This Article Abstract Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Chen, I-F. Articles by Hung, M.-C. Search for Related Content PubMed PubMed Citation Articles by Chen, I-F. Articles by Hung, M.-C.

Departments of ¹ Molecular and Cellular Oncology and ² Breast Medical Oncology, The University of Texas M.D. Anderson Cancer Center, Houston, Texas; ³ Department of Surgery, Kaohsiung Medical University, and ⁴ Department of Medical Research, E-DA Hospital, I-Shou University, Kaohsiung, Taiwan, Republic of China; and ⁵ Cancer Biology Program, Graduate School of Biomedical Science, The University of Texas Health Science Center at Houston, Houston, Texas

Requests for reprints: Mien-Chie Hung, Department of Molecular and Cellular Oncology, Unit 108, The University of Texas M.D. Anderson Cancer Center, 1515 Holcombe Boulevard, Houston, TX 77030. Phone: 713-792-7477; Fax: 713-794-0209. E-mail: mhung{at}mdanderson.org

	Abstract

Top Abstract Introduction Materials and Methods Results and Discussion References

 
IFN-inducible proteins are known to mediate IFN-directed antitumor effects. The human IFN-inducible protein absent in melanoma 2 (AIM2) gene encodes a 39-kDa protein, which contains a 200-amino-acid repeat as a signature of HIN-200 family (hematopoietic IFN-inducible nuclear proteins). Although AIM2 is known to inhibit fibroblast cell growth in vitro, its antitumor activity has not been shown. Here, we showed that AIM2 expression suppressed the proliferation and tumorigenicity of human breast cancer cells, and that AIM2 gene therapy inhibited mammary tumor growth in an orthotopic tumor model. We further showed that AIM2 significantly increased sub-G₁ phase cell population, indicating that AIM2 could induce tumor cell apoptosis. Moreover, AIM2 expression greatly suppressed nuclear factor-B transcriptional activity and desensitized tumor necrosis factor-–mediated nuclear factor-B activation. Together, these results suggest that AIM2 associates with tumor suppression activity and may serve as a potential therapeutic gene for future development of AIM2-based gene therapy for human breast cancer. [Mol Cancer Ther 2006;5(1):1–7]

	Introduction

Top Abstract Introduction Materials and Methods Results and Discussion References

 
IFNs play critical roles in host immune defense signaling pathways and have been successfully used for anticancer therapy (1–3). IFN-inducible proteins are thought to mediate the direct antitumor activity of IFNs (4) and have been used with great success in animal models by cancer gene therapy. The human IFN-inducible proteins IFI16, MNDA, AIM2, and IFIX, and the structurally related murine homologous genes p202, p203, p204, and p205 constitute the family as hematopoietic IFN-inducible nuclear proteins containing a 200-amino-acid repeat, HIN-200 (5). Many HIN-200 proteins have been shown to suppress cell growth (6). For example, murine protein p202 has been shown to suppress tumor growth and metastasis of human breast, pancreatic, and prostate cancer cells (7–10). The antigrowth activity of p202 is partly mediated by up-regulation of p21 and pRb and down-regulation of cyclin-dependent kinase 2 protein kinase activity (11). It has been shown that p202 interacts with several transcription factors, such as nuclear factor-B (NF-B), p50 and p65, activator protein, c-fos, c-jun, and E2F, and modulates their transcriptional activity (12, 13).

The human IFN-inducible protein IFIX1 has been shown to reduce the anchorage-dependent and anchorage-independent growth and the tumorigenicity of breast cancer cells(14). Another human IFN-inducible protein that may associate with tumor suppression activity is AIM2. AIM2 gene was originally identified by subtractive cDNA selection for association with human melanoma tumorigenicity (15) and only shares 31% protein identity with IFIX1 (14). The AIM2 gene contains a site of microsatellite instability that results in gene inactivation in 47.6% of colorectal tumors with high microsatellite instability (16). AIM2 gene mutation has also been found in association with gastric and endometrial cancers (17). These findings, in combination with the observation that AIM2 can be silenced by DNA methylation in its genome in immortalized cells (18), suggest that AIM2 is a tumor suppressor. However, suppression of cancer cell growth and tumorigenicity by AIM2 has not been shown (19).

In this study, we aim to determine whether AIM2 inhibits breast cancer cell growth in vitro and to explore the possibility of using AIM2 for breast cancer gene therapy. We showed that AIM2 DNA liposome treatment not only suppressed breast cancer cell growth in vitro but also inhibited mammary tumor growth in orthotopic tumor models. Our results indicate that AIM2 possesses tumor suppression activity, and that AIM2 transgene expression may be further explored as a novel gene therapy for patients with breast cancer.

	Materials and Methods

Top Abstract Introduction Materials and Methods Results and Discussion References

 
Cell Lines and Plasmids
Human embryonic kidney cells (HEK-293T) and human breast cancer cells (MCF-7, MDA-MB-231, MDA-MB-435, and MDA-MB-453) were obtained from American Type Culture Collection (Manassas, VA) and maintained in DMEM (Hyclone Laboratories, Logan, UT) supplemented with 10% (v/v) fetal bovine serum. The AIM2 expression vector pCMV-Tag-AIM2 (Fig. 1A, top ) was constructed by inserting AIM2 cDNA into pCMV-Tag2C (Flag tag; Stratagene, La Jolla, CA).

View larger version (16K): [in this window] [in a new window]   Figure 1. Cloning and protein expression of AIM2. A, the coding sequence of AIM2 was inserted into pCMV-Tag2C and fused with FLAG tag at the NH2-terminal. pBI-EGFP-Tag-AIM2 was used for selection of Tet-Off gene expression stable transfectants. The tetracycline-responsive element (TRE) is upstream of the minimum CMV promoter (pminiCMV) and the AIM2 gene. SV40 pA, SV40 polyadenylation signal. B, Western blotting was done with whole-cell lysates and using anti-Flag antibody. Left, transient expression of AIM2 in HEK-293T cells was achieved by transfecting cells with the AIM2-expressing vector pCMV-Tag-AIM2 or the control vector pCMV-Tag2C. Right, AIM2 expression in MCF-7 selected stable transfectants #13 and #25. The cells were tested under the culture condition with 2 µg/mL doxycycline (+) or removal of doxycycline (–). ß-Actin was used as a loading control. NS, marks a nonspecific band detected by anti-flag antibody.

Western Blot Analysis
To induce expression of AIM2 protein, DMEM/F-12 medium containing doxycycline was removed from the cell culture plate. The cell culture plate was washed four times with PBS and then supplied with fresh DMEM without doxycycline. Cells were collected at the indicated time points for Western blotting analysis. Protein lysate was prepared using radioimmunoprecipitation assay-B buffer containing 20 mmol/L Na₂PO₄ (pH 7.4), 150 mmol/L NaCl, 1% Triton X-100, 100 mmol/L NaF, 2 mmol/L Na₃VO₄, 5 mmol/L phenylmethylsulfonyl fluoride, 1% aprotinin, and 10 mg/mL leupeptin. The monoclonal antibody specific for Tag, anti-Flag M2 (Sigma), was used to detect AIM2 expression. The cytoplasmic and nuclear extracts were isolated according to the protocol described previously (21). Anti--tubulin monoclonal antibody (Sigma) was used to confirm protein localization.

Cell Growth Analysis
To determine the rate of cell proliferation, 2 x 10⁴ MCF-7 tTA-AIM2 #13 or #25 cells were seeded onto 12-well plates with or without doxycycline. DMEM was replaced with fresh medium with or without doxycycline every 48 hours. At the indicated time, the cells were trypsinized and collected individually from at least two plates. Cells from each plate were counted thrice with a cell counter (Beckman Coulter, Fullerton, CA). The average number of cells from at least three plates was used to determine proliferation rates.

Cell Cycle Analysis
To determine cell cycle distribution, 5 x 10⁵ MCF-7 tTA-Luc and MCF-7 tTA-Tag-AIM2 cells were seeded per each 100-mm plate with or without doxycycline. DMEM was replaced with fresh medium with or without doxycycline every 48 hours. After 5 days, the cells were trypsinized and fixed with 1 mL of 70% ethanol at 4°C overnight. The fixed cells were centrifuged at 1,500 x g for 5 minutes and then resuspended in 1 mL of PBS solution containing 100 µg/mL RNase A (Sigma) and 50 µg/mL propidium iodide (Sigma). The stained cells were subjected to flow cytometry.

Terminal Deoxynucleotidyl Transferase–Mediated Nick End Labeling Assay
AIM2-inducible MCF7 cells were cultured in DMEM/F12 medium with or without doxycycline (2 µg/mL) for 96 hours at 37°C in a humid atmosphere of 5% CO₂/95% air. The cell culture medium was replaced with fresh medium with or without doxycycline every 48 hours. Subsequently, the cells were washed by PBS then fixed by 4% formaldehyde. Apoptotic cells were determined using the Fluorescein DNA Fragmentation Detection kit from Calbiochem (San Diego, CA) following the manufacturer's instructions. The apoptotic cells (bright green signaling) were counted under a microscope. 4',6-Diamidino-2-phenylindole was used for the visualization of both apoptotic and nonapoptotic cells. The apoptotic index was defined by the percentage of green signaling–positive cells among the total number of cells in each sample. Five fields with 100 cells per field were randomly counted for each sample. We counted a minimum of three samples for each group.

Generation of AIM2-Inducible MCF-7 Tumor Xenografts
AIM2-inducible MCF-7 cells (#13) were maintained in DMEM/F-12 medium supplemented with 10% (v/v) fetal bovine serum and 2 µg/mL doxycycline. After 24 hours, the cells were harvested in 0.2 mL of PBS and injected into one of the mammary fat pads within each mouse (10⁷ per mouse). To support the growth of the estrogen-dependent MCF-7 tumors, a 0.72 mg 17ß-estradiol 60-day release pellet (Innovative Research of America, Sarasota, FL) was implanted s.c. into the back of each mouse the day before tumor injection. During this experiment, the mice were fed sucrose water with or without doxycycline and were examined weekly to assess tumor growth.

Colony-Forming Assay
MCF-7, MDA-MB-231, MDA-MB-435, and MDA-MB-453 cells were transfected with the AIM2 expression vector pCMV-Tag-AIM2 or the control vector pCMV-Tag2C using SN liposome and were selected in 500 µg/mL G418. After 3 weeks, the G418-resistant colonies were stained with crystal violet and counted as described previously (9). Experiments were repeated thrice.

Cell Viability Assay
MCF-7, MDA-MB-231, MDA-MB-435, and MDA-MB-453 cells (2 x 10⁵) were plated onto a six-well plate the day before transfection. Cells were transfected with 0.2 µg of CMV-Luc and different amounts of pCMV-Tag2C or pCMV-Tag-AIM2 by using SN as a gene delivery system. The total amount of DNA transfected at each AIM2 dose was kept constant (1.6 µg) by adding an appropriate amount of pCMV-Tag2C vector. At 36 hours after transfection, the cells were harvested, and luciferase activity was determined with a luminometer (Turner Designs Instruments, Sunnyvale, CA). Experiments were done thrice.

AIM2 Gene Therapy
Six-week-old female nude mice (10 mice per group; Charles River Laboratories, Wilmington, MA) were injected with 2 x 10⁶ MDA-MB-435 cells under the second or third left mammary fat pad. After the tumors had grown to 0.5 cm in diameter about 7 days after tumor inoculation, the mice were treated with SN2 liposome (20, 22) with 20 µg of either pCMV-Tag-AIM2 or pCMV-Tag2C twice a week by i.t. injection for 2 weeks. Tumor size was measured in two dimensions with a caliper twice a week and calculated as S²L/2, where S is the shortest diameter of the tumor in millimeters and L is the longest diameter of the tumor in millimeters (Student's t test, day 21, P < 0.01). Animal care and experimental procedures were conducted in compliance with the Institute's Animal Care and Use Committee regulations following NIH guidelines.

Immunostaining
Inducible AIM2 clone (#13) of MCF-7 cells was cultured in a four-well glass chamber overnight. The enhanced GFP (EGFP) expression vector served as a control. Three days after induction, the cells were washed with PBS, fixed with 3% paraformaldehyde in PBS for 30 minutes at room temperature, and washed again with PBS. The primary antibody, anti-Flag antibody (1:100), was incubated with the cells at room temperature for 1 hour. The cells were then washed with PBS and incubated with the secondary antibody conjugated with green fluorescence (FITC) at room temperature for 1 hour. After incubation, the cells were washed with PBS, air-dried, and incubated with the fluorescent dye 4',6-diamidino-2-phenylindole (1:100 in 50% glycerol and PBS). A coverslip was placed on top of the slide for visualization by immunofluorescence microscopy.

Transfection and Luciferase Assay
MDA-MB-435 and MDA-MB-453 cells were transfected with 0.5 µg of B-Luc construct, an IB promoter-driven luciferase gene (9) and 0.1 µg of the internal transfection control, pRL-TK (Promega). NF-B (p65) expression vector has been described previously (23). Forty-eight hours after transfection, the cells were harvested, and luciferase activity was measured using the dual-luciferase reporter assay system (Promega) according to the protocol supplied by the manufacturer. The relative activities were calculated by setting the luciferase activities obtained from transfections without pCMV-Tag-AIM2 at 100%. We did three independent experiments of this assay. Human tumor necrosis factor- (20 ng/mL; R&D Systems, Inc., Minneapolis, MN) was used to stimulate the NF-B transcriptional activity.

	Results and Discussion

Top Abstract Introduction Materials and Methods Results and Discussion References

 
AIM2 Constructs and Generation of Tetracycline-Inducible AIM2 Cell Lines
To investigate whether AIM2 has tumor-suppressing activity, we first generated an AIM2 expression vector. AIM2 cDNA was cloned into transient expression vector pCMV-Tag2C, and the construct was designated pCMV-Tag-AIM2 (Fig. 1A, top). AIM2 protein was readily detected in lysates from HEK-293T cells transiently transfected with AIM2, via Western blot analysis using anti-Flag antibody (Fig. 1B, left). The observed molecular weight of AIM2 was 39 kDa, which was consistent with the predicted size.

We then established MCF-7 transfectants that stably expressed AIM2 under the control of a tetracycline-repressible promoter (Fig. 1A, bottom). Two independent AIM2 expression cell lines (#13 and #25) were selected and expressed high levels of AIM2 after withdrawal of doxycycline (Fig. 1B, right), indicating success of the Tet-off–mediated inducibility. The higher molecular weight bands indicated flag-tagged AIM2, whereas the lower molecular weight bands represent the nonspecific proteins detected by anti-flag antibody.

AIM2 Protein Suppressed Breast Cancer Cell Proliferation and Induced Cell Apoptosis
Next, we aimed to determine whether expression of AIM2 could suppress breast cancer cell proliferation and tumor formation using MCF-7 Tet-Off stable transfectants in the cell growth assay. As shown in Fig. 2A , expression of AIM2 significantly hampered growth of MCF-7 tTA-Tag-AIM2 cells. Suppression of AIM2 expression by doxycycline greatly increased proliferation of MCF-7 tTA-Tag-AIM2 cells in two independent MCF-7 stable lines #13 and #25 (Fig. 2A). These results suggest that AIM2 expression could suppress MCF-7 breast cancer cell growth. As expected, doxycycline has no significant effect on cell growth for the vector control cell line MCF-7 tTA-Luc (Fig. 2A). To elucidate the mechanism by which AIM2 suppressed breast cancer cell proliferation, we did cell cytometry flow analysis to determine the cell population distribution. The results showed that expression of AIM2 by withdrawal of doxycycline increased the sub-G₁ cell population, suggesting that AIM2-expressing cells underwent apoptosis (data not shown). We further confirmed the AIM2-induced cell apoptosis by terminal deoxynucleotidyl transferase–mediated nick end labeling assay (Fig. 2B, left). As expected, expression of AIM2 by withdrawal of doxycycline significantly increased the percentage of apoptotic cells by 5-fold (Fig. 2B, right). The control group of MCF-7 tTA-Luc showed no significant change of apoptotic cell population with or without doxycycline.

View larger version (21K): [in this window] [in a new window]   Figure 2. AIM2 suppresses MCF-7 cell growth in vitro and tumor formation in vivo. A, inducible expression of AIM2 protein retards proliferation of MCF-7 (#13) and (#25) breast cancer cells. The MCF-7 stable line expressing luciferase was used as control. A total of 1 x 104 cells was seeded onto a 12-well plate with (Dox+) or without (Dox–) doxycycline, and the cells were counted at the indicated times. The experiment was done in triplicate, and each sample was counted twice. Points, means; bars, SE. B, AIM2 increases the apoptotic cell population of MCF-7 tTA-AIM2. Left, MCF-7 tTA-AIM2 cells were double stained with 4',6-diamidino-2-phenylindole (nuclear region) and terminal deoxynucleotidyl transferase–mediated nick end labeling (apoptotic cells). The arrows indicate the apoptotic cells. Right, apoptotic cell percentage in MCF-7 tTA-Luc and MCF-7 tTA-AIM2 with doxycycline or without doxycycline. C, the antitumor effect of AIM2 expression by a tetracycline-inducible system in a MCF-7 mammary tumor model. Six-week-old female nude mice (10 mice per group) were inoculated with 2 x 106 MCF-7 (#13) cells, which express the AIM2 gene under the tetracycline regulation system. The mice were fed sucrose water with or without doxycycline (2 µg/mL). Tumor size was measured with a caliper twice a week. Points, means; bars, SD. Inset, tumor incidents of mice bearing the MCF-7 tTA-Luc and MCF-7 tTA-AIM2 cancer cells when feeding sucrose water with doxcycline (Dox+) or without doxcycline (Dox–).

We next examined whether AIM2 also targets other breast cancer cells. We first did a relative cell viability assay using pCMV-Luc cotransfection. Luciferase activity was used as an indication of living cells. As expected, AIM2 expression inhibited the viability of MCF-7, MDA-MB-231, MDA-MB-435, and MDA-MB-453 breast cancer cells in a dose-dependent manner (Fig. 3A ). Consistently, AIM2 inhibited the colony-forming ability of these breast cancer cells (Fig. 3B). AIM2 expression vector pCMV-Tag-AIM2 or the control vector pCMV-Tag2C were transfected into the four human breast cancer cell lines and selected with G418. At 3 weeks, the number of G418-resistant colonies in all four cell lines was at least 85% lower in AIM2-transfected cells than in control vector-transfected cells (Fig. 3B).

View larger version (29K): [in this window] [in a new window]   Figure 3. AIM2 suppresses the viability, colony formation, and tumorigenicity of breast cancer cells. A, AIM2 suppresses cell viability. Different amounts of the AIM2-expressing vector pCMV-Tag-AIM2 were transiently transfected into four breast cancer cell lines together with 0.1 µg of the CMV-Luc reporter plasmid. The total amount of DNA transfected at each AIM2 dose was kept constant (1.6 µg) by adding an appropriate amount of pCMV-Tag2C vector. After 36 h, the cells were lysed, and luciferase activity was measured. The relative activities were calculated by normalizing to the luciferase activities obtained from cells not transfected with AIM2. Columns, means of three independent experiments; bars, SD. B, AIM2 inhibits colony formation of breast cancer cells. Cells from four lines were transfected with pCMV-Tag-AIM2 or the control vector pCMV-Tag2C. Photographs were taken 3 wks after transfection. The colony numbers obtained from pCMV-Tag2C transfection were set at 100%. Columns, means; bars, SD. C, AIM2 is antitumorigenic in a breast cancer cell mammary tumor model. Six-week-old female nude mice (10 mice per group) were inoculated with 2 x 106 MDA-MB-435 cells under the mammary fat pad. Intratumor gene therapy with AIM2 DNA-liposome was started 7 d after tumor inoculation. Tumor size was measured with a caliper twice a week. Points, means; bars, SD. The arrows indicate the day of gene therapy.

AIM2 Suppressed NF-B Transcriptional Activity
We next aimed to elucidate the molecular mechanism by which AIM2 suppresses tumor cell growth. As AIM2 induces apoptosis and NF-B is known to be antiapoptotic, we asked whether AIM2 might induce tumor apoptosis via inhibiting NF-B function (Fig. 2). To this end, we cotransfected pCMV-Tag-AIM2 with a Rel-A (a p65 subunit of NF-B) cDNA expression vector and an NF-B-responsive promoter-reporter construct (B-Luc, an IB promoter-driven luciferase gene) into MDA-MB-435 and MDA-MB-453 cells. The results showed that AIM2 expression greatly repressed NF-B (Rel-A)–activated IB promoter activity in both cell lines (Fig. 4A ). We further tested whether AIM2 expression could affect NF-B–mediated transcription activation in response to tumor necrosis factor- treatment. As expected, B-Luc was readily activated in the presence of tumor necrosis factor-. Importantly, this tumor necrosis factor-–induced transcriptional activation was repressed by AIM2 (Fig. 4B). Together, these results indicate that AIM2 negatively regulates the tumor necrosis factor-/NF-B antiapoptotic pathway, which correlates with the AIM2-induced apoptosis and antitumor activity.

View larger version (27K): [in this window] [in a new window]   Figure 4. AIM2 suppresses NF-B transcriptional activity. A, AIM2 inhibits NF-B transcriptional activity. AIM2 expression vector was cotransfected with B-Luc and p65 into MDA-MB-435 and MDA-MB-453 cells. B-Luc activity without AIM2 suppression was set at 100%. Columns, means of three independent experiments; bars, SD. B, AIM2 desensitizes TNF-–activated NF-B transcriptional activity. TNF-–activated B-Luc activity without AIM2 suppression was set at 100%. AIM2 concentrations were 0.5 and 1.5 µg. Columns, means of three independent experiments; bars, SD.

	Acknowledgments

 
We thank Dr. Hui-Wen Lo at the Department of Molecular and Cellular Oncology and the Department of Scientific Publication at the University of Texas M.D. Anderson Cancer Center for editing the article.

	Footnotes

 
Grant support: Breast Cancer Research Foundation (G.N. Hortobagyi and M-C. Hung), Kaohsiung Medical University (F. Ou-Yang and J-Y. Hung) and M.D. Anderson Cancer Center supporting grant CA16672.

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.

Note: I-F. Chen and F. Ou-Yang contributed equally to this work.

J-Y. Hung is a visiting scientist from the Department of Internal Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan, Republic of China. H. Wang is currently at the Department of Physiology and Biophysics, University of Calgary, Calgary, Alberta, Canada.

Received 8/ 8/05; revised 10/17/05; accepted 11/ 2/05.

	References

Top Abstract Introduction Materials and Methods Results and Discussion References

 

1. Ikeda H, Old LJ, Schreiber RD. The roles of IFN in protection against tumor development and cancer immunoediting. Cytokine Growth Factor Rev 2002;13:95–109.[CrossRef][Medline]
2. Caraglia M, Marra M, Pelaia G, et al. -Interferon and its effects on signal transduction pathways. J Cell Physiol 2005;202:323–35.[CrossRef][Medline]
3. Yoshida J, Mizuno M, Wakabayashi T. Interferon-ß gene therapy for cancer: basic research to clinical application. Cancer Sci 2004;95:858–65.[CrossRef][Medline]
4. Lengyel P. Tumor-suppressor genes: news about the interferon connection. Proc Natl Acad Sci U S A 1993;90:5893–5.[Abstract/Free Full Text]
5. Johnstone RW, Trapani JA. Transcription and growth regulatory functions of the HIN-200 family of proteins. Mol Cell Biol 1999;19:5833–8.[Free Full Text]
6. Asefa B, Klarmann KD, Copeland NG, Gilbert DJ, Jenkins NA, Keller JR. The interferon-inducible p200 family of proteins: a perspective on their roles in cell cycle regulation and differentiation. Blood Cells Mol Dis 2004;32:155–67.[CrossRef][Medline]
7. Ding Y, Wen Y, Spohn B, et al. Proapoptotic and antitumor activities of adenovirus-mediated p202 gene transfer. Clin Cancer Res 2002;8:3290–7.[Abstract/Free Full Text]
8. Wen Y, Giri D, Yan DH, et al. Prostate-specific antitumor activity by probasin promoter-directed p202 expression. Mol Carcinog 2003;37:130–7.[Medline]
9. Wen Y, Yan DH, Wang B, et al. p202, an interferon-inducible protein, mediates multiple antitumor activities in human pancreatic cancer xenograft models. Cancer Res 2001;61:7142–7.[Abstract/Free Full Text]
10. Yan DH, Wen Y, Spohn B, Choubey D, Gutterman JU, Hung MC. Reduced growth rate and transformation phenotype of the prostate cancer cells by an interferon-inducible protein, p202. Oncogene 1999;18:807–11.[CrossRef][Medline]
11. Choubey D, Lengyel P. Binding of an interferon-inducible protein (p202) to the retinoblastoma protein. J Biol Chem 1995;270:6134–40.[Abstract/Free Full Text]
12. Wen Y, Yan DH, Spohn B, Deng J, Lin SY, Hung MC. Tumor suppression and sensitization to tumor necrosis factor -induced apoptosis by an interferon-inducible protein, p202, in breast cancer cells. Cancer Res 2000;60:42–6.[Abstract/Free Full Text]
13. Ma XY, Wang H, Ding B, Zhong H, Ghosh S, Lengyel P. The interferon-inducible p202a protein modulates NF-B activity by inhibiting the binding to DNA of p50/p65 heterodimers and p65 homodimers while enhancing the binding of p50 homodimers. J Biol Chem 2003;278:23008–19.[Abstract/Free Full Text]
14. Ding Y, Wang L, Su LK, et al. Antitumor activity of IFIX, a novel interferon-inducible HIN-200 gene, in breast cancer. Oncogene 2004;23:4556–66.[CrossRef][Medline]
15. DeYoung KL, Ray ME, Su YA, et al. Cloning a novel member of the human interferon-inducible gene family associated with control of tumorigenicity in a model of human melanoma. Oncogene 1997;15:453–7.[CrossRef][Medline]
16. Schulmann K, Brasch FE, Kunstmann E, et al. HNPCC-associated small bowel cancer: clinical and molecular characteristics. Gastroenterology 2005;128:590–9.[CrossRef][Medline]
17. Woerner SM, Benner A, Sutter C, et al. Pathogenesis of DNA repair-deficient cancers: a statistical meta-analysis of putative Real Common Target genes. Oncogene 2003;22:2226–35.[CrossRef][Medline]
18. Kulaeva OI, Draghici S, Tang L, Kraniak JM, Land SJ, Tainsky MA. Epigenetic silencing of multiple interferon pathway genes after cellular immortalization. Oncogene 2003;22:4118–27.[CrossRef][Medline]
19. Cresswell KS, Clarke CJ, Jackson JT, Darcy PK, Trapani JA, Johnstone RW. Biochemical and growth regulatory activities of the HIN-200 family member and putative tumor suppressor protein, AIM2. Biochem Biophys Res Commun 2005;326:417–24.[Medline]
20. Zou Y, Peng H, Zhou B, et al. Systemic tumor suppression by the proapoptotic gene bik. Cancer Res 2002;62:8–12.[Abstract/Free Full Text]
21. Xie Y, Hung MC. Nuclear localization of p185neu tyrosine kinase and its association with transcriptional transactivation. Biochem Biophys Res Commun 1994;203:1589–98.[CrossRef][Medline]
22. Yan DH, Spohn B, Hung MC. Delivery of DNA to tumor cells using cationic liposomes. Methods Mol Biol 2004;245:125–36.[Medline]
23. Shao R, Karunagaran D, Zhou BP, et al. Inhibition of nuclear factor-B activity is involved in E1A-mediated sensitization of radiation-induced apoptosis. J Biol Chem 1997;272:32739–42.[Abstract/Free Full Text]
24. Liao Y, Zou YY, Xia WY, Hung MC. Enhanced paclitaxel cytotoxicity and prolonged animal survival rate by a nonviral-mediated systemic delivery of E1A gene in orthotopic xenograft human breast cancer. Cancer Gene Ther 2004;11:594–602.[Medline]

This article has been cited by other articles:

				Y.-J. Shann, C. Cheng, C.-H. Chiao, D.-T. Chen, P.-H. Li, and M.-T. Hsu Genome-wide mapping and characterization of hypomethylated sites in human tissues and breast cancer cell lines Genome Res., May 1, 2008; 18(5): 791 - 801. [Abstract] [Full Text] [PDF]

				R. Supino, G. Petrangolini, G. Pratesi, M. Tortoreto, E. Favini, L. D. Bo, P. Casalini, E. Radaelli, A. C. Croce, G. Bottiroli, et al. Antimetastatic Effect of a Small-Molecule Vacuolar H+-ATPase Inhibitor in in Vitro and in Vivo Preclinical Studies J. Pharmacol. Exp. Ther., January 1, 2008; 324(1): 15 - 22. [Abstract] [Full Text] [PDF]

				J. A. Ajani New Proposal for Postsurgery Pathologic Staging of Esophageal or Gastroesophageal Junction Adenocarcinoma: Why Bother? J. Clin. Oncol., March 1, 2007; 25(7): 906 - 907. [Full Text] [PDF]

				D. Hoffmann and O. Wildner Enhanced killing of pancreatic cancer cells by expression of fusogenic membrane glycoproteins in combination with chemotherapy. Mol. Cancer Ther., August 1, 2006; 5(8): 2013 - 2022. [Abstract] [Full Text] [PDF]

This Article Abstract Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Chen, I-F. Articles by Hung, M.-C. Search for Related Content PubMed PubMed Citation Articles by Chen, I-F. Articles by Hung, M.-C.