Clinical relevance of serum angiogenic activity in patients with transitional cell carcinoma of the bladder.

Angiogenesis is essential for tumor growth and progression and is mediated by positive and negative regulators of vessel growth. Since angiogenic mediators found in patient serum have been postulated to reflect the angiogenic potential of a malignant tumor, we investigated the angiogenic activity in the serum of patients with transitional cell carcinoma (TCC). The data were correlated to tumor characteristics and the clinical course of the patients. Eighty-one patients with transitional cell carcinoma and 53 control persons were included in the study. Preoperative serum samples were collected and both vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) were quantified by ELISA. Additionally, the serum evoked proliferative activity on human umbilical vein endothelial cells (HUVEC) was evaluated. Data were compared to the clinical course of the patients. Serum of tumor patients significantly enhanced the proliferative capacity of HUVEC, compared to cells grown in standard culture medium (p = 0.0032), but not when compared to serum from control persons. Serum from patients with superficial TCC and well differentiated tumors induced a significantly higher angiogenic response (ANG(hi)) than serum from patients with poorly differentiated and invasive carcinomas (ANG(lo); p = 0.037). VEGF level of ANG(hi) serum was 384.22 +/- 247.76 pg/ml (n = 37) which significantly differed from mean VEGF level detected in ANG(lo) serum (247.72 +/- 211.93 pg/ml, n = 42; p = 0.019). Similarly, mean bFGF levels were 9.58 +/- 5.91 pg/ml in ANG(hi) serum versus 5.74 +/- 3.52 pg/ml) in ANG(lo) serum (p = 0.0043). A negative correlation was established between VEGF/bFGF serum concentration and patient prognosis. The experiments demonstrate a positive correlation between VEGF and bFGF serum level and endothelial proliferation in vitro. The inverse relationship between angiogenic activity and tumor stage might disclose information about angiogenesis and tumor progression in TCC.

Uropharmacology: current and future strategies in the treatment of erectile dysfunction and benign prostate hyperplasia.

Because of the increasing percentage of the world male population suffering from erectile dysfunction (ED) and benign prostate syndrome (BPS), there is a need for new and innovative therapeutic approaches. Pharmacotherapy is an avenue presently being followed in the treatment of both these syndromes. A profound change in the therapy of ED has been obtained through use of the selective phosphodiesterase inhibitor sildenafil. The incidence of surgical intervention in BPS has been reduced by the introduction of uroselective alpha1-receptor antagonists and the new 5alpha-reductase inhibitors (such as finasteride, dutasteride). The investigation of mechanisms in CXC chemokine expression also offers new therapeutic possibilities in these diseases.

Valproic acid induces expression of neutrophil chemoattractants of the CXC chemokine family in endothelial cells.

The branched-chain fatty acid valproate (valproic acid; VPA) displays antitumoral properties by blocking tumor growth, progression and invasion. Recent data have shown that VPA reduces the angiogenic activity of endothelial cells. The object of this study was to investigate whether endothelial modulation might also influence the level of chemotactic mediators. Endothelial cells were isolated from human umbilical cord veins (HUVEC) and treated with VPA-concentrations ranging from 0.125 mM to 1 mM. The mRNA level of CXC-chemokines was investigated by reverse transcriptase-polymerase chain reaction. The proliferative activity of HUVEC was measured as well. VPA evoked a striking increase in the neutrophil chemoattractants CXCL1, CXCL3, CXCL4, CXCL5 and a moderate increase in CXCL6 with maximal effects after a 3-day incubation period. Other CXC-chemokines and CXC-receptors remained unaffected. HUVEC growth was diminished time- and dose-dependently by VPA. We conclude that VPA treatment leads to alterations in the chemokine expression profile of endothelial cells. This might allow more neutrophils to reach the tumor area and trigger cytolysis.

The immunosuppressive drug mycophenolate mofetil impairs the adhesion capacity of gastrointestinal tumour cells.

Immunosuppression correlates with the development and recurrence of cancer. Mycophenolate mofetil (MMF) has been shown to reduce adhesion molecule expression and leucocyte recruitment into the donor organ. We have hypothesized that MMF might also prevent receptor-dependent tumour dissemination. Therefore, we have investigated the effects of MMF on tumour cell adhesion to human umbilical vein endothelial cells (HUVEC) and compared them with the effects on T cell-endothelial cell interactions. Influence of MMF on cellular adhesion to HUVEC was analysed using isolated CD4+ and CD8+ T cells, or WiDr colon adenocarcinoma cells as the model tumour. HUVEC receptors ICAM-1, VCAM-1, E-selectin and P-selectin were detected by flow cytometry, Western blot or Northern blot analysis. Binding activity of T cells or WiDr cells in the presence of MMF were measured using immobilized receptor globulin chimeras. MMF potently blocked both T cell and WiDr cell binding to endothelium by 80%. Surface expression of the endothelial cell receptors was reduced by MMF in a dose-dependent manner. E-selectin mRNA was concurrently reduced with a maximum effect at 1 microm. Interestingly, MMF acted differently on T cells and WiDr cells. Maximum efficacy of MMF was reached at 10 and 1 microm, respectively. Furthermore, MMF specifically suppressed T cell attachment to ICAM-1, VCAM-1 and P-selectin. In contrast, MMF prevented WiDr cell attachment to E-selectin. In conclusion, our data reveal distinct effects of MMF on both T cell adhesion and tumour cell adhesion to endothelial cells. This suggests that MMF not only interferes with the invasion of alloactivated T cells, but might also be of value in managing post-transplantation malignancy.

Robotic-assisted laparoscopic radical cystectomy and intra-abdominal formation of an orthotopic ileal neobladder.

PURPOSE: To describe our technique of robotic-assisted laparoscopic radical cystectomy and intra-abdominal formation of an orthotopic neobladder (Hautmann) for treatment of transitional cell carcinoma of the bladder. METHODS: We describe our surgical technique in the worldwide first attempt to perform a robotic-assisted laparoscopic radical cystectomy and completely intra-abdominal formation of an orthotopic neobladder. The DaVinci System (Intuitive Surgical, Mountain View, CA, USA) was utilized to perform the procedure. RESULTS: Utilizing the DaVinci System the operation could be performed without any complications. Operating time was 8.5 hours, blood loss was 200 ml. The oncologic as well as the functional result of the reservoir were excellent. DISCUSSION: We here demonstrated that sophisticated laparoscopic procedures like the intra-abdominal formation of an orthotopic neobladder are accomplishable with robotic assistance.

[Polymerase chain reaction in the urinary diagnosis of bladder cancer]. / Die Polymerasekettenreaktion (PCR) in der Urindiagnostik des Harnblasenkarzinoms.

The polymerase chain reaction (PCR) is a highly sensitive and specific method for the detection of genetic material. Over the last few years, this method has been used increasingly for the molecular detection of disease. This report demonstrates the use of PCR in the diagnosis of bladder cancer. The principles of the method are shown by describing the detection of p53, CD44 and telomerase, as well as in microsatellite analysis.

Robotic-assisted laparoscopic radical prostatectomy: the Frankfurt technique.

The robotic technique, which was first introduced in laparoscopic heart surgery, has revolutionized laparoscopic surgery over the last 5 years. In May 2000, our department accomplished the first robot assisted laparoscopic radical prostatectomy. Since that time we have performed more than 118 such procedures and several other laparoscopic operations using the robotic technique. We here summarize our experience in robot assisted laparoscopic radical prostatectomy as it has been developed over the past 3 years. Between May 2000 and May 2003, 118 patients with clinically localized prostate cancer were operated using the telerobotic da Vinci Surgical System. Operations were performed with a senior surgeon at the console, assisted by an assistant and a nurse at the operating table. Bilateral pelvic lymph node dissection was undertaken as a first step in all patients. In the initial 60 cases, we investigated different laparoscopic approaches. We used transperitoneal as well as extraperitoneal approaches. For dissection of the prostate we used ascending, descending as well as combined techniques. The combined ascending and descending technique via the transperitoneal route was chosen in 30 patients, and via the extraperitoneal route in seven patients. A modification of the descending Montsouris technique was performed in 81 patients. The robot assisted laparoscopic radical prostatectomy with the da Vinci system has been well standardized. After performing more than 100 radical prostatectomies with this system, we conclude that in our hands the Montsouris technique with only minor adoptions is the most appropriate technique for performing robot assisted radical prostatectomy.

In vivo retinal gene expression in early diabetes.

PURPOSE: Studies have demonstrated a causal role for specific molecules in the pathogenesis of diabetic retinopathy. Among the implicated mediators are growth factors such as vascular endothelial growth factor (VEGF) as well as adhesion molecules and proliferation- and apoptosis-related genes. However, a coordinated large-scale investigation of gene expression in the diabetic retina has not yet been reported. Here the retinal gene expression profile of diabetic and nondiabetic animals using cDNA microarrays were analyzed and compared. METHODS: Long-Evans rats were made diabetic with streptozotocin. Retinal gene expression was analyzed over 3 weeks using high-density nylon filter-based cDNA arrays. Genes were sorted into clusters according to their temporal expression profiles. They were also grouped according to their potential pathophysiological significance. The in vivo gene expression profiles of selected genes were verified via RNase protection assay. RESULTS: The rat GeneFilter contains a total of 5147 genes, of which 1691 are known genes and 3456 are expressed sequence tags (ESTs). On day 3, the expression of 27 known genes was increased by more than twofold. On days 7 and 21, the corresponding numbers were 60 and 12, respectively. A transient upregulation (>2-fold) in expression was seen in 627 of 5147 total genes. A subset of 926 genes exhibited a modest (<2-fold) decrease in expression. No genes showed a greater than twofold decrease in expression. Overall, the identity of the genes that were upregulated suggests that the response of the retina to the diabetic challenge contains an inflammatory component. Moreover, most regulatory activity occurs during the first week of diabetes. CONCLUSIONS: The development of a rational therapy for diabetic retinopathy will be assisted by detailed knowledge regarding the molecular pathophysiology of the disease. Here, an expression profile of an underlying retinal inflammatory process in early diabetes was extracted. Beyond providing insight into the general nature of the response to a pathogenic challenge, gene expression profiling may also allow the efficient identification of potential drug targets and markers for monitoring the course of disease.

Inhibition of inflammatory corneal angiogenesis by TNP-470.

PURPOSE: To determine the efficacy of the angiogenic inhibitor TNP-470 on inflammatory corneal neovascularization. Topical and systemic delivery of the drug were investigated in a murine model as well as inhibition of endothelial cell proliferation in vitro and in vivo. METHODS: The effect of TNP-470 on VEGF- and bFGF-stimulated bovine capillary endothelial (BCE) cell proliferation was evaluated in vitro. Corneal neovascularization was induced in vivo by mechanical debridement of the corneal and limbal epithelium with 0.15 M NaOH on C57BL6 mice. TNP-470 was administered systemically at 30 mg/kg body weight (BW) every other day or topically three times daily in a concentration of 5 ng/ml dissolved in methylcellulose. Vessel length was investigated on day 7. VEGF protein content in murine corneas was analyzed by ELISA on days 2, 4, and 7 of treatment. A modified bromouridine (BrdU) ELISA was used to quantify endothelial cell proliferation. RESULTS: TNP-470 exerted a dose-dependent inhibition of bFGF- and VEGF-induced endothelial cell proliferation in vitro. Both systemic and topical application of TNP-470 led to a significant reduction of inflammatory corneal neovascularization (P < 1 x 10(-5)). BrdU labeling showed that TNP-470 inhibited endothelial cell proliferation. VEGF protein levels were reduced by systemic TNP-470 treatment. CONCLUSIONS: These results suggest that TNP-470 reduces inflammatory corneal angiogenesis by directly inhibiting endothelial cell proliferation. Topical and systemic treatment with TNP-470 reduces VEGF levels that are responsible for vessel growth during the neovascularization process.

Heterogeneity of angiogenic activity in a human liposarcoma: a proposed mechanism for "no take" of human tumors in mice.

BACKGROUND: Tumor cells are known to be heterogeneous with respect to their metastatic activity, proliferation rate, and activity of several enzymes. However, little is known about the heterogeneity of tumor angiogenic activity. We investigated whether heterogeneity of angiogenic activity could be responsible for the well-known observation of "no take" of human tumors transplanted into immunodeficient mice. METHODS: Severe combined immunodeficient (SCID) mice were xenotransplanted subcutaneously with tumor tissue (n = 55) or cell suspension of a human liposarcoma cell line (SW-872) or subclones (n = 28), with varying cell proliferation rates. Xenograft tumor growth was recorded for up to 6 months. Tumor tissues were then removed and analyzed for tumor cell apoptosis, microvessel density, and cell proliferation. All statistical tests were two-sided. RESULTS: Pieces of tumor derived from the parental cell line or its clones gave rise to three kinds of tumors: 1) highly angiogenic and fast-growing (aggressive) tumors, 2) weakly angiogenic and slow-growing tumors, and 3) nonangiogenic and stable tumors. Most tumors retained the original phenotype of their parental tumor. Tumor volume correlated positively with microvessel density (Spearman correlation coefficient [r] =.89; P< or =.0001) and inversely with tumor cell apoptosis (Spearman r = -.68; P =.002). Tumor volume was less strongly but still positively correlated with tumor cell proliferation in vivo (Spearman r =.55; P =.02). CONCLUSIONS: Human liposarcoma cells appear to be heterogeneous in their angiogenic activity. When tumor cells with little or no angiogenic activity are transplanted into SCID mice, a microscopic, dormant tumor results that may not grow further. Because such tiny tumors are neither grossly visible nor palpable, they have previously been called "no take." The finding that an angiogenic tumor can contain subpopulations of tumor cells with little or no angiogenic activity may provide a novel mechanism for dormant micrometastases, late recurrence, and changes in rate of tumor progression.

Sonographic evaluation of orthotopic bladder tumors in mice treated with TNP-470, an angiogenic inhibitor.

RATIONALE AND OBJECTIVES: The purpose of this study was to determine the feasibility of using transabdominal ultrasonography (US) to monitor tumor growth and response to therapy in a mouse model of orthotopic bladder carcinoma. MATERIALS AND METHODS: Human bladder carcinoma cell suspensions were injected into the bladders of 18 SCID mice, allowed to grow for 3 weeks, and monitored weekly with gray-scale US. After 23 days, five animals were treated with TNP-470, an angiogenic inhibitor, and five control animals were treated with saline solution. US images were evaluated for tumor location, size, and neovascularity. All untreated animals (n = 8) were imaged and sacrificed at 25 days. Eight of the treated animals were imaged and sacrificed after 14 days of treatment. US findings for both groups were compared with autopsy findings. RESULTS: While saline-treated tumors continued to grow, the growth of TNP-470-treated tumors was arrested within 7 days of therapy (P < .02). Tumors as small as 1.5 mm were identified prospectively with US. US volume estimates correlated well with autopsy volume measurements (r2 = 1.0, P < .0001). Although tumor neovascularity was identified in every animal, the pattern of neovascularity did not correlate with tumor volume or therapy. CONCLUSION: US can provide accurate intermediate end points for monitoring experimental intraabdominal tumor growth and response to therapy in the mouse model.

Radiation therapy to a primary tumor accelerates metastatic growth in mice.

The surgical removal of a primary tumor can result in the rapid growth of metastases. The production of angiogenesis inhibitors by the primary tumor is one mechanism for the inhibition of metastatic tumor growth. The effect of curative radiotherapy to a primary tumor known to make an inhibitor of angiogenesis and the effects on distant metastases has not been studied. We here show that the eradication of a primary Lewis lung carcinoma (LLC-LM), which is known to generate angiostatin, is followed by the rapid growth of metastases that kill the animal within 18 days after the completion of radiation therapy. The right thighs of C57BL/6 mice (n = 25) were injected s.c. with 1 x 10(6) LLC-LM cells. Animals were randomized to one of five groups: no irradiation, 40 Gy in one fraction, 30 Gy in one fraction, 40 Gy in two 20 Gy fractions, or 50 Gy in five 10 Gy fractions. Tumors were clinically eradicated in each treatment group. All of the surviving animals became dyspneic and were killed within 14-18 days after the completion of radiation therapy. Examination of their lungs revealed >46 (range, 46-62) surface metastases in the treated animals compared with 5 (range, 2-8) in the untreated animals. The lung weights had increased from 0.2 g (range, 0.19-0.22 g) in the controls to 0.58 g (range 0.44-0.84) in the experimental animals. The most effective dose regimen was 10 Gy per fraction for five fractions, and serial experiments were conducted with this fractionation scheme. Complete response of the primary tumor was seen in 25 of 35 (71%) mice. The average weight of the lungs in the nonirradiated animals was 0.22 g (range, 0.19-0.24 g) and in the irradiated animals was 0.66 g (range, 0.61-0.70 g). The average number of surface metastases increased from five per lung (range, 2-13) in the control animals to 53 per lung (range, 46-62) in the irradiated animals. Both differences were statistically significant with P < 0.001. If the nontumor-bearing leg was irradiated or the animals were sham-irradiated, no difference in the number of surface metastases or lung weights was observed between the control group and the treated group. Urinary levels of matrix metalloproteinase 2, the enzyme responsible for angiostatin processing in this tumor model, were measured and correlated with the viability and size of the primary tumor. Administration of recombinant angiostatin prevented the growth of the metastases after the treatment of the primary tumor. In this model, the use of radiation to eradicate a primary LLC-LM tumor results in the growth of previously dormant lung metastases and suggests that combining angiogenesis inhibitors with radiation therapy may control distant metastases.

Effect of antiangiogenic therapy on slowly growing, poorly vascularized tumors in mice.

BACKGROUND: Angiogenesis is essential for tumor growth and progression. Therefore, inhibition of angiogenesis is being studied as a new anticancer therapy. Because cytotoxic chemotherapy is more effective on rapidly growing tumors than on slowly growing tumors, it has been assumed that antiangiogenic therapy will also be effective only on rapidly growing, highly vascularized tumors. We compared the effects of two angiogenesis inhibitors, TNP-470 and angiostatin, on slowly growing, poorly vascularized and rapidly growing, highly vascularized human tumors in mice. METHODS: Slowly growing (RT-4) and rapidly growing (MGH-U1) human bladder carcinoma cell lines were grown in severe combined immunodeficiency mice. Established tumors were treated with one of the two angiogenesis inhibitors. Tumor volumes, vascularity, and proliferation indices were determined. The in vitro effects of TNP-470 and of angiostatin on the proliferation of RT-4 and MGH-U1 cells were also investigated. All statistical tests were two-sided. RESULTS: RT-4 and MGH-U1 tumor growth was statistically significantly inhibited by both angiogenesis inhibitors (P<.001). Both inhibitors decreased the blood vessel density in both tumor types but did not alter the in vivo proliferation indices of the tumors. TNP-470, but not angiostatin, marginally decreased the in vitro proliferation of MGH-U1 cells. CONCLUSION: Slowly growing, poorly vascularized tumors in animal models respond as well as rapidly growing, highly vascularized tumors to therapy with the angiogenesis inhibitors TNP-470 and angiostatin.

Efficacy of antiangiogenic therapy with TNP-470 in superficial and invasive bladder cancer models in mice.

OBJECTIVES: To evaluate the efficacy of antiangiogenic therapy with O-(chloracetyl-carbamoyl) fumagillol (TNP-470) in a superficial and an invasive bladder cancer model in mice. The control of recurrent superficial and metastatic bladder cancer constitutes a major problem in urologic practice. Although the established therapies for these cases (immunotherapy, chemotherapy, and radiation therapy) have improved during the previous decades, further improvement and the reduction of existing side effects are needed. The inhibition of angiogenesis represents a new concept in cancer therapy. METHODS: We evaluated the in vitro effect of TNP-470 on the proliferation of bovine capillary endothelial cells (BCE), the superficial transitional cell carcinoma (TCC) cell line (KK-47), and the invasive TCC cell line (MGH-U1). To evaluate the in vivo effect of TNP-470 on the growth of advanced TCCs, both cell lines were injected subcutaneously into SCID mice. When tumors grew to a size of 100 to 200 mm(3), therapy either with TNP-470 or phosphate-buffered saline was initiated. RESULTS: TNP-470 strongly inhibited endothelial cell proliferation in vitro. The in vitro proliferation of both bladder carcinoma cell lines was also inhibited by TNP-470. However, the doses inhibitory to bladder carcinoma cells were 100-fold higher than the doses that were effective in the inhibition of endothelial cell proliferation. In vivo, TNP-470 significantly inhibited the growth of advanced KK-47 (67%) and MGH-U1 (68%) tumors in SCID mice. CONCLUSIONS: Our results indicate that antiangiogenic therapy with TNP-470 is equally effective in advanced superficial and invasive bladder carcinoma models in mice. When our results are taken together with the reports of other laboratories, TNP-470 appears to be a promising candidate as a tumor suppressor in superficial and invasive bladder cancer.

New molecular mediators in tumor angiogenesis.

Angiogenesis is essential for tumor growth and progression. It has been demonstrated that tumor growth beyond a size 1 to 2 mm(3) requires the induction of new vessels. Angiogenesis is regulated by several endogenous stimulators and inhibitors of endothelial cell migration, proliferation and tube formation. Under physiological conditions these mediators of endothelial cell growth are in balance and vessel growth is limited. In fact, within the angiogenic balance endothelial cell turnover is sufficient to maintain a functional vascular wall but does not allow vessel growth. Tumor growth an progression has successfully been correlated to the serum concentration of angiogenic mediators. Furthermore, the vascular density of tumor tissues could be correlated to the clinical course of the disease in several tumor entities. Within the last years several new mediators of endothelial cell growth have been isolated e.g. angiopoietin 1, angiopoietin 2, midkine, pleiotropin, leptin and maspin. In this review we discuss the mechanisms leading to tumor angiogenesis and describe some of the newer mediators of endothelial cell stimulation and inhibition.

[Pathologic humeral fracture in secondary osteosarcoma of the humerus: a rare complication of osteodystrophia deformans Paget]. / Pathologische Oberarmfraktur beim sekundären Osteosarkom des Humerus: eine seltene Komplikation der Osteodystrophia deformans Paget.

Pathological fracture in histologically proven post-Paget osteosarcoma of the humerus is a rare complication. Due to individual requests as well as age and comorbidity, a course of primary palliative treatment was chosen in the present case. Survival time after diagnosis was 9 months and the patient died of a tumor-independent disease. Even in combined treatment, consisting of surgery and (neo-)adjuvant radio-/chemotherapy, prognosis of osteosarcomas secondary to Paget's disease remains very disappointing. Therefore, in treatment of this highly lethal tumor the patient's individual requests and personal situation often require more consideration than in many other malignancies.