Peroxisome Proliferator-activated Receptor γ Ligands Are Potent Inhibitors of Angiogenesis in Vitro and in Vivo

Abstract

Peroxisome proliferator-activated receptor γ (PPARγ) is a nuclear receptor that functions as a transcription factor to mediate ligand-dependent transcriptional regulation. Activation of PPARγ by the naturally occurring ligand, 15-deoxy-Δ12,14-prostaglandin J2(15d-PGJ2), or members of a new class of oral antidiabetic agents, e.g. BRL49653 and ciglitizone, has been linked to adipocyte differentiation, regulation of glucose homeostasis, inhibition of macrophage and monocyte activation, and inhibition of tumor cell proliferation. Here we report that human umbilical vein endothelial cells (HUVEC) express PPARγ mRNA and protein. Activation of PPARγ by the specific ligands 15d-PGJ2, BRL49653, or ciglitizone, dose dependently suppresses HUVEC differentiation into tube-like structures in three-dimensional collagen gels. In contrast, specific PPARα and -β ligands do not affect tube formation although mRNA for these receptors are expressed in HUVEC. PPARγ ligands also inhibit the proliferative response of HUVEC to exogenous growth factors. Treatment of HUVEC with 15d-PGJ2also reduced mRNA levels of vascular endothelial cell growth factor receptors 1 (Flt-1) and 2 (Flk/KDR) and urokinase plasminogen activator and increased plasminogen activator inhibitor-1 (PAI-1) mRNA. Finally, administration of 15d-PGJ2 inhibited vascular endothelial cell growth factor-induced angiogenesis in the rat cornea. These observations demonstrate that PPARγ ligands are potent inhibitors of angiogenesis in vitro and in vivo, and suggest that PPARγ may be an important molecular target for the development of small-molecule inhibitors of angiogenesis. Peroxisome proliferator-activated receptor γ (PPARγ) is a nuclear receptor that functions as a transcription factor to mediate ligand-dependent transcriptional regulation. Activation of PPARγ by the naturally occurring ligand, 15-deoxy-Δ12,14-prostaglandin J2(15d-PGJ2), or members of a new class of oral antidiabetic agents, e.g. BRL49653 and ciglitizone, has been linked to adipocyte differentiation, regulation of glucose homeostasis, inhibition of macrophage and monocyte activation, and inhibition of tumor cell proliferation. Here we report that human umbilical vein endothelial cells (HUVEC) express PPARγ mRNA and protein. Activation of PPARγ by the specific ligands 15d-PGJ2, BRL49653, or ciglitizone, dose dependently suppresses HUVEC differentiation into tube-like structures in three-dimensional collagen gels. In contrast, specific PPARα and -β ligands do not affect tube formation although mRNA for these receptors are expressed in HUVEC. PPARγ ligands also inhibit the proliferative response of HUVEC to exogenous growth factors. Treatment of HUVEC with 15d-PGJ2also reduced mRNA levels of vascular endothelial cell growth factor receptors 1 (Flt-1) and 2 (Flk/KDR) and urokinase plasminogen activator and increased plasminogen activator inhibitor-1 (PAI-1) mRNA. Finally, administration of 15d-PGJ2 inhibited vascular endothelial cell growth factor-induced angiogenesis in the rat cornea. These observations demonstrate that PPARγ ligands are potent inhibitors of angiogenesis in vitro and in vivo, and suggest that PPARγ may be an important molecular target for the development of small-molecule inhibitors of angiogenesis. Angiogenesis, the formation of new blood vessels, is not only critically involved in a number of normal physiological processes such as embryonic development, ovulation, and wound healing, but is also a critical step in many pathological conditions including solid tumor growth, diabetic retinopathy, and age-related macular degeneration (1Folkman J. Nat. Med. 1995; 1: 27-31Crossref PubMed Scopus (7218) Google Scholar). The complex steps involved in new vessel formation have been well characterized in recent years and involve the degradation of the basement membrane by cellular proteases, the penetration and migration of endothelial cells into the extracellular matrix, endothelial proliferation, and the resultant formation of patent, interconnected vascular networks (2Folkman J. Cancer Res. 1986; 46: 467-473PubMed Google Scholar, 3Liotta L.A. Steeg P.S. Stetler-Stevenson W.G. Cell. 1991; 64: 327-336Abstract Full Text PDF PubMed Scopus (2640) Google Scholar). It is now well established that the progression of solid tumor growth and metastasis is dependent on angiogenesis, with the resultant global effort by many laboratories to identify new angiostatic therapies for the treatment of cancer. Peroxisome proliferator-activated receptors (PPARs) 1The abbreviations used are:PPARs, peroxisome proliferator-activated receptors; HUVEC, human umbilical vein endothelial cells; 15d-PGJ2, 15-deoxy-Δ12,14-prostaglandin J2; RXR, retinoid acid receptor; VEGF, vascular endothelial growth factor; PMA, phorbol myristate acetate; RT-PCR, reverse transcription-polymerase chain reaction; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; BrdUrd, 5′-bromo-2′-deoxyuridine; FBS, fetal bovine serum; bFGF, basic fibroblast growth factor; ITS, insulin, transferrin, and selenium A; uPA, urokinase-type plasminogen activator. are members of the steroid receptor superfamily and, as such, are ligand-activated transcription factors (4Issemann I. Green S. Nature. 1990; 347: 645-650Crossref PubMed Scopus (3047) Google Scholar, 5Schoonjans K. Martin G. Staels B. Auwerx J. Curr. Opin. Lipidol. 1997; 8: 159-166Crossref PubMed Scopus (468) Google Scholar). Three subtypes of PPAR, α, β (also known as δ or NUCI), and γ have been identified and cloned. Like other members of this superfamily, PPARs mediate transcriptional regulation by their central DNA binding domain that recognizes response elements in the promoters of specific target genes (6Kliewer S.A. Umesono K. Noonan D.J. Heyman R.A. Evans R.M. Nature. 1992; 358: 771-774Crossref PubMed Scopus (1521) Google Scholar, 7Palmer C.N.A. Hsu M.H. Griffin K.J. Johnson E.F. J. Biol. Chem. 1995; 270: 16114-16121Abstract Full Text Full Text PDF PubMed Scopus (261) Google Scholar). Activation of PPARγ has been linked to adipocyte differentiation and regulation of glucose homeostasis in rodents and humans (8Spiegelman B.M. Diabetes. 1998; 47: 507-514Crossref PubMed Scopus (1640) Google Scholar). Recent evidence also shows that the natural receptor ligand for PPARγ, 15d-PGJ2 (9Kliewer S.A. Lenhard J.M. Willson T.M. Patel I. Morris D.C. Lehmann J.M. Cell. 1995; 83: 813-819Abstract Full Text PDF PubMed Scopus (1869) Google Scholar, 10Forman B.M. Tontonoz P. Chen J. Brun R.P. Spiegelman B.M. Evans R.M. Cell. 1995; 83: 803-812Abstract Full Text PDF PubMed Scopus (2731) Google Scholar), and synthetic antidiabetic thiazolidinedione drugs (e.g. BRL49653 and ciglitizone), inhibit macrophage and monocyte activation (11Ricote M. Li A.C. Willson T.M. Kelly C.J. Glass C.K. Nature. 1998; 391: 79-82Crossref PubMed Scopus (3260) Google Scholar, 12Jiang C. Ting A.T. Seed B. Nature. 1998; 391: 82-86Crossref PubMed Scopus (539) Google Scholar) and suppress tumor cell growth (14Elstner E. Muller C. Koshizuka K. Williamson E.A. Park D. Asou H. Shintaku P. Said J.W. Heber D. Koeffler H.P. Proc. Natl. Acad. Sci. U. S. A. 1998; 95: 8806-8811Crossref PubMed Scopus (760) Google Scholar, 15Kubota T. Koshizuka K. Williamson E. Asou H. Said J. Holden S. Miyoshi I. Koeffler H.P. Cancer Res. 1998; 58: 3344-3352PubMed Google Scholar, 16Sarraf P. Mueller E. Jones D. King F.J. DeAngelo D.J. Partridge J.B. Holden S.A. Chen L.B. Singer S. Fletcher C. Spiegelman B.A. Nat. Med. 1998; 4: 1046-1052Crossref PubMed Scopus (929) Google Scholar). PPARγ can heterodimerize with at least one other member of the steroid receptor superfamily, retinoid acid receptor (RXR) (6Kliewer S.A. Umesono K. Noonan D.J. Heyman R.A. Evans R.M. Nature. 1992; 358: 771-774Crossref PubMed Scopus (1521) Google Scholar, 17Schulman I.G. Shao G. Heyman R.A. Mol. Cell. Biol. 1998; 18: 3483-3494Crossref PubMed Google Scholar). Specific ligands for the PPARγ and RXR have been shown to act synergistically to induce terminal differentiation of human liposarcoma cells (18Tontonoz P. Singer S. Forman B.M. Sarraf P. Fletcher J.A. Fletcher C.D.M. Brun R.P. Mueller E. Altiok S. Oppenheim H. Evans R.M. Spiegelman B.M. Proc. Natl. Acad. Sci. U. S. A. 1997; 94: 237-241Crossref PubMed Scopus (619) Google Scholar) in vitro and to enhance insulin sensitivity in diabetic animals (19Mukherjee R. Davies P.J.A. Crombie D.J. Bischoff E.D. Cesario R.M. Jow L. Hamam L.G. Boehm M.F. Mondon C.E. Nadzan A.M. Paterniti Jr., J.R. Heyman R.A. Nature. 1997; 386: 407-410Crossref PubMed Scopus (576) Google Scholar). Activation of PPARα has been demonstrated to mediate lipid catabolism. Treatment of animals with PPARα activators results in the proliferation of peroxisomes and the induction of hepatic genes involved in the β-oxidation of fatty acids (20Issemann I. Green S. Nature. 1990; 347: 645-650Crossref PubMed Scopus (2889) Google Scholar). Mice that lack functional PPARα accumulate lipid droplets in their livers (21Lee S.S. Pineau T. Drago J. Lee E.J. Owens J.W. Kroetz D.L. Fernandez-Salguero P.M. Westphal H. Gonzalez F.J. Mol. Cell. Biol. 1995; 15: 3012-3022Crossref PubMed Scopus (1501) Google Scholar). Several selective PPARα activators have been described including WY 14643 and clofibrate (22Forman B.M. Chen J. Evans R.M. Proc. Natl. Acad. Sci. U. S. A. 1997; 94: 4312-4317Crossref PubMed Scopus (1866) Google Scholar). Inhibition of human aortic smooth muscle cell activation by PPARα ligands has also been reported recently (13Staels B. Koenig W. Habib A. Merval R. Lebret M. Torra I.P. Delerive P. A F. Chinetti G. Fruchart J. Najib J. Maclouf J. Tedgui A. Nature. 1998; 393: 790-793Crossref PubMed Scopus (1056) Google Scholar). Although PPARβ has been found to be ubiquitously expressed, specific functions for this receptor are not known at the present time. In this study we report on the potent and novel inhibitory activity of PPARγ ligands on HUVEC differentiation into tube-like structures and proliferation in vitro, and the inhibition of VEGF elicited angiogenesis in vivo. These studies demonstrate that PPARγ is an important molecular target for the development of small molecule inhibitors of angiogenesis, which may be useful therapeutic agents in the treatment of cancer and other vasculoproliferative disorders. PGA1, PGA2, PGB1, PGB2, PGD1, PGD2, PGE1, PGE2, ciglitizone, WY 14643, clofibrate, and 15d-PGJ2 were from Cayman Chemical (Ann Arbor, MI). BRL49653 was kindly synthesized at Genentech by Dr. Jim Marsters (South San Francisco, CA). HUVEC were purchased from Clonetics (San Diego, CA) and maintained in Clonetics EGM medium supplemented with 10% fetal bovine serum (FBS) and endothelial cell growth supplements provided by the company. Type I rat tail collagen and recombinant basic fibroblast growth factor (bFGF) were purchased from Collaborative Research (Becton Dickinson Labware, Bedford, MA). Recombinant VEGF was from Genentech. 10× medium 199 (M199) and PMA were purchased from Sigma. FBS was from Hyclone (Logan, Utah). ITS (insulin, transferrin, and selenium A), trypsin, and versene were from Life Technologies, Inc. The mouse monoclonal antibody to PPARγ (E-8, SC-7273) and goat polyclonal antibodies to PPARα (N-19, SC-1985) and PPARβ (C-20, SC-1983) were purchased from Santa Cruz Biotechnology (Santa Cruz, CA). Magnetic protein A beads (Dynabeads) were from Dynal (Lake Success, NY). Collagen gels were formed by mixing together ice-cold gelation solution (10× M199, H2O, 0.53m NaHCO3, 200 mm l-glutamine, type I collagen, 0.1 m NaOH, 100:27.2:50:10:750:62.5 by volume) and cells in 1× basal medium (see below) at a concentration of 3 × 106 cells/ml at a ratio of 4 volumes gelation solution:1 volume of cells. After gelation at 37 °C for 30 min, the gels were overlaid with 1× basal medium consisting of M199 supplemented with 1% FBS, 1× ITS, 2 mm l-glutamine, 50 μg/ml ascorbic acid, 26.5 mmNaHCO3, 100 units/ml penicillin, and 100 units/ml streptomycin supplemented with 40 ng/ml bFGF, 40 ng/ml VEGF, and 80 nm PMA. All drugs were added to the 1× basal medium immediately after gelation. To quantitate tube formation, the number of tubes per high power (20X) field was determined at 48 h after addition of the basal medium. A tube was defined as an elongated structure comprised of one or more endothelial cells that exceeded 100 μm in length (long axis). Five independent fields separated by 100 μm optical sections were assessed for each well, and the average number of tubes/20X field determined. Cytoxicity was assessed using cell proliferation kit II from Boehringer Mannheim. Synthesis of complementary DNA (cDNA) was performed using 6 μg of total RNA extracted from human liver (positive control) or HUVEC cultured on three-dimensional collagen gels for 24 h by random primer according to the manufacturer's instructions (Stratagene Co., San Diego, CA). Subsequent amplifications of the partial cDNA encoding PPARα, β, and γ were performed using different amounts of reverse transcribed mixture (4 μl for liver α, β, and γ and 5 μl for HUVEC α and β and 6 μl for HUVEC γ) as templates with specific oligonucleotide primers as follows: PPARα sense 5′-CCAGTATTTAGGACGCGGTCC-3′ and antisense 5′-AAGTTCTTCAAGTAGGCCTGCG-3′; PPARβ sense 5′-AACTGCAGATGGGCTGTGAC-3′ and antisense 5′-GTCTCGATGTCGTGGATCAC-3′ and PPARγ sense 5′-TCTCTCCGTAATGGAAGACC-3′ and antisense 5′-GCATTATGAGACATCCCCAC-3′. The expected sizes of PCR products for PPAR α, β, and γ were 492, 484, and for reverse transcription and PCR amplifications were The PCR were to of by at at and at The PCR products were by with has been used to mRNA and has been described in J. K.J. P.M. Scholar, H. F. Park J. J. Biol. Chem. 1997; PubMed Scopus Google Scholar). RNA was extracted from HUVEC cultured in three-dimensional collagen gels or on for in 1× basal medium consisting of M199 supplemented with 1% FBS, 1× ITS, 2 mm l-glutamine, 50 μg/ml ascorbic acid, 26.5 mmNaHCO3, 100 units/ml penicillin, and 100 units/ml streptomycin supplemented with 40 ng/ml bFGF, 40 ng/ml VEGF, and 80 nm PMA. A PCR oligonucleotide primer and an oligonucleotide with a at the and a at the were using the in the The primers and used were as follows: human PPARα primer reverse primer and human PPARβ primer reverse primer and human PPARγ primer reverse primer and human primer reverse primer and human primer reverse primer and human urokinase-type plasminogen activator primer reverse primer and human primer reverse primer and human glyceraldehyde-3-phosphate reverse and RNA was added to a mixture according to the manufacturer's The conditions 1 at 48 °C for 30 min, 1 at °C for min, 40 at °C for at °C for 1 min, and at °C for 2 for of each were by of the total RNA from HUVEC. of was not the to 48 of with drugs and the mRNA of each was to the of in the RNA HUVEC on type I were with ice-cold and by were in ice-cold mm 2 mm 1 mm 40 units/ml μg/ml 1% and 1% to 100 μg of protein was for h at 4 °C with of antibody in a total volume of beads were and the mixture for 2 h at 4 The beads were with μl of and the on the beads were by at 100 °C for 5 in The were by were using the kit HUVEC were on at in Clonetics EGM endothelial cell growth supplements and 10% FBS, 2 mm l-glutamine, 100 units/ml penicillin, and 100 units/ml streptomycin and to for 4 was with 1× basal medium consisting of M199 supplemented with 1% FBS, 1× ITS, 2 mm l-glutamine, 50 μg/ml ascorbic acid, 26.5 mm NaHCO3, 100 units/ml penicillin, and 100 units/ml streptomycin supplemented with 40 ng/ml bFGF, 40 ng/ml VEGF, and 80 nm PMA. were cultured in the medium in the of drugs or for 4 5 μl of was added in a volume of 100 and cells were for was by an kit from Boehringer Mannheim. A was 1 mm from the of the of a the was the the of the A × VEGF with or 15d-PGJ2 was into the of the After the were with were at h for the of and the animals were and with to for of the were of the and for using are expressed as the was performed using of CA). the were using of to was as In this study we used a of in described by Res. PubMed Scopus Google Scholar). In this endothelial cells in three-dimensional collagen comprised of type I collagen in the of VEGF, bFGF, and PMA are in the of endothelial cells 4 the formation of tube-like structures can be at 24 and an interconnected of tube-like structures is at 48 of mRNA and protein the formation of tube-like structures not the for new mRNA and protein It has recently been shown that a number of can as PPAR ligands (8Spiegelman B.M. Diabetes. 1998; 47: 507-514Crossref PubMed Scopus (1640) Google Scholar, S.A. Lenhard J.M. Willson T.M. Patel I. Morris D.C. Lehmann J.M. Cell. 1995; 83: 813-819Abstract Full Text PDF PubMed Scopus (1869) Google Scholar, 10Forman B.M. Tontonoz P. Chen J. Brun R.P. Spiegelman B.M. Evans R.M. Cell. 1995; 83: 803-812Abstract Full Text PDF PubMed Scopus (2731) Google Scholar, B.M. Chen J. Evans R.M. Proc. Natl. Acad. Sci. U. S. A. 1997; 94: 4312-4317Crossref PubMed Scopus (1866) Google Scholar), and activation of PPARs has been linked to functions in a number of cell (8Spiegelman B.M. Diabetes. 1998; 47: 507-514Crossref PubMed Scopus (1640) Google Scholar, M. Li A.C. Willson T.M. Kelly C.J. Glass C.K. Nature. 1998; 391: 79-82Crossref PubMed Scopus (3260) Google Scholar, 12Jiang C. Ting A.T. Seed B. Nature. 1998; 391: 82-86Crossref PubMed Scopus (539) Google Scholar, B. Koenig W. Habib A. Merval R. Lebret M. Torra I.P. Delerive P. A F. Chinetti G. Fruchart J. Najib J. Maclouf J. Tedgui A. Nature. 1998; 393: 790-793Crossref PubMed Scopus (1056) Google Scholar, E. Muller C. Koshizuka K. Williamson E.A. Park D. Asou H. Shintaku P. Said J.W. Heber D. Koeffler H.P. Proc. Natl. Acad. Sci. U. S. A. 1998; 95: 8806-8811Crossref PubMed Scopus (760) Google Scholar, 15Kubota T. Koshizuka K. Williamson E. Asou H. Said J. Holden S. Miyoshi I. Koeffler H.P. Cancer Res. 1998; 58: 3344-3352PubMed Google Scholar, 16Sarraf P. Mueller E. Jones D. King F.J. DeAngelo D.J. Partridge J.B. Holden S.A. Chen L.B. Singer S. Fletcher C. Spiegelman B.A. Nat. Med. 1998; 4: 1046-1052Crossref PubMed Scopus (929) Google Scholar). To PPARs affect HUVEC differentiation, we the of 15d-PGJ2, a naturally occurring ligand of PPARγ on HUVEC tube HUVEC were in three-dimensional collagen gels in medium VEGF, and PMA with addition of 15d-PGJ2 for 48 h and were for tube shown in 1 15d-PGJ2 reduced HUVEC tube The of cells in the gels and of or not inhibitory not observations to a of of and fatty acids for their to inhibit tube formation in three-dimensional collagen gels 1 and and their demonstrated inhibition of tube formation the concentration of 1 to 100 and only inhibitory In contrast, known to cell and PGE2, this concentration the 15d-PGJ2 was the potent of HUVEC tube from that of the the of the not studies J. Biol. Chem. Full Text PDF PubMed Google Scholar, H. P. PubMed Scopus Google Scholar) have demonstrated that can to and in the of serum or to 15d-PGJ2, a natural selective ligand for activation of PPARγ may endothelial was used to demonstrate the of PPAR β, and γ) in HUVEC cultured in the three-dimensional collagen gels. In independent with different HUVEC from different the mRNA of of PPAR were demonstrated 2 using J. K.J. P.M. Scholar, H. F. Park J. J. Biol. Chem. 1997; PubMed Scopus Google Scholar), the mRNA levels of each were determined to be PPARβ PPARα PPARγ observations also that of PPARγ mRNA was not in medium VEGF, and PMA at 24 or 48 h of with 15d-PGJ2 or in HUVEC cultured in three-dimensional collagen gels or on not The of PPARγ protein in HUVEC was by 2 and PPARγ protein levels were not by the growth factor mixture HUVEC were cultured on 2 we were to demonstrate protein of PPARα and -β in HUVEC not The of PPARα mRNA in HUVEC has been reported I. K. S. T. S. Res. 1998; PubMed Scopus Google this study for the the of PPARγ mRNA and protein and PPARβ mRNA by HUVEC. To which PPAR the inhibition of HUVEC tube formation, the of specific PPAR ligands was shown in treatment of HUVEC with the specific PPARγ 15d-PGJ2, BRL49653, and ciglitizone, dose dependently inhibited tube formation with inhibition of and In contrast, the selective for PPARα 14643 and and PPARβ R. A. J. Mol. Biol. 1997; PubMed Scopus Google Scholar) not inhibit tube formation These observations demonstrated that activation of PPARγ results in inhibition of endothelial cell differentiation into tube-like structures in PPARγ ligands also the proliferative response of HUVEC to growth factors. were performed to proliferation of HUVEC cultured on type I in medium VEGF, bFGF, and PMA in the of or PPARγ shown in 3 15d-PGJ2 and BRL49653 dose dependently inhibited HUVEC proliferation with of and In contrast, and not affect HUVEC proliferation not These results demonstrate that PPARγ ligands also growth factor-induced HUVEC proliferation in and are endothelial cell receptors for The of these receptors angiogenesis has been demonstrated by the that functions as a to endothelial cell proliferation and differentiation and that is a critical factor involved in endothelial cell J. M. Nature. 1995; PubMed Scopus Google Scholar, T. 1997; 18: PubMed Scopus Google Scholar, S. J.A. J. Sci. 1998; PubMed Google Scholar). activation of PPARγ and using and mRNA were by the mixture of growth factors in HUVEC in three-dimensional collagen gels A and 15d-PGJ2 inhibited the induction of VEGF receptor mRNA in HUVEC cultured in three-dimensional collagen gels A and were HUVEC were on type I with growth factor It is also well known that the of and their inhibitors activator is with endothelial cell degradation of extracellular and the critical steps of the processes Curr. Opin. Biol. 1997; PubMed Scopus Google Scholar). the of 15d-PGJ2 on of and in three-dimensional collagen gels. Treatment of HUVEC with 15d-PGJ2 reduced mRNA at 4 h and increased at 24 these observations molecular by which PPARγ ligands mediate inhibition of angiogenesis. To the of PPARγ ligand 15d-PGJ2 in vivo, 200 of recombinant VEGF with or μg of 15d-PGJ2 were into the of from the that the of 15d-PGJ2 and VEGF demonstrated a in vessel with the VEGF only from this were in a independent These in in vitro observations that activation of PPARγ results in inhibition of angiogenesis. Recent evidence has demonstrated that PPARγ is expressed in cancer cells and cancer cells (14Elstner E. Muller C. Koshizuka K. Williamson E.A. Park D. Asou H. Shintaku P. Said J.W. Heber D. Koeffler H.P. Proc. Natl. Acad. Sci. U. S. A. 1998; 95: 8806-8811Crossref PubMed Scopus (760) Google Scholar, 16Sarraf P. Mueller E. Jones D. King F.J. DeAngelo D.J. Partridge J.B. Holden S.A. Chen L.B. Singer S. Fletcher C. Spiegelman B.A. Nat. Med. 1998; 4: 1046-1052Crossref PubMed Scopus (929) Google Scholar). Activation of the PPARγ in growth and induction of differentiation of cancer cells and of tumor cell growth of PPARγ as a target for treatment of human In this study we have for the the of PPARγ in human endothelial cells. Activation of PPARγ with naturally occurring ligand or synthetic selective ligands results in potent inhibition of endothelial differentiation into tube-like structures and proliferation in vitro and of angiogenesis in vivo. PPARγ activation also the of at least important genes in the the VEGF receptors and the study a new into the of PPARγ ligands as inhibitors of solid tumor growth and that such drugs may also novel to other disorders. the DNA at Genentech for and