Marker For Metastasis Research Articles

Arginine regulates HSPA5/BiP translation through ribosome pausing in triple-negative breast cancer cells

BackgroundTriple-negative breast cancer (TNBC) is an aggressive subtype of breast cancer with a high mortality rate due to a lack of therapeutic targets. Many TNBC cells are reliant on extracellular arginine for survival and express high levels of binding immunoglobin protein (BiP), a marker of metastasis and endoplasmic reticulum (ER) stress response.MethodsIn this study, the effect of arginine shortage on BiP expression in the TNBC cell line MDA-MB-231 was evaluated. Two stable cell lines were generated in MDA-MB-231 cells: the first expressed wild-type BiP, and the second expressed a mutated BiP free of the two arginine pause-site codons, CCU and CGU, termed G-BiP.ResultsThe results showed that arginine shortage induced a non-canonical ER stress response by inhibiting BiP translation via ribosome pausing. Overexpression of G-BiP in MDA-MB-231 cells promoted cell resistance to arginine shortage compared to cells overexpressing wild-type BiP. Additionally, limiting arginine led to decreased levels of the spliced XBP1 in the G-BiP overexpressing cells, potentially contributing to their improved survival compared to the parental WT BiP overexpressing cells.ConclusionIn conclusion, these findings suggest that the downregulation of BiP disrupts proteostasis during arginine shortage-induced non-canonical ER stress and plays a key role in cell growth inhibition, indicating BiP as a target of codon-specific ribosome pausing upon arginine shortage.

Thyroglobulin immunoassay with a fully automated pretreatment process provides accurate thyroglobulin values in anti-thyroglobulin antibody positive specimens

ObjectivesHuman thyroglobulin (Tg) is widely used as a tumor marker for recurrence and metastasis of differentiated thyroid cancer (DTC). Currently, serum Tg values are measured using second-generation sandwich immunoassays (2nd-IMA). However, interference by endogenous autoantibodies to thyroglobulin (TgAbs) can lead to false-negative results or falsely low Tg values. Here, we describe a new Tg assay using the immunoassay for total antigen including complex via pretreatment (iTACT) method to prevent TgAb interference and compare it with 2nd-IMA. MethodsTg values were evaluated by three assays: iTACT Tg, Elecsys Tg-II, which is a 2nd-IMA, and LC-MS/MS (Liquid chromatography tandem-mass spectrometry). The ratio of Tg values between each assay was then compared to the Tg value by LC-MS/MS and TgAb titer. Tg immunoreactivity was analyzed by size-exclusion chromatography. ResultsCorrelation between iTACT Tg and LC-MS/MS using TgAb-positive specimens was good: Passing–Bablok regression with iTACT Tg = 1.084 × LC-MS/MS + 0.831. Correlation between 2nd-IMA and LC-MS/MS showed a relatively lower slope: 2nd-IMA = 0.747 × LC-MS/MS − 0.518. Thus, Tg values determined by iTACT Tg are equivalent to those of LC-MS/MS regardless of TgAb titer, whereas 2nd-IMA gave lower Tg values due to TgAb interference. Tg-TgAb complexes of various molecular weights were verified by size-exclusion chromatography. Tg values measured by 2nd-IMA fluctuated depending on the molecular weight of the Tg-TgAb complexes, whereas iTACT Tg accurately quantified Tg values regardless of the size of the Tg-TgAb complexes. ConclusionTg values in TgAb-positive specimens were accurately determined by iTACT Tg. TgAb-positive specimens contain Tg-TgAb complexes of various molecular weights that interfere with Tg value determination by 2nd-IMA, whereas iTACT Tg is unaffected by the presence of Tg-TgAb complexes.

ABCA1 transporter promotes the motility of human melanoma cells by modulating their plasma membrane organization

BackgroundMelanoma is one of the most aggressive and deadliest skin tumor. Cholesterol content in melanoma cells is elevated, and a portion of it accumulates into lipid rafts. Therefore, the plasma membrane cholesterol and its lateral organization might be directly linked with tumor development. ATP Binding Cassette A1 (ABCA1) transporter modulates physico-chemical properties of the plasma membrane by modifying cholesterol distribution. Several studies linked the activity of the transporter with a different outcome of tumor progression depending on which type. However, no direct link between human melanoma progression and ABCA1 activity has been reported yet.MethodsAn immunohistochemical study on the ABCA1 level in 110 patients-derived melanoma tumors was performed to investigate the potential association of the transporter with melanoma stage of progression and prognosis. Furthermore, proliferation, migration and invasion assays, extracellular-matrix degradation assay, immunochemistry on proteins involved in migration processes and a combination of biophysical microscopy analysis of the plasma membrane organization of Hs294T human melanoma wild type, control (scrambled), ABCA1 Knockout (ABCA1 KO) and ABCA1 chemically inactivated cells were used to study the impact of ABCA1 activity on human melanoma metastasis processes.ResultsThe immunohistochemical analysis of clinical samples showed that high level of ABCA1 transporter in human melanoma is associated with a poor prognosis. Depletion or inhibition of ABCA1 impacts invasion capacities of aggressive melanoma cells. Loss of ABCA1 activity partially prevented cellular motility by affecting active focal adhesions formation via blocking clustering of phosphorylated focal adhesion kinases and active integrin β3. Moreover, ABCA1 activity regulated the lateral organization of the plasma membrane in melanoma cells. Disrupting this organization, by increasing the content of cholesterol, also blocked active focal adhesion formation.ConclusionHuman melanoma cells reorganize their plasma membrane cholesterol content and organization via ABCA1 activity to promote motility processes and aggressiveness potential. Therefore, ABCA1 may contribute to tumor progression and poor prognosis, suggesting ABCA1 to be a potential metastatic marker in melanoma.

The clinicopathological and prognostic significances of EZH2 expression in urological cancers: A meta‑analysis and bioinformatics analysis.

The Drosophila zeste enhancer homolog 2 gene (enhancer of zeste homolog 2; EZH2) is an important member of the polycomb group (PcG) gene family, which maintains the homologous gene via chromosome modification during embryonic development. EZH2 is overexpressed in various tumors, is closely related to tumor formation and growth, and has a malignant phenotype that promotes tumor cell proliferation, proliferation and metastasis. In the present study, a meta- and bioinformatic analysis was performed using data from multiple online databases until August 30, 2022. EZH2 upregulation was found in kidney, bladder and prostate cancers. EZH2 expression was negatively related to TNM staging and pathological grade in kidney and prostate cancers (P<0.05), as well as invasion depth and pathological grade in bladder cancer. According to the KM-plotter database, EZH2 expression was inversely associated with poor overall survival in patients with kidney clear cell renal cell carcinoma (RCC) and papillary RCC and with favorable survival in bladder cancer. EZH2 expression was negatively related to relapse-free survival in kidney papillary RCC and bladder cancer but positively associated with kidney clear cell RCC. According to GEPIA and UALCAN databases, EZH2 expression was higher in tumor tissue than normal tissue. The TIMER database showed that EZH2 was closely associated with the proportion of seven immune cell infiltrates in kidney, bladder, and prostate cancers. High EZH2 expression may be a potential marker of tumorigenesis and metastasis in patients with urological cancers.

Mutational profiling of true single circulating tumor cells (sCTC) in metastatic lung cancer to determine genomic heterogeneity and drug-resistance signatures.

e21024 Background: Omics of single Circulating tumor cells (CTCs) is highly envisioned though challenging. . Genotyping of TRUE sCTC capture and omics data can determine the molecular heterogeneity of a single cell without the stringent need for deep sequencing compared to tissue or blood (1/100th fold compared to ctDNA). Here, we show the utility of sCTC genome profiling to determine inter and intra-patient mutational heterogeneity and compare the outcome with matched ensembled ctDNA mutational profiles. Methods: OncoRADAR an antibody-affinity-based platform was used to capture live sCTCs without cell fixing. 50 sCTCs (90 %) and clusters (10 %) were isolated from 20 metastatic lung cancer pts for mutational genotyping. Amplified whole genomes were subjected to targeted hybrid-capture comprehensive genome profiling. Libraries prepared using a custom-designed comprehensive gene panel OncoIndx, targeting 600 cancer-relevant exons including MSI and HRR pathway genes, were sequenced in paired-end mode on the Illumina platform. Variant calling was performed using an in-house developed bioinformatics pipeline. Pathway enrichment analysis was performed manually using data enriched from different gene pathway databases. Results: 81 mutations were detected in 56 genes across all sCTCs with pathogenic, likely pathogenic, and unknown significance. The mutational landscape included SNVs (38.6%), splice variants (10%), frameshift mutations (0.8%), and truncations (50.4%). From a subset of patients (10 %), multiple sCTCs showed PTEN variant cooccurring with ESR1 and MET. This suggest a clonal selection and emergence of subclones during progression. Pathways related to drug resistance, migration, deregulated cell cycle, and signaling were significantly enriched in multiple sCTCs when compared to plasma genotyping. Low average TMB (1.89), LOH (3.9) and HRD (14.67%), and high MSI (28.68%) scores were observed. In contrast, matching ctDNA had higher average LOH (18.67), HRD (45.08%), and lower MSI (0.99) scores. Multiple sCTCs from a small fraction of patients had comparable values for genome stability indicators, suggesting maintained genome stability. Genotyping of sCTC among patients showed a high variability compared to comprehensive analysis at the ctDNA level. Conclusions: Targeted gene profiling of true sCTCs revealed multiple variants that remained undetected in ctDNA, enabling a better understanding of tumor heterogeneity. Analysis of multiple sCTCs from the same patient underscores the importance of clonal evolution and plasticity during metastasis. Longitudinal monitoring of sCTCs may reveal personalized markers for disease prevalence, metastasis and relapse. sCTC genotyping may help design personalized therapy with better precision.

An epigenetic signature of advanced colorectal cancer metastasis

Colorectal cancer (CRC) is a leading cause of morbidity and mortality worldwide. The majority of CRC deaths are caused by tumor metastasis, even following treatment. There is strong evidence for epigenetic changes, such as DNA methylation, accompanying CRC metastasis and poorer patient survival. Earlier detection and a better understanding of molecular drivers for CRC metastasis are of critical clinical importance. Here, we identify a signature of advanced CRC metastasis by performing whole genome-scale DNA methylation and full transcriptome analyses of paired primary cancers and liver metastases from CRC patients. We observed striking methylation differences between primary and metastatic pairs. A subset of loci showed coordinated methylation-expression changes, suggesting these are potentially epigenetic drivers that control the expression of critical genes in the metastatic cascade. The identification of CRC epigenomic markers of metastasis has the potential to enable better outcome prediction and lead to the discovery of new therapeutic targets.

Bifunctional electrochemical biosensor based on PB-MXene films for the real-time analysis and detection of living cancer cells

Circulating tumor cells (CTCs) are important prognostic markers for cancer diagnosis and metastasis, and their detection is an important means to detect cancer metastasis. Herein, we construct a novel bifunctional electrochemical biosensor based on the PB-MXene composite films. A simple electrostatic self-assembly approach was employed to prepare a film composed of PB nanocubes on the MXene substrates. Given that the PB is an artificial peroxidase for H2O2 sensing, the PB-MXene films can realize the real-time monitoring of H2O2 secretion from living CTCs. Besides, the anti-CEA attached biosensors can be utilized to quantify the corresponding CTCs. The synergic effects of the MXene with a large specific area and PB with enzyme-free catalysis for H2O2 resulted in PB-MXene films exhibiting high electrocatalytic and low cytotoxicity for both H2O2 sensing and living CTCs capturing. As a result, the biosensor shows a low detection limit of 0.57 μM towards H2O2 with a wide linear range (1 μM to 500 μM), as well as an excellent sensing performance for CTCs (an extremely low detection limit of 9 cells/mL in a wide linear range of 1.3 ×101 to 1.3 ×106 cells/mL). Moreover, the prepared biosensor showed satisfactory stability and anti-interference ability for potential applications in clinical cancer diagnosis and tumor metastasis.

FMO family may serve as novel marker and potential therapeutic target for the peritoneal metastasis in gastric cancer.

To explore the relationship between flavin-containing monooxygenases (FMOs) and peritoneal metastasis (PM) in gastric cancer (GC). TIMER 2.0 was used to perform pan-cancer analysis and assess the correlation between the expression of FMOs and cancers. A dataset from The Cancer Genome Atlas (TCGA) was used to analyze the correlation between FMOs and clinicopathological features of GC. PM is well established as the most common mode of metastasis in GC. To further analyze the correlation between FMOs and PM of GC, a dataset was obtained from the National Center for Biotechnology Information Gene Expression Omnibus (GEO) database. The results were validated by immunohistochemistry. The relationship between FMOs and PM of GC was explored, and a novel PM risk signature was constructed by least absolute shrinkage and selection operator (LASSO) regression analysis. The regression model's validity was tested by multisampling. A nomogram was established based on the model for predicting PM in GC patients. The mechanism of FMOs in GC patients presenting with PM was assessed by conducting Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses in TCGA and GEO datasets. Finally, the potential relationship between FMOs and immunotherapy was analyzed. The pan-cancer analysis in TCGA and GEO datasets showed that FMO1 was upregulated, while FMO2 and FMO4 were downregulated in GC. Moreover, FMO1 and FMO2 correlated positively with the T and N stage of GC in the TCGA dataset. FMO1 and FMO2 expression was a risk factor for GC (hazard ratio: 1.112 and 1.185). The overexpression of FMO1 was significantly correlated with worse disease-free-survival (DFS) and overall survival (OS). However, no relationship was found between FMO2 expression in GC and DFS and OS. PM was highly prevalent among GC patients and typically associated with a worse prognosis. FMO1 was highly expressed in GC with PM. FMO1 and FMO2 were positively correlated with PM in GC. We identified a 12-gene panel for predicting the PM risk signature by LASSO (Area Under Curve (AUC) = 0.948, 95%CI: 0.896-1.000). A 10-gene panel for PM prediction was identified (AUC = 0.932, 95%CI: 0.874-0.990), comprising FMO1 and FMO2. To establish a model for clinical application, a 7-gene panel was established (AUC = 0.927, 95% CI: 0.877-0.977) and successfully validated by multisampling. (AUC = 0.892, 95% CI: 0.878-0.906). GO and KEGG analyses suggest that FMO1 and FMO2 regulate the extracellular matrix and cell adhesion. FMO1 and FMO2 were positively correlated with the immune score of GC, and their expression was associated with the infiltration of immune cells. PM in GC is strongly correlated with FMOs. Overall, FMO1 and FMO2 have huge prospects for application as novel diagnostic and therapeutic targets.

Apelin receptor antagonist boosts dendritic cell vaccine efficacy in controlling angiogenic, metastatic and apoptotic-related factors in 4T1 breast tumor-bearing mice.

Apelin/APJ axis plays a critical role in cancer progression, thus its targeting inhibits tumor growth. However, blocking of Apelin/APJ axis in combination with immunotherapeutic approaches may be more effective. This study aimed to investigate the effects of APJ antagonist ML221 in combination with a DC vaccine on angiogenic, metastatic and apoptotic-related factors in a breast cancer (BC) model. Four groups of female BALB/c mice with 4T1-induced BC were treated with PBS, APJ antagonist ML221, DC vaccine, and "ML221 + DC vaccine". After completion of the treatment, the mice were sacrificed and the serum levels of IL-9 and IL-35 as well as the mRNA expression of angiogenesis (including VEGF, FGF-2, and TGF-β), metastasis (including MMP-2, MMP-9, CXCR4) and apoptosis-related markers (Bcl-2, Bax, Caspase-3) in tumor tissues were determined using ELISA and real-time PCR, respectively. Angiogenesis was also evaluated by co-immunostaining of tumor tissues with CD31 and DAPI. Primary tumor metastasis to the liver was analyzed using hematoxylin-eosin staining. The efficiency of combination therapy with "ML221 + DC vaccine" was remarkably higher than single therapies in preventing liver metastasis compared to the control group. In comparison with the control group, combination therapy could significantly reduce the expression of MMP-2, MMP-9, CXCR4, VEGF, FGF-2, and TGF-β in tumor tissues (P < 0.05). It also decreased the serum level of IL-9 and IL-35 compared with the control group (P < 0.0001). Moreover, vascular density and vessel diameter were significantly reduced in the combination therapy group compared with the control group (P < 0.0001). Overall, our findings demonstrate that combination therapy using a blocker of the apelin/APJ axis and DC vaccine can be considered a promising therapeutic program in cancers.

Association of Serum Leptin With Body Mass Index in Gallbladder Cancer Patients: A Pilot Study.

Leptin has been proposed to be a link between obesity and the increased incidence of various cancers like breast cancer, colon cancer, gastric cancer, etc. The role of leptin in gallbladder cancer is largely undetermined. Moreover, no study has evaluated serum leptin levels and their correlation with clinicopathological characteristics and serum tumour markers in gallbladder cancer (GBC). Therefore, the present study was planned. A cross-sectional study was conducted in a tertiary care hospital in Northern India after obtaining ethical approval from the institution. Forty GBC patients staged as per American Joint Committee on Cancer (AJCC) 8th staging systemwere recruited along with 40 healthy controls. Serum leptin was assayed by sandwich enzyme-linked immunosorbent assay (ELISA) and tumour markers (CA19-9, CEA and CA125) by Chemiluminescence.ROC, Mann Whitney U test, Linear regression and Spearman correlation was performed using Statistical Product and Service Solutions (SPSS) (IBM SPSS Statistics for Windows, Version 25.0, Armonk, NY). BMI was also assessed for both groups. Median BMI for GBC patients was 19.46 (IQR 17.61-22.36). Median serum leptin levels were significantly lower (2.09 (IQR 1.01-7.76) ng/mL) in GBC patients as compared to controls (12.32 (IQR 10.50-14.72) ng/mL). AUC was 0.84 with 100% sensitivity and 75% specificity at 7.57 ng/mL.Serum leptin was not associated with cancer stage, resectability, metastasis, liver infiltration, or tumour markers on linear regression (p=0.74, adjusted R square = -0.07). A significant positive correlation was found between BMI and serum leptin in GBC patients (p=0.00). Lower BMI and relatively lean presentation of GBC patients may account for low serum leptin levels.

Quinacrine inhibits cMET-mediated metastasis and angiogenesis in breast cancer stem cells.

A trans-membrane receptor tyrosine kinase, cMET, belonging to the MET proto-oncogene family, is responsible for cancer metastasis and angiogenesis. But not much is known about the role of cMET in growth and progression of cancer stem cells (CSCs). Earlier studies have shown that Quinacrine (QC), a bioactive agent, has anti-CSCs activity. Here, the role of QC in deregulation of cMET-mediated metastasis and angiogenesis has been systematically evaluated in vitro in highly metastatic breast CSCs (mBCSCs), ex vivo in patient-derived breast cancer stem cells (PDBCSCs) and in vivo in xenograft mice model systems. Cell proliferation, migration, invasion and representative metastasis markers were upregulated in cMET-overexpressed cells and QC exposure inhibited these processes in both mBCSCs and PDBCSCs. Interestingly, metastasis was significantly inhibited by QC in cMET-overexpressed cells but comparatively lesser significant alteration of the process was noted in cMET-silenced cells. Increase in vascularization (in in ovo CAM assay), and cell-cell tube formation (in HUVECs), and enhanced MMP9 and MMP2 enzymatic activities (in gelatin zymography) were noted after cMET overexpression but these processes got reversed after cMET knockdown or QC treatment in cMET-overexpressed cells. QC inhibited angiogenesis significantly in cMET-overexpressed cells, but lesser significant change was observed in cMET-silenced cells. Reduction in tumor volume and decreased expression of metastatic and angiogenic markers were also noted in xenograft mice after QC treatment. Furthermore, QC inhibited cMET activity by dephosphorylation of its tyrosine residues (Y1234 and Y1356) and downregulation of its downstream cascade. Thus, QC inhibited the cMET-mediated metastasis and angiogenesis in in vitro, in ovo, in vivo and ex vivo model systems. Ligand (HGF) binding leads to receptor dimerization and phosphorylation of tyrosine kinase domain of cMET. This activates the cMET signaling cascade. The representative downstream metastasis and angiogenesis-related proteins get upregulated and induce the metastasis and angiogenesis process. But after the QC treatment, cMET get dephosphorylated and inactivated. As a result, the downstream signaling proteins of cMET along with the other representative metastatic and angiogenic factors get downregulated. These lead to inhibition of cMET-mediated metastasis and angiogenesis. (Created with BioRender.com).

MBD2 facilitates tumor metastasis by mitigating DDB2 expression

Despite past extensive studies, the pathoetiologies underlying tumor metastasis remain poorly understood, which renders its treatment largely unsuccessful. The methyl-CpG-binding domain 2 (MBD2), a “reader” to interpret DNA methylome-encoded information, has been noted to be involved in the development of certain types of tumors, while its exact impact on tumor metastasis remains elusive. Herein we demonstrated that patients with LUAD metastasis were highly correlated with enhanced MBD2 expression. Therefore, knockdown of MBD2 significantly attenuated the migration and invasion of LUAD cells (A549 and H1975 cell lines) coupled with attenuated epithelial–mesenchymal transition (EMT). Moreover, similar results were observed in other types of tumor cells (B16F10). Mechanistically, MBD2 selectively bound to the methylated CpG DNA within the DDB2 promoter, by which MBD2 repressed DDB2 expression to promote tumor metastasis. As a result, administration of MBD2 siRNA-loaded liposomes remarkably suppressed EMT along with attenuated tumor metastasis in the B16F10 tumor-bearing mice. Collectively, our study indicates that MBD2 could be a promising prognostic marker for tumor metastasis, while administration of MBD2 siRNA-loaded liposomes could be a viable therapeutic approach against tumor metastasis in clinical settings.

CORRELATION OF PLASMA D-DIMER LEVEL IN FEMALE BREAST CARCINOMA PATIENTS WITH CLINICAL STAGES AND HISTOPATHOLOGICAL PARAMETERS

Background: Breast cancer is one of the leading disease among females in the world. There is a correlation between cancer and hypercoagulation. In a cancer state, there is an increase in the level of cross-linked brin degradation product (d-dimer) which indicates systemic activation of brinolysis and hemostasis. So, there is a relation between an increase in d-dimer value and breast carcinoma.Objective:This study was done in an attempt to evaluate the role of D-dimer in patients with carcinoma breast, in predicting lymph node metastasis in carcinoma patients and to look for the relationship of these markers with histopathologic parameters Method: A prospective observational descriptive study was done in Dr. S N medical college attached hospitals (department of surgery) from Jan 2022 to Jan 2023. Forty-three breast carcinoma patients were included in the study after core needle biopsy conrmation. Plasma d-dimer levels compared with (tumor size, stage, grade, lymphovascular invasion, and lymph node involvement). Data collected and analysed. Results: Forty three patients were studied with age from 21 to 76yrs(mean age-51).left side tumor was more common 52%. D-dimer level was increasing with enlarged tumor size, high stage, high grade, lymphovascular invasion, and lymph node involvement. All are statstacally signicant.(p value: &lt;0.001) Conclusions: Plasma D-dimer level is important preoperative marker with which we can predict the advance stage breast cancer. It is also an important marker of clinical stage progression, lymphovascular invasion, and metastasis . Pre-operative plasma D-dimer level is a safe, cost-effective, and convenient method for the prediction of advanced-stage breast cancer

Clinicopathological and prognostic significance of SPARC expression in gastric cancer: A meta‑analysis and bioinformatics analysis.

Secreted protein acidic and rich in cysteine (SPARC) is a member of the extracellular matrix glycoprotein family that binds to calcium ions. It may bind to a variety of proteins in the extracellular matrix and also compete with cell membrane surface receptors for growth. In the present study, the relationship between SPARC expression in gastric cancer tissues and the clinicopathological characteristics and prognosis of patients with gastric cancer were systematically evaluated. A meta-analysis and bioinformatics analysis were performed using the PubMed, Chinese National Knowledge Infrastructure, Kaplan-Meier (KM)-plotter, The Cancer Genome Atlas (TCGA), Gene Expression Profiling Interactive Analysis (GEPIA), University of ALabama at Birmingham CANcer (UALCAN), Human Protein Atlas (HPA) and Timer databases. SPARC was mainly expressed in tumor mesenchymal cells. The meta-analysis indicated that SPARC expression was higher in gastric cancer tissues than in normal tissues. SPARC was associated with the degree of differentiation and distant metastasis. K-M plotter results indicated that high SPARC expression was negatively associated with overall survival, post-progression survival and progression-free survival rates of patients. According to the Oncomine, GEPIA, UALCAN and HPA databases, SPARC mRNA and protein expression was upregulated in gastric cancer vs. normal tissues and was negatively associated with poor patient prognosis. In the TCGA database, univariate analysis indicated that lymph node metastasis and distant metastasis were associated with the prognosis of patients with gastric cancer. Cox multifactorial analysis suggested that high SPARC expression, age and distant metastasis were important factors affecting the survival time of patients with gastric cancer. Analysis with the Timer database indicated that SPARC was closely associated with the proportion of 7 immune-cell infiltrates in gastric cancer. These findings indicated that high expression of SPARC may be a potential marker of tumorigenesis and metastasis in patients with gastric cancer.

DEC1 is a potential marker of early metastasis in Oral squamous cell carcinoma

ObjectiveThis study aimed to investigate the role of differentiated embryonic-chondrocyte expressed gene 1 (DEC1) in early oral squamous cell carcinoma (OSCC) metastasis. Methods and materialsThis study collected normal oral mucosas (NOM) and OSCC tissues from Xiangya Hospital for immunohistochemistry to detect the expressions of DEC1 and epithelial-mesenchymal transition (EMT) -related molecules. Correlation analysis between the expressions of the cytoplasmic DEC1 and EMT-related molecules was performed. Kaplan-Meier analysis was conducted to estimate Recurrence-free survival (RFS). After knocking down DEC1, cell migration and the expressions of EMT-related molecules were evaluated in HN6 cells by cell scratch assay, qRT-PCR, and western blot. ResultsImmunohistochemistry showed that the subcellular location of DEC1 expression was different between OSCC and NOM tissues. The cytoplasmic expression of DEC1 in OSCC tissues was significantly higher than in NOM tissues, and its expression was highest in early OSCC patients with metastasis. In addition, the cytoplasmic DEC1 was negatively correlated with the E-cadherin and β-catenin, but positively correlated with the N-cadherin in OSCC and NOM tissues. In vitro assays showed that DEC1 knockdown inhibited cell migration and EMT in HN6 cells. ConclusionDEC1 could serve as a potential predictive marker for early OSCC metastasis.

Intratumoral platelet microthrombi in oral squamous cell carcinoma: A marker of lymph node metastasis.

A hypercoagulable state exists in patients with oral squamous cell carcinoma (OSCC), but the role of platelets in the tumour microenvironment has not been explored. This study revealed the status of intratumoral plateletmicrothrombi (PLT-MT) and their clinicopathological relevance and predictive value in OSCC. This study retrospectively evaluated 106 OSCC patients. Tumour and tumour-adjacent tissue specimens were used to stain PLT-MT. Clinicopathological information, patient follow-ups and outcomes and preoperative coagulation and inflammatory hematologic indicators were collected, and their correlation with PLT-MT was analysed. Intratumoral PLT-MT was present in 35 of 106 patients with OSCC who had higher preoperative D-dimer, CRP, FIB and PT levels and lower TT levels. PLT-MT was an independent correlative factor of lymph node metastasis and suggested worse OS in N0 patients. Intratumoral PLT-MT was found in OSCC and was correlated with a hypercoagulable inflammatory state. PLT-MT was an independent marker of lymph node metastasis and showed potential in prognosis prediction.

Abstract 3618: The role of endothelial ACKR1 in triple-negative breast cancer metastasis

Abstract Metastasis is the most common cause of breast cancer mortality. Of the breast cancer subtypes, triple-negative breast cancer (TNBC) is the deadliest due to its increased likelihood to metastasize. Tumor cell extravasation is a critical step of metastasis and allows circulating tumor cells to exit the vasculature and seed distant tissues. Clear understanding of the major regulators of tumor cell extravasation will provide insights into the progression of TNBC metastasis. One potential regulator of TNBC cell extravasation is ACKR1. In many contexts, ACKR1 expression is required in endothelial cells (EC) for leukocyte extravasation. Endothelial ACKR1 binds CXCL2, a promigratory chemokine, and localizes it to EC junctions to guide neutrophils through leukocyte extravasation. CXCL2 expression in TNBC cells is also necessary for tumor cell extravasation from lung microvasculature and for tumor metastasis. Our preliminary data show that ACKR1 is required in at least one stromal cell type for TNBC metastasis from the primary tumor to the lung. These data suggest that endothelial ACKR1-CXCL2 interactions may mediate tumor cell extravasation in TNBC metastasis. Therefore, we hypothesize that endothelial ACKR1 promotes metastasis via retention of CXCL2 at EC junctions, increasing tumor cell chemotaxis and extravasation. To address this hypothesis, we will examine the in vivo significance of endothelial ACKR1 expression using our validated ACKR1 endothelial cell-specific knockout mouse model. We will test the requirement for endothelial ACKR1 for metastasis of orthotopically implanted TNBC tumors to distant sites in the lung and for extravasation of circulating tumor cells into lung tissue. We will determine which steps of extravasation require ACKR1 by evaluating ACKR1-low and ACKR1-overexpressing ECs using an Ibidi flow co-culture system that recapitulates the shear stress conditions of pulmonary microvasculature. We will examine whether these steps are dependent on CXCL2 by introducing CXCL2-neutralizing antibodies to the Ibidi flow system and observing their effects on each extravasation step. Our proposed studies will establish the role of endothelial ACKR1 in TNBC metastatic progression and determine the specific steps of tumor cell extravasation in which endothelial ACKR1 and CXCL2 function. Understanding these processes may guide development of ACKR1 as a prognostic marker for metastasis and can provide mechanistic insight into candidate chemokine and chemokine receptor inhibitors under evaluation for treatment of breast cancer. Citation Format: Samuel Tanner Roach, Chinwe Ewenighi-Amankwah, J Dufraine, L.A. Naiche, Jan K. Kitajewski. The role of endothelial ACKR1 in triple-negative breast cancer metastasis. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3618.

Abstract 2487: Metastasis: Cancer cells that have turned on a Yamanaka-like pluripotency program

Abstract Purpose: To develop therapeutics to treat metastasis of solid tumor cancers. To overcome obstacles to developing anti-metastasis treatments by: 1) developing an animal model that reproducibly mimics cancer metastasis and does so within a reasonable timeframe; and 2) figuring out the basic science that drives metastasis. Experimental: We discovered that all pluripotent human stem cells express a MUC1 cleavage product called MUC1* (muk 1 star). All MUC1 cleavage stops with the onset of differentiation and MUC1 goes back to its quiescent state. Over 75% of solid tumor cancers also express MUC1*. Overexpression of MUC1*, as well as enzymes that cleave MUC1 to MUC1* are predictors of poor prognosis. NME1 is a ligand of MUC1* that is secreted by both embryonic stem cells and cancer cells. As a dimer, NME1 dimerizes the MUC1* extra cellular domain to activate the MAP kinase growth pathway. In an embryo, the more stem cells there are, the more NME1 is secreted and it goes from the active dimer to an inactive hexamer (doesn’t bind MUC1*). The paradox was, “How do stem cells limit self-replication, but cancer cells do not?” Answer: A primitive growth factor, NME7AB, looks like a single chain dimer of NME1, so is always active, is expressed in the earliest, naïve stem cells and in metastatic cancer cells. Unpublished Results: Growing cancer cells in recombinant NME7AB for 10 days transforms them into metastatic cells: 1) become non-adherent; 2) enter dormancy; 3) upregulate metastatic markers by 200-fold; 4) form sub-cu tumors in mice from as few as 50 cells; iv injection leads to total metastasis in 10 days; 5) the percent of injected tumor cells that were first grown in NME7AB determines the growth of the entire tumor. We developed a monoclonal antibody that blocks interaction of NME7AB and MUC1*. This antibody reverses established metastasis of breast cancers in animals. NME7AB should be turned off early in embryogenesis, but is aberrantly re-activated in metastatic cancers. Yamanaka factors OCT4, SOX2 and NANOG bind to the promoters of NME7, MUC1 and MMP16, an enzyme that cleaves MUC1 to MUC1* and exposes the cryptic binding site for NME7AB. Conclusions: Primitive growth factor, NME7AB, drives metastasis of solid tumor cancers. Antibody that disrupts NME7AB-MUC1* interaction reverses metastasis in animals. Citation Format: Cynthia C. Bamdad, Benoit J. Smagghe, Mark G. Carter, Trevor J. Grant, Laura M. Reale, Michael J. Nash, Danica M. Walkley, Jac-Leen S. Nash, Kevin R. Yi, Andrew K. Stewart. Metastasis: Cancer cells that have turned on a Yamanaka-like pluripotency program [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2487.

Engineered Macrophages-Based uPA-Scavenger Load Gemcitabine to Prompt Robust Treating Cancer Metastasis.

The majority of cancer patients die of metastasis rather than primary tumors, and most patients may have already completed the cryptic metastatic process at the time of diagnosis, making them intractable for therapeutic intervention. The urokinase-type plasminogen activator (uPA) system is proved to drive cancer metastasis. However, current blocking agents such as uPA inhibitors or antibodies are far from satisfactory due to poor pharmacokinetics and especially have to face multiplex mechanisms of metastasis. Herein, an effective strategy is proposed to develop a uPA-scavenger macrophage (uPAR-MΦ), followed by loading chemotherapeutics with nanoparticles (GEM@PLGA) to confront cancer metastasis. Interestingly, significant elimination of uPA by uPAR-MΦ is demonstrated by transwell analysis on tumor cells in vitro and enzyme-linked immunosorbent assay detection in peripheral blood of mice with metastatic tumors, contributing to significant inhibition of migration of tumor cells and occurrence of metastatic tumor lesions in mice. Moreover, uPAR-MΦ loaded with GEM@PLGA shows a robust antimetastasis effect and significantly prolonged survival in 4T1-tumor-bearing mice models. This work provides a novel living drug platform for realizing a potent treatment strategy to patients suffering from cancer metastasis, which can be further expanded to handle other tumor metastasis markers mediating cancer metastasis.

Genistein and coumestrol reduce MCF-7 breast cancer cell viability and inhibit markers of preferential metastasis, bone matrix attachment and tumor-induced osteoclastogenesis

The propensity of breast cancer to preferentially metastasize to the skeleton is well known. Once established in bone metastatic breast cancers have a poor prognosis due to their ability to promote extensive bone loss which augments tumor burden. Unfortunately, current anti-resorptive therapies for skeletal metastasis are typically prescribed after secondary tumors have formed and are palliative in nature. One group of compounds with the potential to reduce both tumor burden and osteolysis are phytoestrogens (PE), but the mechanisms mediating a beneficial effect are unclear. Therefore, the current study examined the effect of genistein and coumestrol alone or in combination on breast cancer cell number, expression of mediators of preferential skeletal metastasis, bone matrix attachment and tumor-induced osteoclast formation. Results showed that genistein and coumestrol significantly reduced viable cell number in an estrogen receptor dependent manner (p < 0.05), whereas combinations of PE had no effect. In addition, genistein and coumestrol significantly reduced expression of genes driving epithelial to mesenchymal transition (snail), bone attachment (CXCR4 and integrin αV) and osteolysis (PTHrP and TNF-α). In keeping with this genistein and coumestrol significantly suppressed attachment of breast cancer cells to bone matrix and inhibited tumor and RANKL-induced osteoclast formation. Our data suggests that phytoestrogens not only decrease breast cancer cell viability but also antagonize essential tumor bone interactions that establish and drive the progression of skeletal metastasis.