Project description:Deregulation of Src kinases is associated with cancer. We previously showed that SrcDN conditional expression in MCF7 cells diminished tumorigenesis and causes tumor regression in mice. However, it remained unclear whether SrcDN affected breast cancer stem cell functionality or it reduced tumor mass. Here, we address this question by isolating an enriched population of BCSCs (ESA+-CD44+-CD24-) and the tumor-differentiated cells (ESA+-CD44+-CD24+) from MCF7-Tet-On-SrcDN. ESA+-CD44+-CD24- grew in suspension forming mammospheres, and producing tumors in nude mice, while ESA+-CD44+-CD24+ were poorly/non-tumorigenic. Doxycycline-induction of SrcDN inhibited BCSC tumorigenesis, selfrenewal, and stem-cell markers expression. SrcDN significantly inhibited SFE, and stem-cell markers expression in triple-negative breast cancer (TNBC) MDA-MB-231 and SUM159PT cells. Inducible depletion of c-Src caused similar effects in MDA-MB-231 cells. In MCF7-Tet-On-SrcDN derived mammospheres SrcDN-induction inhibited expression, and activity of hexokinase, pyruvate kinase and lactate dehydrogenase, resulting in diminished glucose consumption and lactate production, which restricted Warburg effect. Thus, c-Src functionality is important for breast cancer stem cell maintenance and renewal, tumorigenicity, and stem cell transcription factor expression, effects linked to glucose metabolism reduction.
Project description:The cancer stem cell model maintains that tumors are organized in a hierarchy driven by tumor initiating cells (TICs), and that patient survival inversely correlates with TIC gene expression. Here we generated a prognostic signature for HER2+ breast cancer from TICs purified from MMTV-Her2/Neu mammary tumors. TICs from this model, identified as Lin-:CD24+:JAG1- at a frequency of 2-5% by serial and single cell transplantation assays, showed elevated expression of proliferation genes and low expression of differentiation genes (compared to non-TIC fraction CD24- of the same tumor). We used microarrays to detect differentially expressed genes in the TIC fraction compared to the non-TIC fraction of the same tumor
Project description:The cancer stem cell model maintains that tumors are organized in a hierarchy driven by tumor initiating cells (TICs), and that patient survival inversely correlates with TIC gene expression. Here we generated a prognostic signature for HER2+ breast cancer from TICs purified from MMTV-Her2/Neu mammary tumors. TICs from this model, identified as Lin-:CD24+:JAG1- at a frequency of 2-5% by serial and single cell transplantation assays, showed elevated expression of proliferation genes and low expression of differentiation genes (compared to non-TIC fraction CD24- of the same tumor). We used microarrays to detect differentially expressed genes in the TIC fraction compared to the non-TIC fraction of the same tumor Cells from each primary tumor were separated by FACS into TIC and non-TIC fractions and total RNA was extracted and hybridized on Affymetrix microarrays.
Project description:Barrett’s esophagus confers significant risk of esophageal adenocarcinoma. We have established the cloning of patient-matched stem cells of Barrett’s, gastric, and esophageal epithelium. Transplantation of transformed Barrett’s stem cells yielded tumors with hallmarks of esophageal adenocarcinoma, whereas transformed esophageal stem cells produced squamous cell carcinomas. These findings define a stem cell target in a precancerous lesion for preemptive therapies.
Project description:Barrett’s esophagus is a precancerous lesion that confers a significant risk of esophageal adenocarcinoma. Strategies for selective eradication of Barrett’s have been stymied by our inability to identify the Barrett’s stem cell. Here we employ novel technologies to clone patient-matched stem cells of Barrett’s, gastric, and esophageal epithelium. Genomic analyses of Barrett’s stem cells reveal a patient-specific mutational spectrum ranging from low somatic variation similar to patient-matched gastric epithelial stem cells to ones marked by extensive heterozygous alteration of genes implicated in tumor suppression, epithelial planarity, and epigenetic regulation. Transplantation of transformed Barrett’s stem cells yields tumors with hallmarks of esophageal adenocarcinoma, whereas transformed esophageal stem cells yield squamous cell carcinomas. Thus Barrett’s develops from cells distinct from local eponymous epithelia, emerges without obvious driver mutations, and likely progresses through and from the generation of dominant clones. These findings define a stem cell target for preemptive therapies of a precancerous lesion.
Project description:Tumorigenic breast cancer cells characterized by CD44 expression and low or undetectable CD24 levels (CD44+/CD24-/low) may be resistant to chemotherapy and therefore responsible for cancer relapse. Paired breast cancer core biopsies before and after neoadjuvant chemotherapy or lapatinib were obtained and as single cell suspensions stained using antibodies against CD24, CD44, and lineage markers, and then analyzed by flow cytometry. Mammosphere (MS) formation in culture was compared before and after treatment. Global gene expression differences between cancer cells bearing CD44+/CD24-/low cells and all other sorted cells, and between cancer MS and the primary bulk invasive cancers were analyzed. We report that CD44+/CD24-/low tumorigenic breast cancer cells were intrinsically chemoresistant ─ chemotherapy led to increased CD44+/CD24-/low cells, increased self-renewal capacity on MS assays, and enhanced tumorigeneicity in immunocompromised SCID/Beige mice. Conversely, in patients with HER2 overexpressing tumors, the EGFR/HER2 tyrosine kinase inhibitor, lapatinib decreased CD44+/CD24-/low cells, with the majority of these patients after conventional therapy achieving pathologic complete response, a validated surrogate marker for long-term survival. Gene transcription pathways that underlie chemoresistant, MS-forming CD44+/CD24-/low cells involve genes belonging to stem cell self-renewal, Wnt signaling, and early development pathways. Keywords: two group comparison