Comparative Proteomics of Colon Cancer Stem Cells and Differentiated Tumor Cells Identifies BIRC6 as a Potential Therapeutic Target

University Medical Center Utrecht, Department of Surgery, Utrecht, The Netherlands.
Molecular & Cellular Proteomics (Impact Factor: 6.56). 07/2011; 10(12):M111.011353. DOI: 10.1074/mcp.M111.011353
Source: PubMed


Patients with liver metastases from colon carcinoma show highly variable responses to chemotherapy and tumor recurrence is frequently observed. Therapy-resistant cancer stem cells have been implicated in drug resistance and tumor recurrence. However, the factors determining therapy resistance and tumor recurrence are poorly understood. The aim of this study was to gain insight into these mechanisms by comparing the proteomes of patient-derived cancer stem cell cultures and their differentiated isogenic offspring. We established colonosphere cultures derived from resection specimens of liver metastases in patients with colon cancer. These colonospheres, enriched for colon cancer stem cells, were used to establish isogenic cultures of stably differentiated nontumorigenic progeny. Proteomics based on one-dimensional gel electrophoresis coupled to nano liquid chromatography tandem MS was used to identify proteome differences between three of these paired cultures. The resulting data were analyzed using Ingenuity Pathway Software. Out of a total data set of 3048 identified proteins, 32 proteins were at least twofold up-regulated in the colon cancer stem cells when compared with the differentiated cells. Pathway analysis showed that "cell death " regulation is strikingly different between the two cell types. Interestingly, one of the top-up-regulated proteins was BIRC6, which belongs to the class of Inhibitor of Apoptosis Proteins. Knockdown of BIRC6 sensitized colon cancer stem cells against the chemotherapeutic drugs oxaliplatin and cisplatin. This study reveals that differentiation of colon cancer stem cells is accompanied by altered regulation of cell death pathways. We identified BIRC6 as an important mediator of cancer stem cell resistance against cisplatin and oxaliplatin. Targeting BIRC6, or other Inhibitors of Apoptosis Proteins, may help eradicating colon cancer stem cells.

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    • "One of the nuclear spheroid proteins that warrants follow-up studies as a candidate prognostic and drug resistance biomarker in CRC is ILF3. ILF3 was also identified as spheroid enriched protein in our previous proteomics screen of single cell-derived CRC spheres [72]. This protein is a double-stranded RNA (dsRNA) binding protein that complexes with other proteins, dsRNAs, small noncoding RNAs, and mRNAs to regulate gene expression and stabilize mRNAs, and has an emerging function in DNA repair. "
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    ABSTRACT: Background Organotypic tumor spheroids, a 3D in vitro model derived from patient tumor material, preserve tissue heterogeneity and retain structural tissue elements, thus replicating the in vivo tumor more closely than commonly used 2D and 3D cell line models. Such structures harbour tumorigenic cells, as revealed by xenograft implantation studies in animal models and maintain the genetic makeup of the original tumor material. The aim of our work was a morphological and proteomic characterization of organotypic spheroids derived from colorectal cancer tissue in order to get insight into their composition and associated biology. Results Morphological analysis showed that spheroids were of about 250 μm in size and varied in structure, while the spheroid cells differed in shape and size and were tightly packed together by desmosomes and tight junctions. Our proteomic data revealed significant alterations in protein expression in organotypic tumor spheroids cultured as primary explants compared to primary colorectal cancer tissue. Components underlying cellular and tissue architecture were changed; nuclear DNA/ chromatin maintenance systems were up-regulated, whereas various mitochondrial components were down-regulated in spheroids. Most interestingly, the mesenchymal cells appear to be substantial component in such cellular assemblies. Thus the observed changes may partly occur in this cellular compartment. Finally, in the proteomics analysis stem cell-like characteristics were observed within the spheroid cellular assembly, reflected by accumulation of Alcam, Ctnnb1, Aldh1, Gpx2, and CD166. These findings were underlined by IHC analysis of Ctnnb1, CD24 and CD44, therefore warranting closer investigation of the tumorigenic compartment in this 3D culture model for tumor tissue. Conclusions Our analysis of organotypic CRC tumor spheroids has identified biological processes associated with a mixture of cell types and states, including protein markers for mesenchymal and stem-like cells. This 3D tumor model in which tumor heterogeneity is preserved may represent an advantageous model system to investigate novel therapeutic approaches.
    Full-text · Article · Jul 2014 · Proteome Science
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    • "One example comes from a comparative proteomic analysis of colon cancer stem cells and differentiated tumor cells, which uncovered a prominently upregulated protein, baculoviral inhibitor of apoptosis protein (IAP) repeat-containing protein 6 (BIRC6), one of the IAPs that may play a crucial role in the chemoresistance of colon cancer stem cells. Subsequent knockdown of the gene resulted in resensitization of the cells to chemotherapy.71 Results from this proteomic study suggest that BIRC6 could be used as a potential therapeutic target to eradicate colon cancer stem cells contributing to colon cancer recurrence. "
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    ABSTRACT: Proteomic approaches are continuing to make headways in cancer research by helping to elucidate complex signaling networks that underlie tumorigenesis and disease progression. This review describes recent advances made in the proteomic discovery of drug targets for therapeutic development. A variety of technical and methodological advances are overviewed with a critical assessment of challenges and potentials. A number of potential drug targets, such as baculoviral inhibitor of apoptosis protein repeat-containing protein 6, macrophage inhibitory cytokine 1, phosphoglycerate mutase 1, prohibitin 1, fascin, and pyruvate kinase isozyme 2 were identified in the proteomic analysis of drug-resistant cancer cells, drug action, and differential disease state tissues. Future directions for proteomics-based target identification and validation to be more translation efficient are also discussed.
    Full-text · Article · Oct 2013 · Drug Design, Development and Therapy
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    • "In-gel digestion and NanoLC-MS/MS analysis was performed as described by van Houdt et al (2011). "
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