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Side population cells in human primary melanoma

ABSTRACT: Melanoma remains the most lethal skin cancer, mainly because of high resistance to therapy. Side population (SP) cells are found in many types of cancer and are usually enriched in therapy-resistant as well as tumorigenic cells. Here, we identified a Hoechst dye-effluxing SP in a large series of human melanoma samples representing different progression phases. The SP size did not change with disease stage but was correlated with the prognostic “Breslow’s depth” in the primary (cutaneous) tumors. When injected into immunodeficient mice, the SP generated larger tumors than the bulk “main population” (MP) melanoma cells in two consecutive generations, and showed tumorigenic capacity at lower cell numbers than the MP. In addition, the SP reconstituted the heterogeneous composition of the human A375 melanoma cell line, and its clonogenic activity was 2.5-fold higher than that of the MP. Gene-expression analysis revealed upregulated expression in the melanoma SP (versus the MP) of genes associated with chemoresistance and anti-apoptosis. Consistent with these molecular characteristics, the SP increased in proportion when A375 cells were exposed to the melanoma standard chemotherapeutic agent dacarbazine, and to the aggravating condition of hypoxia. In addition, the SP showed enhanced expression of genes related to cell invasion and migration, as well as to putative (melanoma) cancer stem cells (CSC) including ABCB1 and JARID1B. ABCB1 immunoreactivity was detected in a number of tumor cells in human melanomas, and in particular in clusters at the invasive front of the primary tumors. Together, our findings support that the human melanoma SP is enriched in tumorigenic and chemoresistant capacity, considered key characteristics of CSC. The melanoma SP may therefore represent an interesting therapeutic target.

ORGANISM(S): Homo sapiens

PROVIDER: GSE48838 | GEO | 2014/05/27

SECONDARY ACCESSION(S): PRJNA212802

REPOSITORIES: GEO

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Project description:Melanoma remains the most lethal skin cancer, mainly because of high resistance to therapy. Side population (SP) cells are found in many types of cancer and are usually enriched in therapy-resistant as well as tumorigenic cells. Here, we identified a Hoechst dye-effluxing SP in a large series of human melanoma samples representing different progression phases. The SP size did not change with disease stage but was correlated with the prognostic “Breslow’s depth” in the primary (cutaneous) tumors. When injected into immunodeficient mice, the SP generated larger tumors than the bulk “main population” (MP) melanoma cells in two consecutive generations, and showed tumorigenic capacity at lower cell numbers than the MP. In addition, the SP reconstituted the heterogeneous composition of the human A375 melanoma cell line, and its clonogenic activity was 2.5-fold higher than that of the MP. Gene-expression analysis revealed upregulated expression in the melanoma SP (versus the MP) of genes associated with chemoresistance and anti-apoptosis. Consistent with these molecular characteristics, the SP increased in proportion when A375 cells were exposed to the melanoma standard chemotherapeutic agent dacarbazine, and to the aggravating condition of hypoxia. In addition, the SP showed enhanced expression of genes related to cell invasion and migration, as well as to putative (melanoma) cancer stem cells (CSC) including ABCB1 and JARID1B. ABCB1 immunoreactivity was detected in a number of tumor cells in human melanomas, and in particular in clusters at the invasive front of the primary tumors. Together, our findings support that the human melanoma SP is enriched in tumorigenic and chemoresistant capacity, considered key characteristics of CSC. The melanoma SP may therefore represent an interesting therapeutic target. Four primary melanomas (subtype SSMM) were dissociated into single cells. From each of these single cell suspensions, side population (SP) and bulk \main population\ (MP) cells were isolated based on their Hoechst-effluxing capability using fluorescence-activated cell sorting. Gene expression of SP cells is analyzed relative to the gene expression of the corresponding MP cells.

Project description:Pituitary adenomas cause significant endocrine and mass-related morbidity. Not much is known about mechanisms underlying pituitary tumor pathogenesis.We searched for a side population (SP) in pituitary tumor, representing cells with high efflux capacity and potentially enriching for cancer/tumor stem cells (CSC/TSC). Human pituitary adenomas contain a SP irrespective of hormonal phenotype. This adenoma SP, as well as the further purified SP (pSP) depleted from endothelial and immune cells, enriches for cells that express ‘tumor stemness’ markers and signaling pathways, including epithelial-mesenchymal transition (EMT)-linked factors. The adenomas were found to contain self-renewing sphere-forming cells, considered a property of TSC. The sphere-initiating cells were retrieved in the pSP.Because benign pituitary adenomas do not grow in vitro and failed to expand in immunodeficient mice, the pituitary tumor cell line AtT20 was further used. We identified a SP in this cell line and found it more tumorigenic than the main population (MP). Of the two EMT-regulatory pathways tested, inhibition of Cxcr4 signaling reduced EMT-linked cell motility in vitro as well as xenograft tumor growth, whereas activation of TGFβ had no effect.The tumor pSP showed upregulated expression of the pituitary stem cell marker SOX2. Pituitaries from dopamine receptor D2 knockout (Drd2-/-) mice bearing prolactinomas, contain more pSP, Sox2+ and colony-forming cells than wildtype glands.In conclusion, we detected a SP in pituitary tumors and identified TSC-associated characteristics. Our study may contribute to a better understanding of pituitary tumor pathogenesis and lead to new therapeutic targets. We determined gene expression profiles for 9 human pituitary adenoma samples (5 x NF-A; 4 x GH-A). The samples were obtained immediately after transsphenoidal resection (by the neurosurgeon Dr. van Loon, Division Neurosurgery, University Hospitals Leuven). Informed consent was received from the patients. Hormonal phenotype was determined by the Department of Imaging and Pathology (University Hospitals Leuven). For each sample, total SP and MP cells were sorted by FACS. Separate assays were run subsequently for the two fractions. As such we present data from 18 microarrays.

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Side population cells in human primary melanoma

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