Nodular basal cell carcinoma is associated with increased hyaluronan homeostasis.

BACKGROUND: Basal cell carcinoma (BCC) is one of the most frequent forms of malignancy in humans. Although BCC is a tumour of low degree of malignancy, if left untreated, it can be locally aggressive, eat away at tissues and cause ulceration. Nodular is the most common subtype of BCC (>50%). Although apparently non-invasive, micronodular, a certain subgroup of nodular, is likely to recur. Glycosaminoglycans (GAGs), such as hyaluronic acid (HA), are extracellular matrix molecules of high importance in malignant transformation, metastasis and other complex remodelling processes. OBJECTIVES: To investigate the expression of GAGs and their metabolizing enzymes in nodular BCC, when compared with adjacent healthy human skin tissue specimens. METHODS: Total GAGs were isolated and purified from nodular BCC and normal adjacent human skin tissue specimens. GAGs were subsequently fractionated by electrophoresis on cellulose acetate membranes and characterized using specific GAG-degrading enzymes. The content of HA in total GAGs was measured using ELISA and the expression of HA synthases (HAS), hyaluronidases (HYAL) and HA receptors (CD44 and receptor hyaluronic acid-mediated motility (RHAMM) was assessed using RT-PCR. RESULTS: Nodular BCC is associated with increased levels of HA concomitant with upregulation of gene expression of HAS3, HYAL3 and RHAMM, when compared with normal adjacent skin. CONCLUSION: These results indicate that HA homeostasis in nodular BCC shows distinct features which may be helpful in understanding the complex behaviour of nodular subtype of BCC, thus eventually leading to new treatment strategies.

Adherence to antiretroviral therapy among HIV seropositive patients in northern Greece. Major factors of influence.

BACKGROUND: Adherence to HIV antiretroviral therapy (ART) is of great importance for reducing viral load and the eventual treatment of the patients and minimizing infectivity. This study aimed to investigate adherence to ART among people living with HIV in northern Greece and investigate the factors influencing adherence to ART. METHODS: A correlational study was performed on a cohort of 112 seropositive individuals (100 men and 12 women) with a mean age of 37.14 years. The simplified medication adherence questionnaire (SMAQ) was used to assess adherence. In addition, the perceived available support questionnaire (PAS) for social support assessment was utilized to evaluate the perceived social support. RESULTS: Approximately 60% of patients were found to be nonadherent to ART. Important factors affecting adherence are educational level, social support, and use of substances. CONCLUSIONS: The results show that a significant proportion of the cohort of patients investigated from northern Greece does not show adherence to ART. Several factors were identified to be of significant influence, which should be taken into consideration by the Greek healthcare providers. HIPPOKRATIA 2020, 24(3): 114-119.

Decreased hyaluronan in airway smooth muscle cells from patients with asthma and COPD.

Glycosaminoglycans (GAG) are essential extracellular matrix molecules which regulate tissue flexibility, a parameter that is reduced in airways of patients with asthma and chronic obstructive pulmonary disease (COPD). We investigated the expression of GAG and their metabolising enzymes in primary human airway smooth muscle cells (ASMC) obtained from healthy donors (controls) and patients with asthma or COPD. Total GAG synthesis was assessed by [(3)H]-glucosamine incorporation. GAG were isolated, purified, fractionated by electrophoresis and characterised using specific GAG-degrading enzymes. Secretion of hyaluronic acid (HA) by ASMC from patients with asthma or COPD was significantly decreased compared with controls. RT-PCR analysis and western blotting revealed that this decrease was associated with a significant reduction in the expression of HA synthase-1 and -2 and a significant increase of hyaluronidase-1. Furthermore, the expression of the HA receptor CD44 was significantly decreased, whereas the receptor for HA-mediated motility was not expressed in asthma or COPD. Our results indicate that there is a decreased expression of HA in asthma and COPD associated with a synergistic regulation of HA metabolising enzymes that may regulate the pathological airway remodelling in these lung diseases.

Extrinsic ageing in the human skin is associated with alterations in the expression of hyaluronic acid and its metabolizing enzymes.

Extrinsic skin ageing or 'photoageing', as opposed to intrinsic skin ageing, is the result of exposure to external factors, mainly ultraviolet irradiation. Glycosaminoglycans (GAG) and particularly hyaluronic acid (HA) are major components of the cutaneous extracellular matrix involved in tissue repair. However, their involvement in extrinsic skin ageing remains elusive. In this study, we investigated the expression of HA and its metabolizing enzymes in photoexposed and photoprotected human skin tissue specimens, obtained from the same patient. Total GAG were isolated, characterized using specific GAG-degrading enzymes and separated by electrophoresis on cellulose acetate membranes and polyacrylamide gels. Quantitation of HA in total GAG was performed using ELISA. Gene expression of hyaluronan synthases (HAS), hyaluronidases (HYAL) and HA receptors CD44 and receptor for HA-mediated motility (RHAMM) was assessed by RT-PCR. We detected a significant increase in the expression of HA, of lower molecular mass, in photoexposed skin as compared with photoprotected skin. This increase was associated with a significant decrease in the expression of HAS1 and an increase in the expression of HYAL1-3. Furthermore, the expression of HA receptors CD44 and RHAMM was significantly downregulated in photoexposed as compared with photoprotected skin. These findings indicate that extrinsic skin ageing is characterized by distinct homoeostasis of HA. The elucidation of the role of HA homoeostasis in extrinsic skin ageing may offer an additional approach in handling cutaneous ageing.

Effects of low-intensity shock wave therapy (LiST) on the erectile tissue of naturally aged rats.

Low-intensity shock wave therapy (LiST) improves erectile function in patients with erectile dysfunction (ED), probably by promoting angiogenesis as suggested by studies on animals with comorbidities as disease associated ED models. We aim to investigate the effects of LiST on erectile tissue of healthy, naturally aged rats. Twelve naturally aged male rats were randomized into two groups: control group (n = 6) and LiST-treatment group (n = 6). Young rats (8 weeks) (n = 6) was also used as control. Each rat in treatment group received 300 shock waves with an energy flux density of 0.09 mJ/mm2 at 2 Hz. Sessions were repeated three times/week for 2 weeks, followed by a 2-week washout period. Real-time RT-PCR for the expressions of vascular endothelial growth factor (VEGF), endothelial nitric oxide synthase (eNOS), nerve growth factor (NGF), neuronal NOS (nNOS), as well as α1 and α2-adrenergic receptors (α1AR, α2AR) was performed, followed by immunohistochemical analysis (IHC) to evaluate protein expression. The expressions of VEGF, eNOS, and α2AR/α1AR ratio were increased after LiST (p = 0.039, p = 0.008, and p = 0.006 respectively). The increase of VEGF, eNOS, and α2AR was confirmed in IHC (p = 0.013, p = 0.092, and p = 0.096, respectively). The increase of VEGF and eNOS seem to play key role in the mechanism of action of LiST, apparently by inducing angiogenesis. The altered expression of α1/α2-adrenergic receptors could indicate a decrease in sympathetic activity. LiST showed to partially reverse changes associated with aging in erectile tissue of rats, which supports future research for ED prevention.

Increased hyaluronic acid content in idiopathic pulmonary arterial hypertension.

Idiopathic pulmonary arterial hypertension (IPAH) is a fatal disease characterised by elevated blood pressure in the pulmonary circulation. Initial vasoconstriction, proliferation of pulmonary arterial smooth muscle cells (PASMC) and increased deposition of extracellular matrix (ECM) contribute to pathological remodelling of pulmonary arterioles in IPAH. Glycosaminoglycans (GAGs), components of the ECM, control cellular proliferation and differentiation, but their expression in IPAH remains elusive. In the present study, GAG expression was investigated in the lungs of patients with IPAH or control transplant donors, and expression and localisation of GAG-metabolising enzymes were analysed in vivo and in vitro. A significant increase in the expression of hyaluronic acid (HA) was detected in IPAH lungs, associated with increased hyaluronan synthase (Has)1 and decreased hyaluronoglucosaminidase 1 gene expression, as assessed by quantitative RT-PCR and Western blotting. HAS1 protein localised to PASMC in vivo and increased HA deposition was observed in remodelled pulmonary arteries in IPAH. Transforming growth factor-beta1, a profibrotic growth factor, led to increased HA secretion and HAS1 expression in primary PASMC. The results demonstrate an increased hyaluronic acid content in idiopathic pulmonary arterial hypertension lungs, associated with increased hyaluronan synthase 1 and decreased hyaluronoglucosaminidase 1 gene expression. Synergistic regulation of glycosaminoglycan-metabolising enzymes in favour of accumulation may, thus, regulate pathological vascular remodelling in idiopathic pulmonary arterial hypertension lungs.

Basement membrane collagen-degrading activity from a malignant tumor is inhibited by anthracycline antibiotics.

Type IV collagenase activity was previously identified and purified to 7500-fold in homogenates from murine Walker 256 carcinoma, using acetylated [3H]-type IV collagen as a substrate. Anthracycline antibiotics daunorubicin, doxorubicin and epirubicin exhibited a non-competitive, reversible inhibition with Ki 92, 49 and 40 microM, respectively. This inhibitory effect, at therapeutically attainable concentrations of the forementioned antineoplastic drugs, may contribute, at least in part, to their antimetastatic properties. The anthracycline derivatives: 4-demethoxydaunorubicin, 4'-iododoxorubicin and 4-demethoxy-3'-deamino-3'-hydroxyepirubicin were without inhibitory effects at comparable concentrations. Other antineoplastic agents, such as belomycin, carmustine, cisplatine, etoposide, methotrexate, mitotane and teniposide did not exhibit any inhibitory effect at concentrations up to 1.0 mM.

Degradation of intact basement membranes by human and murine tumor enzymes.

Homogenates from malignant tumors, obtained from surgery specimens or from transplants of Walker 256 carcinosarcoma in rats, contained an enzyme activity capable of degrading intact 3H-acetylated basement membranes from bovine lens. The enzyme activity from murine tumor was purified about 7500-fold by (NH4)2SO4 fractionation, ion exchange and gel chromatography. The apparent molecular weight of the purified enzyme was approximately 50,000. The rate of degradation of 3H-labelled basement membrane by the murine tumor enzyme was reduced by addition of excess type IV collagen, but not of excess type I, type III or type V collagen. These results suggested specificity of this enzyme for type IV collagen. Inhibitors of serine proteinases, thiol proteinases and soybean trypsin inhibitor were without effect on the enzyme activity. Chelators such as 1,10-phenanthroline or EDTA reduced the activity to control levels, indicating that the enzyme activity was due to a metalloproteinase. Chromatographic and electrophoretic separation of the enzymatic products from 3H-labelled basement membrane and type IV collagen indicated that the enzyme activity was due to a type IV collagenase. The use of basement membrane in the native physiological state as a substrate for the study of basement membrane-degrading activity by homogenates of solid malignant tumors offers an in vitro model for the investigation of the metastatic potential of these tumors.

Tissue structure-specific distribution of glycosaminoglycans in the human penis.

The aim of this work was to isolate and characterise the glycosaminoglycans present in the different tissue structures of the human penis in view of their potentially significant role in the physiology of erection. Penile tissue samples were obtained from patients who underwent penectomy and were subsequently dissected into individual tissue structures. Total glycosaminoglycans were isolated and purified from tunica albuginea, corpora cavernosa and corpus spongiosum, following tissue mincing, ultrasonication, lipid extraction, extensive digestion with pronase and DNase, treatment with alkali-borohydride and ethanol precipitation. Isolated glycosaminoglycans were separated by cellulose acetate electrophoresis and fractionated by anion exchange chromatography on DEAE Sephacel columns. Different glycosaminoglycan fractions were identified using glycosaminoglycan-degrading enzymes of known specificity. Gradient polyacrylamide gel electrophoresis was used to determine the average molecular mass of the glycosaminoglycans. The corpus cavernosum and the corpus spongiosum extracts contained almost twice the amount of glycosaminoglycan-associated uronic acids as compared to the tunical extracts (1.47+/-0.09, and 1.49+/-0.15 as opposed to 0.75+/-0.15 microg/mg dry defatted tissue, respectively; S.E.M., n=5). With the exception of hyaluronic acid, the relative amount of individual glycosaminoglycan types varied significantly among extracts of different origin. Heparan sulphate was more abundant in cavernosal, dermatan sulphate in tunical, and chondroitin-6-sulphate in corpus spongiosum extracts. No structure-specific differences were detected with respect to the molecular mass distribution of each glycosaminoglycan type. Our study shows that the different structures of the human penis produce distinct profiles of glycosaminoglycans, which are well suited to the individual functional characteristics of these structures.

The differential distribution of hyaluronic acid in the layers of human atheromatic aortas is associated with vascular smooth muscle cell proliferation and migration.

Vascular smooth muscle cells (VSMC), under conditions of induced proliferation, similar to those involved in atherosclerosis, secrete an acidic glycan, 82% of which exhibits structural homology with hyaluronic acid (HA), has a molecular mass of 340 kDa (HA-340) and inhibits VSMC proliferation in vitro. In this study, the expression of glycans was investigated in human atheromatic aortas and evidence is presented that a HA molecule, similar to HA-340, is distinctly expressed in all aortic layers. The isolation of the glycans from human aortas was performed after homogenization of the individual aortic layers (atheromatic plaque, tunica intima, tunica media and tunica adventitia), by lipid extraction and extensive digestion with pronase and DNase. The total glycans were purified from the digestion products by gel filtration on Sephadex G-25 and fractionated on a Superose 6 column. Enzymatic treatment of the ensuing glycan fractions with all known glycosaminoglycan-degrading enzymes, followed by electrophoresis on polyacrylamide gradient gels and cellulose acetate membranes, revealed that, in addition to HA, the tunica intima and the atheromatic plaque also contained dermatan sulfate, while the tunica media and the tunica adventitia also contained chondroitin sulfates and heparan sulfate. The highest concentration of the human aorta HA was found in the tunica media, exhibiting a negative concentration gradient from the tunica media to the atheromatic plaque. Investigation of the biological function of the human aorta HA revealed that this molecule acts as a negative regulator on the PDGF-induced VSMC proliferation and as a positive regulator on the PDGF-induced VSMC migration. The differential expression of HA within the aortic layers correlates with the biological function attributed to this acidic glycan and associates it with key events in the progression of atherogenesis.

Inhibition of angiogenesis, tumour growth and metastasis by the NO-releasing vasodilators, isosorbide mononitrate and dinitrate.

1. The effect of the nitric oxide (NO)-producing nitrovasodilators isosorbide mononitrate (ISMN) and isosorbide dinitrate (ISDN) were assessed on (a) the in vivo model of angiogenesis of the chick chorioallantoic membrane (CAM) and (b) on the growth and metastatic properties of the Lewis Lung carcinoma (LLC) in mice. 2. Isosorbide 5-mononitrate (ISMN) and isosorbide dinitrate (ISDN), inhibited angiogenesis in the CAM dose-dependently. ISMN was more potent in inhibiting this process. Both compounds were capable of completely reversing the angiogenic effect of alpha-thrombin. These effects of ISMN and ISDN on angiogenesis were comparable to those previously observed with sodium nitroprusside which generates NO non-enzymatically. 3. Mice, implanted intramuscularly with LLC, received daily i.p. injections of ISMN for 14 days resulting in a significant decrease in the size of the primary tumour and a reduction in the number and size of metastatic foci in the lungs. ISDN had a similar but less pronounced effect than that observed with ISMN. 4. Addition of ISMN or ISDN to cultures of bovine, rabbit and human endothelial cells and to cultures of LLC cells had no effect on their growth characteristics. 5. These results indicate that ISMN and ISDN inhibit angiogenesis and tumor growth and metastasis in an animal tumour model. The possibility should therefore be considered that these nitrovasodilators which are widely used therapeutically and have well characterized pharmacological profiles, may also possess antitumour properties in the clinic.

Androgen-induced sexual dimorphism in high affinity dopamine binding in the brain transcends the hypothalamic-limbic region.

1 High affinity binding of [3H]-dopamine and [3H]-5-hydroxytryptamine ([3H]-5-HT) was measured in membrane fractions prepared from cerebral cortex, amygdala, hypothalamus, thalamus and brain stem of rats of either sex and of rats which had been either neonatally castrated or androgenized. 2 Binding was measured in rats of 8, 20 and 30 days old as well as in adults. 3 [3H]-dopamine bound with approximately 30 nM affinity ahd [3H]-5-HT with approximately 10 nM affinity to all areas of the brain tested. The relative inhibitory effects of haloperidol, apomorphine, cis-flupenthixol, unlabelled dopamine, noradrenaline, spiroperone, (+)-butaclamol, fluphenazine, pimozide and 5-HT on [3H]-dopamine binding in the cerebral cortex was consistent with receptor status for the binding components there as were the relative inhibitory effects of methysergide, dopamine, fluoxetine and ouabain on [3H]-5-HT binding in the fore brain. 4 Neither [3H]-dopamine nor [3H]-5-HT binding varied with the state of the sexual cycle in females. 5 There were no sexual differences in [3H]-5-HT binding in any of the brain areas tested nor was it affected by neonatal androgenization or neonatal castration. 6 [3H]-dopamine binding was greater in the cerebral cortex and amygdala of male than of female rats. These differences could be mimicked artificially by neonatal castration of males (female type development) or neonatal androgenization of females (male type development). Sexual dimorphism did not become overt until 20 days of age and did not extend to hypothalamus, thalamus or brain stem. 7 It is concluded that neonatal sex differences in exposure to steroid hormones has permanent effects on the number of dopamine binding sites in the cerebral cortex and is suggested that this sexual dimorphism extends to the amygdala.

Development of sexual dimorphism in the suprachiasmatic nucleus of the rat.

Synapses develop at similar rates in the suprachiasmatic nucleus of rats of both sexes, but values are higher in male than in female animals from birth to maturity. Male-type development cannot be mimicked by neonatal androgenization but results suggest that female-type development can be induced by neonatal castration of males. The results suggested that both prenatal and postnatal androgens are essential to normal male development.

Suppression of angiogenesis by the antitumor agent titanocene dichloride.

Titanocene dichloride, which is an active antitumor agent against solid but not blood-borne tumors, suppresses angiogenesis and inhibits biosynthesis of collagenous proteins in the in vivo system of the chorioallantoic membrane of the chick embryo. The agent does not affect total protein biosynthesis in the same system. At non-toxic dose regimens titanocene dichloride retards the growth of Walker 256 carcinosarcoma transplants in rats and reduces the number of seeded implants in the mesenteric bed. At concentrations which suppress angiogenesis and inhibit biosynthesis of collagenous proteins, the agent does not affect the viability of Walker 256 carcinosarcoma cells, or the attachment and proliferation of human A549 lung adenocarcinoma or human umbilical vein endothelial cells in culture. It appears that the antitumor activity of titanocene dichloride may be attributed, at least in part, to its ability to suppress angiogenesis.

Corticosteroids inhibit the expression of the vascular endothelial growth factor gene in human vascular smooth muscle cells.

The vascular endothelial growth factor (VEGF) is a specific mitogen for vascular endothelial cells and enhances vascular permeability and edemagenesis. VEGF is also a major regulator of angiogenesis and may be a key target for inhibiting angiogenesis in angiogenesis-associated diseases. Among the extensively studied angiostatic compounds are several corticosteroids when used alone or in combination with heparin. In this study we present evidence for an additional mechanism of action of hydrocortisone, cortisone and dexamethasone in inhibiting edemagenesis or angiogenesis. In cultures of aortic human vascular smooth muscle cells these corticosteroids (1 x 10(-8) to 1 x 10(-12) M) abolished the platelet-derived growth factor-induced (PDGF) expression of the VEGF gene in a dose-dependent manner. In contrast, two precursors of corticosteroids, desoxycorticosterone or pregnenolone, did not affect PDGF-induced VEGF expression. Our findings indicate that the capacity of corticosteroids to reduce edema or to prevent new blood vessel formation may be attributed, at least in part to the ability of these agents to abolish the expression of VEGF.

Expression of proteoglycans and glycosaminoglycans in angiofibroma and fibrous plaque skin lesions from patients with tuberous sclerosis.

Tuberous sclerosis complex (TSC) is a disorder of cell lineage, migration, proliferation and differentiation, characterized by the development of widespread benign hamartomas, which are particularly evident in hamartomatous lesions of the skin. The aim of this study was to investigate differences in gene expression of certain proteoglycans (PGs) and to characterize glycosaminoglycans (GAGs) in tissue specimens of normal skin, fibrous plaques and angiofibromas from patients with TSC. The expression of PG mRNA was determined by semiquantitative RT-PCR analysis. Total GAGs were isolated from tissue specimens after lipid extraction and extensive digestion with Pronase and DNase and characterized by treatment with GAG-degrading enzymes followed by electrophoresis on polyacrylamide gradient gels and cellulose acetate membranes. Normal skin specimens express versican, decorin and aggrecan and contain hyaluronic acid and dermatan sulphate. In angiofibroma specimens aggrecan is not expressed while versican splice variant with two EGF-like domains and decorin are downregulated. Furthermore, angiofibromas differ from normal skin in that they additionally contain keratan, heparan and chondroitin sulphates and do not contain dermatan sulphate. In fibrous plaque specimens gene expression of PGs was similar to that in normal skin, but with respect to GAGs, they contained a single acidic glycan population that did not share common structural features with known GAGs. The variations of the above ECM molecules between normal and TSC skin may be attributed to TSC-related mutations and, overall, support the TSC-associated pathological manifestations of cell migration, proliferation and differentiation.

Isolation and characterization of glycosaminoglycans from human atheromatous vessels.

The complex cascade of events leading to the formation of atheromatous plaques depends on the interaction between several cell types, growth factors, cytokines, and molecules of the extracellular matrix (ECM) (1). Among different molecules of the ECM involved in atherogenesis, the glycosaminoglycan(GAGs) have been reported to contribute to key events leading to the formation of atherosclerotic lesions (2). GAGs are linear acidic polysaccharides of variable length and composition, which occur either in free form or attached to a protein core to form proteoglycans (3). On the basis of their composition, GAGs are grouped into four major categories: hyaluronic acid, heparin and heparan sulfate, chondroitin and dermatan sulfates, and keratan sulfate. ECM GAGs provide structural links between fibrous and cellular elements, contribute to viscoelastic properties, regulate permeability and retention of plasma components within the matrix (2,4), inhibit vascular cell growth (5), affect hemostasis and platelet aggregation (6), and interact with lipoproteins (7) and various growth factors (8,9).

Assay for cell migration and invasion of vascular smooth muscle cells.

Chemotaxis is a critical event in the development of atherosclerotic lesions and in the restenosis that often occurs after surgical intervention and angioplasty (1). Chemokines involved in atherogenesis include colony-stimulating factors (2), oxidized low-density lipoproteins (3), transforming growth factor-ß (4) and fibroblast growth factor (5), which induce chemotaxis of monocytes and endothelial cells. Other growth factors, such as platelet-derived growth factor (PDGF) (6) and insulin-like growth factor-1, induce chemotaxis of vascular smooth muscle cells (VSMCs) (7), which play a major role in the formation of atherosclerotic lesions (8,9). In normal human arteries, VSMC-reside mainly in the tunica media in a quiescent state and express a variety of differentiation-specific genes important to maintain the physiological regulation of vessel tone and blood pressure (10). Under pathological conditions, such as vessel injury or atherosclerotic plaque development, VSMCs become exposed to certain growth factors and cytokines, such as PDGF, which induce a transformation from the contractile to a synthetic state (7,11-13). Only in the latter state, VSMCs acquire the ability to migrate from the tunica media to the tunica intima. In vivo, VSMCs are surrounded by and embedded in a variety of extracellular matrices (ECMs) that must be traversed during migration. In the intact vessel, one of the main ECM barriers to cell movement is the basement membrane (BM) that surrounds each VSMC and separates the VSMC-containing medial cell layer from the endothelium (14). Migrating VSMCs have been shown to digest BM (14,15), an invasion process that is mediated by tightly regulated proteases (16). Within this group of proteases, the family of matrix metalloproteinases (MMPs) are essential for the digestion of ECM components such as collagens, gelatins, or proteoglycans (17,18).

Angiogenesis is associated with collagenous protein synthesis and degradation in the chick chorioallantoic membrane.

Extracts of the chick embryo chorioallantoic membrane (CAM) obtained from 7-20 day old embryos, contained enzyme activity that could degrade type IV collagen. Peak enzyme activity was observed on days 8-10 of embryogenesis, which coincides with the stage of maximum angiogenesis. This activity decreased to lowest values at days 13-15 and increased thereafter up to day 20. Maximum rate of collagen biosynthesis in CAM was observed between days 7 and 10, with a drastic decrease at day 12, when vascular density has reached a plateau. The type IV collagen-degrading activity of CAM was of the metalloprotease type, since it was inhibited by 1,10-phenanthroline and EDTA but was also partially inhibited by serine and thiol protease inhibitors.

Mode of action of razoxane: inhibition of basement membrane collagen-degradation by a malignant tumor enzyme.

Razoxane is an antitumor agent structurally related to EDTA. Its cytotoxic mechanism of action has been shown to involve interference with cell division and reduction of the gross rate of DNA synthesis. In this study we present evidence that razoxane, at therapeutically attainable concentrations, inhibits type IV collagen and basement membrane degradation by an enzyme isolated from malignant tumors. This effect is attributed to the chelating properties of razoxane and may explain the antimetastatic properties, by means of stabilization of tumor blood vessels, which have been associated with the action of the antitumor drug.