Fibroblast Growth Factor 3 Research Articles

Fibroblast growth factor 3, a protein with a dual subcellular fate, is interacting with human ribosomal protein S2

The secreted isoform of fibroblast growth factor 3 (FGF3) induces a mitogenic cell response, while the nuclear form inhibits cell proliferation. Recently, we identified a nucleolar FGF3-binding protein which is implicated in processing of pre-rRNA as a possible target of nuclear FGF3 signalling. Here, we report a second candidate protein identified by a yeast two-hybrid screen for nuclear FGF3 action, ribosomal protein S2, rpS2. Recombinant rpS2 binds to in vitro translated FGF3 and to nuclear FGF3 extracted from transfected COS-1 cells. Characterization of the FGF3 binding domain of rpS2 showed that both the Arg-Gly-rich N-terminal region and a short carboxyl-terminal sequence of rpS2 are necessary for FGF3 binding. Mapping the S2 binding domains of FGF3 revealed that these domains are important for both NoBP and rpS2 interaction. Transient co-expression of rpS2 and nuclear FGF3 resulted in a reduced nucleolar localization of the FGF. These findings suggest that the nuclear form of FGF3 inhibits cell proliferation by interfering with ribosomal biogenesis.

Development of Novel Therapy for Achondroplasia: Use of Parathyroid Hormone

Achondroplasia (ACHs) and thanatophoric dysplasia (TD) are representative diseases of micromelic short stature that are elicited by a constitutively activated FGFR3 caused by point-mutation in the receptor molecule. Based on the mutated point in the receptor, there are several types of diseases of which the severity varies. In the chondrogenic cell line (ATDC5), the signaling cascade in the fibroblast growth factor 3 (FGFR3) was silenced by a overexpression of the mutated FGFR3 that resulted in decreased expression of the mRNA of the parathyroid hormone-related peptide (PTHrP); and also apoptosis was induced. The expression levels of PTHrP were inversely correlated with the severity of the disease; and the replacement of PTHrP prevented the apoptotic changes in the cell lines. In the following studies, however, we selected rhPTH that possesses a high homology and had been clinically used in the treatment of osteoporosis. In the organ culture, rhPTH remarkably elongated the proximal and distal cartilages of the ACH transgenic mice (ACHtg) and expressed collagen X similarly to the wild mice. An in vivo study of rhPTH significantly increased the growth of the long bones of ACHtg in comparison with those of the control (saline). The future prospects of parathyroid hormone (PTH) therapy of ACH are to be discussed.

Retinoblastoma and Hypochondroplasia: A Case Report of Two Germline Mutations Arising Simultaneously

Purpose: To report a rare case of a patient with two germline mutations arising de novo resulting in bilateral retinoblastoma and hypochondroplasia. Design: A brief review about retinoblastoma and hypochondroplasia; a case report with genetic mutational analysis results. Case report: We report a patient manifesting the clinical features of both bilateral retinoblastoma and hypochondroplasia. Genetic analysis revealed two germline mutations, a seven base-pair deletion in exon 12 (G70313-703129del) in one allele of the retinoblastoma gene (RB1) and the N540K (C1620C > A) mutation in one allele of the fibroblast growth factor 3 (FGFR3) gene, a frequent mutation in hypochondroplasia. Neither parent has a personal or family history of cancer or ocular tumors. Only the patient's mother is short in stature, and her genetic analysis revealed no FGFR3 mutations. Conclusions: Although the probability of both germline mutations occurring in a single individual is exceedingly low, the etiology and mechanism are unknown in this patient. To the best of our knowledge, this is the first report of two clinically distinct heritable germline mutations arising de novo in an individual.

Minimal expression of the proto-oncogene int-2 encoded protein in a series of colorectal carcinomas.

Int-2 (fibroblast growth factor-3) is a gene that belongs to the fibroblast growth factor gene family. It has been implicated in the carcinogenesis of several types of cancer, including esophageal squamous cell carcinoma, breast, head, and neck and lung carcinomas; but no firm data on its biological activity regarding neoplasms arising from the glandular epithelia of the gastrointestinal tract exists. In the present immunohistochemical study, we investigated the presence of int-2 encoded protein in a panel of 80 cases of colon carcinoma of various stages, grades and sizes. A sheep antihuman int-2 antibody was applied to paraffin-embedded tissue sections from the tumor samples. The percentage of int-2 immunostaining in the positively stained specimens was evaluated by image analysis. Int-2 was positively detected in only four tumors (i.e. 5% of the cases examined). All immunopositive cases were moderately differentiated tumors; the adjacent mucosa did not express int-2 protein. The relevant patients were male. These findings suggest that the role of int-2 in colorectal carcinogenesis is probably a limited one.

Inducible expression of FGF-3 in mouse mammary gland.

Fibroblast growth factor-3 (FGF-3) is a crucial developmental regulator. Aberrant activation of this gene by mouse mammary tumor virus insertion results in pregnancy-responsive mammary tumorigenesis. To characterize better FGF-3 function in postnatal mammary gland development and cancer initiation/progression, we used a mifepristone (RU486)-inducible regulatory system to express conditionally FGF-3 in the mammary epithelium of transgenic mice. Ectopic overexpression of FGF-3 in pubescent mammary glands elicited severe perturbations in early mammary gland development leading to mammary hyperplasia. Ductal elongation was retarded, multiple cysts persisted in the virgin ducts, and ductal epithelium was expanded and multilayered. The altered ductal architecture and the persistence of hyperplastic multilayered epithelium reflect a defect in growth regulation, which resulted from an imbalance between mitogenic and apoptotic signals. By altering the duration of RU486 treatment, we showed that the persistence of mitogenic signal elicited by FGF-3 is crucial for the initiation, progression, and maintenance of the hyperplastic characteristic of the mammary epithelium. The manifestations elicited by FGF-3 could be reversed by RU486 withdrawal. In addition, synergism between the stimulus from estrogen and FGF-3 mitogenic pathways was evident and likely contributes to the pregnancy-dependent tumorigenesis of FGF-3. Taken together, the mifepristone-inducible regulatory system provides a powerful means for understanding the diverse roles of FGF-3 and its interactions with hormones in mammary gland tumorigenesis.

Up-regulated gene expression of angiogenesis factors in post-chemotherapeutic lung cancer tissues determined by cDNA macroarray

The differential expressions of hundreds of tightly transcriptionally controlled genes in freshly isolated human lung cancers and respective normal lung tissues were analyzed by the cDNA macroarray technique. Three lung cancer patients received pre-operative chemotherapy with cisplatin containing regimens. After chemotherapy, these patients underwent surgery, and poly (A)-RNA expressions of 588 genes in the samples prepared from the lung cancer and normal lung tissues were compared. These expressions of the 588 genes were demonstrated by spotting onto a filter. Histogram analysis of gene expression revealed the tumors to show commonly up-regulated expression of angiogenesis and invasion related genes and adhesion molecules such as fibroblast growth factor 3 (FGFR3), matrix metalloproteinase (MMP)15, 16 and 10, integrin beta 4, integrin alpha 9, endonexin, and several types of collagens. Thus, post-chemotherapeutic tissues from lung cancer parents are characterized by remarkable up-regulation of molecules related to angiogenesis, invasion and adhesion. Tree view showed close clustering of angiogenesis related genes. Furthermore, when the angiogenesis related genes were selected and clustered, they were categorized into three groups depending upon gene expression profiles. These results suggest that angiogenesis related molecules are suitable candidates for target-based therapeutics and angiogenesis inhibitors are expected to be effective in lung cancer patients pretreated with chemotherapy.

A comparative study of the int-2 gene product in primary and secondary parathyroid lesions

The family of fibroblast growth factors stimulates proliferation of cells of mesenchymal, epithelial and neuroectodermal origin. One of the members of this family, the product of proto-oncogene int-2, fibroblast growth factor-3, has been found to stimulate mitosis of parathyroid cells in culture. Primary and secondary hyperparathyroidism have no clear differences with regard to the histopathological features of the diseased parathyroid glands. This study was undertaken in order to determine whether int-2 protein is immunohistochemically expressed in normal and abnormal parathyroid glands and to investigate whether there is a differential expression of the int-2 gene product between primary and secondary parathyroid disease. A sheep anti-human int-2 antibody was applied to tissue sections from 37 samples of primary parathyroid disease (12 sporadic adenomas, 25 hyperplastic glands), from 30 samples of renal hyperparathyroidism, and from seven normal controls. Int-2 immunostaining was evaluated semi-quantitatively. None of the normal parathyroid glands stained positively. Int-2 immunopositive expression was more frequently detected in specimens of uraemic patients than in those of patients with primary parathyroid growth processes (P=0.029). The reason for this differential expression appears to be the higher proportion of oxyphilic cells growing in hyperplastic glands of patients with secondary hyperparathyroidism; the latter cells were specifically found to be int-2 immunoreactive. The int-2 gene product is likely to participate in the proliferation of this parathyroid cell subpopulation.

NoBP, a nuclear fibroblast growth factor 3 binding protein, is cell cycle regulated and promotes cell growth.

Secreted and nuclear forms of fibroblast growth factor 3 (FGF3) have opposing effects on cells. The secreted form stimulates cell growth and transformation, while the nuclear form inhibits DNA synthesis and cell proliferation. By using the yeast two-hybrid system we have identified a nucleolar FGF3 binding protein (NoBP) which coimmunoprecipitated and colocalized with FGF3 in transfected COS-1 cells. Characterization of the NoBP binding domain of FGF3 exactly matched the sequence requirements of FGF3 for its translocation into the nucleoli, suggesting that NoBP might be the nucleolar binding partner of FGF3 essential for its nucleolus localization. Carboxyl-terminal domains of NoBP contain linear nuclear and nucleolar targeting motifs which are capable of directing a heterologous protein beta-galactosidase to the nucleus and the nucleoli. While NoBP expression was detected in all analyzed proliferating established cell lines, NoBP transcription was rapidly downregulated in the promyelocytic leukemia cell line HL60 when induced to differentiate. Analysis on the expression pattern of NoBP mRNA throughout the cell cycle in HeLa cells synchronized by lovastatin demonstrated a substantial upregulation during the late G(1)/early S phase. NoBP overexpression conferred a proliferating effect onto NIH 3T3 cells and can counteract the inhibitory effect of nuclear FGF3, suggesting a role of NoBP in controlling proliferation in cells. We propose that NoBP is the functional target of nuclear FGF3 action.

Induction of inner ear fate by FGF3.

Loss-of-function experiments in avians and mammals have provided conflicting results on the capacity of fibroblast growth factor 3 (FGF3) to act as a secreted growth factor responsible for induction and morphogenesis of the vertebrate inner ear. Using a novel technique for gene transfer into chicken embryos, we have readdressed the role of FGF3 during inner ear development in avians. We find that ectopic expression of FGF3 results in the formation of ectopic placodes which express otic marker genes. The ectopically induced placodes form vesicles which show the characteristic gene expression pattern of a developing inner ear. Ectopic expression of FGF3 also influences the formation of the normal orthotopic inner ear, whereas another member of the FGF family, FGF2, shows no effects on inner ear induction. These results demonstrate that a single gene can induce inner ear fate and reveal an unexpectedly widespread competence of the surface ectoderm to form sensory placodes in higher vertebrates.

Fibroblast growth factor receptor-3 as a marker for precartilaginous stem cells.

The epiphyseal organ contains two kinds of cartilage, articular and growth plate. Both enlarge during the growth phase of life. However, mitosis is not apparent in these tissues. In the current study, a search to trace the reservoirs of stem cells needed for the growth of these cartilages is done. A disorder in which the stem cells responsible for bone growth are mutated is achondroplasia; the mutation resides in the fibroblast growth factor receptor-3. Epiphyses stained with antifibroblast growth factor 3 antibodies reveal clusters of positively stained cells residing in the perichondrial mesenchyme, known as the ring of La Croix. Removal of the ring of La Croix causes a drastic growth arrest in the limbs of rat neonates. Cell cultures derived of the ring of La Croix biopsy specimens show high rates of cell proliferation and cell migration in vitro, in contrast to articular or growth plate derived chondrocytes. These cells stain intensely by antifibroblast growth factor receptor-3 antibodies and antiproliferative cells nuclear antigen, in contrast with articular and epiphyseal chondrocytes. Transfection of cells from the ring La Croix by an adenovirus vector containing the gene encoding for Escherichia coli beta-galactosidase (lacZ), allows tracing of these cells in tissues. Local injections were performed either to the ring of La Croix or to the joint cavity in a guinea pig model. A characteristic distribution was seen after injection. The transfected cells migrated to areas of bone and cartilage formation in the subchondral bone plate and on either side of the growth plate. This labeling and distribution is maintained for as many as 3 months after injection. The cells from the ring of La Croix appear to be responsible for bone growth. Furthermore, perichondrial cells and other precartilaginous cells expressing fibroblast growth factor-3 have been shown to be good cells for implantation to correct defects of articular cartilage.

Involvement of receptor-type tyrosine kinase gene families in cardiac hypertrophy.

The activation of protein tyrosine kinases (PTKs) has been postulated to be involved in cell differentiation and proliferation. To elucidate the involvement of tyrosine kinase genes in normal and pathological conditions, we analysed the expression patterns of receptor-type PTKs in the normal and hypertensive hypertrophied heart in rats. Hypertrophied and normal rat hearts were obtained from hypertensive rats; deoxycorticosterone acetate (DOCA)-salt and 2 kidney-1 clip (2K-1C), and their sham-operated rats, respectively. A reverse transcription-polymerase chain reaction (RT-PCR) was performed using degenerated primers which were designed from highly conserved regions in the catalytic domains of receptor-type PTKs. The PCR products were ligated into a sequence vector, and subcloned by transforming bacteria. To compare the expression level of these PTK mRNAs in the normal and hypertrophied heart, we performed semi-competetive RT-PCR and immunohistochemical and Western blot analyses. Nucleotide sequencing of approximately 80 clones of PTKs revealed 10 receptor-type, five nonreceptor-type and two unknown types in the rat heart. Tie-2/Tek, Ryk, insulin-like growth factor-I receptor were abundantly expressed in the rat heart as members of receptor-type PTKs. Immunohistochemistry and RT-PCR demonstrated the presence of platelet-derived growth factor (PDGF)-alpha receptor, PDGF-beta receptor and fibroblast growth factor-3 receptor in both normal and hypertrophied hearts. We also confirmed the presence of Flt-1, KDR/FIk-1, and their ligand vascular endothelial growth factor, c-Met and its ligand hepatocyte growth factor (HGF), and Tie-1, Tie-2/Tek by immunohistochemistry and RT-PCR. The coexpression of cardiac HGF and c-Met in hypertrophied hearts, especially in 2K-1 C rats, was induced more intensively than that in DOCA-salt rats. These findings suggest that HGF/c-Met interactions may play an important role in cardiac hypertrophy and remodeling, probably as a result of the activation of the local renin-angiotensin system.

Inhibitory effect of suramin in rat models of angiogenesis in vitro and in vivo.

The aim of the present study was to test the ability of the chemotherapeutic agent suramin to inhibit angiogenesis in experimental models in vitro and in vivo. In the culture of rat aortic rings on fibronectin, suramin dose-dependently inhibited vascular cell growth, achieving the maximal effect (mean - 88% versus controls, P < 0.05) at 400 microg/ml. Image analysis showed that suramin could inhibit microvessel sprouting in fibrin from rat aortic rings as evaluated by the ratio between the cellular area and the mean gray value of the sample (sprouting index); suramin at 50 microg/ml significantly reduced the sprouting index from the control value of 0.35+/-0.04 to 0.14+/-0.02 mm2/gray level (P < 0.05). Likewise, the area occupied by cells was 19.2+/-1.8 mm2 as compared with 41.8+/-4.2 mm2 in controls (P < 0.05). In the rat model of neovascularization induced in the cornea by chemical injury, suramin at 1.6 mg/eye per day reduced the length of blood vessels (0.7+/-0.1 mm as compared with 1.5+/-0.1 mm in controls, P < 0.05). In the same model the ratio between the area of blood vessels and the total area of the cornea (area fraction score) was decreased by suramin from 0.19+/-0.02 in controls to 0.03+/-0.003 (P < 0.05). Suramin given i.p. at 30 mg/ kg per day markedly inhibited the neovascularization induced in the rat mesentery by compound 48/80 or conditioned medium from cells secreting the angiogenic protein fibroblast growth factor-3 (FGF-3). The area fraction score in control rats treated with compound 48/80 was 0.31+/-0.03, and this was reduced to 0.07+/-0.01 by suramin (P < 0.05). After i.p. administration of FGF-3 the area fraction score was reduced by suramin from 0.29+/-0.03 to 0.05+/-0.01 (P < 0.05). These results provide evidence that suramin exerts inhibitory effects on angiogenesis in both in vitro and in vivo models.

Evaluation of the tumorigenic and angiogenic potential of human fibroblast growth factor FGF3 in nude mice.

Recently, the expression of fibroblast growth factor 3 (FGF3) was found in 55% of human Kaposi's sarcoma (KS) tumor tissues examined, while almost no expression of FGF3 was found in normal skin. To further these studies, human FGF3 cDNA were constructed by the overlap-extension method. The proteins translated from two FGF3 cDNA, which differ only in the sequences preceding the AUG presumed to be the initiation codon, were shown to have the same molecular mass. This result suggests that translation of human FGF3, which is different from mouse FGF3, begins only at the AUG site. The human FGF cDNA was transfected into NIH3T3 cells. The NIH 3T3 cells transformed by FGF3 were then injected subcutaneously into athymic nude mice. Nodular lesions developed at the injection sites in all seven mice injected with the F3-1 cell clone, which showed high expression of FGF3, and in two out of six mice injected with the F3-2 cell clone, which expressed a low level of FGF3. Histopathological features of these tumors contained fascicles of spindle-shaped cells surrounding irregular endothelial lined vascular clefts, similar to those observed in human KS lesions. Immunohistochemical staining for factor V111 antigen revealed reactivity in multiple areas, especially in abundant vascular structures of the tumor sections examined. The expression of FGF3 together with the FGF receptors FGFR1, FGFR2, and FGFR3, was detected in the mouse tumors by Northern blot analysis. Our results indicate that tumors induced by FGF3-transformed NIH3T3 cells show some similarities to human KS tumors. In conclusion, our results demonstrate the potential tumorigenic and angiogenic role of human FGF3.

Over-expression of epidermal growth factor receptor and c-erbB2/neu but not of int-2 genes in benign prostatic hyperplasia by means of semi-quantitative PCR.

It is well established that activation of cellular genes may trigger uncontrolled cell growth and cancer development. Previous reports suggest that the epidermal growth factor receptor (EGFr), c-erbB2/neu and int-2, fibroblast growth factor-3, may be implicated in the development of benign prostatic hyperplasia (BPH). Using the polymerase chain reaction technique, we have assessed the amplification and expression of these molecular markers in 30 prostate samples from patients with BPH as well as from 5 normal donors. We detected mRNA over-expression of EGFr and c-erbB2/neu in 36% and 63%, respectively, of the BPH samples, but no gene amplification was found. No amplification or over-expression of int-2 was detected in any of the samples analyzed, suggesting that int-2 is not involved in BPH. Our results thus suggest a role for EGFr and c-erbB2/neu but not for int-2 in the development of BPH.

Fibroblast Growth Factor 3, a Protein with Dual Subcellular Localization, Is Targeted to the Nucleus and Nucleolus by the Concerted Action of Two Nuclear Localization Signals and a Nucleolar Retention Signal

The major isoform of fibroblast growth factor 3 (FGF3) is initiated from a CUG codon, and the resultant product is distributed to the nucleus/nucleolus and secretory pathway. This dual subcellular localization is achieved in part by the competing effects of two classical intracellular targeting signals located near the amino terminus. At the extreme amino terminus is a short stretch of 29 amino acids before a signal peptide necessary for translocation into the endoplasmic reticulum, which is next to an adjacent bipartite nuclear localization signal. The carboxyl-terminal region of FGF3 is also implicated in nuclear/nucleolar localization. We describe here the characterization of carboxyl-terminal signals by showing they are capable of directing a heterologous protein, beta-galactosidase, to the nucleus. Furthermore, appending both the amino- and carboxyl-terminal domains onto beta-galactosidase, reproduces the dual subcellular localization properties of FGF3. Nuclear uptake of FGF3 appears to be signal-mediated since it binds to karyopherin alpha, the nuclear localization signal binding subunit of a heterodimeric receptor of the nuclear import machinery. The import of FGF3 into the nucleus is energy-dependent, and the inhibition of this process has demonstrated the importance of the nucleolar retention signal in nucleoplasmic and nucleolar accumulation.

Achondroplasia: recent advances in diagnosis and treatment.

Achondroplasia (ACH) is the most common form of chondrodysplasia in humans. This disorder is inherited as an autosomal dominant trait, though most cases are sporadic. Recent advances in molecular biology have revealed its genetic defect in fibroblast growth factor-3 gene. This may introduce a new diagnostic tool and the classification of ACH and related disorders. Recent molecular engineering techniques have made it possible to provide large amounts of the various kinds of biofactors, such as erythropoietin, granulocyte colony stimulating factor and human growth hormone (GH), for clinical use. In fact, GH has been widely used to treat non-GH-deficient forms of short stature, such as Turner's syndrome, skeletal dysplasia, intrauterine growth retardation, chronic illness and idiopathic short stature, with beneficial effects. This may also be introduced into the medical management of ACH.

The zebrafish Fgf-3 gene: cDNA sequence, transcript structure and genomic organization

We report the isolation and characterization of genomic and cDNA clones encoding zebrafish fibroblast growth factor 3 (FGF3). An initial cDNA clone was generated by PCR amplification using degenerate oligo primers corresponding to a conserved region of protein found in the mouse and human homologues. Screening a cDNA library made from 30–33-h-old zebrafish embryos with this PCR product led to the isolation of two cDNA clones. Sequence analysis of the longest cDNA insert (1810 bp) revealed a 256-amino-acid (aa) orf. The central region, composed of approx. 155 aa, shares 78% identity with the analogous region of Xenopus laevis FGF3 and 72% identity with the product of the more distantly related human gene. However, the N-and C-terminal domains of zebrafish FGF3 are very different from those of other known homologues. The cDNA was used as a probe on genomic DNA to create a physical map of the locus and to isolate a genomic clone encompassing the entire coding region and 5′ sequences. DNA sequencing and RNase protection analyses indicate that zebrafish Fgf-3 (ZFgf-3) is structurally analogous to the mouse gene and regulated through two different promoters. The transcription start point of the proximal promoter aligns to that of mouse promoter P3 and lies within a conserved region of sequence.

Fibroblast growth factor 3 is tumorigenic for mouse mammary cells orthotopically implanted in nude mice

Fibroblast growth factor-3 (Fgf-3) is involved in mouse mammary tumorigenesis since the Fgf-3 gene is a main target for mouse mammary tumor virus (MMTV) insertional activation. Its action has been correlated with the appearance of pregnancy-dependent tumors. We describe here the effects on normal mouse mammary EF43 cells of the short Fgf-3 protein form which enters the secretory pathway. The genes, Fgf-3 AUG or Fgf-4 for comparison, were introduced in the mammary cells by means of retroviral vectors. Fgf-3 expression did not modify EF43 cell morphology, had no effect on growth in soft agar nor on the inhibitory action exerted on cell growth by TGF-beta 1; however, it allowed the cells to grow under low serum conditions in the absence of insulin and EGF. The Fgf-3-expressing cells were not tumorigenic in nude mice when injected s.c., but tumors developed when the cells were implanted in the mammary gland. The tumors appeared after some latency; they had a slow growth phase followed by a phase of increased growth rate. An identical tumoral growth pattern was observed in ovariectomized nude mice. These results show that the secreted Fgf-3 form can initiate tumorigenesis and that the induced tumors are hormone-independent. The mammary-gland environment, however, is required for the EF43 cells to grow and differentiate. During that process, which resembles natural cell growth during mammary-gland development at pregnancy, the cells could pass through a stage which is specifically sensitive to Fgf-3.

Nucleolar association of fibroblast growth factor 3 via specific sequence motifs has inhibitory effects on cell growth.

The dual subcellular fate of fibroblast growth factor 3 (FGF3) is determined by the competing effects of amino-terminal signals for nuclear localization and secretion (P. Kiefer, P. Acland, D. Pappin, G. Peters, and C. Dickson, EMBO J. 13:4126-4136, 1994). Mutation analysis has implicated additional basic domains in the carboxy-terminal region of the protein as necessary for nuclear uptake and the association of FGF3 with the nucleoli. Immunogold electron microscopy shows that FGF3 is predominantly within the dense fibrillar component of the nucleolus. A form of FGF3 that localizes exclusively in the nucleus and nucleolus was generated by removing signals for secretion, and expression of this nonsecreted FGF3 in a mammary epithelial cell line resulted in slowly growing colonies of enlarged cells. Thus, nuclear import and nucleolar association of FGF3 are determined by the concerted interaction of several distinct motifs, and the exclusive production of the nuclear isoform can inhibit DNA synthesis and cell proliferation.

Detection of c-erbB-2/neu and fibroblast growth factor-3/INT-2 but not epidermal growth factor receptor gene amplification in endometrial cancer by differential polymerase chain reaction.

It currently is well established that the activation of protooncogenes could trigger uncontrolled cell growth and cancer development. Although this correlation already has been evidenced in several human tumors, no conclusive studies have related oncogene activation with the development of endometrial neoplasia. Nevertheless, few reports suggest that c-erbB-2/neu, which is a prognostic marker in breast cancer, epidermal growth factor receptor (EGFR), which is overexpressed in a variety of neoplasms, and fibroblast growth factor-3 (FGF-3/INT-2), which has been found to be amplified in breast and ovarian cancer, could be implicated in the development of endometrial adenocarcinoma. Amplification of c-erbB-2/neu, EGFR, and FGF-3/INT-2 was examined in 50 endometrial carcinomas, 10 adenomatous hyperplasias, and 50 normal endometrial samples, using the genomic differential polymerase chain reaction with the single copy reference gene interferon-gamma. Quantitation of the ratios between the amplified bands was assessed by image analysis. c-erbB-2 and FGF-3/INT-2 were amplified in a first group of 7 (14%) and a further group of 7 (14%) patients, respectively, of a total of 50 in whom endometrial cancer had been studied. In the latter seven, a strong correlation between this genetic marker and an advanced disease stage (International Federation of Gynecology and Obstetrics Stage III) was found. In two patients, both genes were amplified. No EGFR gene amplification was detected in any case. c-erbB-2/neu but not EGFR gene amplification was detected and FGF-3/INT-2 amplification and advanced disease were correlated in endometrial cancer.