EGF-containing Fibulin-like Extracellular Matrix Protein Research Articles

Proteo-genomic analyses in relatively lean Chinese adults identify proteins and pathways that affect general and central adiposity levels

Adiposity is an established risk factor for multiple diseases, but the causal relationships of different adiposity types with circulating protein biomarkers have not been systematically investigated. We examine the causal associations of general and central adiposity with 2923 plasma proteins among 3977 Chinese adults (mean BMI = 23.9 kg/m²). Genetically-predicted body mass index (BMI), body fat percentage (BF%), waist circumference (WC), and waist-to-hip ratio (WHR) are significantly (FDR < 0.05) associated with 399, 239, 436, and 283 proteins, respectively, with 80 proteins associated with all four and 275 with only one adiposity trait. WHR is associated with the most proteins (n = 90) after adjusting for other adiposity traits. These associations are largely replicated in Europeans (mean BMI = 27.4 kg/m²). Two-sample Mendelian randomisation (MR) analyses in East Asians using cis-protein quantitative trait locus (cis-pQTLs) identified in GWAS find 30/2 proteins significantly affect levels of BMI/WC, respectively, with 10 showing evidence of colocalisation, and seven (inter-alpha-trypsin inhibitor heavy chain H3, complement factor B, EGF-containing fibulin-like extracellular matrix protein 1, thioredoxin domain-containing protein 15, alpha-2-antiplasmin, fibronectin, mimecan) are replicated in separate MR using different cis-pQTLs identified in Europeans. These findings identified potential novel mechanisms and targets, to our knowledge, for improved treatment and prevention of obesity and associated diseases.

A New Ocular Phenotype Combining Juvenile Glaucoma and Doyne Honeycomb Retinal Dystrophy (Malattia Leventinese) due to a Novel EFEMP1 Pathogenic Variant.

Doyne honeycomb retinal dystrophy (DHRD), also termed malattia leventinese (MLVT), is a dominantly inherited ocular disease characterized by the progressive accumulation of macular and peripapillary drusenoid material beneath the retinal pigment epithelium in the Bruch membrane. In all affected individuals genetically characterized to date, DHRD/MLVT is caused by a single heterozygous p.Arg345Trp missense variant in the EGF-containing fibulin-like extracellular matrix protein 1, EFEMP1. Recently, pathogenic variants in the EFEMP1 gene have also been demonstrated in several families with juvenile or adult-onset hereditary isolated glaucoma. Here, we describe a family featuring a unique phenotype of juvenile glaucoma and DHRD/MLVT caused by a novel EFEMP1 variant. Our results expand both the ocular phenotype associated with EFEMP1 variants and the molecular spectrum causing DHRD by describing the first non-p.Arg345Trp EFEMP1 pathogenic allele.

The extracellular matrix protein EFEMP2 is involved in the response to VHSV infection in the olive flounder Paralichthys olivaceus

The EGF-containing fibulin-like extracellular matrix protein 2 (EFEMP2) is involved in connective tissue development, elastic fiber formation, and tumor growth. In this study, we characterized the cDNA of EFEMP2 (PoEFEMP2), a member of the fibulin family of ECM proteins, in the olive flounder Paralichthys olivaceus. The coding region of PoEFEMP2 encodes a protein that contains six calcium-binding EGF-like (EGF-CA) domains and four complement Clr-like EGF-like (cEGF) domains. PoEFEMP2 shows 67.51–96.77 % similarities to orthologs in a variety of fish species. PoEFEMP2 mRNA was detected in all tissues examined; the highest levels of PoEFEMP2 mRNA expression were observed in the heart, testis, ovary and muscle. The PoEFEMP2 mRNA level increases during early development. In addition, the PoEFEMP2 mRNA level increased at 3 h post-infection (hpi) and decreased from 6 to 48 hpi in flounder Hirame natural embryo (HINAE) cells infected with viral hemorrhagic septicemia virus (VHSV). Disruption of PoEFEMP2 using the clustered regularly interspaced short palindromic repeats/CRISPR-associated-9 (CRISPR/Cas9) system resulted in a significant upregulation of VHSV G mRNA levels and immune-related genes expression in knockout cells. These findings implicate PoEFEMP2 in antiviral responses in P. olivaceus.

Small peptide CSF fingerprint of amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease characterized by abnormal protein aggregation in the motor neurons. Present and earlier proteomic studies to characterize peptides in cerebrospinal fluid (CSF) associated with motoneuron pathology did not target low molecular weight proteins and peptides. We hypothesized that specific changes in CSF peptides or low molecular weight proteins are significantly altered in ALS, and that these changes may support deciphering molecular pathophysiology and even guide approaches towards therapeutic interventions. Cerebrospinal fluid (CSF) from 50 ALS patients and 50 non-ALS controls was collected, centrifuged immediately after collection, aliquoted into polypropylene test tubes, frozen within 30-40 min after the puncture, and stored at -80°C until use. Peptides were sequenced using capillary electrophoresis or liquid chromatography/mass spectrometry (CE-MS/MS or LC-MS/MS). In the CSF of 50 patients and 50 non-ALS controls 33 peptides were found, of which 14 could be sequenced using a non-lytic single-pot proteomic detection method, CE/MS. ALS deregulated peptides vs. controls included Integral membrane protein 2B, Neurosecretory protein VGF, Osteopontin, Neuroendocrine protein 7B2 (Secretogranin-V), EGF-containing fibulin-like extracellular matrix protein 1, Xylosyltransferase 1 XT-1, Chromogranin-A, Superoxide dismutase SOD-1, Secretogranin-1 (Chromogranin B), NR2F2 Nuclear Receptor Subfamily 2 Group F Member 2 and Collagen alpha-1(VII) chain. Most striking deregulations in CSF from ALS patients were found in VGF, Osteopontin, SOD-1 and EFEMP1 peptides. No associations of disease severity, duration and region of onset with sequenced peptides were found.

Recurrent Small Bowel Obstruction and Perforation as a Complication of EFEMP1-Associated Systemic Venous Amyloidosis

Abstract Introduction/Objective Amyloidogenic protein EGF-containing fibulin-like extracellular matrix protein 1 (EFEMP1) is a newly described age-related systemic venous amyloidosis. EFEMP1 venous deposition has recently been reported in postmortem gastrointestinal specimens in which the deceased presented with gastrointestinal bleeding. Methods/Case Report We report a case of a 53-year-old female with long-standing seronegative spondylitis, and multiple recurrent small bowel obstructions and perforations status post surgical interventions who presented with 3 days of generalized abdominal pain, 10/10 in severity with nausea, emesis and absence of bowel movement. Abdominal CT scan confirmed closed loop obstruction in the left hemi-abdomen. An emergent partial duodenectomy was performed with extensive lysis of adhesion which revealed a perforated jejunal diverticulum with abscess formation. Microscopic examination demonstrated the presence of eosinophilic, amorphous, acellular material in the venules of the duodenum wall. Congo red stain demonstrating the presence of amyloid deposits and the specimen was sent for amyloid typing. Liquid chromatography tandem mass spectrometry demonstrated a peptide profile consisting of serum amyloid P, apolipoprotein A4, apolipoprotein E, and abundant peptides associated with EFEMP1/FBLN3 consistent with a recently described age-related venous amyloidosis derived from EFEMP1 protein. Results (if a Case Study enter NA) N/A. Conclusion EFEMP1-associated systemic venous amyloidosis has a predilection for females with a median age of 75 years and almost exclusively occurs in the gastrointestinal tract. We report this case for its novelty, unique clinical presentation and to emphasize awareness of this entity as we learn more about its etiology, presentation, management, and prognostication.

The Diagnostic and Prognostic Significance of EFEMP1 in Breast Cancer: An Immunohistochemistry Study.

EGF-containing fibulin-like extracellular matrix protein 1 (EFEMP1) has been associated to a variety of malignancies. Because EFEMP1 can act as both a tumor suppressor and an oncogene, this study aimed to evaluate the expression of EFEMP1 at mRNA and protein in breast cancer and to ascertain the diagnostic and prognostic value of EFEMP1 in relation to clinical features of breast cancer. Several bioinformatics websites such as GEPIA and Oncomine databases were used to analyze the mRNA level of EFEMP1. Immunohistochemistry assay was used to detect EFEMP1 immunoexpression using tissue microarray (TMA) and clinical breast cancer samples. EFEMP1 was shown to be overexpressed in breast cancer in some study cohorts while being low expressed in others. In TMA, 86 patients (39.1%) with a high H-score and 134 patients (60.9%) with a low H-score had EFEMP1 positive for breast cancer. While HER2 breast cancer and normal breast tissues had the lowest expression of EFEMP1, it was shown to be highly expressed in Luminal B, A, and TNBC. EFEMP1 H-score is associated with tumor stage and indicates poor overall survival in breast cancer. EFEMP1 H-score was high in the clinical tumor tissues compared with adjacent normal tissue (n = 20), therefore, it would to be a sensitive biomarker for breast cancer. EFEMP1 is a key indicator for assessing the clinical prognosis and diagnosis of patients with breast cancer, as evidenced by the higher expression of EFEMP1 in tumor tissue compared to normal tissue and its association with poor overall survival.

TAILS Identifies Candidate Substrates and Biomarkers of ADAMTS7, a Therapeutic Protease Target in Coronary Artery Disease

Loss-of-function mutations in the secreted enzyme ADAMTS7 (a disintegrin and metalloproteinase with thrombospondin motifs 7) are associated with protection for coronary artery disease. ADAMTS7 catalytic inhibition has been proposed as a therapeutic strategy for treating coronary artery disease; however, the lack of an endogenous substrate has hindered the development of activity-based biomarkers. To identify ADAMTS7 extracellular substrates and their cleavage sites relevant to vascular disease, we used TAILS (terminal amine isotopic labeling of substrates), a method for identifying protease-generated neo–N termini. We compared the secreted proteome of vascular smooth muscle and endothelial cells expressing either full-length mouse ADAMTS7 WT, catalytic mutant ADAMTS7 E373Q, or a control luciferase adenovirus. Significantly enriched N-terminal cleavage sites in ADAMTS7 WT samples were compared to the negative control conditions and filtered for stringency, resulting in catalogs of high confidence candidate ADAMTS7 cleavage sites from our three independent TAILS experiments. Within the overlap of these discovery sets, we identified 24 unique cleavage sites from 16 protein substrates, including cleavage sites in EFEMP1 (EGF-containing fibulin-like extracellular matrix protein 1/Fibulin-3). The ADAMTS7 TAILS preference for EFEMP1 cleavage at the amino acids 123.124 over the adjacent 124.125 site was validated using both endogenous EFEMP1 and purified EFEMP1 in a binary in vitro cleavage assay. Collectively, our TAILS discovery experiments have uncovered hundreds of potential substrates and cleavage sites to explore disease-related biological substrates and facilitate activity-based ADAMTS7 biomarker development.

Upregulated Fibulin-1 Increased Endometrial Stromal Cell Viability and Migration by Repressing EFEMP1-Dependent Ferroptosis in Endometriosis.

Endometriosis (EMS) is a prevalent disease in women characterized by the presence of endometrial stroma and glands outside the uterus. Recent studies have showed that EMS is correlated with the resistance of endometrial stromal cells (ESCs) to ferroptosis, an iron-dependent nonapoptotic cell death. Fibulin-1 (FBLN1) is a newly identified target regulated by progesterone in the process of ESC decidualization. However, the role of FBLN1 in regulating ESC ferroptosis and EMS remains unclear. In the present study, the gene expression profiles of GSE58178, GSE23339, and GSE25628 were downloaded from the Gene Expression Omnibus (GEO) database, and the commonly differential genes were identified using Venn diagram analysis. The role of FBLN1 in ESC viability and migration was evaluated using Cell Counting Kit-8, transwell, and western blot analysis. We found that the FBLN1 expression was increased significantly in eutopic and ectopic endometrial tissues of patients with EMS compared with normal endometrium. FBLN1 overexpression in normal ESCs (NESCs) promoted cell viability and migration, whereas FBLN1 inhibition in ectopic ESCs (EESCs) decreased cell viability and migration. Furthermore, FBLN1 inhibition facilitated EESC death by triggering ferroptosis, as evidenced by increased Fe2+, lipid ROS, and malondialdehyde (MDA) level and decreased glutathione peroxidase 4 (GPX4) expression and glutathione (GSH) level. Mechanistically, FBLN1 interacted with EGF-containing fibulin-like extracellular matrix protein 1 (EFEMP1) and increased the protein stability of EFEMP1. More importantly, EFEMP1 silencing repressed the effect of FBLN1 on regulating EESC ferroptosis, death, and migration. Taken together, these results verify the role of the FBLN1/EFEMP1/ferroptosis pathway in the pathogenesis of EMS, and silencing of FBLN1/EFEMP1 might be an effective approach to treat EMS.

EGF‐containing fibulin‐like extracellular matrix protein 1 (EFEMP1) amyloid deposits in the lower rectum from aged patient with bloody stools

EGF‐containing fibulin‐like extracellular matrix protein 1 (EFEMP1) amyloid deposits in the lower rectum from aged patient with bloody stools

The novel form of amyloidosis derived from EGF-containing fibulin-like extracellular matrix protein 1 (EFEMP1) preferentially affects the lower gastrointestinal tract of elderly femalesa.

To characterise the clinicopathological features of amyloidosis due to EGF-containing fibulin-like extracellular matrix protein 1 (EFEMP1), a newly described amyloid type. We identified cases by searching the Mayo Clinic amyloid liquid chromatography and tandem mass spectrometry typing database for specimens with the universal amyloid signature proteins, abundant EFEMP1 spectra and absence of other specific amyloid precursor proteins. We also developed an immunohistochemical stain for EFEMP1 applicable to formalin-fixed tissue sections and performed electron microscopy in one case. We identified 33 specimens from 32 patients with EFEMP1 amyloid. Most patients were female (91%) with a mean age of 75years, and most specimens (94%) were from the bowel. EFEMP1 amyloid was incidentally identified in specimens biopsied/resected for a variety of clinical indications. In bowel specimens, EFEMP1 amyloid involved blood vessels and interstitium of the lamina propria, submucosa and/or muscularis propria. Although the EFEMP1 deposits were weakly to moderately Congo red-positive with absent to weak birefringence, they were strongly positive for EFEMP1 by immunohistochemistry, had the characteristic fibrillar ultrastructure of amyloid and were readily identified by mass spectrometry. EFEMP1 amyloid is a recently described novel amyloid type that predominantly affects the bowel of elderly females. Because EFEMP1 amyloid is only weakly Congo red-positive, it may be overlooked without a high index of suspicion. However, its characteristic microanatomical distribution is highlighted by immunohistochemistry and its identity is readily confirmed by mass spectrometry. Based on its distinctive features, we propose that EFEMP1 amyloidosis be considered a new amyloid type.

EFEMP2 Inhibits Breast Cancer Invasion And Metastasis In Vitro And In Vivo.

BackgroundEGF-containing fibulin-like extracellular matrix protein 2 (EFEMP2) is an extracellular matrix (ECM) glycoprotein, which is regarded as potential prognostic biomarkers in some carcinoma. Little is known about the association of EFEMP2 and breast cancer.MethodsEFEMP2 expressions in normal breast tissue, benign fibroadenoma, breast cancer, the normal mammary epithelial cell line, and 4 different invasive breast cancer cell lines were evaluated by immunohistochemistry (IHC) or immunocytochemistry (ICC) and real time quantitative reverse transcriptase-polymerase chain reaction (RT-qPCR). Expression and prognostic value of EFEMP2 in breast cancer were verified by the Public databases (Oncomine and Kaplan-Meier plotter database). Lentiviral vector with EFEMP2 cDNA was constructed and used to infect breast cancer cell lines to investigate the effects of EFEMP2 on the biological behavior of breast cancer cells by functional in vitro and in vivo assays.ResultsDown-regulated EFEMP2 expression was found in breast cancer tissues and cells, and low expression of EFEMP2 was associated with poor prognosis in patients with breast cancer. Analysis by the Public database leaded to the same conclusion. Up-regulated EFEMP2 expression significantly hampered the invasion and metastasis abilities of breast cancer cells and the process of epithelial interstitial transformation (EMT) via the Wnt/β-catenin pathway.ConclusionEFEMP2 expression was lower in breast cancer and closely related to the prognosis of patients, its anti-oncogenic roles indicated the underlying therapeutic target for the future treatment of breast cancer.

The anti-cancer effect of ZR30 protein via targeting extracellular signal proteins of different cell subpopulations of glioma

Objective: To investigate the roles and anti-cancer mechanism of artificially synthesized EGF-containing fibulin-like extracellular matrix protein (EFEMP1) derived tumor suppressor ZR30 protein in glioma (GBM). Methods: ZR30 protein were in vitro expressed using a wheat germ cell-free system. GBM cell lines (U251, U251NS, and U87) were cultured for 2-3 days in the presence or absence of ZR30 treatment. MMP-2 level was detected by gelatin zymography assay, moreover, the expression of EGFR, Notch-1 and p-Akt/Akt levels were determined by western blot. Additionally, MTT assay was used to measure ZR30's effect on the cell proliferation of U251 and U251NS cells. Furthermore, pre-mixed U251-GFP and U251NS-RFP cells (1∶9) were injected into the brain of nude mice, and then ZR30 or PBS was injected into the intra-tumor after 10 and 21 days, respectively. Then DNA was extracted from the right brain of nude mice in each group. Comparative quantitative polymerase chain reaction (CQ-PCR) was used to examine the copy numbers of human gene hSPAG16, mouse gene mSpag16, GFP and RFP. The survival status of each group of nude mice was also observed. Results: The levels of activated MMP-2 in U87 and U251 cells were lower after 10, 50 and 100 ng/ml ZR30 treatment for 2-3 days. Western blot analysis showed that ZR30 treatment reduced the expression of EGFR, Notch-1 and p-Akt/Akt in U251 cells, and inhibited Notch-1 and p-Akt/Akt expression in U251NS cells, and then decreased the response of U251 cells to EGF stimulation. Moreover, ZR30 inhibited the cell proliferation of U251 and U251NS two days after exposure. The in vivo orthotopic GBM xenografts were successfully constructed. CQ-PCR results indicated that the hSPAG16/mSpag16 ratios of mice in PBS group and ZR30 treatment groups at 180, 700, and 1 800 ng dosages were 3.67±2.82, 1.18±0.97, 1.75±1.55 and 1.38±1.17, respectively, and ZR30 treatment groups showed significantly lower ratios than the PBS group (P<0.05 for all). Correspondingly, the ratios of GFP/RFP in each group were 1.97±0.80, 1.97±0.85, 1.48±0.71 and 1.73±0.77, respectively, showing no statistical significance (P>0.05 for all). When treatment was performed 10 d after cell implantation, and the median survival time of mice in PBS group and ZR30 group was 40.5 days and 59.0 days, respectively. When treatment was performed 21 d after cell implantation, the median survival time of mice in PBS group and ZR30 group was extended to 57.0 days and 74.5 days, respectively. The median survival time of ZR30 treatment groups significantly prolonged (P<0.05 for all). Conclusions: ZR30 inhibits in vitro cell growth, invasion, angiogenesis and stemness maintenance in glioma via suppressing activated MMP-2, EGFR, p-Akt/Akt and Notch-1 proteins. In vivo, ZR30 markedly increased survival of mice harboring glioma xenografts, even for only one intra-tumoral injection at the time of early tumor formation. Overall, the in vivo and in vitro experiments supported the therapeutic potential of ZR30 for GBM.

A dual-type L2 11–88 peptide from HPV types 16/18 induced strong immune responses and broad spectrum cross-reactive antibodies in mice

The fusion between human tumorigenic cells and normal human diploid fibroblasts results in non-tumorigenic hybrid cells, suggesting a dominant role for tumor suppressor genes in the generated hybrid cells. After long-term cultivation in vitro, tumorigenic segregants may arise. The loss of tumor suppressor genes on chromosome 11q13 has been postulated to be involved in the induction of the tumorigenic phenotype of human papillomavirus (HPV)18-positive cervical carcinoma cells and their derived tumorigenic hybrid cells after subcutaneous injection in immunocompromised mice. The aim of this study was the identification of novel cellular genes that may contribute to the suppression of the tumorigenic phenotype of non-tumorigenic hybrid cells in vivo. We used cDNA microarray technology to identify differentially expressed cellular genes in tumorigenic HPV18-positive hybrid and parental HeLa cells compared to non-tumorigenic HPV18-positive hybrid cells. We detected several as yet unknown cellular genes that play a role in cell differentiation, cell cycle progression, cell−cell communication, metastasis formation, angiogenesis, antigen presentation, and immune response. Apart from the known differentially expressed genes on 11q13 (e.g., phosphofurin acidic cluster sorting protein 1 (PACS1) and FOS ligand 1 (FOSL1 or Fra-1)), we detected novel differentially expressed cellular genes located within the tumor suppressor gene region (e.g., EGF-containing fibulin-like extracellular matrix protein 2 (EFEMP2) and leucine rich repeat containing 32 (LRRC32) (also known as glycoprotein-A repetitions predominant (GARP)) that may have potential tumor suppressor functions in this model system of non-tumorigenic and tumorigenic HeLa x fibroblast hybrid cells.

EFEMP2 Mediates GALNT14-Dependent Breast Cancer Cell Invasion

N-Acetylgalactosaminyltransferase-14 (GALNT14) is a member of acetylgalactosaminyltransferases family. We have shown that GALNT14 could promote breast cancer cell invasion. However, the underlying molecular mechanism is unclear. Here, using yeast two hybrid, we find that EGF-containing fibulin-like extracellular matrix protein 2 (EFEMP2) interacts with GALNT14. Both in vitro and in vivo binding assays show that EFEMP2 is associated with GALNT14. Moreover, we find that GALNT14 mediates glycosylation of EFEMP2. EFEMP2 significantly increased the invasion ability of breast cancer cells including MCF-7 and MBA-MD-231 cells, and this phenomenon is suppressed by knockdown expression of GALNT14. In addition, the GALNT14-dependent O-glycosylation of EFEMP-2 regulates the stability of EFEMP-2 protein in breast cancer cells. Taken together, our results demonstrate a novel molecular mechanism underlying breast cancer invasion.

Electrochemical detection of methylated DNA on a microfluidic chip with nanoelectrokinetic pre-concentration

DNA methylation is considered to be a promising marker for the early diagnosis and prognosis of cancer. However, direct detection of the methylated DNAs in clinically relevant samples is still challenging because of its extremely low concentration (~fM). Here, an integrated microfluidic chip is reported, which is capable of pre-concentrating the methylated DNAs using ion concentration polarization (ICP) and electrochemically detecting the pre-concentrated DNAs on a single chip. The proposed chip is the first demonstration of an electrochemical detection of both level and concentration of the methylated DNAs by integrating a DNA pre-concentration unit without gene amplification. Using the proposed chip, 500 fM to 500 nM of methylated DNAs is pre-concentrated by almost 100-fold in 10 min, resulting in a drastic improvement of the electrochemical detection threshold down to the fM level. The proposed chip is able to measure not only the DNA concentration, but also the level of methylation using human urine sample by performing a consecutive electrochemical sensing on a chip. For clinical application, the level as well as the concentration of methylation of glutathione-S transferase-P1 (GSTP1) and EGF-containing fibulin-like extracellular matrix protein 1 (EFEMP1), which are known to be closely associated with prostate cancer diagnosis, are electrochemically detected in human urine spiked with these genes. The developed chip shows a limit of detection (LoD) of 7.9 pM for GSTP1 and 11.8 pM for EFEMP1 and is able to detect the level of methylation in a wide range from 10% to 100% with the concentration variation from 50 pM to 500 nM.

Human fibulin-3 protein variant expresses anti-cancer effects in the malignant glioma extracellular compartment in intracranial xenograft models.

BackgroundDecades of cytotoxic and more recently immunotherapy treatments for malignant glioma have had limited success due to dynamic intra-tumoral heterogeneity. The dynamic interplay of cancer cell subpopulations has been found to be under the control of proteins in the cancer microenvironment. EGF-containing fibulin-like extracellular matrix protein (EFEMP1) (also fibulin-3) has the multiple functions of suppressing cancer growth and angiogenesis, while promoting cancer cell invasion. EFEMP1-derived tumor suppressor protein (ETSP) retains EFEMP1’s anti-growth and anti-angiogenic functions while actually inhibiting cancer cell invasion.MethodsIn this study, we examined the therapeutic effect on glioblastoma multiforme (GBM) of an in vitro synthesized protein, ZR30, which is based on the sequence of ETSP, excluding the signaling peptide.ResultsZR30 showed the same effects as ETSP in blocking EGFR/NOTCH/AKT signaling pathways, when applied to cultures of multiple GBM cell lines and primary cultures. ZR30’s inhibition of MMP2 activation was shown not only for GBM cells, but also for other types of cancer cells having overexpression of MMP2. A significant improvement in survival of mice with orthotopic human GBM xenografts was observed after a single, intra-tumoral injection of ZR30. Using a model mimicking the intra-tumoral heterogeneity of GBM with cell subpopulations carrying different invasive and proliferative phenotypes, we demonstrated an equal and simultaneous tumor suppressive effect of ZR30 on both tumor cell subpopulations, with suppression of FOXM1 and activation of SEMA3B expressions in the xenografts.ConclusionOverall, the data support a complementary pleiotrophic therapeutic effect of ZR30 acting in the extracellular compartment of GBM.

Evaluating EFEMP1 in Cerebrospinal Fluid and Serum as a Potential Diagnosis Biomarker for Meningiomas.

The aim of this study was to determine the cerebrospinal fluid (CSF) and serum levels of EGF containing fibulin-like extracellular matrix protein 1 (EFEMP1) in the patients with meningiomas and explore its potential as a biomarker. Forty-five patients with meningioma, 11 of whom underwent meningioma resection, as well as 30 healthy controls were enrolled in this study. CSF and blood samples were collected preoperatively and postoperatively. Expression levels of EFEMP1 were measured by enzyme-linked immunosorbent assay (ELISA). Receiver operating characteristic (ROC) curves were used to evaluate its discriminant ability. CSF EFEMP1 levels were significantly higher in the CSF samples (p < 0.0001) and serum samples (p = 0.0056) of meningioma patients compared to controls. To distinguish meningioma patients from controls by CSF and serum EFEMP1 levels, ROC/AUC analysis indicated an AUC of 0.945 (sensitivity 0.933; specificity 0.833) and an AUC of 0.674 (sensitivity 0.867; specificity 0.400), respectively. Moreover, the postoperative CSF levels of EFEMP1 were significantly decreased compared to the preoperative levels (p < 0.0001). The present study suggested that elevated EFEMP1 levels might be a novel diagnostic biomarker for meningioma patients.

Weaponizing human EGF-containing fibulin-like extracellular matrix protein 1 (EFEMP1) for 21st century cancer therapeutics.

De-regulated EFEMP1 gene expression in solid tumors has been widely reported with conflicting roles. We dissected EFEMP1 to identify domains responsible for its cell context-dependent dual functions, with the goal being to construct an EFEMP1-derived tumor-suppressor protein (ETSP) that lacked tumor-promoting function. Exon/intron boundaries of EFEMP1 were used as boundaries of functional modules in constructing EFEMP1 variants, with removal of various module(s), and/or mutating an amino acid residue to convert a weak integrin binding-site into a strong one. A series of in vitro assays on cancerous features, and subcutaneous and intracranial xenograft-formation assays, were carried out for effects from overexpression of wild-type and variant forms of EFEMP1 in two glioma subpopulations characterized as tumor mass-forming cells (TMCs) or stem-like tumor initiating cells (STICs), where EFEMP1 showed cellcontext- dependent dual functions. One of the EFEMP1 variants was identified as the sought-after ETSP, which had a stronger tumor-suppression function in TMCs by targeting EGFR and angiogenesis, and a new tumor-suppression function in STICs by targeting NOTCH signaling and MMP2-mediated cell invasion. Therefore, ETSP may form the basis for further important research to develop a novel cancer therapy to treat many types of cancer by its tumor suppressor effect in the extracellular matrix compartment.

Overexpression of SPARC correlates with poor prognosis in patients with cervical carcinoma and regulates cancer cell epithelial-mesenchymal transition.

Secreted protein acidic and rich in cysteine (SPARC) is associated with the progression of numerous types of cancer. However, the role of SPARC in the progression of cervical cancer has not yet been adequately elucidated. In the current study, reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and immunohistochemistry were employed to evaluate the mRNA and protein expression of SPARC in normal cervical tissue, cervical intraepithelial neoplasia (CIN) and cervical cancer. In addition, three epithelial-mesenchymal transition (EMT) markers (E-cadherin, N-cadherin and vimentin) were detected by immunohistochemistry in the same specimens, and an enzyme-linked immunosorbent assay was conducted to detect the serum levels of SPARC in patients with cervical neoplasia. In highly invasive subclones of human cervical carcinoma cells, HeLa-1 and SiHa-1, lentiviral transfections were performed and RT-qPCR and western blot were used to investigate the effects of downregulated EGF-containing fibulin-like extracellular matrix protein 1 on the expression of E-cadherin, N-cadherin and vimentin. The results revealed that, in cervical carcinoma tissue, SPARC expression was significantly upregulated in a manner that positively correlated with N-cadherin and vimentin expression, and negatively correlated with E-cadherin expression. SPARC overexpression and high serum levels were significantly associated with the progression of cervical cancer and adverse prognosis of cervical cancer patients. Downregulation of SPARC can markedly reduce the expression of N-cadherin and vimentin and increase the expression of E-cadherin. Thus, overexpression of SPARC is significantly associated with poor prognostic clinicopathological characteristics in cervical carcinoma, and may be important in EMT. The results of the current study suggest that SPARC may be a potential therapeutic option for individuals diagnosed with cervical carcinoma.

Association between genetic variants of EGF-containing fibulin-like extracellular matrix protein1 gene and sporadic breast cancer in a Chinese Han population.

Genetic susceptibility of breast cancer has been shown to be modulated by inheritance of polymorphic genes. EGF-containing fibulin-like extracellular matrix protein 1 (EFEMP1) gene played an important role in many tumors, including lung cancer, hepatic carcinoma, and prostate cancer. In addition, it was importantly downexpressed in breast cancer. The present research aimed to assess the association between genetic variations of EFEMP1 and breast cancer risk. The authors genotyped 11 common tagging SNPs with an array platform including 960 cases and 972 cancer-free controls of Chinese women, according to the HapMap database based on the pairwise linkage disequilibrium (LD) r² threshold of 0.8, minor allele frequency of 0.05. Three SNPs were significant associated with breast cancer (rs3791679, p = 0.016, OR = 1.21, 95% CI = 1.04-1.41; rs1346786, p = 0.005, OR = 1.31, 95% CI = 1.08 -1.59; rs727878, p = 0.002, OR = 1.29, 95%CI = 1.10-1.51). Multivariate logistic regression analysis revealed that, compared with wild-type carriers in a dominant model, a significantly increased breast cancer risk was associated with the three identified risk SNPs. Among the selected tagging SNPs, three haplotype blocks were identified, and the results of haplotype analysis were consistent with the single-locus analysis. The haplotype 'GG' in block 1 and haplotype 'AG' in block 2 were significantly associated with breast cancer, and had a 54% and 28% increased breast cancer risk respectively, compared with their corresponding noncarriers. The present results suggested that the polymorphisms of EFEMP1 gene were associated with breast cancer and might contribute to the susceptibility of the progression of breast cancer in Chinese Han women. Individuals with the risk alleles might increase the risk of breast cancer.