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Altered expression of insulin receptor isoforms in breast cancer.

ABSTRACT: PURPOSE: Insulin-like growth factor (IGF) signaling through human insulin receptor isoform A (IR-A) contributes to tumorigenesis and intrinsic resistance to anti-IGF1R therapy. In the present study, we (a) developed quantitative TaqMan real time-PCR-based assays (qRT-PCR) to measure human insulin receptor isoforms with high specificity, (b) evaluated isoform expression levels in molecularly-defined breast cancer subtypes, and (c) identified the IR-A:IR-B mRNA ratio as a potential biomarker guiding patient stratification for anti-IGF therapies. EXPERIMENTAL DESIGN: mRNA expression levels of IR-A and IR-B were measured in 42 primary breast cancers and 19 matched adjacent normal tissues with TaqMan qRT-PCR assays. The results were further confirmed in 165 breast cancers. The tumor samples were profiled using whole genome microarrays and subsequently subtyped using the PAM50 breast cancer gene signature. The relationship between the IR-A:IR-B ratio and cancer subtype, as well as markers of proliferation were characterized. RESULTS: The mRNA expression levels of IR-A in the breast tumors were similar to those observed in the adjacent normal tissues, while the mRNA levels of IR-B were significantly decreased in tumors. The IR-A:IR-B ratio was significantly higher in luminal B breast cancer than in luminal A. Strong concordance between the IR-A:IR-B ratio and the composite Oncotype DX proliferation score was observed for stratifying the latter two breast cancer subtypes. CONCLUSIONS: The reduction in IR-B expression is the key to the altered IR-A:IR-B ratio observed in breast cancer. The IR-A:IR-B ratio may have biomarker utility in guiding a patient stratification strategy for an anti-IGF therapeutic.

SUBMITTER: Huang J

PROVIDER: S-EPMC3202518 | biostudies-literature | 2011

REPOSITORIES: biostudies-literature

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Altered expression of insulin receptor isoforms in breast cancer.

Huang Jiaqi J Morehouse Chris C Streicher Katie K Higgs Brandon W BW Gao Jin J Czapiga Meggan M Boutrin Anmarie A Zhu Wei W Brohawn Philip P Chang Yong Y Viner Jaye J LaVallee Theresa T Richman Laura L Jallal Bahija B Yao Yihong Y

PloS one 20111026 10

<h4>Purpose</h4>Insulin-like growth factor (IGF) signaling through human insulin receptor isoform A (IR-A) contributes to tumorigenesis and intrinsic resistance to anti-IGF1R therapy. In the present study, we (a) developed quantitative TaqMan real time-PCR-based assays (qRT-PCR) to measure human insulin receptor isoforms with high specificity, (b) evaluated isoform expression levels in molecularly-defined breast cancer subtypes, and (c) identified the IR-A:IR-B mRNA ratio as a potential biomarke ...[more]

PMID: 22046260

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Project description:IntroductionWe sought to determine the level of protein expression of the critical components of the insulin-like growth factor receptor (IGFR) pathway and to evaluate their prognostic significance across the different early breast cancer subtypes.Patients and methodsArchival tumor tissue from 1,021 women with early, node positive breast cancer, who were prospectively evaluated within two randomized clinical trials, was used to construct tissue microarrays that were stained for hormone receptors (HR), Ki67, HER2, epidermal growth factor receptor (EGFR) and cytokeratins 5/6, to classify tumors into five immunophenotypical subgroups. Immunohistochemical (IHC) expression of IGF1R-alpha and beta subunits, IGF2R and IGF-binding protein 2 (IGFBP2) was assessed using the immunoreactive score (IRS). Repeated internal cross-validation was performed to examine the statistical validity of the cut off points for all biomarkers.ResultsAfter a median follow-up time of 105.4 months, overall 370 women (36.2%) had relapsed and 270 (26.4%) had died. Tumors expressing IGF1R-alpha above the median IRS were significantly more frequently HR positive (luminal A+B+HER2), as compared to HER2-enriched and triple negative ones (p<0.001 for both comparisons). IGF2R was overexpressed significantly more frequently in HR negative tumors (p?=?0.001) and had an inverse correlation with all other biomarkers. Patients with luminal A and B tumors with high IGF1R-alpha and negative EGFR expression (N?=?190) had significantly higher 4-year survival rates, as compared to the rest (log-rank p?=?0.046), as did patients with luminal A and B tumors with high IGF1R-alpha and low IGF2R expression, as compared to the rest (N?=?91), (log-rank p?=?0.035). After adjustment for significant variables, patients in the latter group had a relative 45% reduction in the risk of death, as compared to the rest (p?=?0.035).ConclusionAberrant expression of components of the IGF1R pathway is associated with better clinical outcomes in women with luminal A and B, node positive, early breast cancer.

Project description:An increase in the expression of estrogen receptors (ER) and the expanded population of ER-positive cells are two common phenotypes of breast cancer. Detection of the aberrantly expressed ERα in breast cancer is carried out using ERα-antibodies and radiolabelled ligands to make decisions about cancer treatment and targeted therapy. Capitalizing on the beneficial advantages of aptamer over the conventional antibody or radiolabelled ligand, we have identified a DNA aptamer that selectively binds and facilitates the detection of ERα in human breast cancer tissue sections. The aptamer is identified using the high throughput sequencing assisted SELEX screening. Biophysical characterization confirms the binding and formation of a thermodynamically stable complex between the identified DNA aptamer (ERaptD4) and ERα (Ka = 1.55±0.298×108 M(-1); ΔH = 4.32×104±801.1 cal/mol; ΔS = -108 cal/mol/deg). Interestingly, the specificity measurements suggest that the ERaptD4 internalizes into ERα-positive breast cancer cells in a target-selective manner and localizes specifically in the nuclear region. To harness these characteristics of ERaptD4 for detection of ERα expression in breast cancer samples, we performed the aptamer-assisted histochemical analysis of ERα in tissue samples from breast cancer patients. The results were validated by performing the immunohistochemistry on same samples with an ERα-antibody. We found that the two methods agree strongly in assay output (kappa value = 0.930, p-value <0.05 for strong ERα positive and the ERα negative samples; kappa value = 0.823, p-value <0.05 for the weak/moderate ER+ve samples, n = 20). Further, the aptamer stain the ERα-positive cells in breast tissues without cross-reacting to ERα-deficient fibroblasts, adipocytes, or the inflammatory cells. Our results demonstrate a significant consistency in the aptamer-assisted detection of ERα in strong ERα positive, moderate ERα positive and ERα negative breast cancer tissues. We anticipate that the ERaptD4 aptamer targeting ERα may potentially be used for an efficient grading of ERα expression in cancer tissues.

Project description:BackgroundThe development of predictive biomarkers for IGF targeted anti-cancer therapeutics remains a critical unmet need. The insulin receptor A isoform (InsR-A) has been identified as a possible biomarker candidate but quantification of InsR-A in widely available formalin fixed paraffin embedded (FFPE) tissues is complicated by its similarities with the metabolic signaling insulin receptor isoform B (InsR-B). In the present study, qPCR based assays specific for InsR-A, InsR-B and IGF-1R were developed for use in FFPE tissues and tested for feasible use in clinical archived FFPE estrogen receptor (ER)+and ER- breast cancer tumors.DesignFFPE compatible primer sets were designed with amplicon sizes of less than 60 base pairs and validated for target specificity, assay repeatability and amplification efficiency. FFPE tumors from ER+ (n=83) and ER-(n=64) primary untreated breast cancers, and ER+ hormone refractory (HR ER+) (n=61) breast cancers were identified for feasibility testing. The feasible use of InsR-A and InsR-B qPCRs were tested using all tumor groups and the feasibility of IGF-1R qPCR was determined using HR ER+ tumors.ResultsAll qPCR assays were highly reproducible with amplification efficiencies between 96-104% over a 6 log range with limits of detection of 4 or 5 copies per reaction. Greater than 90% of samples were successfully amplified using InsR-A, InsR-B or IGF-1R qPCR primer sets and greater than 88% of samples tested amplified both InsR isoforms or both isoforms and IGF-1R. InsR-A was the predominant isoform in 82% ER+, 68% ER- and 100% HR ER+ breast cancer. Exploratory analyses demonstrated significantly more InsR-A expression in ER+ and HR ER+ groups compared to InsR-B (ER+ p<0.05, HR ER+ p<0.0005) and both groups had greater InsR-A expression when compared to ER- tumors (ER+ p<0.0005, HR ER+ p<0.05). IGF-1R expression of HR ER+ tumors was lower than InsR-A (p<0.0005) but higher than InsR-B (p<0.0005). The InsR-B expression of HR ER+ tumors was significantly reduced compared other tumor subgroups (ER+ and ER-, p<0.0005) and lead to a significant elevation of HR ER+ InsR-A: InsR-B ratios (ER+ and ER-, p<0.0005).ConclusionsThe validated, highly sensitive InsR-A and InsR-B qPCR based assays presented here are the first to demonstrate the feasible amplification of InsR isoforms in FFPE tissues. Quantification data generated from this feasibility study indicating InsR-A is more predominant than InsR-B in breast cancer support the use of these assays for further investigation of InsR-A and InsR-B as predictive biomarkers for IGF targeted therapeutics.

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Altered expression of insulin receptor isoforms in breast cancer.

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Altered expression of insulin receptor isoforms in breast cancer.

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