Molecular Cloning of HumanGRB-7Co-amplified withCAB1and c-ERBB-2in Primary Gastric Cancer

Kishi, Tatsuya; Sasaki, Hiroki; Akiyama, Nobu; Ishizuka, Tomoki; Sanada, Hiromi; Aizawa, Shigeo; Sügimura, Takashi; Terada, Masaaki

doi:10.1006/bbrc.1997.6218

Cited by 46 publications

(34 citation statements)

References 22 publications

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“…LIF gene expression occurs at di erent stages of development, usually at very low levels (data not shown). Comparing with a standard cDNA library screening method, our modi®ed cDNA selection method was shown to be more e ective for cloning a cDNA derived from a transcript whose expression level was too low to be detected by Northern blot analysis (Ide et al, 1997;Kishi et al, 1997;. Thus, our modi®ed cDNA selection method was used for isolation of a rat LIF cDNA clone containing complete coding sequence.…”

Section: Resultsmentioning

confidence: 99%

Molecular cloning and functional analysis of cDNA encoding a rat leukemia inhibitory factor: towards generation of pluripotent rat embryonic stem cells

Takahama¹,

Ochiya²,

Sasaki³

et al. 1998

Oncogene

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Embryonic stem (ES) cells are pluripotent cell lines established directly from the early embryo. Maintenance of the stem-cell phenotype of ES cells in vitro requires the presence of a feeder layer of ®broblasts or of a soluble factor, di erentiation inhibitory activity (DIA) such as leukemia inhibitory factor (LIF). Here we report the cloning of complete rat LIF cDNA and its nucleotide sequence so as to facilitate studies of rat ES cell technologies on tumor biology. The nucleotide sequence of the rat LIF cDNA indicated that the rat LIF has 91% amino acid sequence identity with murine LIF. The cloned rat LIF cDNA has a putative biological activity as a di erentiation-inducing factor on the murine myeloid leukemia cell line M1 cells. Culture supernatant of the rat LIF cDNA-transduced rat ®broblast cell line could maintain the stem-cell phenotype of rat ES cells which showed alkaline phosphatase activity, and this e ect was much stronger than that by murine LIF. The availability of rat LIF which shows DIA will assist the in vitro analysis of rat ES cells, and culture of these cells is a route for the generation of gene targeting in rat.

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Section: Resultsmentioning

confidence: 99%

Molecular cloning and functional analysis of cDNA encoding a rat leukemia inhibitory factor: towards generation of pluripotent rat embryonic stem cells

Takahama¹,

Ochiya²,

Sasaki³

et al. 1998

Oncogene

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“…Grb7 is found coamplified with ErbB2 in breast, esophageal, and gastric adenocarcinoma cells (Stein et al, 1994;Kishi et al, 1997;Tanaka et al, 1998). This coamplification nicely correlates with the enhanced invasive capacity of these tumor cells, suggesting that Grb7 plays a role in ErbB2-mediated cell motility and extramucosa tumor cell invasion (Tanaka et al, 1997).…”

Section: Introductionmentioning

confidence: 81%

The adaptor Grb7 is a novel calmodulin-binding protein: functional implications of the interaction of calmodulin with Grb7

Sánchez-Torres

Carpio

et al. 2005

Oncogene

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We demonstrate using Ca 2 þ -dependent calmodulin (CaM)-affinity chromatography and overlay with biotinylated CaM that the adaptor proteins growth factor receptor bound (Grb)7 and Grb7V (a naturally occurring variant lacking the Src homology 2 (SH2) domain) are CaM-binding proteins. Deletion of an amphiphilic basic amino-acid sequence (residues 243-256) predicted to form an a-helix located in the proximal region of its pleckstrin homology (PH) domain demonstrates the location of the CaM-binding domain. This site is identical in human and rodents Grb7, and shares great homology with similar regions of Grb10 and Grb14, and the Mig10 protein from Caenorhabditis elegans. We show that Grb7 and Grb7V are present in the cytosol and bound to membranes, while the deletion mutants (Grb7D and Grb7VD) have less capacity to be associated to membranes. Grb7D maintains in part the capacity to bind phosphoinositides, and CaM competes for phosphoinositide binding. Activation of ErbB2 by heregulin b1 decreases the pool of Grb7 associated to membranes. The cell-permeable CaM antagonist W7 (N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide), but not the CaM-dependent protein kinase II inhibitor KN93, prevents this effect. Highly specific cell-permeable CaM inhibitory peptides decrease the association of Grb7 to membranes. This suggests that CaM regulates the intracellular mobilization of Grb7 in living cells. Direct interaction between enhanced yellow fluorescent protein (EYFP)-Grb7 and enhanced cyan fluorescent protein (ECFP)-CaM chimeras at the plasma membrane of living cells was demonstrated by fluorescence resonance energy transfer (FRET). The FRET signal dramatically decreased in cells loaded with a cellpermeable Ca 2 þ chelator, and was significantly attenuated when enhanced yellow fluorescent protein-Grb7 chimera (EYFP-Grb7)D instead of EYFP-Grb7 was used. Finally, we show that conditioned media from cells transiently transfected with Grb7D and Grb7VD lost its angiogenic activity, in contrast to those from cells transiently transfected with their wild-type counterparts.

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“…Grb7 is an especially promising cancer target as the encoding gene maps closely to the ErbB2 gene on human chromosome 17q12 and is found co-amplified and overexpressed in a subset of breast, esophageal and gastric cancers (Stein et al, 1994;Kishi et al, 1997;Tanaka et al, 1997;Fiddes et al, 1998;Kauraniemi et al, 2001;Varis et al, 2002;Andrechek et al, 2003;Bieche et al, 2003). Analysis of chromosome 17q12 in a variety of different breast cancer cell lines Grb7 was found to have a high DNA copy number with a high level of ErbB2 co-expression (Kauraniemi et al, 2001).…”

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confidence: 99%

Combination treatment with Grb7 peptide and Doxorubicin or Trastuzumab (Herceptin) results in cooperative cell growth inhibition in breast cancer cells

et al. 2007

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Grb7 has potential importance in the progression of cancer. We have previously identified a novel peptide that binds to the SH2 domain of Grb7 and inhibits its association with several different receptor tyrosine kinases. We have synthesised the Grb7 peptide, G7-18NATE, with two different cell penetrating peptides, Penetratin and Tat. In this study, we have shown that both Penetratin-and Tat-conjugated G7-18NATE peptides are able to inhibit the proliferation of SK-BR-3, ZR-75-30, MDA-MB-361 and MDA-MB-231 breast cancer cells. There was no significant effects on breast cancer MCF-7cells, non-malignant MCF 10A or 3T3 cells. In addition, there was no significant inhibition of proliferation by Penetratin or Tat alone or by their conjugates with arbitrary peptide sequence in any of the cell lines tested. We determined the EC 50 of G7-18NATE-P peptide for SK-BR-3 cell proliferation to be 7.663 Â 10 À6 M. Co-treatment of G7-18NATE-P peptide plus Doxorubicin in SK-BR-3 breast cancer cells resulted in an additional inhibition of proliferation, resulting in 56 and 84% decreases in the Doxorubicin EC 50 value in the presence of 5 Â 10 À6 and 1.0 Â 10 À5 M G7-18NATE-P peptide, respectively. Importantly, the co-treatment with Doxorubicin and the delivery peptide did not change the Doxorubicin EC 50 . Since Grb7 associates with ErbB2, we assessed whether the peptide inhibitor would have a combined effect with a molecule that targets ErbB2, Herceptin. Co-treatment of Herceptin plus 1.0 Â 10 À5 M G7-18NATE-P peptide in SK-BR-3 cells resulted in a 46% decrease in the Herceptin EC 50 value and no decrease following the co-treatment with Herceptin and penetratin alone. This Grb7 peptide has potential to be developed as a therapeutic agent alone, in combination with traditional chemotherapy, or in combination with other targeting molecules.

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Molecular Cloning of HumanGRB-7Co-amplified withCAB1and c-ERBB-2in Primary Gastric Cancer

Cited by 46 publications

References 22 publications

Molecular cloning and functional analysis of cDNA encoding a rat leukemia inhibitory factor: towards generation of pluripotent rat embryonic stem cells

Molecular cloning and functional analysis of cDNA encoding a rat leukemia inhibitory factor: towards generation of pluripotent rat embryonic stem cells

The adaptor Grb7 is a novel calmodulin-binding protein: functional implications of the interaction of calmodulin with Grb7

Combination treatment with Grb7 peptide and Doxorubicin or Trastuzumab (Herceptin) results in cooperative cell growth inhibition in breast cancer cells

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