Sialidase NEU3 defines invasive potential of human glioblastoma cells by regulating calpain-mediated proteolysis of focal adhesion proteins.

BACKGROUND: Glioblastoma multiforme is one of the most malignant tumors of the human central nervous system characterized by high degree of invasiveness. Focusing on this invasive nature, we investigated whether ganglioside-specific sialidase NEU3 might be involved, because gangliosides are major components of brain tissues, and cell surface sialic acids, as target residues of sialidase catalysis, are thought to be closely related to cell invasion. METHODS: NEU3 mRNA levels of human glioblastoma specimens were evaluated by quantitative RT-PCR. Human glioblastoma cell lines, U251, A172, and T98G were used for cell invasion and migration by transwell and cell scratching assay. The molecules involved in the signaling cascade were investigated by western blot and immunofluorescent microscopy. RESULTS: NEU3 expression was down-regulated in human glioblastoma specimens. In the human glioblastoma cell lines, NEU3 overexpression reduced invasion and migration by promoting the assembly of focal adhesions through reduced calpain-dependent proteolysis, but NEU3 silencing resulted in accelerating cell invasion via disassembly of focal adhesions. In NEU3-silenced cells, elevation of calpain activity and GM3 accumulation were observed, as results of reduced sialidase hydrolysis, localization of calpain and GM3 at the cell lamellipodium being demonstrated by immunofluorescence microscopy. CONCLUSION: Sialidase NEU3 was found to exert a great influence on cell invasion in regulation of calpain activity and focal adhesion disassembly and consequent invasive potential of glioblastoma cells. GENERAL SIGNIFICANCE: This first demonstration of sialidase involvement in invasive potential of gliolastoma cells may point to NEU3 as an attractive treatment target of human gliomas.

DUSP13B/TMDP inhibits stress-activated MAPKs and suppresses AP-1-dependent gene expression.

The dual-specificity phosphatase (DUSP) 13 gene encodes two atypical DUSPs, DUSP13B/TMDP and DUSP13A/MDSP using alternative exons. DUSP13B protein is most highly expressed in testis, particularly in spermatocytes and round spermatids of the seminiferous tubules, while that of DUSP13A is restricted to skeletal muscle. Here, we show that DUSP13B inactivated MAPK activation in the order of selectivity, JNK = p38>ERK in cells, while DUSP13A did not show MAPK phosphatase activity. Reporter gene analysis showed that DUSP13B had significant inhibitory effect on AP-1-dependent gene expression, but DUSP13A did not. To our knowledge, DUSP13B is the first identified testis-specific phosphatase that inhibits stress-activated MAPKs. These data suggest an important role for DUSP13B in protection from external stress during spermatogenesis.

CDC25A mRNA levels significantly correlate with Ki-67 expression in human glioma samples.

Cell division cycle 25 (CDC25) phosphatases are cell-cycle regulatory proteins which are overexpressed in a significant number of human cancers. This study evaluated the role of CDC25 phosphatases in human glioma proliferation. Upregulation of CDC25A was observed in human glioma specimens and human glioma cell lines. Comparison of expression levels of CDC25A and CDC25B messenger ribonucleic acid (RNA) to Ki-67 labeling index in glioma tissues found that Ki-67 labeling index was significantly correlated with the expression of CDC25A, but not with that of CDC25B. Depletion of CDC25A by small interfering RNA and inhibition of CDC25 suppressed cell proliferation and induced apoptosis in glioma cell lines, indicating that CDC25A is a potential target for the development of new therapy for glioma.

PKM1 Confers Metabolic Advantages and Promotes Cell-Autonomous Tumor Cell Growth.

Expression of PKM2, which diverts glucose-derived carbon from catabolic to biosynthetic pathways, is a hallmark of cancer. However, PKM2 function in tumorigenesis remains controversial. Here, we show that, when expressed rather than PKM2, the PKM isoform PKM1 exhibits a tumor-promoting function in KRASG12D-induced or carcinogen-initiated mouse models or in some human cancers. Analysis of Pkm mutant mouse lines expressing specific PKM isoforms established that PKM1 boosts tumor growth cell intrinsically. PKM1 activated glucose catabolism and stimulated autophagy/mitophagy, favoring malignancy. Importantly, we observed that pulmonary neuroendocrine tumors (NETs), including small-cell lung cancer (SCLC), express PKM1, and that PKM1 expression is required for SCLC cell proliferation. Our findings provide a rationale for targeting PKM1 therapeutically in certain cancer subtypes, including pulmonary NETs.

Drug delivery of oral anti-cancer fluoropyrimidine agents.

INTRODUCTION: Sixty years since its introduction, 5-FU still forms the core of chemotherapy regimens for many types of malignancies. 5-FU is a time-dependent drug but is rapidly degraded in plasma by dihydropyrimidine dehydrogenase (DPD). Although originally developed in an intravenous form, 5-FU oral prodrugs were developed with the goal of improving efficacy and minimizing toxicity as well as to capitalize on the advantages of oral drug administration. The inactive 5-FU prodrug is gradually converted into the active form in the systemic circulation. UFT, S-1, and capecitabine are oral 5-FU prodrugs currently in clinical use. However, the efficacy of 5-FU can be further improved by its combination with DPD inhibitors and biochemical modulators, such as uracil and leucovorin, in addition to modifying administration schedules. Areas covered: We focused on the drug delivery of oral 5-FU prodrugs, their pharmacokinetics, and the development of DPD inhibitors. Since oral 5-FU prodrugs have been formulated into combination drugs, we also discussed the regulatory approval of combination drugs. Expert opinion: Many regimens that include intravenously administered 5-FU can be replaced by oral 5-FU prodrugs. Patients would benefit from development of combination 5-FU oral prodrug formulations and its associated path through the combination drug regulatory approval process.

Loss of protein phosphatase 6 in mouse keratinocytes increases susceptibility to ultraviolet-B-induced carcinogenesis.

We previously reported that deficiency in the gene encoding the catalytic subunit of protein phosphatase 6 (Ppp6c) predisposes mouse skin tissue to papilloma formation initiated by DMBA. Here, we demonstrate that Ppp6c loss acts as a tumor promoter in UVB-induced squamous cell carcinogenesis. Following UVB irradiation, mice with Ppp6c-deficient keratinocytes showed a higher incidence of skin squamous cell carcinoma than did control mice. Time course experiments showed that following UVB irradiation, Ppp6c-deficient keratinocytes upregulated expression of p53, PUMA, BAX, and cleaved caspase-3 proteins. UVB-induced tumors in Ppp6c-deficient keratinocytes exhibited a high frequency of both p53- and γH2AX-positive cells, suggestive of DNA damage. Epidemiological and molecular data strongly suggest that UVB from sunlight induces p53 gene mutations in keratinocytes and is the primary causative agent of human skin cancers. Our analysis suggests that PP6 deficiency underlies molecular events that drive outgrowth of initiated keratinocytes harboring UVB-induced mutated p53. Understanding PP6 function in preventing UV-induced tumorigenesis could suggest strategies to prevent and treat this condition.

Treatment results of glioblastoma during the last 30 years in a single institute.

Treatment results of glioblastoma (GB) during the last 30 years in Tohoku University were analyzed to identify any improvements in patient outcome in all 332 histologically proven cases of newly diagnosed GB treated consecutively in our department between 1982 and 2011. These 30 years was divided into 5 treatment eras, Group 1 (1982-1988, without preoperative evaluation by magnetic resonance [MR] imaging, n = 46), Group 2 (1989-1996, with preoperative MR imaging, n = 41), Group 3 (1997-1999, additionally underwent intraoperative functional brain mapping and neuronavigation system, n = 38), Group 4 (2000-August 2006, underwent 30 Gy of whole brain radiation followed by 30 Gy of extended local accelerated hyperfractionated radiation therapy, n = 96), and Group 5 (September 2006-2011, adjuvant usage of temozolomide [TMZ], n = 111). Overall survival (OS) was calculated from the date of surgery to the death from any cause. The median survival time/2-year OS/5-year OS of Groups 1 to 5 were 10.7 months/10.9%/0%, 17.3 months/26.2%/6.9%, 15.9 months/23.7%/5.3%, 20.1 months/34.8%/15.5%, and 20.9 months/45.5%/19.7%. The prognosis for patients with GB improved significantly after the introduction of MR imaging. Younger GB, defined as patients aged below 60 years, or total tumor resection with all ages in Group 5 had 5-year 0S of 31.0% and 30.1%, respectively. The prognosis of GB was improved significantly after the introduction of TMZ for elderly GB, recursive partitioning analysis class 5, or totally resected GB. Introduction of MR imaging and TMZ, and total resection of the tumor were important in the improvement of outcome for patients with GB.

Novel function of MKP-5/DUSP10, a phosphatase of stress-activated kinases, on ERK-dependent gene expression, and upregulation of its gene expression in colon carcinomas.

Mitogen-activated protein kinase phosphatase 5 (MKP-5)/DUSP10 acts as a phosphatase of stress-activated kinases (JNK and p38), but its activity towards ERK has not been demonstrated. In the present study we observed that MKP-5 interacts with ERK, retains it in the cytoplasm, suppresses its activation and downregulates ERK-dependent transcription. These data suggested a novel MKP-5 function as a scaffold protein for the ERK pathway. We analyzed MKP-5 gene expression in several tumors, and found that it is frequently upregulated in colorectal but not in lung and breast cancer, suggesting its association with the malignant phenotype of colon cancer.

Therapeutic Efficacy of Adoptive Cell Transfer on Survival of Patients with Glioblastoma Multiforme: Case Reports.

Glioblastoma multiforme (GBM), which occurs mostly in individuals over the age of 40, accounts for 12-15% of all primary brain tumors. Patients with GBM have a poor prognosis, even after aggressive upfront therapies. The present study documents that in 5 of these patients, the use of a novel immunotherapeutic approach combined with standard initial therapies resulted in a prolonged survival of over 3 years, which is significantly longer than the expected survival time with conventional therapies. During the course of intravenous cell-transfer immunotherapy, axial magnetic resonance images of the tumor region were monitored for over 5 years. The discontinuation of adoptive transfer regimens resulted in the rapid deterioration of patients with development of Gd-enhancing regions, indicating the initiation of tumor recurrence. Among patients with recurrence, the reinstatement of adoptive cell regimens with more frequent cell-transfers resulted in an apparent re-regression of tumors. Significantly longer survival times were seen in patients receiving transferred autologous lymphoid cells which were expanded in vitro, and which had a considerable proportion of gammadeltaT cells. We conclude that immunotherapy, combined with standard treatment, plays a significant role in the management of GBM patients and provides patients with a better prognosis.

Anaplastic ganglioglioma with sarcomatous component: an immunohistochemical study and molecular analysis of p53 tumor suppressor gene.

The present case report describes a case of ganglioglioma with a distinct sarcomatous component in the left temporal lobe of a 59-year-old Japanese man. Neoplastic neuroglial tissue contained both benign and anaplastic glial components with a MIB-1 labeling index of 0.1% and 12.0%, respectively. Sarcomatous tissue adjacent to the anaplastic glial tissue was dominated by pleomorphic fibroblastic cells with a MIB-1 labeling index of 10.8%. They were immunoreactive for smooth muscle actin, type IV collagen, and alpha 1 antitrypsin, but not for desmin and CD34. Interestingly, some of the sarcomatous cells were double-positive for smooth muscle actin and GFAP. The p53 protein had accumulated in the anaplastic astrocytes and sarcomatous cells, but direct DNA sequencing of PCR products failed to detect any mutation in the p53 gene (from exon 4 to exon 10).