Background: Loss of NF1 gene function predisposes individuals to develop NF1-associated tumors, for which there are no known biomarkers/therapeutic targets. Results: TCTP was up-regulated in NF1-associated tumors and enhanced their growth via its positive feedback to the mTOR signaling, which was inhibited by artesunate/rapamycin. Conclusion: TCTP is a novel target functionally implicated in NF1 tumorigenesis. Significance: TCTP could serve as a biomarker/therapeutic target for NF1-associated tumors.

The blood-brain barrier (BBB) is formed by brain capillary endothelial cells linked together via complex tight junctions, and serves to prevent entry of drugs into the brain. Multiple transporters are expressed at the BBB, where they control exchange of materials between the circulating blood and brain interstitial fluid, thereby supporting and protecting the CNS. An understanding of the BBB is necessary for efficient development of CNS-acting drugs and to identify potential drug targets for treatment of CNS diseases. Quantitative targeted proteomics can provide detailed information on protein expression levels at the BBB. The present review highlights the latest applications of quantitative targeted proteomics in BBB research, specifically to evaluate species and in vivo-in vitro differences, and to reconstruct in vivo transport activity. Such a BBB quantitative proteomics approach can be considered as pharmacoproteomics.

Neurofibromatosis type 1 (NF1) tumor suppressor gene product, neurofibromin, functions in part as a Ras-GAP, and though its loss is implicated in the neuronal abnormality of NF1 patients, its precise cellular function remains unclear. To study the molecular mechanism of NF1 pathogenesis, we prepared NF1 gene knockdown (KD) PC12 cells, as a NF1 disease model, and analyzed their molecular (gene and protein) expression profiles with a unique integrated proteomics approach, comprising iTRAQ, 2D-DIGE, and DNA microarrays, using an integrated protein and gene expression analysis chart (iPEACH). In NF1-KD PC12 cells showing abnormal neuronal differentiation after NGF treatment, of 3198 molecules quantitatively identified and listed in iPEACH, 97 molecules continuously up-or down-regulated over time were extracted. Pathway and network analysis further revealed overrepresentation of calcium signaling and transcriptional regulation by glucocorticoid receptor (GR) in the up-regulated protein set, whereas nerve system development was overrepresented in the down-regulated protein set. The novel up-regulated network we discovered, "dynein IC2-GR-COX-1 signaling," was then examined in NF1-KD cells. Validation studies confirmed that NF1 knockdown induces altered splicing and phosphorylation patterns of dynein IC2 isomers, up-regulation and accumulation of nuclear GR, and increased COX-1 expression in NGF-treated cells. Moreover, the neurite retraction phenotype observed in NF1-KD cells was significantly recovered by knockdown of the dynein IC2-C isoform and COX-1. In addition, dynein IC2 siRNA significantly inhibited nuclear translocation and accumulation of GR and up-regulation of COX-1 expression. These results suggest that dynein IC2 up-regulates GR nuclear translocation and accumulation, and subsequently causes increased COX-1 expression, in this NF1 disease model. Our integrated proteomics strategy, which combines multiple approaches, demonstrates that NF1-related neural abnormalities are, in part, caused by upregulation of dynein IC2-GR-COX-1 signaling, which may be a novel therapeutic target for NF1. Neurofibromatosis type 1 (NF1)1 is an autosomal dominantly inherited disorder with an estimated prevalence of 1 in 3000 people (1). The hallmarks of NF1 include development of benign tumors of the peripheral nervous system and increased risk of malignancies. The phenotype of NF1 is highly variable, with several organ systems affected including the skin, bones, irises, and central and peripheral nervous systems. The effects on the nervous system are manifested in multiple neurofibroma, gliomas, and learning disabilities.The NF1 gene is located on chromosome 17q11.2 and encodes a large protein of 2818 amino acids, neurofibromin (2). Because the great majority of NF1 gene mutations frequently found in NF1 patients disturb the expression of intact neurofibromin, functional disruption of neurofibromin is potentially relevant to the expression of some or all of the abnormalities that occur in NF1 patients (3). A region centered 1...

Glioma initiating cells (GICs) are considered responsible for the therapeutic resistance and recurrence of malignant glioma. To clarify the molecular mechanism of GIC maintenance/differentiation, we established GIC clones having the potential to differentiate into malignant gliomas, and subjected to DNA microarray/iTRAQ based integrated proteomics. 21,857 mRNAs and 8,471 proteins were identified and integrated into a gene/protein expression analysis chart. Gene Ontology analysis revealed that the expression of cell adhesion molecules, including integrin subfamilies, such as α2 and αV, and extracellular matrices (ECMs), such as collagen IV (COL4), laminin α2 (LAMA2), and fibronectin 1 (FN), was significantly upregulated during serum-induced GIC differentiation. This differentiation process, accompanied by the upregulation of MAPK as well as glioma specific proteins in GICs, was dramatically accelerated in these ECM (especially FN)-coated dishes. Integrin αV blocking antibody and RGD peptide significantly suppressed early events in GIC differentiation, suggesting that the coupling of ECMs to integrin αV is necessary for GIC differentiation. In addition, the expression of integrin αV and its strong ligand FN was prominently increased in glioblastomas developed from mouse intracranial GIC xenografts. Interestingly, during the initial phase of GIC differentiation, the RGD treatment significantly inhibited GIC proliferation and raised their sensitivity against anti-cancer drug temozolomide (TMZ). We also found that combination treatments of TMZ and RGD inhibit glioma progression and lead the longer survival of mouse intracranial GIC xenograft model. These results indicate that GICs induce/secrete ECMs to develop microenvironments with serum factors, namely differentiation niches that further stimulate GIC differentiation and proliferation via the integrin recognition motif RGD. A combination of RGD treatment with TMZ could have the higher inhibitory potential against the glioma recurrence that may be regulated by the GICs in the differentiation niche. This study provides a new perspective for developing therapeutic strategies against the early onset of GIC-associated glioma.

We identified a cDNA encoding a putative cytosolic sulfotransferase (SULT) by searching the expressed sequence tag database of Bombyx mori, and subsequently obtained the full-length cDNA for this gene via rapid amplification of cDNA ends (RACE). We designated this gene bmST1, and showed by sequence analysis that it belongs to a novel SULT family. The tissue specificity of bmST1 mRNA expression was examined in fifth instar larvae by reverse transcriptase-polymerase chain reaction (RT-PCR), and transcripts were detectable in the silk gland, gut, fat body, and Malpighian tube. A recombinant form of bmST1 was then expressed using a gluthathione S-transferase (GST) gene fusion system, and it was purified from Escherichia coli. Purified bmST1 did not exhibit sulfating activity toward SULT substrates such as 4-nitrophenol, vanillin, hydroxysteroids, or monoamines. Surprisingly, however, recombinant bmST1 showed considerable activity toward 4-nitrocatechol and also gallate esters, although the catechins are not sulfated by this enzyme.