Radiation induces an inflammatory response that results in STAT3-dependent changes in cellular plasticity and radioresistance of breast cancer stem-like cells.

Purpose: Pro-inflammatory cytokines within the tumor microenvironment, such as IL-6, contribute to the maintenance of stem cells and promote their survival following treatment. The IL-6/STAT3 pathway is a key regulator of genes involved in cancer progression. Activation of STAT3 promotes expansion of cancer stem cells in triple negative breast cancer. Radiation has also been shown to expand cancer stem cell populations and can induce stemness in nonstem cells. However, the role of IL-6/STAT3 in radiation-induced changes in cellular plasticity is unclear.Materials and methods: Expression and secretion of IL-6 from triple-negative breast cancer cell lines SUM159PT and MDA-MB-231 were determined after radiation treatment by real-time PCR and ELISA. Activation of STAT3 after radiation was determined by western blotting. Changes in cellular plasticity induced by radiation were determined by examining ALDEFLUOR activity, gene expression analysis of aldehyde dehydrogenase isoforms and mammosphere forming assays with and without the addition of STAT3 inhibitors. To determine the effect of radiation on nonstem cell populations, experiments were also carried out in ALDEFLUOR sorted cells.Results: Radiation induced an inflammatory response in both cell lines that resulted in activation of STAT3. Additionally, radiation induced a stem-like state as evidenced by an increased activity and expression of the ALDH isoforms ALDH1A1 and ALDH1A3, and increased self-renewal capabilities. Radiation increased ALDH activity and self-renewal in non-stem cell (ALDH-) populations, suggesting radiation-induced cellular reprograming. However, inhibition of STAT3 blocked the radiation-induced stem-like state in both ALDEFLUOR positive and negative populations, and enhanced radiosensitivity.Conclusions: Radiation-induced changes in cellular plasticity are STAT3 dependent and may be a potential target to reduce radioresistance in TNBC and improve treatment outcome.

The Impact of Radiation on the Tumor Microenvironment: Effect of Dose and Fractionation Schedules.

In addition to inducing lethal DNA damage in tumor and stromal cells, radiation can alter the interactions of tumor cells with their microenvironment. Recent technological advances in planning and delivery of external beam radiotherapy have allowed delivery of larger doses per fraction (hypofractionation) while minimizing dose to normal tissues with higher precision. The effects of radiation on the tumor microenvironment vary with dose and fractionation schedule. In this review, we summarize the effects of conventional and hypofractionated radiation regimens on the immune system and tumor stroma. We discuss how these interactions may provide therapeutic benefit in combination with targeted therapies. Understanding the differential effects of radiation dose and fractionation can have implications for choice of combination therapies.

A Strategy toward Icetexane Natural Products.

Icetexane diterpenoids are richly complex polycyclic natural products that have been described with a variety of biological activities. We report here a general synthetic approach toward the 6-7-6 tricyclic core structure of these interesting synthetic targets based on a two-step enolate alkylation and ring-closing metathesis reaction sequence.

Co-activation of Hedgehog and Wnt signaling pathways is associated with poor outcomes in triple negative breast cancer.

Hedgehog (HH) and Wnt pathway activation have been implicated in poor prognosis of breast cancer. Crosstalk between these two pathways has been demonstrated to be important in breast cancer progression, however the association between these two pathways and breast cancer survival rate is unknown. The present study comprised a cohort of 36 patients with triple negative breast cancer (TNBC) to investigate co-activation of HH and canonical Wnt pathway in association to patient outcome. All patients had varying degrees of cytoplasmic sonic HH and glioma-associated oncogene homolog (Gli)-1 staining, which positively correlated with tumor stage. Nuclear ß-catenin was additionally correlated to tumor stage. A significant association was observed between nuclear Gli-1 and nuclear ß-catenin. Co-activation of HH and Wnt pathways was associated with poorer prognosis in TNBC patients resulting in a greater risk of early recurrence and decreased overall survival rate compared with patients with only one pathway activated. Therefore, the combined activation status of the HH and Wnt pathways may be a useful prognostic marker for TNBC patients at risk for early recurrence.

The Multifaceted Roles of B Cells in Solid Tumors: Emerging Treatment Opportunities.

The influence of tumor infiltrating lymphocytes on tumor growth and response to therapy is becoming increasingly apparent. While much work has focused on the role of T cell responses in anti-tumor immunity, the role of B cells in solid tumors is much less understood. Tumor infiltrating B cells have been found in a variety of solid tumors, including breast, ovarian, prostate, melanoma, and colorectal cancer. The function of B cells in solid tumors is controversial, with many studies reporting a pro-tumor effect, while other studies demonstrate a role for B cells in the anti-tumor immune response. In this review, we discuss the prognostic ability of B cells in solid tumors as well as the mechanisms by which B cells can either promote or suppress anti-tumor immunity. Additionally, we review current therapeutic strategies that may target both pro- and anti-tumor B cells.

Obesity and Breast Cancer: Do Age, Race and Subtype Matter?

Obesity rates within the United States are on the rise. Obesity is a known risk factor for various diseases, including cancer. Numerous studies have linked obesity to the incidence and treatment outcomes of breast cancer. However, the risk of obesity may vary between breast cancer subtypes and different racial or age groups. In this article, we review the literature regarding the impact of obesity on incidence and response for different subtypes of breast cancer within different population groups.

Wound healing and cancer stem cells: inflammation as a driver of treatment resistance in breast cancer.

The relationship between wound healing and cancer has long been recognized. The mechanisms that regulate wound healing have been shown to promote transformation and growth of malignant cells. In addition, chronic inflammation has been associated with malignant transformation in many tissues. Recently, pathways involved in inflammation and wound healing have been reported to enhance cancer stem cell (CSC) populations. These cells, which are highly resistant to current treatments, are capable of repopulating the tumor after treatment, causing local and systemic recurrences. In this review, we highlight proinflammatory cytokines and developmental pathways involved in tissue repair, whose deregulation in the tumor microenvironment may promote growth and survival of CSCs. We propose that the addition of anti-inflammatory agents to current treatment regimens may slow the growth of CSCs and improve therapeutic outcomes.

Activation of Inflammatory Responses Correlate With Hedgehog Activation and Precede Expansion of Cancer Stem-Like Cells in an Animal Model of Residual Triple Negative Breast Cancer after Neoadjuvant Chemotherapy.

Triple Negative Breast Cancer (TNBC) is characterized as a lack of expression of the hormonal receptors, estrogen and progesterone, and Human epidermal growth factor receptor 2 (HER2) and as such is unresponsive to current targeted therapy. Resistance of breast cancers to treatment is thought to be due to a sub-population of tumor cells called Breast Cancer Stem Cells (BCSCs) and contributes to poor prognosis and increased risk of recurrence. Previously, we have shown that hedgehog activation is induced by chemotherapy and promotes expansion of a stem-like population in breast cancer cell lines. In addition, chemotherapy can induce an inflammatory response and inflammatory factors can lead to activation of Hedgehog (HH) at sites of tissue injury. Therefore, we wanted to investigate how chemotherapy altered hedgehog signaling and correlated with the release of inflammatory cytokines in a mouse model of breast cancer. Patient derived triple negative breast tumor bearing mice were treated with weekly doses of docetaxel. Following treatment, tumor volume decreased reaching a nadir around 15 days after the start of treatment and increased back to pre-treatment size 35-39 days post treatment. Immunohistochemical staining of mice tumors revealed that Sonic hedgehog and nuclear Gli-1 expression transiently increased following docetaxel treatment, reached peak expression at day 8, and subsequently decreased to almost pre-treatment levels following regrowth of the tumor. Similarly, Interleukin 6 (IL-6) and Interleukin 8 (IL-8) expression transiently increased, peaked around day 8, and decreased upon tumor regrowth, however, remained above pre-treatment levels. Expression of the stem cell marker ALDH1A3 proceeded activation of hedgehog signaling and expression of inflammatory cytokines, increasing around day 15 post treatment and continued to be elevated during tumor regrowth. Thus, chemotherapy treatment resulted in activation of the hedgehog pathway and release of inflammatory cytokines leading to long-term expansion of ALDH1A3 positive stem cells, which can contribute to the regrowth of the tumor and promote resistance to treatment.

Taxane-induced hedgehog signaling is linked to expansion of breast cancer stem-like populations after chemotherapy.

Recurrence of breast cancer after chemotherapy is thought to arise from resistant breast cancer stem cells which are eventually able to repopulate the tumor. The Hedgehog (HH) signaling pathway has been shown to regulate the proliferation and survival of breast cancer stem cells, and has been shown to promote resistance to chemotherapy through the activation of multi-drug resistance and pro survival pathways. Here we report that exposure of heterogenous breast cancer cell lines to docetaxel (DOC) resulted in release of Sonic Hedgehog ligand (SHH) and activation of the HH pathway as evidenced by increased expression and nuclear translocation of the downstream effector Gli-1 at 4-24 h after DOC treatment. This activation had little effect on the bulk of the tumor cell population as inhibition of HH signaling failed to increase apoptosis in response to DOC. However, HH pathway activation was required for clonogenic growth of cell lines after DOC. Increases in stemness markers as well as mammosphere formation were observed after treatment with DOC suggesting an increase in the breast cancer stem cell populations. These increases were similar to that of cell lines cultured in the presence of recombinant SHH and could be eliminated by co-treatment with HH inhibitors. These results suggest that HH pathway activation induced by DOC treatment does not have a chemosensitizing effect on the heterogeneous tumor population, but may be required for survival and expansion of breast cancer stem cells after chemotherapy.

Immunohistochemical analysis of aldehyde dehydrogenase isoforms and their association with estrogen-receptor status and disease progression in breast cancer.

In many types of tumors, especially breast tumors, aldehyde dehydrogenase (ALDH) activity has been used to identify cancer stem-like cells within the tumor. The presence and quantity of these cells are believed to predict the response of tumors to chemotherapy. Therefore, identification and eradication of these cells would be necessary to cure the patient. However, there are 19 different ALDH isoforms that could contribute to the enzyme activity. ALDH1A1 and ALDH1A3 are among the isoforms mostly responsible for the increased ALDH activity observed in these stem-like cells, although the main isoforms vary in different tissues and tumor types. In the study reported here, we attempted to determine if ALDH1A1 or ALDH1A3, specifically, correlate with tumor stage, grade, and hormone-receptor status in breast-cancer patients. While there was no significant correlation between ALDH1A1 and any of the parameters tested, we were able to identify a positive correlation between ALDH1A3 and tumor stage in triple-negative cancers. In addition, ALDH1A3 was negatively correlated with estrogen-receptor status. Our data suggest that ALDH1A3 could be utilized as a marker to identify stem-like cells within triple-negative tumors.

Loss of focal adhesion kinase enhances endothelial barrier function and increases focal adhesions.

OBJECTIVE: To determine the role of FAK in the regulation of endothelial barrier function. METHODS: Stable FAK knockdown HLEC were generated by lentiviral infection of FAK shRNA. Measurements of isometric tension and transendothelial electrical resistance were performed. RESULTS: A FAK knockdown human pulmonary endothelial cell line was generated by lentiviral infection with FAK shRNA and resulted in greater than 90% reduction in FAK protein with no change in Pyk2 protein. Loss of FAK altered cell morphology and actin distribution in both pre- and post-confluent endothelial cells. Large, polygonal shaped endothelial cells with randomly organized stress fibers were identified in pre-confluent cultures, while in confluent monolayers, endothelial cells were irregularly shaped with actin bundles present at cell margins. An increase in the number and size of vinculin plaques was detected in FAK-depleted cells. FAK knockdown monolayers generated a greater transendothelial electrical resistance than controls. Thrombin treatment induced similar changes in TER in both FAK knockdown and control cell lines. FAK-depleted endothelial cells developed a higher stable basal isometric tension compared to control monolayers, but the increase in tension stimulated by thrombin does not differ between the cell lines. Basal myosin II regulatory light chain phosphorylation was unaltered in FAK-depleted cells. In addition, loss of FAK enhanced VE-cadherin localization to the cell membrane without altering VE-cadherin protein levels. CONCLUSIONS: The loss of FAK in endothelial cells enhanced cell attachment and strengthened cell-cell contacts resulting in greater basal tension leading to formation of a tighter endothelial monolayer.