[Expression and clinical implication of platelet-derived growth factor-D and platelet-derived growth factor-beta in childhood IgA nephropathy].

OBJECTIVE: To investigate the clinical implication of platelet-derived growth factor (PDGF)-D and PDGF-beta in IgA nephropathy in childhood. METHODS: Forty-seven children with IgA nephropathy and 26 controls were enrolled for study, and their serum, urine and renal biopsy specimens were examined. The patients were divided into control group [including serum, urine specimens of 13 healthy children and 13 renal biopsy samples of non-IgA nephropathy in children], mild proliferation (MP) group (13 patients), focal proliferation (FP) group (19 patients), and proliferation sclerosis (PS) group (15 patients). Enzyme-linked immunosorbent assay (ELISA) and immunohistochemistry were used to determine contents of PDGF-D, PDGF-beta and PDGF-B in blood, urine and renal tissues. The levels of 24-hour urinary protein excretion, serum albumin (Alb), serum blood urea nitrogen (BUN) and creatinine (Cr) were also determined. RESULTS: Compared with control group, levels of PDGF-D and PDGF-B were progressively elevated in blood and urine of IgA nephropathy children with increase in severity of glomerular damage (all P<0.01). Serum as well as urinary PDGF-D and PDGF-B levels were positively correlated with 24-hour urinary protein excretion (PDGF-D blood: r=0.546, urine: r=0.760; PDGF-B blood: r=0.634, urine: r=0.577, respectively, P<0.01), while negatively correlated with serum Alb levels in IgA nephropathy patients (PDGF-D blood: r=-0.649, urine: r=-0.528; PDGF-B blood: r=-0.613, urine: r=-0.531, respectively, P<0.01). Contents of PDGF-D and PDGF-beta in renal tissue were much higher than those of control group (P<0.01). Along with the increase in severity of glomerular pathology, their contents increased gradually. PDGF-B was only significantly expressed in renal tissue in FP group and PS group. CONCLUSION: PDGF-D might significantly enhance the development of mesangial proliferation and tubulointerstitial fibrosis. In comparison with PDGF-B, PDGF-D appears to reflect more sensitive to the severity and prognosis of IgA nephropathy.

Prostaglandin E2 and collagenase production by fibroblasts and synovial cells is regulated by urine-derived human interleukin 1 and inhibitor(s).

Interleukin 1 (IL-1) possesses multiple biological activities that may be blocked selectively by different inhibitors. Some known inhibitors block the lymphocyte activating factor (LAF/IL-1) but not the mononuclear cell factor (MCF/IL-1) measured by its capacity to stimulate prostaglandin E2 (PGE2) and collagenase production. The presence of IL-1 in vivo may be difficult to detect due to the presence of inhibitor(s) and the level of the inhibitor(s) may vary depending upon pathological conditions. We have found that urine from three patients with monocytic leukemia (M5) contained high levels of inhibitor(s) of MCF/IL-1, whereas urine of normal subjects did not contain significant amounts. Urine from two patients with other blood neoplasic diseases also contained little inhibitory activity. The MCF/IL-1 inhibitor(s), which also acts on human recombinant IL-1 beta, is approximately 25-35 kD, is not retained on concanavalin A-Sepharose column and can be partially destroyed with urea and boiling. At this stage of the purification the fraction containing the MCF/IL-1 inhibitor(s) also inhibits the LAF/IL-1 assay. However, this inhibitor(s) is probably distinct from other inhibitors already described.

Identification of the lymphokine soluble immune response suppressor in urine of nephrotic children.

Patients with minimal change nephrotic syndrome (MCNS) frequently have suppressed in vivo and in vitro immune responsiveness of uncertain etiology. Because increased suppressor cell activity has been associated with this disease, urines from MCNS patients were screened for activity of the lymphokine soluble immune response suppressor (SIRS), a product of concanavalin A- or interferon-activated suppressor T cells. Urines from untreated MCNS patients suppressed polyclonal plaque-forming cell responses of cultured splenocytes. This suppressive activity was identified as human SIRS by the following functional and physical criteria: molecular weight estimated by gel filtration; kinetics of suppression; inhibition of suppression by catalase, levamisole, and 2-mercaptoethanol; abrogation of activity by acid or protease treatment; elution pattern on high performance liquid chromatography; and cross-reactivity with monoclonal antimurine SIRS antibodies. Suppressive activity disappeared from urine after initiation of treatment but before remission of symptoms. Urines were tested from 11 patients with MCNS, all of whom excreted SIRS. In addition, two nephrotic patients with acute glomerulonephritis and three nephrotic patients with membranoproliferative disease excreted SIRS, but other nephrotics and all nonnephrotic patients did not. These results indicate that excretion of SIRS occurs in certain cases of nephrotic syndrome and that the presence of SIRS in the urine is not accounted for solely by the presence of proteinuria or nephrosis. Serum from four nephrotic patients also contained SIRS, whereas neither serum nor urine from six normal subjects contained SIRS activity. The systemic presence of SIRS in these four patients, and the identification of SIRS in urines from a larger group of patients, suggest a possible role for SIRS in the suppressed immune responses often found in nephrotic syndrome.

Vascular permeability factor (vascular endothelial growth factor) in guinea pig and human tumor and inflammatory effusions.

Vascular permeability factor (VPF), also known as vascular endothelial growth factor, is a dimeric M(r) 34,000-42,000 glycoprotein that possesses potent vascular permeability-enhancing and endothelial cell-specific mitogenic activities. It is synthesized by many rodent and human tumor cells and also by some normal cells. Recently we developed a sensitive and specific time-resolved immunofluorometric assay for quantifying VPF in biological fluids. We here report findings with this assay in guinea pigs and patients with both malignant and nonmalignant effusions. Line 1 and line 10 tumor cells were injected into the peritoneal cavities of syngeneic strain 2 guinea pigs, and ascitic fluid, plasma, and urine were collected at various intervals. Within 2 to 4 days, we observed a time-dependent, parallel increase in VPF, ascitic fluid volume, and tumor cell numbers in animals bearing either tumor line; in contrast, VPF was not detected in plasma or urine, even in animals with extensive tumor burdens. However, low levels of VPF were detected in the inflammatory ascites induced by i.p. oil injection. In human studies, high levels of VPF (> 10 pM) were measured in 21 of 32 effusions with cytology-documented malignant cells and in only seven of 35 effusions without cytological evidence of malignancy. Thus, VPF levels in human effusions provided a diagnostic test for malignancy with a sensitivity of 66% and a specificity of 80% (perhaps as high as 97% in that six of the seven cytology-negative patients with VPF levels > 10 pM had cancer as determined by other criteria). As in the animal tumor models, VPF was not detected in serum or urine obtained from patients with or without malignant ascites. Many nonmalignant effusions contained measurable VPF but, on average, in significantly smaller amounts than were found in malignant effusions. VPF levels in such fluids correlated strongly (p = 0.59, P < 0.001) with monocyte and macrophage content. Taken together, these data relate ascitic fluid accumulation to VPF concentration in a well-defined animal tumor system and demonstrate, for the first time, the presence of VPF in human malignant effusions. It is likely that VPF expression by tumor and mononuclear cells contributes to the plasma exudation and fluid accumulation associated with malignant and certain inflammatory effusions. The VPF assay may prove useful for cancer diagnosis as a supplement to cytology, especially in tumors that grow in the pleural lining but not as a suspension in the effusions that they induce.

Vascular endothelial growth factor and basic fibroblast growth factor urine levels as predictors of outcome in hormone-refractory prostate cancer patients: a cancer and leukemia group B study.

Better prognostic markers are needed for hormone-refractory prostate cancer (HRPC) patients. No single biochemical or clinical parameter can reliably predict patient response to therapy or rapidity of disease progression. Peptide factors involved in major cancer growth pathways, such as tumor angiogenesis, are attractive candidates as markers of low- and high-risk HRPC patients. We analyzed prospectively collected urine specimens from 100 of 390 HRPC patients undergoing therapy with the growth factor antagonist suramin as part of CALGB 9480. Levels of vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) were assessed from day 1 of therapy (D1) and day 29 (D29) urine samples from this subset of 100 randomly selected patients. Growth factor levels were determined by standardized ELISA microtiter plate assays from a commercial (bFGF) or proprietary (VEGF) source. Pretreatment urine VEGF levels were predictive of survival. In univariate analysis, patients whose baseline urine VEGF level was < or =28 pg/ml (the median level) had an average survival of 17 months; those with baseline VEGF >28 pg/ml had a significantly shorter survival of 10 months (P = 0.024). This difference corresponded to a 60% increased risk of dying for the higher urine VEGF patients (hazard ratio, 1.62; P = 0.03) and remained significant in multivariate analysis (hazard ratio, 1.72, P = 0.02). No significant correlations between urine bFGF level or change in bFGF levels and survival were found. These results support the notion that certain peptide growth factor-mediated, mitogenic pathways are important in HRPC and that their levels can predict outcome.

Effects of the cyclooxygenase inhibitor, piroxicam, on tumor response, apoptosis, and angiogenesis in a canine model of human invasive urinary bladder cancer.

The mechanisms by which cyclooxygenase inhibitors exert antitumor effects are not completely defined but are postulated to involve antiangiogenic effects and induction of apoptosis. In this study, we determined the effects of the cox inhibitor, piroxicam, on tumor response, apoptotic index, proliferative index, cyclooxygenase-2 expression, prostaglandin E(2) concentration, tumor microvessel density, and urine basic fibroblast growth factor and vascular endothelial growth factor concentrations in pet dogs with naturally occurring invasive transitional cell carcinoma of the urinary bladder. Piroxicam caused reduction in tumor volume in 12 of 18 dogs, and this was strongly associated with induction of apoptosis (Fisher's exact test P < 0.015) and reduction in urine basic fibroblast growth factor concentration.

High glucose induces platelet-derived growth factor-C via carbohydrate response element-binding protein in glomerular mesangial cells.

Persistent high concentration of glucose causes cellular stress and damage in diabetes via derangement of gene expressions. We previously reported high glucose activates hypoxia-inducible factor-1αand downstream gene expression in mesangial cells, leading to an extracellular matrix expansion in the glomeruli. A glucose-responsive transcription factor carbohydrate response element-binding protein (ChREBP) is a key mediator for such perturbation of gene regulation. To provide insight into glucose-mediated gene regulation in mesangial cells, we performed chromatin immunoprecipitation followed byDNAmicroarray analysis and identified platelet-derived growth factor-C (PDGF-C) as a novel target gene of ChREBP In streptozotocin-induced diabetic mice, glomerular cells showed a significant increase inPDGF-C expression; the ratio ofPDGF-C-positive cells to the total number glomerular cells demonstrated more than threefold increase when compared with control animals. In cultured human mesangial cells, high glucose enhanced expression ofPDGF-C protein by 1.9-fold. Knock-down of ChREBPabrogated this induction response. UpregulatedPDGF-C contributed to the production of typeIVand typeVIcollagen, possibly via an autocrine mechanism. Interestingly, urinaryPDGF-C levels in diabetic model mice were significantly elevated in a fashion similar to urinary albumin. Taken together, we hypothesize that a high glucose-mediated induction ofPDGF-C via ChREBPin mesangial cells contributes to the development of glomerular mesangial expansion in diabetes, which may provide a platform for novel predictive and therapeutic strategies for diabetic nephropathy.

Phase I clinical trial of recombinant human endostatin administered as a short intravenous infusion repeated daily.

PURPOSE: To perform a phase I trial of recombinant human endostatin (rhEndostatin; EntreMed, Rockville, MD) given as a daily 20-minute intravenous (IV) injection in adult patients with refractory solid tumors. PATIENTS AND METHODS: The daily dose was increased from 15 to 240 mg/m(2) by a factor of 100% in cohorts of three patients. In the absence of dose-limiting toxicity, uninterrupted treatment was continued until the tumor burden increased by more than 50% from baseline. Correlative studies included dynamic contrast-enhanced magnetic resonance imaging of tumor blood flow, urinary vascular endothelial growth factor and basic fibroblast growth factor levels, rhEndostatin serum pharmacokinetics, and monitoring of circulating antibodies to rhEndostatin. RESULTS: There were no notable treatment related toxicities among 15 patients receiving a total of 50 monthly cycles of rhEndostatin. One patient with a pancreatic neuroendocrine tumor had a minor response and two patients showed disease stabilization. Linearity in the pharmacokinetics of rhEndostatin was indicated by dose-proportionate increases in the area under the curve for the first dose and the peak serum concentration at steady state. Daily systemic exposure to rhEndostatin in patients receiving 240 mg/m(2)/d was approximately 50% lower than that provided by the therapeutically optimal dose in preclinical studies. CONCLUSION: rhEndostatin administered as a 20-minute daily IV injection at doses up to 240 mg/m(2) showed no significant toxicities. Evidence of clinical benefit was observed in three patients. Due to high variability between the peak and trough serum concentrations associated with the repeated short IV infusion schedule, daily serum drug levels only briefly exceeded concentrations necessary for in vitro antiangiogenic effects.

Nuclease-resistant nucleic acid ligands to vascular permeability factor/vascular endothelial growth factor.

BACKGROUND: Vascular permeability factor/vascular endothelial growth factor (VPF/VEGF) is a potent inducer of new blood vessel growth (angiogenesis) that contributes to the pathology of many angiogenesis-associated disease states such as psoriasis, rheumatoid arthritis and cancer. Few molecular entities capable of binding to VPF/VEGF with high affinity and specificity have been described to date. RESULTS: Nuclease-resistant 2'-amino-2'-deoxypyrimidine nucleotide RNA (2'-aminopyrimidine RNA) ligands that bind to VPF/VEGF with high affinity have been identified by iterative rounds of affinity-selection/amplification from two independent random libraries. The sequence information that confers high affinity binding to VPF/VEGF is contained in a contiguous stretch of 24 nucleotides, 5'-CCCUGAUGGUAGACGCCGGGGUG-3' (2'-aminopyrimidine nucleotides are designated with italic letters). Of the 14 ribopurines in this minimal ligand, 10 can be substituted with the corresponding 2'-O-methylpurine nucleotides without a reduction in binding affinity to VPF/VEGF. In fact, the 2'-O-methyl substitution at permissive positions leads to a approximately 17-fold improvement in the binding affinity to VPF/VEGF. The higher affinity results from the reduction in the dissociation rate constant of the 2'-O-methyl-substituted RNA ligand from the protein compared to the unsubstituted ligand. The 2'-O-methyl-substituted minimal ligand, which folds into a bulged hairpin motif, is also more thermally stable than the unsubstituted ligand. Nuclease resistance of the ligand is further improved by the 2'-O-methyl substitutions and the addition of short phosphorothioate caps to the 3'- and 5'-ends. CONCLUSIONS: We have used the SELEX (systematic evolution of ligands by exponential enrichment) process in conjunction with post-SELEX modifications to define a highly nuclease-resistant oligonucleotide that binds to VPF/VEGF with high affinity and specificity.

Are basic fibroblast growth factor and vascular endothelial growth factor prognostic indicators in pediatric patients with malignant solid tumors?

Angiogenesis plays an important role in the growth, progression, and metastasis of solid tumors. Among angiogenic factors, basic fibroblast growth factor (bFGF) and vascular endothelial growth factor (VEGF) appear to be useful markers in adults with cancer. The aim of this pilot study was to determine the levels of VEGF in serum and bFGF in serum and urine of children with solid tumor at diagnosis (as measured by ELISA), and to investigate whether these parameters provide prognostic information. Forty consecutive patients with different types of cancer were prospectively included in this study. Median values of all studied angiogenic factors were higher in patients than in controls (n = 40), and the differences were statistically significant for bFGF in serum and urine: 10 versus 3 pg/ml (P = 0.0004) and 6406 versus 0 pg/g of creatinine (P < 0.0001), respectively. Among patients, median serum values of bFGF and VEGF were higher in children with metastatic disease (n = 14) than in those with localized disease (n = 26). The difference was statistically significant for serum bFGF: 17.5 versus 6 pg/ml (P = 0.02). Serum angiogenic factor levels correlated with outcome. The estimated event-free survival at 3 years was 79% for patients with normal bFGF values (n = 13) versus 42% (n = 26; P = 0.02) for those with high levels, and 71% in case of normal VEGF values (n = 20) versus 38% (n = 19; P = 0.04) for those with high levels. No benefit of normal urinary bFGF values was observed. Our results provide a rationale for exploring the clinical interest of bFGF and VEGF measurements in body fluids of a larger group of children with cancer.

A phase II study of razoxane, an antiangiogenic topoisomerase II inhibitor, in renal cell cancer with assessment of potential surrogate markers of angiogenesis.

Renal cell carcinoma (RCC) is an angiogenic tumor resistant to standard cytotoxic chemotherapeutic agents. Although often responsive to immunomodulatory agents including interleukin 2 and IFN-alpha, the overall results in randomized Phase III studies are disappointing with only modest improvements in overall survival. This Phase II study evaluated the efficacy and tolerability of razoxane, an antiangiogenic topoisomerase II inhibitor, in 40 patients (32 men, 8 women; age: range, 31-76 years; median, 58 years) with inoperable RCC. Twenty patients received razoxane 125 mg p.o., twice a day for 5 days each week for 8 weeks (one cycle). This was repeated in patients with stable disease (StD), but was discontinued after 16 weeks if there was no evidence of an objective response. Because minimal toxicity was seen, subsequent patients (n = 20) were treated until progressive disease (PD) was documented. Of 38 evaluable patients, 11 (29%) had StD for a minimum of 4 months, and the remainder had PD. Median overall survival was 7.3 months. Duration of survival was significantly better in patients with StD compared with those with PD (P = 0.003). The effect of treatment on six potential surrogate serum/plasma (vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), urokinase plasminogen activator soluble receptor (uPAsr), E-selectin, vascular cell adhesion molecule-1 (VCAM-1) and von Willebrand's factor (vWF) and two urinary (VEGF and bFGF) markers of angiogenesis was evaluated before and after 1 cycle of treatment. Pretreatment serum VEGF and E-selectin levels above the median value were associated with a poor prognosis. Serum VCAM-1 levels and urinary VEGF levels rose significantly after one cycle in patients with PD but not in those with StD. Serum VEGF, bFGF, VCAM-1 and vWF, plasma uPAsr and urinary bFGF levels were significantly higher in PD patients compared with StD patients before and/or after 1 cycle of treatment. In conclusion, razoxane is an antiangiogenic agent that has minimal toxicity and that requires further evaluation in combination with other active agents in the treatment of RCC. Surrogate serum and urinary markers of angiogenesis may have a role to play in predicting disease response and overall survival in RCC.

Results of a Phase I dose-escalating study of the antiangiogenic agent, SU5416, in patients with advanced malignancies.

SU5416 is a small molecule antiangiogenic agent that inhibits vascular endothelial growth factor (VEGF) stimulation of the KDR tyrosine kinase receptor. In this Phase I dose escalation trial, a weekly dose schedule of SU5416 was tested whereby an initial 5-day loading dose was followed by weekly maintenance infusions. The start dose was 20 mg/m(2) for the loading dose followed by 65 mg/m(2) for the weekly infusions. Dose escalations occurred at 33% until a final dose of 65 mg/m(2) (loading dose) and 190 mg/m(2) (weekly infusion) was obtained. Twenty-two patients were treated at five dose levels; tumor types included gastrointestinal (8), breast (3), lung (4), sarcoma (2), and other (5). The most common serious drug-related toxicity was headache, often associated with nausea and vomiting. Grade 1 and 2 toxicities included headache, nausea, vomiting, asthenia, pain at the infusion site, phlebitis, change in voice, and fevers. Of 19 evaluable patients, 4 obtained clinical benefit as defined by tumor regression (1) or disease stabilization for at least 12 weeks (3). Pharmacokinetic data revealed that the weekly infusion schedule prevented the reported 50-60% induction in SU5416 clearance observed with either daily or twice weekly dosing. Higher baseline levels of urine VEGF were observed in the 4 patients who gained clinical benefit, suggesting this may be a useful marker for predicting response to anti-VEGF therapies. Our results suggest that a weekly schedule of SU5416 shows signs of biological activity and is well tolerated at doses up to 145 mg/m(2).

Effects of the cyclooxygenase inhibitor, piroxicam, in combination with chemotherapy on tumor response, apoptosis, and angiogenesis in a canine model of human invasive urinary bladder cancer.

The objectives of this study were: (a) to determine the antitumor activity and toxicity of a cyclooxygenase inhibitor (piroxicam) combined with cisplatin chemotherapy in dogs with naturally-occurring, invasive transitional cell carcinoma (TCC) of the urinary bladder; and (b) to determine the effects of this treatment on prostaglandin E(2) concentration, tumor cell proliferation and apoptosis, and angiogenesis. Pet dogs with naturally-occurring invasive TCC underwent complete tumor staging before and after 10 weeks of piroxicam/cisplatin treatment. Prostaglandin E(2) concentrations were determined by immunoassay in snap-frozen tumor tissues. Apoptosis (terminal deoxynucleotidyl transferase-mediated nick end labeling assay), proliferation (proliferating cell nuclear antigen), and microvessel density were determined in formalin-fixed tissues. Urine basic fibroblast growth factor and vascular endothelial cell growth factor concentrations were determined by immunoassay. Partial remission (> or =50% reduction in tumor volume) was noted in 6 of 12 dogs treated with piroxicam/cisplatin. Renal toxicity was dose-limiting. Apoptotic index doubled with treatment in 11 of 12 dogs but was not associated with tumor response. Proliferative index decreased in five dogs, and tumor decreased in size in three of the five dogs. Change in urine basic fibroblast growth factor and vascular endothelial cell growth factor was associated with tumor response. microvessel density was not associated with tumor response. In conclusion, piroxicam/cisplatin had antitumor activity against canine TCC, a disease that closely mimics human invasive urinary bladder cancer. Strategies to prevent renal toxicity of this protocol are needed. Induction of tumor apoptosis and reduction in angiogenic factor concentrations were observed, but additional studies are needed to further define the mechanisms of the antitumor activity of piroxicam/cisplatin.

HILDA/LIF urinary excretion during acute kidney rejection.

Recently, a new lymphokine called HILDA (human interleukin for DA cells) has been described and cloned. This cytokine, initially described to be produced by alloreactive T lymphocyte clones grown from a rejected human kidney allograft, is identical to other factors termed D-factor, differentiation-inducing factor, differentiation inhibitory activity, hepatocyte-stimulating factor III, and leukemia inhibitory factor. HILDA/LIF induces various effects on neural, hemopoietic, embryonic cells as well as on bone remodeling and acute phase protein synthesis in hepatocyte. In this study we demonstrate the presence of HILDA/LIF in the urine but not in the serum of kidney graft recipients during acute rejection episodes, whereas this lymphokine was detectable neither in the serum nor in the urine of kidney transplanted patients with stable renal function. These data reinforce the notion of a possible role for this lymphokine in the inflammatory and/or the immune response.

One-year enzyme-linked immunosorbent assay follow-up of human interleukin for Da cells/leukemia inhibitory factor in blood and urine of 22 kidney transplant recipients.

The cytokine human interleukin for Da cells/leukemia inhibitory factor (HILDA/LIF) exerts multiple biological effects in vitro. In mice, high circulating levels of HILDA/LIF induce a wide range of pathophysiological events, some of them closely involved with immunological and inflammatory responses. Using a sandwich ELISA recognizing the natural human HILDA/LIF molecule with a threshold of 50 pg/ml in urine and 150 pg/ml in plasma, we monitored the urine and plasma HILDA/LIF levels of 22 patients in their first year after a kidney transplant. HILDA/LIF urine excretion is increased during acute rejection, and infections also trigger heavy HILDA/LIF plasma concentrations or urine excretion. In addition, this study raises the question of HILDA/LIF involvement in post-kidney-transplant phenomena such as hypercalcemia, osteoporosis, or the reversal of anemia.

HILDA/LIF, G.CSF, IL-1 beta, IL-6, and TNF alpha production during acute rejection of human kidney allografts.

HILDA/LIF, a recently described glycoprotein, has been characterized from supernatants of alloreactive T cell clones (CD4 and CD8) extracted from a human rejected kidney graft. This suggests a possible role for HILDA/LIF in the rejection process. In order to further investigate this possible role and the role of other cytokines in allograft rejection, we tested HILDA/LIF, G.CSF, IL-6, TNF alpha, and IL-1 beta in supernatants of cultured mononucleated cells from patients during rejection and from stable grafted patients. In addition, we also tested HILDA/LIF in urine of the same patients. No significant differences were directly observed in the production of HILDA/LIF, TNF alpha, and IL-1 beta in supernatants from mononucleated cells between rejecting and stable patients. However, when antibodies were used to block the TNF alpha and the IL-1 beta receptors, an increase of both cytokines was detected in cells from rejecting patients suggesting that an over-expression of both receptors and cytokines occurred during rejection. A significant increase was also observed for both G.CSF and IL-6 during the rejection compared to stable grafts. In addition, HILDA/LIF was detected in urine of patients during rejection and not in urine of stable patients, suggesting that this cytokine may indeed play a role in rejection.

Factor with erythroid burst-promoting activity in human urine unlike other hematopoietic growth factors.

A factor with burst-promoting activity (BPA) stimulates the formation of erythroid bursts in the presence of erythropoietin, acting on early erythroid progenitor cells (erythroid burst-forming units, or BFU-E). Here we investigated the biological properties of this factor partially purified from the urine of anemic patients. The human urinary factor did not cause the formation of late erythroid progenitor cells (erythroid colony-forming units, or CFU-E) or enhance such colony formation in the presence of erythropoietin. Thus, the urinary factor was a different substance from erythroid potentiating activity and from activin, which act on both BFU-E and CFU-E. The urinary factor promoted the colony formation of BFU-E from both humans and mice, but the human hematopoietic growth factors such as recombinant interleukin-3, interleukin-6, granulocyte-macrophage colony-stimulating factor, and macrophage colony-stimulating factor did not stimulate the formation of BFU-E derived colonies from mice. The results suggested that the factor in the urine of anemic patients was different from the hematopoietic growth factors identified so far.

Clinical pharmacokinetics, pharmacodynamics and metabolism of the novel matrix metalloproteinase inhibitor ABT-518.

PURPOSE: To investigate the pharmacokinetics, pharmacodynamics and metabolism of the novel matrix metalloproteinase (MMP) inhibitor ABT-518. METHODS: Plasma and urine samples were obtained from six patients included in a phase I trial in which ABT-518 was given once daily via the oral route. Samples were analyzed by LC-MS/MS, ELISA and immunocapture assay. The pharmacokinetics of the parent compound and of detectable metabolites were calculated. RESULTS: After a single dose of ABT-518 peak plasma levels were reached within 4-8 h. ABT-518 had an estimated clearance (Cl/F) of approximately 3 l/h, an estimated volume of distribution (V/F) of over 70 l and a terminal half-life (T(1/2)) of 20 h. At least six different metabolites were formed. Pharmacodynamic analysis for angiogenic growth factors (bFGF and VEGF) showed plasma and urine levels in the picogram range and for total MMP-9 and MMP-2 or MMP-9 activity showed plasma and urine levels in the nanogram range. CONCLUSIONS: The MMP inhibitor ABT-518 is extensively metabolized in humans. No significant correlations between pharmacokinetics and pharmacodynamics could be established.

Clinical significance of urinary vascular endothelial growth factor in patients with superficial bladder tumors.

Vascular endothelial growth factor (VEGF) is an important mediator of tumor angiogenesis and has been shown to be excreted in the urine of bladder cancer patient. The goal of this study was to evaluate urinary VEGF levels of patients with superficial bladder transitional cell carcinoma (TCC) and to determine its predictive value for recurrence. Pre-operative urinary VEGF levels were determined in 31 patients with superficial bladder TCC and 10 control patients. A quantitative enzyme immunoassay was used to measure urinary VEGF levels and the urine VEGF concentration was corrected by the creatinine concentration in a 24-h urine specimen. The corrected urinary VEGF levels were higher in patients than controls (p=0.003). Ten of 31 patients had TCC recurrences during this study. Corrected urinary VEGF levels were significantly higher in recurrent vs. non-recurrent patients (p=0.001). A cut-off value of 0.32 (corrected urinary VEGF levels) was valuable for predicting recurrences in this prospective study. However, there was no statistical correlation between VEGF levels and tumor stage (Ta or T1), tumor size or tumor grade. Pre-operative urinary VEGF levels are associated with a risk of recurrence in patient with superficial bladder TCC. Quantification of urinary VEGF may prove to be a valuable, non-invasive indicator of carcinoma recurrence in patients with superficial bladder TCC. Urinary VEGF may be a therapeutic target for intravesical therapy. However, because of the small number of cases, further studies with larger number of patients will be needed to clarify this issue.

Elevated vascular endothelial growth factor levels in the urine of patients with minimal-change nephrotic syndrome.

BACKGROUND: Vascular endothelial growth factor (VEGF) is a selective endothelial mitogen and vascular permeability factor (VPF), that is mainly produced by activated monocytes/macrophages and T cells. To our knowledge, very little is known about the involvement of VEGF in minimal-change nephrotic syndrome (MCNS). The aim here was to define further the involvement of VEGF in MCNS. PATIENTS AND METHODS: Urine samples were obtained from 20 MCNS patients. The disease controls included 20 patients with IgA nephropathy (IgAN). The samples were assayed for VEGF protein by a sandwich enzyme-linked immunosorbent assay (ELISA). RESULTS: Compared with normal controls, markedly increased urinary levels of VEGF were detected in both MCNS and IgAN patients with the nephrotic syndrome (NS). The urinary VEGF (uVEGF) levels correlated with the degree ofproteinuria in MCNS and IgAN patients. Moreover, when individual MCNS patients were followed through their clinical illness, uVEGF levels were increased during the active phase and decreased as the patients went into remission. Our main concern is to distinguish between two possibilities: Increases in uVEGF excretion might indeed relate to specific glomerular pathology and thus have a pathophysiological role. Alternatively, uVEGF may be derived from the circulation and as such may be nothing more than an assay for proteinuria. In fact, given the strict correlation between uVEGF excretion and the amount of proteinuria, the second possibility appears quite conceivable. CONCLUSION: Therefore, this may be a coincidental finding which has no bearing on the pathophysiology of MCNS.