Animals ; Bufo bufo ; Caspase 3 ; genetics ; metabolism ; Female ; Immunohistochemistry ; Male ; Proto-Oncogene Proteins c-bcl-2 ; genetics ; metabolism ; Proto-Oncogene Proteins c-kit ; genetics ; metabolism ; Spinal Cord ; metabolism ; Spinal Cord Injuries ; metabolism ; bcl-2-Associated X Protein ; genetics ; metabolism

Gastrointestinal stromal tumors (GISTs) of the gallbladder are representative of an extremely rare group of tumors. We have encountered a patient with a malignant GIST of the gallbladder and presented it with a review of some articles. A 72-yr-old woman initially presented with right upper quadrant abdominal pain, fever and chills. Emergency cholecystectomy was performed under the impression of gallbladder empyema. Liver metastasis was found at 7 months postoperatively and the patient expired 9 months after the surgery. At the time of cholecystectomy, the gallbladder showed a necrotic serosal surface with an irregular thickened wall. A mass, 6 cm in length and 3 cm in width, encircled the whole wall of the neck and upper body of the gallbladder. Microscopic findings revealed frequent mitotic figures (more than 20/50 HPF) and tumor necrosis. Hyperchromatic, pleomorphic and spindle shaped neoplastic cells that were arranged in a pattern of short fascicles infiltrated the entire layer of the gallbladder. The tumor cells were immunoreactive for CD117 antigen (c-kit protein) and vimentin. They were negative for desmin, smooth muscle actin and S-100 protein. Mutations of the c-kit proto-oncogene were not found in this case. These findings were sufficient to provide enough clinical, histopathological and immunohistochemicalevidence in diagnosing our case as a malignant GIST.
Aged ; Female ; Gallbladder Neoplasms/genetics/metabolism/*pathology ; Gastrointestinal Stromal Tumors/genetics/metabolism/*pathology ; Humans ; Proto-Oncogene Protein c-kit/genetics/metabolism

This study was aimed to investigate the expression level of SHP-1 and C-kit genes in acute leukemia HL-60 cells and effect of inhibitor As2O3 demethylation on SHP-1 and C-kit genes expression. RT-PCR was used to detect the expression level of SHP-1 and C-kit mRNA in drug-treated cell group and control group. The methylation specific PCR (MSP) was applied to measure the methylation status of SHP-1 gene in HL-60 cells. The results showed that after being treated with As2O3 the recovery of SHP-1 gene expression was observed in HL-60 cells in which SHP-1 mRNA originally did not expressed, meanwhile the expression level of C-kit mRNA in HL-60 cells with high expression decreased. When HL-60 cells were treated with As2O3 of 1.0, 2.5, 5.0 µmol/L, the demethylation effects was enhanced, the expression of SHP-1 mRNA displayed an ascending tendency, and expression of C-kit mRNA showed an descending tendency in dose-dependent manner (P < 0.05). It is concluded that the absence of SHP-1 mRNA expression in HL-60 cells and recovery of expression after treatment with As2O3 suggest the hypermethylation of SHP-1 gene related with pathogenesis of leukemia, and the abnormal increase of C-kit mRNA expression maybe exist in formation of leukemia. The effect of As2O3 on expression of SHP-1 and C-kit shows dose-dependency, the higher the As2O3 concentration, the higher the SHP-1 expression and the lower the C-kit expression, moreover, the effect of As2O3 shows time-dependency in specific concentration. The SHP-1 mRNA expression negatively relates with C-kit mRNA expression, suggesting that the decrease or absence of SHP-1 expression in leukemia cells weakens the negative regulation on C-kit signaling pathway, thus plays a role in the formation of leukemia.
Arsenicals ; metabolism ; pharmacology ; DNA Methylation ; Gene Expression Regulation, Leukemic ; HL-60 Cells ; Humans ; Oxides ; metabolism ; pharmacology ; Protein Tyrosine Phosphatase, Non-Receptor Type 6 ; genetics ; Proto-Oncogene Proteins c-kit ; genetics

OBJECTIVETo explore the effect of Imatinib mesylate on proliferation, differentiation and apoptosis of leukemic Kasumi-1 cells bearing c-kit mutation.

METHODSKasumi-1 cells were treated with Imatinib at different concentrations in culture. Cell proliferation was assayed by MTT assay, expressions of c-kit antigen, surface myeloid antigen and cell cycle by flow cytometry, cell apoptosis by annexin V staining and agarose gel electrophoresis. Western blot was used to analyze the level of c-kit protein tyrosine phosphorylation.

RESULTSImatinib treatment caused a time- and dose-dependent inhibition of the cell proliferation, with a 72 h IC50 of 4.45 micromol/L. Imatinib treatment induced a decrease in the mean fluorescence value of c-kit antigen, a progressive decline in S-phase cell fraction and an increase in G0/G1 cells. Treatment with 5.00 micromol/L of imatinib for 72 h induced an increase in expression of myeloid surface protein CD11, CD13 and CD15, and for 24 h induced an increase in early apoptosis cells [from 9.04% to 86.84% (P < 0.05)]. The apoptosis ladder was observed on agarose gel electrophoresis on 5-day treatment. Tyrosine phosphorylation level of c-kit protein was decreased by Imatinib treatment.

CONCLUSIONTyrosine kinase inhibitor Imatinib mesylate treatment could inhibit proliferation of Kasumi-1 cells which bear a c-kit mutation, induce differentiation, apoptosis and G0/G1 cells accumulation.

Apoptosis ; drug effects ; genetics ; Benzamides ; Cell Differentiation ; drug effects ; genetics ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Humans ; Imatinib Mesylate ; Mutation ; Piperazines ; pharmacology ; Protein-Tyrosine Kinases ; antagonists & inhibitors ; Proto-Oncogene Proteins c-kit ; genetics ; metabolism ; Pyrimidines ; pharmacology

This study was aimed to investigate the expression level of SHP-1 and C-kit genes in acute leukemia HL-60 cells and effect of 5-aza-CdR demethylation on expression of SHP-1 and C-kit genes. RT-PCR was used to detect the mRNA expression level of SHP-1 and C-kit mRNA in HL-60 cells of the drug-treated group and control group.The methylation specific PCR (MSP) was applied to measure the methylation status of SHP-1 and C-kit genes in HL-60 cells.The results showed that after being treated with 5-aza-CdR, the recovery of SHP-1 gene expression was observed in HL-60 cells in which SHP-1 mRNA originally was not expressed. Meanwhile, the high expression level of C-kit mRNA in HL-60 cells was decreased. When HL-60 cells were treated with 0, 0.5, 1.0, 2.0 µmol/L 5-aza-CdR, the demethylation effect was enhanced, the expression of SHP-1 mRNA displayed an ascending tendency, and the expression of C-kit mRNA showed an descending tendency in dose-dependent manner (P < 0.05) . It is concluded that the absence of SHP-1 mRNA expression in HL-60 cells and recovery of expression after treatment with 5-aza-CdR suggest that the hypermethylation of SHP-1 gene relates with pathogenesis of leukemia, and the abnormal increase of C-kit mRNA expression maybe exist in formation of leukemia. The effect of 5-aza-CdR on expression of SHP-1 and C-kit shows dose-dependency, the higher the 5-aza-CdR concentration, the higher the SHP-1 expression and the lower the C-kit expression, moreover, the effect of 5-aza-CdR shows time-dependency in specific concentration.The SHP-1 mRNA expression negatively correlates with C-kit mRNA expression, suggesting that the decrease or absence of SHP-1 expression in leukemia cells weakens the negative regulation on C-kit signaling pathway, thus plays a role in the formation of leukemia.
Azacitidine ; pharmacology ; DNA Methylation ; drug effects ; HL-60 Cells ; Humans ; Leukemia ; genetics ; metabolism ; Protein Tyrosine Phosphatase, Non-Receptor Type 6 ; genetics ; Proto-Oncogene Proteins c-kit ; genetics ; RNA, Messenger

BACKGROUNDThe addition of anti-human epidermal growth factor receptor 2 (HER2)-targeted drugs, such as trastuzumab, lapatinib, and trastuzumab emtansine (T-DM1), to chemotherapy significantly improved prognosis of HER2-positive breast cancer patients. However, it was confused that metastatic patients vary in the response of targeted drug. Therefore, methods of accurately predicting drug response were really needed. To overcome the spatial and temporal limitations of biopsies, we aimed to develop a more sensitive and less invasive method of detecting mutations associated with anti-HER2 therapeutic response through circulating-free DNA (cfDNA).

METHODSFrom March 6, 2014 to December 10, 2014, 24 plasma samples from 20 patients with HER2-positive metastatic breast cancer who received systemic therapy were eligible. We used a panel for detection of hot-spot mutations from 50 oncogenes and tumor suppressor genes, and then used targeted next-generation sequencing (NGS) to identify somatic mutation of these samples in those 50 genes. Samples taken before their first trastuzumab administration and subsequently proven with clinical benefit were grouped into sensitive group. The others were collected after disease progression of the trastuzumab-based therapy and were grouped into the resistant group.

RESULTSA total of 486 single-nucleotide variants from 46 genes were detected. Of these 46 genes, phosphatidylinositol 4,5-bisphosphate 3-kinase catalytic subunit alpha (PIK3CA), proto-oncogene c-Kit (KIT), and tumor protein p53 (TP53) were the most common mutated genes. Seven genes, including epidermal growth factor receptor (EGFR), G protein subunit alpha S (GNAS), HRas proto-oncogene (HRAS), mutL homolog 1 (MLH1), cadherin 1 (CDH1), neuroblastoma RAS viral oncogene homolog (NRAS), and NOTCH1, that only occurred m utations in the resistant group were associated with the resistance of targeted therapy. In addition, we detected a HER2 S855I mutation in two patients who had persistent benefits from anti-HER2 therapy.

CONCLUSIONTargeted NGS of cfDNA has potential clinical utility to detect biomarkers from HER2-targeted therapies.

Adolescent ; Adult ; Aged ; Biomarkers, Tumor ; genetics ; Breast Neoplasms ; genetics ; metabolism ; Cadherins ; genetics ; Chromogranins ; genetics ; Class I Phosphatidylinositol 3-Kinases ; Drug Resistance, Neoplasm ; genetics ; Female ; GTP-Binding Protein alpha Subunits, Gs ; genetics ; Humans ; Male ; Middle Aged ; Mutation ; genetics ; Phosphatidylinositol 3-Kinases ; genetics ; Proto-Oncogene Proteins c-kit ; genetics ; Receptor, ErbB-2 ; metabolism ; Receptor, Notch1 ; genetics ; Tumor Suppressor Protein p53 ; genetics ; Young Adult

Cyclin-Dependent Kinase 6 ; metabolism ; Gene Expression Regulation, Neoplastic ; Genes, p53 ; Humans ; Killer Cells, Natural ; pathology ; Lymphoma, T-Cell ; genetics ; metabolism ; pathology ; Mutation ; Nose Neoplasms ; genetics ; metabolism ; pathology ; Protein Tyrosine Phosphatase, Non-Receptor Type 6 ; Proto-Oncogene Proteins c-kit ; metabolism ; fas Receptor ; genetics ; metabolism

The aim of study is to investigate the expression of hematopoietic cell phosphatase (SHP-1) gene and c-kit pro-oncogene in acute leukemia (AL) and its impact on prognosis in AL. Semi-quantity reverse transcriptase-polymerase chain reaction (RT-PCR) was used to detect the expression of SHP-1 mRNA and c-kit mRNA in 60 AL patients and 33 normal controls (NC). The results showed that the positive rates of SHP-1 expression from high to low level were found orderly in complete remission group, newly diagnosed group and relapsed group, there was significance difference between each group and NC group (P < 0.05). The positive rates of c-kit expression were opposite order in each groups as compared with SHP-1. there was also significance difference between each group and NC group (P < 0.05). The positive rate of SHP-1 and c-kit expressions in AML was higher than that in ALL (P < 0.05), there was negative correlation between expressions of SHP-1 and c-kit (r = -0.502, P < 0.05); The difference between the complete remission rate in SHP-1 positive and in SHP-1 negative patients from 30 newly diagnosed AML patients was significant (P < 0.05), the same result was found between c-kit(+) complete remission and c-kit(-) complete remission. It is concluded that SHP-1 gene is a potentially anti-oncogene and inhibits the growing of tumor by negatively modulating c-kit gene. Simultaneous detection of SHP-1 and c-kit gene may act as a factor for predicting prognosis in AL.
Adolescent ; Adult ; Aged ; Female ; Humans ; Leukemia, Myeloid, Acute ; genetics ; metabolism ; Male ; Middle Aged ; Polymerase Chain Reaction ; Precursor Cell Lymphoblastic Leukemia-Lymphoma ; genetics ; metabolism ; Prognosis ; Protein Tyrosine Phosphatase, Non-Receptor Type 6 ; biosynthesis ; genetics ; metabolism ; Proto-Oncogene Proteins c-kit ; biosynthesis ; genetics

The hematopoietic system of the mouse arises from extraembryonic mesoderm that migrate through primitive streak to the presumptive yolk sac at day 7.0 of gestation. However, the mechanisms regulating mesoderm commitment to hematopoietic lineages remain poorly understood. Previous studies demonstrated that the development kinetics and growth factor responsiveness of hematopoietic precursors derived from embryonic stem cells (ES cells) is similar to that found in the yolk sac, indicating that the onset of hematopoiesis within the embryoid bodies (EBs) parallels that found in the embryo. Furthermore, in vitro differentiation of ES cells to hematopoietic cells is valuable for establishment of therapeutic clone against a variety of hematological disorders. Despite the identification of multipotential hematopoietic progenitors in EBs, a subset of more primitive progenitors, identical to the high proliferative potential colony-forming cells (HPP-CFC) derived from human and murine hematopoietic tissues, have not been clearly identified regarding particular their replating potential in vitro. HPP-CFC is among the most primitive hematopoietic multipotent precursors cultured in vitro. In this study, our aim was to investigate the in vitro and in vivo hematopoietic capacity of HPP-CFC within the day 12 EBs, rather than the expansion of more committed progenitors. In this study the HPP-CFC could be detected within EBs differentiated for 5 to 14 days of murine ES cells, but the development dynamics of the HPP-CFC differed greatly among distinct serum lots. Qualitatively HPP-CFC is capable of forming secondary colonies. As to our expectation the ES cells-derived HPP-CFC demonstrated similar regeneration capacity to those from yolk sac, giving rise to secondary granulocyte, erythrocyte, macrophage and mast cells, however largely differed from the counterparts of adult bone marrow. In addition, by RT-PCR ES cells-derived HPP-CFC were found to express transcription factors associated closely with stem cell proliferation including SCL, GATA-2 and AML1 as well as various receptors of hematopoietic growth factors such as c-kit, GM-CSF receptor and interleukin 3 receptor et al. Finally, in order to understand the in vivo hematopoietic capacity of the ES cells-derived HPP-CFC, spleen colony-forming unit (CFU-S) assay was performed. Nevertheless, typical CFU-S was not observed after transplantation of the day 12 EB cells or HPP-CFC colonies into lethally irradiated adult murine. In conclusion the HPP-CFC differentiated from murine ES cells displayed robust hematopoietic activity in vitro, however their in vivo reconstitution ability was not detected. The difference between in vitro and in vivo hematopoietic activities of ES cells-derived primitive hematopoietic precursors deserves further investigation.
Animals ; Basic Helix-Loop-Helix Transcription Factors ; genetics ; Cell Differentiation ; genetics ; physiology ; Colony-Forming Units Assay ; Core Binding Factor Alpha 2 Subunit ; genetics ; Embryonic Stem Cells ; cytology ; GATA2 Transcription Factor ; genetics ; Hematopoietic Stem Cells ; cytology ; metabolism ; Humans ; Mice ; Proto-Oncogene Proteins ; genetics ; Proto-Oncogene Proteins c-kit ; genetics ; Receptors, Granulocyte-Macrophage Colony-Stimulating Factor ; genetics ; Receptors, Interleukin-3 ; genetics ; Reverse Transcriptase Polymerase Chain Reaction ; T-Cell Acute Lymphocytic Leukemia Protein 1

Over the past 60 years, investigators of basic science, pathology, and clinical medicine have studied gastrointestinal stromal tumor (GIST) and made minor advances in patient care. Recent discoveries have led to an understanding of the biological role of KIT and platelet-derived growth factor receptor-α in GIST and the development of the tyrosine kinase inhibitor imatinib mesylate (Gleevec, formerly STI-571), one of the most exciting examples of targeted therapy to date. The success of targeted therapy in GIST has lead to new developments in our understanding of the medical and surgical management of the disease. Intense study of GIST may lead to new paradigms in the management of cancer.
Antineoplastic Agents ; therapeutic use ; Benzamides ; Combined Modality Therapy ; Drug Delivery Systems ; Gastrointestinal Neoplasms ; drug therapy ; genetics ; pathology ; surgery ; Gastrointestinal Stromal Tumors ; drug therapy ; genetics ; pathology ; surgery ; Humans ; Imatinib Mesylate ; Mutation ; Piperazines ; therapeutic use ; Protein Kinase Inhibitors ; therapeutic use ; Proto-Oncogene Proteins c-kit ; genetics ; metabolism ; Pyrimidines ; therapeutic use ; Receptor, Platelet-Derived Growth Factor alpha ; genetics ; metabolism