© The Author 2006. Published by Oxford University Press.
EDITORIAL |
Choking Hypoxia-Inducible Factor 1
: A Novel Mechanism for Connective Tissue Growth Factor Inhibition of Angiogenesis
Affiliations of authors: Molecular Oncology Laboratory, National Cancer Research Institute, Genoa, Italy (FT, AA); University of Insubria, Varese, Italy (DMN)
Correspondence to: Adriana Albini, PhD, Ist. Nazionale per la Ricerca sul Cancro, Molecular Oncology, Largo Rosanna Benzi, 10, Genova 16132, Italy (e-mail: adriana.albini{at}istge.it).
Among the driving forces that orchestrate oncogenic transformation, aberrant tumor angiogenesis has evolved as a process assured by the conscription of various signaling pathways with redundant functions. These pathways converge on the same task: to produce more vessels, without ensuring correct structural constraints and proper function. Different cellular components in tissues are equipped with proangiogenic molecular machinery, as demonstrated by studies on the role of inflammatory cells in tumor promotion that have driven the development of new therapies for cancer prevention and treatment (1,2).
Transcriptional reprogramming controlled by dysregulated signaling pathways is one of the mechanisms that are the basis for the cellular adaptations necessary for cancer establishment. In particular, one master regulator, the hypoxia-inducible transcription factor 1 (HIF-1) complex, which controls tissue homeostasis, seems to take center stage as a pervasive instigator of transformation for almost all cellular compartments in tissues.
HIF-1, primarily recognized as a mediator of adaptive responses to tissue oxygenation, integrates diverse information related to energy status, glucose, and iron metabolism, as well as growth factor signaling in normoxia and hypoxia (3,4). Furthermore, recent evidence indicates that HIF-1–activated metabolic responses are essential for the recruitment and infiltration of myeloid cells at early stages of inflammation (5). The decision to constitutively activate HIF-1
as a means for coping with stressful or threatening local conditions in the microenvironment carries the high cost of increased risk of neoplastic transformation. HIF-1 also contributes to define an invasive, lethal cancer phenotype by elevating the metabolic resources of cells through increased glucose uptake, glycolysis, and angiogenesis (6).
The HIF-1 subunit of the heterodimeric transcriptional complex HIF-1 is the labile regulatory element destined to ubiquitination and default destruction in the absence of alerting signals from the microenvironment. Proteins of the von Hippel-Lindau (pVHL)–ubiquitin–proteasome pathway, isoforms of prolyl hydroxylases (PHDs), and the acetyltransferase ARD1 eliminate necessary HIF-1 under normoxia in normal cells. Hydroxylation of HIF-1 at proline residues 402 and 564 in the oxygen-dependent degradation (ODD) domain by PHDs targets HIF-1 to pVHL in the presence of dioxygen (7). ARD1 marks HIF-1 for proteasomal degradation by acetylating the Lys 532 residue in the ODD domain, thus inducing a conformational change that increases binding to pVHL (8).
Apparently, the list of molecules that are involved in the HIF-1 pathway is far from complete. In this issue of the Journal, Chang et al. (9) provide mechanistic insight of how connective tissue growth factor (CTGF/CCN2) exerts a potent growth-inhibiting and antimetastatic effect through angiogenesis repression by controlling HIF-1 degradation.
CTGF is a cysteine-rich secretory protein belonging to the CCN (from CYR61, CTGF, and NOV) family first discovered in endothelial cells. It works as a multifunctional growth factor in wound repair, inflammation, cell adhesion, chemotaxis, apoptosis, tumor growth, and fibrotic disorders (10). This panoply of functions for one protein reveals the complex duty of matrix components that are delegated to the microenvironmental control of cell proliferation and differentiation at the whole-organ level. What Chang et al. unraveled is that CTGF drives a negative modulation of HIF-1 through ARD1.
The authors began by studying xenograft tumors of human lung adenocarcinoma cells transfected with CTGF. Although cell proliferation in culture was only marginally affected, growth of xenografts of CTGF overexpressing clones was reduced, a phenomenon that was directly associated with reduced microvessel density in the xenografts. Conversely, when CTGF was silenced, these xenografts grew faster and vascularization was increased. The results of the straightforward experimental approach used to single out the key molecules in the angiogenic process were not surprising: vascular endothelial growth factor A (VEGF-A), the major angiogenic factor, turned out to be the most prominent mediator strongly depending on CTGF.
Starting from the notion that HIF-1 is the major transcriptional regulator of VEGF, Chang et al. demonstrated by genetic manipulation that CTGF controls HIF-1 expression. The human lung adenocarcinoma cell line used in this part of the study (CL1-5) constitutively expresses high levels of HIF-1, as frequently occurs in transformed cells. Chang et al. found that disruption of HIF-1 protein stability is the mode of action of CTGF in this tumor type. Indeed, they found not only that HIF-1 expression and transcriptional activity were dramatically reduced in CTGF transfectants, whereas the opposite effect was seen in CTGF-silenced cells, but what's more, that CTGF increased the interaction of HIF-1 with pVHL. This effect appeared to be the consequence of a strong induction of ARD1 mRNA, leading to increased acetylation of HIF-1 at Lys 532 in CTGF transfectants. Accordingly, introduction of antisense ARD1 into CTGF-transfected cells restored HIF-1 expression.
A highlight of this finding is that three other negative regulators of HIF-1—pVHL, p53, and PTEN (phosphatase and tensin homologue deleted on chromosome 10)—are tumor suppressors. Importantly, ARD1 is a candidate negative regulator of HIF-1 under normoxic conditions, and its apparent accessory role has been questioned (8). Because CTGF has been shown to respond to metabolic alterations, such as hyperglycemia in diabetes (11), it is possible that different microenvironmental stimuli other than oxygen tension can recruit CTGF to the HIF-1 pathway to fine-tune its biologic activity in physiologic conditions.
Importantly, the negative control operated by CTGF on HIF-1 was sufficient to abolish both the angiogenic and the metastatic potential of CL1–5 cells in vivo and to increase survival time in mice with metastatic disease. Moreover, the authors showed an inverse association between CTGF expression and microvessel density in human lung adenocarcinoma samples.
VEGF as a representative biomarker of tumor angiogenesis has been variably implicated in lung adenocarcinoma progression (12); however, clinical application of VEGF inhibitors as single agents in solid tumors has not shown remarkable success (13), as they work in combination with standard chemotherapy (14). This finding is not unexpected, since neither VEGF alone nor HIF-1 itself are the unique effectors of tumor angiogenesis. The HIF-1–mediated antiangiogenic activity of CTGF reported by Chang et al. is striking; a previous study (15) showed that knocking down HIF-1 in colon cancer cells, despite halving VEGF expression levels, proved not to be antiangiogenic per se due to a compensatory overexpression of interleukin 8 driven by NF-
B. So it is possible that effective global antiangiogenic control could be more efficiently obtained by targeting master regulators of angiogenesis, including HIF-1, through activation of subtle posttranslational mechanisms.
Whether the findings reported by Chang et al. are restricted to lung adenocarcinoma remains an open question. In fact, the role of CTGF emerging from this and previous work by the same group (16) is in striking contrast with its assignment to a bone metastasis gene expression signature in breast cancer cells (17) or with a proangiogenic activity registered in other systems (see references in Chang's report, this issue (16)).
Intriguingly, CTGF seems to be a hypoxia-responsive gene, because CTGF mRNA stabilization by hypoxia has been reported in human chondrosarcoma cells (18), whereas HIF-1 can induce CTGF expression in normal mouse primary tubular epithelial cells (19). The reduction in HIF-1 protein stability by CTGF now described by Chang et al. suggests the existence of a complex feedback mechanism that, when subjected to tissue specific modulation, could account for the heterogeneous and even contrasting functions of CTGF in different organs.
In this regard, it is interesting that CTGF shares many functions, including stimulation of extracellular matrix deposition, with another prototypic two-faced janus molecule, transforming growth factor 
(TGF-). TGF- can exert both positive and negative roles in cancer (20); often associated with angiogenesis, induction of other members of this family has been associated with inhibition of angiogenesis (21). Cross-talk between TGF- and the HIF-1 can lead to a certain degree of cooperation in some systems (22). HIF-1 also has contradictory and even paradoxic roles—it can induce tumor cell death (23).
What are the factors that permit one molecule to have opposite functions in different tumor contexts? A cell- and tissue-specific threshold of susceptibility to microenvironmental changes could signal the accomplishment of diversified adaptive programs based on the modulation of key genes and proteins endowed with a dual function in the regulatory network. The elucidation of the interplay between central and tributary pathways in tumor progression, such as those highlighted here for the putative metastasis suppressor gene CTGF, will help in deciding which molecular target in the network will more specifically affect aberrant tumor angiogenesis without compromising the physiological process. Targeting the tumor microenvironment (24), in addition to the tumor cells and/or endothelial cells themselves (25), may permit important improvements to clinical outcomes in the future.
REFERENCES
(1) Coussens LM, Werb Z. Inflammation and cancer. Nature 2002;420:860–7.[CrossRef][Medline]
(2) Albini A, Tosetti F, Benelli R, Noonan DM. Tumor inflammatory angiogenesis and its chemoprevention. Cancer Res 2005;65:10637–41.
(3) Brahimi-Horn MC, Pouyssegur J. The hypoxia-inducible factor and tumor progression along the angiogenic pathway. Int Rev Cytol 2005;242:157–213.[ISI][Medline]
(4) Esteban MA, Maxwell PH. HIF, a missing link between metabolism and cancer. Nat Med 2005;11:1047–8.[CrossRef][ISI][Medline]
(5) Cramer T, Yamanishi Y, Clausen BE, Forster I, Pawlinski R, Mackman N, et al. HIF-1alpha is essential for myeloid cell-mediated inflammation. Cell 2003;112:645–57.[CrossRef][ISI][Medline]
(6) Semenza GL. Hypoxia, clonal selection, and the role of HIF-1 in tumor progression. Crit Rev Biochem Mol Biol 2000;35:71–103.[CrossRef][ISI][Medline]
(7) Semenza GL. HIF-1, O(2), and the 3 PHDs: how animal cells signal hypoxia to the nucleus. Cell 2001;107:1–3.[CrossRef][ISI][Medline]
(8) Jeong JW, Bae MK, Ahn MY, Kim SH, Sohn TK, Bae MH, et al. Regulation and destabilization of HIF-1alpha by ARD1-mediated acetylation. Cell 2002;111:709–20.[CrossRef][ISI][Medline]
(9) Chang C-C, Lin M-T, Lin B-R, Jeng Y-M, Chen S-T, Chu C-Y, et al. Effect of connective tissue growth factor on hypoxia-inducible factor 1 degradation and tumor angiogenesis. J Natl Cancer Inst 2006;98:984–95.
(10) Brigstock DR. The connective tissue growth factor/cysteine-rich 61/nephroblastoma overexpressed (CCN) family. Endocr Rev 1999;20:189–206.
(11) Lam S, van der Geest RN, Verhagen NA, van Nieuwenhoven FA, Blom IE, Aten J, et al. Connective tissue growth factor and IGF-I are produced by human renal fibroblasts and cooperate in the induction of collagen production by high glucose. Diabetes 2003;52:2975–83.
(12) Herbst R, Hidalgo M, Pierson A, Holden S, Bergen M, Eckhardt S. Angiogenesis inhibitors in clinical development for lung cancer. Semin Oncol 2002;29(1 Suppl 4):66–77.[CrossRef][ISI][Medline]
(13) Kerbel R, Folkman J. Clinical translation of angiogenesis inhibitors. Nat Rev Cancer 2002;2:727–39.[CrossRef][ISI][Medline]
(14) Ferrara N, Kerbel RS. Angiogenesis as a therapeutic target. Nature 2005;438:967–74.[CrossRef][Medline]
(15) Mizukami Y, Jo WS, Duerr EM, Gala M, Li J, Zhang X, et al. Induction of interleukin-8 preserves the angiogenic response in HIF-1alpha-deficient colon cancer cells. Nat Med 2005;11:992–7.[ISI][Medline]
(16) Chang CC, Shih JY, Jeng YM, Su JL, Lin BZ, Chen ST, et al. Connective tissue growth factor and its role in lung adenocarcinoma invasion and metastasis. J Natl Cancer Inst 2004;96:364–75.
(17) Kang Y, Siegel PM, Shu W, Drobnjak M, Kakonen SM, Cordon-Cardo C, et al. A multigenic program mediating breast cancer metastasis to bone. Cancer Cell 2003;3:537–49.[CrossRef][ISI][Medline]
(18) Kondo S, Kubota S, Mukudai Y, Moritani N, Nishida T, Matsushita H, et al. Hypoxic regulation of stability of connective tissue growth factor/CCN2 mRNA by 3’-untranslated region interacting with a cellular protein in human chondrosarcoma cells. Oncogene 2006;25:1099–110.[CrossRef][ISI][Medline]
(19) Higgins DF, Biju MP, Akai Y, Wutz A, Johnson RS, Haase VH. Hypoxic induction of CTGF is directly mediated by Hif-1. Am J Physiol Renal Physiol 2004;287:F1223–32.
(20) Pinkas J, Teicher BA. TGF-beta in cancer and as a therapeutic target. Biochem Pharmacol 2006 Mar 10 [Epub ahead of print].
(21) Ferrari N, Pfeffer U, Dell'Eva R, Ambrosini C, Noonan DM, Albini A. The transforming growth factor-beta family members bone morphogenetic protein-2 and macrophage inhibitory cytokine-1 as mediators of the antiangiogenic activity of N-(4-hydroxyphenyl)retinamide. Clin Cancer Res 2005;11:4610–9.
(22) Sanchez-Elsner T, Botella LM, Velasco B, Corbi A, Attisano L, Bernabeu C. Synergistic cooperation between hypoxia and transforming growth factor-beta pathways on human vascular endothelial growth factor gene expression. J Biol Chem 2001;276:38527–35.
(23) Piret JP, Mottet D, Raes M, Michiels C. Is HIF-1alpha a pro- or an anti-apoptotic protein? Biochem Pharmacol 2002;64:889–92.[CrossRef][ISI][Medline]
(24) Joyce JA. Therapeutic targeting of the tumor microenvironment. Cancer Cell 2005;7:513–20.[CrossRef][ISI][Medline]
(25) Tosetti F, Ferrari N, De Flora S, Albini A. Angioprevention: angiogenesis is a common and key target for cancer chemopreventive agents. FASEB J 2002;16:2–14.
CiteULike 
Connotea 
Del.icio.us What's this?
This article has been cited by other articles:
|
|
 |
|
  E. J. Kuiper, P. Roestenberg, C. Ehlken, V. Lambert, H. B. van Treslong-de Groot, K. M. Lyons, H.-J. T. Agostini, J.-M. Rakic, I. Klaassen, C. J.F. Van Noorden, et al. Angiogenesis Is Not Impaired in Connective Tissue Growth Factor (CTGF) Knock-out Mice J. Histochem. Cytochem., November 1, 2007; 55(11): 1139 - 1147. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||