Advertisement
help button home button The Oncologist
HOME HELP CONTACT US SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow eLetters: Submit a response to this article
Right arrow Purchase Article
Right arrow View Shopping Cart
Right arrow Alert me when this article is cited
Right arrow Alert me when eLetters are posted
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow E-mail this article link to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow Reprints/Permissions
Citing Articles
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Conti, C. J.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Conti, C. J.
The Oncologist, Vol. 7, Suppl 3, 4-11, August 2002
© 2002 AlphaMed Press

Vascular Endothelial Growth Factor: Regulation in the Mouse Skin Carcinogenesis Model and Use in Antiangiogenesis Cancer Therapy

Claudio J. Conti

Department of Carcinogenesis, University of Texas M.D. Anderson Cancer Center, Science Park—Research Division, Smithville, Texas, USA

Correspondence: Claudio J. Conti, D.V.M., Ph.D., M. D. Anderson Cancer Center, Park Road 1-C, Smithville, Texas 78957, USA. Telephone: 512-237-9428; Fax: (512) 237-9421; e-mail: sa83125{at}odin.mdacc.tmc.edu


   LEARNING OBJECTIVES
Top
 Learning Objectives
Abstract
 Introduction
 Mechanisms of Tumor...
 Angiogenesis Stimuli
 Targeting Angiogenesis in the...
 Conclusions
 References
 
After completing this course, the reader will be able to:

  1. Describe mechanisms of tumor neovascularization.
  2. Identify possible targets for cancer therapy in the biologic pathways that control angiogenesis.
  3. Identify some of the drugs that are presently used in preclinical or clinical trials, as well as their targets.

Access and take the CME test online and receive one hour of AMA PRA category 1 credit at CME.TheOncologist.com


   ABSTRACT
Top
 Learning Objectives
 Abstract
Introduction
 Mechanisms of Tumor...
 Angiogenesis Stimuli
 Targeting Angiogenesis in the...
 Conclusions
 References
 
Of the various mechanisms responsible for tumor neovascularization, the angiogenesis process, in particular vascular endothelial growth factor (VEGF), is described here as a target for cancer therapy. While hypoxia is a trigger of tumor angiogenesis, various alterations in oncogenes and tumor suppressor genes also have been reported to induce VEGF expression in tumors. The regulation of VEGF has been investigated in chemically induced mouse squamous cell carcinoma of the skin. In this cancer model, VEGF expression appears to be dependent on ras oncogene activation as well as the epidermal growth factor receptor. Thus, in addition to VEGF, oncogene signaling pathways may be relevant targets in antiangiogenesis cancer therapies.

The central role of VEGF in angiogenesis has led to the development of several drugs targeting the pathway of this growth factor. The present paper provides an overview of these drugs and their stage of development. In the near future, clinical trials using anti-VEGF drugs and other antiangiogenic agents, such as endostatin and angiostatin, will yield valuable information about their potential for cancer therapy.

Key Words. Tumor angiogenesis • Vascular endothelial growth factor • ras oncogene • Squamous skin carcinoma • Therapy-related cancer • Clinical trials


   INTRODUCTION
Top
 Learning Objectives
 Abstract
 Introduction
Mechanisms of Tumor...
 Angiogenesis Stimuli
 Targeting Angiogenesis in the...
 Conclusions
 References
 
Normal tissue vasculature cannot support tumor growth beyond 2 mm in diameter [1]. Formation of new vessels is needed for tumor development as well as invasion and metastasis [1]. Tumor neovascularization is an interesting target for chemotherapy because, in general, adult endothelial cells are quiescent except for the menstrual cycle [2]. Furthermore, this process is regulated by cell-surface receptors and extracellular factors, which means that drugs can interfere with neovascularization without the need for cellular uptake. The factors regulating neovascularization, e.g., vascular endothelial growth factor (VEGF) and fibroblast growth factor (FGF) [3], are highly specific for endothelial cells and thus chemotherapeutics directed at these factors are expected to have few side effects.


   MECHANISMS OF TUMOR NEOVASCULARIZATION
Top
 Learning Objectives
 Abstract
 Introduction
 Mechanisms of Tumor...
Angiogenesis Stimuli
 Targeting Angiogenesis in the...
 Conclusions
 References
 
Neovascularization of tumors can occur via different mechanisms, depending on the tumor type, the stage of tumor development, and the anatomic localization. These different mechanisms, as well as their potential as targets for cancer therapy, are described in this section.

Vessel Co-Option
According to this concept, tumors take up existing blood vessels from surrounding normal tissues and use these for their initial growth. Once the tumor reaches a critical size, these vessels will be insufficient to support tumor growth, and anoxia will ensue. This then will induce true angiogenesis with the generation of de novo vessels. Vessel co-option has been demonstrated in experimental models where tumor cells are injected [4]. This mechanism may apply to lymphomas where tumor cells can move and develop along vessels. Vessel co-option is probably irrelevant in the great majority of solid tumors, in particular carcinomas, where cells grow in a mass toward the stroma and cannot move individually. Thus, vessel co-option is not a likely target for cancer therapy.

Vasculogenesis
Vasculogenesis is the mechanism of blood vessel formation in the embryo: hematopoietic precursor cells line blood vessels and develop into blood islands [5]. Centrally located cells develop into blood cells, whereas peripheral cells develop into endothelial cells [5]. Recent studies have shown a similar process in tumors in experimental animal models, in which precursor cells from the bone marrow circulate, aggregate, and mesh into new blood vessels [6]. The importance of postnatal vasculogenesis to human tumors is unknown and therefore its relevance as a target for cancer therapy remains undecided.

Angiogenesis
This process is probably the essential mechanism for tumor neovascularization, and drugs inhibiting angiogenesis are currently being evaluated in clinical trials. Angiogenesis consists of several steps [7]. Endothelial cells that form the wall of existing small blood vessels are activated, degrade the extracellular matrix, migrate through the matrix, and proliferate. The new endothelial cells organize into hollow tubes, which ultimately anastomose to form new capillaries.


   ANGIOGENESIS STIMULI
Top
 Learning Objectives
 Abstract
 Introduction
 Mechanisms of Tumor...
 Angiogenesis Stimuli
Targeting Angiogenesis in the...
 Conclusions
 References
 
Angiogenesis is induced when the balance between proangiogenic and antiangiogenic factors is disturbed. In tumors, this imbalance may be caused by either hypoxia (low oxygen tension) or genetic alterations that activate oncogenes and/or inactivate tumor suppressor genes [8] (Fig. 1Go).



View larger version (20K):
[in this window]
[in a new window]
  		Figure 1. Schematic representation of the triggering mechanisms in tumor angiogenesis: inactivated tumor suppressor genes/activated oncogenes versus hypoxia. In cell culture, these mechanisms can act independently. In human tumors, both mechanisms may modulate angiogenesis [8].

 
Hypoxia
Using oxygen that diffuses from existing vessels, a tumor grows until oxygen demand exceeds supply. The ensuing hypoxia induces tumor cells to generate, via both increased transcription (regulated by the hypoxia-induced transcription factor HIF1a) and increased mRNA stabilization, a series of proangiogenic factors that function as highly specific growth factors/survival factors for endothelial cells [9]. Among these factors are VEGF, FGF, transforming growth factor ß, and tumor necrosis factor {alpha}. VEGF and FGF are considered the most important mediators in tumor angiogenesis. VEGF (VEGF-A) induces proliferation of vascular endothelial cells [10–13], promotes survival of endothelial cells in newly formed vessels [14–17], and increases blood vessel permeability [18, 19]. Through increased blood vessel permeability, VEGF allows creation of an extravascular fibrin gel that supports endothelial cell growth [20, 21]. This permits increased tumor growth resulting in hypoxia, which in turn leads to VEGF production (Fig. 1Go).

Oncogenes and Tumor Suppressor Genes
In many tumors, angiogenesis is induced not by hypoxia, but by genetic alterations. The loss of function of tumor suppressor genes such as VHL, p53, and p16INK4a, or the activation of oncogenes including ras, raf, HER2/erbB2 (neu), and src, results in increased expression and/or secretion of VEGF [22–27] (Fig. 1Go). The role of mutant H-ras in skin tumor angiogenesis has been studied in our laboratory in collaboration with Drs. José Jorcano, Fernando Larcher, Llanos Casanova et al. (Centro de Investigaciones Energéticas, Medioambientales y Tecnológicas [CIEMAT], Madrid, Spain) and will be described in more detail in the next section.

ras Induction of Angiogenesis in Skin Tumors
The mouse skin carcinogenesis model, in which mice are treated with the genotoxic 7,12-dimethylbenz[a]anthracene followed by multiple topical applications of 12-O-tetradecanoylphorbol-13-acetate as a promoter, allows observation of the sequential stages of tumor development [28, 29]. Hyperplastic skin gives rise to papillomas, which are benign exophytic lesions. These lesions eventually transform into squamous cell carcinoma (SCC), which is practically identical to human head and neck SCC and human skin SCC. The initiating mutation in this model is an A->T transversion in codon 61 of the H-ras gene [30, 31]. Larcher et al. showed an upregulation of VEGF mRNA and protein in papillomas and a further increase in SCC [22]. They confirmed mutant H-ras involvement in the VEGF upregulation by showing that transduction of keratinocytes with this activated oncogene in normal oxygen conditions markedly induced VEGF mRNA levels [22]. This finding is consistent with other reports demonstrating ras-dependence of VEGF induction in human colon carcinoma cells, H-ras transformed rat intestinal epithelial cells, and NIH 3T3 cells stably transfected with v-H-ras or v-raf [32, 33]. Furthermore, increased expression of the epidermal growth factor receptor (EGFR) in SCC has been found in association with H-ras activation [34–36].

In collaboration with our laboratory, Dr. Jorcano and colleagues studied the role of EGFR in mutant H-ras-induced angiogenesis. An ex vivo experiment with tumor cells carrying mutant H-ras and transfected with dominant-negative (DN) EGFR showed that DN EGFR cells formed smaller tumors in nude mice compared with control cells [37]. In transgenic mice that harbor DN EGFR under control of the keratin 5 promoter, chemical induction of skin tumors resulted in small papillomas containing narrow blood vessels, whereas the papillomas from control animals displayed large vessels [37]. These experiments demonstrate that a functional EGFR is required for mutant H-ras induction of angiogenesis in skin tumors; because mutant H-ras is a downstream effector of EGFR, this may appear paradoxical. However, a recent report suggested cell type– specific modes of VEGF regulation by the mutant ras oncogene [38]. The requirement for a functional EGFR for angiogenesis induction by mutant H-ras is interesting in light of ongoing clinical trials targeting the EGFR, which thus may target angiogenesis indirectly.

Analysis of vasculature in the sequential skin tumor stages by immunohistochemical staining of the endothelial cell marker CD31 [39] has shown that blood vessel density increases considerably in very early papillomas [40]. In more advanced papillomas as well as SCC, blood vessel density stabilizes but the size of the blood vessels increases [40]. Large blood vessels also have been observed following hyperproliferative stimuli in the dermis of transgenic mice that express mouse VEGF120 (1 of 3 VEGF isoforms that is not bound to extracellular proteins, but is freely diffusible) under control of the keratin 6 promoter [41]. Endothelial cell proliferation is detectable in the vessels present in papillomas and SCC, indicating that these cells are proliferating. Although various markers have been reported that are expressed differentially in proliferating versus quiescent endothelial cells in vivo [42–53], no markers expressed exclusively in tumor endothelium have been detected. In hyperplastic skin, VEGF production by keratinocytes has been demonstrated by immunohistochemistry; VEGF production increases in papillomas and stabilizes in more advanced stages (Franco and Conti, unpublished results). We believe that the increase in VEGF in SCC, which was previously detected by Western blot analysis [41], reflects the tumor cell mass rather than the amount produced by individual cells. Interestingly, immunohistochemical detection of VEGF in SCC shows no association with anoxic areas, confirming that angiogenesis in these tumors is not induced by hypoxia/anoxia. Demonstrating proof of principle by deletion of VEGF function in vivo has not been feasible. VEGF-knockout mice are lethal [54–56] and only recently a few conditional VEGF-knockout mice, with abrogated VEGF expression in the epidermis, were obtained in our laboratory.


   TARGETING ANGIOGENESIS IN THE CLINIC
Top
 Learning Objectives
 Abstract
 Introduction
 Mechanisms of Tumor...
 Angiogenesis Stimuli
 Targeting Angiogenesis in the...
Conclusions
 References
 
VEGF
The central role of VEGF in angiogenesis has been clearly demonstrated [57 and references therein]. Consequently, the use of an anti-VEGF antibody and small molecule tyrosine kinase inhibitors to interfere with VEGF signaling is being investigated in clinical trials in cancer patients. Table 1Go shows a list of drugs that are being developed to target VEGF function [58–87]. These agents, like other antiangiogenic drugs, are cytostatic to the tumor cells. Therefore, several of the anti-VEGF drugs that currently are being tested in ongoing clinical trials as single agents, also are being tested in combination with conventional cytotoxic chemotherapeutics, such as cisplatin, carboplatin, gemcitabine, fluorouracil, paclitaxel, and docetaxel (see also: http://www.cancer.gov/search/clinical_trials/).


View this table:
[in this window]
[in a new window]
  		Table 1. Drugs that interfere with the VEGF pathway
 
Antiangiogenic Factors
Proteolysis is an important component of angiogenesis. It is involved in degradation of the basement membrane and the extracellular matrix, release of growth factors bound to the matrix (including VEGF and FGF, which are bound to extracellular proteins), and remodeling of the matrix. Finally, Folkman and colleagues showed that tumors themselves secrete antiangiogenic factors that are generated upon proteolysis of large precursor proteins [88]. The endogenous antiangiogenic factors angiostatin and endostatin are proteolytic fragments of plasminogen and collagen VIII, respectively [88, 89]. Angiostatin and endostatin inhibit proliferation and migration of endothelial cells, and induce endothelial cell apoptosis [88, 90–94]. Although the mechanism of action of endostatin is still unclear, the actions of angiostatin are most likely mediated via a cell surface ATP synthase [90]. Both antiangiogenic factors currently are undergoing testing in phase I clinical trials.


   CONCLUSIONS
Top
 Learning Objectives
 Abstract
 Introduction
 Mechanisms of Tumor...
 Angiogenesis Stimuli
 Targeting Angiogenesis in the...
 Conclusions
References
 
The relevance of angiogenesis to tumor development is unquestioned and a number of clinical trials are ongoing, testing agents that directly target angiogenesis, such as anti-VEGF drugs and endogenous antiangiogenic factors.

The angiogenesis studies in the mouse skin carcinogenesis model described in this article suggest that inhibition of oncogene signal transduction pathways may indirectly inhibit angiogenesis. Indeed, signal transduction inhibitors are thought to indirectly downregulate VEGF expression in tumor cells and thus block angiogenesis [95]. Angiogenesis also may be targeted indirectly in other therapies. The cyclin-dependent kinase inhibitor flavopiridol has been shown to decrease VEGF production [96], while conventional chemotherapy has been shown to have antiangiogenic effects [95, 97].

In the near future, the outcome of ongoing clinical trials will give us more insights into the potential of antiangiogenic approaches to treat cancer.


   ACKNOWLEDGMENT
Top
 Learning Objectives
 Abstract
 Introduction
 Mechanisms of Tumor...
 Angiogenesis Stimuli
 Targeting Angiogenesis in the...
 Conclusions
 References
 
The author acknowledges Anita Frijhoff (University of Texas, M.D. Anderson Cancer Center) as technical writer for editing the manuscript as well as recompiling literature and clinical trial data for this article. He also recognizes Margaret Hayes for transcription and Drs. Adrian Senderowicz (NIDCR, NIH) and Marcela Franco (University of Texas, M.D. Anderson Cancer Center) for helpful discussion.


   REFERENCES
Top
 Learning Objectives
 Abstract
 Introduction
 Mechanisms of Tumor...
 Angiogenesis Stimuli
 Targeting Angiogenesis in the...
 Conclusions
 References
 

  1. Folkman J. Tumor angiogenesis: therapeutic implications. N Engl J Med 1971;285:1182–1186.
  2. Hyder SM, Stancel GM. Regulation of angiogenic growth factors in the female reproductive tract by estrogens and progestins. Mol Endocrinol 1999;13:806–811.[Free Full Text]
  3. Cross MJ, Claesson-Welsh L. FGF and VEGF function in angiogenesis: signalling pathways, biological responses and therapeutic inhibition. Trends Pharmacol Sci 2001;22:201–207.[CrossRef][Medline]
  4. Holash J, Maisonpierre PC, Compton D et al. Vessel cooption, regression, and growth in tumors mediated by angiopoietins and VEGF. Science 1999;284:1994–1998.[Abstract/Free Full Text]
  5. Risau W, Flamme I. Vasculogenesis. Annu Rev Cell Dev Biol 1995;11:73–91.[CrossRef][Medline]
  6. Asahara T, Masuda H, Takahashi T et al. Bone marrow origin of endothelial progenitor cells responsible for postnatal vasculogenesis in physiological and pathological neovascularization. Circ Res 1999;85:221–228.[Abstract/Free Full Text]
  7. Risau W. Mechanisms of angiogenesis. Nature 1997; 386:671–674.[CrossRef][Medline]
  8. Rak J, Yu JL, Klement G et al. Oncogenes and angiogenesis: signaling three-dimensional tumor growth. J Investig Dermatol Symp Proc 2000;5:24–33.[CrossRef][Medline]
  9. Dachs GU, Tozer GM. Hypoxia modulated gene expression: angiogenesis, metastasis and therapeutic exploitation. Eur J Cancer 2000;36:1649–1660.
  10. Ferrara N, Davis-Smyth T. The biology of vascular endothelial growth factor. Endocr Rev 1997;18:4–25.[Abstract/Free Full Text]
  11. Ferrara N, Henzel WJ. Pituitary follicular cells secrete a novel heparin-binding growth factor specific for vascular endothelial cells. Biochem Biophys Res Commun 1989;161:851–858.[CrossRef][Medline]
  12. Conn G, Bayne ML, Soderman DD et al. Amino acid and cDNA sequences of a vascular endothelial cell mitogen that is homologous to platelet-derived growth factor. Proc Natl Acad Sci USA 1990;87:2628–2632.[Abstract/Free Full Text]
  13. Plouet J, Schilling J, Gospodarowicz D. Isolation and characterization of a newly identified endothelial cell mitogen produced by AtT-20 cells. EMBO J 1989;8:3801–3806.[Medline]
  14. Alon T, Hemo I, Itin A et al. Vascular endothelial growth factor acts as a survival factor for newly formed retinal vessels and has implications for retinopathy of prematurity. Nat Med 1995;1:1024–1028.[CrossRef][Medline]
  15. Gerber HP, Dixit V, Ferrara N. Vascular endothelial growth factor induces expression of the antiapoptotic proteins Bcl-2 and A1 in vascular endothelial cells. J Biol Chem 1998;273:13313–13316.[Abstract/Free Full Text]
  16. Benjamin LE, Golijanin D, Itin A et al. Selective ablation of immature blood vessels in established human tumors follows vascular endothelial growth factor withdrawal. J Clin Invest 1999;103:159–165.[Medline]
  17. Yuan F, Chen Y, Dellian M et al. Time-dependent vascular regression and permeability changes in established human tumor xenografts induced by an antivascular endothelial growth factor/vascular permeability factor antibody. Proc Natl Acad Sci USA 1996;93:14765–14770.[Abstract/Free Full Text]
  18. Senger DR, Galli SJ, Dvorak AM et al. Tumor cells secrete a vascular permeability factor that promotes accumulation of ascites fluid. Science 1983;219:983–985.[Abstract/Free Full Text]
  19. Dvorak HF, Brown LF, Detmar M et al. Vascular permeability factor/vascular endothelial growth factor, microvascular hyperpermeability, and angiogenesis. Am J Pathol 1995;146:1029–1039.[Abstract]
  20. Dvorak HF. Tumors: wounds that do not heal. Similarities between tumor stroma generation and wound healing. N Engl J Med 1986;315:1650–1659.[Medline]
  21. Dvorak HF, Harvey VS, Estrella P et al. Fibrin containing gels induce angiogenesis. Implications for tumor stroma generation and wound healing. Lab Invest 1987;57:673–686.[Medline]
  22. Larcher F, Robles AI, Duran H et al. Up-regulation of vascular endothelial growth factor/vascular permeability factor in mouse skin carcinogenesis correlates with malignant progression state and activated H-ras expression levels. Cancer Res 1996;56:5391–5396.[Abstract/Free Full Text]
  23. Kaelin WG, Iliopoulos O, Lonergan KM et al. Functions of the von Hippel-Lindau tumour suppressor protein. J Intern Med 1998;243:535–539.[CrossRef][Medline]
  24. Chiarugi V, Magnelli L, Gallo O. Cox-2, iNOS and p53 as play-makers of tumor angiogenesis (review). Int J Mol Med 1998;2:715–719.[Medline]
  25. Harada H, Nakagawa K, Iwata S et al. Restoration of wild-type p16 down-regulates vascular endothelial growth factor expression and inhibits angiogenesis in human gliomas. Cancer Res 1999;59:3783–3789.[Abstract/Free Full Text]
  26. Rak J, Filmus J, Finkenzeller G et al. Oncogenes as inducers of tumor angiogenesis. Cancer Metastasis Rev 1995;14:263–277.[CrossRef][Medline]
  27. Petit AM, Rak J, Hung MC et al. Neutralizing antibodies against epidermal growth factor and ErbB-2/neu receptor tyrosine kinases down-regulate vascular endothelial growth factor production by tumor cells in vitro and in vivo: angiogenic implications for signal transduction therapy of solid tumors. Am J Pathol 1997;151:1523–1530.[Abstract]
  28. DiGiovanni J. Multistage carcinogenesis in mouse skin. Pharmacol Ther 1992;54:63–128.[CrossRef][Medline]
  29. Yuspa SH, Dlugosz AA. Cutaneous carcinogenesis: natural and experimental. In: Goldsmith LA, ed. Physiology, Biochemistry, and Molecular Biology of the Skin. Vol 2. 2nd ed. New York: Oxford University Press, 1991;1365-1402.
  30. Quintanilla M, Brown K, Ramsden M et al. Carcinogen-specific mutation and amplification of Ha-ras during mouse skin carcinogenesis. Nature 1986;322:78–80.[Medline]
  31. Brown K, Buchmann A, Balmain A. Carcinogen-induced mutations in the mouse c-Ha-ras gene provide evidence of multiple pathways for tumor progression. Proc Natl Acad Sci USA 1990;87:538–542.[Abstract/Free Full Text]
  32. Rak J, Mitsuhashi Y, Bayko L et al. Mutant ras oncogenes upregulate VEGF/VPF expression: implications for induction and inhibition of tumor angiogenesis. Cancer Res 1995;55:4575–4580.[Abstract/Free Full Text]
  33. Grugel S, Finkenzeller G, Weindel K et al. Both v-Ha-Ras and v-Raf stimulate expression of the vascular endothelial growth factor in NIH 3T3 cells. J Biol Chem 1995;270:25915–25919.[Abstract/Free Full Text]
  34. Rho O, Beltran LM, Gimenez-Conti IB et al. Altered expression of the epidermal growth factor receptor and transforming growth factor-alpha during multistage skin carcinogenesis in SENCAR mice. Mol Carcinog 1994;11:19–28.[Medline]
  35. Dlugosz AA, Hansen L, Cheng C et al. Targeted disruption of the epidermal growth factor receptor impairs growth of squamous papillomas expressing the v-ras(Ha) oncogene but does not block in vitro keratinocyte responses to oncogenic ras. Cancer Res 1997;57:3180–3188.[Abstract/Free Full Text]
  36. Hansen LA, Woodson 2nd RL, Holbus S et al. The epidermal growth factor receptor is required to maintain the proliferative population in the basal compartment of epidermal tumors. Cancer Res 2000;60:3328–3332.[Abstract/Free Full Text]
  37. Casanova ML, Larcher F, Casanova B et al. A critical role for ras-mediated, epidermal growth factor receptor-dependent angiogenesis in mouse skin carcinogenesis. Cancer Res 2002;62:3402–3407.[Abstract/Free Full Text]
  38. Rak J, Mitsuhashi Y, Sheehan C et al. Oncogenes and tumor angiogenesis: differential modes of vascular endothelial growth factor up-regulation in ras-transformed epithelial cells and fibroblasts. Cancer Res 2000;60:490–498.[Abstract/Free Full Text]
  39. Risau W. Differentiation of endothelium. FASEB J 1995;9:926–933.[Abstract]
  40. Bolontrade MF, Stern MC, Binder RL et al. Angiogenesis is an early event in the development of chemically induced skin tumors. Carcinogenesis 1998;19:2107–2113.[Abstract/Free Full Text]
  41. Larcher F, Murillas R, Bolontrade M et al. VEGF/VPF overexpression in skin of transgenic mice induces angiogenesis, vascular hyperpermeability and accelerated tumor development. Oncogene 1998;17:303–311.[CrossRef][Medline]
  42. Oono T, Specks U, Eckes B et al. Expression of type VI collagen mRNA during wound healing. J Invest Dermatol 1993;100:329–334.[CrossRef][Medline]
  43. Niquet J, Represa A. Entactin immunoreactivity in immature and adult rat brain. Brain Res Dev Brain Res 1996;95:227–233.[Medline]
  44. Lee WS, Jain MK, Arkonac BM et al. Thy-1, a novel marker for angiogenesis upregulated by inflammatory cytokines. Circ Res 1998;82:845–851.[Abstract/Free Full Text]
  45. Iurlaro M, Loverro G, Vacca A et al. Angiogenesis extent and expression of matrix metalloproteinase-2 and -9 correlate with upgrading and myometrial invasion in endometrial carcinoma. Eur J Clin Invest 1999;29:793–801.[CrossRef][Medline]
  46. St. Croix B, Rago C, Velculescu V et al. Genes expressed in human tumor endothelium. Science 2000;289:1197–1202.[Abstract/Free Full Text]
  47. Carson-Walter EB, Watkins DN, Nanda A et al. Cell surface tumor endothelial markers are conserved in mice and humans. Cancer Res 2001;61:6649–6655.[Abstract/Free Full Text]
  48. Sato TN, Tozawa Y, Deutsch U et al. Distinct roles of the receptor tyrosine kinases Tie-1 and Tie-2 in blood vessel formation. Nature 1995;376:70–74.[CrossRef][Medline]
  49. Puri MC, Rossant J, Alitalo K et al. The receptor tyrosine kinase TIE is required for integrity and survival of vascular endothelial cells. EMBO J 1995;14:5884–5891.[Medline]
  50. Jones N, Master Z, Jones J et al. Identification of Tek/Tie2 binding partners. Binding to a multifunctional docking site mediates cell survival and migration. J Biol Chem 1999;274:30896–30905.[Abstract/Free Full Text]
  51. Kim I, Kim HG, Moon S-O et al. Angiopoietin-1 induces endothelial cell sprouting through the activation of focal adhesion kinase and plasmin secretion. Circ Res 2000;86:952–959.[Abstract/Free Full Text]
  52. Koblizek TI, Weiss C, Yancopoulos GD et al. Angiopoietin-1 induces sprouting angiogenesis in vitro. Curr Biol 1998;8:529–532.[CrossRef][Medline]
  53. Witzenbichler B, Maisonpierre PC, Jones P et al. Chemotactic properties of angiopoietin-1 and -2, ligands for the endothelial-specific receptor tyrosine kinase Tie2. J Biol Chem 1998;273:18514–18521.[Abstract/Free Full Text]
  54. Carmeliet P, Ferreira V, Breier G et al. Abnormal blood vessel development and lethality in embryos lacking a single VEGF allele. Nature 1996;380:435–439.[CrossRef][Medline]
  55. Ferrara N, Carver-Moore K, Chen H et al. Heterozygous embryonic lethality induced by targeted inactivation of the VEGF gene. Nature 1996;380:439–442.[CrossRef][Medline]
  56. Carmeliet P, Ng YS, Nuyens D et al. Impaired myocardial angiogenesis and ischemic cardiomyopathy in mice lacking the vascular endothelial growth factor isoforms VEGF164 and VEGF188. Nat Med 1999;5:495–502.[CrossRef][Medline]
  57. Ferrara N. VEGF: an update on biological and therapeutic aspects. Curr Opin Biotechnol 2000;11:617–624.[CrossRef][Medline]
  58. Gordon MS, Margolin K, Talpaz M et al. Phase I safety and pharmacokinetic study of recombinant human anti-vascular endothelial growth factor in patients with advanced cancer. J Clin Oncol 2001;19:843–850.[Abstract/Free Full Text]
  59. Chen HX, Gore-Langton RE, Cheson BD. Clinical trials referral resource: current clinical trials of the anti-VEGF monoclonal antibody bevacizumab. Oncology (Huntingt) 2001;15:1017, 1020, 1023-1026.[Medline]
  60. Chen Y, Wiesmann C, Fuh G et al. Selection and analysis of an optimized anti-VEGF antibody: crystal structure of an affinity-matured Fab in complex with antigen. J Mol Biol 1999;293:865–881.[CrossRef][Medline]
  61. Prewett M, Huber J, Li Y et al. Antivascular endothelial growth factor receptor (fetal liver kinase 1) monoclonal antibody inhibits tumor angiogenesis and growth of several mouse and human tumors. Cancer Res 1999;59:5209–5218.[Abstract/Free Full Text]
  62. Witte L, Hicklin DJ, Zhu Z et al. Monoclonal antibodies targeting the VEGF receptor-2 (Flk1/KDR) as an anti-angiogenic therapeutic strategy. Cancer Metastasis Rev 1998;17:155–161.[CrossRef][Medline]
  63. Brekken RA, Overholser JP, Stastny VA et al. Selective inhibition of vascular endothelial growth factor (VEGF) receptor 2 (KDR/Flk-1) activity by a monoclonal anti-VEGF antibody blocks tumor growth in mice. Cancer Res 2000;60:5117–5124.[Abstract/Free Full Text]
  64. Kunkel P, Ulbricht U, Bohlen P et al. Inhibition of glioma angiogenesis and growth in vivo by systemic treatment with a monoclonal antibody against vascular endothelial growth factor receptor-2. Cancer Res 2001;61:6624–6628.[Abstract/Free Full Text]
  65. Takayama K, Ueno H, Nakanishi Y et al. Suppression of tumor angiogenesis and growth by gene transfer of a soluble form of vascular endothelial growth factor receptor into a remote organ. Cancer Res 2000;60:2169–2177.[Abstract/Free Full Text]
  66. Mori A, Arii S, Furutani M et al. Soluble Flt-1 gene therapy for peritoneal metastases using HVJ-cationic liposomes. Gene Ther 2000;7:1027–1033.[CrossRef][Medline]
  67. Mahasreshti PJ, Navarro JG, Kataram M et al. Adenovirus-mediated soluble FLT-1 gene therapy for ovarian carcinoma. Clin Cancer Res 2001;7:2057–2066.[Abstract/Free Full Text]
  68. Machein MR, Risau W, Plate KH. Antiangiogenic gene therapy in a rat glioma model using a dominant-negative vascular endothelial growth factor receptor 2. Hum Gene Ther 1999;10:1117–1128.[CrossRef][Medline]
  69. Parry TJ, Bouhana KS, Blanchard KS et al. Ribozyme pharmacokinetic screening for predicting pharmacodynamic dosing regimens. Curr Issues Mol Biol 2000;2:113–118.[Medline]
  70. Weng DE, Usman N. Angiozyme: a novel angiogenesis inhibitor. Curr Oncol Rep 2001;3:141–146.[Medline]
  71. Weng DE, Weiss P, Kellackey C et al. Angiozyme pharmacokinetic and safety results: a phase I/II study in patients with refractory solid tumors. Proc Am Soc Clin Oncol 2001;20(pt 1):99a.
  72. Fong TA, Shawver LK, Sun L et al. SU5416 is a potent and selective inhibitor of the vascular endothelial growth factor receptor (Flk-1/KDR) that inhibits tyrosine kinase catalysis, tumor vascularization, and growth of multiple tumor types. Cancer Res 1999;59:99–106.[Abstract/Free Full Text]
  73. Mendel DB, Laird AD, Smolich BD et al. Development of SU5416, a selective small molecule inhibitor of VEGF receptor tyrosine kinase activity, as an anti-angiogenesis agent. Anticancer Drug Des 2000;15:29–41.[Medline]
  74. Via LE, Gore-Langton RE, Pluda JM. Clinical trials referral resource. Current clinical trials administering the antiangiogenesis agent SU5416. Oncology (Huntingt) 2000;14:1312, 1315-1316, 1321-1323.[Medline]
  75. Salzberg M, Pless M, Rochlitz C et al. A phase I and pharmacokinetic study of single oral administration of SU5416 in patients with advanced solid tumors. Proc Am Soc Clin Oncol 2001;20(pt 1):98a.
  76. Fiedler WM, Tinnefeld H, Mende T et al. A phase II study with SU5416 in patients with C-Kit positive AML. Proc Am Soc Clin Oncol 2001;20(pt 1):288a.
  77. Kindler HL, Vogelzang NJ, Chien K et al. SU5416 in malignant mesothelioma: a University of Chicago phase II consortium study. Proc Am Soc Clin Oncol 2001;20(pt 1):341a.
  78. Gajewski TF, Flickinger S, Heimann R et al. Phase II study of the anti-angiogenic Flk-1 tyrosine kinase inhibitor SU5416 in melanoma: initial results. Proc Am Soc Clin Oncol 2001;20(pt 1):360a.
  79. Eng C, Kindler HL, Stadler WM et al. SU5416 in advanced colorectal cancer (CRC): a University of Chicago phase II consortium study. Proc Am Soc Clin Oncol 2001;20(pt 2):116b.
  80. Laird AD, Vajkoczy P, Shawver LK et al. SU6668 is a potent antiangiogenic and antitumor agent that induces regression of established tumors. Cancer Res 2000;60:4152–4160.[Abstract/Free Full Text]
  81. Rosen LS, Rosen PJ, Kabbinavar F et al. Phase I experience with SU6668, a novel multiple receptor tyrosine kinase inhibitor in patients with advanced malignancies. Proc Am Soc Clin Oncol 2001;20(pt 1):97a.
  82. Wood JM, Bold G, Buchdunger E et al. PTK787/ZK 222584, a novel and potent inhibitor of vascular endothelial growth factor receptor tyrosine kinases, impairs vascular endothelial growth factor-induced responses and tumor growth after oral administration. Cancer Res 2000;60:2178–2189.[Abstract/Free Full Text]
  83. Traxler P, Bold G, Buchdunger E et al. Tyrosine kinase inhibitors: from rational design to clinical trials. Med Res Rev 2001;21:499–512.[CrossRef][Medline]
  84. Thomas A, Morgan B, Drevs J et al. Pharmacodynamic results using dynamic contrast enhanced magnetic resonance imaging, of 2 phase 1 studies of the VEGF Inhibitor PTK787/ZK 222584 in patients with liver metastases from colorectal cancer. Proc Am Soc Clin Oncol 2001;20(pt 1):71a.
  85. Drevs J, Mross K, Fuxius S. A phase-1 dose-escalating and pharmacokinetic (PK) study of the VEGF-receptor-inhibitor PTK787/ZK222584 (PTK/ZK) in patients with liver metastasis of advanced cancer. Proc Am Soc Clin Oncol 2001;(pt 1):100a.
  86. Wedge SR, Ogilvie DJ, Dukes M et al. ZD4190: an orally active inhibitor of vascular endothelial growth factor signaling with broad-spectrum antitumor efficacy. Cancer Res 2000;60:970–975.[Abstract/Free Full Text]
  87. Wedge SR, Ogilvie DJ. Inhibition of VEGF signal transduction. Identification of ZD4190. Adv Exp Med Biol 2000;476:307–310.[Medline]
  88. O’Reilly MS, Holmgren L, Shing Y et al. Angiostatin: a novel angiogenesis inhibitor that mediates the suppression of metastases by a Lewis lung carcinoma. Cell 1994;79:315–328.[CrossRef][Medline]
  89. O’Reilly MS, Boehm T, Shing Y et al. Endostatin: an endogenous inhibitor of angiogenesis and tumor growth. Cell 1997;88:277–285.[CrossRef][Medline]
  90. Moser TL, Stack MS, Asplin I et al. Angiostatin binds ATP synthase on the surface of human endothelial cells. Proc Natl Acad Sci USA 1999;96:2811–2816.[Abstract/Free Full Text]
  91. Claesson-Welsh L, Welsh M, Ito N et al. Angiostatin induces endothelial cell apoptosis and activation of focal adhesion kinase independently of the integrin-binding motif RGD. Proc Natl Acad Sci USA 1998;95:5579–5583.[Abstract/Free Full Text]
  92. Lucas R, Holmgren L, Garcia I et al. Multiple forms of angiostatin induce apoptosis in endothelial cells. Blood 1998;92:4730–4741.[Abstract/Free Full Text]
  93. Taddei L, Chiarugi P, Brogelli L et al. Inhibitory effect of full-length human endostatin on in vitro angiogenesis. Biochem Biophys Res Commun 1999;263:340–345.[CrossRef][Medline]
  94. Dhanabal M, Ramchandran R, Waterman MJ et al. Endostatin induces endothelial cell apoptosis. J Biol Chem 1999;274:11721–11726.[Abstract/Free Full Text]
  95. Kerbel RS, Viloria-Petit A, Klement G et al. ‘Accidental’ anti-angiogenic drugs. Anti-oncogene directed signal transduction inhibitors and conventional chemotherapeutic agents as examples. Eur J Cancer 2000;36:1248–1257.
  96. Melillo G, Sausville EA, Cloud K et al. Flavopiridol, a protein kinase inhibitor, down-regulates hypoxic induction of vascular endothelial growth factor expression in human monocytes. Cancer Res 1999;59:5433–5437.[Abstract/Free Full Text]
  97. Miller KD, Sweeney CJ, Sledge Jr. GW Redefining the target: chemotherapeutics as antiangiogenics. J Clin Oncol 2001;19:1195–1206.[Abstract/Free Full Text]
Received April 25, 2002; accepted for publication June 26, 2002.





This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow eLetters: Submit a response to this article
Right arrow Purchase Article
Right arrow View Shopping Cart
Right arrow Alert me when this article is cited
Right arrow Alert me when eLetters are posted
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow E-mail this article link to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow Reprints/Permissions
Citing Articles
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Conti, C. J.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Conti, C. J.

HOME HELP CONTACT US SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
THE ONCOLOGIST STEM CELLS CME ALPHAMED PRESS JOURNALS