Expression of HIF-1α and Markers of Angiogenesis Are Not Significantly Different in Triple Negative Breast Cancer Compared to Other Breast Cancer Molecular Subtypes: Implications for Future Therapy

Lamis Yehia, Fouad Boulos, Mark Jabbour, Ziyad Mahfoud, Najla Fakhruddin, Marwan El-Sabban, Lamis Yehia, Fouad Boulos, Mark Jabbour, Ziyad Mahfoud, Najla Fakhruddin, Marwan El-Sabban

Abstract

Introduction: Triple negative breast cancer lacks estrogen, progesterone and epidermal growth factor receptors rendering it refractory to available targetedtherapies. TNBC is associated with central fibrosis and necrosis, both indicators of tumor hypoxia. Hypoxia inducible factor 1α is up-regulated under hypoxia and its expression is associated with induction of angiogenesis resulting in proliferation, aggressive tumor phenotype and metastasis. In this study we evaluate the potential use of HIF-1α as aTNBC-specific marker.

Methods: 62 TNBC, 64 HER2+, and 64 hormone-receptors positive breast cancer cases were evaluated for central fibrosis and necrosis, HIF-1α, HIF-1β, VEGFR3, CD31 expression and microvessel density. RNA extraction from paraffin-embedded samples, followed by quantitative real-time polymerase chain reaction (qRT-PCR) evaluation of HIF-1α and VEGF transcripts was performed on 54 cases (18 from each subtype).

Results: HIF-1α protein was expressed in 35.5% TNBC, 45.3% HER2+and 25.0% ER+/PR+ (p = 0.055; χ2 test). PCRanalysis of subgroup of breast cancers, 84.2% expressed HIF-1α protein and its transcripts, while only 66.7% expressed VEGF transcripts simultaneously with the HIF-1α protein and its transcripts. Central fibrosis and necrosis was highest in TNBC (p = 0.015; χ2 test), while MVD was comparable among all groups (p = 0.928; χ2 test). VEGFR3 was highest in TNBC expressing HIF-1α. HIF-1β protein was expressed in 32.0% of HIF-1α(+), and in (44.3%) of HIF-1α(-) breast cancer cases (p = 0.033; χ2 test). Moreover, HIF-1α expression in cases with central fibrosis and necrosis was highest in the HER2+ followed by the TNBC (p = 0.156; χ2 test).

Conclusions: A proportion of TNBC express HIF-1α but not in a significantly different manner from other breast cancer subtypes. The potential of anti-HIF-1α targeted therapy is therefore not a candidate for exclusive use in TNBC, but should be considered in all breast cancers, especially in the setting of clinically aggressive or refractory disease.

Conflict of interest statement

Competing Interests: "The authors have declared that no competing interests exist."

Figures

Fig 1. HIF-1α fold increase mRNA expression…
Fig 1. HIF-1α fold increase mRNA expression in TNBC, HER2+, ER+/PR+.
Fig 2. The percentage of TNBC and…
Fig 2. The percentage of TNBC and Her2 + cases with positivenuclearexpression of HIF-1α that expressed HIF-1α and VEGF transcripts fold change.
Fig 3. HIF-1αexpression in breast ductal carcinoma…
Fig 3. HIF-1αexpression in breast ductal carcinoma cells with negative (A) and weak (1+ and 2+) expression (B & C) (400x magnification).
Fig 4. HIF-1β nuclear immunoexpression at various…
Fig 4. HIF-1β nuclear immunoexpression at various intensities showing negative staining (A), weak (1+) staining (B), moderate (2+) staining (C) and strong (3+) staining (D).

References

    1. Hockel M, Schlenger K, Aral B, Mitze M, Schaffer U, Vaupel P (1996) Association between tumor hypoxia and malignant progression in advanced cancer of the uterine cervix. Cancer Res 56: 4509–4515.
    1. Hockel M, Schlenger K, Hockel S, Aral B, Schaffer U, Vaupel P (1998) Tumor hypoxia in pelvic recurrences of cervical cancer. Int J Cancer 79: 365–369.
    1. Brizel DM, Scully SP, Harrelson JM, Layfield LJ, Bean JM, Prosnitz LR, et al. (1996) Tumor oxygenation predicts for the likelihood of distant metastases in human soft tissue sarcoma. Cancer Res 56: 941–943.
    1. Semenza GL (2000) HIF-1: using two hands to flip the angiogenic switch. Cancer Metastasis Rev 19: 59–65.
    1. Bos R, van Diest PJ, de Jong JS, van der Groep P, van der Valk P, van der Wall E, et al. (2005) Hypoxia-inducible factor-1alpha is associated with angiogenesis, and expression of bFGF, PDGF-BB, and EGFR in invasive breast cancer. Histopathology 46: 31–36.
    1. Wong CC, Gilkes DM, Zhang H, Chen J, Wei H, Chaturvedi P, et al. (2011) Hypoxia-inducible factor 1 is a master regulator of breast cancer metastatic niche formation. Proc Natl Acad Sci U S A 108: 16369–16374. 10.1073/pnas.1113483108
    1. Mimeault M, Batra SK (2013) Hypoxia-inducing factors as master regulators of stemness properties and altered metabolism of cancer- and metastasis-initiating cells. J Cell Mol Med 17: 30–54. 10.1111/jcmm.12004
    1. Sorlie T, Perou CM, Tibshirani R, Aas T, Geisler S, Johnsen H, et al. (2001) Gene expression patterns of breast carcinomas distinguish tumor subclasses with clinical implications. Proc Natl Acad Sci U S A 98: 10869–10874.
    1. Turner NC, Reis-Filho JS (2006) Basal-like breast cancer and the BRCA1 phenotype. Oncogene 25: 5846–5853.
    1. van der Groep P, Bouter A, Menko FH, van der Wall E, van Diest PJ (2008) High frequency of HIF-1alpha overexpression in BRCA1 related breast cancer. Breast Cancer Res Treat 111: 475–480.
    1. Haffty BG, Yang Q, Reiss M, Kearney T, Higgins SA, Weidhaas J, et al. (2006) Locoregional relapse and distant metastasis in conservatively managed triple negative early-stage breast cancer. J Clin Oncol 24: 5652–5657.
    1. Harris LN, Broadwater G, Lin NU, Miron A, Schnitt SJ, Cowan D, et al. (2006) Molecular subtypes of breast cancer in relation to paclitaxel response and outcomes in women with metastatic disease: results from CALGB 9342. Breast Cancer Res 8: R66
    1. Bauer KR, Brown M, Cress RD, Parise CA, Caggiano V (2007) Descriptive analysis of estrogen receptor (ER)-negative, progesterone receptor (PR)-negative, and HER2-negative invasive breast cancer, the so-called triple-negative phenotype: a population-based study from the California cancer Registry. Cancer 109: 1721–1728.
    1. Brown NS, Bicknell R (2001) Hypoxia and oxidative stress in breast cancer. Oxidative stress: its effects on the growth, metastatic potential and response to therapy of breast cancer. Breast Cancer Res 3: 323–327.
    1. Carey LA, Dees EC, Sawyer L, Gatti L, Moore DT, Collichio F, et al. (2007) The triple negative paradox: primary tumor chemosensitivity of breast cancer subtypes. Clin Cancer Res 13: 2329–2334.
    1. Dent R, Trudeau M, Pritchard KI, Hanna WM, Kahn HK, Sawka CA, et al. (2007) Triple-negative breast cancer: clinical features and patterns of recurrence. Clin Cancer Res 13: 4429–4434.
    1. Morris GJ, Naidu S, Topham AK, Guiles F, Xu Y, MacCue P, et al. (2007) Differences in breast carcinoma characteristics in newly diagnosed African-American and Caucasian patients: a single-institution compilation compared with the National Cancer Institute's Surveillance, Epidemiology, and End Results database. Cancer 110: 876–884.
    1. Rakha EA, El-Sayed ME, Green AR, Lee AH, Robertson JF, Ellis IO (2007) Prognostic markers in triple-negative breast cancer. Cancer 109: 25–32.
    1. Tischkowitz M, Brunet JS, Begin LR, Huntsman DG, Cheang MC, et al. (2007) Use of immunohistochemical markers can refine prognosis in triple negative breast cancer. BMC Cancer 7: 134
    1. Reis-Filho JS, Tutt AN (2008) Triple negative tumours: a critical review. Histopathology 52: 108–118. 10.1111/j.1365-2559.2007.02889.x
    1. Carvalho FM, Bacchi LM, Santos PP, Bacchi CE (2010) Triple-negative breast carcinomas are a heterogeneous entity that differs between young and old patients. Clinics (Sao Paulo) 65: 1033–1036.
    1. Nielsen TO, Hsu FD, Jensen K, Cheang M, Karaca G, Hu Z, et al. (2004) Immunohistochemical and clinical characterization of the basal-like subtype of invasive breast carcinoma. Clin Cancer Res 10: 5367–5374.
    1. Ishihara A, Tsuda H, Kitagawa K, Yoneda M, Shiraishi T (2009) Morphological characteristics of basal-like subtype of breast carcinoma with special reference to cytopathological features. Breast Cancer 16: 179–185. 10.1007/s12282-009-0108-x
    1. Tsuda H, Takarabe T, Hasegawa T, Murata T, Hirohashi S (1999) Myoepithelial differentiation in high-grade invasive ductal carcinomas with large central acellular zones. Hum Pathol 30: 1134–1139.
    1. Tsuda H, Takarabe T, Hasegawa F, Fukutomi T, Hirohashi S (2000) Large, central acellular zones indicating myoepithelial tumor differentiation in high-grade invasive ductal carcinomas as markers of predisposition to lung and brain metastases. Am J Surg Pathol 24: 197–202.
    1. Lakhani SR, Reis-Filho JS, Fulford L, Penault-Llorca F, van der Vijver M, Parry S, et al. (2005) Prediction of BRCA1 status in patients with breast cancer using estrogen receptor and basal phenotype. Clin Cancer Res 11: 5175–5180.
    1. Livasy CA, Karaca G, Nanda R, Tretiakova MS, Olopade OI, Moore DT, et al. (2006) Phenotypic evaluation of the basal-like subtype of invasive breast carcinoma. Mod Pathol 19: 264–271.
    1. Fulford LG, Easton DF, Reis-Filho JS, Sofronis A, Gillett CE, Lakhani SR, et al. (2006) Specific morphological features predictive for the basal phenotype in grade 3 invasive ductal carcinoma of breast. Histopathology 49: 22–34.
    1. Hammond ME, Hayes DF, Dowsett M, Allred DC, Hagerty KL, Badve S, et al. (2010) American Society of Clinical Oncology/College Of American Pathologists guideline recommendations for immunohistochemical testing of estrogen and progesterone receptors in breast cancer. J Clin Oncol 28: 2784–2795. 10.1200/JCO.2009.25.6529
    1. Wolff AC, Hammond ME, Schwartz JN, Hagerty KL, Allred DC, Cote RJ, et al. (2007) American Society of Clinical Oncology/College of American Pathologists guideline recommendations for human epidermal growth factor receptor 2 testing in breast cancer. J Clin Oncol 25: 118–145.
    1. Bos R, van der Groep P, Greijer AE, Shvarts A, Meijer S, Pinedo HM, et al. (2003) Levels of hypoxia-inducible factor-1alpha independently predict prognosis in patients with lymph node negative breast carcinoma. Cancer 97: 1573–1581.
    1. Mouawad R, Spano JP, Comperat E, Capron F, Khayat D (2009) Tumoural expression and circulating level of VEGFR-3 (Flt-4) in metastatic melanoma patients: correlation with clinical parameters and outcome. Eur J Cancer 45: 1407–1414. 10.1016/j.ejca.2008.12.015
    1. dos Santos M, Mercante AM, Louro ID, Goncalves AJ, de Carvalho MB, da Silva EH, et al. (2012) HIF1-alpha expression predicts survival of patients with squamous cell carcinoma of the oral cavity. PLoS One 7: e45228 10.1371/journal.pone.0045228
    1. Elston CW, Ellis IO (1991) Pathological prognostic factors in breast cancer. I. The value of histological grade in breast cancer: experience from a large study with long-term follow-up. Histopathology 19: 403–410.
    1. De Francesco EM, Lappano R, Santolla MF, Marsico S, Caruso A, Magiolini M (2013) HIF-1alpha/GPER signaling mediates the expression of VEGF induced by hypoxia in breast cancer associated fibroblasts (CAFs). Breast Cancer Res 15: R64
    1. Huang LE, Gu J, Schau M, Bunn HF (1998) Regulation of hypoxia-inducible factor 1alpha is mediated by an O2-dependent degradation domain via the ubiquitin-proteasome pathway. Proc Natl Acad Sci U S A 95: 7987–7992.
    1. Forsythe JA, Jiang BH, Iyer NV, Agani F, Leung SW, Koos RD, et al. (1996) Activation of vascular endothelial growth factor gene transcription by hypoxia-inducible factor 1. Mol Cell Biol 16: 4604–4613.
    1. Chappuis PO, Nethercot V, Foulkes WD (2000) Clinico-pathological characteristics of BRCA1- and BRCA2-related breast cancer. Semin Surg Oncol 18: 287–295.
    1. Yan M, Rayoo M, Takano EA, Investigators KC, Fox SB (2009) BRCA1 tumours correlate with a HIF-1alpha phenotype and have a poor prognosis through modulation of hydroxylase enzyme profile expression. Br J Cancer 101: 1168–1174. 10.1038/sj.bjc.6605287
    1. Tan EY, Yan M, Campo L, Han C, Takano E, Turley H, et al. (2009) The key hypoxia regulated gene CAIX is upregulated in basal-like breast tumours and is associated with resistance to chemotherapy. Br J Cancer 100: 405–411. 10.1038/sj.bjc.6604844
    1. Choi J, Jung WH, Koo JS (2013) Metabolism-related proteins are differentially expressed according to the molecular subtype of invasive breast cancer defined by surrogate immunohistochemistry. Pathobiology 80: 41–52. 10.1159/000339513
    1. Toikkanen S, Helin H, Isola J, Joensuu H (1992) Prognostic significance of HER-2 oncoprotein expression in breast cancer: a 30-year follow-up. J Clin Oncol 10: 1044–1048.
    1. Giatromanolaki A, Koukourakis MI, Simopoulos C, Polychronidis A, Gatter KC, Harris AL, et al. (2004) c-erbB-2 related aggressiveness in breast cancer is hypoxia inducible factor-1alpha dependent. Clin Cancer Res 10: 7972–7977.
    1. Laughner E, Taghavi P, Chiles K, Mahon PC, Semenza GL (2001) HER2 (neu) signaling increases the rate of hypoxia-inducible factor 1alpha (HIF-1alpha) synthesis: novel mechanism for HIF-1-mediated vascular endothelial growth factor expression. Mol Cell Biol 21: 3995–4004.
    1. Konecny GE, Meng YG, Untch M, Wang HJ, Bauerfeind I, Epstein M, et al. (2004) Association between HER-2/neu and vascular endothelial growth factor expression predicts clinical outcome in primary breast cancer patients. Clin Cancer Res 10: 1706–1716.
    1. Whelan KA, Schwab LP, Karakashev SV, Franchetti L, Johannes GJ, Seagroves TN, et al. (2013) The oncogene HER2/neu (ERBB2) requires the hypoxia-inducible factor HIF-1 for mammary tumor growth and anoikis resistance. J Biol Chem 288: 15865–15877. 10.1074/jbc.M112.426999
    1. Sotiriou C, Neo SY, McShane LM, Korn EL, Long PM, Jazaeri A, et al. (2003) Breast cancer classification and prognosis based on gene expression profiles from a population-based study. Proc Natl Acad Sci U S A 100: 10393–10398.
    1. Bos R, Zhong H, Hanrahan CF, Mommers EC, Semenza GL, Pinedo HM, et al. (2001) Levels of hypoxia-inducible factor-1 alpha during breast carcinogenesis. J Natl Cancer Inst 93: 309–314.
    1. Kurebayashi J, Otsuki T, Moriya T, Sonoo H (2001) Hypoxia reduces hormone responsiveness of human breast cancer cells. Jpn J Cancer Res 92: 1093–1101.
    1. Stoner M, Saville B, Wormke M, Dean D, Burghardt R, Safe S (2002) Hypoxia induces proteasome-dependent degradation of estrogen receptor alpha in ZR-75 breast cancer cells. Mol Endocrinol 16: 2231–2242.
    1. Kronblad A, Hedenfalk I, Nilsson E, Pahlman S, Landberg G (2005) ERK1/2 inhibition increases antiestrogen treatment efficacy by interfering with hypoxia-induced downregulation of ERalpha: a combination therapy potentially targeting hypoxic and dormant tumor cells. Oncogene 24: 6835–6841.
    1. Tredan O, Galmarini CM, Patel K, Tannock IF (2007) Drug resistance and the solid tumor microenvironment. J Natl Cancer Inst 99: 1441–1454.
    1. Yamamoto Y, Ibusuki M, Okumura Y, Kawasoe T, Kai K, Iyama K, et al. (2008) Hypoxia-inducible factor 1alpha is closely linked to an aggressive phenotype in breast cancer. Breast Cancer Res Treat 110: 465–475.
    1. Cho J, Bahn JJ, Park M, Ahn W, Lee YJ (2006) Hypoxic activation of unoccupied estrogen-receptor-alpha is mediated by hypoxia-inducible factor-1 alpha. J Steroid Biochem Mol Biol 100: 18–23.
    1. Uzzan B, Nicolas P, Cucherat M, Perret GY (2004) Microvessel density as a prognostic factor in women with breast cancer: a systematic review of the literature and meta-analysis. Cancer Res 64: 2941–2955.
    1. Xia Y, Choi HK, Lee K (2012) Recent advances in hypoxia-inducible factor (HIF)-1 inhibitors. Eur J Med Chem 49: 24–40. 10.1016/j.ejmech.2012.01.033
    1. . DIG-HIF1 Pharmacodynamic Trial in Newly Diagnosed Operable Breast Cancer.
    1. Onitilo AA, Engel JM, Greenlee RT, Mukesh BN (2009) Breast cancer subtypes based on ER/PR and Her2 expression: comparison of clinicopathologic features and survival. Clin Med Res 7: 4–13. 10.3121/cmr.2009.825

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