A pilot study investigating the influence of dietary boron levels on osteoporosis in postmenopausal women.

This study aimed to evaluate the impact of dietary boron on osteoporosis in postmenopausal Jordanian women. Sixty-six women diagnosed with osteoporosis were recruited and data on personal information, dietary habits, medical history, and lifestyle were collected. Bone mineral density, serum calcium, and serum vitamin D measurements were obtained from patient records. This study showed a strong correlation between boron intake and bone mineral density in these women with osteoporosis and a negative correlation between boron intake and serum calcium (p < .05). However, no significant correlation was found between boron intake and various parameters such as serum vitamin D, dietary habits, body mass index (BMI), waist circumference (WC), age of menopause, cases of hysterectomy or oophorectomy, location of fractures, education level, social status, smoking, and physical activity (p > .05). A significant link was found between boron intake and bone mineral density highlighting the importance of nutritional and lifestyle factors affecting bone health. Further research on the specific impact of boron is warranted to better inform dietary interventions for osteoporosis prevention and management.

Monocytic MDSCs exhibit superior immune suppression via adenosine and depletion of adenosine improves efficacy of immunotherapy.

Immune checkpoint inhibitor (ICI) therapy is effective against many cancers for a subset of patients; a large percentage of patients remain unresponsive to this therapy. One contributing factor to ICI resistance is accumulation of monocytic myeloid-derived suppressor cells (M-MDSCs), a subset of innate immune cells with potent immunosuppressive activity against T lymphocytes. Here, using lung, melanoma, and breast cancer mouse models, we show that CD73-expressing M-MDSCs in the tumor microenvironment (TME) exhibit superior T cell suppressor function. Tumor-derived PGE2, a prostaglandin, directly induces CD73 expression in M-MDSCs via both Stat3 and CREB. The resulting CD73 overexpression induces elevated levels of adenosine, a nucleoside with T cell-suppressive activity, culminating in suppression of antitumor CD8+ T cell activity. Depletion of adenosine in the TME by the repurposed drug PEGylated adenosine deaminase (PEG-ADA) increases CD8+ T cell activity and enhances response to ICI therapy. Use of PEG-ADA can therefore be a therapeutic option to overcome resistance to ICIs in cancer patients.

Exosomes from GM-CSF expressing embryonic stem cells are an effective prophylactic vaccine for cancer prevention.

The antigenic similarity between embryos and tumors has raised the idea of using embryonic material as a preventative vaccine against neoplastic disease. Indeed, we have previously reported that a vaccine comprises allogeneic murine embryonic stem cells (ESCs) and murine fibroblasts expressing GM-CSF (to amplify immune responses) successfully blocks the outgrowth of an implantable cancer (Lewis lung carcinoma; LLC) and lung tumors generated in mice using a combination of a mutagen followed by chronic pulmonary inflammation. However, such a vaccine is obviously impractical for application to humans. The use of fibroblasts to generate GM-CSF is needlessly complicated, and intact whole ESCs carry the hazard of generating embryomas/teratomas. Here, we report the successful application of an alternative prophylactic vaccine comprises exosomes derived from murine ESCs engineered to produce GM-CSF. Vaccination of mice with these exosomes significantly slowed or blocked the outgrowth of implanted LLC while control exosomes lacking GM-CSF were ineffective. Examination of tumor-infiltrating immune cells from mice vaccinated with the GM-CSF-expressing exosomes showed robust tumor-reactive CD8+ T effector responses, Th1 cytokine responses, and higher CD8+ T effector/CD4+CD25+Foxp3+ T regulatory cell ratio in the tumors. We conclude that a similar vaccine derived from GM-CSF- expressing human ESCs can be employed as a preventative vaccine for humans with an increased risk of developing cancer.

Lunasin is a novel therapeutic agent for targeting melanoma cancer stem cells.

Recent studies provide compelling evidence that melanoma is initiated and maintained by a small population of malignant cells called cancer-initiating cells (CICs) that exhibit stem-cell-like properties. Observations that CICs have a distinct biology when compared to that of the bulk tumor cells and, importantly, are resistant to chemotherapies and radiation, suggest that CICs are involved in invasion, metastasis, and ultimately relapse. Lunasin, a bioactive peptide present in soybean, has both chemopreventive activity and chemotherapeutic activity against multiple cancer types. In this study, we tested the potential of Lunasin to specifically target CICs in melanoma tumor cell populations. In vitro studies using human melanoma cell lines showed that Lunasin treatment decreased the size of a subpopulation of melanoma cells expressing the surrogate CIC marker, Aldehyde Dehydrogenase, concomitant with a reduction in the ability to form colonies in soft agar assays, and reduced tumor growth in mouse xenografts. Similarly, Lunasin inhibited colony formation by isolated melanoma CICs in soft agar and reduced oncosphere formation in vitro and substantially inhibited tumor growth in mouse xenografts. Mechanistic studies revealed that Lunasin treatment of isolated melanoma CICs induced expression of the melanocyte-associated differentiation markers Tyrosinase and Microphthalmia-associated Transcription Factor concomitant with reduced expression of the stemness factor NANOG. These findings document for the first time that Lunasin has significant therapeutic activity against melanoma by specifically targeting melanoma CICs, and inducing a more differentiated, non-CIC phenotype. Thus, Lunasin may represent a novel therapeutic option for both chemoresistant and advanced metastatic melanoma management.

MIF Is Necessary for Late-Stage Melanoma Patient MDSC Immune Suppression and Differentiation.

Highly aggressive cancers "entrain" innate and adaptive immune cells to suppress antitumor lymphocyte responses. Circulating myeloid-derived suppressor cells (MDSC) constitute the bulk of monocytic immunosuppressive activity in late-stage melanoma patients. Previous studies revealed that monocyte-derived macrophage migration inhibitory factor (MIF) is necessary for the immunosuppressive function of tumor-associated macrophages and MDSCs in mouse models of melanoma. In the current study, we sought to determine whether MIF contributes to human melanoma MDSC induction and T-cell immunosuppression using melanoma patient-derived MDSCs and an ex vivo coculture model of human melanoma-induced MDSC. We now report that circulating MDSCs isolated from late-stage melanoma patients are reliant upon MIF for suppression of antigen-independent T-cell activation and that MIF is necessary for maximal reactive oxygen species generation in these cells. Moreover, inhibition of MIF results in a functional reversion from immunosuppressive MDSC to an immunostimulatory dendritic cell (DC)-like phenotype that is at least partly due to reductions in MDSC prostaglandin E(2) (PGE(2)). These findings indicate that monocyte-derived MIF is centrally involved in human monocytic MDSC induction/immunosuppressive function and that therapeutic targeting of MIF may provide a novel means of inducing antitumor DC responses in late-stage melanoma patients.

5-Aza-2-deoxycytidine and trichostatin A increase COUP-TFII expression in antiestrogen-resistant breast cancer cell lines.

COUP-TFII is reduced in endocrine-resistant breast cancer cells and is negatively associated with tumor grade. Transient re-expression of COUP-TFII restores antiestrogen sensitivity in resistant LCC2 and LCC9 cells and repression of COUP-TFII results in antiestrogen-resistance in MCF-7 endocrine-sensitive cells. We addressed the hypothesis that reduced COUP-TFII expression in endocrine-resistant breast cancer cells results from epigenetic modification. The NR2F2 gene encoding COUP-TFII includes seven CpG islands, including one in the 5' promoter and one in exon 1. Treatment of LCC2 and LCC9 endocrine-resistant breast cancer cells with 5-aza-2'-deoxycytidine (AZA), a DNA methyltransferase (DNMT) inhibitor, +/- trichostatin A (TSA), a histone deacetylase (HDAC) inhibitor, increased COUP-TFII suggesting that the decrease in COUP-TFII is mediated by epigenetic changes. Methylation-specific PCR (MSP) revealed higher methylation of NR2F2 in the first exon in LCC2 and LCC9 cells compared to MCF-7 cells and AZA reduced this methylation. Translational importance is suggested by Cancer Methylome System (CMS) analysis revealing that breast tumors have increased COUP-TFII (NR2F2) promoter and gene methylation versus normal breast.

ß-D-glucan inhibits endocrine-resistant breast cancer cell proliferation and alters gene expression.

Endocrine therapies have been successfully used for breast cancer patients with estrogen receptor α (ERα) positive tumors, but ~40% of patients relapse due to endocrine resistance. ß-glucans are components of plant cell walls that have immunomodulatory and anticancer activity. The objective of this study was to examine the activity of ß-D-glucan, purified from barley, in endocrine-sensitive MCF-7 versus endocrine-resistant LCC9 and LY2 breast cancer cells. ß-D-glucan dissolved in DMSO but not water inhibited MCF-7 cell proliferation in a concentration-dependent manner as measured by BrdU incorporation with an IC50 of ~164 ± 12 µg/ml. ß-D-glucan dissolved in DMSO inhibited tamoxifen/endocrine-resistant LCC9 and LY2 cell proliferation with IC50 values of 4.6 ± 0.3 and 24.2 ± 1.4 µg/ml, respectively. MCF-10A normal breast epithelial cells showed a higher IC50 ~464 µg/ml and the proliferation of MDA-MB-231 triple negative breast cancer cells was not inhibited by ß-D-glucan. Concentration-dependent increases in the BAX/BCL2 ratio and cell death with ß-D-glucan were observed in MCF-7 and LCC9 cells. PCR array analysis revealed changes in gene expression in response to 24-h treatment with 10 or 50 µg/ml ß-D-glucan that were different between MCF-7 and LCC9 cells as well as differences in basal gene expression between the two cell lines. Select results were confirmed by quantitative real-time PCR demonstrating that ß-D-glucan increased RASSF1 expression in MCF-7 cells and IGFBP3, CTNNB1 and ERß transcript expression in LCC9 cells. Our data indicate that ß-D-glucan regulates breast cancer-relevant gene expression and may be useful for inhibiting endocrine-resistant breast cancer cell proliferation.

Reduced expression of miR-200 family members contributes to antiestrogen resistance in LY2 human breast cancer cells.

INTRODUCTION: The role of miRNAs in acquired endocrine-resistant breast cancer is not fully understood. One hallmark of tumor progression is epithelial-to-mesenchymal transition (EMT), characterized by a loss of cell adhesion resulting from reduced E-cadherin and increased cell mobility. miR-200 family members regulate EMT by suppressing expression of transcriptional repressors ZEB1/2. Previously we reported that the expression of miR-200a, miR-200b, and miR-200c was lower in LY2 endocrine-resistant, mesenchymal breast cancer cells compared to parental, endocrine sensitive, epithelial MCF-7 breast cancer cells. Here we investigated the regulation of miR-200 family members and their role in endocrine-sensitivity in breast cancer cells. RESULTS: miR-200 family expression was progressively reduced in a breast cancer cell line model of advancing endocrine/tamoxifen (TAM) resistance. Concomitant with miR-200 decrease, there was an increase in ZEB1 mRNA expression. Overexpression of miR-200b or miR-200c in LY2 cells altered cell morphology to a more epithelial appearance and inhibited cell migration. Further, miR-200b and miR-200c overexpression sensitized LY2 cells to growth inhibition by estrogen receptor (ER) antagonists TAM and fulvestrant. Knockdown of ZEB1 in LY2 cells recapitulated the effect of miR-200b and miR-200c overexpression resulting in inhibition of LY2 cell proliferation by TAM and fulvestrant, but not the aromatase inhibitor exemestane. Demethylating agent 5-aza-2'-deoxycytidine (5-aza-dC) in combination with histone deacetylase inhibitor trichostatin A (TSA) increased miR-200b and miR-200c in LY2 cells. Concomitant with the increase in miR-200b and miR-200c, ZEB1 expression was decreased and cells appeared more epithelial in morphology and were sensitized to TAM and fulvestrant inhibition. Likewise, knockdown of ZEB1 increased antiestrogen sensitivity of LY2 cells resulting in inhibition of cell proliferation. CONCLUSIONS: Our data indicate that reduced miRNA-200b and miR-200c expression contributes to endocrine resistance in breast cancer cells and that the reduced expression of these miR-200 family members in endocrine-resistant cells can be reversed by 5-aza-dC+TSA.

DHEA metabolites activate estrogen receptors alpha and beta.

Dehydroepiandrosterone (DHEA) levels were reported to associate with increased breast cancer risk in postmenopausal women, but some carcinogen-induced rat mammary tumor studies question this claim. The purpose of this study was to determine how DHEA and its metabolites affect estrogen receptors α or ß (ERα or ERß)-regulated gene transcription and cell proliferation. In transiently transfected HEK-293 cells, androstenediol, DHEA, and DHEA-S activated ERα. In ERß transfected HepG2 cells, androstenedione, DHEA, androstenediol, and 7-oxo DHEA stimulated reporter activity. ER antagonists ICI 182,780 (fulvestrant) and 4-hydroxytamoxifen, general P450 inhibitor miconazole, and aromatase inhibitor exemestane inhibited activation by DHEA or metabolites in transfected cells. ERß-selective antagonist R,R-THC (R,R-cis-diethyl tetrahydrochrysene) inhibited DHEA and DHEA metabolite transcriptional activity in ERß-transfected cells. Expression of endogenous estrogen-regulated genes: pS2, progesterone receptor, cathepsin D1, and nuclear respiratory factor-1 was increased by DHEA and its metabolites in an ER-subtype, gene, and cell-specific manner. DHEA metabolites, but not DHEA, competed with 17ß-estradiol for ERα and ERß binding and stimulated MCF-7 cell proliferation, demonstrating that DHEA metabolites interact directly with ERα and ERßin vitro, modulating estrogen target genes in vivo.