The liver is the commonest site for metastases in colorectal carcinoma; other isolated sites are considered extremely rare. 5-fluorouracil (5-FU) is the backbone of treatment for metastatic colorectal carcinoma (mCRC) and without it survival may be significantly reduced. It is primarily metabolised by dihydropyrimidine dehydrogenase (DPD). Testing for DPD deficiency is not a routine practice and toxicity will only manifest following drug challenge. There are limited standardised treatment guidelines in managing patients with severe drug reactions following 5-FU exposure. We describe a delayed presentation of life-threatening DPD deficiency in a patient with colorectal carcinoma and mediastinal lymphadenopathy. We describe our experience with chemotherapy in this difficult clinical scenario and highlight the importance of histological confirmation in unusual sites of metastatic disease.

Enhancer-derived long non-coding RNAs CCAT1 and CCAT2 at rs6983267 has limited predictability for early stage colorectal carcinoma metastasis

Up-regulation of long non-coding RNAs (lncRNAs), colon-cancer associated transcript (CCAT) 1 and 2, was associated with worse prognosis in colorectal cancer (CRC). Nevertheless, their role in predicting metastasis in early-stage CRC is unclear. We measured the expression of CCAT1, CCAT2 and their oncotarget, c-Myc, in 150 matched mucosa-tumour samples of early-stage microsatellite-stable Chinese CRC patients with definitive metastasis status by multiplex real-time RT-PCR assay. Expression of CCAT1, CCAT2 and c-Myc were significantly up-regulated in the tumours compared to matched mucosa (p < 0.0001). The expression of c-Myc in the tumours was significantly correlated to time to metastasis [hazard ratio = 1.47 (1.10–1.97)] and the risk genotype (GG) of rs6983267, located within CCAT2. Expression of c-Myc and CCAT2 in the tumour were also significantly up-regulated in metastasis-positive compared to metastasis-negative patients (p = 0.009 and p = 0.04 respectively). Nevertheless, integrating the expression of CCAT1 and CCAT2 by the Random Forest classifier did not improve the predictive values of ColoMet19, the mRNA-based predictor for metastasis previously developed on the same series of tumours. The role of these two lncRNAs is probably mitigated via their oncotarget, c-Myc, which was not ranked high enough previously to be included in ColoMet19.

Risk Factors for Lymph Node Metastasis in Pathological T1b Colorectal Cancer.

The rate of lymph node metastasis (LNM) of colorectal carcinoma (CRC) with a submucosal (SM) invasion depth of 1000 µm or more can reach 12.5%, which is the most common reason for additional resection in daily practice. Other studies have reported that the rate of LNM is less than 2%, regardless of the depth of invasion, if the lesions show good histology, lymphovascular infiltration is negative, and tumor budding is limited. The purpose of this study was to investigate new risk factors for LNM in T1b colorectal cancer. The 239 patients who were diagnosed with pathological T1b CRC after colorectal surgical resection at the Osaka Police Hospital in Japan between January 2008 and December 2018 were retrospectively reviewed in this study. The LNM rate was 11.3% (27/239). The variables identified as being significant factors using multivariate analysis were: i) lymphatic invasion (Ly)-positive [odds ratio (OR)=5.97; 95% confidence interval (CI)=2.27-15.74], ii) female gender (OR=3.49; 95%CI=1.38-8.85), and iii) left-sided colorectal involvement (OR=4.98; 95%CI=1.22-20.39). If none of these risk factors were present with T1b, the LNM rate was 0% (0/28). Ly-positive, female gender, and left-sided colorectal involvement could be risk factors for LNM in T1b CRC.

Penile Metastasis of Colorectal Cancer Mimicking Priapism with Acute Urinary Retention: A Rare Case

Background: Penile metastasis of colorectal carcinoma is a rare phenomenon in clinical setting. They normally manifest as penile lesion and acute urinary retention. However, presentation of priapism is exceedingly rare. Aims: Discussion of this rare presentation as well as the diagnostic processes and subsequent management. Case Presentation: A 54-year-old male with a history of colorectal cancer presents with acute urinary retention. Examination of the patient demonstrates a semi-erect penis, with multiple palpable nodules on the shaft and penile meatus. Histological and imaging findings indicate penile metastasis of colorectal cancer. Conclusion: Biopsy via cystoscopy is used to obtain definitive diagnosis of penile metastasis. Urinary drainage followed by further cancer intervention or palliative care is crucial for effective management.

Correlation of C-arm CT acquired parenchymal blood volume (PBV) with 99mTc-macroaggregated albumin (MAA) SPECT/CT for radioembolization work-up

SPECT/CT with 99mTc-macroaggregated albumin (MAA) is generally used for diagnostic work-up prior to transarterial radioembolization (TARE) to exclude shunts and to provide additional information for treatment stratification and dose calculation. C-arm CT is used for determination of lobular vascular supply and assessment of parenchymal blood volume (PBV). Aim of this study was to correlate MAA-uptake and PBV-maps in hepatocellular carcinoma (HCC) and hepatic metastases of the colorectal carcinoma (CRC). 34 patients underwent a PBV C-arm CT immediately followed by 99mTc-MAA injection and a SPECT/CT acquisition after 1 h uptake. MAA-uptake and PBV-maps were visually assessed and semi-quantitatively analyzed (MAA-tumor/liver-parenchyma = MAA-TBR or PBV in ml/100ml). In case of a poor match, tumors were additionally correlated with post-TARE 90Y-Bremsstrahlung-SPECT/CT as a reference. 102 HCC or CRC metastases were analyzed. HCC presented with significantly higher MAA-TBR (7.6 vs. 3.9, p<0.05) compared to CRC. Tumors showed strong intra- and inter-individual dissimilarities between TBR and PBV with a weak correlations for capsular HCCs (r = 0.45, p<0.05) and no correlation for CRC. The demarcation of lesions was slightly better for both HCC and CRC in PBV-maps compared to MAA-SPECT/CT (exact match: 52%/50%; same intensity/homogeneity: 38%/39%; insufficient 10%/11%). MAA-SPECT/CT revealed a better visual correlation with post-therapeutic 90Y-Bremsstrahlung-SPECT/CT. The acquisition of PBV can improve the detectability of small intrahepatic tumors and correlates with the MAA-Uptake in HCC. The results indicate that 99mTc-MAA-SPECT/CT remains to be the superior method for the prediction of post-therapeutic 90Y-particle distribution, especially in CRC. However, intra-procedural PBV acquisition has the potential to become an additional factor for TARE planning, in addition to improving the determination of segment and tumor blood supply, which has been demonstrated previously.

Genotype-phenotype associations in colorectal adenocarcinomas and their matched metastases

Although primary colorectal carcinomas (CRCs) frequently share genetic alterations with their metastases, morphologic surrogates reflecting the genotype contexture of metastases remain largely unknown. We investigated phenotype/genotype associations in paired primary and metastatic colorectal adenocarcinomas from 75 patients. Thirty-three (44%) metastatic lesions were synchronous and 42 (56%) were metachronous. Tumor budding, micronecrosis, and tumor-infiltrating lymphocyte (TIL) density were compared with matched next-generation sequencing genotypes. Micronecrosis in the primary were significantly associated with nodal status (P=0.0054) and with micronecrosis in metastatic sites (P=0.0216), particularly in metachronous metastases (P=0.0033). With a 57-gene panel, one or more mutations were identified in 64 (85.3%) cases. In metastases, high (brisk) TILs were associated with overall mutational burden (P=0.0058) and with mutations in EGF (P=0.0325), RAS genes (P=0.0043), and MMR genes (P=0.0069), whereas high-level micronecrosis correlated with mutations in APC (P=0.0004) and MSH6 (P=0.0385) genes. Genomic alterations were shared in 90.1% of primary/metastatic pairs, but clonality of the same mutation was shared in only 57.1% of paired lesions. Compared with synchronous, metachronous metastases had more private clonal alterations (P=0.0291); in this group, clonal alterations coincided with brisk TILs (P=0.0334) and high micronecrosis (P=0.0133). High TILs in metastatic lesions were predictive of favorable overall survival (log-rank P=0.044). The observed phenotype/genotype associations favor the clonal evolution model in CRC metastases that seems accompanied by intense host immune response. If the role of micronecrosis and brisk TILs in metachronous metastases is validated in larger studies, these histologic parameters will be worth adding in the armamentarium for the evaluation of metastatic CRC.

Clinical significance of immunohistochemical lymphovascular evaluation to determine additional surgery after endoscopic submucosal dissection for colorectal T1 carcinoma.

The Japanese Society for Cancer of the Colon and Rectum (JSCCR) guidelines indicate lymphovascular invasion-evaluated by hematoxylin and eosin (HE) staining-as a surgical requirement after endoscopic submucosal dissection (ESD) in T1 colorectal carcinoma (CRC) patients; however, immunohistochemical evaluation may be superior. This study aimed to clarify the significance of immunohistochemical lymphovascular evaluation as an indicator for additional surgery of T1 CRC after ESD, and assessed the guidelines' adequacy, even when evaluating through immunostaining. Patients with T1 CRC who underwent ESD were enrolled across three institutions between January 2012 and December 2017. Immunohistochemical lymphovascular evaluation was performed. Clinicopathological features, pathological evaluations, and surgery indications were recorded. Univariate and multivariate logistic regression identified risk factors for lymph node (LN) metastasis of T1 CRC after ESD. Among 370 patients with T1 CRC, recurrence, 5-year overall survival, and 5-year disease specific survival rates were 1.6%, 94.6%, and 99.5%, respectively. Six patients (1.6%) experienced recurrence, five of whom underwent additional surgery. Those with no risk factors did not exhibit recurrence. A total of 215 (58.1%) patients underwent additional surgery after ESD, 21 (9.7%) of whom exhibited LN metastasis. Among 16 patients who underwent additional surgery due to lymphovascular invasion, three (18.8%) had LN metastasis. Multivariate logistic regression analysis identified lymphatic invasion as a significant risk factor for LN metastasis (odds ratio 3.9, 95% confidence interval 1.0-14.6, P = 0.0421). The JSCCR guidelines have clinical validity, and immunohistochemical lymphatic evaluation findings potentially predict LN metastasis for T1 CRC after ESD.

The Long Noncoding RNA LOC441461 (STX17-AS1) Modulates Colorectal Cancer Cell Growth and Motility.

Simple SummaryLong noncoding RNA dysfunction is crucial for colorectal carcinoma (CRC) development. Whether the dysfunction of LOC441461, a novel lncRNA, can regulate cancer-related signaling pathways in cancer progression remains unclear. Here, we uncover the oncogenic role of LOC441461 in colon cancer cell growth and motility and identify a novel mechanism for LOC441461 knockdown-induced suppression of cancer motility through modulating Ras homolog family member A (RhoA)/Rho-associated protein kinase (ROCK) activity. This is the first report that LOC441461 knockdown impairs cell cycle progression and accelerates the apoptosis of colon cancer cells following chemotherapy drug treatment. The results suggest that LOC441461 expression confers drug sensitivity in colon cancer by inducing apoptosis. Our findings offer new insight into LOC441461 regulation and provide an application for colon cancer therapy in the future.Colorectal carcinoma (CRC) is one of the most prevalent cancers worldwide and has a high mortality rate. Long noncoding RNAs (lncRNAs) have been noted to play critical roles in cell growth; cell apoptosis; and metastasis in CRC. This study determined that LOC441461 expression was significantly higher in CRC tissues than in adjacent normal mucosa. Pathway enrichment analysis of LOC441461-coexpressed genes revealed that LOC441461 was involved in biological functions related to cancer cell growth and motility. Knockdown of the LOC441461 expression significantly suppressed colon cancer cell growth by impairing cell cycle progression and inducing cell apoptosis. Furthermore, significantly higher LOC441461 expression was discovered in primary colon tumors and metastatic liver tumors than in the corresponding normal mucosa, and LOC441461 knockdown was noted to suppress colon cancer cell motility. Knockdown of LOC441461 expression suppressed the phosphorylation of MLC and LIMK1 through the inhibition of RhoA/ROCK signaling. Overall, LOC441461 was discovered to play an oncogenic role in CRC cell growth and motility through RhoA/ROCK signaling. Our findings provide new insights into the regulation of lncRNAs and their application in the treatment of colon cancer

The role of Hepatocyte nuclear factor 4 alpha HNF4α and Homeobix protein CDX2 immunomarkers in the diagnosis of metastatic colorectal carcinoma

Background: Detection of the primary site of metastatic carcinoma of unknown origin is necessary to help in the choice of the treatment. CDX2 is routinely used in metastatic adenocarcinomas cases for identifying the gastrointestinal origin. HNF4α is a new immunohistochemical marker with a few studies showing that it is expressed in a majority of colorectal adenocarcinomas. Aim: To determine the expression of HNF4α and CDX2 in metastatic carcinoma cases for detection of gastrointestinal origin. Material and methods: We assessed HNF4α and CDX2 in 60 metastatic carcinoma cases in different organs, which were diagnosed retrospectively. HNF4-α and CDX-2 expressions were evaluated by light microscopic examination of all tissue sections by two different pathologists. The cut-off for positivity of HNF4α and CDX-2 was 1% of stained nucleus of metastatic tumor cells. The sensitivity, specificity, negative predictive value (NPV), positive predictive value (PPV) and accuracy of two markers and P value were calculated. Results: The calculated sensitivity, specificity, positive predictive value, negative predictive value and accuracy of HNF4α were 90%, 82.5%, 72%, 94.3% and 85% respectively for diagnosis of colorectal carcinoma metastasis from other metastatic carcinomas. The sensitivity, specificity, positive predictive value, negative predictive value and accuracy of CDX2 were 80%, 72.5%, 59.3%, 87.9% and 75% respectively. The P-value for comparing between their expression in metastatic adenocarcinoma from lower GIT and each other groups is less than 0.05. Conclusions: We concluded that HNF4α can be used as a supplementary marker with CDX2 to identify metastatic colorectal adenocarcinoma. HNF4α expression is a highly specific and sensitive marker of colorectal origin.

Colorectal Cancer Stem Cells in the Progression to Liver Metastasis

Colorectal carcinoma (CRC) is a leading cause of cancer mortality. Tumorigenesis is a dynamic process wherein cancer stem cells (CSCs) and their microenvironment promote initiation, progression, and metastasis. Metastatic colonization is an inefficient process that is very complex and is poorly understood; however, in most cases, metastatic disease is not curable, and resistance mechanisms tend to develop against conventional treatments. An understanding of the underlying mechanisms and factors that contribute to the development of metastasis in CRC can aid in the search for specific therapeutic targets for improving standard treatments. In this review, we summarize current knowledge regarding tumor biology and the use of stroma cells as prognostic factors and inflammatory inducers associated with the use of tumor microenvironments as a promoter of cancer metastasis. Moreover, we look into the importance of CSC, pericytes, and circulating tumor cells as mechanisms that lead to liver metastasis, and we also focus on the cellular and molecular pathways that modulate and regulate epithelial–mesenchymal transition. Finally, we discuss a novel therapeutic target that can potentially eliminate CSCs as a CRC treatment.

The Protective Role of Decorin in Hepatic Metastasis of Colorectal Carcinoma.

Decorin, the prototype member of the small leucine-rich proteoglycan gene family of extracellular matrix (ECM) proteins, acts as a powerful tumor suppressor by inducing the p21Waf1/Cip1 cyclin-dependent kinase inhibitor, as well as through its ability to directly bind and block the action of several tyrosine kinase receptors. Our previous studies suggested that the lack of decorin promotes hepatic carcinogenesis in mice. Based on this, we set out to investigate whether excess decorin may protect against the liver metastases of colon carcinoma. We also analyzed the effect of decorin in tissue microarrays of human colon carcinoma liver metastasis and examined whether the tumor cells can directly influence the decorin production of myofibroblasts. In humans, low levels of decorin in the liver facilitated the development of colon carcinoma metastases in proportion with more aggressive phenotypes, indicating a possible antitumor action of the proteoglycan. In vitro, colon carcinoma cells inhibited decorin expression in LX2 hepatic stellate cells. Moreover, liver-targeted decorin delivery in mice effectively attenuated metastasis formation of colon cancer. Overexpressed decorin reduced the activity of multiple receptor tyrosine kinases (RTKs) including the epidermal growth factor receptor (EGFR), an important player in colorectal cancer (CRC) pathogenesis. Downstream of that, we observed weakened signaling of ERK1/2, PLCγ, Akt/mTOR, STAT and c-Jun pathways, while p38 MAPK/MSK/CREB and AMPK were upregulated culminating in enhanced p53 function. In conclusion, decorin may effectively inhibit metastatic tumor formation in the liver.

Abstract 3535: Whole genome sequencing of single circulating tumor cells from localized high-risk prostate cancer patients: Laser microdissection and 3Dtelomere profile

Abstract Advances in the development of precision medicine for prostate cancer patients rely on accurate identification of the genomic features underlying a patient's tumor. Genomic analysis of circulating tumor cells (CTCs) could provide a unique and accessible representation of tumor diversity, however for the use of CTCs as a less-invasive sampling of prostate cancer and as biomarkers for patient stratification and selection of targeted therapy, complex platforms are necessary to ensure an efficient enrichment (isolation), detection (identification-imaging) and characterization strategies (molecular profiles). In this study, we combined a filtration-based CTC isolation technology with prostate cancer cell-specific antibodies to identify the prostate cancer CTCs. After identification, we performed the 3D telomere profiling prior to laser microdissection for whole genome sequencing of singles CTCs from 11 localized high-risk prostate cancer patient samples (36 CTCs in total). We identified unique and common somatic single nucleotide variants (SSNVs) and copy number alterations (CNAs) that can be used to predict high-risk lethal prostate cancer and treatment response in patients with clinically localized high- risk prostate cancers. The distribution of telomeres intensities show a range between 0 to 190.000 (a.u) and telomeres parameters were similar as described previously for high risk patients by our group. We analyzed 36 CTCs from 11 different patient samples and compared with normal lymphocytes from the same patient using whole genome sequencing (WGS). We identified 184 common mutated genes in all CTCs and two mutated genes with the same base pair (bp) mutation (PMS2 and EIF2AK4). We also identified 503 consensus mutations found in all patient but not in all CTCs and 1728 unique mutations found in all or any CTCs of only one patient, showing the heterogeneity that arises in single cells. The list of genes affected are all involved in potential pathways for tumor development such as immune system, signal transduction, homeostasis, gene expression, chromatin organization, DNA repair, cell cycle, programmed cell death, cell-cell communication and others. MCPH1, also known as BRIT1, is a key regulatory gene of the DNA damage response pathway and was mutated in all CTCs from prostate patients. MCPH1 deficiency promotes genomic instability and increases cancer susceptibility. The PMS2 gene is a member of a set of genes known as the mismatch repair (MMR) genes which promotes also genomic Instability. Those mutations can be closely linked with the genomic instability we found assessing the telomere profiles. The CDH17 gene is a member of the cadherin superfamily and in contrast to classic cadherins, such as E- and N-cadherin, LI-cadherin possess biological functions distinct from classic cadherins and reduced expression of LI-cadherin is closely associated with tumor progression and lymph node metastasis of human colorectal carcinoma. These mutations were also found in all CTCs. All this information is needed for individualized therapy and predict drug resistance, phenotypic transition which is heterogeneous from cell to cell from patient to patient. Citation Format: Aline Rangel Pozzo. Whole genome sequencing of single circulating tumor cells from localized high-risk prostate cancer patients: Laser microdissection and 3Dtelomere profile [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3535.