Project description:Background: Interval breast cancers can occur through failure to detect an abnormality at the time of screening (missed interval cancer), or as a new event after a negative screen (true interval cancer). The development and progression of true interval tumors (TIBC) is known to be different than screen-detected tumors (SDBC). However, much work still needs to be done to understand the biological characteristics and clinical behaviour of these TIBC. Objectives: To characterize the gene expression profile in TIBC and SDBC aimed to identify biological markers that may be associated with the emergence of symptomatic breast cancer in the screening interval. Material and Methods: An unsupervised exploratory gene expression profile analysis was performed among 10 samples (discovery set, TIBC=5 and SDBC=5) using Affymetrix Human Gene 1.0 ST arrays and interpreted by Ingenuity Pathway Analysis. Differential expression of selected genes was confirmed in validation series of 91 patients (TIBC=12 and SDBC=79) by immunohistochemistry and 24 patients (TIBC=8 and SDBC=16) by RT-qPCR, expanding the analysis to other genes in same pathway (mTOR, 4E-BP1, eIF-4G and S6). Results: Exploratory gene expression analysis identified 1060 transcripts with a p value <0.05 and 132 transcripts with an adjusted p value <0.01 difference between TIBC and SDBC samples. Based on biological implications in breast cancer, four genes were selected for further validation: CP (ceruloplasmin) and RPS6KB2 (ribosomal protein S6 kinase, 70kDa, polypeptide 2) both up-regulated in TIBC vs SDBC and PTEN (phosphatase and tensin homolog) and TGFBR3 (transforming growth factor beta receptor III), down-regulated in TIBC vs SDBC. Their differential expression was confirmed by RT-qPCR and immunohistochemistry, suggesting mTOR pathway overexpression in TIBC at both mRNA and protein level. Further expanded analysis by immunohistochemistry for mTOR pathway activation, including expression of phosphorylated forms of mTOR, 4E-BP1, eIF-4G, RPS6KB2 and S6, confirmed the upregulation of this pathway in TIBC. Conclusions: TIBC and SDBC shows differential expression profile both at the gene and protein levels. The mTOR signaling is significantly upregulated in TIBC compared with SDBC, suggesting an enhanced aggressiveness of TIBC. Besides, CP may also represent novel immunohistochemical marker helpful in distinguishing between TIBC and SDBC. Further studies with larger sets of patients are guaranteed to verify these findings associated with TIBC.

Project description:Background: Interval breast cancers can occur through failure to detect an abnormality at the time of screening (missed interval cancer), or as a new event after a negative screen (true interval cancer). The development and progression of true interval tumors (TIBC) is known to be different than screen-detected tumors (SDBC). However, much work still needs to be done to understand the biological characteristics and clinical behaviour of these TIBC. Objectives: To characterize the gene expression profile in TIBC and SDBC aimed to identify biological markers that may be associated with the emergence of symptomatic breast cancer in the screening interval. Material and Methods: An unsupervised exploratory gene expression profile analysis was performed among 10 samples (discovery set, TIBC=5 and SDBC=5) using Affymetrix Human Gene 1.0 ST arrays and interpreted by Ingenuity Pathway Analysis. Differential expression of selected genes was confirmed in validation series of 91 patients (TIBC=12 and SDBC=79) by immunohistochemistry and 24 patients (TIBC=8 and SDBC=16) by RT-qPCR, expanding the analysis to other genes in same pathway (mTOR, 4E-BP1, eIF-4G and S6). Results: Exploratory gene expression analysis identified 1060 transcripts with a p value <0.05 and 132 transcripts with an adjusted p value <0.01 difference between TIBC and SDBC samples. Based on biological implications in breast cancer, four genes were selected for further validation: CP (ceruloplasmin) and RPS6KB2 (ribosomal protein S6 kinase, 70kDa, polypeptide 2) both up-regulated in TIBC vs SDBC and PTEN (phosphatase and tensin homolog) and TGFBR3 (transforming growth factor beta receptor III), down-regulated in TIBC vs SDBC. Their differential expression was confirmed by RT-qPCR and immunohistochemistry, suggesting mTOR pathway overexpression in TIBC at both mRNA and protein level. Further expanded analysis by immunohistochemistry for mTOR pathway activation, including expression of phosphorylated forms of mTOR, 4E-BP1, eIF-4G, RPS6KB2 and S6, confirmed the upregulation of this pathway in TIBC. Conclusions: TIBC and SDBC shows differential expression profile both at the gene and protein levels. The mTOR signaling is significantly upregulated in TIBC compared with SDBC, suggesting an enhanced aggressiveness of TIBC. Besides, CP may also represent novel immunohistochemical marker helpful in distinguishing between TIBC and SDBC. Further studies with larger sets of patients are guaranteed to verify these findings associated with TIBC. 5 TIBC samples where compared with 5 SDBC

Project description:BACKGROUND:Heterogeneity of immune gene expression patterns of luminal breast cancer (BC), which is clinically heterogeneous and overall considered as low immunogenic, has not been well studied especially in non-European populations. Here, we aimed at characterizing the immune gene expression profile of luminal BC in an Asian population and associating it with patient characteristics and tumor genomic features. METHODS:We performed immune gene expression profiling of tumor and adjacent normal tissue in 92 luminal BC patients from Hong Kong using RNA-sequencing data and used unsupervised consensus clustering to stratify tumors. We then used luminal patients from The Cancer Genome Atlas (TCGA, N?=?564) and a Korean breast cancer study (KBC, N?=?112) as replication datasets. RESULTS:Based on the expression of 130 immune-related genes, luminal tumors were stratified into three distinct immune subtypes. Tumors in one subtype showed higher level of tumor-infiltrating lymphocytes (TILs), characterized by T cell gene activation, higher expression of immune checkpoint genes, higher nonsynonymous mutation burden, and higher APOBEC-signature mutations, compared with other luminal tumors. The high-TIL subtype was also associated with lower ESR1/ESR2 expression ratio and increasing body mass index. The comparison of the immune profile in tumor and matched normal tissue suggested a tumor-derived activation of specific immune responses, which was only seen in high-TIL patients. Tumors in a second subtype were characterized by increased expression of interferon-stimulated genes and enrichment for TP53 somatic mutations. The presence of three immune subtypes within luminal BC was replicated in TCGA and KBC, although the pattern was more similar in Asian populations. The germline APOBEC3B deletion polymorphism, which is prevalent in East Asian populations and was previously linked to immune activation, was not associated with immune subtypes in our study. This result does not support the hypothesis that the germline APOBEC3B deletion polymorphism is the driving force for immune activation in breast tumors in Asian populations. CONCLUSION:Our findings suggest that immune gene expression and associated genomic features could be useful to further stratify luminal BC beyond the current luminal A/B classification and a subset of luminal BC patients may benefit from checkpoint immunotherapy, at least in Asian populations.

Project description:Normal-appearing epithelium of cancer patients can harbor occult genetic abnormalities. Data comprehensively comparing gene expression between histologically normal breast epithelium of breast cancer patients and cancer-free controls are limited. The present study compares global gene expression between these groups. We performed microarrays using RNA from microdissected histologically normal terminal ductal-lobular units (TDLU) from 2 groups: (i) cancer normal (CN) (TDLUs adjacent to untreated ER1 breast cancers (n = 14)) and (ii) reduction mammoplasty (RM) (TDLUs of age-matched women without breast disease (n = 15)). Cyber-T identi?ed differentially expressed genes. Quantitative RT-PCR (qRT-PCR), immunohistochemistry (IHC), and comparison to independent microarray data including 6 carcinomas in situ (CIS), validated the results. Gene ontology (GO), UniProt and published literature evaluated gene function. About 127 probesets, corresponding to 105 genes, were differentially expressed between CN and RM (p < 0.0009, corresponding to FDR <0.10). 104/127 (82%) probesets were also differentially expressed between CIS and RM, nearly always (102/104 (98%)) in the same direction as in CN vs. RM. Two-thirds of the 105 genes were implicated previously in carcinogenesis. Overrepresented functional groups included transcription, G-protein coupled and chemokine receptor activity, the MAPK cascade and immediate early genes. Most genes in these categories were under-expressed in CN vs. RM. We conclude that global gene expression abnormalities exist in normal epithelium of breast cancer patients and are also present in early cancers. Thus, cancer-related pathways may be perturbed in normal epithelium. These abnormalities could be markers of disease risk, occult disease, or the tissue’s response to an existing tumor. Experiment Overall Design: 29 samples from histologically normal microdissected breast epithelium are included in this series. 14 samples are from epithelium adjacent to a breast tumor, 15 samples were obtained from patients undergoing reduction mammoplasty without apparent breast cancer

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Gene expression profiling in true interval breast cancer reveals overactivation of mTOR signalling pathway

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