Biomedical subjects

G E Gallick

Publications and source records attributed to G E Gallick.

At least 37 records · Page 2Linked to original sources

PubMed1993

Transforming growth factor-alpha production and autoinduction in a colorectal carcinoma cell line (DiFi) with an amplified epidermal growth factor receptor gene.

The DiFi colorectal carcinoma cell line, derived from a patient with familial adenomatous polyposis, was examined for gene expression and production of the autocrine growth factor transforming growth factor alpha (TGF-alpha) and for epidermal growth factor receptor (EGFR) gene expression and gene copy number. DiFi cells expressed TGF-alpha transcripts as identified on Northern (RNA) blots. Addition of TGF-alpha (10 ng/ml) or EGF (10 ng/ml) to DiFi cell cultures (lacking EGF or serum) up-regulated DiFi cell basal TGF-alpha mRNA levels, suggesting that autoinduction of TGF-alpha occurs in these cells. DiFi cell cultures in log phase growth secreted measurable amounts of TGF-alpha (347 pg/10(6) cells/24 h) into their culture medium, as determined by radioimmunoassay. DiFi cells showed strong overexpression of the EGFR gene on Northern blots relative to three other colon cancer cell lines examined. Immunoperoxidase staining showed enhanced EGFR expression in a cell subpopulation among the original (uncultured) ascitic fluid cells from which the DiFi cell line was established. This cell subpopulation was observed to expand dramatically between passages 1 and 25. Immune complex kinase assay of DiFi cells showed that EGFR were functional as determined by their ability to autophosphorylate. The EGFR gene in these cells was not found to be rearranged or genetically altered using Southern blot analysis. Dot blot analysis of DiFi cell DNA revealed EGFR gene amplification in the range of 60-80 copies/cell, which is approximately twice the copy number seen in A-431 epidermoid carcinoma cells. To our knowledge DiFi cells represent the first example of EGFR gene amplification in a colorectal adenocarcinoma. Because DiFi colorectal cancer cells uniquely show production and auto-induction of TGF-alpha in addition to amplification and overexpression of the EGFR gene, these cells represent a valuable tool for studying the role(s) of the EGFR in the regulation of tumor cell growth.

Blotting, Northern↗

PubMed1993

Down-modulation of epidermal growth factor receptor accompanies TNF-induced differentiation of the DiFi human adenocarcinoma cell line toward a goblet-like phenotype.

Although the biologic response modifier tumor necrosis factor-alpha (TNF) is a known differentiation inducer in hematopoietic cells, its role in differentiation of other tissue types has yet to be elucidated. In the studies presented here, TNF treatment of the human rectal adenocarcinoma cell line, DiFi, elicits characteristics of early stage differentiating, mucin-producing colonocytes. Not only are TNF-treated DiFi cells growth-inhibited by TNF, but they also display a unique morphology. Additionally, TNF treatment of DiFi cells enhances > fivefold the expression of high molecular weight mucin glycoproteins, as measured by [125I]-wheat germ agglutinin (WGA) binding and the human milk fat globule-1 (HMFG-1) anti-MUC1 antibody reactivity. The induction of these differentiation characteristics correlates with novel alterations in epidermal growth factor receptor (EGF-R). Following 5-day TNF treatment of DiFi cultures, EGF receptor levels, kinase autophosphorylation activity, and receptor tyrosine phosphorylation are reduced by > fourfold. The establishment of a model system in which goblet-like cell characteristics and alterations in a growth factor receptor can be induced in vitro may be potentially useful in studying the underlying mechanisms of colonic epithelial cell proliferation and differentiation.

Adenocarcinoma↗

PubMed1993

Site-specific differences in pp60c-src activity in human colorectal metastases.

The c-src proto-oncogene has been implicated in the progression of primary human colorectal carcinoma to hepatic metastasis. To determine if increased pp60c-src tyrosine kinase activity is a colon-specific phenomenon present in colorectal metastases to all sites, the pp60c-src-specific kinase activity of noncolon tumor metastases to the liver was compared to that of colorectal liver metastases. Activity of extrahepatic colon carcinoma metastases was compared to that of colorectal liver metastases as well as that of normal colonic mucosa. The specific activity of pp60c-src in multiple synchronous metastases from colon carcinoma was also examined. Tyrosine kinase activity was determined by immune complex kinase assay; protein levels were determined by immunoblotting. Specific activity was calculated for each group by dividing the total activity by protein level. Colon carcinoma metastases to the liver had significantly (P < 0.04) increased pp60c-src activity with an average 2.2-fold increase over normal mucosa. In contrast, noncolon tumor metastases to the liver showed minimal pp60c-src kinase activity. Extrahepatic colorectal metastases demonstrated significantly increased (P < 0.005) pp60c-src activity with an average 12.7-fold increase over normal mucosa. When compared to colon liver metastases, extrahepatic colorectal tumor metastases show a significant difference in activity (P < 0.05) with an average 5.7-fold increase. Examination of multiple synchronous colon carcinoma metastases confirmed these results. In summary, we conclude that (1) the activation of pp60c-src between primary tumors and metastases is specific to colon metastases, and (2) although pp60c-src activity is significantly increased in colorectal metastases, site-specific differences in the magnitude of activity are evident.

Adenocarcinoma↗

PubMed1993

Increase in activity and level of pp60c-src in progressive stages of human colorectal cancer.

Activation of the tyrosine kinase of the c-src gene product, pp60c-src, has been shown to occur in nearly every primary colorectal carcinoma, and is found as early as in polyps of high malignant potential. However, no studies have addressed potential pp60c-src changes which occur during progression. To examine this question, we have studied kinase activity and protein levels in 7 colonic polyps, 19 primary lesions, and 19 liver metastases relative to normal colonic mucosa. Significant increases in tyrosine kinase activity were seen as early as in colonic polyps of high malignant potential. Further increases were observed in activity and level in primary tumors. However, the greatest increases in activity and protein levels were observed in liver metastases. Additionally, six metastatic lesions were obtained in which synchronous primary tumor was resected. In each of these liver metastases, pp60c-src activity and level were significantly increased relative to the corresponding primary tumor, as well as to normal colonic mucosa. Our results demonstrate that progression of colon primary tumors to liver metastases correlates with increased pp60c-src kinase activity and protein level.

Adenocarcinoma↗

PubMed1992

Autocrine growth stimulation by transforming growth factor-alpha in human non-small cell lung cancer.

We studied the biological response to and production of transforming growth factor-alpha (TGF-alpha) by the non-small cell lung carcinoma (NSCLC) clonal cell lines H226b, H322a, H460a, H596b. Each of these cell lines expressed epidermal growth factor receptor (EGFR) as determined by [125I]EGF competitive binding and Scatchard analysis and by phosphorylation. The receptors were functionally active as determined in immune complex kinase assays. H322a, H226b, H460a, and H596b cells showed stimulated [3H]thymidine (Thd) uptake in response to TGF-alpha. Exogenously added TGF-alpha increased colony formation in soft agar for three of the cell lines in media containing serum. All cell lines expressed TGF-alpha detected by immunohistochemistry and TGF-alpha mRNA, although to differing degrees. Cell lysates and spent media competed for EGFR binding with EGF, thus demonstrating production of TGF-alpha-like activity. The anti-TGF-alpha monoclonal antibody AB-3 inhibited the uptake of [3H]Thd by proliferating H322a and H226b cells but not H460a and H596b cells. No inhibition occurred with MOPC21 antibody and inhibition was completely reversed by addition of TGF-alpha to the culture. Suramin inhibited cell proliferation and [3H]Thd uptake by all cell lines. Inhibition of H460a and H596b cells was reversed with exogenous TGF-alpha but not PDGF. Our data suggests that TGF-alpha is a mediator of autocrine growth stimulation for NSCLC cells, and that for some NSCLC cells cytoplasmic binding of receptor and ligand is the primary mechanism for autocrine growth stimulation.

Animals↗

PubMed1992

Growth inhibition of human colorectal carcinoma cell lines by tumor necrosis factor-alpha correlates with reduced activity of pp60c-src.

One molecular alteration that has been observed in the majority of colorectal carcinomas is the activation of pp60c-src kinase. To address the role of pp60c-src in growth control of colon carcinoma cell lines, the effects of the biologic response modifier, tumor necrosis factor (TNF)-alpha were studied on the established HT29 colorectal carcinoma cell line and two clonal variants derived from the parental line. In one clone, HT29-A34, approximately 55% growth inhibition was observed following a 120-h incubation period with 10(3) U/ml TNF. In this TNF-sensitive cell line, pp60c-src immune complex kinase activity was reduced 3.9-fold accompanied by only a slight reduction in pp60c-src protein levels. Growth inhibition and decreased pp60c-src kinase activity correlated in a dose-dependent manner. The HT29-A14 TNF-resistant clone was not growth-inhibited by TNF, and no changes in pp60c-src were observed following treatment. Growth inhibition also correlated with reduced pp60c-src kinase specific activity in a number of other established colon carcinoma cell lines that were TNF-sensitive. In TNF-resistant colon carcinoma cell lines, pp60c-src kinase activity and levels remained unchanged. When changes in specific activity of pp60c-src were observed in sensitive cells, they occurred after decreased [3H]-thymidine uptake was observed. Therefore, these changes in pp60c-src activity are not the earliest event in TNF-induced growth inhibition. Nevertheless, our results suggest that modulation of pp60c-src kinase activity may be important in growth control of colorectal carcinoma cell lines.

Adenocarcinoma↗

PubMed1992

Suppression by retinoic acid of epidermal growth factor receptor autophosphorylation and glycosylation in cultured human head and neck squamous carcinoma cells.

The epidermal growth factor receptor (EGF-R) gene is overexpressed or amplified in various human squamous cell carcinomas, including those of the head and neck (HNSCC). Earlier we found that beta-all-trans-retinoic acid (RA) inhibited the growth and suppressed the aberrant squamous cell differentiation of several cultured HNSCC cell lines. Here we examined the effects of RA on the expression and function of EGF-R in two HNSCC cell lines, 1483 and 183, which exhibit distinct states of squamous cell differentiation, EGF-R mRNA levels, and responses to the growth inhibitory effects of RA. Treatment with RA (1 microM, 7 days) of the RA-sensitive 1483 cells decreased the level of EGF-R mRNA two- to four-fold and the binding of 125I-EGF to the cell surface by 30%-35%. In contrast, RA treatment of the 183 cells did not alter the EGF-R mRNA level or the binding of 125I-EGF. Other effects of RA on EGF-R structure and function were similar in both cell lines. RA did not alter the amount of immunoprecipitable [35S]methionine-labeled cellular EGF-R, 125I-cell surface labeled EGF-R, EGF-R internalization, or transforming growth factor alpha (TGF-alpha) mRNA. More important, RA treatment of both cell lines decreased EGF-R autophosphorylation activity detected in immune-complex-kinase assay by about three- and five-fold in the 1483 and 183 cells, respectively. Likewise, RA decreased the glycosylation of EGF-R in both cell lines. In the 1483 cells, RA suppressed the incorporation of either glucosamine or fucose by about 50%, whereas in the 183 cells RA suppressed the incorporation of fucose by about 80%. These results demonstrate that RA can modify the structure of the EGF-R by decreasing its glycosylation and suggest that these changes may suppress the autophosphorylation activity of the receptor kinase. The RA-induced changes in EGF-R do not correlate with the effect of RA on the growth of the cells but may be related to the suppression of squamous cell differentiation in the 1483 cells.

Carcinoma, Squamous Cell↗

PubMed1992

Transfection of activated c-H-rasEJ/pSV2neo or pSV2neo genes into rat mammary cells: rapid stimulation of clonal diversification of spontaneous metastatic and cell-surface properties.

We examined whether the conversion of benign rat mammary cells to metastatic cells by transfection of c-H-rasEJ/pSV2neo or control pSV2neo genes results in rapid stimulation of diversification of cellular phenotypes. Transfection of c-H-rasEJ into twice-cloned, stable (greater than 1 year) rat mammary MTC.4 cells, followed quickly by cell cloning, revealed differences in transfected oncogene copy numbers and expression of p21rasEJ. No correlation between c-H-rasEJ copy numbers or the cellular amounts of p21rasEJ or total p21ras and spontaneous metastatic potentials was found. By subcloning transfected cells as soon as possible after gene transfer, we found some rearrangements and amplifications of c-H-rasEJ and heterogeneous spontaneous metastatic potentials. In addition, the expression of the mammary tumor metastasis-associated cell-surface glycoprotein gp580 on untransfected and transfected MTC.4 cells indicated that the cell populations of higher metastatic potential were also more diverse in their cell-to-cell antigen expression than untransfected or non-metastatic, transfected MTC.4 cells. In contrast, the expression of a putative metastasis-suppressor gene, nm23, was unchanged after transfection and subcloning. Control pSV2neo transfections or calcium phosphate treatment alone also resulted in the generation of cellular heterogeneity, although at an apparently lower frequency than c-H-rasEJ transfections, suggesting that transfection of activated, dominantly acting oncogenes, or in some cases control genes, can result in destabilization of transfected cells, rapid diversification and generation of heterogeneity in growth rate, spontaneous metastatic potential and antigen expression.

Animals↗

PubMed1991

Cellular growth response to epidermal growth factor in colon carcinoma cells with an amplified epidermal growth factor receptor derived from a familial adenomatous polyposis patient.

The receptor binding and cellular growth responses to exogenous epidermal growth factor (EGF) were studied using the DiFi cell line established from a familial adenomatous polyposis patient. The number of cell membrane EGF receptors on DiFi cells, as measured by competitive radioligand binding assays and Scatchard analysis of 125I-EGF binding isotherms, was calculated to be 4.8 x 10(6) receptors/cell. An acid prewash step performed prior to ligand binding assays did not reveal additional receptor numbers. A single, low-affinity receptor population was identified by Scatchard analysis, with an apparent Kd of 4.6 nM. This result was confirmed by radioligand binding studies performed in the presence and absence of the receptor-antagonist monoclonal antibody 528 IgG that binds predominantly to the low-affinity form of the EGF receptor. DiFi cells at 50-60% confluence, when exposed to 50 nM exogenous EGF, exhibited a rapid but partial (30%) reduction in their cell membrane-associated receptor, characteristic of sequestration. Exposure of DiFi cells to 50 nM EGF for longer periods of time (4 h) did not result in any further reduction in EGF-receptor number. The cellular growth response of DiFi cells to exogenous EGF was studied in monolayer cultures as well as in a soft agarose assay. Inhibition of soft agar colony formation was observed at exogenous EGF concentrations greater than 1.7 nM, and inhibition of monolayer growth occurred at EGF concentrations greater than 1 nM. In immune complex kinase assays, the DiFi receptor showed similar specific activity to that from the well-characterized A431 cell line. Additionally, phosphorylation of the receptor on tyrosine was qualitatively similar to that of A431 cells, further suggesting that the DiFi receptors identified by EGF-binding studies were biologically functional.

Adenomatous Polyposis Coli↗

PubMed1991

Susceptibility to ras oncogene transformation is coregulated with signal transduction through growth factor receptors.

The human teratocarcinoma cell line PA-1 was derived from culturing ascites fluid cells from a patient with an ovarian germ line tumor. We previously described a non-neoplastic variant cloned from the PA-1 human teratocarcinoma cell line, clone 6, which at passage 40 was resistant to transformation by activated ras oncogenes. However, these cells could be transformed by a plasmid containing both myc and ras. Another PA-1 cell variant, clone 1, isolated at passage 63 and used 50 passages later becomes tumorigenic in nude mice after transfection with an activated ras oncogene (Tainsky et al., Anticancer Res., 8, 899-914, 1988). We report here that the progression from ras resistance to ras susceptibility occurs in both clone 1 and clone 6 cells during 25 passages in culture. In the presence of epidermal growth factor, transforming growth factor-alpha, and basic fibroblast growth factor, the ras-transformable cells exhibit anchorage independent growth, whereas the ras-resistant cells can not be growth stimulated by these growth factors. Similarly, ornithine decarboxylase (ODC) activity was inducible in ras susceptible and ras transformed cells by these growth factors, but not in the ras resistant cells. These differences are not due to the level and activity of epidermal growth factor receptor or to the level of expression of 25 proto-oncogenes.

Blotting, Northern↗

PubMed1991

Effect of herbimycin A on growth and pp60c-src activity in human colon tumor cell lines.

The effect of herbimycin A, an ansamycin antibiotic which inhibits cellular transformation by retroviral tyrosine kinases, on the monolayer growth of seven colon tumor cell lines and one cell line established from normal colonic mucosa, CCL239, was examined. Each colon tumor cell line tested showed dose-dependent growth inhibition in response to herbimycin A. A 125ng ml-1 dose of the antibiotic caused greater than 40% growth inhibition in all colon tumor cell lines after two cell doublings. In contrast, at similar herbimycin A concentrations only 12% inhibition was observed in 'normal' CCL239 cells. No major morphologic changes were observed at the light microscopic level in any of the tumor cell lines or CCL239 cells in response to treatment with herbimycin A. Studies using the HT29 colon adenocarcinoma cell line showed dose-dependent inactivation of pp60c-src by herbimycin A, resulting in decreased autophosphorylation, enolase phosphorylation and steady-state levels, which correlated with cellular growth inhibition. Herbimycin A-induced reductions in pp60c-src kinase activity preceded changes in pp60c-src steady-state levels. Growth and pp60c-src inhibition were reversible following removal of herbimycin A from cell culture media. Our results suggest that regulation of pp60c-src tyrosine kinase activity may be important in growth control of colon tumor cells.

Adenocarcinoma↗

PubMed1990

Lack of correlation between intercellular junctional communication, p21rasEJ expression, and spontaneous metastatic properties of rat mammary cells after transfection with c-H-rasEJ or neo genes.

The loss of intercellular junctional communication between rat 13762NF mammary carcinoma cells and their spontaneous metastatic potentials from the mammary fat pad show a high degree of correlation. We examined a stable, benign, completely junctionally coupled cell clone (MTC.4) of this system after calcium phosphate-mediated transfection with c-H-rasEJ/pSV2neo and control pSV2neo-containing plasmids. There was a good correlation between the copy numbers of c-H-rasEJ incorporated into MTC.4 cells and their contents of p21rasEJ; however, there was not always a correspondence between spontaneous metastatic potential and copy number of c-H-rasEJ or amount of p21rasEJ. After c-H-rasEJ/pSV2neo transfection, some MTC.4 cells lost intercellular junctional communication and became spontaneously metastatic, although some nonmetastatic transfectants also had low percentages of junctionally coupled cells. One of the control pSV2neo transfectants also became metastatic and lost intercellular junctional coupling, and calcium phosphate treatment itself resulted in increased growth rates at mammary fat pad sites and a marginal increase in incidence of spontaneous metastases, both of which preceded loss of intercellular junctional coupling in some cells. Examination of 12 subclones derived from two cloned transfectants, however, revealed a poor correlation between spontaneous metastatic potential and intercellular junctional coupling. The results suggest that loss of junctional communication between cells is often but not always associated with the progression of cells from benign to metastatic states.

Animals↗

PubMed1989

Tumor necrosis factor modulates epidermal growth factor receptor phosphorylation and kinase activity in human tumor cells. Correlation with cytotoxicity.

Tumor necrosis factor (TNF) is a cytokine which induces cytotoxicity in some but not all tumor cells. Initial studies of five tumor cell lines demonstrated that TNF was able to rapidly (within 30 min) modulate tyrosine protein kinase activity of epidermal growth factor (EGF) receptors on tumor cell lines which were sensitive to the cytotoxic effects of TNF but not alter EGF receptor kinase activity in TNF-resistant tumor cells. Two tumor cell lines (ME-180 cervical carcinoma and T24 bladder carcinoma) which have been shown to express similar TNF-binding characteristics but differ in their sensitivity to the cytotoxic actions of TNF were chosen for further characterization. Treatment of TNF-sensitive ME-180 cells with 1 nM TNF resulted in a 3-fold stimulation of EGF receptor tyrosine protein kinase activity within 10 min which correlated with increased phosphorylation of EGF receptor protein itself. In addition, dose-response studies indicate that similar concentrations of TNF modulate both ME-180 cell growth and EGF receptor kinase activity. Treatment of TNF-resistant T24 cells showed that TNF had no significant effect on their growth, EGF receptor tyrosine protein kinase activity, or phosphorylation of EGF receptor protein although EGF receptor kinase activity was stimulated by EGF. Quantitation of receptors expressed on the surface of ME-180 and T24 cells demonstrated a 3-fold difference between the number of EGF-binding sites on T24 (100,000) versus ME-180 cells (300,000), suggesting the relative abundance of EGF receptor does not solely account for differential effects of TNF on EGF receptor activation in these two cell lines. Phosphoamino acid analysis of EGF receptor from 32P-equilibrated ME-180 cells demonstrated that TNF-induced phosphorylation of amino acids which was quantitatively similar to that of EGF but distinct from the effects of phorbol ester. However, unlike EGF, TNF was unable to stimulate EGF receptor kinase activity in ME-180 cell lysates. The kinetics of EGF receptor activation and the metabolic consequence of activation of EGF receptor activity by TNF appear to be distinct from those induced by EGF. These results suggest that TNF-induced modulation of EGF receptor occurs through a unique mechanism and may play a role in the cytotoxic actions of TNF.

Carcinoma, Squamous Cell↗

PubMed1989

Epidermal growth factor receptor protein-tyrosine kinase activity in human cell lines established from squamous carcinomas of the head and neck.

Two cell lines established from tumors of the head and neck area at different clinical stages were found to differ in the expression and in the tyrosine kinase activity of the epidermal growth factor (EGF) receptor. Cell line 183A was derived from an early-stage tumor and cell line 1483 was derived from a tumor that had metastasized to lymph nodes. The 1483 cells displayed a higher plating efficiency and clonogenicity in soft agar, suggesting a more tumorigenic phenotype over the 183A cells. Analyses of EGF receptor levels by using R1 anti-EGF receptor serum indicated that the 1483 cells expressed 5-fold more receptor than did the 183A cells. EGF receptors isolated from each cell line were active for kinase activity in an immune complex kinase assay, using monoclonal R1 anti-EGF receptor antibody. The autophosphorylation activity of both receptors was stimulated by addition of EGF to isolated membrane preparations and intact cells, although the EGF receptor of the 1483 cells was much less responsive to EGF than the receptor from 183A cells. In addition, the 1483 receptor consistently incorporated about twice as much phosphate as did the 183A receptor in an immune complex kinase assay. These data suggest that the basal tyrosine kinase activity of the EGF receptor from 1483 cells may be more active than the EGF receptor kinase from 183 cells.

Adenosine Triphosphate↗

PubMed1989

Early events in the antiproliferative action of tumor necrosis factor are similar to the early events in epidermal growth factor growth stimulation.

The process of TNF-induced cytotoxicity is complex but appears to be mediated through a TNF-specific cell surface receptor. Recent evidence suggests that TNF action on tumor cells may be antagonized by epidermal growth factor (EGF) and other EGF-receptor modulatory peptides implicating a role for EGF-R in the process of TNF-induced cytotoxicity. In the present report, we investigated the biochemical actions of TNF on several biochemical events known to occur in the process of EGF signal transduction in intact cells. The actions of TNF were compared directly to those of EGF in both TNF-sensitive and -resistant tumor cell lines. In TNF-sensitive ME-180 cervical carcinoma cells, TNF (20 ng/ml) stimulated the tyrosine protein kinase activity of the EGF-receptor (EGF-R) fivefold when measured by receptor autophosphorylation in an immune complex kinase assay. TNF activation of EGF-R kinase activity in ME-180 was measurable 10 min after TNF incubation and enzymatic activity remained elevated 20 min after TNF addition. Activation of the receptor by TNF correlated with increased 32P incorporation into EGF-R protein when receptor was immunoprecipitated from 32P-equilibrated cells following a 20 min incubation with TNF. Acid hydrolysis of EGF-R protein isolated from TNF-treated ME-180 cells demonstrates an increase in the phosphotyrosine content of EGF-R when compared to receptor isolated from untreated cells. The results suggest that TNF increased EGF-R tyrosine protein kinase activity and the state of EGF-receptor tyrosine phosphorylation in a manner similar to that reported for EGF. However, TNF does not appear to be structurally related to EGF since TNF was unable to directly activate EGF-R when incubated with extensively washed immunoprecipitates of EGF-R. In TNF-resistant T24 bladder carcinoma cells, TNF failed to alter EGF-R tyrosine protein kinase activity although both EGF and phorbol ester were shown to modulate the enzymatic activity of the receptor in these cells. These results indicate that the ability of TNF to modulate EGF-R kinase in target cells may correlate with its cytotoxic actions on TNF-sensitive tumor cells. Other biochemical activities associated with the induction or regulation of cellular growth were examined in TNF- or EGF-treated tumor cells. EGF stimulated a rapid 8-16-fold increase in the expression of the proto-oncogene c-myc when analyzed by dot-blot analysis of total cellular RNA or Northern blot hybridization of polyadenylated RNA. TNF treatment failed to alter c-myc expression in ME-180 cells when analyzed by either technique.(ABSTRACT TRUNCATED AT 400 WORDS)

Cell Division↗

PubMed1988

Establishment and characterization of two new squamous cell carcinoma cell lines derived from tumors of the head and neck.

Two human cell lines were established from untreated squamous cell carcinomas of the head and neck. Line 183 was derived from a head and neck squamous cell carcinoma of the tonsil and 1483 from a head and neck squamous cell carcinoma of the retromolar trigone. Both lines grow in a cobblestone pattern demonstrating their epithelial heritage. Immunofluorescence studies and one-dimensional polyacrylamide gel electrophoresis indicated that both lines contain cytokeratins. Line 1483 is more aggressive in nude mice, has a higher efficiency for anchorage-independent growth, expresses p21ras (product of the ras oncogene) at a higher level, and is more aneuploid than 183. 1483 also grows as a multicellular tumor spheroid. Line 1483, which was established from the primary tumor of a patient with nodal metastasis, thus displays more progressed characteristics than line 183, which was established from a patient with no clinically positive nodes.

Aged↗

PubMed1988

Amplified and overexpressed epidermal growth factor receptor gene in uncultured primary human breast carcinoma.

We analyzed the epidermal growth factor receptor gene using a complementary DNA probe of the epidermal growth factor receptor gene in 21 uncultured primary breast carcinomas and found that the gene was amplified in three of these tumors. We further demonstrated by immunohistochemistry using a monoclonal antibody to the epidermal growth factor receptor that the receptor protein product of this gene was overexpressed and displayed elevated kinase activity. Our data indicate that one of the molecular mechanisms for overexpression of epidermal growth factor receptor in human breast cancer is epidermal growth factor receptor gene amplification without rearrangement in a subset of tumors.

Adult↗

PubMed1988

Gene expression and tumor cell escape from host effector mechanisms in murine large cell lymphoma.

Using in vivo selection methods, we obtained metastatic sublines of the murine RAW117 large cell lymphoma that form multiple liver metastases. The highly metastatic subline RAW117-H10 has a low number of gp70 molecules expressed at the cell surface and low cytostatic sensitivity to activated syngeneic macrophages. This subline was infected with endogenous RNA tumor virus isolated from a high virus-expressing RAW117-P subline of low metastatic potential. After superinfection the H10 subline gradually increased its expression of cell surface gp70 and showed enhanced sensitivity to macrophage-mediated cytostasis, suggesting that gp70 might be involved in host macrophage-mediated surveillance. Culture of RAW117-P and H10 cells in media conditioned by activated macrophages indicated that parental cells are severely growth inhibited in a dose dependent fashion while H10 cells showed almost no effect. Examination of differentially expressed genes in the highly metastatic RAW117-H10 cells by analysis of RNA blots indicated that a mitochondrial gene was expressed at a level that was approximately 10 times higher in H10 cells than in parental cells. This gene was identified as ND5, which codes for a subunit of NADH dehydrogenase (complex I of the mitochondrial electron transport chain); this complex is the target for an activated macrophage-released cytostatic factor. Among other possibilities, the results are consistent with the suggestion that highly metastatic RAW117 cells may escape macrophage surveillance by decreasing the synthesis of specific cell-surface receptors for cytostatic molecules and increasing the synthesis of specific cellular targets for such molecules.

Animals↗