Comparative pharmacokinetics of rVIII-SingleChain and octocog alfa (Advate(®) ) in patients with severe haemophilia A.

BACKGROUND: rVIII-SingleChain, a novel recombinant factor VIII (rFVIII), has been designed as a B-domain truncated construct with covalently bonded heavy and light chains, aiming to increase binding affinity to von Willebrand factor (VWF). Preclinical studies confirmed greater affinity for VWF, giving improved pharmacokinetic and pharmacodynamic properties compared with full-length rFVIII. AIM: To investigate the pharmacokinetics of rVIII-SingleChain and compare them against those of full-length rFVIII. METHODS: This study enrolled 27 patients with severe haemophilia A in the AFFINITY clinical trial programme. After a 4-day washout period, all patients received a single infusion of 50 IU kg(-1) octocog alfa (Advate(®) ); after a ≥4-day postinfusion washout period, they received a single infusion of 50 IU kg(-1) rVIII-SingleChain. Blood samples for pharmacokinetic assessments of each product were collected before infusion (predose) and at 0.5, 1, 4, 8, 10, 24, 32, 48 and 72 h postinfusion for both products. RESULTS: rVIII-SingleChain had a longer mean half-life (t1/2 ) (14.5 vs. 13.3 h), lower mean clearance (CL) (2.64 vs. 3.68 mL h(-1) kg(-1) ), higher mean residence time (20.4 vs. 17.1 h) and larger mean AUCinf (2090 vs. 1550 IU?h dL(-1) ) than octocog alfa, respectively. The mean AUCinf after rVIII-SingleChain infusion was ~35% larger than after octocog alfa. A similar pattern was observed for AUC0-last . No serious adverse events or inhibitors were reported. CONCLUSIONS: rVIII-SingleChain has a favourable pharmacokinetic profile compared with octocog alfa and was well tolerated. The prolonged t1/2 , larger AUC and reduced CL of rVIII-SingleChain may permit longer dosing intervals, thereby improving patient adherence to prophylactic treatment.

Longer-acting clotting factor concentrates for hemophilia.

Hemophilia, when severe, leads to spontaneous life-threatening bleeding episodes. Current therapy requires frequent intravenous infusions. Most patients must limit their physical activities to avoid bleeding when the factor activity levels are below normal. In 2014, new therapeutic factor VIII and IX products were approved in Canada and the U.S. Over the next couple of years, other new factor products will likely be approved. These new factors have been engineered to have improved pharmacokinetic properties, including extended half-life in circulation, thus providing major therapeutic advances for patients with hemophilia. In the completed clinical trials, over 700 patients have successfully used these longer acting products regularly for more than one year. These promising new therapies should allow patients with hemophilia to use fewer infusions to prevent spontaneous bleeding or to treat bleeding episodes, and to provide appropriate clotting factor levels for different physical activities.

Safety and pharmacokinetics of a recombinant factor VIII with pegylated liposomes in severe hemophilia A.

BACKGROUND: BAY 79-4980 is a sucrose-formulated recombinant factor VIII (rFVIII-FS) combined with pegylated liposomes to prolong activity. OBJECTIVES: To investigate the safety, tolerability, bioavailability, pharmacokinetics and pharmacodynamics of a single administration of BAY 79-4980 compared with standard rFVIII-FS in patients with severe hemophilia A. METHODS: This randomized, double-blind study consisted of two crossover substudies comparing two doses of liposomal rFVIII-FS with standard rFVIII-FS. Males (12-60 years) with severe hemophilia A received a single infusion of standard rFVIII-FS (35 IU kg(-1)) followed by a single infusion of BAY 79-4980 (13 or 22 mg kg(-1) pegylated liposomes) or vice versa, with 12 observation days and a 2-day washout period between treatments. RESULTS: Twenty-six subjects were enrolled at two centers. No serious adverse events were reported. Transient increases in complement C3a, but not CH50, were seen in subjects receiving both the low- and high-liposome-dose BAY 79-4980. Mild transient elevations of total and low-density lipoprotein cholesterol were observed. There were no clinically significant differences in clotting or laboratory parameters or in pharmacokinetic behavior between BAY 79-4980 and standard rFVIII-FS. The number of subjects with spontaneous bleeds on days 1-14 postinfusion was low, and group comparisons were inconclusive. CONCLUSIONS: Single-dose administration of BAY 79-4980 is well tolerated in patients with severe hemophilia A. Plasma pharmacokinetics of FVIII cannot explain the extended protection from bleeding observed previously with BAY 79-4980. Further studies of efficacy and long-term safety of chronic administration are planned.

Osteonectin/SPARC polymorphisms in Caucasian men with idiopathic osteoporosis.

UNLABELLED: Animal models suggest a role for osteonectin/SPARC in determination of bone mass. We found haplotypes consisting of three single nucleotide polymorphisms (SNPs) in the 3' untranslated region (UTR) of the osteonectin gene are associated with bone density in Caucasian men with idiopathic osteoporosis. INTRODUCTION: Osteonectin is a matricellular protein regulating matrix assembly, osteoblast differentiation, and survival. Animal studies indicate that osteonectin is essential for normal bone mass. The 3' UTR is a regulatory region controlling mRNA stability, trafficking, and translation, and we determined whether osteonectin 3' UTR haplotypes could be associated with bone mass and/or idiopathic osteoporosis. METHODS: Single strand conformation polymorphism and allele-specific PCR analysis were used to assess alleles at osteonectin cDNA bases 1046, 1599, and 1970, using genomic DNA from middle-aged Caucasian men with idiopathic, low turnover osteoporosis (n = 56) and matched controls (n = 59). Bone density was measured by DXA at spine, hip and radius. Allele and haplotype frequencies were analyzed by Chi square analysis and Fisher's exact test. RESULTS: Five common osteonectin 3' UTR haplotypes were identified. The frequency of one haplotype (1046C-1599C-1970T) was higher in controls compared with patients, and this haplotype was also associated with higher bone densities at multiple sites in patients. In contrast, a second haplotype (1046C-1599G-1970T) was associated with lower bone densities in patients at multiple sites. CONCLUSIONS: Osteonectin regulates skeletal remodeling and bone mass in animals, and haplotypes in the 3' UTR of this gene are associated with bone density in Caucasian men with idiopathic osteoporosis.

Liposomal approach towards the development of a longer-acting factor VIII.

Prevention of spontaneous bleeding in patients with severe haemophilia A usually requires therapeutic infusions every 2-3 days because of the short half-life of factor VIII (FVIII). Longer-acting FVIII products that require less frequent infusions would be beneficial and might obviate the need for central catheters in most patients. Liposomal formulation can enhance the efficacy of some therapeutic products. The incorporation of high-molecular weight polyethylene glycol (PEG) can extend the circulatory half-life of the liposome. These combined approaches led to the development of BAY 79-4,980, a PEG-containing liposomal version of Kogenate FS (rFVIII-FS). Results from preclinical models and early clinical trials have shown that BAY 79-4,980 prolongs the time to the next bleed. Further clinical evaluation of the efficacy and long-term safety of BAY 79-4,980 are planned.

Reformulated BeneFix: efficacy and safety in previously treated patients with moderately severe to severe haemophilia B.

BeneFix, the only recombinant factor IX (FIX), has been reformulated. The reformulation involves a change in diluent and allows for more concentrated infusions of recombinant FIX. A double-blind, randomized, pharmacokinetic (PK) crossover study demonstrated that reformulated BeneFix was bioequivalent to original BeneFix and follow-up PK evaluation after 6 months of treatment demonstrated the PK stability of reformulated BeneFix after multiple exposures. Favourable efficacy and safety profiles, consistent with those already well-established for original BeneFix, were observed: 81.1% of haemorrhages resolved with only a single infusion; 85.3% of initial treatment response ratings were Excellent or Good; more than half of the subjects using reformulated BeneFix for routine prophylaxis (11 of 17, 64.7%) had no spontaneous haemorrhages during their 6-12 month course of prophylactic treatment, with an overall spontaneous bleeding rate of 0.72 year(-1); and for the single surgical procedure (knee washing), treatment was rated Useful. In addition, there was no FIX inhibitor development, allergic-type manifestations, or thrombogenic complications with more than 1100 infusions (nearly 5.2 million IUs) administered in this trial. All efficacy and safety outcomes from this study were achieved with more concentrated recombinant protein infusions than that possible with original BeneFix, and utilization of the 2000 IU per vial dosage strength, newly introduced with the reformulated product, was high (>62%). The reformulation of BeneFix allows smaller delivery volumes and an increased choice of dosage strengths without altering the PK properties (including incremental recovery and half-life) or the established efficacy and safety profile of recombinant FIX.

Sucrose formulated recombinant human antihemophilic factor VIII is safe and efficacious for treatment of hemophilia A in home therapy--International Kogenate-FS Study Group.

To add an increased level of safety to antihemophilic factor replacement therapy, a full-length, recombinant Factor VIII (rFVIII) product has been developed without human-derived plasma proteins during purification and formulation and using an additional solvent/detergent viral inactivation step. This first clinical trial of a sucrose-formulated full-length rFVIII (rFVIII-FS) was conducted in previously treated patients (> or = 100 prior exposure days) with severe (<2% FVIII) hemophilia A in North America (NA) and Europe (EU). Pharmacokinetic profiles for rFVIII-FS were compared with those of currently licensed rFVIII product (Kogenate) in 35 patients. Safety and efficacy during home therapy were evaluated in 71 patients. The new formulation displayed a pharmacokinetic profile similar to that of rFVIII. Patients on home therapy received a cumulative total of 11,867 exposure days, 12,546 infusions, and 22,443,694 IU of rFVIII-FS. Of 2585 bleeds, 93.5% were treated with 1-2 infusions and 80.5% of responses were rated as excellent or good. No evidence of de novo inhibitor formation was observed. Only 0.27% of infusions were associated with any drug-related adverse event. Except for an episode of intermittent chest pain with palpitations which ceased after treatment with analgesics, associated adverse events were mild or moderate. Overall, rFVIII-FS provided excellent hemostatic control, was well-tolerated, and caused no significant adverse effects, thus demonstrating safety and efficacy for treatment of bleeds in patients with hemophilia A.

Outcome of radiotherapy for acromegaly using normalization of insulin-like growth factor I to define cure.

Radiation therapy (RT) has traditionally been considered a useful additional therapy for patients with acromegaly not achieving biochemical remission after surgery. However, recent evidence has suggested that RT is not curative in most patients with acromegaly when normalization of the serum insulin-like growth factor I (IGF-I) level is used to define remission. Therefore, we evaluated the success of RT based on IGF-I level in the 47 patients who received RT as part of their treatment from the cohort of 161 patients with acromegaly seen by us between 1981 and 1999. Four patients in whom no post-RT IGF-I level was available were excluded from the analysis. Of the remaining 43 patients, 32 patients received external beam RT, 6 received fractionated stereotactic radiosurgery, 4 received gamma-knife RT, and 1 received proton beam RT. The most recent IGF-I levels in these 43 patients, obtained a mean of 5.2 yr post-RT (range, 0.8-13.2 yr), were compared to age-adjusted normal ranges. IGF-I levels were normal in 17 patients (39.5%) without the addition of medical therapy. The percentage of patients with a normal IGF-I level generally increased with time post-RT; 27% of patients less than 6 yr post-RT, but 69.2% of patients 6 yr or more post-RT had normal IGF-I levels. Using the more traditional criterion for cure, a random GH measurement, 74% of patients had a GH level below 5 ng/mL, and 44% had a GH level below 2.5 ng/mL and would have been considered in remission based on these criteria. We conclude that with time RT remains a useful adjunctive treatment for many patients with acromegaly. RT should be considered along with appropriate medical therapy in selected patients who do not achieve normalization of IGF-I level after surgery or for those resistant to medical therapy.

Safety and efficacy of solvent/detergent-treated antihaemophilic factor with an added 80 degrees C terminal dry heat treatment in patients with haemophilia A.

Plasma-derived factor VIII concentrates remain an important resource for haemophilia A patients. To improve the safety of these preparations, various methods of viral removal and inactivation have been used that are designed to eliminate both enveloped and non-enveloped viruses. There have been rare reports that some viral inactivation processes altered the immunogenicity of some concentrates, leading to the development of factor VIII inhibitors in previously treated haemophilia A patients. This study evaluated the safety, efficacy and lack of neo-antigenicity of a highly purified factor VIII preparation which undergoes both solvent/detergent treatment and final dry heat treatment at 80 degrees C for 72 h. The study included: (i) a single blind, single-dose crossover pharmacokinetic study in 18 previously treated patients, comparing sibling lots of the unheated preparation (Koate(R)-HP) and the heat-treated preparation (Koate(R)-DVI), and (ii) an extended home treatment programme for 36 patients at two haemophilia treatment centres primarily to assess immunogenicity. Clinical parameters were assessed at regular intervals. The results confirm that Koate(R)-HP and Koate(R)-DVI are bioequivalent, and that Koate(R)-DVI is safe and efficacious for treatment of acute bleeding episodes and for surgery. Furthermore, the heat-treated preparation is not associated with the development of inhibitors in previously treated patients.

Treatment of acromegaly with the growth hormone-receptor antagonist pegvisomant.

BACKGROUND: Patients with acromegaly are currently treated with surgery, radiation therapy, and drugs to reduce hypersecretion of growth hormone, but the treatments may be ineffective and have adverse effects. Pegvisomant is a genetically engineered growth hormone-receptor antagonist that blocks the action of growth hormone. METHODS: We conducted a 12-week, randomized, double-blind study of three daily doses of pegvisomant (10 mg, 15 mg, and 20 mg) and placebo, given subcutaneously, in 112 patients with acromegaly. RESULTS: The mean (+/-SD) serum concentration of insulin-like growth factor I (IGF-I) decreased from base line by 4.0+/-16.8 percent in the placebo group, 26.7+/-27.9 percent in the group that received 10 mg of pegvisomant per day, 50.1+/-26.7 percent in the group that received 15 mg of pegvisomant per day, and 62.5+/-21.3 percent in the group that received 20 mg of pegvisomant per day (P<0.001 for the comparison of each pegvisomant group with placebo), and the concentrations became normal in 10 percent, 54 percent, 81 percent, and 89 percent of patients, respectively (P<0.001 for each comparison with placebo). Among patients treated with 15 mg or 20 mg of pegvisomant per day, there were significant decreases in ring size, soft-tissue swelling, the degree of excessive perspiration, and fatigue. The score fortotal symptoms and signs of acromegaly decreased significantly in all groups receiving pegvisomant (P< or =0.05). The incidence of adverse effects was similar in all groups. CONCLUSIONS: On the basis of these preliminary results, treatment of patients who have acromegaly with a growth hormone-receptor antagonist results in a reduction in serum IGF-I concentrations and in clinical improvement.

Evaluation of disease status with sensitive measures of growth hormone secretion in 60 postoperative patients with acromegaly.

Traditionally, suppression of GH measured by polyclonal RIA to less than 2.0 microg/L after oral glucose was accepted as evidence of remission after transsphenoidal surgery for acromegaly. Recently, with newer, more sensitive GH assays, a cut-off of less than 1.0 microg/L has been suggested. With the development of accurate insulin-like growth factor I (IGF-I) and IGF-binding protein-3 (IGFBP-3) assays, additional tools are now available for assessing postoperative GH secretion. There has, however, never been a systematic comparison of sensitive GH, IGF-I, and IGFBP-3 assays in defining disease status in a large cohort of postoperative patients with acromegaly. Therefore, we evaluated how the use of modern assays impacts on our assessment of disease activity in these patients. Sixty postoperative subjects with acromegaly and 25 age-matched healthy subjects were evaluated with nadir GH levels after 100 g oral glucose as well as baseline IGF-I and IGFBP-3 levels. GH was assayed by polyclonal RIA, sensitive immunoradiometric assay (IRMA), and highly sensitive enzyme-linked immunosorbent assay. The mean nadir GH determined by IRMA was 0.09 +/- 0.004 microg/L in the healthy subjects, with the upper limit of the normal nadir being 0.14 microg/L (mean + 2 SD). Subjects with acromegaly were divided into those with active disease (n = 22), defined by elevated IGF-I levels, and those in remission (n = 38), defined by normal IGF-I levels. GH determined by IRMA failed to suppress into the normal range defined by our healthy subjects in all patients with active disease; nadir GH determined by IRMA ranged from 0.33-5.0 microg/L in this group. In 50% of the active group, nadir GH levels determined by IRMA were less than 1.0 microg/L, a GH nadir previously considered normal by strict criteria. When nadir GH levels in the subjects with active disease were measured by polyclonal RIA, there was overlap with the range of RIA values in the healthy subjects. Thus, the IRMA was superior to the RIA in that the overlap between these two groups was eliminated. Subjects with acromegaly in remission included those with normal GH suppression (n = 23; mean nadir GH by IRMA, 0.10 +/- 0.006 microg/L) and others with abnormal GH suppression by IRMA (n = 15; mean nadir GH by IRMA, 0.35 +/- 0.07 microg/L). The latter group may have persistent GH dysregulation detected by the sensitive IRMA. GH levels measured by enzyme-linked immunosorbent assay confirmed the IRMA results. IGFBP-3 levels were significantly higher in subjects with active acromegaly (4940 +/- 301 microg/L) vs. those in healthy subjects (2887 +/- 153 microg/L; P < 0.0001) and those in the subjects in remission (2966 microg/L; P < 0.0001). IGFBP-3 levels correlated overall with IGF-I levels (r = 0.765; P < 0.0001), but IGFBP-3 levels were not predictive of disease status because 32% of the subjects with active acromegaly had normal IGFBP-3 levels. In addition, failure of GH to suppress adequately was not associated with a higher IGFBP-3 level among the subjects in remission. These data indicate that the IRMA is superior to the RIA in distinguishing between patients with active disease (defined by elevated IGF-I levels) and healthy subjects. We also show that GH levels after oral glucose measured with highly sensitive GH assays can be much lower in subjects with active disease than previously believed; values less than 1.0 microg/L may be found in up to 50% of patients. In addition, in 39% of patients in apparent remission with normal IGF-I levels, GH determined by highly sensitive assays fails to suppress normally; it remains to be determined whether these patients are at higher risk for recurrence of active disease.

Novel vascular sealing device for closure of percutaneous vascular access sites.

To investigate the hemostatic capabilities of a novel vascular sealing device consisting of a balloon catheter and procoagulant, vascular sheaths were placed percutaneously in the femoral arteries of dogs. The sealing device was evaluated using the balloon catheter alone in six femoral arteries and with the addition of a procoagulant, in 21 femoral arteries. The balloon catheter alone was successfully deployed in six of six femoral arteries achieving immediate hemostasis. In a second study in which the procoagulant was delivered following balloon placement, the sealing device was successfully deployed and hemostasis was achieved in 20 of 21 attempts (95%) despite removal of the balloon catheter. In a subset of fully anticoagulated animals, hemostasis was achieved in the sealing device-treated arteries at 6.5+/-3.4 minutes, but in none of the controls (P < 0.001). This novel vascular sealing device successfully achieves rapid hemostasis in normal and anticoagulated dogs following percutaneous vascular procedures.

Polylysine enhances cationic liposome-mediated transfection of the hepatoblastoma cell line Hep G2.

Plasmid DNA condensed by polylysine enhanced cationic-liposome-mediated transfection of Hep G2 cells. The luciferase expression plasmid pCMVL was complexed with the polycation poly-L-lysine and mixed with liposomes that contained a 1:1 molar ratio of the cationic lipid 1,2-dioleoyloxy-3-trimethyl-ammoniumpropane, with the neutral phospholipid 1,2-di-(cis-9-octadecenoyl)-sn-glycero-3-phosphoethanolamine. Polylysine enhanced cationic-liposome-mediated transfection of the hepatoblastoma cell line Hep G2 9-fold compared with pCMVL complexed alone with liposomes. The ratio of cationic to anionic charge of the polylysine-pCMVL complexes, and the quantity of cationic liposomes, are important determinants for optimal transfection of Hep G2 cells.

Induction of human factor VIII inhibitors in rats by immunization with human recombinant factor VIII: a small animal model for humans with high responder inhibitor phenotype.

Hemophilia A is a clotting disorder that is due to reduced or absent coagulation factor VIII (FVIII) activity. In approximately 25% of people with severe hemophilia A, standard treatment with intravenous plasma-derived or recombinant FVIII (rFVIII) induces anti-FVIII antibodies that inhibit FVIII activity (inhibitors). We describe the development of a rat model to study the formation of inhibitors. Immunization of rats with human rFVIII in adjuvant induced an anti-human rFVIII antibody response characteristic of an anti-FVIII inhibitor response in hemophilia A patients. The rats exhibited a rapid, polyclonal secondary antibody response to human rFVIII. These antibodies were reactive against epitopes located in the heavy and light chains. All the rFVIII-immunized rats developed antibodies against the FVIII C2 domain, a region of major reactivity in hemophilia A patients with inhibitors. Furthermore, competition ELISAs demonstrated that rat and human anti-FVIII antibodies recognized identical or overlapping epitopes of the FVIII molecule. The rat anti-FVIII antibodies also functioned as human FVIII inhibitors with titers ranging from 120 to 2048 Bethesda Units (B.U.). We propose that this rat model may be useful to investigate immune responses to FVIII and may lead to better therapies for FVIII inhibitors.

Vascular endothelial growth factor causes endothelial proliferation after vascular injury.

Vascular endothelial growth factor was infused into rat carotid arteries for 3 minutes immediately after endothelial denudation by balloon injury. Endothelial proliferation was determined by immunohistochemical labelling of proliferating cell nuclear antigen using Häutchen preparations. The proliferation index, or number of proliferating cells/total cells, measured at 25.5 or 30 hours was markedly increased after infusion of vascular endothelial growth factor. In addition, the total number of proliferating cells increased with increasing doses up to 100 micrograms total dose per infusion. These data indicate that infusion of vascular endothelial growth factor increases endothelial cell proliferation after mechanical denudation injury of the vascular wall.

Gene expression following direct injection of DNA into liver.

The liver is an attractive target tissue for gene therapy. Current approaches for hepatic gene delivery include retroviral and adenoviral vectors, liposome/DNA, and peptide/DNA complexes. This study describes a technique for direct injection of DNA into liver that led to significant gene expression. Gene expression was characterized in both rats and cats following injection of plasmid DNA encoding several different proteins. Luciferase activity was measured after injection of plasmid DNA encoding the luciferase gene (pCMVL), beta-galactosidase (beta-Gal) activity was evaluated in situ using plasmid DNA encoding Lac Z (pCMV beta), and serum concentration of secreted human alpha-1-antitrypsin was measured following injection of plasmid DNA encoding this protein (pRC/CMV-sHAT). Several variables, including injection technique, DNA dose, and DNA diluent, were investigated. Direct injection of pCMVL resulted in maximal luciferase expression at 24-48 hr. beta-Gal staining demonstrated that the majority of transfected hepatocytes were located near the injection site. Significant concentrations of human alpha-1-antitrypsin were detected in the serum of animals injected with pRC/CMV-sHAT. These findings demonstrate the general principle that direct injection of plasmid DNA into liver can lead to significant gene expression.

Dexamethasone enhancement of gene expression after direct hepatic DNA injection.

The critical physiological functions of the liver make hepatocytes important targets for therapeutic gene delivery. This study reports significant gene expression following direct injection of plasmid DNA into the livers of rats and cats. Transfection was characterized using luciferase and Lac Z expression from plasmids with the cytomegalovirus immediate early promoter/enhancer (CMV IE) or the Rous sarcoma virus long terminal repeat (RSV LTR). Dexamethasone treatment enhanced and prolonged transfected gene expression, possibly by activating gene expression. Southern analysis of total DNA extracted from liver at various times following injection detected persistent unintegrated plasmid DNA which maintained a prokaryotic methylation pattern. This study demonstrates the feasibility of direct DNA injection in the experimental analysis of hepatic gene expression in vivo.

Effect of cilazapril on the proliferative response after vascular damage.

Cilazapril, a novel long-acting inhibitor of angiotensin-converting enzyme, markedly suppressed the proliferative response and neointima formation after balloon catheter-induced injury of the carotid artery in a rat model of angioplasty. The reduction in neointima was dose-dependent, required sustained high levels of enzyme inhibition, and was significantly greater in animals treated starting 6 days prior to the procedure than in animals starting treatment the day of catheterization. In experiments with vascular smooth-muscle cells (SMC) in culture, the addition of angiotensin II reduces increased mRNA levels for several growth factors and extracellular matrix proteins. Here we report that Ang II selectively induces mRNA for thrombospondin I, but not for thrombospondin II. Under selected conditions SMC can be induced to proliferate after exposure to Ang II, in vitro and in vivo. Using neutralizing anti transforming growth factor beta (TGF-beta) antibodies we found that Ang II stimulation of proliferation was threefold greater when the anti-TGF-beta was added to the cultures. We suggest (a) that an important effect of Ang II during the proliferative response is the induction of thrombospondin I, which is required for matrix interactions during the formation of neointima, and (b) that, among the complex array of growth factors potentially active in vivo, TGF-beta may be an important negative regulatory factor that limits the proliferative response and prevents restenosis in most cases of angioplasty.