Project description:Ductal carcinoma in situ (DCIS) is a nonobligate precursor of invasive breast cancer. Its biological features, particularly its intratumoral heterogeneity, remain obscure. Moreover, mechanism of lymph node metastasis is unclear. To address this deficiency, we performed single-cell transcriptome profiling of DCIS, invasive ductal carcinoma (IDC) and lymph node metastasis. Single-cell transcriptome analysis revealed that breast cancer exhibits intratumoral heterogeneity at the transcriptional level, defining specific functions, and that DCIS has similar heterogeneity to IDC.
Project description:Metastasis remains the leading cause of death in breast cancer. However, little is known about the dynamic changes during the dissemination of breast cancer. Here, we generate single-cell RNA and spatial transcriptome of primary tumors and paired metastatic lymph nodes in 4 breast cancer patients. We identified a disseminated cancer cell cluster with high levels of oxidative phosphorylation (OXPHOS). We also noticed the transition between glycolysis and OXPHOS when dissemination initiates. Furthermore, this distinct cell cluster is distributed along the tumor’s leading edge.
Project description:In the present manuscript we show that T cells within tumors are highly radio-resistant compared to T cells in lymphoid organs. To better understand this different response to ionizing radiation (IR), gene expression is compared for T cells from lymph nodes and those from transplantable MC38 tumors, in mice treated or not with whole body irradiation. We find that T cells from tumors and lymph nodes respond similarly to IR in terms of gene expression, and this response is dominated by p53 signaling. In contrast, gene expression at baseline (i.e. in the absence of radiation) shows profound differences between T cells from tumors and T cells from lymph nodes, and suggests transcriptomic reprogramming of the intratumoral T cells in the tumor microenvironment .
Project description:As the differentiation state of the cells is affected by epigenetic modification regulation, the abnormal epigenetic modification is an important factor leading to the intratumoral heterogeneity. To further understand of the intratumoral epigenetic heterogeneity, we performed Reduced Representation Bisulfite Sequencing and RNA sequencing for multiple regions within a breast tumor and exhibited the epigenetic maps of different regions. We detected intratumoral heterogeneity-related molecular features and further explored the correlation between tumor heterogeneity and tumor hypoxic microenvironment.
Project description:As the differentiation state of the cells is affected by epigenetic modification regulation, the abnormal epigenetic modification is an important factor leading to the intratumoral heterogeneity. To further understand of the intratumoral epigenetic heterogeneity, we performed Reduced Representation Bisulfite Sequencing and RNA sequencing for multiple regions within a breast tumor and exhibited the epigenetic maps of different regions. We detected intratumoral heterogeneity-related molecular features and further explored the correlation between tumor heterogeneity and tumor hypoxic microenvironment.
Project description:To obtain more information about the lymph node metastasis of breast cancer cells, we selected the matched primary breast cancer (PC), positive lymph nodes (PL), and negative lymph nodes (NL) of the same patient to perform integrated analysis. The PC, PL, NL samples were analysed with single-cell RNA sequencing.
Project description:To obtain more information about the lymph node metastasis of breast cancer cells, we selected the matched positive lymph nodes (PL), and negative lymph nodes (NL) of the same patient to perform integrated analysis. The PL, NL samples were analysed with single-cell ATAC sequencing.
Project description:Background Breast cancer patients who present in the early stage of disease are affected by metastasis to the axillary group of lymph nodes. The first among this group that is affected is called as sentinel lymph node, and metastasis to this lymph node is crucial for the staging of cancer and the quality of surgical intervention. Sentinel Lymph Node Biopsy (SLNB) that is currently used to assess lymph node metastasis is neither sensitive, nor specific, is time-consuming, thereby necessitating the identification of novel biomarkers that can flag sentinel lymph node metastasis. Methods Breast cancer patients were screened, and those with early stage were recruited in the study. Surgical resection of the breast was followed by identification of sentinel lymph nodes by methylene fluorescent technique. Histo-pathology of fresh frozen section biopsy was used as the gold standard to assign the clinical phenotypes of metastatic (SLNM+) and non-metastatic sentinel lymph nodes (SLNM-). Discovery phase of the experiment included isobaric Tags for Relative and Absolute Quantitation (iTRAQ) technique comprising of six comparative experiments coupled with mass spectrometric analysis on Orbitrap Fusion to identify differentially expressed proteins on Proteome Discoverer 2.4. Functional enrichment and pathway analyses of differentially regulated genes was carried out in DAVID functional annotation tool. Validation was done by ELISA and protein concentrations were used to estimate the ROC for computing diagnostic parameters. Results Based on MS/MS spectra there were 2396 unique protein groups and 81 differentially expressed proteins comparing SLNB + and SLNB -. Nineteen proteins up-regulated, and eight proteins that were down regulated in SLNB+ as compared to SLNB-. Bioinformatic analysis showed the implication of extra cellular matrix proteins and ECM-receptor interaction pathways to be implicated in lymph node metastasis. ELISA confirmed the up-regulation of caveolin 1, collagen α-1, desmin, fibrillin-1, and microfibrillar associated glycoprotein 4 in metastatic, as compared to non-metastatic lymph nodes. These proteins are known to be integral in tumorogenesis, cell proliferation, invasion, cell survival and anti-apoptosis. These proteins have 80%-100%, of sensitivity and specificity to differentiate the two clinical phenotypes. Conclusion Identified extra cellular matrix protein biomarkers have requisite diagnostic parameters to be developed as a translational tool to assess the status of sentinel lymph nodes during mastectomy procedure to guide surgical therapy of axillary lymph nodes in early breast oncology.
Project description:Analysis of purified immune and breast tumor cells from three major compartments where cancer and immune cells interact: primary tumor, tumor draining lymph nodes (tumor invaded or tumor free), and peripheral blood. The results suggests that node-positive patients’ immune regulation and functionality is down-regulated compared to node-negative patients.
Project description:Analysis of purified immune and breast tumor cells from three major compartments where cancer and immune cells interact: primary tumor, tumor draining lymph nodes (tumor invaded or tumor free), and peripheral blood. The results suggests that node-positive patients’ immune regulation and functionality is down-regulated compared to node-negative patients. CD45+ Immune and ESA+ tumor cells were purified from breast cancer patients' primary tumor, tumor-draining lymph node, and peripheral blood (ficoll) and placed onto Agilent microarrays using the dye-swap method. A universal human reference was used as a reference for the patient samples.
