Expression Of COL17A1 Research Articles

Glomerular Endothelial Cell Receptor Adhesion G-Protein-Coupled Receptor F5 (ADGRF5) and the Integrity of the Glomerular Filtration Barrier.

Background: Glomerular endothelial cells are recognized to be important for maintaining the glomerular filtration barrier. ADGRF5, an adhesion G protein-coupled receptor, has been suggested to be involved in endothelial cell function. However, the role of ADGRF5 in the glomerular filtration barrier integrity remains elusive. Methods: Cellular expression of ADGRF5 in mouse glomerulus was determined by histological analyses. The impact of ADGRF5 deletion on the glomerular morphology, kidney function, and glomerular endothelial gene/protein expression was then analyzed using ADGRF5 knockout (Adgrf5 −/−) mice and human primary glomerular endothelial cells. Results: ADGRF5 was specifically expressed in the capillary endothelial cells within the glomerulus. Adgrf5 −/− mice developed albuminuria and impaired kidney function with morphological defects in the glomeruli, namely glomerular hypertrophy, glomerular basement membrane splitting and thickening, diaphragmed fenestration and detachment of the glomerular endothelial cells, and mesangial interposition. These defects were accompanied by the altered expression of genes responsible for glomerular basement membrane organization (type IV collagens and laminins) and Krüppel-like factor 2 (Klf2) in glomerular endothelial cells. Moreover, ADGRF5 knockdown decreased COL4A3 and COL4A4 expression and increased KLF2 expression in human primary glomerular endothelial cells. Conclusions: The loss of ADGRF5 resulted in altered gene expression in glomerular endothelial cells, and perturbed the structure and permselectivity of the glomerular filtration barrier.

Genetic reprogramming with stem cells regenerates glomerular epithelial podocytes in Alport syndrome.

Glomerular filtration relies on the type IV collagen (ColIV) network of the glomerular basement membrane, namely, in the triple helical molecules containing the α3, α4, and α5 chains of ColIV. Loss of function mutations in the genes encoding these chains (Col4a3, Col4a4, and Col4a5) is associated with the loss of renal function observed in Alport syndrome (AS). Precise understanding of the cellular basis for the patho-mechanism remains unknown and a specific therapy for this disease does not currently exist. Here, we generated a novel allele for the conditional deletion of Col4a3 in different glomerular cell types in mice. We found that podocytes specifically generate α3 chains in the developing glomerular basement membrane, and that its absence is sufficient to impair glomerular filtration as seen in AS. Next, we show that horizontal gene transfer, enhanced by TGFβ1 and using allogenic bone marrow-derived mesenchymal stem cells and induced pluripotent stem cells, rescues Col4a3 expression and revive kidney function in Col4a3-deficient AS mice. Our proof-of-concept study supports that horizontal gene transfer such as cell fusion enables cell-based therapy in Alport syndrome.

Strategy for the Optimization of Read-Through Therapy for Junctional Epidermolysis Bullosa with COL17A1 Nonsense Mutation

The read-through therapy suppresses premature termination codons and induces read-through activity consequently restoring missing proteins. Aminoglycosides are widely studied as read-through drugs in different human genetic disorders including hereditary skin diseases. Our previous work revealed that aminoglycosides have effect on COL17A1 nonsense mutations and represent a therapeutic option to alleviate disease severity. However, the amount of restored type XVII collagen (C17) in C17 deficient junctional epidermolysis bullosa (JEB-C17) keratinocytes was less than 1% relative to normal keratinocytes and was achieved only after high dose gentamicin treatment, which induced deep transcriptional changes. Therefore, in this study, we designed a strategy for the read-through therapy to challenge with aminoglycosides limitation factors in clinical use and the chronic inflammation in JEB-C17 patients. We developed TRID-C5 containing low dosage of aminoglycosides, CC-90009, NMDI-14, melatonin and apocynin that was able to induce about 20% of missing C17 without cell toxicity and effect on in vitro wound closure. TRID-C5 significantly induced COL17A1 expression and reverted the proinflammatory phenotype of JEB-C17 keratinocytes. Evaluation of this drug cocktail regarding its stability, penetration and efficacy as a topical treatment remains to be determined. TRID-C5 might represent an improved therapeutic strategy for JEB and for other genetic skin disorders.

Collagen XVII regulates tumor growth in pancreatic cancer through interaction with the tumor microenvironment.

Expression of the gene for collagen XVII (COL17A1) in tumor tissue is positively or negatively associated with patient survival depending on cancer type. High COL17A1 expression is thus a favorable prognostic marker for breast cancer but unfavorable for pancreatic cancer. This study explored the effects of COL17A1 expression on pancreatic tumor growth and their underlying mechanisms. Analysis of published single-cell RNA-sequencing data for human pancreatic cancer tissue revealed that COL17A1 was expressed predominantly in cancer cells rather than surrounding stromal cells. Forced expression of COL17A1 did not substantially affect the proliferation rate of the mouse pancreatic cancer cell lines KPC and AK4.4 in vitro. However, in mouse homograft tumor models in which KPC or AK4.4 cells were injected into syngeneic C57BL/6 or FVB mice, respectively, COL17A1 expression promoted or suppressed tumor growth, respectively, suggesting that the effect of COL17A1 on tumor growth was influenced by the tumor microenvironment. RNA-sequencing analysis of tumor tissue revealed effects of COL17A1 on gene expression profiles (including the expression of genes related to cell proliferation, the immune response, Wnt signaling, and Hippo signaling) that differed between C57BL/6-KPC and FVB-AK4.4 tumors. Our data thus suggest that COL17A1 promotes or suppresses cancer progression in a manner dependent on the interaction of tumor cells with the tumor microenvironment.

COL17A1 facilitates tumor growth and predicts poor prognosis in pancreatic cancer

ObjectiveThis study aimed to determine the role of COL17A1 in tumor progression and predict the prognosis of pancreatic cancer (PC). MethodsRNA-seq data from The Cancer Genome Atlas and Genotype-Tissue Expression were analyzed using bioinformatics methods. “Limma” package was used to screen differentially expressed genes (DEGs). Prognostic-associated data were further analyzed using univariate Cox regression and verified using the GSE28375 and GSE62452 datasets. Protein-protein interaction (PPI) network analysis was integrated to screen for hub genes. In vitro quantitative real-time PCR (qPCR) and western blotting were used to detect gene expression. The functional attributes of PC cells were verified by wound healing assays, migration and invasion assays, Cell Counting Kit 8 (CCK8), and 5-ethynyl-2′-deoxyuridine (EdU) assay. ResultsOn analyzing PC data, 4637 DEGs were identified. Of these, 2399 genes were upregulated and 2238 were downregulated. Through PPI network analysis, we identified that COL17A1 expression was highly correlated with poor prognosis of patients with PC. Functional attribute assays in the in vitro study showed that COL17A1 knockdown inhibited PC cell proliferation, migration, and invasion. ConclusionsAccording to our results, COL17A1 promotes PC cell proliferation, migration, and invasion mediated by the epithelial-mesenchymal transition (EMT) pathway. Thus, COL17A1 could be used as a prognostic marker in PC.

Integrated Analysis of ECT2 and COL17A1 as Potential Biomarkers for Pancreatic Cancer.

Background Pancreatic cancer (PC) is a malignant tumor of the digestive tract. It presents with atypical clinical symptoms and lacks specific diagnostic indicators. This study is aimed at exploring the potential biomarkers of PC. Methods TCGA database pancreatic cancer dataset was normalized and used to identify differentially expressed genes (DEGs). Survival, independent prognostic, and clinical correlation analyses were performed on DEGs to screen for key genes. DNA methylation, mutation, and copy number variation (CNV) analyses were used to analyze genetic variants in key genes. GSEA was performed to explore the functional enrichment of the key genes. Based on the expression of key genes, construction of a competing endogenous RNA (ceRNA) network, analysis of the tumor microenvironment (TME), and prediction of chemotherapeutic drug sensitivity were performed. Furthermore, the GEO database was used to validate the reliability of key genes. Results Two key genes (ECT2 and COL17A1) were identified, which were highly expressed in PC. The mRNA expression of ECT2 and COL17A1 was associated with DNA methylation and CNV. The cell cycle, proteasome, and pathways in cancer were enriched in the high-COL17A1 and ECT2 groups. The TME results showed that immune scores were decreased in the high-ECT2 group. CeRNA network results showed that there were eleven miRNAs were involved in the regulation of ECT2 and COL17A1. Moreover, pRRophetic analysis showed that 20 chemotherapeutic drugs were associated with ECT2 and COL17A1 expression. Conclusions Collectively, ECT2 and COL17A1 may be potential biomarkers for PC, providing a new direction for clinical diagnosis and treatment.

The CD44/COL17A1 pathway promotes the formation of multilayered, transformed epithelia

At the early stage of cancer development, oncogenic mutations often cause multilayered epithelial structures. However, the underlying molecular mechanism still remains enigmatic. By performing a series of screenings targeting plasma membrane proteins, we have found that collagen XVII (COL17A1) and CD44 accumulate in RasV12-, Src-, or ErbB2-transformed epithelial cells. In addition, the expression of COL17A1 and CD44 is also regulated by cell density and upon apical cell extrusion. We further demonstrate that the expression of COL17A1 and CD44 is profoundly upregulated at the upper layers of multilayered, transformed epithelia invitro and invivo. The accumulated COL17A1 and CD44 suppress mitochondrial membrane potential and reactive oxygen species (ROS) production. The diminished intracellular ROS level then promotes resistance against ferroptosis-mediated cell death upon cell extrusion, thereby positively regulating the formation of multilayered structures. To further understand the functional role of COL17A1, we performed comprehensive metabolome analysis and compared intracellular metabolites between RasV12 and COL17A1-knockout RasV12 cells. The data imply that COL17A1 regulates the metabolic pathway from the GABA shunt to mitochondrial complex I through succinate, thereby suppressing the ROS production. Moreover, we demonstrate that CD44 regulates membrane accumulation of COL17A1 in multilayered structures. These results suggest that CD44 and COL17A1 are crucial regulators for the clonal expansion of transformed cells within multilayered epithelia, thus being potential targets for early diagnosis and preventive treatment for precancerous lesions.

COL4A3 expression in asthmatic epithelium depends on intronic methylation and ZNF263 binding.

BackgroundReduction of COL4A3, one of the six isoforms of collagen 4, in asthmatic airways results in increased inflammation and angiogenesis, implicating it as a central part of asthma pathogenesis. However, to date, the path underlying these diminished COL4A3 levels has been elusive. This study investigated a possible mechanism underlying the reduction of COL4A3 expression.MethodsBronchial biopsies of 76 patients with asthma and 83 controls were subjected to RNA-sequencing and DNA methylation bead arrays to identify expression and methylation changes. The binding of ZNF263 was analysed by chromatin-immunoprecipitation sequencing coupled with quantitative (q)PCR. Effects of ZNF263 silencing, using small interfering RNA, on the COL4A3 expression were studied using qPCR.ResultsCOL4A3 expression was significantly reduced in bronchial biopsies compared to healthy controls, whereas DNA methylation levels at cg11797365 were increased. COL4A3 expression levels were significantly low in asthmatics without inhaled corticosteroid (ICS) use, whereas the expression was not statistically different between asthmatics using ICS and controls. Methylation levels at cg11797365 in vitro were increased upon consecutive rhinovirus infections.ConclusionOur data indicate an epigenetic modification as a contributing factor for the loss of COL4A3 expression in asthmatic airway epithelium.

Differential expression of COL4A3 and collagen in upward and downward progressing types of nasopharyngeal carcinoma.

Upward (local growth and invasion of the base of skull), downward (distant metastasis) and mixed progressing types of nasopharyngeal carcinoma (NPC) have been identified and are distinctly different with respect to clinical symptoms, therapeutic strategies and prognosis. The present study aimed to identify the genetic difference and collagen expression levels in the upward and downward progressing types of NPC. Whole exon sequencing (WES) was used to detect genes differentially mutated between the upward and downward progressing types of NPC. Collagen deposition in the upward and downward progressing types of NPC was determined using Masson trichromatic staining, while the protein expression level of COL4A3 was detected using immunohistochemistry. Survival analysis was also performed using the Kaplan-Meier Plotter database to examine the role of COL4A3 expression level in the prognosis of head and neck squamous cell carcinoma. Knockdown of COL4A3 was performed using short interfering (si)RNA-COL4A3 in a 5-8F NPC cell line. Reverse transcription-quantitative PCR and western blot analyses were utilized to analyze the mRNA and protein expression levels of COL4A3, respectively. The roles of COL4A3 in the migration and invasion of the 5-8F cell line were examined using wound-healing Transwell and Matrigel assays, respectively. A total of 21 genes were differentially mutated between the upward and downward progressing types of NPC. The COL4A3 was investigated further, as it was found to be associated with extracellular matrix deposition and cancer metastasis. The COL4A3 gene was markedly downregulated in the downward progressing type compared with that in the upward progressing type (2.161±1.306 vs. 5.077±3.619; P<0.05). In addition, the deposition of collagen in the downward progressing type was also significantly decreased compared with that in the upward progressing type (5.63±6.83 vs. 10.94±9.60; P<0.05). Kaplan-Meier analysis indicated that high expression level of COL4A3 was positively associated with a favorable prognosis of head and neck squamous cell carcinoma (HR, 0.69; 95% CI, 0.49- 0.97; P=0.031). To confirm the role of COL4A3, the expression level of COL4A3 was knocked down using siRNA in the 5-8F cell line and the results showed that the invasion and migration was significantly increased when the expression of COL4A3 was inhibited (P<0.0001). In conclusion, the gene mutation patterns were significantly different between the upward and downward progressing types of NPC. In addition, the expression level of the COL4A3 gene was decreased in the downward progressing type, which might promote NPC metastasis through the downregulation of extracellular collagen expression.

High Expression of COL17A1 Predicts Poor Prognosis and Promotes the Tumor Progression via NF-κB Pathway in Pancreatic Adenocarcinoma.

COL17A1 (collagen type XVII alpha 1 chain) is known to be upregulated and has a prognostic role in many malignancies, as well as contributing to cell proliferation, apoptosis, and invasion. However, little knowledge is available on the expression and prognostic value of COL17A1 in pancreatic adenocarcinoma (PDAC). In our study, we searched the public database and found that mRNA and protein levels of COL17A1 are commonly upregulated in PDAC tissues. The immunohistochemical analysis conducted by us revealed enhanced expression of COL17A1 protein in 169 PDAC samples compared with that in 67 adjacent normal tissues. We also observed a significantly positive correlation between COL17A1 expression and lymph node metastasis (p < 0.0001), TNM clinical stage (p < 0.0001), and pathology differentiation (p < 0.01). The KM-plot results indicated that PDAC patients with a high COL17A1 expression have a poorer overall survival (p < 0.001) than those with a low COL17A1 expression. The result of the Cox regression analysis of multivariate data suggested COL17A1 is an independent prognostic indicator of PDAC patients' overall survival. CCK-8, wound healing, and transwell assays suggested that COL17A1 knockdown markedly inhibited tumor proliferation and invasion in PDAC cells, and cells with COL17A1 overexpression had a prominently higher proliferative and invasive capacity. Knockdown of COL17A1 significantly upregulated the apoptosis rate. We deduce that upregulated COL17A1 activated the NF-κB pathway in PDAC cells. In summary, our studies showed the prognostic value of COL17A1 in PDAC and that COL17A1 may act as a molecular therapeutic target for PDAC treatment.

Corneal recurrent erosions dystrophies in Sweden

AbstractCorneal dystrophies are a heterogeneous group of genetic disorders. We aimed to identify genetic cause of a dominantly inherited Epithelial Recurrent Erosion Dystrophy (ERED)‐like disease common in Northern Sweden. Examinations and interviews were performed at the Ophthalmology Clinics of the University Hospital of Umeå and Skellefteå Hospital. A total of 20 patients from Västerbotten were extensively examined and blood samples for genetic analysis collected. Four unrelated families were subject to genealogical studies. Known genes causative of corneal dystrophies were excluded by array‐based allele specific primer extension. Haplotype assessment was done by array genotyping and identification of potentially causative genomic variants by whole exome sequencing (WES). The dystrophy common in Västerbotten has three phases; recurrent erosions during childhood, alleviated symptoms in the higher teens and finally, after the age 40, decreased visual acuity. Haplotype analysis and WES revealed a novel mutation, c.2816C&gt;T, p.T939I, in the COL17A1 gene coding collagen type XVII alpha1. It appeared to be a founder mutation that segregated with disease in a genealogically expanded pedigree dating back 200 years to a common ancestor, Theodor. Notably, the patients called the disease ‘Theodor's eyes’, but Theodor being unknown to them. Furthermore, COL17A1 expression in cornea was demonstrated by RT‐PCR and immunohistochemistry. We identified COL17A1 mutation as a cause of ERED, revealing a novel corneal disease in Sweden, and highlighting the necessity of COL17A1 screening in corneal dystrophies with erosions and no known genetic defect. Our genealogical analysis identified a common ancestor, Theodor, living 200 years ago.

Key genes associated with pancreatic cancer and their association with outcomes: A bioinformatics analysis.

Pancreatic cancer is a highly malignant neoplastic disease of the digestive system. In the present study, the dataset GSE62165 was downloaded from the Gene Expression Omnibus (GEO) database. GSE62165 contained the data of 118 pancreatic ductal adenocarcinoma samples (38 early‑stage tumors, 62 lymph node metastases and 18 advanced tumors) and 13 control samples. Differences in the expression levels of genes between normal tissues and early‑stage tumors were investigated. A total of 240 differentially expressed genes (DEGs) were identified using Rsoftware3.5 (137 upregulated genes and 103 downregulated genes). Then, the differentially expressed genes were subjected to Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analysis. The following 18 core genes were identified using Cytoscape, based on the protein‑interaction network of DEGs determined using the online tool STRING: EGF, ALB, COL17A1, FN1, TIMP1, PLAU, PLA2G1B, IGFBP3, PLAUR, VCAN, COL1A1, PNLIP, CTRL, PRSS3, COMP, CPB1, ITGA2 and CEL. The pathways of the core genes were primarily associated with pancreatic secretion, protein digestion and absorption, and focal adhesion. Finally, survival analyses of core genes in pancreatic cancer were conducted using the UALCAN online database. It was revealed that PLAU and COL17A1 were significantly associated with poor prognosis (P<0.05). The expression levels of genes in primary pancreatic cancer tissues were then compared; only one gene, COL17A1, was identified to be significantly differentially expressed. Finally, another dataset from GEO, GSE28735, was analyzed to verify the upregulated expression of COL17A1. Taken together, the results of the present study have indicated that the expression of COL17A1 gene may be associated with the occurrence and development of pancreatic cancer.

Stem cell competition orchestrates skin homeostasis and ageing

Stem cells underlie tissue homeostasis, but their dynamics during ageing-and the relevance of these dynamics to organ ageing-remain unknown. Here we report that the expression of the hemidesmosome component collagen XVII (COL17A1) by epidermal stem cells fluctuates physiologically through genomic/oxidative stress-induced proteolysis, and that the resulting differential expression of COL17A1 in individual stem cells generates a driving force for cell competition. In vivo clonal analysis in mice and in vitro 3D modelling show that clones that express high levels of COL17A1, which divide symmetrically, outcompete and eliminate adjacent stressed clones that express low levels of COL17A1, which divide asymmetrically. Stem cells with higher potential or quality are thus selected for homeostasis, but their eventual loss of COL17A1 limits their competition, thereby causing ageing. The resultant hemidesmosome fragility and stem cell delamination deplete adjacent melanocytes and fibroblasts to promote skin ageing. Conversely, the forced maintenance of COL17A1 rescues skin organ ageing, thereby indicating potential angles for anti-ageing therapeutic intervention.

Urine-derived podocytes-lineage cells: A promising tool for precision medicine in Alport Syndrome.

Alport Syndrome (ATS) is a rare genetic disorder caused by collagen IV genes mutations, leading to glomerular basement membrane damage up to end-stage renal disease. Podocytes, the main component of the glomerular structure, are the only cells able to produce all the three collagens IV alpha chains associated with ATS and thus, they are key players in ATS pathogenesis. However, podocytes-targeted therapeutic strategies have been hampered by the difficulty of non-invasively isolating them and transcripts-based diagnostic approaches are complicated by the inaccessibility of other COL4 chains-expressing cells. We firstly isolated podocyte-lineage cells from ATS patients' urine samples, in a non-invasive way. RT-PCR analysis revealed COL4A3, COL4A4, and COL4A5 expression. Transcripts analysis on RNA extracted from patient's urine derived podocyte-lineage cells allowed defining the pathogenic role of intronic variants, namely one mutation in COL4A3 (c.3882+5G>A), three mutations in COL4A4 (c.1623+2T>A, c.3699_3706+1del, c.2545+143T>A), and one mutation in COL4A5 (c.3454+2T>C). Therefore, our cellular model represents a novel tool, essential to unequivocally prove the effect of spliceogenic intronic variants on transcripts expressed exclusively at a glomerular level. This process is a key step for providing the patient with a definite molecular diagnosis and with a proper recurrence risk. The established system also opens up the possibility of testing personalized therapeutic approaches on disease-relevant cells.

205 Altering the splice pattern of COL17A1 with antisense oligonucleotides

Junctional epidermolysis bullosa (JEB) is a devastating disease of the skin and mucous membranes, characterized by blistering and erosions upon minor mechanical friction. Genotypically, mutations within the type XVII collagen gene underlie this phenotype. In this project, we targeted a cryptic splice site arising due to a mutation (c. 380-1 G>A) at the COL17A1 intron 6 / exon 7 junction. Utilization of this splice site leads to the loss of 16 nucleotides at the mRNA level. Subsequently, the reading frame is shifted so that a premature stop codon is generated within exon 9 and the mRNA is degraded by nonsense-mediated decay. Using three distinct antisense oligonucleotides (ASOs), we aimed to either restore the wild type splice pattern of COL17A1 or achieve skipping of exon 7, which would lead to a slightly truncated, but potentially functional type XVII collagen, since the reading frame remains intact. We transfected patient keratinocytes with either of the ASOs and analyzed overall COL17A1 expression, as well as the generation of alternative splicing products. Amplification and sequencing of the COL17A1 mRNA from exon 5 to exon 9 showed skipping of exon 7 as the most prominent result of ASO treatments in JEB keratinocytes. Additionally, alleles with a restored wild type splice pattern, as well as such showing concomitant skipping of exons 6 and 7, were identified. SqRT-PCR confirmed an increase in overall COL17A1 expression. Furthermore, full length (180 kDa) type XVII collagen was detected upon ASO treatment using Western blot analysis. Taken together, we conclude that targeting a cryptic splice site with ASOs may hold therapeutic potential for JEB and possibly other forms of EB with similar types of mutations.

A PTEN-COL17A1 fusion gene and its novel regulatory role in Collagen XVII expression and GBM malignance.

Collagen XVII expression has recently been demonstrated to be correlated with the tumor malignance. While Collagen XVII is known to be widely distributed in neurons of the human brain, its precise role in pathogenesis of glioblastoma multiforme (GBM) is unknown. In this study, we identified and characterized a new PTEN-COL17A1 fusion gene in GMB using transcriptome sequencing. Although fusion gene did not result in measurable fusion protein production, its presence is accompanied with high levels of COL17A1 expression, revealed a novel regulatory mechanism of Collagen XVII expression by PTEN-COL17A1 gene fusion. Knocked down Collagen XVII expression in glioma cell lines resulted in decreased tumor invasiveness, along with significant reduction of MMP9 expression, while increased Collagen XVII expression promotes invasive activities of glioma cells and associated with GBM recurrences. Together, our results uncovered a new PTEN-COL17A1 fusion gene and its novel regulatory role in Collagen XVII expression and GBM malignance, and demonstrated that COL17A1 could serve as a useful prognostic biomarker and therapeutic targets for GBM.

Identification of a novel p53 target, COL17A1, that inhibits breast cancer cell migration and invasion.

p53 mutation is a marker of poor prognosis in breast cancers. To identify downstream targets of p53, we screened two transcriptome datasets, including cDNA microarrays of MCF10A breast epithelial cells with wild-type p53 or p53-null background, and RNA sequence analysis of breast invasive carcinoma. Here, we unveil ten novel p53 target candidates that are up-regulated after the induction of p53 in wild-type cells. Their expressions are also high in breast invasive carcinoma tissues with wild-type p53. The GO analysis identified epidermis development and ectoderm development, which COL17A1 participates, as significantly up-regulated by wild-type p53. The COL17A1 expressions increased in a p53-dependent manner in human breast cells and mouse mammary tissues. Reporter assay and ChIP assay identified intronic p53-binding sequences in the COL17A1 gene. The MDA-MB-231 cells that genetically over-express COL17A1 gene product exhibited reduced migration and invasion in vitro. Similarly, COL17A1 expression was decreased in metastatic tumors compared to primary tumors and normal tissues, even from the same patients. Moreover, high COL17A1 expression was associated with longer survival of patients with invasive breast carcinoma. In conclusion, we revealed that COL17A1 is a novel p53 transcriptional target in breast tissues that inhibits cell migration and invasion and is associated with better prognosis.

Evidence for Activation of the Unfolded Protein Response in Collagen IV Nephropathies

Thin-basement-membrane nephropathy (TBMN) and Alport syndrome (AS) are progressive collagen IV nephropathies caused by mutations in COL4A3/A4/A5 genes. These nephropathies invariably present with microscopic hematuria and frequently progress to proteinuria and CKD or ESRD during long-term follow-up. Nonetheless, the exact molecular mechanisms by which these mutations exert their deleterious effects on the glomerulus remain elusive. We hypothesized that defective trafficking of the COL4A3 chain causes a strong intracellular effect on the cell responsible for COL4A3 expression, the podocyte. To this end, we overexpressed normal and mutant COL4A3 chains (G1334E mutation) in human undifferentiated podocytes and tested their effects in various intracellular pathways using a microarray approach. COL4A3 overexpression in the podocyte caused chain retention in the endoplasmic reticulum (ER) that was associated with activation of unfolded protein response (UPR)-related markers of ER stress. Notably, the overexpression of normal or mutant COL4A3 chains differentially activated the UPR pathway. Similar results were observed in a novel knockin mouse carrying the Col4a3-G1332E mutation, which produced a phenotype consistent with AS, and in biopsy specimens from patients with TBMN carrying a heterozygous COL4A3-G1334E mutation. These results suggest that ER stress arising from defective localization of collagen IV chains in human podocytes contributes to the pathogenesis of TBMN and AS through activation of the UPR, a finding that may pave the way for novel therapeutic interventions for a variety of collagenopathies.

High COL4A3 expression correlates with poor prognosis after cisplatin plus gemcitabine chemotherapy in non-small cell lung cancer

In this study, we investigated whether COL4A3 mRNA expression levels were associated with clinical outcomes after treatment with a combination of gemcitabine (Gem)/cis-diamminedichloroplatinum(II) (CDDP) regimen for patients with advanced stage non-small cell lung cancer (NSCLC). Response and survival were correlated with the level of COL4A3 expression in 58 patients with advanced (stage IIIb or IV) NSCLC treated as part of a multicenter randomized trial with Gem 1,250mg/m(2) on days 1 and 8 plus CDDP 100mg/m(2) on day 1 every 3weeks. mRNA was isolated from paraffin-embedded pretreatment primary tumor specimens, and relative expression levels of COL4A3/β-actin were measured using a quantitative reverse transcription-PCR (Taqman) system. COL4A3 expression was detectable in all tumors. There were no significant differences in COL4A3 levels by gender, age, performance status, weight loss, or tumor stage. The overall response rate was 45.8%. There were no significant associations between COL4A3 expression and response. Median overall survival was significantly longer in patients with low COL4A3 expression tumors compared to patients with high expression tumors. COL4A3 expression, Eastern Cooperative Oncology Group performance status, and presence of weight loss were significant prognostic factors for survival in a Cox proportional hazards multivariable analysis. These data suggest that COL4A3 expression is a predictive factor for survival after CDDP/Gem therapy in advanced NSCLC. Although there was a trend toward decreased response with high COL4A3 mRNA levels, this difference failed to reach statistical significance. This result may reflect the impact of Gem and the requirement for COL4A3 expression for CDDP/Gem synergism or may be attributable to the relatively small patient sample size in this study. Prospective studies of COL4A3 as a predictive marker for activity of CDDP-based regimens in NSCLC are warranted.

COL4A3 expression correlates with pathogenesis, pathologic behaviors, and prognosis of gastric carcinomas

COL4A3 protein belongs to type IV collagen family and is closely linked to kidney diseases and cancer. To clarify the roles of COL4A3 in gastric carcinogenesis and subsequent progression, its expression was examined by immunohistochemistry on tissue microarrays containing gastric carcinomas, adjacent intestinal metaplasia, pure intestinal metaplasia, and gastritis. Gastric carcinoma tissue and cell lines were studied for COL4A3 expression by Western blotting and reverse transcription-polymerase chain reaction. We found that COL4A3 was differentially expressed in GES-1, AGS, BGC-823, GT-3 TKB, HGC-27, KATO-III, MGC-803, MKN28, MKN45, SCH, SGC-7901, and STKM-2 at both messenger RNA and protein levels. Carcinomas showed statistically lower COL4A3 expression than matched nonneoplastic mucosa (P < .05). Expression was strong in intestinal metaplasia in comparison with gastritis and carcinoma (P < .05). There was greater COL4A3 expression in carcinoma than gastritis (P < .05). Expression of COL4A3 protein was positively correlated with tumor size, lymphatic invasion, venous invasion, and TNM stage (P < .05). There was more COL4A3 expression in diffuse than in intestinal-type carcinomas regardless of invasion into the muscularis propria (P < .05). Histologically, all signet ring cell (n = 43) and mucinous (n = 12) carcinomas showed COL4A3 expression. Kaplan-Meier analysis indicated that COL4A3 expression was negatively associated with a favorable prognosis of overall, advanced, and intestinal-type gastric carcinomas (P < .05). Aberrant COL4A3 expression might play an important role in the pathogenesis and subsequent progression of gastric carcinoma. COL4A3 overexpression might be used as a marker of gastric intestinal metaplasia and mucinous and signet ring cell carcinoma.