miR-let-7f-1 regulates SPARC mediated cisplatin resistance in medulloblastoma cells.

Our previous studies indicate that Secreted Protein Acidic and Rich in Cysteine (SPARC) expression suppressed medulloblastoma tumor growth in vitro and in vivo. Here we sought to determine the effect of SPARC expression in medulloblastoma cells to chemotherapeutic agents. In this study, we show that SPARC expression induces cisplatin resistance in medulloblastoma cells. We also demonstrate that the autophagy was involved in SPARC expression mediated resistance to cisplatin. Suppression of autophagy by either autophagy inhibitor, 3-methyladenosine (3MA) or Atg5 siRNA enhanced cisplatin sensitivity in SPARC expressed cells. Further, SPARC expression suppressed miR-let-7f-1 expression which resulted in disrupted repression of High Mobility Group Box 1 (HMGB1), a critical regulator of autophagy. We also show that HMGB1 is a direct target of miR-let-7f-1 and forced expression of HMGB1 cDNA enhanced cisplatin sensitivity in SPARC expressed cells. In summary, our results suggest that SPARC modulates cisplatin resistance by modulating the Let-7f-1 miRNA/HMGB1 axis in medulloblastoma cells.

Molecular mechanisms underlying the divergent roles of SPARC in human carcinogenesis.

Communication between the cell and its surrounding environment, consisting of proteinaceous (non-living material) and extracellular matrix (ECM), is important for biophysiological and chemical signaling. This signaling results in a range of cellular activities, including cell division, adhesion, differentiation, invasion, migration and angiogenesis. The ECM non-structural secretory glycoprotein called secreted protein, acidic and rich in cysteine (SPARC), plays a significant role in altering cancer cell activity and the tumor's microenvironment (TME). However, the role of SPARC in cancer research has been the subject of controversy. This review mainly focuses on recent advances in understanding the contradictory nature of SPARC in relation to ECM assembly, cancer cell proliferation, adhesion, migration, apoptosis and tumor growth.

MicroRNA 203 Modulates Glioma Cell Migration via Robo1/ERK/MMP-9 Signaling.

Glioblastoma (GBM) is the most common and malignant primary adult brain cancer. Allelic deletion on chromosome 14q plays an important role in the pathogenesis of GBM, and this site was thought to harbor multiple tumor suppressor genes associated with GBM, a region that also encodes microRNA-203 (miR-203). In this study, we sought to identify the role of miR-203 as a tumor suppressor in the pathogenesis of GBM. We analyzed the miR-203 expression data of GBM patients in 10 normal and 495 tumor tissue samples derived from The Cancer Genome Atlas data set. Quantitative real-time PCR and in situ hybridization in 10 high-grade GBM and 10 low-grade anaplastic astrocytoma tumor samples showed decreased levels of miR-203 expression in anaplastic astrocytoma and GBM tissues and cell lines. Exogenous expression of miR-203 using a plasmid expressing miR-203 precursor (pmiR-203) suppressed glioma cell proliferation, migration, and invasion. We determined that one relevant target of miR-203 was Robo1, given that miR-203 expression decreased mRNA and protein levels as determined by RT-PCR and Western blot analysis. Moreover, cotransfection experiments using a luciferase-based transcription reporter assay have shown direct regulation of Robo1 by miR-203. We also show that Robo1 mediates miR-203 mediated antimigratory functions as up-regulation of Robo1 abrogates miR-203 mediated antimigratory effects. We also show that miR-203 expression suppressed ERK phosphorylation and MMP-9 expression in glioma cells. Furthermore, we demonstrate that miR-203 inhibits migration of the glioma cells by disrupting the Robo1/ERK/MMP-9 signaling axis. Taken together, these studies demonstrate that up-regulation of Robo1 in response to the decrease in miR-203 in glioma cells is responsible for glioma tumor cell migration and invasion.

Blockade of SOX4 mediated DNA repair by SPARC enhances radioresponse in medulloblastoma.

SPARC is a matricellular glycoprotein and a putative radioresistance-reversal-gene. We therefore explored the possibility of SPARC expression on medulloblastoma radiosensitivity in vitro and in vivo. The combined treatment of the SPARC and irradiation resulted in increased cell death when compared to cells treated with irradiation alone in vitro and in vivo. SPARC expression prior to irradiation suppressed checkpoints-1,-2 and p53 phosphorylation and DNA repair gene XRCC1. We also demonstrate that SPARC expression suppressed irradiation induced SOX-4 mediated DNA repair. These results provide evidence of the anti-tumor effect of combining SPARC with irradiation as a new therapeutic strategy for the treatment of medulloblastoma.

SPARC expression induces cell cycle arrest via STAT3 signaling pathway in medulloblastoma cells.

Dynamic cell interaction with ECM components has profound influence in cancer progression. SPARC is a component of the ECM, impairs the proliferation of different cell types and modulates tumor cell aggressive features. We previously reported that SPARC expression significantly impairs medulloblastoma tumor growth in vivo. In this study, we demonstrate that expression of SPARC inhibits medulloblastoma cell proliferation. MTT assay indicated a dose-dependent reduction in tumor cell proliferation in adenoviral mediated expression of SPARC full length cDNA (Ad-DsRed-SP) in D425 and UW228 cells. Flow cytometric analysis showed that Ad-DsRed-SP-infected cells accumulate in the G2/M phase of cell cycle. Further, immunoblot and immunoprecipitation analyses revealed that SPARC induced G2/M cell cycle arrest was mediated through inhibition of the Cyclin-B-regulated signaling pathway involving p21 and Cdc2 expression. Additionally, expression of SPARC decreased STAT3 phosphorylation at Tyr-705; constitutively active STAT3 expression reversed SPARC induced G2/M arrest. Ad-DsRed-SP significantly inhibited the pre-established orthotopic tumor growth and tumor volume in nude-mice. Immunohistochemical analysis of tumor sections from mice treated with Ad-DsRed-SP showed decreased immunoreactivity for pSTAT3 and increased immunoreactivity for p21 compared to tumor section from mice treated with mock and Ad-DsRed. Taken together our studies further reveal that STAT3 plays a key role in SPARC induced G2/M arrest in medulloblastoma cells. These new findings provide a molecular basis for the mechanistic understanding of the effects of SPARC on medulloblastoma tumor cell proliferation.

SPARC stimulates neuronal differentiation of medulloblastoma cells via the Notch1/STAT3 pathway.

Secreted protein acidic and rich in cysteine (SPARC) participates in the regulation of morphogenesis and cellular differentiation through its modulation of cell-matrix interactions. We previously reported that SPARC expression significantly impairs medulloblastoma tumor growth in vivo. In this study, we show that adenoviral-mediated overexpression of SPARC cDNA (Ad-DsRed-SP) elevated the expression of the neuronal markers NeuN, nestin, neurofilament, and MAP-2 in medulloblastoma cells and induced neuron-like differentiation. SPARC overexpression decreased STAT3 phosphorylation; constitutive expression of STAT3 reversed SPARC-mediated expression of neuronal markers. We also show that Notch signaling is suppressed in the presence of SPARC, as well as the Notch effector basic helix-loop-helix (bHLH) transcription factor hairy and enhancer of split 1 (HES1). Notch signaling was found to be responsible for the decreased STAT3 phosphorylation in response to SPARC expression. Furthermore, expression of SPARC decreased the production of interleukin 6 (IL-6) and supplemented IL-6-abrogated, SPARC-mediated suppression of Notch signaling and expression of neuronal markers. Immunohistochemical analysis of tumor sections from mice treated with Ad-DsRed-SP showed increased immunoreactivity for the neuronal markers and a decrease in Notch1 expression and phosphorylation of STAT3. Taken together, our results suggest that SPARC induces expression of neuronal markers in medulloblastoma cells through its inhibitory effect on IL-6-regulated suppression of Notch pathway-mediated STAT3 signaling, thus giving further support to the potential use of SPARC as a therapeutic candidate for medulloblastoma treatment. Findings show that SPARC-induced neuronal differentiation can sensitize medulloblastoma cells for therapy.