Hannele Tapiovaara

Production of an active urokinase by leukemia cells: a novel distinction from cell lines of solid tumors

A new screening test is described which enabled rapid determination of the proportion of single-chain and two-chain urokinase produced in the culture supernatants of 18 human cell lines. A clear distinction was found between two groups of cell lines: cells derived from ten solid tumors produced almost exclusively single-chain proenzyme, while the majority of the enzyme found in cultures of eight leukemia cell lines was in the active, two-chain form.

REGULATION OF THE PERICELLULAR ACTIVATION OF PLASMINOGEN AND ITS ROLE IN TISSUE‐DESTRUCTIVE PROCESSES

While it is generally recognized today that cells use specific adhesion proteins such as fibronectin and laminin, both in cell adhesion and migration, it is equally clear that pericelldar proteolysis is critically involved in the invasion of malignant and certain normal cells through the extracellular matrices, both interstitial connective tissue matrices and basement membranes. There is interesting new evidence suggesting that a key component in pericellular proteolysis, i.e. the urokinase-receptor complex, may have a role also in both cell adhesion and cell migration. In addition there is direct evidence that adhesion proteins of pericellular matrix can provide a molecular link between the membrane receptor proteins, known as integrins, and plasminogen.

PLASMINOGEN ACTIVATION ON TUMOR CELL SURFACE AND ITS INVOLVEMENT IN HUMAN LEUKEMIA

Publisher Summary This chapter focuses on the plasminogen activation on tumor cell surface and its involvement in human leukemia. The function of plasminogen activators (PAS) and plasmin in the adult organism has been mainly related to fibrinolysis, tissue remodeling, cell migration, and tissue destruction. Associated with plasmin activity is the dissolution of blood clots, inflammation, tumor cell dissemination, wound healing, angiogenesis, and trophoblast invasion. In addition to directly cleaving components of the extracellular matrix, plasmin is able to regulate various functions through the activation of prohormones and latent growth factors. The invasive capacity of tumor cells requires proteolytic activity; therefore, plasmin, together with other tissue-destructive enzymes, has been implicated in the property of tumor cells to metastasize. Both plasminogen and PAS bind to cell surfaces, and this allows plasmin generation at restricted areas of the cell surface and the cell-matrix interface. High levels of urokinase plasminogen activator (uPA) and/or its inhibitor plasminogen activator inhibitor type 1, in tumor tissue seem to indicate an adverse prognosis for patients with cancer, as was first shown in the case of breast cancer. The components of plasminogen activation described in the chapter include plasminogen, urokinase-type plasminogen activator, and tissue-type plasminogen activator. Plasminogen activation is regulated both by inhibitors of PAS and by inhibitors of plasmin. Plasminogen activator inhibitors (PAIs) are specific, fast-acting inhibitors of PAS. Four inhibitors are known, PAI-1, PAI-2, PAI-3, and PAI-4 (protease nexin). The α 2 MR, which is identical to the low-density lipoprotein receptor related protein (LRP), has been reported to mediate the internalization and degradation of tPA, tPA-PAI-1 complexes, uPA-PAL1 complexes, and uPA-protease nexin complexes. Although plasminogen activation is needed in the process of matrix destruction, it is still unknown whether the activity of uPA or merely the uPA antigen and/or uPAR is involved in the process of cell migration.

Interferons and retinoids enhance and dexamethasone suppresses urokinase-mediated plasminogen activation in promyelocytic leukemia cells

All-trans retinoic acid (RA) has been successfully used in the treatment of patients with acute promyelocytic leukemia (APL). It induces differentiation of APL cells and reduces the bleeding tendency in APL patients. It has been proposed that plasminogen activation could affect the fibrinolytic balance in patients with leukemia. In our earlier study we found that treatment of APL cells with RA results in changes in urokinase (uPA) production. As interferons (IFNs) and dexamethasone can be used together with RA in the treatment of patients with APL, we have now studied the effects of RA together with IFNs and dexamethasone on the plasminogen activation cascade of these cells, including measurement of plasmin generation and uPA receptor (uPAR), using enzyme immunoassays, fluorescence-activated cell sorter analysis and RNA extraction with Northern blotting. Our main results were: (1) plasmin was formed on the surface of APL cells; (2) RA stimulated transiently plasmin generation and increased uPAR mRNA level; (3) IFNs α and γ potentiated RA in its effects on uPA and plasmin activities and on uPAR level; (4) dexamethasone suppressed totally the effect of RA on uPA induction and plasminogen activation; and (5) IFNs and dexamethasone alone did not have potent effects on plasminogen activation. These results may assist in the design of therapy for APL patients.

Cell Surface Plasminogen Activation

The assumption that the plasminogen activation system, through a breakdown of extracellular matrix proteins, plays a role in invasiveness and destruction of normal tissue during growth of malignant tumors is supported by a variety of findings. These include a close correlation between transformation of cells with oncogenic viruses and synthesis of urokinase-type plasminogen activator (u-PA), the finding that u-PA is involved in tissue destruction in many nonmalig-nant conditions, and the immunohistochemical localization of u-PA in invading areas of tumors (for reviews, see Dan0 et al. 1985; Saksela 1985). Further support for this hypothesis has come from studies with anticatalytic antibodies to u-PA in model systems for invasion and metastasis. Such antibodies were found to decrease metastasis to the lung from a human u-PA-producing tumor, HEp-3, transplanted onto the chorioallantoic membrane of chicken embryos (Ossowski and Reich 1983; Ossowski 1988), penetration of amniotic membranes by B16 melanoma cells (Mignatti et al. 1986), basement membrane invasion by several human and murine cell lines of neoplastic origin (Reich et al. 1988), and formation of lung metastasis after intravenous injection of B16 melanoma cells in mice (Hearing et al. 1988). In some of these studies (Mignatti et al. 1986; Reich et al. 1988), a plasmin-catalyzed activation of procollagenases (see Tryggvason et al. 1987) appeared to be a crucial part of the effect of plasminogen activation.

α2‐Macroglobulin in the Regulation of Pericellular Plasminogen Activation of Human Tumor Cells

Increased activation of plasminogen is a close correlate of the malignant invasive cellular phenotype and is also involved in various tissue-destructive processes. Of the several extracellular proteolytic pathways, it is the plasminogen activation cascade, along with basement membrane collagenases, that represents, quantitatively at least, the principal enzyme system of matrix degradation in both malignant invasion and other tissue-destructive processes. In the following, we first summarize the mechanisms of tumor cell-associated plasminogen activation and then consider the regulatory role of alpha-2-macroglobulin ( a 2 M ) in these processes.