Napoleone Ferrara

University of California, San Diego, San Diego, California, United States

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Publications (300)3041.98 Total impact

  • Junji Moriya, Napoleone Ferrara
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    ABSTRACT: Diabetes is a risk factor for the development of cardiovascular diseases with impaired angiogenesis. We have previously shown that platelet-derived growth factor C (PDGF-C) and its receptor, PDGF receptor α (PDGFR-α) were downregulated in ischemic limbs of diabetic mice, although the underlying mechanisms remained elusive. Protein kinase C (PKC) is a family of serine/threonine kinases and is known to be involved in angiogenesis. The purpose of this study is to elucidate the mechanisms of how PDGF-C/PDGFR-α axis is impaired in diabetes. Human umbilical vein endothelial cells (HUVECs) and human cardiac microvascular endothelial cells (HMVECs) cultured in normoglycemic or hyperglycemic conditions were examined. We also examined the effects of PKC inhibition on the PDGF-C/PDGFR-α axis in endothelial cells exposed to hyperglycemia. Hyperglycemia inhibited proliferation and decreased viability of both HUVECs and HMVECs. Hyperglycemic endothelial cells exhibited decreased PDGFR-α expression both at messenger RNA (mRNA) and protein levels, while there was no significant change in expression of PDGF-C. We also found that expression of PKC-α, one of the PKC isoforms, was increased in hyperglycemic endothelial cells and that inhibition of PKC upregulated PDGFR-α expression in these cells. Phosphorylation of extracellular signal-regulated kinase (ERK) and Akt induced by PDGF-C was significantly attenuated in hyperglycemic endothelial cells, whereas inhibition of PKC effectively reversed these inhibitory effects. Moreover, inhibition of PKC also promoted angiogenesis induced by PDGF-C in hyperglycemic endothelial cells, which was not observed in vascular endothelial growth factor-A (VEGF-A)-induced angiogenesis. These findings suggest that downregulation of the PDGF-C/PDGFR-α axis is involved in impaired angiogenesis of hyperglycemia through upregulation of PKC. Targeting PKC to restore PDGF-C signaling might be a novel therapeutic strategy for the treatment of vascular complications in diabetes.
    Cardiovascular Diabetology 12/2015; 14(1). DOI:10.1186/s12933-015-0180-9 · 3.71 Impact Factor
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    ABSTRACT: Cancer pain is a debilitating disorder and a primary determinant of the poor quality of life. Here, we report a non-vascular role for ligands of the Vascular Endothelial Growth Factor (VEGF) family in cancer pain. Tumor-derived VEGF-A, PLGF-2, and VEGF-B augment pain sensitivity through selective activation of VEGF receptor 1 (VEGFR1) expressed in sensory neurons in human cancer and mouse models. Sensory-neuron-specific genetic deletion/silencing or local or systemic blockade of VEGFR1 prevented tumor-induced nerve remodeling and attenuated cancer pain in diverse mouse models in vivo. These findings identify a therapeutic potential for VEGFR1-modifying drugs in cancer pain and suggest a palliative effect for VEGF/VEGFR1-targeting anti-angiogenic tumor therapies. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.
    Cancer cell 06/2015; 27(6):780-796. DOI:10.1016/j.ccell.2015.04.017 · 23.89 Impact Factor
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    ABSTRACT: Because VEGFA has been implicated in follicle development, the objective of this study was to determine the effects of granulosa- and germ cell-specific VEGFA loss on ovarian morphogenesis, function, and female fertility. pDmrt1-Cre mice were mated to floxed VEGFA mice to develop granulosa-/germ cell-specific knockouts (pDmrt1-Cre;Vegfa-/-). The time from mating to first parturition was increased when pDmrt1-Cre;Vegfa-/- females were mated to control males (P = 0.0008) and tended to be longer for heterozygous females (P < 0.07). Litter size was reduced for pDmrt1-Cre;Vegfa-/- females (P < 0.007). The time between the first and second parturitions was also increased for heterozygous females (P < 0.04) and tended to be increased for pDmrt1-Cre;Vegfa-/- females (P < 0.07). pDmrt1-Cre;Vegfa-/- females had smaller ovaries (P < 0.04), reduced plasma estradiol (P < 0.007), fewer developing follicles (P < 0.008) and tended to have fewer corpora lutea (P < 0.08). Expression of Igf1r was reduced (P < 0.05); expression of Foxo3a tended to be increased (P < 0.06); and both Fshr (P < 0.1) and Sirt6 tended to be reduced (P < 0.06) in pDmrt1-Cre;Vegfa-/- ovaries. To compare VEGFA knockouts, we generated Amhr2-Cre;Vegfa-/- mice that required more time from mating to first parturition (P < 0.003) with variable ovarian size. Both lines had more apoptotic granulosa cells, and vascular staining did not appear different. Taken together these data indicate that the loss of all VEGFA isoforms in granulosa/germ cells (proangiogenic and antiangiogenic) causes subfertility by arresting follicular development, resulting in reduced ovulation rate and fewer pups per litter.
    PLoS ONE 02/2015; 10(2):e0116332. DOI:10.1371/journal.pone.0116332 · 3.53 Impact Factor
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    ABSTRACT: Blood vessels serve as key regulators of organogenesis by providing oxygen, nutrients and molecular signals. During limb development, programmed cell death (PCD) contributes to separation of the digits. Interestingly, prior to the onset of PCD, the autopod vasculature undergoes extensive patterning that results in high interdigital vascularity. Here, we show that in mice, the limb vasculature positively regulates interdigital PCD. In vivo, reduction in interdigital vessel number inhibited PCD, resulting in syndactyly, whereas an increment in vessel number and distribution resulted in elevation and expansion of PCD. Production of reactive oxygen species (ROS), toxic compounds that have been implicated in PCD, also depended on interdigital vascular patterning. Finally, ex vivo incubation of limbs in gradually decreasing oxygen levels led to a correlated reduction in both ROS production and interdigital PCD. The results support a role for oxygen in these processes and provide a mechanistic explanation for the counterintuitive positive role of the vasculature in PCD. In conclusion, we suggest a new role for vascular patterning during limb development in regulating interdigital PCD by ROS production. More broadly, we propose a double safety mechanism that restricts PCD to interdigital areas, as the genetic program of PCD provides the first layer and vascular patterning serves as the second. © 2015. Published by The Company of Biologists Ltd.
    Development 01/2015; 142(4). DOI:10.1242/dev.120279 · 6.27 Impact Factor
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    ABSTRACT: Although it has been documented that the nervous and the vascular systems share numerous analogies and are closely intermingled during development and pathological processes, interactions between the two systems are still poorly described. In the present study, we investigated whether Vascular Endothelial Growth Factor (VEGF), which is a key regulator of vascular development, also modulates neuronal developmental processes. We report that VEGF enhances the GABA/glycinergic but not glutamatergic synaptic activity in embryonic spinal motoneurons (MNs), without affecting MNs excitability. In response to VEGF, the frequency of these synaptic events but not their amplitude was increased. Blocking endogenous VEGF led to an opposite effect by decreasing frequency of synaptic events. We found that this effect occurred specifically at early developmental stages (E13.5 and E15.5) and vanished at the prenatal stage E17.5. Furthermore, VEGF was able to increase Vesicular Inhibitory Amino-Acid Transporter (VIAAT) density at the motoneuron membrane. Inhibition of single VEGF receptors did not modify electrophysiological parameters indicating receptor combinations or an alternative pathway. Altogether, our findings identify VEGF as a modulator of the neuronal activity during synapse formation and highlight a new ontogenic role for this angiogenic factor in the nervous system. © 2014 Wiley Periodicals, Inc. Develop Neurobiol, 2014.
    Developmental Neurobiology 11/2014; 74(11). DOI:10.1002/dneu.22187 · 4.19 Impact Factor
  • Cancer Research 10/2014; 74(20 Supplement):IA17-IA17. DOI:10.1158/1538-7445.PEDCAN-IA17 · 9.28 Impact Factor
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    ABSTRACT: The objectives of this study were to evaluate the relative binding and potencies of three inhibitors of vascular endothelial growth factor A (VEGF), used to treat neovascular age-related macular degeneration, and assess their relevance in the context of clinical outcome. Ranibizumab is a 48 kDa antigen binding fragment, which lacks a fragment crystallizable (Fc) region and is rapidly cleared from the systemic circulation. Aflibercept, a 110 kDa fusion protein and bevacizumab, a 150 kDa monoclonal antibody, each contain an Fc region. Binding affinities were determined using Biacore analysis. Competitive binding by sedimentation velocity analytical ultracentrifugation (SV-AUC) was used to support the binding affinities determined by Biacore of ranibizumab and aflibercept to VEGF. A bovine retinal microvascular endothelial cell (BREC) proliferation assay was used to measure potency. Biacore measurements were format dependent, especially for aflibercept, suggesting that biologically relevant, true affinities of recombinant VEGF (rhVEGF) and its inhibitors are yet to be determined. Despite this assay format dependency, ranibizumab appeared to be a very tight VEGF binder in all three formats. The results are also very comparable to those reported previously1-3. At equivalent molar ratios ranibizumab was able to displace aflibercept from preformed aflibercept-VEGF complexes in solution as assessed by SV-AUC, whereas aflibercept was not able to significantly displace ranibizumab from preformed ranibizumab-VEGF complexes. Ranibizumab, aflibercept, and bevacizumab showed dose-dependent inhibition of BREC proliferation induced by 6 ng/ml VEGF, with average IC50 values of 0.088 ± 0.032 nM, 0.090 ± 0.009 nM, and 0.500 ± 0.091 nM, respectively. Similar results were obtained with 3ng/mL VEGF. In summary Biacore studies and SV-AUC solution studies show that aflibercept does not bind with higher affinity than ranibizumab to VEGF as recently reported 4 and both inhibitors appeared to be equipotent with respect to their ability to inhibit VEGF function. 1. Ferrara, N.; Damico, L.; Shams, N.; Lowman, H.; Kim, R. Development of ranibizumab, an anti-vascular endothelial growth factor antigen binding fragment, as therapy for neovascular age-related macular degeneration. Retina 2006, 26, (8), 859-70. 2. Lowe, J.; Araujo, J.; Yang, J.; Reich, M.; Oldendorp, A.; Shiu, V.; Quarmby, V.; Lowman, H.; Lien, S.; Gaudreault, J.; Maia, M. Ranibizumab inhibits multiple forms of biologically active vascular endothelial growth factor in vitro and in vivo. Exp Eye Res 2007, 85, (4), 425-30. 3. Presta, L. G.; Chen, H.; O'Connor, S. J.; Chisholm, V.; Meng, Y. G.; Krummen, L.; Winkler, M.; Ferrara, N. Humanization of an anti-vascular endothelial growth factor monoclonal antibody for the therapy of solid tumors and other disorders. Cancer Res 1997, 57, (20), 4593-9. 4.Papadopoulos, N.; Martin, J.; Ruan, Q.; Rafique, A.; Rosconi, M. P.; Shi, E.; Pyles, E. A.; Yancopoulos, G. D.; Stahl, N.; Wiegand, S. J. Binding and neutralization of vascular endothelial growth factor (VEGF) and related ligands by VEGF Trap, ranibizumab and bevacizumab. Angiogenesis 2012, 15, (2), 171-85.
    Molecular Pharmaceutics 08/2014; 11(10). DOI:10.1021/mp500160v · 4.79 Impact Factor
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    ABSTRACT: Cardiac and craniofacial developmental programs are intricately linked during early embryogenesis, which is also reflected by a high frequency of birth defects affecting both regions. The molecular nature of the crosstalk between mesoderm and neural crest progenitors and the involvement of endothelial cells within the cardio-craniofacial field are largely unclear. Here we show in the mouse that genetic ablation of vascular endothelial growth factor receptor 2 (Flk1) in the mesoderm results in early embryonic lethality, severe deformation of the cardio-craniofacial field, lack of endothelial cells and a poorly formed vascular system. We provide evidence that endothelial cells are required for migration and survival of cranial neural crest cells and consequently for the deployment of second heart field progenitors into the cardiac outflow tract. Insights into the molecular mechanisms reveal marked reduction in Transforming growth factor beta 1 (Tgfb1) along with changes in the extracellular matrix (ECM) composition. Our collective findings in both mouse and avian models suggest that endothelial cells coordinate cardio-craniofacial morphogenesis, in part via a conserved signaling circuit regulating ECM remodeling by Tgfb1.
    07/2014; 3(8). DOI:10.1242/bio.20148078
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    ABSTRACT: Clinical Relevance Angiogenesis and arteriogenesis after ischemia are attenuated in most diabetic patients, although the precise mechanisms remain unclear. Platelet-derived growth factors (PDGFs) have a variety of functions on many cell types, and PDGF-C stimulates angiogenesis and revascularizes ischemic tissues. This study indicates the role for PDGF-C as a critical regulator of impaired angiogenesis of diabetes and suggests that PDGF-C might be a novel target for the treatment of ischemic cardiovascular diseases in diabetes.
    Journal of Vascular Surgery 05/2014; 59(5):1402–1409.e4. DOI:10.1016/j.jvs.2013.04.053 · 2.98 Impact Factor
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    Junji Moriya, Napoleone Ferrara
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    ABSTRACT: Inhibition of the vascular endothelial growth factor (VEGF)-VEGF receptor 2 (VEGFR2) signaling axis may play a role in endothelial dysfunction and serious vascular complications associated with diabetes. In this issue, Warren et al. identified a ligand-independent, receptor tyrosine kinase-independent VEGFR2 signaling pathway that is responsible for impaired responses to VEGF in diabetic endothelial cells. Reactive oxygen species (ROS) resulting from the hyperglycemic status promoted activation and subsequent degradation of VEGFR2 in a ligand-independent manner. Consequently, VEGF-VEGFR2 signaling was inhibited due to depletion of VEGFR2 at the cell surface. Activation of this ligand-independent, ROS-induced VEGFR2 signaling was mediated by the Src family of kinases and occurred in the Golgi compartment in the endothelial cells. Blocking ROS production by antioxidants effectively reversed VEGFR2 deficiency at the cell surface in hyperglycemia. These findings suggest that ROS-induced VEGFR2 signaling might be a promising new target for the treatment of vascular diseases in diabetes.
    Science Signaling 01/2014; 7(307):pe1. DOI:10.1126/scisignal.2004996 · 7.65 Impact Factor
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    ABSTRACT: Angiogenesis defines the process in which new vessels grow from existing vessels. Using the mouse retina as a model system, we show that cysteine-rich motor neuron 1 (Crim1), a type I transmembrane protein, is highly expressed in angiogenic endothelial cells. Conditional deletion of the Crim1 gene in vascular endothelial cells (VECs) causes delayed vessel expansion and reduced vessel density. Based on known Vegfa binding by Crim1 and Crim1 expression in retinal vasculature, where angiogenesis is known to be Vegfa dependent, we tested the hypothesis that Crim1 is involved in the regulation of Vegfa signaling. Consistent with this hypothesis, we showed that VEC-specific conditional compound heterozygotes for Crim1 and Vegfa exhibit a phenotype that is more severe than each single heterozygote and indistinguishable from that of the conditional homozygotes. We further showed that human CRIM1 knockdown in cultured VECs results in diminished phosphorylation of VEGFR2, but only when VECs are required to rely on an autocrine source of VEGFA. The effect of CRIM1 knockdown on reducing VEGFR2 phosphorylation was enhanced when VEGFA was also knocked down. Finally, an anti-VEGFA antibody did not enhance the effect of CRIM1 knockdown in reducing VEGFR2 phosphorylation caused by autocrine signaling, but VEGFR2 phosphorylation was completely suppressed by SU5416, a small-molecule VEGFR2 kinase inhibitor. These data are consistent with a model in which Crim1 enhances the autocrine signaling activity of Vegfa in VECs at least in part via Vegfr2.
    Development 12/2013; 141(2). DOI:10.1242/dev.097949 · 6.27 Impact Factor
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    ABSTRACT: Vascular endothelial growth factor A (VEGFA) isoform treatment has been demonstrated to alter spermatogonial stem cell (SSC) homeostasis. Therefore, we generated pDmrt1-Cre;Vegfa(-/-) (KO) mice by crossing pDmrt1-Cre mice to floxed Vegfa mice to test whether loss of all VEGFA isoforms in Sertoli and germ cells would impair spermatogenesis. When first mated, KO males took 14 days longer to get control females pregnant (P < 0.02) and tended to take longer for all subsequent parturition intervals (9 days; P < 0.07). Heterozygous males sired fewer pups per litter (P < 0.03) and after the first litter took 10 days longer (P < 0.05) to impregnate females suggesting a more progressive loss of fertility. Reproductive organs were collected from 6-month-old male mice. There were fewer sperm per tubule in the corpus epididymides (P < 0.001) and fewer ZBTB16-stained undifferentiated spermatogonia (P < 0.003) in the testes of KO males. Testicular mRNA abundance for Bcl2 (P < 0.02), Bcl2:Bax (P < 0.02), Neurog3 (P < 0.007), and Ret was greater (P = 0.0005), tended to be greater for Sin3a and tended to be reduced for total Foxo1 (P < 0.07) in KO males. Immunofluorescence for CD31 and VE-Cadherin showed no differences in testis vasculature; however, CD31-positive staining was evident in undifferentiated spermatogonia only in KO testes. Therefore, loss of VEGFA isoforms in Sertoli and germ cells alters genes necessary for long-term maintenance of undifferentiated spermatogonia, ultimately reducing sperm numbers and resulting in subfertility.
    Endocrinology 10/2013; 154(12). DOI:10.1210/en.2013-1363 · 4.64 Impact Factor
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    ABSTRACT: The features and regulation of uterine angiogenesis and vascular remodeling during pregnancy are poorly defined. Here we show that dynamic and variable decidual angiogenesis (sprouting, intussusception, and networking), and active vigorous vascular remodeling such as enlargement and elongation of "vascular sinus folding" (VSF) and mural cell drop-out occur distinctly in a spatiotemporal manner in the rapidly growing mouse uterus during early pregnancy-just after implantation but before placentation. Decidual angiogenesis is mainly regulated through VEGF-A secreted from the progesterone receptor (PR)-expressing decidual stromal cells which are largely distributed in the anti-mesometrial region. In comparison, P4 -PR-regulated VEGF-A-VEGFR2 signaling, ligand-independent VEGFR3 signaling, and uterine natural killer cells positively and coordinately regulate enlargement and elongation of VSF. During the postpartum period, Tie2 signaling could be involved in vascular maturation at the endometrium in a ligand-independent manner, with marked reduction of VEGF-A, VEGFR2 and PR expressions. Overall, we show that two key vascular growth factor receptors - VEGFR2 and Tie2 - strikingly but differentially regulate decidual angiogenesis and vascular remodeling in rapidly growing and regressing uteri in an organotypic manner.
    EMBO Molecular Medicine 09/2013; 5(9). DOI:10.1002/emmm.201302618 · 8.25 Impact Factor
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    ABSTRACT: Although angiogenesis inhibitors have provided substantial clinical benefit as cancer therapeutics, their use is limited by resistance to their therapeutic effects. While ample evidence indicates that such resistance can be influenced by the tumor microenvironment, the underlying mechanisms remain incompletely understood. Here, we have uncovered a paracrine signaling network between the adaptive and innate immune systems that is associated with resistance in multiple tumor models: lymphoma, lung and colon. Tumor-infiltrating T helper type 17 (TH17) cells and interleukin-17 (IL-17) induced the expression of granulocyte colony-stimulating factor (G-CSF) through nuclear factor κB (NF-κB) and extracellular-related kinase (ERK) signaling, leading to immature myeloid-cell mobilization and recruitment into the tumor microenvironment. The occurrence of TH17 cells and Bv8-positive granulocytes was also observed in clinical tumor specimens. Tumors resistant to treatment with antibodies to VEGF were rendered sensitive in IL-17 receptor (IL-17R)-knockout hosts deficient in TH17 effector function. Furthermore, pharmacological blockade of TH17 cell function sensitized resistant tumors to therapy with antibodies to VEGF. These findings indicate that IL-17 promotes tumor resistance to VEGF inhibition, suggesting that immunomodulatory strategies could improve the efficacy of anti-angiogenic therapy.
    Nature medicine 08/2013; DOI:10.1038/nm.3291 · 28.05 Impact Factor
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    ABSTRACT: The phosphatidylinositol 3-kinase (PI3K) pathway is a central mediator of vascular endothelial growth factor (VEGF)-driven angiogenesis. The discovery of small molecule inhibitors that selectively target PI3K or PI3K and mammalian target of rapamycin (mTOR) provides an opportunity to pharmacologically determine the contribution of these key signaling nodes in VEGF-A-driven tumor angiogenesis in vivo. This study used an array of micro-vascular imaging techniques to monitor the antivascular effects of selective class I PI3K, mTOR, or dual PI3K/mTOR inhibitors in colorectal and prostate cancer xenograft models. Micro-computed tomography (micro-CT) angiography, dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI), vessel size index (VSI) MRI, and DCE ultrasound (DCE-U/S) were employed to quantitatively evaluate the vascular (structural and physiological) response to these inhibitors. GDC-0980, a dual PI3K/mTOR inhibitor, was found to reduce micro-CT angiography vascular density, while VSI MRI demonstrated a significant reduction in vessel density and an increase in mean vessel size, consistent with a loss of small functional vessels and a substantial antivascular response. DCE-MRI showed that GDC-0980 produces a strong functional response by decreasing the vascular permeability/perfusion-related parameter, K (trans). Interestingly, comparable antivascular effects were observed for both GDC-980 and GNE-490 (a selective class I PI3K inhibitor). In addition, mTOR-selective inhibitors did not affect vascular density, suggesting that PI3K inhibition is sufficient to generate structural changes, characteristic of a robust antivascular response. This study supports the use of noninvasive microvascular imaging techniques (DCE-MRI, VSI MRI, DCE-U/S) as pharmacodynamic assays to quantitatively measure the activity of PI3K and dual PI3K/mTOR inhibitors in vivo.
    Neoplasia (New York, N.Y.) 07/2013; 15(7):694-711. DOI:10.1593/neo.13470 · 5.40 Impact Factor
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    ABSTRACT: Our goal was to develop a potent humanized antibody against mouse/human CXCL12. This report summarized its in vitro and in vivo activities. Cell surface binding and cell migration assays were used to select neutralizing hamster antibodies, followed by testing in several animal models. Mab 30D8 was selected for humanization based on its in vitro and in vivo activities. 30D8, a hamster antibody against mouse and human CXCL12α, CXCL12β and CXCL12γ, was shown to dose-dependently block CXCL12αbinding to CXCR4 and CXCR7, and CXCL12α-induced Jurkat cell migration in vitro. Single agent activity in both primary tumor growth and/or lung metastasis was demonstrated in several xenograft and orthotopic models. 30D8 alone significantly ameliorated arthritis in a mouse collagen-induced arthritis model (CIA). Combination with a TNFα antagonist was additive. In addition, 30D8 inhibited 50% of laser-induced choroidal neovascularization (CNV) in mice. Humanized 30D8 (hu30D8) showed similar in vitro and in vivo activities as the parental hamster antibody. A crystal structure of the hu30D8 Fab/ CXCL12α complex in combination with mutational analysis revealed a "hot spot" around residues Asn44/ Asn45 of CXCL12α and part of the RFFESH region required for CXCL12α binding to CXCR4 and CXCR7. Finally, hu30D8 exhibited fast clearance in cynomolgus monkeys but not in rats. CXCL12 is an attractive target for treatment of cancer and inflammation-related diseases; hu30D8 is suitable to test this hypothesis in humans.
    Clinical Cancer Research 06/2013; 19(16). DOI:10.1158/1078-0432.CCR-13-0943 · 8.19 Impact Factor
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    ABSTRACT: PURPOSE: The aim of this study was to identify conserved pharmacodynamic and potential predictive biomarkers of response to anti-VEGF therapy using gene expression profiling in preclinical tumor models and in patients. EXPERIMENTAL DESIGN: Surrogate markers of VEGF inhibition (VEGF dependent genes or VDV) were identified by profiling gene expression changes induced in response to VEGF blockade in preclinical tumor models and in human biopsies from patients treated with anti-VEGF monoclonal antibodies. The potential value of VDV genes as candidate predictive biomarkers was tested by correlating high or low VDV gene expression levels in pre-treatment clinical samples with the subsequent clinical efficacy of bevacizumab (anti-VEGF) containing therapy. RESULTS: We show that VEGF-dependent vasculature (VDV) genes, including direct and more distal VEGF downstream endothelial targets, enable detection of VEGF signaling inhibition in mouse tumor models and human tumor biopsies. Retrospective analyses of clinical trial data indicates that patients with higher VDV expression in pre-treatment tumor samples exhibited improved clinical outcome when treated with bevacizumab containing therapies. CONCLUSIONS: In this work we identified surrogate markers (VDV genes) for in vivo VEGF signaling in tumors and showed clinical data supporting a correlation between pre-treatment VEGF bioactivity and the subsequent efficacy of anti-VEGF therapy. We propose that VDV genes are candidate biomarkers with the potential to aid the selection of novel indications as well as patients likely to respond to anti-VEGF therapy. The data presented here define a diagnostic biomarker hypothesis based on translational research that warrants further evaluation in additional retrospective and prospective trials.
    Clinical Cancer Research 05/2013; 19(13). DOI:10.1158/1078-0432.CCR-12-3635 · 8.19 Impact Factor
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    ABSTRACT: The vascular endothelial growth factor (VEGF) signaling pathway plays a pivotal role in normal development and also represents a major therapeutic target for tumors and intraocular neovascular disorders. The VEGF receptor tyrosine kinases promote angiogenesis by phosphorylating downstream proteins in endothelial cells. We applied a large-scale proteomic approach to define the VEGF-regulated phosphoproteome and its temporal dynamics in human umbilical vein endothelial cells and then used siRNA (small interfering RNA) screens to investigate the function of a subset of these phosphorylated proteins in VEGF responses. The PI3K (phosphatidylinositol 3-kinase)-mTORC2 (mammalian target of rapamycin complex 2) axis emerged as central in activating VEGF-regulated phosphorylation and increasing endothelial cell viability by suppressing the activity of the transcription factor FoxO1 (forkhead box protein O1), an effect that limited cellular apoptosis and feedback activation of receptor tyrosine kinases. This FoxO1-mediated feedback loop not only reduced the effectiveness of mTOR inhibitors at decreasing protein phosphorylation and cell survival but also rendered cells more susceptible to PI3K inhibition. Collectively, our study provides a global and dynamic view of VEGF-regulated phosphorylation events and implicates the mTORC2-FoxO1 axis in VEGF receptor signaling and reprogramming of receptor tyrosine kinases in human endothelial cells.
    Science Signaling 04/2013; 6(271):ra25. DOI:10.1126/scisignal.2003572 · 7.65 Impact Factor
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    ABSTRACT: Granulocyte-colony stimulating factor (G-CSF) promotes mobilization of CD11b(+)Gr1(+) myeloid cells and has been implicated in resistance to anti-VEGF therapy in mouse models. High G-CSF production has been associated with a poor prognosis in cancer patients. Here we show that activation of the RAS/MEK/ERK pathway regulates G-CSF expression through the Ets transcription factor. Several growth factors induced G-CSF expression by a MEK-dependent mechanism. Inhibition of G-CSF release with a MEK inhibitor markedly reduced G-CSF production in vitro and synergized with anti-VEGF antibodies to reduce CD11b(+)Ly6G(+) neutrophil mobilization and tumor growth and led to increased survival in animal models of cancer, including a genetically engineered mouse model of pancreatic adenocarcinoma. Analysis of biopsies from pancreatic cancer patients revealed increased phospho-MEK, G-CSF, and Ets expression and enhanced neutrophil recruitment compared with normal pancreata. These results provide insights into G-CSF regulation and on the mechanism of action of MEK inhibitors and point to unique anticancer strategies.
    Proceedings of the National Academy of Sciences 03/2013; 110(15). DOI:10.1073/pnas.1303302110 · 9.81 Impact Factor

Publication Stats

77k Citations
3,041.98 Total Impact Points


  • 2013–2014
    • University of California, San Diego
      • Department of Pathology
      San Diego, California, United States
  • 2011–2013
    • Cincinnati Children's Hospital Medical Center
      Cincinnati, Ohio, United States
    • Università degli Studi di Bari Aldo Moro
      Bari, Apulia, Italy
  • 2012
    • Novartis Institutes for BioMedical Research
      Cambridge, Massachusetts, United States
    • Weizmann Institute of Science
      • Department of Molecular Genetics
  • 1998–2012
    • Genentech
      • Department of Molecular Oncology
      San Francisco, California, United States
  • 2006
    • University of Toronto
      • Laboratory Medicine Program
      Toronto, Ontario, Canada
  • 2005
    • A.C.O. San Filippo Neri
      Roma, Latium, Italy
    • Harvard Medical School
      • Department of Cell Biology
      Boston, MA, United States
  • 2004
    • Unité Inserm U1077
      Caen, Lower Normandy, France
    • Harvard University
      • Department of Molecular and Cell Biology
      Cambridge, Massachusetts, United States
  • 2003
    • Samuel Lunenfeld Research Institute
      Toronto, Ontario, Canada
  • 1994–2003
    • University of California, San Francisco
      • • Center for Reproductive Sciences
      • • Department of Surgery
      • • Department of Obstetrics, Gynecology and Reproductive Sciences
      San Francisco, California, United States
  • 1994–1997
    • St. Elizabeth's Medical Center
      Boston, Massachusetts, United States
  • 1994–1996
    • Tufts University
      • Department of Medicine
      Georgia, United States
  • 1988
    • Cancer Research Institute
      New York, New York, United States