Endocrinology, doi:10.1210/en.2006-0926
Endocrinology Vol. 148, No. 3 932-935
Copyright © 2007 by The Endocrine Society
The Paired Box-8/Peroxisome Proliferator-Activated Receptor-
Oncogene in Thyroid Tumorigenesis
Honey V. Reddi,
Bryan McIver,
Stefan K. G. Grebe and
Norman L. Eberhardt
Department of Medicine (H.V.R., B.M., N.L.E.), Division of Endocrinology, and Departments of Laboratory Medicine and Pathology (S.K.G.G.) and Biochemistry and Molecular Biology (N.L.E.), Mayo Clinic and Foundation, Rochester, Minnesota 55905
Address all correspondence and requests for reprints to: Norman L. Eberhardt, Ph.D., Department of Medicine/Division of Endocrinology, 200 First Street SW, Mayo Clinic, Rochester, Minnesota 55905. E-mail: eberhardt{at}mayo.edu.
 |
Abstract
|
|---|
The American Cancer Society estimates 30,180 new cases of thyroid cancer in the United States in 2006. Of all thyroid cancers, 15–20% are follicular thyroid carcinoma (FTC), making this the second most common thyroid malignancy (after papillary carcinoma). A proportion of FTC has been found to be associated with a chromosomal translocation, t (2, 3)(q13;p25), which fuses the thyroid-specific transcription factor paired box-8 with the peroxisome proliferator-activated receptor-
nuclear receptor, a ubiquitously expressed transcription factor. This fusion event causes expression of a paired box-8/peroxisome proliferator-activated receptor- fusion protein (PPFP). PPFP is detected in approximately 30% of FTC. In this report we review data on the role of PPFP in FTC, its mechanism of oncogenesis, and PPFP targeting as a strategy in thyroid cancer treatment.
|
Introduction
|
|---|
FOLLICULAR CELL-DERIVED thyroid carcinoma comprises several morphological subtypes: papillary (PTC), follicular (FTC), Hürthle-cell (HCC), and anaplastic (ATC) carcinomas. These subtypes are phenotypically distinct and exhibit extremes of malignant potential, from relatively indolent (PTC) to highly aggressive (ATC). FTC might arise in an adenoma-carcinoma sequence from follicular adenoma (FA), some of which are cytologically difficult to distinguish from FTC (1). The pathogenesis of HCC remains unclear, and there is controversy whether it represents a distinct morphotype or evolves on the background of PTC or FTC. FTC accounts for approximately 10–20% of all thyroid cancers and up to 40% of cause-specific deaths. ATC is the rarest but most aggressive thyroid cancer, with more than 90% 1 yr mortality, and probably arises from preexisting PTC, FTC, or HCC by acquisition of p53 mutation/inactivation, impaired Rb pathway function, or abnormalities in ß-catenin signaling (1).
No signature oncogenic pathways have been identified in FTC until recently. RAS mutations are found in a subset of these tumors, but they have not been tied conclusively to a particular signaling pathway. However, the last 6 yr have seen the identification and partial molecular-mechanistic examination of a novel oncogenic fusion gene (2) in FTC. The resulting fusion protein, paired box-8 (PAX8)/peroxisome proliferator-activated receptor (PPAR)- fusion protein (PPFP) has been identified in almost 50% of FTC and a much smaller proportion of its putative precursor lesion, FA. It is absent in PTC and HCC, suggesting that PPFP may represent an early FTC-specific oncogene (2, 3, 4, 5, 6, 7). A point to note is that in FTC, PAX8/PPAR fusion and Ras gene activations rarely overlap in the same tumor, providing additional evidence that PPAR-driven tumors represent a separate subset of tumors.
The absence of specific molecular markers available for the detection, diagnosis, or prognosis of FTC and the presence of PAX8/PPAR rearrangements in a major proportion of FTC raises the possibility that study of this gene could provide insight into the main oncogenic pathways guiding FTC pathogenesis, ultimately improving the diagnosis, staging and treatment of FTC.
|
PPFP: A Putative Oncogene for FTC
|
|---|
PPFP is a somatic tumor genetic rearrangement, wherein most of the (long) q-arm of chromosome 2 is translocated to the (short) p-arm of chromosome 3, whereas in exchange the 3p25 terminal region is translocated to 2q13. This translocation creates a fusion transcript wherein the 5'-region of the thyroid-specific transcription factor pax8 gene (2q13) is fused in frame with exon1 of the PPAR gene (3p25), a member of the thyroid hormone-steroid hormone nuclear receptor superfamily (2). The pax8 promoter, which is active in thyroid follicular cells, appears to drive expression of PPFP (8).
The predicted fusion protein and its putative functional domain structure are depicted in Fig. 1
. PAX8 is a member of the paired-box family of transcription factors, which is necessary for normal thyroid development (9), whereas PPAR is involved in adipocyte differentiation and lipid and carbohydrate metabolism as well as cell proliferation and differentiation (reviewed in Ref. 10).
PPFP encompasses an N-terminal PAX8 fragment, which lacks only the essential PAX8 transactivation domain (11, 12), whereas the PPAR component is apparently full length, with no reported mutations, deletions, or other alterations. To determine how PPFP might function as an oncogene, it is important to understand whether these domains are functionally intact or sterically hindered in this abnormal protein.
Various studies have demonstrated PPFP expression in about 25–70% FTC (a cumulative prevalence of 
30%), using either immunohistochemistry or RT-PCR (reviewed in Ref. 13), supporting the initial findings of Kroll et al. (2), that among malignant thyroid tumors, PPFP expression is a common and specific feature of FTC. Small, but significant rates of PPFP expression have been seen in FA at a lower frequency than FTC, suggesting that PPFP may be involved early in the neoplastic process, possibly even at the premalignant level. Whereas immunohistochemistry appears to be the most widely used method of PPFP detection, RT-PCR is used to actually detect the fusion points in the tumors because break points in PPFP may be localized not only after exon 9 but also after exons 7 and 8 of PAX8. Attempts to correlate PPFP expression in FTC with clinical presentation and outcome variables have generated conflicting results, although there is some suggestion that PPFP might characterize a less aggressive FTC subtype (14).
With regard to PPFP’s oncogenic mechanism, studies in our laboratory have demonstrated that transient or constitutive expression of full-length PPFP in immortalized human thyrocytes generated a growth advantage, compared with cells expressing control vector (15), supporting an oncogenic role for PPFP, an observation that has since been reproduced by others (16). We believe that the impact of PPFP on the observed increase in cell numbers is mediated through a substantially reduced rate of apoptosis and to a lesser effect on accelerated cell cycle kinetics (15). In addition to increased cell growth, PPFP expression was also associated with a significant increase in both the rate of soft agar colony formation (15), arguing strongly that PPFP is a true transforming agent in these cells. In general, these properties induced by an oncogene are useful markers of malignant potential.
No studies have yet demonstrated a primary transforming capacity for PPFP in vitro or in vivo. All in vitro experiments have been conducted in cell lines that are already immortalized (15, 16). Whether some preexisting oncogenic milieu is required for PPFP oncogene action remains unknown. Development of an animal model with thyroid expression of PPFP should clarify whether PPFP is sufficient on its own to precipitate follicular neoplasia or whether additional steps are required in that process.
|
Mechanism(s) of PPFP Action
|
|---|
Chromosomal translocations creating fusion genes could cause oncogenesis through a variety of mechanisms. The translocation could generate an abnormal pathogenic fusion protein, functioning in its own right as an entirely novel oncogene. Alternatively, deregulation of the genes involved in the translocation might abrogate their endogenous function, leading to oncogenesis, if one of these genes is a tumor suppressor. A third alternative, seen most prominently in PTC, is the overexpression of a protooncogene caused by alteration of a gene promoter system.
One possible mechanism of PPFP function could be modulation of the downstream pathways of one or other of its independent components, PAX8 and PPAR. PAX8 is required for normal thyroid development and is involved in the maintenance of differentiated follicular cell function. However, as is typical for genes from the pax family, its expression is tightly regulated and controlled, so aberrant expression of PPFP might have oncogenic effects, simply by down-regulating endogenous PAX8 expression, akin to that observed for many other fusion genes from the pax family (17).
PPAR, on the other hand, is a nuclear hormone receptor transcription factor that is expressed at very low levels in the thyroid and has no as yet identified function in that organ. In vitro studies indicate that PPFP function is mediated at least in part by inhibition of wild-type PPAR function (2, 15, 16) or down-regulation of wild-type PPAR expression (18). Data indicate that although ligand binding and activation of the ligand-gated activation function 2 domain increase the transcriptional activity of PPAR, these same processes also induce ubiquitination and subsequent proteosomal degradation (19), providing a negative feedback system to balance PPAR activity. It is possible that PPFP interferes in this delicate balance, pushing the cell toward proliferation (Fig. 2).
Gene transcription by PPAR requires the formation of a heterodimer with retinoic X receptor (RXR), followed by binding to a PPAR response element. Transactivation of the heterodimer is activated by ligands of PPAR, RXR, or both. Because PPFP has the ability to independently bind RXR (16), it is possible that PPFP competes with wild-type PPAR for RXR, DNA binding sites, or both, thus preventing wild-type PPAR-initiated transcriptional regulation (Fig. 2
). In addition, the PPFP-RXR complex might still recruit cofactors, which are bound, but do not initiate transactivation, further diminishing the necessary resources for wild-type PPAR action.
Analysis of gene expression array data of FTC that express PPFP demonstrated that these cancers have a distinct transcriptional signature (20, 21, 22). PPFP expression up-regulates genes associated with signal transduction, cell growth, and translational control, whereas a large number of ribosomal protein and translational associated genes are concurrently underexpressed. With respect to its ability to function as a novel protein, Giordano et al. (20) demonstrated that PPFP has unique transcriptional activities. They also demonstrated that PPFP has the potential to function in ways qualitatively similar to PAX8 or PPAR, depending on the promoter and cellular environment (20). PPFP has been shown to disrupt normal transcriptional pathways of PAX8 in a cell type-specific manner (16), presumably through some form of negative feedback. Based on these observations, the hypothesis that PPFP functions primarily through the control of PPAR requires reevaluation. Further analysis of the downstream regulatory pathways of PPFP will be essential as we begin to design strategies to intervene in the fundamental mechanisms of FTC.
|
PPFP as a Therapeutic Target
|
|---|
Currently therapeutic options for patients diagnosed with FTC include surgery followed in most cases by radioactive iodine treatment and thyroid hormone-suppressive therapy. These methods, however, are ineffective in the treatment of patients with metastatic cancer, in whom traditional chemotherapy is also usually ineffective (23). Consequently, novel therapeutic approaches are needed for these patients.
Molecular therapeutic approaches for FTC might be designed to target PPFP, keeping in mind the minimal knowledge available regarding PPFP-mediated oncogenesis (13). Because modulation of PAX8 and PPAR transcriptional pathways appears to be PPFP’s primary mode of action, specific intervention might target one or more of those pathways, raising the possibility for highly selective, targeted therapy. Modulators of the PPAR pathway (retinoic acid derivatives as well as thiazolidenediones) are already in clinical use for other diseases, generating hope that rapid progress can be made in the development of effective therapies that target the PPFP effect. PPAR agonists have been shown to be modestly effective against ATC cell lines in vitro (24), raising the level of optimism for their function in FTC. The effect of PPFP on PAX8 might be altered by redifferentiation therapy with retinoic acid, an approach that has been tried in the past and found to be marginally useful (25). PPFP itself could be directly targeted using small-molecule compounds or monoclonal humanized antibodies against unique epitopes of the protein. The discovery of PPFP in FTC has not only generated hope for the early diagnosis of FTC but also gives us the opportunity to develop effective strategies based on an understanding of the disease process.
|
Footnotes
|
|---|
This work was supported by National Institutes of Health Grant CA080117 and the Mayo Foundation.
First Published Online August 31, 2006
Abbreviations: ATC, Anaplastic thyroid carcinoma; FA, follicular adenoma; FTC, follicular thyroid carcinoma; HCC, Hürthle-cell carcinoma; PAX8, paired box-8; PPAR, peroxisome proliferator-activated receptor; PPFP, PAX8/PPAR fusion protein; PTC, papillary thyroid carcinoma; RXR, retinoic X receptor.
Received July 11, 2006.
Accepted for publication August 24, 2006.
|
References
|
|---|
- Goellner JR, Gharib H, Grant CS, Johnson DA 1987 Fine needle aspiration cytology of the thyroid, 1980 to 1986. Acta Cytol 31:587–590[Medline]
- Kroll TG, Sarraf P, Pecciarini L, Chen CJ, Mueller E, Spiegelman BM, Fletcher JA 2000 PAX8-PPAR1 fusion oncogene in human thyroid carcinoma. Science 289:1357–1360[Abstract/Free Full Text]
- Nikiforova MN, Lynch RA, Biddinger PW, Alexander EK, Dorn GW, Tallini G, Kroll TG, Nikiforov YE 2003 RAS point mutations and PAX8-PPAR rearrangement in thyroid tumors: evidence for distinct molecular pathways in thyroid follicular carcinoma. J Clin Endocrinol Metab 88:2318–2326[Abstract/Free Full Text]
- Nikiforova MN, Biddinger PW, Caudill CM, Kroll TG, Nikiforov YE 2002 PAX8-PPAR rearrangement in thyroid tumors: RT-PCR and immunohistochemical analyses. Am J Surg Pathol 26:1016–1023[CrossRef][Medline]
- Marques AR, Espadinha C, Catarino AL, Moniz S, Pereira T, Sobrinho LG, Leite V 2002 Expression of PAX8-PPAR1 rearrangements in both follicular thyroid carcinomas and adenomas. J Clin Endocrinol Metab 87:3947–3952[Abstract/Free Full Text]
- French CA, Alexander EK, Cibas ES, Nose V, Laguette J, Faquin W, Garber J, Moore Jr F, Fletcher JA, Larsen PR, Kroll TG 2003 Genetic and biological subgroups of low-stage follicular thyroid cancer. Am J Pathol 162:1053–1060[Abstract/Free Full Text]
- Thomas GA, Vecchio G, Williams GH, Fusco A, Chiappetta G, Pozcharskaya V, Bogdanova TI, Demidchik EP, Cherstvoy ED, Voscoboinik L, Tronko ND, Carss A, Bunnell H, Tonnachera M, Parma J, Dumont JE, Keller G, Hofler H, Williams ED 2000 High prevalence of RET/PTC rearrangements in Ukrainian and Belarussian post-Chernobyl thyroid papillary carcinomas: a strong correlation between RET/PTC3 and the solid-follicular variant. Br J Cancer 82:315–322[CrossRef][Medline]
- Mascia A, Nitsch L, Di Lauro R, Zannini M 2002 Hormonal control of the transcription factor Pax8 and its role in the regulation of thyroglobulin gene expression in thyroid cells. J Endocrinol 172:163–176[Abstract]
- Pasca di Magliano M, Di Lauro R, Zannini M 2000 Pax8 has a key role in thyroid cell differentiation. Proc Natl Acad Sci USA 97:13144–13149[Abstract/Free Full Text]
- Fajas L, Debril MB, Auwerx J 2001 PPAR: an essential role in metabolic control. Nutr Metab Cardiovasc Dis 11:64–69[Medline]
- Poleev A, Okladnova O, Musti AM, Schneider S, Royer-Pokora B, Plachov D 1997 Determination of functional domains of the human transcription factor PAX8 responsible for its nuclear localization and transactivating potential. Eur J Biochem 247:860–869[Medline]
- Poleev A, Wendler F, Fickenscher H, Zannini MS, Yaginuma K, Abbott C, Plachov D 1995 Distinct functional properties of three human paired-box-protein, PAX8, isoforms generated by alternative splicing in thyroid, kidney and Wilms’ tumors. Eur J Biochem 228:899–911[Medline]
- McIver B, Grebe SK, Eberhardt NL 2004 The PAX8/PPAR fusion oncogene as a potential therapeutic target in follicular thyroid carcinoma. Curr Drug Targets Immune Endocr Metabol Disord 4:221–234[CrossRef][Medline]
- Sahin M, Allard BL, Yates M, Powell JG, Wang XL, Hay ID, Zhao Y, Goellner JR, Sebo TJ, Grebe SK, Eberhardt NL, McIver B 2005 PPAR staining as a surrogate for PAX8/PPAR fusion oncogene expression in follicular neoplasms: clinicopathological correlation and histopathological diagnostic value. J Clin Endocrinol Metab 90:463–468[Abstract/Free Full Text]
- Gregory Powell J, Wang X, Allard BL, Sahin M, Wang XL, Hay ID, Hiddinga HJ, Deshpande SS, Kroll TG, Grebe SK, Eberhardt NL, McIver B 2004 The PAX8/PPAR fusion oncoprotein transforms immortalized human thyrocytes through a mechanism probably involving wild-type PPAR inhibition. Oncogene 23:3634–3641[CrossRef][Medline]
- Au AY, McBride C, Wilhelm Jr KG, Koenig RJ, Speller B, Cheung L, Messina M, Wentworth J, Tasevski V, Learoyd D, Robinson BG, Clifton-Bligh RJ 2006 PAX8-peroxisome proliferator-activated receptor (PPAR) disrupts normal PAX8 or PPAR transcriptional function and stimulates follicular thyroid cell growth. Endocrinology 147:367–376[Abstract/Free Full Text]
- Barr FG 1997 Chromosomal translocations involving paired box transcription factors in human cancer. Int J Biochem Cell Biol 29:1449–1461[CrossRef][Medline]
- Fuhrer D 2001 A nuclear receptor in thyroid malignancy: is PAX8/PPAR the Holy Grail of follicular thyroid cancer? Eur J Endocrinol 144:453–456[CrossRef][Medline]
- Hauser S, Adelmant G, Sarraf P, Wright HM, Mueller E, Spiegelman BM 2000 Degradation of the peroxisome proliferator-activated receptor is linked to ligand-dependent activation. J Biol Chem 275:18527–18533[Abstract/Free Full Text]
- Giordano TJ, Au AYM, Kuick R, Thomas DG, Rhodes DR, Wilhelm KG, Vinco M, Misek DE, Sanders D, Zhu ZW, Ciampi R, Hanash S, Chinnaiyan A, Clifton-Bligh RJ, Robinson BG, Nikiforov YE, Koenig RJ 2006 Delineation, functional validation, and bioinformatic evaluation of gene expression in thyroid follicular carcinomas with the PAX8-PPAR translocation. Clin Cancer Res 12:1983–1993[Abstract/Free Full Text]
- Lui WO, Foukakis T, Liden J, Thoppe SR, Dwight T, Hoog A, Zedenius J, Wallin G, Reimers M, Larsson C 2005 Expression profiling reveals a distinct transcription signature in follicular thyroid carcinomas with a PAX8-PPAR fusion oncogene. Oncogene 24:1467–1476[CrossRef][Medline]
- Lacroix L, Lazar V, Michiels S, Ripoche H, Dessen P, Talbot M, Caillou B, Levillain JP, Schlumberger M, Bidart JM 2005 Follicular thyroid tumors with the PAX8-PPAR1 rearrangement display characteristic genetic alterations. Am J Pathol 167:223–231[Abstract/Free Full Text]
- Karavitaki N, Vlassopoulou V, Tzanela M, Tzavara I, Thalassinos N 2002 Recurrent and/or metastatic thyroid cancer: therapeutic options. Expert Opin Pharmacother 3:939–947[CrossRef][Medline]
- Copland JA, Marlow LA, Kurakata S, Fujiwara K, Wong AK, Kreinest PA, Williams SF, Haugen BR, Klopper JP, Smallridge RC 2006 Novel high-affinity PPAR agonist alone and in combination with paclitaxel inhibits human anaplastic thyroid carcinoma tumor growth via p21WAF1/CIP1. Oncogene 25:2304–2317[CrossRef][Medline]
- Park JW, Clark OH 2004 Redifferentiation therapy for thyroid cancer. Surg Clin North Am 84:921–943[CrossRef][Medline]
Top
Abstract
Introduction
