Socs-1 gene methylation in cancer
a gene methylation and cancer technology, applied in the field of epigenetic markers indicative of cancer, can solve the problems of unregulated cell proliferation, physical damage to the dna in an individual, and the cell containing the damaged dna often dies, so as to facilitate the entry of polypeptides, facilitate transport, and facilitate cell permeability.
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example 1
Methylation of SOCS-1 is Associated with Hepatocellular Carcinoma
[0126]This example demonstrates that hypermethylation of the SOCS-1 gene and consequent decreased expression of SOCS-1 correlate with unregulated growth of hepatocellular carcinoma (HCC) cells, and that agents that circumvent the decreased SOCS-1 expression induce apoptosis of the HCC cells.
Methods
Cell Lines and Tissue Samples
[0127]Human hepatocellular carcinoma cell (HCC) lines SNU-182, SNU-423, SNU-387, SNU-398, SNU-449, SNU-475, and PLC / PRF / 5 were obtained from American Type Culture Collection. GM06061C was obtained from NIGMS repository. HuH-1, HuH-4, HuH-7 and Hep3B were obtained from Japanese Culture Collection. The cells were grown in DMEM or RPMI 1640 supplemented with 10% fetal bovine serum (FBS) for isolation of DNA and RNA. Primary hepatocarcinoma samples and adjacent normal liver tissue are described by Rashid et al. (Brit. J. Cancer 80:59-66, 1999).
Northern Blot Analysis
[0128]Poly(A) RNA of HCC cell lines ...
example 2
Methylation of SOCS-1 is Associated with Multiple Myeloma
[0166]This Example demonstrates that SOCS-1 gene methylation silencing also occurs in multiple myeloma (MM) cells.
Methods
Multiple Myeloma Patient and Control Samples
[0167]After informed consent was given, bone marrow (BM) specimens were aspirated during routine clinical assessment of 35 mM patients, who presented at the University Hospital Aachen, Germany between 1995 and 2000. MM diagnosis and staging classification were made in accordance with standard criteria (Durie, Sem. Oncol. 13:300-309, 1986). Two control bone marrow aspirates were obtained from patients with non-metastatic solid tumors or malignant lymphoma without bone marrow infiltration or hematopoietic dysfunction as part of the routine staging procedure. Peripheral blood (PB) samples were collected from five healthy volunteers. Mononuclear cells from BM and PB were separated by density gradient centrifugation prior to further analysis. Details of the lymphoma sam...
example 3
Methylation of SOCS-1 is Associated with Leukemia
[0186]This Example demonstrate that hypermethylation of the SOCS-1 gene also is associated with acute leukemias.
Methods
Primary Leukemia Samples
[0187]Adult leukemia were obtained from the University Hospital Aachen, Germany. The pediatric have previously been described for inactivation at the INK4A and INK4B locus (Herman et al., supra, 1997).
Cell Culture and Drug Treatment
[0188]HL60 cells were obtained from the American Type Culture Collection and KG1a cells from the German Collection of Microorganisms and Cell Cultures. Both cell lines were routinely cultured in IMDM (Invitrogen Corp.) with 20% FCS (Gemini Bio-Products). In order to assess the sensitivity to AG490, HL60 and KG1a cells were incubated with or without a final concentration of 50 μM AG490 (Sigma) for 96 hr prior to analysis.
Flow Cytometric Analysis of Apoptosis
[0189]The percentage of apoptotic cells was determined by annexin V-binding and counterstaining with propidium i...
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