Involvement of VEGF and its receptors in ascites tumor formation

Vascular endothelial growth factor (VEGF) has potent endothelial cell mitotic and vascular permeability activity. Several reports have suggested that VEGF may be one of the major factors regulating ascites formation, although no quantitative and systematic analyses have been carried out. To determine the role of VEGF in ascites formation, we examined the expression of VEGF in 13 mouse ascites tumors (5 sarcomas, 3 carcinomas, and 5 hematopoietic malignancies). We found that significant amounts (6-850 ng/mL) of biologically active VEGF accumulated in the ascites fluid of all 13 tumors, particularly in tumors of sarcoma and carcinoma origin (430 - 234 ng/mL). The microvessel densities in the peritoneal walls of tumor-bearing mice, which are significantly higher than those in healthy mice, basically correlated with but did not parallel VEGF concentrations, suggesting the existence of an additional modulator(s) of the angiogenic process. Administration of anti-mouse VEGF-neutralizing antibody to mice bearing the carcinoma-derived ascites tumor MM2 suppressed ascites accumulation, tumor growth, and tendency to bleed. These results directly demonstrate the crucial role of VEGF in carcinoma-derived ascites tumor formation in vivo.     

Japanese Orthopaedic Association Back Pain Evaluation Questionnaire. Part 2. Verification of its reliability

Background The project to develop a new Japanese Orthopaedic Association (JOA) score rating system for low back disorders, the JOA Back Pain Evaluation Questionnaire (JOABPEQ), is currently in progress. Part 1 of the study selected 25 “candidate” items for use on the JOABPEQ. The purpose of this current Part 2 of the study was to verify the reliability of the questionnaire. Methods A total of 161 patients with low-back disorders of any type participated in the study. Each patient was interviewed twice at an interval of 2 weeks using the same questionnaire. The reliability of the questionnaire was evaluated by determining the extension of the kappa and weighted kappa coefficients. Results Both kappa and weighted kappa were more than 0.50 for all but one item, which was 0.48. The lower 95% confidence interval exceeded 0.4 in all but two items, which was 0.39. This implied that the test–retest reliability of JOABPEQ was acceptable as a measure of outcome. Conclusions The tentative questionnaire of the JOABPEQ with 25 items was confirmed to be reliable enough to describe the quality of life of patients who suffer low back disorders. This report was composed by the Subcommittee on Low Back Pain and Cervical Myelopathy Evaluation of the Clinical Outcome Committee of the Japanese Orthopaedic Association     

Hypoxia and low-nutrition double stress induces aggressiveness in a murine model of melanoma

Antiangiogenic therapy is a potent cancer treatment, however, the possibility of recurrence and resistance to this approach remains. Here we show that hypoxia and low-nutrition double-deprivation stress induces reversible tumor aggressiveness. In a stress-cycle-dependent manner, murine melanoma cells showed morphological changes, up-regulated phospho-Akt, and abnormal regulation of multiple genes including fibroblast growth factor-21, a metabolic regulator, resulting in increased cell proliferation in vitro , and increased tumorigenesis and invasive potential in vivo . In this system, altered cellular metabolism participates in the adaptation of tumor to the double-deprivation stress. Our results suggest the targeting of a minor population of cancer cells resistant to both hypoxia and low nutrition to be an effective new antitumor strategy in combination with antiangiogenic therapy. ( Cancer Sci 2009; 100: 844–851)     

Inhibition of choroidal neovascularization by blocking vascular endothelial growth factor receptor tyrosine kinase

Purpose To investigate the role played by receptors of vascular endothelial growth factors, Flt-1 and KDR/Flk-1, on an experimental model of choroidal neovascularization (CNV). Methods The vascular endothelial growth factor-A (VEGF-A) receptor-specific tyrosine kinase inhibitor SU5416 was administered to a laser-induced mouse model of CNV. The formation of CNV and the degree of vascular permeability in Flt-1 tyrosine kinase domain-deficient mice were also investigated. Results SU5416 reduced vascularity and vascular endothelial cell proliferation, and promoted endothelial cell apoptosis within CNV. Furthermore, the formation of CNV and the degree of vascular permeability were significantly reduced in Flt-1 tyrosine kinase domain-deficient mice, and this effect was enhanced by the administration of SU5416. Conclusions Both Flt-1 and KDR/Flk-1 have a significant role in CNV formation. Suppression of apoptosis may be involved in the process.     

Transient evoked otoacoustic emissions in newborn infants: Effects of ear asymmetry, gender, and age

Our aim was to examine the effects of gender, ear asymmetry, and age of infants on various parameters of transient evoked otoacoustic emissions (TEOAEs). Three hundred thirty-two infants (181 males, 151 females) were tested using the ILO292 Otodynamics Analyzer (Otodynamics Ltd, England) as a screening procedure. The subjects were divided into two age groups: group 1, newborn infants prior to hospital discharge (mean age of 4 days), and group 2, infants at the 1-month-old health checkup (mean age of 35 days). Responses to TEOAE stimuli were recorded at 1.0, 1.5, 2.0, 3.0, and 4.0 kHz. There were significant effects of gender and ear (left/right) on the signal-to-noise ratio, response level, and whole-wave and band reproducibility values in TEOAEs. The right ear had higher values of whole-wave reproducibility, response level, signal-to-noise ratio, and band reproducibility than the left ear. Females displayed higher whole-wave reproducibility, response level, band reproducibility, and signal-to-noise ratio values than males. There was no significant difference in response level, signal-to-noise ratio, and band reproducibility between the two age groups. The findings of the present investigation may contribute toward future improvements in neonatal hearing screening based on the community.     

A phase II study of uracil-tegafur plus doxorubicin and prognostic factors in patients with unresectable biliary tract cancer

Purpose

The purpose of this study was to clarify the safety and efficacy of combination chemotherapy of uracil-tegafur (UFT) and doxorubicin (UFD regimen), and to identify the prognostic factors in patients with unresectable advanced biliary tract cancer who received systemic chemotherapy.

Methods

Patients with histologically or cytologically confirmed, measurable biliary tract cancer, including intrahepatic or extrahepatic cholangiocarcinoma, gallbladder cancer, and ampullary cancer, who were not suitable candidates for surgery, were eligible for the study. Patients received oral UFT at 300 mg/m² per day divided into two doses on days 1–14 and intravenous doxorubicin at 30 mg/m² on day 1. This cycle was repeated every 21 days. The relationship between the patient characteristics and the prognosis was examined. Univariate and multivariate analyses were conducted to identify the prognostic factors associated with survival.

Results

Sixty-one patients from 12 institutions were enrolled in the late phase II study between April 2005 and March 2006. Of the 61 patients, 4 patients had partial responses, for an objective response rate of 6.6% (95% CI: 1.8–15.9%); 28 patients had stable disease, 27 had progressive diseases, and 2 patients were not evaluated. The median progression-free survival was 1.6 months, and the overall median survival time was 6.5 months. In the 85 patients who received this UFD chemotherapy in previous and late phase II studies, multivariate analysis revealed the ECOG performance status 1 (P = 0.001), gallbladder as the primary cancer site (P = 0.014), T-factor 4 of the TNM classification (P = 0.035), and elevated serum lactate dehydrogenase levels (P = 0.043) as being associated with a significantly shorter survival.

Conclusions

Combination chemotherapy of UFT and doxorubicin had minimum activity against advanced biliary tract cancer. Performance status was identified as the most important prognostic factor in patients who received systemic chemotherapy.     

Growth inhibition of AML cells with specific chromosome abnormalities by monoclonal antibodies to receptors for vascular endothelial growth factor

By using neutralizing monoclonal antibodies to vascular endothelial growth factor receptor type 1 (VEGFR1) and VEGFR2, we have shown that acute myelogenous leukemia (AML) cells with specific chromosome abnormalities are dependent on VEGF/VEGFR system. AML with t(8;21) is the most dependent subtype on VEGF with both VEGFR1 and VEGFR2. t(15;17)AML cells depend on VEGF with VEGFR1. AML cells with 11q23 abnormalities showed variable dependence on VEGF. The growth of t(11;19)AML cells are most extensively inhibited by anti-VEGFR1 antibody. Then, the growth of Kasumi-1, a t(8;21) cell line was suppressed by either anti-VEGFR1 antibody (p = 0.0022) or anti-VEGFR2 antibody (p = 0.0029) in a dose-dependent manner. The growth of NB4, a t(15;17) cell line was more potently suppressed by anti-VEGFR1 antibody (p = 0.0111) than by anti-VEGFR2 antibody (p = 0.0477). These results are quite concordant with the results of clinical samples with t(8;21) or t(15;17). In addition, anti-VEGFR2 monoclonal antibody significantly potentiated the growth inhibitory effect of idarubicin for Kasumi-1. As for downstream signals, we have shown that VEGFR2 transduce growth and survival signals through phosphorylation of Akt and MEK in leukemia cells (Kasumi-1). However, VEGFR1 transduce growth and survival signals through pathways other than MEK and Akt (NB4), although Akt phosphorylation may account for some of the VEGFR1 signals (Kasumi-1). Finally, our data suggested that autocrine pathway of VEGF and VEGFRs observed in AML cells with specific chromosomal translocations have contributed to leukemogenesis as activated signaling of receptor tyrosine kinase.     

M-CSF inhibition selectively targets pathological angiogenesis and lymphangiogenesis

Antiangiogenic therapy for the treatment of cancer and other neovascular diseases is desired to be selective for pathological angiogenesis and lymphangiogenesis. Macrophage colony-stimulating factor (M-CSF), a cytokine required for the differentiation of monocyte lineage cells, promotes the formation of high-density vessel networks in tumors and therefore possesses therapeutic potential as an M-CSF inhibitor. However, the physiological role of M-CSF in vascular and lymphatic development, as well as the precise mechanisms underlying the antiangiogenic effects of M-CSF inhibition, remains unclear. Moreover, therapeutic potential of M-CSF inhibition in other neovascular diseases has not yet been evaluated. We used osteopetrotic (op/op) mice to demonstrate that M-CSF deficiency reduces the abundance of LYVE-1⁺ and LYVE1⁻ macrophages, resulting in defects in vascular and lymphatic development. In ischemic retinopathy, M-CSF was required for pathological neovascularization but was not required for the recovery of normal vasculature. In mouse osteosarcoma, M-CSF inhibition effectively suppressed tumor angiogenesis and lymphangiogenesis, and it disorganized extracellular matrices. In contrast to VEGF blockade, interruption of M-CSF inhibition did not promote rapid vascular regrowth. Continuous M-CSF inhibition did not affect healthy vascular and lymphatic systems outside tumors. These results suggest that M-CSF–targeted therapy is an ideal strategy for treating ocular neovascular diseases and cancer.Pathological angiogenesis is a hallmark of cancer and various inflammatory conditions including ocular neovascular diseases (1, 2). Recent studies have led to the discovery of a growing number of antiangiogenic molecules, some of which are already being evaluated in clinical trials (1, 2). The most established approach for limiting pathological angiogenesis involves blockade of the vascular endothelial growth factor (VEGF) pathway (3–5); however, several studies have shown that VEGF blockade damages healthy vessels and results in toxic side effects (6, 7) and that interrupting the VEGF blockade induces rapid vascular regrowth in tumors (8). Clinical trials involving patients with neovascular age-related macular degeneration have shown that VEGF blockade has limited effectiveness in some patients (5). Therefore, several groups have explored other targets that could potentially be combined with VEGF blockade (9–11). In particular, tumor-associated hematopoietic cells are thought to be a promising therapeutic target, especially neutralizing Bv8, which promotes both tissue-specific angiogenesis and hematopoietic cell mobilization (10).Macrophage CSF (M-CSF) (12) is a cytokine required for the differentiation of monocyte lineage cells (e.g., tissue macrophages, osteoclasts, and microglia) during development (13, 14). Although greatly reduced numbers of monocyte lineage cells are present in osteopetrotic (op/op) mutant mice (i.e., mice that possess a mutant csf-1 gene encoding M-CSF) (13), the potential effects on angiogenesis and lymphangiogenesis have not been evaluated. Recent studies suggest that M-CSF plays a key role in the formation of high-density vessel networks and acts as an “angiogenic switch” in a mouse model of mammary tumors (15, 16). The inhibition of M-CSF by antisense oligonucleotides, small interfering RNAs, or blocking antibodies suppresses the growth of human mammary tumor xenografts in mice (17, 18). Clodronate liposome, a reagent for depleting macrophages, also inhibits tumor growth (19). However, the mechanisms underlying the role of M-CSF in tumor progression, particularly with regards to tumor lymphangiogenesis and tumor selectivity, have not been fully dissected.Osteosarcoma, the most common primary bone tumor, is defined as a malignant tumor derived from mesenchymal or stromal cells with highly metastatic capacities, particularly in the lung and liver (20). Multimodal treatment, often consisting of aggressive chemotherapy combined with radical surgical resection (e.g., limb amputation), has traditionally been the mainstay of osteosarcoma management; however, the prognosis of osteosarcoma patients has not improved significantly in recent years (20). Lack of an appropriate animal model for human osteosarcoma has hampered the development of an effective antiosteosarcoma therapy.In this study, we used op/op mice to demonstrate that M-CSF contributes to both vascular and lymphatic development. Furthermore, our data show that M-CSF is required for retinal pathological neovascularization but not for the maintenance of stable adult vasculature or lymphatics. Using a newly established mouse model of osteosarcoma, we show that M-CSF inhibition selectively suppresses tumor angiogenesis and lymphangiogenesis. In contrast to VEGF blockade, the interruption of M-CSF inhibition does not promote rapid tumor regrowth. These findings indicate that M-CSF–targeted therapy is an ideal strategy for the treatment of ocular neovascular diseases and cancer.