Nature Of Triple Negative Breast Cancer Research Articles

Time and Space Dual-Blockade Strategy for Highly Invasive Nature of Triple-Negative Breast Cancer in Enhanced Sonodynamic Therapy Based on Fe-MOF Nanoplatforms.

Triple-negative breast cancer (TNBC), due to its high malignant degree and strong invasion ability, leads to poor prognosis and easy recurrence, so effectively curbing the invasion of TNBC is the key to obtaining the ideal therapeutic effect. Herein, a therapeutic strategy is developed that curbs high invasions of TNBC by inhibiting cell physiological activity and disrupting tumor cell structural function to achieve the time and space dual-blockade. The time blockade is caused by the breakthrough of the tumor-reducing blockade based on the ferroptosis process and the oxidation-toxic free radicals generated by enhanced sonodynamic therapy (SDT). Meanwhile, alkyl radicals from 2,2'-azobis[2-(2-imidazolin-2-yl)propane] dihydrochloride (AIPH) and 1 O2 attacked the organelles of tumor cells under ultrasound (US), reducing the physiological activity of the cells. The attack of free radicals on the cytoskeleton, especially on the proteins of F-actin and its assembly pathway, achieves precise space blockade of TNBC. The damage to the cytoskeleton and the suppression of the repair process leads to a significant decline in the ability of tumor cells to metastasize and invade other organs. In summary, the FTM@AM nanoplatforms have a highly effective killing and invasion inhibition effect on invasive TNBC mediated by ultrasound, showcasing promising clinical transformation potential.

Therapeutic Role of Tamoxifen for Triple-Negative Breast Cancer: Leveraging the Interaction Between ERβ and Mutant p53.

The absence of effective therapeutic targets and aggressive nature of triple-negative breast cancer (TNBC) renders this disease subset difficult to treat. Although estrogen receptor beta (ERβ) is expressed in TNBC, studies on its functional role have yielded inconsistent results. However, recently, our preclinical studies, along with other observations, have shown the potential therapeutic utility of ERβ in the context of mutant p53 expression. The current case study examines the efficacy of the selective estrogen receptor modulator tamoxifen in p53-mutant TNBC with brain metastases. Significant increase in ERβ protein expression and anti-proliferative interaction between mutant p53 and ERβ were observed after cessation of tamoxifen therapy, with significant regression of brain metastases. This case study provides supporting evidence for the use of tamoxifen in p53-mutant, ERβ+TNBC, especially in the setting of brain metastasis.

Abstract 1541: LncRNA MALAT1 as a potential therapeutic target in triple negative breast cancer

Abstract In the United States 1 in 8 women will be diagnosed with breast cancer within their lifetime, accounting for nearly 30% of cancer diagnoses annually. Although improvements have been made in early diagnoses and treatment, the heterogeneity of breast cancer often makes treatment of the disease a challenge. Triple negative breast cancer (TNBC) ranks as the 5th leading cause of cancer related death in women, while also being more prevalent in younger African American and Hispanic premenopausal women. The invasive nature of TNBC increases the risk of metastasis and recurrence, and patients will often acquire resistance to existing treatment options, emphasizing the need for new breast cancer therapies. A novel therapeutic approach is the inhibition of the highly abundant long noncoding RNA (lncRNA), metastasis-associated lung adenocarcinoma transcript (MALAT1) using an antisense oligonucleotide (ASO). MALAT1 has been shown to be upregulated in several breast cancer subtypes, increasing the migratory and invasive properties of tumor cells. Using a MALAT1 ASO in our preclinical syngeneic p53 null TNBC tumor mouse models, we were able to significantly knockdown MALAT1 RNA expression and observed a delay in primary tumor growth as well as an improvement in tumor response when used in combination with the chemotherapy drugs carboplatin or docetaxel. Additionally, immunophenotyping of tumor infiltrating lymphocytes show that MALAT1 inhibition alters the tumor microenvironment with a significant decrease in immunosuppressive CD206+ Arginase+ macrophages and myeloid derived suppressor cells (MDSC) as well as an increase in cytotoxic CD8+ T cells. The immuno-stimulatory effects of MALAT1 inhibition highlight the benefit of a MALAT1 ASO therapeutic and its potential to be used in combination with current chemotherapies and/or developing immunotherapies. Citation Format: Oluwatoyosi Adewunmi, Jeffrey Rosen. LncRNA MALAT1 as a potential therapeutic target in triple negative breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1541.

Stromal Characteristics and Impact on New Therapies for Metastatic Triple-Negative Breast Cancer.

Simple SummaryIn this review, we summarize the recent outcomes from clinical trials with new agents for metastatic triple-negative breast cancer, specifically focusing on immunotherapies targeting the PD-1/PD-L1 pathway and antibody–drug conjugates. In addition to the clinical data supporting these therapies, we review the impact of the tumor microenvironment on the effectiveness of these therapies. The heterogenous nature of triple-negative breast cancer (TNBC) is an underlying factor in therapy resistance, metastasis, and overall poor patient outcome. The lack of hormone and growth factor receptors lends to the use of chemotherapy as the first-line treatment for TNBC. However, the failure of chemotherapy demonstrates the need to develop novel immunotherapies, antibody–drug conjugates (ADCs), and other tumor- and stromal-targeted therapeutics for TNBC patients. The potential for stromal-targeted therapy is driven by studies indicating that the interactions between tumor cells and the stromal extracellular matrix (ECM) activate mechanisms of therapy resistance. Here, we will review recent outcomes from clinical trials targeting metastatic TNBC with immunotherapies aimed at programed death ligand–receptor interactions, and ADCs specifically linked to trophoblast cell surface antigen 2 (Trop-2). We will discuss how biophysical and biochemical cues from the ECM regulate the pathophysiology of tumor and stromal cells toward a pro-tumor immune environment, therapy resistance, and poor TNBC patient outcome. Moreover, we will highlight how ECM-mediated resistance is motivating the development of new stromal-targeted therapeutics with potential to improve therapy for this disease.

New and Emerging Targeted Therapies for Advanced Breast Cancer.

In the United States, breast cancer is among the most frequently diagnosed cancers in women. Breast cancer is classified into four major subtypes: human epidermal growth factor receptor 2 (HER2), Luminal-A, Luminal-B, and Basal-like or triple-negative, based on histopathological criteria including the expression of hormone receptors (estrogen receptor and/or progesterone receptor) and/or HER2. Primary breast cancer treatments can include surgery, radiation therapy, systemic chemotherapy, endocrine therapy, and/or targeted therapy. Endocrine therapy has been shown to be effective in hormone receptor-positive breast cancers and is a common choice for adjuvant therapy. However, due to the aggressive nature of triple-negative breast cancer, targeted therapy is becoming a noteworthy area of research in the search for non-endocrine-targets in breast cancer. In addition to HER2-targeted therapy, other emerging therapies include immunotherapy and targeted therapy against critical checkpoints and/or pathways in cell growth. This review summarizes novel targeted breast cancer treatments and explores the possible implications of combination therapy.

Abstract PO-150: Comparison of RUNX1, RUNX2, RUNX3 and CBFβ gene expression in breast tumors Indicate ethnic differences and similarities by receptor status

Abstract The RUNX family of transcription factors RUNX1, RUNX2 and RUNX3 encode nuclear proteins with a Runt DNA-binding domain that functions in a complex with their obligate partner core-binding factor CBFβ to regulate gene expression. RUNX genes are known for paradoxical roles, with oncogenic as well as tumor suppressor functions, dependent on context. Interestingly, RUNX2 has been shown to play a potentially important functional role in Triple-Negative Breast Cancer (TNBC), and RUNX1 expression associates with poor prognosis in TNBC in a cohort of women of European descent. Women of African descent experience more aggressive TNBC with higher mortality rates than their women of European ancestry counterparts. TNBC is the most difficult breast cancer subtype to treat globally and has a poor prognosis and low survival which remains an important clinical challenge. There is a paucity of genomics studies interrogating TNBC of diverse populations. Hence, RNAseq datasets of 782 breast cancer patients were leveraged from a web-based platform -CbioPortal and published literature. 685 patients were queried for the RUNX1, RUNX2, RUNX3, and CBFβ genes, and an additional 97 patient data were harnessed from research publications. The cohort includes 595 (70%) patients of European ancestry (EA), 90 (11%) of African descent (AD), both from The Cancer Genome Atlas (TCGA), and 97 (13%) patients indigenous African from Nigeria (IAN). The queried genes were compared for race/ethnic disparity by estrogen receptor status (er). Mutual exclusivity was analyzed and P-values were determined by Fisher's extract test with the null hypothesis that a pair of genes alteration frequency of occurrence is proportional to uncorrelated occurrence in each gene. The queried genes enrichment in TP53, PIK3CA, CDH1, MAP3K1, and GATA3 between patients of European ancestry and African descent were analyzed and the Kaplan-Meier plots with P-value from the Long rank test were determined. The queried genes were altered in 69 patients (12%) of EA and 9 patients of AD due to unavailability of data on RUNX2 and RUNX 3 in IAN only RUNX1 and CBfβ was altered in 4 (4%) patients of 97 patients. Ethnic disparities and similarities were indicated in the queried altered genes in RUNX1 and RUNX 2 and showed similar expression in ER-negative correlating with the TNBC poor prognosis however RUNX3 genes were silenced in that of AD and was expressed in EA. They were differences in mutual exclusivity and co-occurrence but were not statistically significant, this may be due to the small data size. Interestingly, there was statistically significant equal gene enrichment of RUNX1 and CBfβ in CDH1mutations of both EA and AD. Larger cohorts in particular the AD cohort is needed, to further elucidate some of these borderline findings. In conclusion, our findings are insightful towards deciphering the characteristic aggressive nature of TNBC in ethnic disparity. Citation Format: Uzoamaka A. Okoli. Comparison of RUNX1, RUNX2, RUNX3 and CBFβ gene expression in breast tumors Indicate ethnic differences and similarities by receptor status [abstract]. In: Proceedings of the AACR Virtual Conference: 14th AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2021 Oct 6-8. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2022;31(1 Suppl):Abstract nr PO-150.

Circular RNA-associated ceRNA network involved in HIF-1 signalling in triple-negative breast cancer: circ_0047303 as a potential key regulator.

The aggressive and highly metastatic nature of triple‐negative breast cancer (TNBC) causes patients to suffer from the poor outcome. HIF‐1 signalling pathway is a prominent pathway that contributes to angiogenesis and metastasis progression in tumours. On the contrary, the undeniable importance of circular RNAs (circRNAs) as multifunctional non‐coding RNAs (ncRNAs) has been identified in breast cancer. These ncRNAs owing to their high stability and specificity have been becoming a hotspot in cancer researches. circRNAs act as competing endogenous RNAs (ceRNAs) and compete with mRNAs for shared miRNAs, thus modulate gene expression. Since the most dysregulated biological functions in TNBC are associated with cellular invasion, understanding the molecular pathogenesis of these processes is a crucial step towards the development of new treatment approaches. The purpose of this study is to undermine the circRNA‐associated ceRNA network involved in HIF‐1 signalling in TNBC using an integrative bioinformatics approach. In the next step, the novel circ_0047303‐mediated ceRNA regulatory axes have been extracted and validated across TNBC samples. We show that circ_0047303 has the highest degree in the circRNA‐associated ceRNA network and shows a significant up‐expression in TNBC. Moreover, our results suggest that circ_0047303 could mediate the upregulation of key angiogenesis‐related genes, including HIF‐1, EIF4E2 and VEGFA in TNBC through sponging the tumour‐suppressive miRNAs. The circ_0047303 could be a promising molecular biomarker and/or therapeutic target for TNBC.

Precision Medicine and Triple-Negative Breast Cancer: Current Landscape and Future Directions.

Simple SummaryThe implementation of precision medicine will revolutionize cancer treatment paradigms. Notably, this goal is not far from reality: genetically similar cancers can be treated similarly. The heterogeneous nature of triple-negative breast cancer (TNBC) made it a suitable candidate to practice precision medicine. Using TNBC molecular subtyping and genomic profiling, a precision medicine-based clinical trial is ongoing. This review summarizes the current landscape and future directions of precision medicine and TNBC.Triple-negative breast cancer (TNBC) is an aggressive and heterogeneous subtype of breast cancer associated with a high recurrence and metastasis rate that affects African-American women disproportionately. The recent approval of targeted therapies for small subgroups of TNBC patients by the US ‘Food and Drug Administration’ is a promising development. The advancement of next-generation sequencing, particularly somatic exome panels, has raised hopes for more individualized treatment plans. However, the use of precision medicine for TNBC is a work in progress. This review will discuss the potential benefits and challenges of precision medicine for TNBC. A recent clinical trial designed to target TNBC patients based on their subtype-specific classification shows promise. Yet, tumor heterogeneity and sub-clonal evolution in primary and metastatic TNBC remain a challenge for oncologists to design adaptive precision medicine-based treatment plans.

Abstract P4-08-03: Tumor suppressors p53 and ARF control ADAR1-driven tumorigenicity in triple negative breast cancer

Abstract Research Overview and Objective Triple-negative breast cancer (TNBC) accounts for one-fifth of the breast cancer patient population. The heterogeneous nature of TNBC and lack of options for targeted therapy make its treatment a constant challenge. The co-deficiency of tumor suppressors p53 and ARF is a significant genetic signature enriched in TNBC, but it is not yet clear how TNBC is regulated by this genetic alteration. Methods To answer this question, we established p53/ARF-defective murine embryonic fibroblast (MEF) to study the molecular and phenotypic consequences in vitro. Moreover, transgenic mice were generated to investigate the effect of p53/ARF deficiency on mammary tumor development in vivo. Results Increased transformation capability was observed in p53/ARF-defective cells, and formation of aggressive mammary tumors was also seen in p53−/−ARF−/− mice. RNA-editing enzyme ADAR1 was identified as a potential mediator for the elevated oncogenic potential. Interestingly, we found that the overexpression of ADAR1 is also prevalent in human TNBC cell lines and patient specimen. Using short hairpin RNA (shRNA) to reduce ADAR1 expression abrogated the oncogenic potential of human TNBC cell lines, while non-TNBC cells are less susceptible. Different levels of RNA editing of known ADAR1 targets were detected in shRNA-treated human TNBC cell lines, suggesting that ADAR1-mediated RNA editing contributes to TNBC pathogenesis. Implication/Discussion These results indicate critical roles played by the tumor suppressors p53 and ARF in the pathogenesis of TNBC, partially through affecting ADAR1-mediated RNA editing. Further understanding of this pathway could shed light on potential vulnerabilities of TNBC and inform the development of personalized therapies based on patients’ genetic signiatures. Citation Format: Che-Pei Kung, Catherine E Kuzmicki, Emily A Bross, Sua Ryu, Eric Freeman, Thwisha Sabloak, Emily R Bramel, Leonard B Maggi Jr, Jason D Weber. Tumor suppressors p53 and ARF control ADAR1-driven tumorigenicity in triple negative breast cancer [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P4-08-03.

Flubendazole elicits anti-metastatic effects in triple-negative breast cancer via STAT3 inhibition.

Tumor metastasis remains the cause of 90% of cancer-related deaths. Cancer stem cells (CSC) are thought to be responsible for the aggressive and metastatic nature of triple-negative breast cancers (TNBC), and new therapeutic strategies are being devised to target them. Flubendazole (FLU) is a widely used anthelmintic agent that also exhibits anticancer activity in several cancer types. The aim of this study was to characterize the mechanism of action of FLU on breast cancer stem cell (BCSC)-like properties and metastasis in TNBC. FLU treatment caused a significant induction of apoptosis, accompanied by G2/M phase accumulation, caspase-3/-7 activation and the dysregulation of STAT3 activation in TNBC cells. The latter phenomenon was associated with impairment of cancer stem-like traits, concomitant with a reduction in the CD24low /CD44high , CD24high /CD49fhigh subpopulation, ALDH1 activity and mammosphere formation. The BCSC-enriched populations exhibited enhanced metastasis with higher STAT3 activation, while FLU administration inhibited tumor growth, angiogenesis and lung and liver metastasis, coinciding with decreased MMP-2 and MMP-9 levels in circulating blood. FLU kills not only rapid proliferating tumor cells but also effectively eradicates BCSC-like cells in vitro and in vivo. Our findings warrant further investigation of FLU as a treatment for metastatic TNBC.

Microarray-based SNP genotyping to identify genetic risk factors of triple-negative breast cancer (TNBC) in South Indian population.

In the view of aggressive nature of Triple-Negative Breast cancer (TNBC) due to the lack of receptors (ER, PR, HER2) and high incidence of drug resistance associated with it, a case-control association study was conducted to identify the contributing genetic risk factors for Triple-negative breast cancer (TNBC). A total of 30 TNBC patients and 50 age and gender-matched controls of Indian origin were screened for 9,00,000 SNP markers using microarray-based SNP genotyping approach. The initial PLINK association analysis (p<0.01, MAF 0.14-0.44, OR 10-24) identified 28 non-synonymous SNPs and one stop gain mutation in the exonic region as possible determinants of TNBC risk. All the 29 SNPs were annotated using ANNOVAR. The interactions between these markers were evaluated using Multifactor dimensionality reduction (MDR) analysis. The interactions were in the following order: exm408776>exm1278309>rs316389>rs1651654>rs635538>exm1292477. Recursive partitioning analysis (RPA) was performed to construct decision tree useful in predicting TNBC risk. As shown in this analysis, rs1651654 and exm585172 SNPs are found to be determinants of TNBC risk. Artificial neural network model was used to generate the Receiver operating characteristic curves (ROC), which showed high sensitivity and specificity (AUC-0.94) of these markers. To conclude, among the 9,00,000 SNPs tested, CCDC42 exm1292477, ANXA3 exm408776, SASH1 exm585172 are found to be the most significant genetic predicting factors for TNBC. The interactions among exm408776, exm1278309, rs316389, rs1651654, rs635538, exm1292477 SNPs inflate the risk for TNBC further. Targeted analysis of these SNPs and genes alone also will have similar clinical utility in predicting TNBC.

Abstract B50: Elucidating the roles of Kaiso in the metastasis of triple-negative breast cancer cells

Abstract The triple negative breast cancer (TNBC) subtype represent a subset of breast tumours that lack expression of the estrogen-receptor, progesterone-receptor and human epidermal growth factor receptor-2. Interestingly, TNBC is more prevalent in young women of African ancestry (WAA) than other ethnicities, and WAA have a high mortality despite a low BC incidence compared to Caucasian women. This high mortality rate is attributed to the highly aggressive and metastatic nature of triple-negative tumors and limited treatment options or targeted-therapies. The metastatic nature of TNBC suggests a deregulation of the epithelial-to-mesenchymal (EMT) pathway that is often implicated in tumor progression. During EMT, cells downregulate the tumor suppressor and cell adhesion protein E-cadherin and re-express mesenchymal proteins such as N-cadherin and vimentin, which endow the cells with increased motility. During the course of our characterization of the unique transcription factor Kaiso that was implicated as a potential regulator of E-cadherin, we found that increased nuclear expression of Kaiso is significantly correlated with basal/TNBCs. However, little is known about the role of Kaiso in EMT or TNBC metastasis. Thus, the goal of our research is to elucidate the function of Kaiso in TNBC metastasis. To gain insight into Kaiso's role in TNBC metastasis we generated stable Kaiso-depleted TNBC cells (MDA-MB-231) using a Kaiso specific shRNA. Stable Kaiso-depleted cells were then analyzed for the gene expression profile of key EMT markers and for their migratory and invasive capacities in vitro using wound healing and Matrigel invasion assays. We found that Kaiso is highly expressed at both the transcript and protein level in breast tumor cells and tissues. Kaiso-depletion reduced the migratory and invasive capacities of TNBC cells, and induced the down-regulation of potent E-cadherin regulators including Slug and ZEB1 at both the transcript and protein levels. Interestingly, we observed that Kaiso-depletion also resulted in the downregulation of TGFβR1 and TGFβR2; key effectors of the TGFβ signaling pathway. Since TGFβ signaling is a potent inducer of EMT in late stage breast cancer via up-regulation of Snail and/or Slug and ZEB1/2, our results suggests that Kaiso may be promoting TNBC metastasis via the TGFβ signaling pathway. Kaiso's expression might thus be a useful biomarker for TNBC prognosis. Citation Format: Blessing I. Bassey, Catherine Crawford-Brown, Robin Hallet, Juliet Daniel. Elucidating the roles of Kaiso in the metastasis of triple-negative breast cancer cells. [abstract]. In: Proceedings of the Seventh AACR Conference on The Science of Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; Nov 9-12, 2014; San Antonio, TX. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2015;24(10 Suppl):Abstract nr B50.

Abstract P6-08-01: Endothelial-Like Phenotype of Triple-Negative Breast Carcinoma Cells and Implications for New Molecular Targets

Abstract Background. Triple Negative Breast Cancers (TNBCs) still represent a question mark in breast cancer biology and a primary issue in clinics. A promising approach for an efficient targeted treatment of TNBCs seems to be represented by antiangiogenic therapies. Methods. Effects of different compounds on proliferation of TNBC was evaluated in vivo and in vitro respectively by xenograft tumor volume analysis and SRB assay. Vasculogenic Mimicry (VM) properties were evaluated in vitro and in vivo respectively by tube formation assay and Transmission Electron Microscopy (TEM). Results. Treatment of triple negative (TN) MDA-MB-231 xenografts with Sunitinib induced tumor regression, versus only a slight growth inhibition in tumors derived from MCF7 luminal cell line, whereas Bevacizumab determined only a modest decrease in tumor growth in both models. Accordingly, the efficacy of Sunitinib in blocking in vitro growth of breast cancer cell lines was higher in MDA-MB-231 cells in comparison with MCF7 cells (IC50 at 72h in MDA-MB-231 was 5 uM vs 25 uM in MCF7, as evaluated by SRB), and sensitivity to Bevacizumab was comparable between the two different cell lines. Investigating the undifferentiated nature of TNBCs, we observed that these tumors present an endothelial like phenotype and behavior, as supported respectively by the expression of endothelial markers, and the formation of vascular-like channels in vitro. In fact, all six TN cell lines evaluated (MDA-MB-231, MDA-MB-157, MDA-MB-468, BT-549, BT-20 and HCC1937) were able to form vascular channels when seeded on the murine tumor-derived basement membrane (Matrigel), whereas luminal (MCF7, T47D and ZR-75-1) and HER2- positive (MDA-MB-361, BT474 and SKBr3) breast carcinoma cell lines did not exhibit vascular structures. Then, we evaluated the VM in xenograft tumors derived from MDA-MB-231, MCF7 and MDA-MB-361 using transmission electron microscopy (TEM). MDA-MB-231 xenografts displayed channel-like structures formed by tumor cells encompassing erytrocytes, whereas in MCF7 and MDA-MB-361 xenografts the endothelial lining delimiting blood vessels was clearly visible. Notably, blood vessels surrounded by tumor cells were also identified in human TN specimens processed for TEM, and these structures were significantly more frequent in TN compared to non-TN tumors. 60% reduction in TN vascular channel formation in vitro by an anti-bFGF monoclonal antibody along with no effect using anti-VEGF antibody indicated that TN breast carcinoma cells can generate vascular channels through bFGF-mediated pathway. Silencing of different receptors involved in bFGF signal (i.e. FGFR2 and PDGFR β) abrogated VM in TN cells. Finally, TNBC cells able to perform vascular-like channels were found to express significantly higher (p= 0.003) levels of FGFR related genes, described associated with basal-like BC aggressiveness, compared to all the other tested cell lines. Conclusions. Our findings point to the possibility that TNBC cells are maintained by proangiogenic signals and that increased sensitivity to Sunitinib probably relies on the specific impairment of PDGFRβ and FGFR2-mediated pathways, which might represent a possible specific therapeutic target. Partially supported by AIRC and Italian Bureau of Health. Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P6-08-01.

Differential Wnt pathway activation in triple negative breast cancers comparative to HER2 and hormone positive breast cancers identified from formalin-fixed, paraffin-embedded tissues.

Abstract Abstract #43 Background: Patients afflicted with triple negative (TN) breast cancer (BC) are typically younger and have the worst prognosis of any BC subtype as a result of its aggressive and invasive phenotype. TN BC has been well studied with respect to the associative and prognostic values of its mRNA expression profiles. We have used the Illumina DASL (cDNA mediated, Annealing, Selection, Extension, and Ligation) platform to characterize 502 genes from 96 BC patients sourced from formalin-fixed, paraffin-embedded (FFPE) tissues. RNA extracted from FFPE tissues is highly fragmented and typically unsuited for expression platforms, but recent advances have allowed for accurate expression profiling of RNA from FFPE tissues. This 96 patient cohort included 21 TN BC patients identified by immunohistochemistry (IHC); differential analysis of TN expression was integrated with known pathway mechanisms and identified activation of the Wnt pathway in TN tumors.&amp;#x2028; Material and Methods: The Illumina DASL assay was used to measure mRNA expression levels from BC patients obtained from St. Mary's Hospital, Montreal, Quebec. RNA was extracted from three 5µm tissue sections from FFPE blocks using the RNA High Pure Kit (Roche). Differential mRNA regulation was identified by Significance Analysis of Microarrays (SAM) software using a false discover rate (FDR, q-value &amp;lt; 0.01) or a Bonferroni corrected two-tailed student's t-test (p-value &amp;lt; 0.05). IHC concordance was measured by mRNA expression fold-change for positive versus negative IHC categories.&amp;#x2028; Results: 165 samples from 96 patients were characterized for mRNA expression on 1,488 probes across 502 genes in the DASL assay. Quality control limited the analysis to 150 samples from 91 patients over 500 genes. Comparison of the TN subtype relative to HER2 positive and hormone positive breast carcinomas identified up-regulation [fold-change] of MMP7 [2.9], MMP9 [2.0], MMP1 [1.8], NOTCH1 [1.5], FZD7 [1.5], PPARD [1.3], CCND2 [1.2], MET [1.3], MYC [1.5], and MYCN [2.0] in the TN subtype. Most of these genes are either a component or downstream target of the Wnt signaling pathway. Down-regulated relevant genes included ESR1 [-8.4], PGR [-3.9], ERBB2 [-3.4] as expected, as well as ERRB3, ERBB4, GLI3, TFF1, LAF4/AFF3, and AR.&amp;#x2028; Discussion: Down-regulation of TFF1 suggests inhibition of the ERK/MAPK pathways which have complex effects on cell-cell adhesion. In combination with over-expression of MMPs 1, 7, and 9, these changes may contribute to the invasive nature of TN BCs. GLI3 down-regulation may attenuate Hedgehog signaling potentially contributing to Wnt activation, further leading to cell migration, invasion, and decrease in cell adhesion to cognate integrin-matrix interaction Up-regulation of both the FZD7 receptor as well as the Wnt targets MMP7, MYC, MYCN, PPARD, and CCND2 strongly suggest Wnt pathway activation in the TN subtype. Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 43.