Beta-blockade enhances anthracycline control of metastasis in triple-negative breast cancer.

Beta-adrenergic blockade has been associated with improved cancer survival in patients with triple-negative breast cancer (TNBC), but the mechanisms of these effects remain unclear. In clinical epidemiological analyses, we identified a relationship between beta-blocker use and anthracycline chemotherapy in protecting against TNBC progression, disease recurrence, and mortality. We recapitulated the effect of beta-blockade on anthracycline efficacy in xenograft mouse models of TNBC. In metastatic 4T1.2 and MDA-MB-231 mouse models of TNBC, beta-blockade improved the efficacy of the anthracycline doxorubicin by reducing metastatic development. We found that anthracycline chemotherapy alone, in the absence of beta-blockade, increased sympathetic nerve fiber activity and norepinephrine concentration in mammary tumors through the induction of nerve growth factor (NGF) by tumor cells. Moreover, using preclinical models and clinical samples, we found that anthracycline chemotherapy up-regulated ß2-adrenoceptor expression and amplified receptor signaling in tumor cells. Neurotoxin inhibition of sympathetic neural signaling in mammary tumors using 6-hydroxydopamine or genetic deletion of NGF or ß2-adrenoceptor in tumor cells enhanced the therapeutic effect of anthracycline chemotherapy by reducing metastasis in xenograft mouse models. These findings reveal a neuromodulatory effect of anthracycline chemotherapy that undermines its potential therapeutic impact, which can be overcome by inhibiting ß2-adrenergic signaling in the tumor microenvironment. Supplementing anthracycline chemotherapy with adjunctive ß2-adrenergic antagonists represents a potential therapeutic strategy for enhancing the clinical management of TNBC.

Stress-driven lymphatic dissemination: An unanticipated consequence of communication between the sympathetic nervous system and lymphatic vasculature.

Chronic stress drives cancer progression, but the routes of metastasis are unclear. We recently demonstrated that chronic stress activates a neural-inflammatory signaling axis to remodel lymphatic vasculature and increase lymph flow. This unanticipated crosstalk between stress and the lymphatic system provides pathways of tumor cell dissemination and accelerates metastasis.

ß2-Adrenoceptors on tumor cells play a critical role in stress-enhanced metastasis in a mouse model of breast cancer.

Chronic stress accelerates metastasis - the main cause of death in cancer patients - through the activation of ß-adrenoceptors (ßARs). We have previously shown that ß2AR signaling in MDA-MB-231(HM) breast cancer cells, facilitates invadopodia formation and invasion in vitro. However, in the tumor microenvironment where many stromal cells also express ßAR, the role of ß2AR signaling in tumor cells in metastasis is unclear. Therefore, to investigate the contribution of ß2AR signaling in tumor cells to metastasis in vivo, we used RNA interference to generate MDA-MB-231(HM) breast cancer cells that are deficient in ß2AR. ß2AR knockdown in tumor cells reduced the proportion of cells with a mesenchymal-like morphology and, as expected, reduced tumor cell invasion in vitro. Conversely, overexpression of ß2AR in low metastatic MCF-7 breast cancer cells induced an invasive phenotype. Importantly, we found that knockdown of ß2AR in tumor cells significantly reduced the impact of stress on metastasis in vivo. These findings highlight a crucial role for ß2AR tumor cell signaling in the adverse effects of stress on metastasis, and indicate that it may be necessary to block ß2AR on tumor cells to fully control metastatic progression.

Chronic stress in mice remodels lymph vasculature to promote tumour cell dissemination.

Chronic stress induces signalling from the sympathetic nervous system (SNS) and drives cancer progression, although the pathways of tumour cell dissemination are unclear. Here we show that chronic stress restructures lymphatic networks within and around tumours to provide pathways for tumour cell escape. We show that VEGFC derived from tumour cells is required for stress to induce lymphatic remodelling and that this depends on COX2 inflammatory signalling from macrophages. Pharmacological inhibition of SNS signalling blocks the effect of chronic stress on lymphatic remodelling in vivo and reduces lymphatic metastasis in preclinical cancer models and in patients with breast cancer. These findings reveal unanticipated communication between stress-induced neural signalling and inflammation, which regulates tumour lymphatic architecture and lymphogenous tumour cell dissemination. These findings suggest that limiting the effects of SNS signalling to prevent tumour cell dissemination through lymphatic routes may provide a strategy to improve cancer outcomes.

ß2-adrenoceptor signaling regulates invadopodia formation to enhance tumor cell invasion.

INTRODUCTION: For efficient metastatic dissemination, tumor cells form invadopodia to degrade and move through three-dimensional extracellular matrix. However, little is known about the conditions that favor invadopodia formation. Here, we investigated the effect of ß-adrenoceptor signaling - which allows cells to respond to stress neurotransmitters - on the formation of invadopodia and examined the effect on tumor cell invasion. METHODS: To characterize the molecular and cellular mechanisms of ß-adrenergic signaling on the invasive properties of breast cancer cells, we used functional cellular assays to quantify invadopodia formation and to evaluate cell invasion in two-dimensional and three-dimensional environments. The functional significance of ß-adrenergic regulation of invadopodia was investigated in an orthotopic mouse model of spontaneous breast cancer metastasis. RESULTS: ß-adrenoceptor activation increased the frequency of invadopodia-positive tumor cells and the number of invadopodia per cell. The effects were selectively mediated by the ß2-adrenoceptor subtype, which signaled through the canonical Src pathway to regulate invadopodia formation. Increased invadopodia occurred at the expense of focal adhesion formation, resulting in a switch to increased tumor cell invasion through three-dimensional extracellular matrix. ß2-adrenoceptor signaling increased invasion of tumor cells from explanted primary tumors through surrounding extracellular matrix, suggesting a possible mechanism for the observed increased spontaneous tumor cell dissemination in vivo. Selective antagonism of ß2-adrenoceptors blocked invadopodia formation, suggesting a pharmacological strategy to prevent tumor cell dissemination. CONCLUSION: These findings provide insight into conditions that control tumor cell invasion by identifying signaling through ß2-adrenoceptors as a regulator of invadopodia formation. These findings suggest novel pharmacological strategies for intervention, by using ß-blockers to target ß2-adrenoceptors to limit tumor cell dissemination and metastasis.

Neural Regulation of Pancreatic Cancer: A Novel Target for Intervention.

The tumor microenvironment is known to play a pivotal role in driving cancer progression and governing response to therapy. This is of significance in pancreatic cancer where the unique pancreatic tumor microenvironment, characterized by its pronounced desmoplasia and fibrosis, drives early stages of tumor progression and dissemination, and contributes to its associated low survival rates. Several molecular factors that regulate interactions between pancreatic tumors and their surrounding stroma are beginning to be identified. Yet broader physiological factors that influence these interactions remain unclear. Here, we discuss a series of preclinical and mechanistic studies that highlight the important role chronic stress plays as a physiological regulator of neural-tumor interactions in driving the progression of pancreatic cancer. These studies propose several approaches to target stress signaling via the ß-adrenergic signaling pathway in order to slow pancreatic tumor growth and metastasis. They also provide evidence to support the use of ß-blockers as a novel therapeutic intervention to complement current clinical strategies to improve cancer outcome in patients with pancreatic cancer.

Chronic stress accelerates pancreatic cancer growth and invasion: a critical role for beta-adrenergic signaling in the pancreatic microenvironment.

Pancreatic cancer cells intimately interact with a complex microenvironment that influences pancreatic cancer progression. The pancreas is innervated by fibers of the sympathetic nervous system (SNS) and pancreatic cancer cells have receptors for SNS neurotransmitters which suggests that pancreatic cancer may be sensitive to neural signaling. In vitro and non-orthotopic in vivo studies showed that neural signaling modulates tumour cell behavior. However the effect of SNS signaling on tumor progression within the pancreatic microenvironment has not previously been investigated. To address this, we used in vivo optical imaging to non-invasively track growth and dissemination of primary pancreatic cancer using an orthotopic mouse model that replicates the complex interaction between pancreatic tumor cells and their microenvironment. Stress-induced neural activation increased primary tumor growth and tumor cell dissemination to normal adjacent pancreas. These effects were associated with increased expression of invasion genes by tumor cells and pancreatic stromal cells. Pharmacological activation of ß-adrenergic signaling induced similar effects to chronic stress, and pharmacological ß-blockade reversed the effects of chronic stress on pancreatic cancer progression. These findings indicate that neural ß-adrenergic signaling regulates pancreatic cancer progression and suggest ß-blockade as a novel strategy to complement existing therapies for pancreatic cancer.

Lymphovascular and neural regulation of metastasis: shared tumour signalling pathways and novel therapeutic approaches.

The progression of cancer is supported by a wide variety of non-neoplastic cell types which make up the tumour stroma, including immune cells, endothelial cells, cancer-associated fibroblasts and nerve fibres. These host cells contribute molecular signals that enhance primary tumour growth and provide physical avenues for metastatic dissemination. This article provides an overview of the role of blood vessels, lymphatic vessels and nerve fibres in the tumour microenvironment and highlights the interconnected molecular signalling pathways that control their development and activation in cancer. Further, this article highlights the known pharmacological agents which target these pathways and discusses the potential therapeutic uses of drugs that target angiogenesis, lymphangiogenesis and stress-response pathways in the different stages of cancer care.

PEGylation of interferon α2 improves lymphatic exposure after subcutaneous and intravenous administration and improves antitumour efficacy against lymphatic breast cancer metastases.

The efficacy of protein-based therapeutics with indications in the treatment of lymphatic diseases is expected to be improved by enhancing lymphatic disposition. This study was therefore aimed at examining whether PEGylation can usefully be applied to improve the lymphatic uptake of interferon α2 and whether this ultimately translates into improved therapeutic efficacy against lymph-resident cancer. The lymphatic pharmacokinetics of interferon α2b (IFN, 19kDa) and PEGylated interferon α2b (IFN-PEG12, 31kDa) or α2a (IFN-PEG40, 60kDa) was examined in thoracic lymph duct cannulated rats. IFN was poorly absorbed from the SC injection site (Fabs 36%) and showed little uptake into lymph after SC or IV administration (≤1%). In contrast, IFN-PEG12 was efficiently absorbed from the SC injection site (Fabs 82%) and approximately 20% and 8% of the injected dose was recovered in thoracic lymph over 30h after SC or IV administration respectively. IFN-PEG40, however, was incompletely absorbed from the SC injection site (Fabs 23%) and showed similar lymphatic access after SC administration to IFN-PEG12 (21%). The recovery of IFN-PEG40 in thoracic lymph after IV administration, however, was significantly greater (29%) when compared to IV IFN-PEG12. The anti-tumour efficacy of interferon against axillary metastases of a highly lymph-metastatic variant of human breast MDA-MB-231 carcinoma was significantly increased by SC administration of lymph-targeted IFN-PEG12 when compared to the administration of IFN on the ipsilateral side to the axillary metastasis. Optimal PEGylation may therefore represent a viable approach to improving the lymphatic disposition and efficacy of therapeutic proteins against lymphatic diseases.