Inhibition of iNOS as a novel effective targeted therapy against triple-negative breast cancer

Abstract

IntroductionTriple-negative breast cancer (TNBC) is an aggressive form of breast cancer with no effective targeted therapy. Inducible nitric oxide synthase (iNOS) is associated with poor survival in patients with breast cancer by increasing tumor aggressiveness. This work aimed to investigate the potential of iNOS inhibitors as a targeted therapy for TNBC. We hypothesized that inhibition of endogenous iNOS would decrease TNBC aggressiveness by reducing tumor initiation and metastasis through modulation of epithelial-mesenchymal transition (EMT)-inducing factors.MethodsiNOS protein levels were determined in 83 human TNBC tissues and correlated with clinical outcome. Proliferation, mammosphere-forming efficiency, migration, and EMT transcription factors were assessed in vitro after iNOS inhibition. Endogenous iNOS targeting was evaluated as a potential therapy in TNBC mouse models.ResultsHigh endogenous iNOS expression was associated with worse prognosis in patients with TNBC by gene expression as well as immunohistochemical analysis. Selective iNOS (1400 W) and pan-NOS (L-NMMA and L-NAME) inhibitors diminished cell proliferation, cancer stem cell self-renewal, and cell migration in vitro, together with inhibition of EMT transcription factors (Snail, Slug, Twist1, and Zeb1). Impairment of hypoxia-inducible factor 1α, endoplasmic reticulum stress (IRE1α/XBP1), and the crosstalk between activating transcription factor 3/activating transcription factor 4 and transforming growth factor β was observed. iNOS inhibition significantly reduced tumor growth, the number of lung metastases, tumor initiation, and self-renewal.ConclusionsConsidering the effectiveness of L-NMMA in decreasing tumor growth and enhancing survival rate in TNBC, we propose a targeted therapeutic clinical trial by re-purposing the pan-NOS inhibitor L-NMMA, which has been extensively investigated for cardiogenic shock as an anti-cancer therapeutic.Electronic supplementary materialThe online version of this article (doi:10.1186/s13058-015-0527-x) contains supplementary material, which is available to authorized users.

Pharmacological Inhibition of NOS Activates ASK1/JNK Pathway Augmenting Docetaxel-Mediated Apoptosis in Triple-Negative Breast Cancer

“…In vitro, iNOS inhibition diminished cell proliferation, cancer stem cell self-renewal, and cell migration in TNBC cell lines. These effects have been replicated in corresponding cell lines in in vivo xenografts (16). In addition, mutations in RPL39 (A14V) and MLF2 (R158W) have been shown to enhance migration in in vitro experiments (17).…”

“…Regimen treatment design followed three, 2-week cycles of docetaxel (20 or 33 mg/kg intraperitoneal on day 1) and NOS inhibition therapy from days 2 to 6 and 9 to 13 [L-NMMA (400 mg/kg oral gavage on days 2 and 9, 200 mg/kg on days 3 to 6 and 10 to 13) þ amlodipine (10 mg/ kg intraperitoneal injection on days 2 to 6 and 9 to 13]. A Caþ channel blocker, amlodipine, was administrated to counteract the effects of NOS inhibition on blood pressure as previously described (16).…”

“…NO and reactive NO-derived species have been implicated in the modulation of carcinogenesis and as a critical determinant of oxidative stress in cells (12,13). In TNBC, increased iNOS expression is related to tumor grade, aggressiveness, and poor prognosis (14)(15)(16). In vitro, iNOS inhibition diminished cell proliferation, cancer stem cell self-renewal, and cell migration in TNBC cell lines.…”

Are NHE1 and inducible nitric oxide synthase involved in human ovarian cancer?

“…Since in several human diseases the NOS activity and pH are altered and affect a variety of processes including tumour cells metabolism [1,3,4,11,12,14], and expression and activity of H + exchangers (mainly NHE1) are increased in tumour cells [4][5][6][7][8], we propose that human ovarian cancer cells will show a potential functional link between increase in NO synthesis and low pHo, but higher pHi (Figure 1). The role of NHE1 seems crucial in this phenomenon, which also generates a pro-inflammatory environment maintaining elevated iNOS expression and NO synthesis.…”

“…To date, expression of the inducible NOS (iNOS) isoform is upregulated and correlates with poor prognosis in breast cancer [11]. Other reports show that NO promotes human ovarian cancer cell growth and inhibits mitochondrial oxidative phosphorylation [12].…”