Proteins Found in the Triple-Negative Breast Cancer Secretome and Their Therapeutic Potential.

The cancer secretome comprises factors secreted by tumors, including cytokines, growth factors, proteins from the extracellular matrix (ECM), proteases and protease inhibitors, membrane and extracellular vesicle proteins, peptide hormones, and metabolic proteins. Secreted proteins provide an avenue for communication with other tumor cells and stromal cells, and these in turn promote tumor growth and progression. Breast cancer is the most commonly diagnosed cancer in women in the US and worldwide. Triple-negative breast cancer (TNBC) is characterized by its aggressiveness and its lack of expression of the estrogen receptor (ER), progesterone receptor (PR), and HER2, making it unable to be treated with therapies targeting these protein markers, and leaving patients to rely on standard chemotherapy. In order to develop more effective therapies against TNBC, researchers are searching for targetable molecules specific to TNBC. Proteins in the TNBC secretome are involved in wide-ranging cancer-promoting processes, including tumor growth, angiogenesis, inflammation, the EMT, drug resistance, invasion, and development of the premetastatic niche. In this review, we catalog the currently known proteins in the secretome of TNBC tumors and correlate these secreted molecules with potential therapeutic opportunities to facilitate translational research.

The Impact of Recent State and Local Minimum Wage Increases on Nursing Facility Employment.

Various U.S. states and municipalities raised their mandated minimum wages between 2017 and 2019. In some areas, minimum wages became high enough to bind for more professional workers, such as lower paid staff at nursing facilities. We add to the small prior literature on the effects of minimum wages on nursing facility staffing using novel establishment-level data on daily hours worked; these data allow us to examine changes in staffing hours along margins previously unexplored in the minimum wage literature. We find no evidence that minimum wage increases reduced hours worked among lower-paid nurses in nursing facilities. In contrast, we find that increases in state and local minimum wages increased hours worked per resident day by nursing assistants; increases occurred for the average of all days throughout the month and on weekend days. We also find that a higher minimum wage increased the share of days in the month that facilities meet at least 75% of the minimum recommended levels of staffing for nursing assistants. These results lessen concerns that minimum wage hikes may reduce the quality of resident care at nursing facilities.

Do financial incentives matter? Effects of Medicare price shocks on skilled nursing facility care.

Skilled nursing facility (SNF) spending has been one of the fastest growing categories of Medicare spending over the past few decades, and reductions in SNF payments are often recommended as part of Medicare cost containment efforts. Using a quasi-experiment resulting from a policy-driven and facility-specific Medicare payment change, we provide new evidence on how Medicare payment changes affect the amount of SNF care provided to Medicare patients. Specifically, we examine a one-time, plausibly exogenous change in the hospital wage index, an area-level adjustment to SNF payments that affected the majority of SNFs nationwide. Using a panel dataset of SNFs, we model the effects of these payment changes on more than 12,000 SNFs across the United States. We find that increases in Medicare payment rates to SNFs increased the total number of Medicare resident days at SNFs. Specifically, a 5% payment increase raised Medicare resident days by 2.33% at facilities with a 10% Medicare share relative to 0%. Further, the effects were asymmetric: Although Medicare payment increases affected Medicare days, payment decreases did not. Our results have important implications for policies that alter the Medicare base payment rates to SNFs and other health care providers.

Medicare hospital payment adjustments and nursing wages.

Despite the importance of the nursing profession for healthcare delivery, costs, and quality, there is relatively little research on how provider payments to hospitals affect the labor market for nurses. This study deals with the hospital wage index (HWI) adjustment to Medicare hospital payments, an area-level adjustment intended to compensate hospitals in high-cost labor markets. Since the HWI adjustment is based on hospital-reported labor costs, some argue that it incentivizes hospitals in concentrated markets to pay higher wages to nurses and other workers (the "circularity" critique). We investigate this critique using market-level data on the relative wages reported by nurses and hospital-level data on the average hourly wage for healthcare workers. For identification, we exploit a 2005 change in the geographic area used to define labor markets, which resulted in exogenous changes in the ability of some hospitals to influence their area's wage index. We find that worker-reported relative nurse wages and hospital-reported healthcare worker wages are higher in some locations where hospitals experienced increased opportunities to game the circularity of the wage index, but these effects appear to be driven by pre-existing wage growth. Medicare's HWI adjustment method does not appear to suffer from inefficiency due to circularity.

Does Formal Employment Reduce Informal Caregiving?

Using the Survey of Income and Program Participation, we examine the impact of formal employment on informal caregiving. We instrument for individual work hours with state unemployment rates. We find that, among women of prime caregiving ages (40-64 years), working 10% more hours per week reduces the probability of providing informal care by about 2 percentage points. The effects are stronger for more time-intensive caregiving and if care recipients are household members. Our results imply that work-promoting policies have the unintended consequence of reducing informal caregiving in an aging society. Copyright © 2015 John Wiley & Sons, Ltd.

P190B RhoGAP Regulates Chromosome Segregation in Cancer Cells.

Rho GTPases are overexpressed and hyperactivated in many cancers, including breast cancer. Rho proteins, as well as their regulators and effectors, have been implicated in mitosis, and their altered expression promotes mitotic defects and aneuploidy. Previously, we demonstrated that p190B Rho GTPase activating protein (RhoGAP) deficiency inhibits ErbB2-induced mammary tumor formation in mice. Here we describe a novel role for p190B as a regulator of mitosis. We found that p190B localized to centrosomes during interphase and mitosis, and that it is differentially phosphorylated during mitosis. Knockdown of p190B expression in MCF-7 and Hela cells increased the incidence of aberrant microtubule-kinetochore attachments at metaphase, lagging chromosomes at anaphase, and micronucleation, all of which are indicative of aneuploidy. Cell cycle analysis of p190B deficient MCF-7 cells revealed a significant increase in apoptotic cells with a concomitant decrease in cells in G1 and S phase, suggesting that p190B deficient cells die at the G1 to S transition. Chemical inhibition of the Rac GTPase during mitosis reduced the incidence of lagging chromosomes in p190B knockdown cells to levels detected in control cells, suggesting that aberrant Rac activity in the absence of p190B promotes chromosome segregation defects. Taken together, these data suggest that p190B regulates chromosome segregation and apoptosis in cancer cells. We propose that disruption of mitosis may be one mechanism by which p190B deficiency inhibits tumorigenesis.

P190B RhoGAP has pro-tumorigenic functions during MMTV-Neu mammary tumorigenesis and metastasis.

INTRODUCTION: Rho GTPases are overexpressed and hyperactivated in human breast cancers. Deficiency of p190B RhoGAP, a major inhibitor of the Rho GTPases, inhibits mouse mammary tumor virus long terminal repeat (MMTV)-Neu/ErbB2 mammary tumor formation and progression in part through effects within the stromal environment, suggesting that p190B function is pro-tumorigenic. To further investigate the potential pro-tumorigenic actions of p190B, we examined the effects of exogenous p190B expression within the mammary epithelium on MMTV-Neu tumor formation and progression. METHODS: Tetracycline-regulatable p190B transgenic mice were bred to MMTV-Neu mice, and the effects of exogenous p190B expression on tumor latency, multiplicity, growth rates, angiogenesis, and metastasis were examined. The effects of exogenous p190B expression on cell-matrix adhesion and invasion were tested using non-transformed primary mammary epithelial cells (MECs). Rho GTPase activity, oxidative stress as an indicator of reactive oxygen species (ROS) production, and downstream signaling pathways were analyzed. RESULTS: Altered p190B expression resulted in a 2-fold increase in tumor multiplicity and a 3-fold increase in metastases compared to control mice indicating that exogenous p190B expression in the mammary epithelium promotes MMTV-Neu mammary tumor formation and progression. Interestingly, non-transformed primary MECs expressing exogenous p190B displayed increased adhesion to laminin and type IV collagen and formed invasive structures in a three-dimensional culture assay. Ras related C3 botulinum toxin 1 (Rac1)-GTP levels were elevated in p190B transgenic tumors whereas Ras homologous A (RhoA) and cell division cycle 42 (Cdc42)-GTP levels were not significantly altered. Rac1 activity affects production of ROS, which regulate transformation, metastasis, and oxidative stress. Protein carbonylation, which is indicative of oxidative stress, was elevated 1.75-fold in p190B transgenic tumors as compared to control tumors suggesting that exogenous p190B expression may affect Rac1-dependent ROS production. CONCLUSIONS: These studies indicate that paradoxically, p190B RhoGAP, a major inhibitor of the Rho GTPases in vitro, has pro-tumorigenic functions that enhance MMTV-Neu induced mammary tumor formation and metastasis. Furthermore, exogenous p190B expression enhances cell adhesion and invasion, which may facilitate metastasis. Rac1 activity and oxidative stress are elevated in tumors expressing exogenous p190B suggesting that p190B may promote tumorigenesis through a Rac1/ROS dependent mechanism.

Iejimalides A and B inhibit lysosomal vacuolar H+-ATPase (V-ATPase) activity and induce S-phase arrest and apoptosis in MCF-7 cells.

Iejimalides are novel macrolides that are cytostatic or cytotoxic against a wide range of cancer cells at low nanomolar concentrations. A recent study by our laboratory characterized the expression of genes and proteins that determine the downstream effects of iejimalide B. However, little is known about the cellular target(s) of iejimalide or downstream signaling that lead to cell-cycle arrest and/or apoptosis. Iejimalides have been shown to inhibit the activity of vacuolar H(+)-ATPase (V-ATPase) in osteoclasts, but how this inhibition may lead to cell-cycle arrest and/or apoptosis in epithelial cells is not known. In this study, MCF-7 breast cancer cells were treated with iejimalide A or B and analyzed for changes in cell-cycle dynamics, apoptosis, lysosomal pH, cytoplasmic pH, mitochondrial membrane potential, and generation of reactive oxygen species. Both iejimalides A and B sequentially neutralize the pH of lysosomes, induce S-phase cell-cycle arrest, and trigger apoptosis in MCF-7 cells. Apoptosis occurs through a mechanism that involves oxidative stress and mitochondrial depolarization but not cytoplasmic acidification. These data confirm that iejimalides inhibit V-ATPase activity in the context of epithelial tumor cells, and that this inhibition may lead to a lysosome-initiated cell death process.

Haploinsufficiency for p190B RhoGAP inhibits MMTV-Neu tumor progression.

INTRODUCTION: Rho signaling regulates key cellular processes including proliferation, survival, and migration, and it has been implicated in the development of many types of cancer including breast cancer. P190B Rho GTPase activating protein (RhoGAP) functions as a major inhibitor of the Rho GTPases. P190B is required for mammary gland morphogenesis, and overexpression of p190B in the mammary gland induces hyperplastic lesions. Hence, we hypothesized that p190B may play a pivotal role in mammary tumorigenesis. METHODS: To investigate the effects of loss of p190B function on mammary tumor progression, p190B heterozygous mice were crossed with an MMTV-Neu breast cancer model. Effects of p190B deficiency on tumor latency, multiplicity, growth, preneoplastic progression and metastasis were evaluated. To investigate potential differences in tumor angiogenesis between the two groups, immunohistochemistry to detect von Willebrand factor was performed and quantified. To examine gene expression of potential mediators of the angiogenic switch, an angiogenesis PCR array was utilized and results were confirmed using immunohistochemistry. Finally, reciprocal transplantation of tumor fragments was performed to determine the impact of stromal deficiency of p190B on tumor angiogenesis. RESULTS: P190B deficiency reduced tumor penetrance (53% of p190B+/-Neu mice vs. 100% of p190B+/+Neu mice formed tumors) and markedly delayed tumor onset by an average of 46 weeks. Tumor multiplicity was also decreased, but an increase in the number of preneoplastic lesions was detected indicating that p190B deficiency inhibited preneoplastic progression. Angiogenesis was decreased in the p190B heterozygous tumors, and expression of a potent angiogenic inhibitor, thrombospondin-1, was elevated in p190B+/-Neu mammary glands. Transplantation of p190B+/-Neu tumor fragments into wild-type recipients restored tumor angiogenesis. Strikingly, p190B+/+Neu tumor fragments were unable to grow when transplanted into p190B+/-Neu recipients. CONCLUSIONS: These data suggest that p190B haploinsufficiency in the epithelium inhibits MMTV-Neu tumor initiation. Furthermore, p190B deficiency in the vasculature is responsible, in part, for the inhibition of MMTV-Neu tumor progression.

Effects of Iejimalide B, a marine macrolide, on growth and apoptosis in prostate cancer cell lines.

Iejimalide B, a marine macrolide, causes growth inhibition in a variety of cancer cell lines at nanomolar concentrations. We have investigated the effects of Iejimalide B on cell cycle kinetics and apoptosis in the p53+/AR+ LNCaP and p53-/AR- PC-3 prostate cancer cell lines. Iejimalide B, has a dose and time dependent effect on cell number (as measured by crystal violet assay) in both cell lines. In LNCaP cells Iejimalide B induces a dose dependent G0/G1 arrest and apoptosis at 48 h (as measured by Apo-BrdU staining). In contrast, Iejimalide B initially induces G0/G1 arrest followed by S phase arrest but does not induce apoptosis in PC-3 cells. qPCR and Western analysis suggests that Iejimalide B modulates the steady state level of many gene products associated with cell cycle (including cyclins D, E, and B and p21(waf1/cip1)) and cell death (including survivin, p21B and BNIP3L) in LNCaP cells. In PC-3 cells Iejimalide B induces the expression of p21(waf1/cip1), down regulates the expression of cyclin A, and does not modulate the expression of the genes associated with cell death. Comparison of the effects of Iejimalide B on the two cell lines suggests that Iejimalide B induces cell cycle arrest by two different mechanisms and that the induction of apoptosis in LNCaP cells is p53-dependent.

Total synthesis of iejimalide B. An application of the Shiina macrolactonization.

The potent anticancer compound iejimalide B (1) was prepared by a total synthesis through a strategy that features Julia olefinations, Wittig olefinations, a Carreira enantioselective alkynylation, a Heck reaction, a Marshall propargylation reaction, a Stille coupling, and a Shiina macrolactonization.