scholarly journals Simultaneous Quantitative Detection of Relevant Biomarkers in Breast Cancer by Quantitative Real-Time PCR

2006 ◽  
Vol 21 (1) ◽  
pp. 30-39 ◽  
Author(s):  
M. Labuhn ◽  
V. Vuaroqueaux ◽  
F. Fina ◽  
A. Schaller ◽  
I. Nanni-Metellus ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Mrna Expression ◽  
Real Time ◽  
Real Time Pcr ◽  
Quantitative Detection ◽  
Mrna Levels ◽  
Quantitative Real Time Pcr ◽  
Expression Levels ◽  
Qrt Pcr ◽  
Mrna Expression Levels

The assessment of ERα, PgR and HER2 status is routinely performed today to determine the endocrine responsiveness of breast cancer samples. Such determination is usually accomplished by means of immunohistochemistry and in case of HER2 amplification by means of fluorescent in situ hybridization (FISH). The analysis of these markers can be improved by simultaneous measurements using quantitative real-time PCR (Qrt-PCR). In this study we compared Qrt-PCR results for the assessment of mRNA levels of ERα, PgR, and the members of the human epidermal growth factor receptor family, HER1, HER2, HER3 and HER4. The results were obtained in two independent laboratories using two different methods, SYBR Green I and TaqMan probes, and different primers. By linear regression we demonstrated a good concordance for all six markers. The quantitative mRNA expression levels of ERα, PgR and HER2 also strongly correlated with the respective quantitative protein expression levels prospectively detected by EIA in both laboratories. In addition, HER2 mRNA expression levels correlated well with gene amplification detected by FISH in the same biopsies. Our results indicate that both Qrt-PCR methods were robust and sensitive tools for routine diagnostics and consistent with standard methodologies. The developed simultaneous assessment of several biomarkers is fast and labor effective and allows optimization of the clinical decision-making process in breast cancer tissue and/or core biopsies.

scholarly journals Synaptotagmin 13 Is Highly Expressed in Estrogen Receptor-Positive Breast Cancer

2021 ◽  
Vol 28 (5) ◽  
pp. 4080-4092
Author(s):  
Takahiro Ichikawa ◽  
Masahiro Shibata ◽  
Takahiro Inaishi ◽  
Ikumi Soeda ◽  
Mitsuro Kanda ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Estrogen Receptor ◽  
Mrna Expression ◽  
Cell Lines ◽  
Mrna Levels ◽  
Pcr Array ◽  
Array Analysis ◽  
Expression Levels ◽  
Er Positive ◽  
Mrna Expression Levels

Background: Accumulating evidence indicates tumor-promoting roles of synaptotagmin 13 (SYT13) in several cancers; however, no studies have investigated its expression in breast cancer (BC). This study aimed to clarify the significance of SYT13 in BC. Methods: SYT13 mRNA expression levels were evaluated in BC cell lines. Polymerase chain reaction (PCR) array analysis was conducted to determine the correlation between expression levels of SYT13 and other tumor-associated genes. Then, the association of SYT13 expression levels in the clinical BC specimens with patients’ clinicopathological factors was evaluated. These findings were subsequently validated using The Cancer Genome Atlas (TCGA) database. Results: Among 13 BC cell lines, estrogen receptor (ER)-positive cells showed higher SYT13 mRNA levels than ER-negative cells. PCR array analysis revealed positive correlations between SYT13 and several oncogenes predominantly expressed in ER-positive BC, such as estrogen receptor 1, AKT serine/threonine kinase 1, and cyclin-dependent kinases 4. In 165 patients, ER-positive specimens exhibited higher SYT13 mRNA expression levels than ER-negative specimens. The TCGA database analysis confirmed that patients with ER-positive BC expressed higher SYT13 levels than ER-negative patients. Conclusion: This study suggests that SYT13 is highly expressed in ER-positive BC cells and clinical specimens, and there is a positive association of SYT13 with the ER signaling pathways.

p29ING4 Regulates the Production of Pro-Angiogenic Molecules by Human Myeloma Cells in Normoxic and Hypoxic Conditions Being Involved in Myeloma-Induced Angiogenesis.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 515-515
Author(s):  
Sara Tagliaferri ◽  
Francesca Morandi ◽  
Paolo Lunghi ◽  
Simona Colla ◽  
Mirca Lazzaretti ◽  
...  
Keyword(s):  
Tumor Suppressor ◽  
Mrna Expression ◽  
Real Time ◽  
Real Time Pcr ◽  
Interleukin 8 ◽  
Mrna Levels ◽  
Quantitative Real Time Pcr ◽  
Angiogenic Switch ◽  
Hypoxic Conditions

Abstract Multiple myeloma (MM) cells produce several angiogenic molecules as VEGF, Angiopoietin-1 (Ang-1), interleukin-8 (IL-8) and osteopontin (OPN), however the molecular mechanisms underlying the angiogenic switch are not completely elucidated. The candidate tumor suppressor gene inhibitor of growth family member 4 (p29ING4) has been recently implicated in solid tumors as a repressor of angiogenesis and tumor growth through the suppression of angiogenic related molecules including interleukin-8 (IL-8) and the hypoxia inducible factor (HIF)-1 alpha. In this study we investigate the potential involvement of p29ING4 in the angiogenic switch in MM. First using quantitative real time PCR we compared p29ING4 with VEGF, Ang-1, IL-8 and OPN mRNA levels in eight human myeloma cell lines (HMCLs). A significantly negative correlation was observed between ING4 and IL-8 and a trend of correlation with OPN. Following we transfected HMCLs JJN3, OPM-2 and RPMI-8226 with specific siRNA to completely block the expression of p29ING4 checking the effect on the expression and production of the myeloma-related angiogenic molecules VEGF, Ang-1, IL-8 and OPN by quantitative real time PCR and ELISA assay. p29ING4 suppression in HMCLs did not affect VEGF and Ang-1 production but induced a strong up-regulation of IL-8 mRNA and IL-8 protein secretion. Similarly an induction of OPN mRNA expression as well as OPN secretion was induced by siRNA anti-p29ING4. Moreover conditioned media of HMCLs transfected with siRNA anti p29ING4 significantly increased vessel formation in an experimental in vitro model of angiogenesis (ANGIO kit) as compared to controls. Further we investigate the role of p29ING4 in the production of angiogenic molecule by MM cells in hypoxic condition compared to normoxic one as well as its potential relationship with HIF-1alpha system. Hypoxia induced HIF-1alpha expression at nuclear level and its activity in HMCLs and p29ING4 suppression by siRNA further induced HIF-1alpha transcriptional activity with an increase of its target gene Nip-3 in HMCLs. In turn the block of HIF1-alpha by specific siRNA up-regulated p29ING4 and suppressed IL-8 and OPN mRNA expression by HMCLs suggesting a relationship between p29ING4 and HIF-1alpha activity. These in vitro observations have been extended in vivo by the finding of a significant correlation between bone marrow (BM) plasma IL-8 levels and p29ING4 mRNA expression in purified MM cells obtained from 40 newly diagnosed MM patients (R=−0.58 Spearman’s 2-tailed test: p=0.04), consistently MM patients with higher BM IL-8 levels have a significantly lower p29ING4 mRNA levels. Similarly MM patients positive for OPN expression with high OPN BM levels had a significant lower p29ING4 levels (p=0.02). Finally we found that MM patients with high microvalscular density (MVD>30) have significant lower p29ING4 levels as compared to those with low MVD (<30) (p=0.04) and that MM patients with histological high grade had significant lower p29ING4 mRNA levels (Mann-Whitney 2-tailed: p=0.05). In conclusion, our data indicate that the tumor suppressor p29ING4 regulate the production of angiogenic molecules by MM cells both in normoxic and hypoxic conditions being involved in MM-induced angiogenesis and potentially in tumor progression.

Evaluation of thymidylate synthase and ERCC1 mRNA levels as predictive markers in colorectal cancer patients treated with S-1 and oxaliplatin

2009 ◽  
Vol 27 (15_suppl) ◽  
pp. e15071-e15071
Author(s):  
H. Kuramochi ◽  
K. Hayashi ◽  
G. Nakajima ◽  
H. Kamikozuru ◽  
M. Yamamoto
Keyword(s):  
Colorectal Cancer ◽  
Mrna Expression ◽  
Real Time ◽  
Cancer Patients ◽  
Thymidylate Synthase ◽  
Excision Repair ◽  
Mrna Levels ◽  
Expression Levels ◽  
Colorectal Cancer Patients ◽  
Mrna Expression Levels

e15071 Background: Oxaliplatin has been widely used for the treatment of colorectal cancer. The mechanism of action of platinum compounds such as oxaliplatin is to bind to a DNA molecule in the form of a platinum-DNA-adduct. Excision repair cross complementation group 1 (ERCC1), which plays a major role in the nucleotide excision pathway, has a polymorphism in codon 118, and is reported to be associated with a resistance to platinum-based therapy. Thymidylate synthase (TS) and dehydropyrimidine dehydrogenase (DPD) are key enzymes of 5-FU metabolism and are well known to be associated with a response to 5-FU-based therapy. Methods: Twenty-one colorectal cancer patients (male:female = 7:14; median age, 65) treated with a combination of oxaliplatin and S-1 as a first-line therapy were analyzed for ERCC1 codon 118 polymorphism and the mRNA expression levels of TS, ERCC1, and DPD. Formalin-fixed paraffin- embedded surgical specimens were used and t-RNA and DNA were extracted. The mRNA expression levels were measured using real-time RT-PCR, and the polymorphism was analyzed using the allelic discrimination method together with real-time PCR. Results: No correlation was observed between ERCC1 codon118 polymorphism and any response to the chemotherapy. ERCC1 mRNA levels tended to be higher in the patients with wild-type homozygous alleles in codon 118 than in those with at least one mutant allele(1.19 vs.0.68: p= 0.15). Patients with both high TS and ERCC1 mRNA levels showed a significantly lower response rate than the others (25% vs. 67%, p=0.02). No relationship was seen between DPD mRNA expression levels and the response. Conclusions: The mRNA expression levels of TS and ERCC1 appear to be useful markers for the treatment of S-1 and oxaliplatin. No particular usefulness of ERCC1 codon 118 polymorphism was verified. No significant financial relationships to disclose.

scholarly journals Druggable Nucleolin Identifies Breast Tumours Associated with Poor Prognosis That Exhibit Different Biological Processes

Cancers ◽  
2018 ◽  
Vol 10 (10) ◽  
pp. 390 ◽  
Author(s):  
Flora Nguyen Van Long ◽  
Audrey Lardy-Cleaud ◽  
Susan Bray ◽  
Sylvie Chabaud ◽  
Thierry Dubois ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Mrna Expression ◽  
Expression Profiles ◽  
Disease Free Survival ◽  
Mrna Levels ◽  
Breast Cancers ◽  
Breast Tumours ◽  
Expression Levels ◽  
Cancer Management ◽  
Mrna Expression Levels

Background: Nucleolin (NCL) is a multifunctional protein with oncogenic properties. Anti-NCL drugs show strong cytotoxic effects, including in triple-negative breast cancer (TNBC) models, and are currently being evaluated in phase II clinical trials. However, few studies have investigated the clinical value of NCL and whether NCL stratified cancer patients. Here, we have investigated for the first time the association of NCL with clinical characteristics in breast cancers independently of the different subtypes. Methods: Using two independent series (n = 216; n = 661), we evaluated the prognostic value of NCL in non-metastatic breast cancers using univariate and/or multivariate Cox-regression analyses. Results: We reported that NCL mRNA expression levels are markers of poor survivals independently of tumour size and lymph node invasion status (n = 216). In addition, an association of NCL expression levels with poor survival was observed in TNBC (n = 40, overall survival (OS) p = 0.0287, disease-free survival (DFS) p = 0.0194). Transcriptomic analyses issued from The Cancer Genome Atlas (TCGA) database (n = 661) revealed that breast tumours expressing either low or high NCL mRNA expression levels exhibit different gene expression profiles. These data suggest that tumours expressing high NCL mRNA levels are different from those expressing low NCL mRNA levels. Conclusions: NCL is an independent marker of prognosis in breast cancers. We anticipated that anti-NCL is a promising therapeutic strategy that could rapidly be evaluated in high NCL-expressing tumours to improve breast cancer management.

scholarly journals Kallikrein-related peptidases in triple negative breast cancer: prognostic value of quantitatively assessed KLK8, KLK10 and KLK11 mRNA expression

2021 ◽  
Author(s):  
Yueyang Liu ◽  
Sarah Preis ◽  
Geng Xiaocong ◽  
Weiwei Gong ◽  
Marion Kiechle ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Mrna Expression ◽  
Triple Negative Breast Cancer ◽  
Triple Negative ◽  
Cox Regression ◽  
Prognostic Biomarker ◽  
Disease Free Survival ◽  
Mrna Levels ◽  
Expression Levels ◽  
Mrna Expression Levels

Abstract Background. Triple-negative breast cancer (TNBC) is a breast cancer subtype with poor prognosis and limited targeted therapy options. Multiple KLKs have been described to play key roles in carcinogenesis and metastasis of breast cancer. Purpose. In the present study, the clinical significance of KLK8, KLK10, and KLK11 mRNA expression in tumor tissue of TNBC patients was investigated. Methods. The mRNA expression levels of KLK8, 10, and 11 were quantified by quantitative PCR and their prognostic values were analyzed in a large, well-characterized TNBC cohort (n = 123). Results. Significantly positive correlations were observed between all three KLK mRNA levels indicating coordinate expression of these proteases in TNBC. In univariate analyses, both elevated KLK8, KLK10 as well as all combinations of the three factors (KLK8 + KLK10, KLK8 + KLK11, KLK10 + KLK11, KLK8 + KLK10 + KLK11) were significantly associated with shortened disease-free survival (DFS), while high mRNA levels of KLK11, as well as KLK10 + KLK11 were significantly associated with shortened overall survival (OS). In multivariate Cox regression analyses, KLK10 and all combined factors remained unfavorable independent predictive markers for DFS, while high KLK11 mRNA expression represented an unfavorable independent predictor for OS. Conclusions. Increased KLK8, KLK10, and KLK11 mRNA expression levels are associated with unfavorable prognosis in triple-negative breast cancer. The combination of KLK8 + KLK10 + KLK11 may represent a stronger prognostic biomarker for DFS than KLK8, KLK10, KLK11 alone, or other combinations thereof, whereas KLK11 mRNA expression is an independent prognostic biomarker for OS in TNBC patients.

scholarly journals Selection of reference genes for quantitative real-time PCR analysis in cucumber (Cucumis sativus L.), pumpkin (Cucurbita moschata Duch.) and cucumber–pumpkin grafted plants

PeerJ ◽  
2019 ◽  
Vol 7 ◽  
pp. e6536 ◽  
Author(s):  
Li Miao ◽  
Xing Qin ◽  
Lihong Gao ◽  
Qing Li ◽  
Shuzhen Li ◽  
...  
Keyword(s):  
Gene Expression ◽  
Real Time ◽  
Reference Gene ◽  
Real Time Pcr ◽  
Reference Genes ◽  
Stable Expression ◽  
Quantitative Real Time Pcr ◽  
Graft Union ◽  
Expression Levels ◽  
Qrt Pcr

Background Quantitative real-time PCR (qRT-PCR) is a commonly used high-throughput technique to measure mRNA transcript levels. The accuracy of this evaluation of gene expression depends on the use of optimal reference genes. Cucumber–pumpkin grafted plants, made by grafting a cucumber scion onto pumpkin rootstock, are superior to either parent plant, as grafting conveys many advantages. However, although many reliable reference genes have been identified in both cucumber and pumpkin, none have been obtained for cucumber–pumpkin grafted plants. Methods In this work, 12 candidate reference genes, including eight traditional genes and four novel genes identified from our transcriptome data, were selected to assess their expression stability. Their expression levels in 25 samples, including three cucumber and three pumpkin samples from different organs, and 19 cucumber–pumpkin grafted samples from different organs, conditions, and varieties, were analyzed by qRT-PCR, and the stability of their expression was assessed by the comparative ΔCt method, geNorm, NormFinder, BestKeeper, and RefFinder. Results The results showed that the most suitable reference gene varied dependent on the organs, conditions, and varieties. CACS and 40SRPS8 were the most stable reference genes for all samples in our research. TIP41 and CACS showed the most stable expression in different cucumber organs, TIP41 and PP2A were the optimal reference genes in pumpkin organs, and CACS and 40SRPS8 were the most stable genes in all grafted cucumber samples. However, the optimal reference gene varied under different conditions. CACS and 40SRPS8 were the best combination of genes in different organs of cucumber–pumpkin grafted plants, TUA and RPL36Aa were the most stable in the graft union under cold stress, LEA26 and ARF showed the most stable expression in the graft union during the healing process, and TIP41 and PP2A were the most stable across different varieties of cucumber–pumpkin grafted plants. The use of LEA26, ARF and LEA26+ARF as reference genes were further verified by analyzing the expression levels of csaCYCD3;1, csaRUL, cmoRUL, and cmoPIN in the graft union at different time points after grafting. Discussion This work is the first report of appropriate reference genes in grafted cucumber plants and provides useful information for the study of gene expression and molecular mechanisms in cucumber–pumpkin grafted plants.

scholarly journals Exploring the metastatic role of the inhibitor of apoptosis BIRC6 in Breast Cancer

2021 ◽  
Author(s):  
Santiago Manuel Gomez Bergna ◽  
Abril Marchesini ◽  
Leslie Cinthya Amoros Morales ◽  
Paula Nazarena Arrias ◽  
Hernan Gabriel Farina ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Gene Ontology ◽  
Mrna Expression ◽  
Differential Expression ◽  
Expression Patterns ◽  
Mrna Levels ◽  
Expression Levels ◽  
Apoptosis Protein ◽  
Mrna Expression Levels

Breast cancer is the most common cancer as well as the first cause of death by cancer in women worldwide. BIRC6 (baculoviral IAP repeat-containing protein 6) is a member of the inhibitors of apoptosis protein family thought to play an important role in the progression or chemoresistance of many cancers. The aim of the present work was to investigate the role of apoptosis inhibitor BIRC6 in breast cancer, focusing particularly on its involvement in the metastatic cascade. We analyzed BIRC6 mRNA expression levels and Copy Number Variations (CNV) in three breast cancer databases from The Cancer Genome Atlas (TCGA) comparing clinical and molecular attributes. Genomic analysis was performed using CBioportal platform while transcriptomic studies (mRNA expression levels, correlation heatmaps, survival plots and Gene Ontology) were performed with USC Xena and R. Statistical significance was set at p values less than 0.05. Our analyses showed that there was a differential expression of BIRC6 in cancer samples when compared to normal samples. CNV that involve amplification and gain of BIRC6 gene were correlated with negative hormone receptor tumors, higher prognostic indexes, younger age at diagnosis and both chemotherapy and radiotherapy administration. Transcriptomic and gene-ontology analyses showed that, in conditions of high BIRC6 mRNA levels, there are differential expression patterns in apoptotic, proliferation, and metastatic pathways. In summary, our in silico analyses suggest that BIRC6 exhibits an antiapoptotic, pro-proliferative and an apparent pro-metastatic role and could be a relevant molecular target for treatment of Breast Cancer tumors.

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