Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P00750 (PLA)
 16,800 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Extracellular matrix (ECM), prepared from chick embryo fibroblasts, contains fibronectin as the major structural protein along with collagen and other polypeptides as less abundant protein components. When Rous sarcoma virus-transformed chick embryo fibroblasts are cultured on the ECM in the presence of the tumor promoter tetradecanoyl phorbol acetate, the transformed cells lose their characteristic rounded morphology and align on and within the ECM fibrillar network. This restrictive aspect of ECM is only temporary, however, and with time (24-72 h) the transformed cells progressively degrade the ECM fibers and resume their rounded appearance. The matrix degradation can be monitored by employing biosynthetically radiolabeled ECM. The addition of purified chicken plasminogen to the Rous sarcoma virus-transformed chick embryo fibroblast cultures enhances the rate and extent of ECM degradation, due to the elevated levels in the transformed cultures of plasminogen activator. Plasminogen-dependent and -independent degradation of ECM has been characterized with regard to sensitivity to various natural and synthetic protease inhibitors and to the requirement of cell/ECM contact. Plasminogen-dependent degradation of ECM occurs rapidly when ECM and cells are in contact or separated, whereas plasminogen-independent degradation is greatly reduced when ECM and cells are separated, which suggests that cell surface-associated proteolytic enzymes are involved. A possible role in ECM degradation has been indicated for cysteine proteases, metallo enzymes, and plasminogen activator, the latter as both a zymogen activator and a direct catalytic mediator.
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PMID:The extracellular matrix of normal chick embryo fibroblasts: its effect on transformed chick fibroblasts and its proteolytic degradation by the transformants. 299 35

The tumor induced RBC cytotoxicity assay has been used to explore the mechanism by which Rous sarcoma virus mutant transformed chick embryo fibroblasts and mouse 3T3 cells cause the cytolysis of RBC in vitro. All Rous sarcoma virus and viral mutant transformed cells were cytolytic for RBC. Three mutant viruses, tsGl251, rASV1702, and rASV157, appeared to cause quantitatively less cytolysis after transformation of chick embryo fibroblasts than other virally transformed cells. This decreased cytolytic activity may be correlated with decreased in vivo tumorigenicity. Temperature sensitive mutant transformed chick embryo fibroblasts and mouse 3T3, which were phenotypically normal and which secrete little if any plasminogen activator at non-permissive temperatures, were cytolytic at non-permissive temperatures. In addition, inhibitors of plasminogen activator and plasmin were ineffective inhibitors of cytotoxicity. The only effective inhibitor of cytotoxicity for both transformed chick embryo fibroblasts and mouse 3T3 cells was leupeptin. In Rous sarcoma virus transformed mouse 3T3 cells, the leupeptin inhibitable enzyme appears to be a plasma membrane enzyme.
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PMID:Role of proteases in red blood cell target cell destruction by cells transformed by Rous sarcoma virus mutants. 300 9

A monoclonal antibody has been raised against the serine protease, plasminogen activator (PA) produced by Rous sarcoma virus-transformed chick embryo fibroblasts (RSVCEF), and selected for its ability to inhibit the catalytic activity of PA. The high specificity and anticatalytic nature of the antibody has allowed probing of the direct role of PA in cellular behavior. Microgram quantities of monoclonal IgG inhibit the overgrowth and the morphological changes associated with RSVCEF transformation and the degradation of extracellular matrix mediated by RSVCEF, indicating a catalytic role for PA in these cellular processes. Specific cleavage of extracellular matrix proteins by immunoaffinity-purified PA in the complete absence of plasminogen demonstrates a direct catalytic involvement of PA in matrix degradation.
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PMID:An anticatalytic monoclonal antibody to avian plasminogen activator: its effect on behavior of RSV-transformed chick fibroblasts. 301 Dec 82

A pBR322::Rous sarcoma virus(RSV)-based shuttle vector was used to insert fused genes, composed of the amino-terminal portion of the bacterial chloramphenicol-acetyltransferase gene (cat) and the entire coding region for the C-terminally derived light (L) chain of human tissue-type plasminogen activator (t-PA) cDNA. Cotransfection of rat 3Y1 cells with pRSVneo DNA and pRSVcat/t-PA DNA yielded stably integrated G418-resistant transfectants which contain unrearranged copies of pRSVcat/t-PA DNA. These transfectants synthesize cat/t-PA L-chain mRNA, apparently correctly initiated and terminated. With the help of an enzyme-linked immunosorbent assay (ELISA), it is demonstrated that these cells produce human t-PA antigen. Furthermore, pRSVcat/t-PA L-chain cDNA-containing rat 3Y1 cells synthesize a plasminogen-dependent amidolytic activity which is suppressed by specific anti-human t-PA antibodies. This activity cannot be stimulated by fibrin, a property displayed by native t-PA. It is concluded that the t-PA L-chain cDNA contains the complete genetic information for the plasminogen activator activity.
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PMID:Functional analysis of the human tissue-type plasminogen activator protein: the light chain. 308 18

The results of four different assay methods showed that both normal and malignant plasminogen activator-secreting cells deposited substantial amounts of this protease on tissue-culture substrata, including collagen coatings. The cells studied were Rous sarcoma virus (RSV)-transformed vole fibroblasts, a malignant neural cell line (NG108-15) capable of neurite formation, and normal mouse-regenerating sensory neurons. Deposited plasminogen activator was detected by a fibrin overlay assay at sites from which cells growing on coverslips had been gently dislodged, showing that active enzyme is left beneath cells and in the immediate pericellular area. For neuronal cells, fibrinolytic zones were detected not only at the previous positions of cell bodies but also along the terrain conditioned by neurite extension, suggesting that a trail of plasminogen activator is left behind during growth cone movement. Substratum-bound enzyme could be solubilized in buffers containing sodium dodecyl sulfate (SDS) or Triton X-100 and demonstrated by zymography following electrophoresis or assayed for amidolytic activity with a chromogenic substrate (Kabi S-2251). The results suggest that plasminogen activator may be considered a component of substrate-adhesion material. Secretory proteases deposited directly on matrix molecules would seem strategically positioned to participate in local degradation of components of the extracellular environment.
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PMID:Normal and malignant cells, including neurons, deposit plasminogen activator on the growth substrata. 352 28

An avian retrovirus containing only the v-mil oncogene (PA200-MH2) was analyzed for its ability to induce a transformed phenotype in chicken embryo fibroblasts. Infected cultures exhibited an altered morphology, disarranged actin cable filaments, and a decrease in the amount of cell surface fibronectin. In addition, these cells showed a high level of plasminogen activator protease activity and were also capable of growth in low serum concentrations. In contrast, PA200-MH2 was very inefficient at inducing foci under agar and colonies in semisolid medium relative to the Mill Hill 2 and Rous sarcoma viruses. This inefficiency was further reflected in vivo by the total inability of PA200-MH2 to induce wing tumors in young birds. However, 40% of the birds inoculated in the wing web with PA200-MH2-infected cells did develop slow-growing tumors at the site of injection, with no evidence of hematopoietic involvement. Our results indicate that the v-mil oncogene is transforming both in vitro and in vivo and that each of the oncogenes in the Mill Hill 2 virus, v-mil and v-myc, can independently transform fibroblasts. These data suggest that v-mil is functionally related to its homologous murine counterpart, v-raf, which also transforms fibroblasts.
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PMID:Fibroblast transformation parameters induced by the avian v-mil oncogene. 357 49

Chicken embryo cells infected with partial transformation mutants of Rous sarcoma virus were tested for tumor-forming ability in chickens and in nude mice. Cells transformed by each of these partial transformation mutants display different combinations of transformation parameters. They therefore present a potentially favorable system for analyzing which properties of transformed cells are necessary for tumor formation. We found that the relative tumorigenicity of the virus mutants was generally similar in chickens and in nude mice, except that certain temperature-conditional mutants appeared to be sensitive to the differences in body temperature of the two experimental animals. (The body temperature of nude mice is 4 to 5 degrees C lower than that of chickens). Thus, the nude mouse appears to be a suitable system for testing the tumorigenicity of transformed chicken cells. Because mice are nonpermissive for Rous sarcoma virus infection and replication, it was possible to recover the transformed chicken cells from the tumors in this host and to determine what phenotypic changes they had undergone during tumor development. We also examined the relationship between various cellular properties of the virus-infected chicken cells in vitro and their tumorigenicity in nude mice. The combined results of these two studies indicated that anchorage independence and plasminogen activator production were highly correlated with the tumor-forming ability of these cells, whereas loss of fibronectin did not correlate with tumorigenicity. Furthermore, the inability of the least tumorigenic virus mutant to stimulate the phosphorylation of a 36,000-Mr target of pp60src raises the possibility that the 36,000-Mr protein plays a role in tumor formation.
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PMID:Tumorigenicity of partial transformation mutants of Rous sarcoma virus. 617 71

We have examined the phosphorylation state of five proteins known to become phosphorylated on tyrosine during transformation by Rous sarcoma virus by using cells infected with a panel of partially transforming mutant viruses. Situations of viral mutant and growth temperature were found in which phosphorylation of some proteins occurred more extensively than that of others, indicating that mutations in the src gene had affected the specificity of pp60src for some of its substrates as well as affecting the activity of the enzyme. To obtain insight into the biological functions of these phosphorylations, comparisons were made between the degree of phosphorylation of these proteins and the expression of various indicators of the transformed phenotype. The data suggest that phosphorylation of proteins l, p, and q (Mr of 46,000, 39,000 and 28,000, respectively) is not sufficient to induce changes in adhesiveness, hexose transport or morphology. The phosphorylation of protein p or l or total phosphotyrosine content correlated well with the production of plasminogen activator, and the phosphorylation of proteins l and q correlated well with increased hexose transport. However, even when good correlations were observed, significant exceptions were sometimes noted. It thus remains possible that some phosphorylations on tyrosine observed in Rous sarcoma virus-transformed cells are not causally related to the expression of the measured parameters of transformation.
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PMID:Phosphotyrosine-containing proteins and expression of transformation parameters in cells infected with partial transformation mutants of Rous sarcoma virus. 618 22

The mechanism by which Rous sarcoma virus transforms cells is better understood at the molecular level than that of any other oncogenic agent. The gene (src) responsible for transformation has been identified and its nucleotide sequence has been determined. The transforming protein (pp60src) has been identified and an enzymatic activity assigned to it. The unusual enzymatic activity of pp60src (phosphorylation of proteins on tyrosine) has allowed us to identify a large number of putative targets of this protein. And genetic evidence indicates that the phosphorylation of various targets is responsible for generating the various manifestations of the transformed phenotype. What can this model system contribute to understanding of hereditary large bowel cancer? First of all, it provides an intellectual paradigm for analyzing the mechanism by which a single autosomal dominant gene can alter the metabolism and regulatory behavior of a cell. A cellular homolog of src or of some other onc gene could be responsible for hereditary colon cancer. Second, it provides a model for understanding why some "markers" of malignancy are not invariably associated with cancer: since the oncogenic protein can interact with a variety of primary targets giving rise to the various parameters of transformation, not every sort of biological effect need be necessary for malignancy. Third, it points out that the various syndromes which constitute hereditary colon cancer may well be due to a single gene: since mutations in the src gene are capable of generating a variety of distinct phenotypic alterations in infected cells, different from that generated by the wild-type virus, it certainly is conceivable that different alleles of a single transforming gene could give rise to the different types of hereditary colon cancer. Whether this is the explanation for the various forms of hereditary colon cancer, or whether they result from the activities of several different onc genes can only be determined by identification of the gene(s) at the molecular level. Finally, this model system has provided information which may prove useful in improving the specificity of cancer chemotherapy. Since production of plasminogen activator seems to correlate well with growth in soft agar and tumorigenicity, an anti-cancer prodrug which is activated specifically by cells producing plasminogen activator might be selectively toxic to malignant cells. We have in fact synthesized such drugs and shown them to be selectively toxic in vitro to malignant cells (Carl et al 1980). In vivo tests of these agents are in progress.
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PMID:Cultured cells transformed by Rous sarcoma virus: a genetically defined model and its phenotype. 619 Jan 85

Many tumors contain elevated levels of plasminogen activator and thus produce elevated levels of the protease plasmin in the milieu of the tumor. We have hypothesized, therefore, that it should be possible to prepare peptidyl prodrug derivatives of anticancer drugs that would be locally activated by tumor-associated plasmin. As an initial test of this hypothesis, we synthesized the peptidyl prodrugs of the anticancer drugs (alpha S, 5S)-alpha-amino-3-chloro-4,5-dihydro-5-isoxazoleacetic acid (acivicin, AT-125) and N,N-bis(2-chloroethyl)-p-phenylenediamine (phenylenediamine mustard) by mixed anhydride coupling of the parent drug with the protected tripeptide, Boc-D-Val-Leu-Lys(Boc)-OH, followed by deprotection with trifluoroacetic acid. The prodrugs showed an increased selective in vitro cytotoxicity for Rous sarcoma virus transformed chicken embryo fibroblasts (which produce elevated levels of plasminogen activator) compared to nontransformed fibroblasts (which produce low levels of plasminogen activator). In the presence of the plasmin inhibitor, p-nitrophenyl p'-guanidinobenzoate at 2 micrograms/mL, the selectivity of the phenylenediamine mustard prodrug was reduced, but there was no effect on the cytotoxicity of the free drug. Furthermore, the prodrug analogue D-valylleucyl-D-lysylphenylenediamine mustard (in which L-Lys has been replaced by D-Lys) was inactive. Finally, the prodrug derivative of acivicin did not display selective toxicity for transformed cells when the cells were cultured in plasminogen-free medium. These results suggest that plasmin hydrolysis is necessary for the activation of the prodrugs. The prodrugs were tested in vivo for antitumor activity. The prodrug of acivicin, like acivicin itself, was inactive against the B16 melanoma, a murine tumor that produces high levels of plasminogen activator. This prodrug was active against the M5076 carcinoma, a tumor that displays only moderate levels of plasminogen activator; however, despite the fact that the prodrug was 2- to 3-fold less toxic on a molar basis than acivicin, there was no evidence of an increased therapeutic index. The prodrug of phenylenediamine mustard was also slightly less toxic than the parent drug, but again there was no evidence for an improved therapeutic index against the B16 tumor.
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PMID:Plasmin-activated prodrugs for cancer chemotherapy. 1. Synthesis and biological activity of peptidylacivicin and peptidylphenylenediamine mustard. 622 Oct 99


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