| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
Departments of 1 Head and Neck Surgery, 2 Biostatistics and Applied Mathematics, and 3 Cancer Biology, University of Texas M.D. Anderson Cancer Center, Houston, Texas
Requests for reprints: Jeffrey N. Myers, Department of Head and Neck Surgery, University of Texas M.D. Anderson Cancer Center, Unit 441, 1515 Holcombe Boulevard, Houston, TX 77030-4009. Phone: 713-792-6920; Fax: 713-794-4662. E-mail: jmyers{at}mdanderson.org
 |
Abstract |
|---|
 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
6- and 8-fold, respectively, compared with the control group. The microvessel density within the ATC xenografts was decreased by >80% in the mice treated with AEE788 alone and in combination with paclitaxel compared with the control group. Lastly, immunofluorescence microscopy showed the inhibition of EGFR autophosphorylation on the tumor cells as well as the inhibition of VEGFR-2 autophosphorylation on tumor endothelium. Considering the fact that curative options seldom exist for patients with ATC, concurrent inhibition of EGFR and VEGFR tyrosine kinases seems to be a valid and promising anticancer strategy for these patients.
Key Words: preclinical study • MAPK • angiogenesis
|
Introduction |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
1% of all new malignant diseases in the United States (1). Relatively high cure rates can be achieved in well-differentiated thyroid carcinomas, such as papillary and follicular thyroid carcinomas. However, anaplastic thyroid carcinoma (ATC) is one of the most aggressive human malignancies known and carries a grave prognosis (2). Although ATC accounts for only 1.6% of all thyroid cancers, the median overall survival following diagnosis is only 6 months (3, 4). The disease is usually well advanced by the time of diagnosis as evidenced by the average presenting tumor size of 8 cm. Ninety percent of the patients have extraglandular spread at the time of diagnosis and 75% of the patients develop distant metastasis (5, 6). Because ATC is such an aggressive disease, it is staged by the American Joint Commission on Cancer as stage IV regardless of tumor size, cervical lymph node status, or metastatic status (7). Although the treatment of ATC is frequently multidisciplinary, there is no effective cure. This may be due in part to the rarity of this disease (and hence the lack of sufficient research effort). Nevertheless, the inadequacy of the current available treatment options suggests an urgent need for development of novel treatment strategies.
Targeted molecular therapy for cancer is rapidly becoming accepted as an established treatment strategy and has shown promising results in several types of cancer (8). In particular, preclinical and clinical studies of targeted molecular therapy against epidermal growth factor receptor (EGFR) have shown promising results against lung and head and neck cancer (9, 10). EGFR is also frequently overexpressed in various types of thyroid carcinomas (11–16) and this overexpression has been shown to correlate with poor prognosis in several studies (17, 18). Recent work from our laboratory has shown that EGFR is overexpressed in ATC tumors as well (19). Consistent with these observations, inhibition of EGFR by using an anti-EGFR antibody and a small-molecule inhibitor of EGFR tyrosine kinase, AG 1478, has shown antiproliferative effects against thyroid carcinoma cell lines in vitro (20, 21). Despite observations that suggest EGFR as a valid therapeutic target in thyroid cancer, this molecular targeted approach has not been studied much in animal models.
It is well established that the activation of tumor-expressed EGFR frequently results in increased production of VEGF by the tumor cells (22–25). VEGF, a potent proangiogenic molecule, is critical in establishing the ingrowth of tumor vasculature (26). Furthermore, blockade of EGFR has been shown to inhibit tumor-associated angiogenesis in addition to its direct cytotoxic effects on the tumor cells (22, 23, 27, 28). Specific inhibitors of EGFR, such as PKI166 and ZD1839, have been shown to inhibit both neovascularization in rat cornea model (29) and tumor-associated angiogenesis in an animal model of renal cell carcinoma (30). Apoptosis of tumor-associated endothelial cells has also been shown in response to EGFR inhibition with small-molecule inhibitors (31). Therefore, it has been postulated that dual blockade of EGFR and vascular endothelial growth factor receptor (VEGFR) will lead to greater inhibition of tumor-associated angiogenesis compared with inhibition of VEGFR alone (32).
In this article, we report that NVP-AEE788 (AEE788, Novartis Pharma AG, Basel, Switzerland), a recently described dual specific inhibitor of EGFR and VEGFR tyrosine kinases (KDR and Flt-1; ref. 32), inhibited the proliferation and induced apoptosis of ATC cell lines in vitro. Furthermore, the administration of AEE788 to athymic nude mice bearing ATC xenografts inhibited both ATC tumor growth and tumor-associated angiogenesis.
|
Materials and Methods |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
Cell Lines and Culture Conditions
ATC cell lines C643 and K-4 were used. These cell lines were obtained from Sai-Ching Yeung, M.D., Ph.D. (Department of Endocrine Neoplasia and Hormonal Disorders, University of Texas M.D. Anderson Cancer Center). The cells were grown in RPMI 1640 supplemented with 10% fetal bovine serum, penicillin, sodium pyruvate, and nonessential amino acids. Adherent monolayer cultures were maintained on plastic and incubated at 37°C in 5% CO2 and 95% air. The cultures were free of Mycoplasma species. The cultures were maintained no longer than 12 weeks after recovery from frozen stocks.
Reagents
AEE788 was generously provided by Novartis Pharma. For in vitro administration, AEE788 was dissolved in DMSO (Sigma-Aldrich Corp., St. Louis, MO) to a concentration of 20 mmol/L and further diluted to appropriate final concentration in RPMI 1640 with 10% fetal bovine serum. DMSO in the final solution did not exceed 0.1% v/v. For in vivo testing, AEE788 was dissolved in N-methylpyrrolidone and polyethylene glycol 300 1:9 (v/v). The AEE788 solution was prepared just before it was given to the mice. Paclitaxel (Mead Johnson, Princeton, NJ) was diluted 1:6 in Hank's medium for i.p. injections. Propidium iodide and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) were both purchased from Sigma-Aldrich.
Western Immunoblotting
To show that AEE788 is able to inhibit the autophosphorylation EGFR in vitro, Western immunoblotting was done. All cells were incubated in serum-free medium for 24 hours. Cells were incubated with AEE788 for 1 hour at concentrations varying from 0.01 to 20 µmol/L before addition of EGF (40 ng/mL) for 15 minutes. The cells were then washed with PBS and lysis buffer was added [1% Triton X-100, 20 mmol/L Tris (pH 8.0), 137 mmol/L NaCl, 10% glycerol (v/v), 2 mmol/L EDTA, 1 mmol/L phenylmethylsulfonyl fluoride, 20 mmol/L aprotinin-leupeptin-trypsin inhibitor, 2 mmol/L sodium orthovanadate]. The cells were scraped and centrifuged to remove insoluble proteins. The samples were diluted in sample buffer (10% SDS, 0.5 mmol/L Tris-HCl (pH 6.8), 1 mol/L DTT, 10% (v/v) glycerol, 1% bromophenol blue) and boiled. The proteins (100 µg) were resolved by PAGE and electrophoretically transferred onto nitrocellulose membranes. The membranes were blocked with 5% (w/v) nonfat milk in 0.1% Tween 20 (v/v) in TBS, probed with rabbit monoclonal anti-EGFR antibody (1:2,000, Upstate Biotechnology, Inc., Lake Placid, NY) in 1% nonfat milk, and incubated with peroxidase-conjugated sheep anti-rabbit IgG antibody (1:2,000, Amersham Life Science, Inc., Arlington Heights, IL) in 1% nonfat milk. The blots were also probed with rabbit anti-phosphorylated EGFR (pEGFR) antibody (Tyr1068, Cell Signaling, Beverly, MA) diluted 1:1,000 in 1% nonfat milk and incubated with peroxidase-conjugated sheep anti-rabbit IgG antibody (1:2,000). Phosphorylated mitogen-activated protein kinase (pMAPK) rabbit antibodies (Cell Signaling) were used at a concentration of 1:3,000 with anti-rabbit IgG at 1:2,000 as the secondary antibody. Anti-MAPK mouse antibody (Cell Signaling) was used at 1:2,000 dilutions. Anti-ß-actin was used at a 1:1,000 concentration followed by horseradish peroxidase–conjugated donkey anti-rabbit IgG (1:2,000) in 1% nonfat milk. Protein bands were visualized using the Enhanced Chemiluminescence Plus Western blotting detection system (Amersham Life Science).
Measurement of Cell Proliferation
To test the ability of AEE788 to inhibit the proliferation of K-4 and C643 ATC cell lines in vitro, we used a MTT-based assay. Cells (2,000 per well) were grown in RPMI 1640 supplemented with 10% fetal bovine serum in 96-well tissue culture plates. After 24 hours, the cells were treated with various concentrations of AEE788 (0–6 µmol/L) in RPMI 1640 supplemented with 2% fetal bovine serum. Because of the concern that the DMSO in AEE788 preparation may affect the experiments, the concentrations of DMSO in all the wells were adjusted to a common concentration. However, this concentration of DMSO was always <2 µL/mL. To measure the number of metabolically active cells after a 3-day incubation period, we used a MTT assay as measured by a 96-well microtiter plate reader (MR-5000, Dynatech Laboratories, Inc., Chantilly, VA) at an absorbance of 570 nm.
Measurement of Cell Death
To measure cell death, K-4 and C643 cells were plated at a density of 2 x 105 cells/well in 38 mm2 six-well plates (Costar, Cambridge, MA) and maintained for 24 hours before treatment with AEE788. After 24 hours, AEE788 was added in various concentrations in RPMI 1640 supplemented with 2% fetal bovine serum. After 48 hours of treatment with AEE788, the extent of cell death was determined by propidium iodide staining of hypodiploid DNA. The concentrations of DMSO in all the wells were adjusted to a common concentration. Furthermore, this concentration was always kept <2 µg/mL. The treated cells were resuspended in a Nicoletti buffer (50 µg/mL propidium iodide, 0.1% sodium citrate, 0.1% Triton X-100) for 20 minutes at 4°C. Cells were then analyzed by flow cytometry and the sub-G0/G1 fraction was measured.
Immunohistochemistry on Murine Tumor Tissue Sections
Immunohistochemistry was done with the following antibodies: rabbit anti-pEGFR (Tyr1068), rabbit anti-EGFR (Santa Cruz Biotechnology, Santa Cruz, CA), rabbit anti-pMAPK, rabbit anti-MAPK, rabbit anti-phosphorylated VEGFR-2 (pVEGFR-2; Tyr1045, Santa Cruz Biotechnology), and rat anti-mouse CD31-platelet/endothelial cell adhesion molecule 1 (PharMingen, San Diego, CA). For EGFR staining, paraffin-embedded sections were first dewaxed in xylene. Excess xylene was removed by washing the slides in ethanol. After treating the tissue with pepsin for 20 minutes at 37°C, the slides were washed thrice with PBS. Endogenous blocking was done with 3% H2O2 followed by protein blocking using 5% horse serum with 1% goat serum (protein-blocking solution). After washing the slides in PBS, the primary antibody (1:200 dilution) was added for 18 hours at 4°C. The slides were then washed with PBS thrice, blocked again with protein-blocking solution for 1 hour, and incubated with horseradish peroxidase–conjugated anti-rabbit antibody at 1:200 dilutions for 1 hour at room temperature. The slides were washed again in PBS thrice and then incubated with 3,3'-diaminobenzidine tetrahydrochloride (DAB) for 10 minutes. After the excess DAB was washed off, counterstaining was done with Gill's No. 3 hematoxylin.
For staining with antibodies against pEGFR, pMAPK, total MAPK, pVEGFR-2, and CD31-platelet/endothelial cell adhesion molecule 1, frozen tumors were sectioned (8–10 mm thick), mounted on positively charged Superfrost slides (Fisher Scientific, Houston, TX), air dried for 30 minutes, and fixed in cold acetone for 10 minutes. The slides were washed thrice with PBS (pH 7.5), blocked for 20 minutes at room temperature in PBS supplemented with 1% normal goat serum and 5% normal horse serum (protein-blocking solution), and incubated with primary antibody (1:50 dilution for pEGFR, pMAPK, and total MAPK antibodies, 1:400 dilution for pVEGFR-2, and 1:800 dilution for CD31-platelet/endothelial cell adhesion molecule 1 antibody) for 18 hours at 4°C. The samples were then washed thrice for 3 minutes and blocked with protein-blocking solution for 10 minutes. For pEGFR, pMAPK, total MAPK, and pVEGFR-2, the samples were then incubated with Alexa Fluor 594–conjugated goat anti-rabbit IgG at 1:400 (Molecular Probes, Eugene, OR) for 1 hour at room temperature in the dark. Samples were again washed thrice for 5 minutes and then counterstained with 300 µg/mL Hoechst stain for 1 to 2 minutes at room temperature. The slides were again washed and then mounted using propyl gallate. CD31-platelet/endothelial cell adhesion molecule 1 staining was visualized using DAB chromogen. For this purpose, the sections were washed with PBS and incubated with blocking solution for 5 minutes. The sections were then incubated with anti-rat secondary antibody conjugated with horseradish peroxidase (1:800 dilution) for 1 hour at room temperature. The slides were washed again in PBS thrice and then incubated with DAB for 10 minutes. After the excess DAB was washed off, counterstaining was done with Gill's No. 3 hematoxylin.
To assess the degree of intratumoral apoptosis, terminal deoxynucleotidyl transferase–mediated dUTP nick end labeling (TUNEL) staining was done using an apoptosis detection kit (Promega, Madison, WI) with the following modifications: tissues were fixed with 4% paraformaldehyde (methanol free) for 10 minutes at room temperature, washed twice with PBS for 5 minutes, and then incubated with 0.2% Triton X-100 for 15 minutes at room temperature. Then, the tissue sections were incubated with reaction buffer containing 44 µL equilibration buffer, 5 µL nucleotide mix, and 1 µL terminal deoxynucleotidyl transferase at 37°C for 1 hour. The reaction was terminated by immersing the samples in 2x SCC for 15 minutes. The samples were then washed thrice for 5 minutes with PBS to remove unincorporated fluorescein-dUTP.
Immunofluorescence microscopy was done using a Zeiss Axioplan2 microscope (Carl Zeiss, Thornwood, NY) equipped with a 100 W HBO mercury bulb and filter sets (Chroma, Inc., Brattleboro, VT) to individually capture red and blue fluorescent images. Images were captured using a C5810 Hamamatsu color chilled three-chip charge-coupled device camera (Hamamatsu, Japan) and digitized using Optimas imaging software (Silver Spring, MD). To image the DAB-stained paraffin section, the stained sections were examined in a Microphot-FX microscope (Nikon, Melville, NY) equipped with a three-chip charge-coupled device color video camera (Model DXC990, Sony Corp., Tokyo, Japan).
For quantitative analysis of TUNEL staining, the labeled cells were counted in five random 0.159 mm2 fields (x100 magnification) per slide from a total of five slides per study group. To quantify microvessel density (MVD), the labeled endothelial cells were counted from five random 0.159 mm2 fields (x100 magnification) per slide from a total of five slides per study group.
The photomontages were prepared using Photoshop software (Adobe Systems, Inc., San Jose, CA).
In vivo s.c. Xenografts
The K-4 cell line was selected for this part of the study because of its high tumorigenicity in nude mice. K-4 cells were harvested from subconfluent cultures by trypsinization and washed. K-4 cells (2.5 x 106) were injected s.c. into each mouse on the right flank under direct visualization using a 30 gauge needle. The tumors were allowed to develop during the following 7 days. After the development of palpable tumors, the mice were randomized into four groups of 10 mice per each group, and the drugs were given as follows: (a) AEE788 via p.o. gavages at 50 mg/kg thrice weekly, (b) paclitaxel via i.p. injection at 200 µg/injection once weekly, (c) both AEE788 via p.o. gavages at 50 mg/kg thrice weekly and paclitaxel via i.p. injection at 200 µg/injection once weekly, and (d) 250 µL N-methylpyrrolidone and polyethylene glycol 300 1:9 (v/v) given via p.o. gavage and PBS given i.p. as placebo.
Twice weekly, the mice were weighed and tumor sizes were measured. The volumes of the tumors were determined using the formula: V = (
/6)WL2, where W and L represent measurements in two dimensions, with L being the smaller dimension. Our animal protocol required that the mice be sacrificed if they became moribund or lost >20% of their weight. However, all the mice survived until the end of the 6-week treatment period without weight loss of >20% or morbidity. Therefore, all the mice were sacrificed at the end of the treatment period. When the mice were killed at the end of 6-week treatment period, AEE788 and paclitaxel were given 2 hours before the sacrifice.
For immunohistochemical and routine H&E staining, one part of the tumor was fixed in formalin and embedded in paraffin. Another part was embedded in OCT compound (Miles, Inc., Elkhart, IN), rapidly frozen in liquid nitrogen, and stored at –80°C.
Statistical Analysis
The nonparametric Wilcoxon rank-sum test was used to assess differences in mouse tumor volume between each treatment group and the control group. To check for a difference between the combination treatment and the single-agent groups, we ran a repeated-measures regression model with treatment, time, and treatment by time interaction to compare the tumor volumes observed for the group receiving AEE788 plus paclitaxel with the group receiving paclitaxel alone. This methodology was also used to compare tumor volumes in the group receiving AEE788 plus paclitaxel to the group receiving AEE788 alone. TUNEL staining results and MVD counts were compared by a two-sample Student's t test. Best-fit curves were generated for the MTT and propidium iodide assays and used to determine the concentration at which 50% of the drug effect (IC50) were exhibited.
|
Results |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
|
40% and 30%, respectively, at the IC50. These MTT data were also confirmed by counting the cells at 72 hours after treatment with AEE788 (data not shown).
|
35% for both cell lines and the maximal apoptosis of 70% required relatively high concentration of >20 µmol/L AEE788.
AEE788 Inhibits ATC Xenograft Growth In vivo
With respect to the xenografts assay, both AEE788 and paclitaxel were able to inhibit the growth of ATC xenografts generated with K-4 cell line in nude mice. However, the highest growth inhibition was achieved by the coadministration of AEE788 and paclitaxel (Fig. 3). At the end of the 42-day treatment period, the mice treated with paclitaxel, AEE788, and AEE788 plus paclitaxel showed 44%, 58%, and 69% decreases, respectively, in the mean estimated tumor volume compared with the control group (P < 0.05). The difference in tumor volume for the treatment groups was statistically significant compared with the control group starting on treatment day 12. The mean tumor volume of the group receiving the combination treatment was also compared with that of the groups receiving AEE788 alone or paclitaxel alone. The difference in mean tumor volumes of the paclitaxel treatment group compared with combination treatment group became statistically significant after treatment day 23 (P < 0.05). However, there were no statistically significant differences in the mean tumor volume between the mice treated with AEE788 alone and those treated with both AEE788 and paclitaxel.
|
AEE788 Inhibits EGFR Autophosphorylation and Induces Apoptosis of K-4 Tumor Cells In vivo
To show in vivo inhibition of EGFR autophosphorylation by AEE788, immunohistochemical staining was done using antibodies specific to EGFR and tyrosine-phosphorylated EGFR (Fig. 4). Tumors from all the study groups showed similar levels of total EGFR expression, whereas only tumors from control mice or mice treated with paclitaxel stained positive for pEGFR. To determine whether the blockade of EGFR down-regulated any elements of downstream signaling pathways, tumors from the study groups were stained with antibodies specific to MAPK and pMAPK (Fig. 4). Although the level of MAPK expression was similar throughout the study groups, pMAPK staining was absent in tumors of mice treated with AEE788 alone or in combination with paclitaxel.
|
|
|
Lastly, MVD was determined by staining tumor sections for CD31-specific antibodies (Fig. 5). Treatment with paclitaxel did not affect the tumor MVD significantly compared with the control group. However, treatment with AEE788 alone or in combination with paclitaxel resulted in >80% inhibition of tumor-associated angiogenesis (Table 1; P < 0.001).
|
Discussion |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
The therapeutic potential of EGFR inhibition in thyroid carcinoma has been shown in several in vitro studies (20, 21, 35). Gabler et al. (20) showed that a monoclonal antibody against EGFR inhibited the in vitro proliferation of papillary thyroid carcinoma cell lines. More importantly, Yin et al. (35) showed that inhibition of EGFR tyrosine kinase with apigenin, a flavonoid compound, resulted in apoptosis of ATC cell lines in vitro. Despite the evidence that EGFR inhibition may be a valid therapeutic strategy against ATC, only few studies that examined the in vivo effects of EGFR inhibition on thyroid carcinoma have not been reported. In this report, we showed that the blockade of EGFR signaling pathway using AEE788, a novel, dual specific inhibitor of EGFR and VEGFR tyrosine kinases, resulted in in vitro inhibition of proliferation and induction of apoptosis of ATC cells. The inhibition of EGFR resulted in down-regulation of downstream signaling pathways as evidenced by the in vitro and in vivo inhibition of MAPK activation. These effects were maintained in vivo where AEE788 induced apoptosis of the tumor cells and significantly inhibited the growth of ATC xenografts in nude mice.
Administration of AEE788 also significantly decreased tumor-associated angiogenesis. The decrease in MVD in tumors of mice treated AEE788 can be attributed to several mechanisms. First, it has been shown previously in several organ systems and tumor types that activation of EGFR signaling pathway results in up-regulation of VEGF production by the target cells (22, 27, 36). Conversely, EGFR blockade using EGFR monoclonal antibody C225 has been shown to down-regulate the production of VEGF by tumor cells and decreased the MVD in nude mice xenografts of transitional cell carcinoma (27), renal cell carcinoma (28), and colorectal carcinoma (37). Similarly, we have found that AEE788 decreases the in vitro VEGF production by ATC cell lines.4 AEE788 was able to inhibit tumor-expressed EGFR and this may partly account for the decreased MVD in tumors of mice treated with this compound.
The decreased angiogenesis in tumors of mice treated with AEE788 can also be attributed to the inhibition of endothelial cell–expressed EGFR. As shown by immunofluorescent microscopy, the administration of AEE788 efficiently inhibited the autophosphorylation of EGFR on the tumor endothelium. Recent results have suggested that blockade of endothelial cell–expressed EGFR leads to arrest of human umbilical vascular endothelial cells in G1 phase and inhibits endothelial cell proliferation in vivo (29, 30). In a study by Hirata et al. (29), gefinitib, an inhibitor of EGFR tyrosine kinase, completely inhibited neoangiogenesis in vivo in rat cornea model of angiogenesis. Furthermore, a recent preclinical study showed that blockade of endothelial cell–expressed EGFR using PKI166 resulted in endothelial cell apoptosis in pancreatic carcinoma (31).
Lastly, the antiangiogenic property of AEE788 can be attributed to its ability to inhibit endothelial cell–expressed VEGFR tyrosine kinase. The critical need for VEGFR activation in endothelial cell proliferation is well documented (38, 39). Furthermore, antiangiogenic strategies using a monoclonal antibody against VEGF have shown promising results in vivo against nude mice xenografts of various thyroid carcinomas, including ATC (40, 41). In our study, we showed via immunofluorescent microscopy that the administration of AEE788 efficiently inhibited the autophosphorylation of VEGFR-2 on tumor endothelium.
The concept of dual inhibition of EGFR and VEGFR is not new and has been evaluated in animal models of colon (37), gastric (42), and pancreatic carcinomas (43). However, these studies used the administration of two agents targeting each receptor separately. The major advantages of AEE788 are that it is able as a single agent to efficiently block of both EGFR and VEGFR tyrosine kinases and that it is orally given and well tolerated. Although the double specificity of AEE788 may raise the concern of possible, nonspecific interaction of this compound with other tyrosine kinases, in vitro studies by Traxler et al. have confirmed the affinity of the inhibitor to be highly selective for EGFR and VEGFR tyrosine kinases (32).
In this study, the in vivo antitumor efficacy of AEE788 was evaluated as a single agent therapy as well as in combination with paclitaxel. EGFR blockade has shown synergistic effect on tumor cells when combined with radiotherapy and other DNA damaging agents, such as irinotecan (44, 45). Although the animals treated with AEE788 and paclitaxel had statistically significant reduction in tumor volume compared with those that received paclitaxel alone, similar statistical significance could not be shown for mice treated with AEE788 alone when compared with the combination treatment group. However, toward the end of the treatment period, there was a separation of tumor growth curve between mice treated with AEE788 alone and those treated in combination with paclitaxel. With longer study period, this separation most likely would have reached statistical significance. Given the significant antitumor activity of AEE788 against ATC when given in combination with paclitaxel, its role in multimodal therapy seems promising and remains to be further elucidated.
In conclusion, ATC is a highly aggressive disease with poor prognosis that warrants the development of novel treatment strategies. AEE788, an orally given inhibitor of EGFR and VEGFR kinases, has shown significant antitumor activity against ATC cell lines in vitro and against ATC xenografts in vivo in nude mice. In addition, AEE788 efficiently inhibited tumor-associated angiogenesis and decreased the intratumoral MVD. The inhibition of overall tumor growth in mice treated with AEE788 is most likely the result of its direct cytotoxicity on the tumor cells as well as the suppression of tumor-associated angiogenesis. Considering the fact that curative options seldom exist for patients with ATC, concurrent inhibition of EGFR and VEGFR tyrosine kinases seems to be a valid and promising anticancer strategy for these patients.
|
Footnotes |
|---|
The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
Received 11/ 1/04; revised 1/21/05; accepted 2/21/05.
|
References |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
and epidermal growth factor receptor in thyroid tumors. In Vivo 1992;6:291–6.
Song B. Immunohistochemical demonstration of epidermal growth factor receptors and ceruloplasmin in thyroid disease. Acta Pathol Jpn 1991;41:333–43.
Akslen LA, Varhaug JE. Oncoproteins and tumor progression in papillary thyroid carcinoma: presence of epidermal growth factor receptor, c-erbB-3 protein, estrogen receptor related protein, p21-ras protein, and proliferation indicators in relation to tumor recurrences and patient survival. Cancer 1995;76:1643–54.[CrossRef][Medline]
Chen BK, Ohtsuki Y, Furihata M, et al. Co-overexpression of p53 protein and epidermal growth factor receptor in human papillary thyroid carcinomas correlates with lymph node metastasis, tumor size and clinicopathologic stage. Int J Oncol 1999;15:893–8.[Medline]
Schiff BA, McMurphy AB, Jasser SA, et al. Epidermal growth factor receptor (EGF-R) is overexpressed in anaplastic thyroid cancer, and the EGF-R inhibitor gefinitib ("Iressa", ZD1839) inhibits the growth of anaplastic thyroid cancer. Clin Cancer Res 2004;10:8594–602.This article has been cited by other articles:
|
|
 |
|
  M. Croyle, N. Akeno, J. A. Knauf, D. Fabbro, X. Chen, J. E. Baumgartner, H. A. Lane, and J. A. Fagin RET/PTC-Induced Cell Growth Is Mediated in Part by Epidermal Growth Factor Receptor (EGFR) Activation: Evidence for Molecular and Functional Interactions between RET and EGFR Cancer Res., June 1, 2008; 68(11): 4183 - 4191. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 F. Gomez-Rivera, A. A. Santillan-Gomez, M. N. Younes, S. Kim, D. Fooshee, M. Zhao, S. A. Jasser, and J. N. Myers The Tyrosine Kinase Inhibitor, AZD2171, Inhibits Vascular Endothelial Growth Factor Receptor Signaling and Growth of Anaplastic Thyroid Cancer in an Orthotopic Nude Mouse Model Clin. Cancer Res., August 1, 2007; 13(15): 4519 - 4527. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. Kim, Y. D. Yazici, G. Calzada, Z.-Y. Wang, M. N. Younes, S. A. Jasser, A. K. El-Naggar, and J. N. Myers Sorafenib inhibits the angiogenesis and growth of orthotopic anaplastic thyroid carcinoma xenografts in nude mice Mol. Cancer Ther., June 1, 2007; 6(6): 1785 - 1792. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. M. Wiseman, H. Masoudi, P. Niblock, D. Turbin, A. Rajput, J. Hay, S. Bugis, D. Filipenko, D. Huntsman, and B. Gilks Anaplastic Thyroid Carcinoma: Expression Profile of Targets for Therapy Offers New Insights for Disease Treatment Ann. Surg. Oncol., February 1, 2007; 14(2): 719 - 729. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 C. S. Mitsiades, V. Kotoula, V. Poulaki, E. Sozopoulos, J. Negri, E. Charalambous, G. Fanourakis, G. Voutsinas, S. Tseleni-Balafouta, and N. Mitsiades Epidermal Growth Factor Receptor as a Therapeutic Target in Human Thyroid Carcinoma: Mutational and Functional Analysis J. Clin. Endocrinol. Metab., September 1, 2006; 91(9): 3662 - 3666. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Kim, C. N. Prichard, M. N. Younes, Y. D. Yazici, S. A. Jasser, B. N. Bekele, and J. N. Myers Cetuximab and Irinotecan Interact Synergistically to Inhibit the Growth of Orthotopic Anaplastic Thyroid Carcinoma Xenografts in Nude Mice Clin. Cancer Res., January 15, 2006; 12(2): 600 - 607. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
| Cancer Research | Clinical Cancer Research |
| Cancer Epidemiology Biomarkers & Prevention | Molecular Cancer Therapeutics |
| Molecular Cancer Research | Cancer Prevention Research |
| Cancer Prevention Journals Portal | Cancer Reviews Online |
| Annual Meeting Education Book | Meeting Abstracts Online |
