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RNAscope® ISH to detect breast cancer-related markers

RNA expression of ERBB2 gene in human breast cancer tissue using RNAscope<sup>®</sup> 2.5 HD Assay Brown RNA in situ hybridization (ISH)

RNA expression of ERBB2 gene in human breast cancer tissue using RNAscope® 2.5 HD Assay Brown

Expression of HOTAIR in human breast cancer tissue using RNAscope<sup>®</sup> 2.5 HD Assay Brown RNA in situ hybridization (ISH)

Expression of HOTAIR in human breast cancer tissue using RNAscope® 2.5 HD Assay Brown

Expression of HOTAIR in human breast cancer tissue using RNAscope<sup>®</sup> 2.5 HD Assay Brown RNA in situ hybridization (ISH)

Expression of HOTAIR in human breast cancer tissue using RNAscope® 2.5 HD Assay Brown

Expression of GAS5 in human breast cancer tissue using RNAscope<sup>®</sup> 2.5 HD Assay Brown RNA in situ hybridization (ISH)

Expression of GAS5 in human breast cancer tissue using RNAscope® 2.5 HD Assay Brown

Expression of PVT1 in human breast cancer tissue using RNAscope<sup>®</sup> 2.5 HD Assay Brown RNA in situ hybridization (ISH)

Expression of PVT1 in human breast cancer tissue using RNAscope® 2.5 HD Assay Brown

Expression of TNFA in human breast cancer tissue using RNAscope<sup>®</sup> 2.5 HD Assay Brown RNA in situ hybridization (ISH)

Expression of TNFA in human breast cancer tissue using RNAscope® 2.5 HD Assay Brown

Breast cancer is a complex disease with great clinical and molecular heterogeneity that requires accurate assessment of known breast cancer biomarkers, such as HER2, estrogen receptor (ER), and progesterone receptor (PgR), as well as the discovery of emerging potential targets, for the precise treatment of breast tumors.

The RNAscope® in situ hybridization assay offers a very reliable and robust method for the detection and validation of breast cancer-related markers within the tissue environment and can be performed on routinely available FFPE samples. Several studies have been published demonstrating the application of the RNAscope® ISH assay in breast-cancer research:

  • Resolution of equivocal and heterogeneous cases of HER2 expression
  • Patient stratification for treatment regimens
  • Validation of transcriptome data
  • Sub-cellular localization of RNA in breast cancer
  • Detection of lncRNAs in breast cancer
  • Preclinical drug efficacy for breast cancer treatment
  • Assessment of potential prognostic markers for breast cancer

Resolution of equivocal and heterogeneous cases of HER2 expression: HER2, a proto-oncogene encoding the HER2 tyrosine kinase receptor, is amplified in 10 to 20% of breast cancers, leading to an aggressive tumor phenotype associated with reduced survival and high metastatic potential. While standard HER2 testing methods have been developed, including IHC for protein detection and FISH for DNA detection, these methods are often hampered by equivocal testing and intratumoral heterogeneity. Wang et al. developed a fully automated, quantitative ISH assay using the RNAscope® technology to quantify single-cell HER2 mRNA levels in invasive breast carcinomas. This assay was able to resolve the HER2 amplification status in problematic, heterogeneous cases or cases that were not resolvable by standard methods. 
Automated Quantitative RNA in Situ Hybridization for Resolution of Equivocal and Heterogeneous ERBB2 (HER2) Status in Invasive Breast Carcinoma

Patient-stratification for treatment regimens: Accurate assessment of HER2 status is necessary to recommend therapy for patients who are most likely to benefit from anti-HER2 treatment such as trastuzumab (Herceptin) and minimize unnecessary overtreatment and exposure to side effects. Vassilakopoulou et al. used the RNAscope® assay to assess HER2 mRNA as a predictor of benefit from trastuzumab-based chemotherapy and to correlate it to HER2 protein and gene levels. HER2 mRNA levels were significantly associated with time to progression and overall survival in a trastuzumab-treated metastatic cohort. Therefore, in situ assessment of HER2 mRNA has the potential to identify breast cancer patients who may benefit from trastuzumab treatment.
In situ Quantitative Measurement of HER2mRNA Predicts Benefit from Trastuzumab-Containing Chemotherapy in a Cohort of Metastatic Breast Cancer Patients

Validation of transcriptome data and sub-cellular localization of lncRNA in breast cancer: Recent research has been focusing on long non-coding RNAs (lncRNAs) as potential biomarkers and therapeutic targets to combat triple-negative breast cancer (TNBC). A previous study using lncRNA microarrays performed on stage III breast cancer tissue and adjacent non-cancerous tissue identified 2839 lncRNAs differentially expressed between breast cancer and adjacent normal tissues. In this study Lin et al. selected LINK-A from these lncRNAs due to its frequently elevated expression in TNBC patients. RNAscope® ISH confirmed the array results, showing that expression of LINK-A was significantly increased in TNBC tissues compared with normal breast tissues and that its sub-cellular localization was in the cytoplasm or close to the cellular membrane. 
The LINK-A lncRNA activates normoxic HIF1α signalling in triple-negative breast cancer

Preclinical drug efficacy for breast cancer treatment: FGFR2 gene amplification is found in 4% of triple-negative breast cancers (TNBC) and may promote breast tumorigenicity by maintaining breast tumor-initiating cells, making FGFR2 a potential therapeutic target. In this study, Sommer et al. demonstrate the preclinical efficacy of BAY1187982, a novel FGFR2-antibody drug conjugate (ADC). A positive correlation was observed between FGFR2 gene amplification and mRNA expression (shown by RNAscope® ISH) and antitumor activity, making FGFR2 mRNA and FGFR2 gene amplification potential selection markers for patient stratification during clinical development of the FGFR2-ADC. 
Preclinical efficacy of the auristatin-based antibody-drug conjugate BAY 1187982 for the treatment of FGFR2-positive solid tumors

Assessment of HOTAIR as a prognostic breast cancer biomarker for breast cancer: Metastasis of breast tumors to the lymph nodes is an important means of tumor dissemination and the number of involved lymph nodes is a strong indicator of whether or not the cancer is metastatic. High expression of the large intergenic non-coding RNA (lincRNA) HOTAIR has been shown to be a significant predictor of poor prognosis and metastasis in breast carcinomas. Using RNAscope® ISH to assess whether HOTAIR expression could be used as a surrogate to assess nodal metastases in breast cancer patients, Gökmen-Polar et al. showed that the prognostic role of HOTAIR is restricted to ER-/lymph node+ tumors, where its expression could be used as a potential prognostic marker to identify patients at greater risk for poor overall survival.
Prognostic Impact of HOTAIR Expression is Restricted to ER-Negative Breast Cancers

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