Journal of the National Cancer Institute,
Vol. 97, No. 21, 1611-1615, November 2, 2005
Effect of Gamma-Linolenic Acid on the Transcriptional Activity of the Her-2/neu (erbB-2) Oncogene
Javier A. Menendez, Luciano Vellon, Ramon Colomer, Ruth Lupu
Affiliations of authors: Department of Medicine, Evanston Northwestern Healthcare Research Institute, Evanston, IL (JAM, LV, RL); Department of Medicine, Northwestern University Feinberg School of Medicine, Chicago, IL (JAM, LV, RL); Robert H. Lurie Comprehensive Cancer Center of Northwestern University, Chicago, IL (JAM, RL); Medical Oncology, Institut Catala d'Oncologia, Hospital Universitari Dr. Josep Trueta, Girona, Spain (RC)
The -6 polyunsaturated fatty acid -linolenic acid (GLA; 18:3n-6), which is found in several plant oils and is used as an herbal medicine, has antitumor activity in vitro. We examined the effect of GLA on the expression of the Her-2/neu (erbB-2) oncogene, which is involved in development of numerous types of human cancer. Flow cytometric and immunoblotting analyses demonstrated that GLA treatment substantially reduced Her-2/neu protein levels in the Her-2/neu–overexpressing cell lines BT-474, SK-Br3, and MDA-MB-453 (breast cancer), SK-OV3 (ovarian cancer), and NCI-N87 (gastrointestinal tumor derived). GLA exposure led to a dramatic decrease in Her-2/neu promoter activity and a concomitant increase in the levels of polyomavirus enhancer activator 3 (PEA3), a transcriptional repressor of Her-2/neu, in these cell lines. In transient transfection experiments, a Her-2/neu promoter bearing a PEA3 site–mutated sequence was not subject to negative regulation by GLA in Her-2/neu–overexpressing cell lines. Concurrent treatments of Her-2/neu–overexpressing cancer cells with GLA and the anti–Her-2/neu antibody trastuzumab led to synergistic increases in apoptosis and reduced growth and colony formation.
The oil from seeds of the evening primrose is used in traditional medicine as a treatment for a variety of chronic diseases. This oil (and that from seeds of borage and black currant) contains -linolenic acid (GLA), a member of the -6 family of polyunsaturated fatty acids. GLA exerts selective cytotoxic effects on cancer cells without affecting normal cells (1–5). In addition, exogenous supplementation with GLA sensitizes breast cancer cells to antimitotic drugs and endocrine therapies (6–9). A recent phase II study suggested that GLA may have activity against endocrine-sensitive breast cancer with low systemic toxicity (10), and GLA treatments have led to some tumor responses in other advanced solid malignancies (11–15). Although enhanced lipid peroxidation has been proposed as the main mechanism of action of GLA (1–5), the ultimate molecular pathways underlying GLA's anticancer actions remain largely unknown. A novel molecular explanation concerning the anticancer actions of GLA may relate to its ability to specifically regulate oncoproteins. We recently reported that exogenous supplementation of cultured breast cancer cells with GLA significantly diminished proteolytic cleavage of the extracellular domain of the Her-2/neu–coded p185Her-2/neu tyrosine kinase oncoprotein and, consequently, its activation (16). Considering that activation and overexpression of Her-2/neu oncogene are crucial for the etiology, progression, and cell sensitivity to various treatments in 30% of breast carcinomas (17–32), these findings showed a previously unrecognized mechanism by which GLA might regulate breast cancer cell growth, metastasis formation, and response to chemotherapy and endocrine therapy. However, two main questions remained to be addressed: (1) Does GLA-induced deactivation of p185Her-2/neu relate to GLA-induced changes in Her-2/neu gene expression? (2) Is the ability of GLA to regulate Her-2/neu oncogene a common mechanism of GLA's action against other types of cancer or is it restricted to breast cancer?
To characterize the effects of GLA on the expression of Her-2/neu oncogene, we first treated BT-474 and SK-Br3 breast cancer cells, which naturally contain Her-2/neu oncogene amplification (33,34), with GLA (10 µg/mL for 48 hours). In flow cytometry analyses, levels of cell surface–associated Her-2/neu protein were substantially lower in GLA-treated cells than in vehicle-treated cells (Fig. 1, A). Similarly, immunoblot analysis indicated that GLA treatment led to a substantial reduction in Her-2/neu protein levels in both cell lines (Fig. 1, B).
Although Her-2/neu overexpression was originally attributed solely to erbB-2 gene amplification, an elevation in Her-2/neu mRNA levels per gene copy is also observed in all cell lines that exhibit gene amplification (35). We used reporter gene expression and reverse transcription–polymerase chain reaction (RT-PCR) analyses to characterize the effects of GLA on the transcription of the Her-2/neu gene. Treatment of BT-474 and SK-Br3 cells that had been transfected with a construct containing a luciferase reporter gene driven by a wild-type Her-2/neu promoter fragment with GLA (10 µg/mL for 48 hours) led to a strong reduction in reporter gene expression in both lines (Table 1).
Notes
This work was supported by the Basic, Clinical and Translational Award (BRCTR0403141) from the Susan G. Komen Breast Cancer Foundation (United States), the Breast Cancer Concept Award (BC033538) from the Department of Defense (United States), and by the Special Program For Research Excellence in Breast Cancer Career Development award P50CA89018-03 (to J. A. Menendez) and the RO1 Project of the Special Program For Research Excellence in Breast Cancer Career Development awards P50CA89018–03 (to R. Lupu).
The authors wish to thank Prof. Mien-Chie Hung (The University of Texas M. D. Anderson Cancer Center) for kindly providing Her-2/neu promoter constructs. Trastuzumab was kindly provided by the Evanston Northwestern Healthcare Hospital Pharmacy (Evanston, IL).
Funding to pay the Open Access publication charges for this article was provided by the Northwestern University Breast Cancer Special Program for Research Excellence (SPORE).
References
(1) Horrobin DF. Unsaturated lipids and cancer. In: Horrobin DF, editor. New approaches to cancer treatment. Edinburgh (Scotland): Churchill; 1994. p. 1–29.
(2) Das UN. Gamma-linolenic acid, arachidonic acid, and eicosapentaenoic acid as potential anticancer drugs. Nutrition 1990;6:429–34.[ISI][Medline]
(3) Jiang WG, Bryce RP, Horrobin DF. Essential fatty acids: molecular and cellular basis of their anti-cancer action and clinical implications. Crit Rev Oncol Hematol 1998;27:179–209.[ISI][Medline]
(4) Begin ME, Ells G, Das UN, Horrobin DF. Differential killing of human carcinoma cells supplemented with n-3 and n-6 polyunsaturated fatty acids. J Natl Cancer Inst 1986;77:1053–62.[ISI][Medline]
(5) Begin ME, Ells G, Horrobin DF. Polyunsaturated fatty acid-induced cytotoxicity against tumor cells and its relationship to lipid peroxidation. J Natl Cancer Inst 1988;80:188–94.[Abstract]
(6) Menendez JA, del Mar Barbacid M, Montero S, Sevilla E, Escrich E, Solanas M, et al. Effects of gamma-linolenic acid and oleic acid on paclitaxel cytotoxicity in human breast cancer cells. Eur J Cancer 2001;37:402–13.[ISI][Medline]
(7) Menendez JA, Ropero S, del Barbacid MM, Montero S, Solanas M, Escrich E, et al. Synergistic interaction between vinorelbine and gamma-linolenic acid in breast cancer cells. Breast Cancer Res Treat 2002;72:203–19.[ISI][Medline]
(8) Menendez JA, Ropero S, Lupu R, Colomer R. Omega-6 polyunsaturated fatty acid gamma-linolenic acid (18:3n-6) enhances docetaxel (Taxotere) cytotoxicity in human breast carcinoma cells. Oncol Rep 2004;11:1241–52.[ISI][Medline]
(9) Menendez JA, Colomer R, Lupu R. Omega-6 polyunsaturated fatty acid gamma-linolenic acid (18:3n-6) is a selective estrogen-response modulator in human breast cancer cells: gamma-linolenic acid antagonizes estrogen receptor-dependent transcriptional activity, transcriptionally represses estrogen receptor expression and synergistically enhances tamoxifen and ICI 182,780 (Faslodex) efficacy in human breast cancer cells. Int J Cancer 2004;109:949–54.[CrossRef][ISI][Medline]
(10) Kenny FS, Pinder SE, Ellis IO, Gee JM, Nicholson RI, Bryce RP, et al. Gamma linolenic acid with tamoxifen as primary therapy in breast cancer. Int J Cancer 2000;85:643–48.[CrossRef][ISI][Medline]
(11) van der Merwe CF, Booyens J, Joubert HF, van der Merwe CA. The effect of gamma-linolenic acid, an in vitro cytostatic substance contained in evening primrose oil, on primary liver cancer: a double-blind placebo controlled trial. Prostaglandins Leukot Essent Fatty Acids 1990;40:199–202.[CrossRef][ISI][Medline]
(12) Fearon KC, Falconer JS, Ross JA, Carter DC, Hunter JO, Reynolds PD, et al. An open-label phase I/II dose escalation study of the treatment of pancreatic cancer using lithium gammalinolenate. Anticancer Res 1996;16:867–74.[ISI][Medline]
(13) Harris NM, Crook TJ, Dyer JP, Solomon LZ, Bass P, Cooper AJ, Birch BR. Intravesical meglumine gamma-linolenic acid in superficial bladder cancer: an efficacy study. Eur Urol 2002;42:39–42.[CrossRef][ISI][Medline]
(14) Harris NM, Anderson WR, Lwaleed BA, Cooper AJ, Birch BR, Solomon LZ. Epirubicin and meglumine gamma-linolenic acid: a logical choice of combination therapy for patients with superficial bladder carcinoma. Cancer 2003;97:71–8.[CrossRef][ISI][Medline]
(15) Das UN. Occlusion of infusion vessels on gamma-linolenic acid infusion. Prostaglandins Leukot Essent Fatty Acids 2004;70:23–32.[CrossRef][ISI][Medline]
(16) Menendez JA, Ropero S, Lupu R, Colomer R. Dietary fatty acids regulate the activation status of Her-2/neu (c-erbB-2) oncogene in breast cancer cells. Ann Oncol 2004;15:1719–21.[Free Full Text]
(17) Slamon DJ, Clark GM, Wong SG, Levin WJ, Ullrich A, McGuire WL. Human breast cancer: correlation of relapse and survival with amplification of the HER-2/neu oncogene. Science 1987;235:177–82.[ISI][Medline]
(18) Slamon DJ, Godolphin W, Jones LA, Holt JA, Wong SG, Keith DE, et al. Studies of the HER-2/neu proto-oncogene in human breast and ovarian cancer. Science 1989;244:707–12.[ISI][Medline]
(19) Yarden Y. Biology of HER2 and its importance in breast cancer. Oncology 2001;61 Suppl 2:1–13.
(20) Ross JS, McKenna BJ. The HER-2/neu oncogene in tumors of the gastrointestinal tract. Cancer Invest 2001;19:554–68.[CrossRef][ISI][Medline]
(21) Xu R, Perle MA, Inghirami G, Chan W, Delgado Y, Feiner H. Amplification of Her-2/neu gene in Her-2/neu-overexpressing and -nonexpressing breast carcinomas and their synchronous benign, premalignant, and metastatic lesions detected by FISH in archival material. Mod Pathol 2002;15:116–24.[CrossRef][ISI][Medline]
(22) Hoque A, Sneige N, Sahin AA, Menter DG, Bacus JW, Hortobagyi GN, et al. Her-2/neu gene amplification in ductal carcinoma in situ of the breast. Cancer Epidemiol Biomarkers Prev 2002;11:587–90.[Abstract/Free Full Text]
(23) Tan M, Yao J, Yu D. Overexpression of the c-erbB-2 gene enhanced intrinsic metastasis potential in human breast cancer cells without increasing their transformation abilities. Cancer Res 1997;57:1199–205.[Abstract]
(24) Eccles SA. The role of c-erbB-2/HER2/neu in breast cancer progression and metastasis. J Mammary Gland Biol Neoplasia 2001;6: 393–406.[CrossRef][ISI][Medline]
(25) Chen X, Yeung TK, Wang Z. Enhanced drug resistance in cells coexpressing ErbB2 with EGF receptor or ErbB3. Biochem Biophys Res Commun 2000;277:757–63.[CrossRef][ISI][Medline]
(26) Orr MS, O'Connor PM, Kohn KW. Effects of c-erbB2 overexpression on the drug sensitivities of normal human mammary epithelial cells. J Natl Cancer Inst 2000;92:987–94.[Abstract/Free Full Text]
(27) Nelson NJ. Can HER2 status predict response to cancer therapy? J Natl Cancer Inst 2000;92:366–67.[Free Full Text]
(28) Valagussa P. HER2 status: a statistician's view. Ann Oncol 2001;12 Suppl 1:S29–S34.[Medline]
(29) Yamauchi H, Stearns V, Hayes DF. The role of c-erbB-2 as a predictive factor in breast cancer. Breast Cancer 2001;8:171–83.[Medline]
(30) Yu D, Hung MC. Role of erbB2 in breast cancer chemosensitivity. Bioessays 2000;22: 673–80.[CrossRef][ISI][Medline]
(31) Yu D, Hung MC. Overexpression of ErbB2 in cancer and ErbB2-targeting strategies. Oncogene 2000;19:6115–21.[CrossRef][ISI][Medline]
(32) Dowsett M. Overexpression of HER-2 as a resistance mechanism to hormonal therapy for breast cancer. Endocr Relat Cancer 2001;8:191–95.[Abstract/Free Full Text]
(33) Jarvinen TA, Tanner M, Rantanen V, Barlund M, Borg A, Grenman S, et al. Amplification and deletion of topoisomerase IIalpha associate with ErbB-2 amplification and affect sensitivity to topoisomerase II inhibitor doxorubicin in breast cancer. Am J Pathol 2000;156:839–47.[Abstract/Free Full Text]
(34) Yakes FM, Chinratanalab W, Ritter CA, King W, Seelig S, Arteaga CL. Herceptin-induced inhibition of phosphatidylinositol-3 kinase and Akt Is required for antibody-mediated effects on p27, cyclin D1, and antitumor action. Cancer Res 2002;62:4132–41.[Abstract/Free Full Text]
(35) Kraus MH, Popescu NC, Amsbaugh SC, King CR. Overexpression of the EGF receptor-related proto-oncogene erbB-2 in human mammary tumor cell lines by different molecular mechanisms. EMBO J 1987;6:605–10.[Abstract]
(36) Xing X, Wang SC, Xia W, Zou Y, Shao R, Kwong KY, et al. The ets protein PEA3 suppresses HER-2/neu overexpression and inhibits tumorigenesis. Nat Med 2000;6:189–95.[CrossRef][ISI][Medline]
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(39) Carter P, Presta L, Gorman CM, Ridgway JB, Henner D, Wong WL, et al. Humanization of an anti-p185HER2 antibody for human cancer therapy. Proc Natl Acad Sci USA 1992;89:4285–9.[Abstract/Free Full Text]
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(41) Slamon DJ, Leyland-Jones B, Shak S, Fuchs H, Paton V, Bajamonde A, et al. Use of chemotherapy plus a monoclonal antibody against HER2 for metastatic breast cancer that overexpresses HER2. N Engl J Med 2001;344:783–92.[Abstract/Free Full Text]
(42) Bosher JM, Williams T, Hurst HC. The developmentally regulated transcription factor AP-2 is involved in c-erbB-2 overexpression in human mammary carcinoma. Proc Natl Acad Sci USA 1995;92:744–7.[Abstract/Free Full Text]
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(47) Nagata Y, Lan KH, Zhou X, Tan M, Esteva FJ, Sahin AA, et al. PTEN activation contributes to tumor inhibition by trastuzumab, and loss of PTEN predicts trastuzumab resistance in patients. Cancer Cell 2004;6:117–27.[CrossRef][ISI][Medline]
Manuscript received December 21, 2004; revised September 6, 2005; accepted September 8, 2005.
Javier A. Menendez, Luciano Vellon, Ramon Colomer, Ruth Lupu
Affiliations of authors: Department of Medicine, Evanston Northwestern Healthcare Research Institute, Evanston, IL (JAM, LV, RL); Department of Medicine, Northwestern University Feinberg School of Medicine, Chicago, IL (JAM, LV, RL); Robert H. Lurie Comprehensive Cancer Center of Northwestern University, Chicago, IL (JAM, RL); Medical Oncology, Institut Catala d'Oncologia, Hospital Universitari Dr. Josep Trueta, Girona, Spain (RC)
The -6 polyunsaturated fatty acid -linolenic acid (GLA; 18:3n-6), which is found in several plant oils and is used as an herbal medicine, has antitumor activity in vitro. We examined the effect of GLA on the expression of the Her-2/neu (erbB-2) oncogene, which is involved in development of numerous types of human cancer. Flow cytometric and immunoblotting analyses demonstrated that GLA treatment substantially reduced Her-2/neu protein levels in the Her-2/neu–overexpressing cell lines BT-474, SK-Br3, and MDA-MB-453 (breast cancer), SK-OV3 (ovarian cancer), and NCI-N87 (gastrointestinal tumor derived). GLA exposure led to a dramatic decrease in Her-2/neu promoter activity and a concomitant increase in the levels of polyomavirus enhancer activator 3 (PEA3), a transcriptional repressor of Her-2/neu, in these cell lines. In transient transfection experiments, a Her-2/neu promoter bearing a PEA3 site–mutated sequence was not subject to negative regulation by GLA in Her-2/neu–overexpressing cell lines. Concurrent treatments of Her-2/neu–overexpressing cancer cells with GLA and the anti–Her-2/neu antibody trastuzumab led to synergistic increases in apoptosis and reduced growth and colony formation.
The oil from seeds of the evening primrose is used in traditional medicine as a treatment for a variety of chronic diseases. This oil (and that from seeds of borage and black currant) contains -linolenic acid (GLA), a member of the -6 family of polyunsaturated fatty acids. GLA exerts selective cytotoxic effects on cancer cells without affecting normal cells (1–5). In addition, exogenous supplementation with GLA sensitizes breast cancer cells to antimitotic drugs and endocrine therapies (6–9). A recent phase II study suggested that GLA may have activity against endocrine-sensitive breast cancer with low systemic toxicity (10), and GLA treatments have led to some tumor responses in other advanced solid malignancies (11–15). Although enhanced lipid peroxidation has been proposed as the main mechanism of action of GLA (1–5), the ultimate molecular pathways underlying GLA's anticancer actions remain largely unknown. A novel molecular explanation concerning the anticancer actions of GLA may relate to its ability to specifically regulate oncoproteins. We recently reported that exogenous supplementation of cultured breast cancer cells with GLA significantly diminished proteolytic cleavage of the extracellular domain of the Her-2/neu–coded p185Her-2/neu tyrosine kinase oncoprotein and, consequently, its activation (16). Considering that activation and overexpression of Her-2/neu oncogene are crucial for the etiology, progression, and cell sensitivity to various treatments in 30% of breast carcinomas (17–32), these findings showed a previously unrecognized mechanism by which GLA might regulate breast cancer cell growth, metastasis formation, and response to chemotherapy and endocrine therapy. However, two main questions remained to be addressed: (1) Does GLA-induced deactivation of p185Her-2/neu relate to GLA-induced changes in Her-2/neu gene expression? (2) Is the ability of GLA to regulate Her-2/neu oncogene a common mechanism of GLA's action against other types of cancer or is it restricted to breast cancer?
To characterize the effects of GLA on the expression of Her-2/neu oncogene, we first treated BT-474 and SK-Br3 breast cancer cells, which naturally contain Her-2/neu oncogene amplification (33,34), with GLA (10 µg/mL for 48 hours). In flow cytometry analyses, levels of cell surface–associated Her-2/neu protein were substantially lower in GLA-treated cells than in vehicle-treated cells (Fig. 1, A). Similarly, immunoblot analysis indicated that GLA treatment led to a substantial reduction in Her-2/neu protein levels in both cell lines (Fig. 1, B).
Although Her-2/neu overexpression was originally attributed solely to erbB-2 gene amplification, an elevation in Her-2/neu mRNA levels per gene copy is also observed in all cell lines that exhibit gene amplification (35). We used reporter gene expression and reverse transcription–polymerase chain reaction (RT-PCR) analyses to characterize the effects of GLA on the transcription of the Her-2/neu gene. Treatment of BT-474 and SK-Br3 cells that had been transfected with a construct containing a luciferase reporter gene driven by a wild-type Her-2/neu promoter fragment with GLA (10 µg/mL for 48 hours) led to a strong reduction in reporter gene expression in both lines (Table 1).
Notes
This work was supported by the Basic, Clinical and Translational Award (BRCTR0403141) from the Susan G. Komen Breast Cancer Foundation (United States), the Breast Cancer Concept Award (BC033538) from the Department of Defense (United States), and by the Special Program For Research Excellence in Breast Cancer Career Development award P50CA89018-03 (to J. A. Menendez) and the RO1 Project of the Special Program For Research Excellence in Breast Cancer Career Development awards P50CA89018–03 (to R. Lupu).
The authors wish to thank Prof. Mien-Chie Hung (The University of Texas M. D. Anderson Cancer Center) for kindly providing Her-2/neu promoter constructs. Trastuzumab was kindly provided by the Evanston Northwestern Healthcare Hospital Pharmacy (Evanston, IL).
Funding to pay the Open Access publication charges for this article was provided by the Northwestern University Breast Cancer Special Program for Research Excellence (SPORE).
References
(1) Horrobin DF. Unsaturated lipids and cancer. In: Horrobin DF, editor. New approaches to cancer treatment. Edinburgh (Scotland): Churchill; 1994. p. 1–29.
(2) Das UN. Gamma-linolenic acid, arachidonic acid, and eicosapentaenoic acid as potential anticancer drugs. Nutrition 1990;6:429–34.[ISI][Medline]
(3) Jiang WG, Bryce RP, Horrobin DF. Essential fatty acids: molecular and cellular basis of their anti-cancer action and clinical implications. Crit Rev Oncol Hematol 1998;27:179–209.[ISI][Medline]
(4) Begin ME, Ells G, Das UN, Horrobin DF. Differential killing of human carcinoma cells supplemented with n-3 and n-6 polyunsaturated fatty acids. J Natl Cancer Inst 1986;77:1053–62.[ISI][Medline]
(5) Begin ME, Ells G, Horrobin DF. Polyunsaturated fatty acid-induced cytotoxicity against tumor cells and its relationship to lipid peroxidation. J Natl Cancer Inst 1988;80:188–94.[Abstract]
(6) Menendez JA, del Mar Barbacid M, Montero S, Sevilla E, Escrich E, Solanas M, et al. Effects of gamma-linolenic acid and oleic acid on paclitaxel cytotoxicity in human breast cancer cells. Eur J Cancer 2001;37:402–13.[ISI][Medline]
(7) Menendez JA, Ropero S, del Barbacid MM, Montero S, Solanas M, Escrich E, et al. Synergistic interaction between vinorelbine and gamma-linolenic acid in breast cancer cells. Breast Cancer Res Treat 2002;72:203–19.[ISI][Medline]
(8) Menendez JA, Ropero S, Lupu R, Colomer R. Omega-6 polyunsaturated fatty acid gamma-linolenic acid (18:3n-6) enhances docetaxel (Taxotere) cytotoxicity in human breast carcinoma cells. Oncol Rep 2004;11:1241–52.[ISI][Medline]
(9) Menendez JA, Colomer R, Lupu R. Omega-6 polyunsaturated fatty acid gamma-linolenic acid (18:3n-6) is a selective estrogen-response modulator in human breast cancer cells: gamma-linolenic acid antagonizes estrogen receptor-dependent transcriptional activity, transcriptionally represses estrogen receptor expression and synergistically enhances tamoxifen and ICI 182,780 (Faslodex) efficacy in human breast cancer cells. Int J Cancer 2004;109:949–54.[CrossRef][ISI][Medline]
(10) Kenny FS, Pinder SE, Ellis IO, Gee JM, Nicholson RI, Bryce RP, et al. Gamma linolenic acid with tamoxifen as primary therapy in breast cancer. Int J Cancer 2000;85:643–48.[CrossRef][ISI][Medline]
(11) van der Merwe CF, Booyens J, Joubert HF, van der Merwe CA. The effect of gamma-linolenic acid, an in vitro cytostatic substance contained in evening primrose oil, on primary liver cancer: a double-blind placebo controlled trial. Prostaglandins Leukot Essent Fatty Acids 1990;40:199–202.[CrossRef][ISI][Medline]
(12) Fearon KC, Falconer JS, Ross JA, Carter DC, Hunter JO, Reynolds PD, et al. An open-label phase I/II dose escalation study of the treatment of pancreatic cancer using lithium gammalinolenate. Anticancer Res 1996;16:867–74.[ISI][Medline]
(13) Harris NM, Crook TJ, Dyer JP, Solomon LZ, Bass P, Cooper AJ, Birch BR. Intravesical meglumine gamma-linolenic acid in superficial bladder cancer: an efficacy study. Eur Urol 2002;42:39–42.[CrossRef][ISI][Medline]
(14) Harris NM, Anderson WR, Lwaleed BA, Cooper AJ, Birch BR, Solomon LZ. Epirubicin and meglumine gamma-linolenic acid: a logical choice of combination therapy for patients with superficial bladder carcinoma. Cancer 2003;97:71–8.[CrossRef][ISI][Medline]
(15) Das UN. Occlusion of infusion vessels on gamma-linolenic acid infusion. Prostaglandins Leukot Essent Fatty Acids 2004;70:23–32.[CrossRef][ISI][Medline]
(16) Menendez JA, Ropero S, Lupu R, Colomer R. Dietary fatty acids regulate the activation status of Her-2/neu (c-erbB-2) oncogene in breast cancer cells. Ann Oncol 2004;15:1719–21.[Free Full Text]
(17) Slamon DJ, Clark GM, Wong SG, Levin WJ, Ullrich A, McGuire WL. Human breast cancer: correlation of relapse and survival with amplification of the HER-2/neu oncogene. Science 1987;235:177–82.[ISI][Medline]
(18) Slamon DJ, Godolphin W, Jones LA, Holt JA, Wong SG, Keith DE, et al. Studies of the HER-2/neu proto-oncogene in human breast and ovarian cancer. Science 1989;244:707–12.[ISI][Medline]
(19) Yarden Y. Biology of HER2 and its importance in breast cancer. Oncology 2001;61 Suppl 2:1–13.
(20) Ross JS, McKenna BJ. The HER-2/neu oncogene in tumors of the gastrointestinal tract. Cancer Invest 2001;19:554–68.[CrossRef][ISI][Medline]
(21) Xu R, Perle MA, Inghirami G, Chan W, Delgado Y, Feiner H. Amplification of Her-2/neu gene in Her-2/neu-overexpressing and -nonexpressing breast carcinomas and their synchronous benign, premalignant, and metastatic lesions detected by FISH in archival material. Mod Pathol 2002;15:116–24.[CrossRef][ISI][Medline]
(22) Hoque A, Sneige N, Sahin AA, Menter DG, Bacus JW, Hortobagyi GN, et al. Her-2/neu gene amplification in ductal carcinoma in situ of the breast. Cancer Epidemiol Biomarkers Prev 2002;11:587–90.[Abstract/Free Full Text]
(23) Tan M, Yao J, Yu D. Overexpression of the c-erbB-2 gene enhanced intrinsic metastasis potential in human breast cancer cells without increasing their transformation abilities. Cancer Res 1997;57:1199–205.[Abstract]
(24) Eccles SA. The role of c-erbB-2/HER2/neu in breast cancer progression and metastasis. J Mammary Gland Biol Neoplasia 2001;6: 393–406.[CrossRef][ISI][Medline]
(25) Chen X, Yeung TK, Wang Z. Enhanced drug resistance in cells coexpressing ErbB2 with EGF receptor or ErbB3. Biochem Biophys Res Commun 2000;277:757–63.[CrossRef][ISI][Medline]
(26) Orr MS, O'Connor PM, Kohn KW. Effects of c-erbB2 overexpression on the drug sensitivities of normal human mammary epithelial cells. J Natl Cancer Inst 2000;92:987–94.[Abstract/Free Full Text]
(27) Nelson NJ. Can HER2 status predict response to cancer therapy? J Natl Cancer Inst 2000;92:366–67.[Free Full Text]
(28) Valagussa P. HER2 status: a statistician's view. Ann Oncol 2001;12 Suppl 1:S29–S34.[Medline]
(29) Yamauchi H, Stearns V, Hayes DF. The role of c-erbB-2 as a predictive factor in breast cancer. Breast Cancer 2001;8:171–83.[Medline]
(30) Yu D, Hung MC. Role of erbB2 in breast cancer chemosensitivity. Bioessays 2000;22: 673–80.[CrossRef][ISI][Medline]
(31) Yu D, Hung MC. Overexpression of ErbB2 in cancer and ErbB2-targeting strategies. Oncogene 2000;19:6115–21.[CrossRef][ISI][Medline]
(32) Dowsett M. Overexpression of HER-2 as a resistance mechanism to hormonal therapy for breast cancer. Endocr Relat Cancer 2001;8:191–95.[Abstract/Free Full Text]
(33) Jarvinen TA, Tanner M, Rantanen V, Barlund M, Borg A, Grenman S, et al. Amplification and deletion of topoisomerase IIalpha associate with ErbB-2 amplification and affect sensitivity to topoisomerase II inhibitor doxorubicin in breast cancer. Am J Pathol 2000;156:839–47.[Abstract/Free Full Text]
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Manuscript received December 21, 2004; revised September 6, 2005; accepted September 8, 2005.