Dexamethasone Interferes with Autophagy and Affects Cell Survival in Irradiated Malignant Glioma Cells

Objective@#: Radiation is known to induce autophagy in malignant glioma cells whether it is cytocidal or cytoprotective.Dexamethasone is frequently used to reduce tumor-associated brain edema, especially during radiation therapy. The purpose of the study was to determine whether and how dexamethasone affects autophagy in irradiated malignant glioma cells and to identify possible intervening molecular pathways. @*Methods@#: We prepared p53 mutant U373 and LN229 glioma cell lines, which varied by phosphatase and tensin homolog (PTEN) mutational status and were used to make U373 stable transfected cells expressing GFP-LC3 protein. After performing cell survival assay after irradiation, the IC50 radiation dose was determined. Dexamethasone dose (10 µM) was determined from the literature and added to the glioma cells 24 hours before the irradiation. The effect of adding dexamethasone was evaluated by cell survival assay or clonogenic assay and cell cycle analysis. Measurement of autophagy was visualized by western blot of LC3-I/LC3-II and quantified by the GFP-LC3 punctuated pattern under fluorescence microscopy and acridine orange staining for acidic vesicle organelles by flow cytometry. @*Results@#: Dexamethasone increased cell survival in both U373 and LN229 cells after irradiation. It interfered with autophagy after irradiation differently depending on the PTEN mutational status : the autophagy decreased in U373 (PTEN-mutated) cells but increased in LN229 (PTEN wild-type) cells. Inhibition of protein kinase B (AKT) phosphorylation after irradiation by LY294002 reversed the dexamethasone-induced decrease of autophagy and cell death in U373 cells but provoked no effect on both autophagy and cell survival in LN229 cells. After ATG5 knockdown, radiation-induced autophagy decreased and the effect of dexamethasone also diminished in both cell lines. The diminished autophagy resulted in a partial reversal of dexamethasone protection from cell death after irradiation in U373 cells; however, no significant change was observed in surviving fraction LN229 cells. @*Conclusion@#: Dexamethasone increased cell survival in p53 mutated malignant glioma cells and increased autophagy in PTENmutant malignant glioma cell but not in PTEN-wildtype cell. The difference of autophagy response could be mediated though the phosphatidylinositol 3-kinase/AKT/mammalian target of rapamycin signaling pathway.

Preclinical development of a humanized neutralizing antibody targeting HGF

Hepatocyte growth factor (HGF) and its receptor, cMET, play critical roles in cell proliferation, angiogenesis and invasion in a wide variety of cancers. We therefore examined the anti-tumor activity of the humanized monoclonal anti-HGF antibody, YYB-101, in nude mice bearing human glioblastoma xenografts as a single agent or in combination with temozolomide. HGF neutralization, The extracellular signal-related kinases 1 and 2 (ERK1/2) phosphorylation, and HGF-induced scattering were assessed in HGF-expressing cell lines treated with YYB-101. To support clinical development, we also evaluated the preclinical pharmacokinetics and toxicokinetics in cynomolgus monkeys, and human and cynomolgus monkey tissue was stained with YYB-101 to test tissue cross-reactivity. We found that YYB-101 inhibited cMET activation in vitro and suppressed tumor growth in the orthotopic mouse model of human glioblastoma. Combination treatment with YYB-101 and temozolomide decreased tumor growth and increased overall survival compared with the effects of either agent alone. Five cancer-related genes (TMEM119, FST, RSPO3, ROS1 and NBL1) were overexpressed in YYB-101-treated mice that showed tumor regrowth. In the tissue cross-reactivity assay, critical cross-reactivity was not observed. The terminal elimination half-life was 21.7 days. Taken together, the in vitro and in vivo data demonstrated the anti-tumor efficacy of YYB-101, which appeared to be mediated by blocking the HGF/cMET interaction. The preclinical pharmacokinetics, toxicokinetics and tissue cross-reactivity data support the clinical development of YYB-101 for advanced cancer.

Dynamic relocalization of NHERF1 mediates chemotactic migration of ovarian cancer cells toward lysophosphatidic acid stimulation

NHERF1/EBP50 (Na⁺/H⁺ exchanger regulating factor 1; Ezrin-binding phosphoprotein of 50 kDa) organizes stable protein complexes beneath the apical membrane of polar epithelial cells. By contrast, in cancer cells without any fixed polarity, NHERF1 often localizes in the cytoplasm. The regulation of cytoplasmic NHERF1 and its role in cancer progression remain unclear. In this study, we found that, upon lysophosphatidic acid (LPA) stimulation, cytoplasmic NHERF1 rapidly translocated to the plasma membrane, and subsequently to cortical protrusion structures, of ovarian cancer cells. This movement depended on direct binding of NHERF1 to C-terminally phosphorylated ERM proteins (cpERMs). Moreover, NHERF1 depletion downregulated cpERMs and further impaired cpERM-dependent remodeling of the cell cortex, suggesting reciprocal regulation between these proteins. The LPA-induced protein complex was highly enriched in migratory pseudopodia, whose formation was impaired by overexpression of NHERF1 truncation mutants. Consistent with this, NHERF1 depletion in various types of cancer cells abolished chemotactic cell migration toward a LPA gradient. Taken together, our findings suggest that the high dynamics of cytosolic NHERF1 provide cancer cells with a means of controlling chemotactic migration. This capacity is likely to be essential for ovarian cancer progression in tumor microenvironments containing LPA.

Radiation-Induced Autophagy Contributes to Cell Death and Induces Apoptosis Partly in Malignant Glioma Cells / Journal of the Korean Cancer Association, 대한암학회지

PURPOSE: Radiation-induced autophagy has been shown to play two different roles, in malignant glioma (MG) cells, cytocidal or cytoprotective. However, neither the role of radiation-induced autophagy for cell death nor the existence of autophagy-induced apoptosis, a well-known cell-death pathway after irradiation, has been verified yet. MATERIALS AND METHODS: We observed both temporal and dose-dependent response patterns of autophagy and apoptosis to radiation in MG cell lines. Additionally, we investigated the role of autophagy in apoptosis through knockdown of autophagy-related proteins. RESULTS: Autophagic activity measured by staining of acidic vesicle organelles and Western blotting of LC-3 protein increased in proportion to radiation dose from day 1 to 5 after irradiation. Apoptosis measured by annexin-V staining and Western blotting of cleaved poly(ADP-ribose) polymerase demonstrated relatively late appearance 3 days after irradiation that increased for up to 7 days. Blocking of pan-caspase (Z-VAD-FMK) did not affect apoptosis after irradiation, but silencing of Atg5 effectively reduced radiation-induced autophagy, which decreased apoptosis significantly. Inhibition of autophagy in Atg5 knockdown cells was shown to be beneficial for cell survival. Stable transfection of GFP-LC3 cells was observed after irradiation. Annexin-V was localized in cells bearing GFP-LC3 punctuated spots, indicating autophagy in immunofluorescence. Some of these punctuated GFP-LC3 bearing cells formed conglomerated spots and died in final phase. CONCLUSION: These findings suggest that autophagy appears earlier than apoptosis after irradiation and that a portion of the apoptotic population that appears later is autophagy-dependent. Thus, autophagy is a pathway to cell death after irradiation of MG cells.

Roles of Long Non-Coding RNAs on Tumorigenesis and Glioma Development

More than 98% of eukaryotic transcriptomes are composed of non-coding RNAs with no functional protein-coding capacity. Those transcripts also include tens of thousands of long non-coding RNAs (lncRNAs) which are emerging as key elements of cellular homeostasis, essentially tumorigenesis steps. However, we are only beginning to understand the nature and extent of the involvement of lncRNAs on tumorigeneis. Here, we highlight recent progresses that have identified a myriad of molecular functions on tumorigenesis for several lncRNAs including metastasis-associated lung adenocarcinoma transcript 1 (MALAT1), prostate cancer associated non-coding RNA 1 (PRNCR1), prostate cancer gene expression marker 1 (PCGEM1), H19, and homeobox transcript antisense intergenic RNA (HOTAIR), and several new lncRNAs for glioma development. Potential therapeutic approaches for the lncRNAs in various human diseases are also discussed.

A case of mucinous borderline ovarian tumor associated with undifferentiated sarcoma with angiosarcoma-like appearance / 대한산부인과학회지

The ovarian mucinous tumor associated with true sarcoma is very rare. Four cases of true sarcoma: fibrosarcoma, undifferentiated sarcoma, rhabdomyosarcoma, angiosarcoma in ovarian mucinous tumor have been reported. We present a case of mucinous borderline tumor associated with undifferentiated sarcoma with angiosarcoma-like appearance in 80-year-old woman.

Two Cases of Granulosa Cell Tumor of the Ovary / 대한산부인과학회지

Granulosa cell tumors are relatively low-grade malignancies accounting for about 1 to 2% of all primary ovarian neoplasms and have an indolent growth pattern. There are two types of tumors, adult type granulosa cell tumor (AGCT) and juvenile type granulosa cell tumor (JGCT), and each tumor reveals different clinical or histopathological features. The clinical manifestations are mostly associated with estrogen produced by tumor, which are vaginal bleeding or menstrual irregularity in AGCT and precocious puberty in JGCT. Although most patients are diagnosed in early stage with favorable prognosis, some recur after several years. So, continuous follow up is required. Recently, we experienced two cases of adult type granulosa cell tumor and report with a brief review of literatures.

A Case of Variants of Turner syndrome Showing 46,X,inv(Y)/45,X Karyotype with Y Chromosome microdeletion / 대한산부인과학회지

46,X,inv(Y)/45,X mosaicism is a extremely rare sex chromosomal disorder. We experienced an unusual mosaic Turner syndrome case in a 29-years old Korean woman with a phenotypic female, primary amenorrhea and immature secondary sexual characteristics. Cytogenetic analysis including GTG banding revealed 46,X,inv(Y)(q11.2q12?)[15]/45,X[35] mosaicism, and X/Y chromosome Fluorescence in situ hybridization (FISH) analysis result was ish (SRY-,DYZ3-)[16]/ ish der(Y)del(Y)(q12)inv(Y)(p11.3q12)(SRY sp, DYZ3-)[4] and its meaning was coexistence of microdeletion and inversion of Y chromosome. To our knowledge, this karyotype may be a very rare variant of Turner syndrome, and we report this case with brief review of related literature.

A Case of Mature cystic teratoma in Omentum / 대한산부인과학회지

The mature cystic teratoma of the omentum is a very rare tumor. It is generally believed that autoamputation and reimplantation of an ovarian tumor is the most common etiology of omental teratoma. Abdominal pain is the main presenting symptom of these tumors. A 41-year-old woman was admitted for pelvic mass. At laparotomy, a 8.5 x 7.0 x 7.5 cm mass was found in the pelvic cavity, attached to the omentum with adhesion to bladder. The histopathologic diagnosis was mature cystic teratoma in conjunction with the cyst wall is diffusely necrotic and calcified with hairs. The absence of the left ovary suggested that the tumor underwent autoamputation and reimplantation on the omentum. We report the case with a brief review of literature.

Inositol 5'-phosphatase, SHIP1 interacts with phospholipase C-gamma1 and modulates EGF-induced PLC activity

Phospholipase C-gamma1, containing two SH2 and one SH3 domains which participate in the interaction between signaling molecules, plays a significant role in the growth factor-induced signal transduction. However, the role of the SH domains in the growth factor-induced PLC-gamma1 regulation is unclear. By peptide-mass fingerprinting analysis, we have identified SHIP1 as the binding protein for the SH3 domain of PLC-gamma1. SHIP1 was co-immunoprecipitated with PLC-gamma1 and potentiated EGF-induced PLC-gamma1 activation. However, inositol 5'-phosphatase activity of SHIP1 was not required for the potentiation of EGF-induced PLC-gamma1 activation. Taken together, these results suggest that SHIP1 may function as an adaptor protein which can potentiate EGF-induced PLC-gamma1 activation without regards to its inositol 5'-phosphatase activity.

Secondary Abdominal Pregnancy after Tubal Reanastomosis: Case Report / 대한산부인과학회잡지

Abdominal pregnancy is a very rare and life threatening varient of ectopic pregnancy with high maternal mortality and perinatal mortality. A 26-year-old woman, gravida 2, para 0, abortion 2, visited our emergency department for amenorrhea for 11 weeks and lower abdominal pain. Diagnostic transvaginal ultrasonographic finding suggested ruptured ectopic pregnancy. Serum Hemoglobin level was 8.1 mg/dL, and Hematocrit value was 25.2%. On laparatomy about 2.000 mL of blood was filled in the abdomen and severe adhesion was found on right adnexa, posterior cul-de-sac and omentum. A live fetus was attached to uterus. After adhesiolysis, we removed conceptus from uterine surface. We performed subtotal hysterectomy and excised right fallopian tube. 10 pints of packed red blood cell and 3 pints of fresh frozen plasma were given to the patient during and after the operation. Patient recovered postoperatively without any complications and discharged at postoperative seventh day. We experienced a case of first trimester secondary abdominal pregnancy after tubal reanastomosis and reported it with brief of a literature review.

Two Cases of Endometriosis at the Subcutaneous Tissue of the Abdominal Scar after Cesarean Section / 대한산부인과학회잡지

Endometriosis is generally confined to the pelvic viscera and the peritoneum, but it can proliferate in other areas like pleura, skin, extremities, lung, gallbladder, stomach, kidney and surgical scar. Scar endometriosis usually occurs in the surgical scar of previous cesarean sections, hysterotomy and episiotomy. The occurrence of endometrioma in cesarean scar is an infrequent event, usually presenting as a tender abdominal wall mass. We report two cases of abdominal wall endometrioma after cesarean section, which is presented with a brief review of the literature.