Ultrastructural detection of environmental nanoparticles in circulating blood

The increase in the incidence of acute myeloid leukemia (AML) may suggest a possible environmental etiology. PM2.5 was declared by the International Agency for Research on Cancer (WHO organ) a Class I carcinogen. To date, no reports have focused on particulate environmental pollution together with AML. The study investigated the presence and composition of particulate matter in circulating blood with a Environmental Scanning Electron Microscope coupled with an Energy Dispersive Spectroscope for the elemental analysis of the samples. 38 peripheral blood samples, 19 AML cases and 19 healthy controls, were analysed. A significant overload of particulate matter-derived nanoparticles linked or aggregated to blood components was found in AML patients, while almost absent in matched healthy controls. Two tailed Student’s t-test, MANOVA and Principal Component Analysis indicated that the total numbers of aggregates and particles were statistically different between cases and controls (MANOVA,

Benda-BEAM High-Dose Therapy Prior to Auto-SCT is Effective in Resistant/Relapsed DLBCL

Abstract Background: The most important drawback of clinical trials of high-dose therapy (HDT) followed by autologous stem cell transplant (ASCT) in lymphomas is the high heterogeneity of histological entities. Therefore, the statistical power is reduced, and data are not conclusive. We previously demonstrated the safety of a new conditioning regimen with bendamustine, etoposide, cytarabine, and melphalan (BeEAM) prior to ASCT in resistant/relapsed lymphoma patients. This combination of drugs was able to induce a high CR rate in a population that did not have an opportunity of being cured with other therapies. However, that study enrolled both Hodgkin and non-Hodgkin lymphoma patients. Aims: We designed a phase II study to evaluate the efficacy of the BeEAM conditioning in resistant/relapsed diffuse large B-cell non-Hodgkin lymphoma (DLBCL) patients. Patients and methods: The study was registered at European Union Drug Regulating Authorities Clinical Trials (EudraCT) N. 2011-001246-14. Until now, 61 patients (median age 54 years, range 19-69) with resistant/relapsed DLBCL were enrolled. The primary end-point of the study is to evaluate the 1-year complete remission rate. Results: Briefly, 46/61 patients had advanced stage disease (III-IV); 20 were primary refractory and 41 had relapsed after a median number of 2 lines of therapy (range: 1-3). Twenty-one patients had 1 or more relevant comorbidities (range: 1- 5). 30 patients were in II or subsequent CR after salvage therapy, whereas 27 were in PR and 4 had stable or progressive disease. A median number of 5.72x106 CD34+/kg cells (range 2.21-10.60) collected from peripheral blood was reinfused to patients. All patients engrafted, with a median time to ANC>0.5x109/l of 10 days. Median times to achieve a platelet count >20x109/l and >50x109/l were 12 and 17 days respectively. Twenty-two out of 61 patients presented a fever of unknown origin (36%), whereas 24 patients (39%) presented a clinically documented infection. All patients received G-CSF after transplant for a median time of 8 days (range: 8-13). One patient died due to an incomplete hematological recovery after transplant, producing an overall transplant related mortality of 2.7%. Fifty-seven patients are evaluable for response: 48/57 (84%) obtained a CR, 3/57 (5%) a PR, whereas 6/57 (11%) did not respond to therapy. After a median follow-up of 10.5 months after transplant (range 3-37), 6/57 (11%) patients were refractory, 12/57 (21%) relapsed and 39/57 (68%) are still alive, in continuous CR. Conclusion: Our clinical trial was designed to closely resemble real-world treatment for these patients. During the study, we transplanted a similar proportion of the patients that would have received ASCT in a real-world scenario. While we cannot make sound comparisons without head-to-head trials, results from previous studies using HDT regimens in DLCBL have not been as encouraging as ours. Accordingly, our data preliminary provide the evidence that the Benda-BEAM regimen is safe and has promising high efficacy in resistant-relapsed aggressive DLBCL patients. Acknowledgments: The study was supported in part by AIL Pesaro Onlus. Mundipharma Italy is grateful acknowledged for providing Bendamustine free of charge. Disclosures Patriarca: Janssen-Cilag, Celgene, Merck Sharp & Dohme: Honoraria. Zinzani:Gilead: Membership on an entity's Board of Directors or advisory committees; Takeda: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees; J&J: Membership on an entity's Board of Directors or advisory committees; Pfizer: Membership on an entity's Board of Directors or advisory committees

Prolonged exposure to (R)-bicalutamide generates a LNCaP subclone with alteration of mitochondrial genome.

Advanced prostate cancers, initially sensitive to androgen deprivation therapy, frequently progress to the castration-resistant prostate cancer phenotype (CRPC) through mechanisms not yet fully understood. In this study we investigated mitochondrial involvement in the establishment of refractoriness to hormone therapy in two human prostate cancer cell lines. Our data seem to indicate that the androgen-independent phenotype in our experimental model is due, at least in part, to alterations in mitochondrial dynamics and to a breakdown in the Drp-1-mediated mitochondrial network

Prolonged exposure to (R)-bicalutamide generates a LNCaP subclone with alteration of mitochondrial genome

Advanced prostate cancers, initially sensitive to androgen deprivation therapy, frequently progress to the castration-resistant prostate cancer phenotype (CRPC) through mechanisms not yet fully understood. In this study we investigated mitochondrial involvement in the establishment of refractoriness to hormone therapy. Two human prostate cancer cell lines were used, the parental LNCaP and the resistant LNCaP-Rbic, the latter generated after continuous exposure to 20 μM of (R)-bicalutamide, the active enantiomer of Casodex®. We observed a significant decrease in mtDNA content and a lower expression of 8 mitochondria-encoded gene transcripts involved in respiratory chain complexes in both cell lines. We also found that (R)-bicalutamide differentially modulated dynamin-related protein (Drp-1) expression in LNCaP and LNCaP-Rbic cells. These data seem to indicate that the androgen-independent phenotype in our experimental model was due, at least in part, to alterations in mitochondrial dynamics and to a breakdown in the Drp-1-mediated mitochondrial network

Effect of Small Molecules Modulating Androgen Receptor (SARMs) in Human Prostate Cancer Models

<div><p>The management of hormone-refractory prostate cancer represents a major challenge in the therapy of this tumor, and identification of novel androgen receptor antagonists is needed to render treatment more effective. We analyzed the activity of two novel androgen receptor antagonists, (<i>S</i>)-<b>11</b> and (<i>R</i>)-<b>9</b>, in <i>in vitro</i> and <i>in vivo</i> experimental models of hormone-sensitive or castration-resistant prostate cancer (CRPC). <i>In vitro</i> experiments were performed on LNCaP, LNCaP-AR, LNCaP-Rbic and VCaP human prostate cancer cells. Cytotoxic activity was assessed by SRB and BrdU uptake, AR transactivation by luciferase reporter assay and PSA levels by Real Time RT-PCR and ELISA assays. Cell cycle progression-related markers were evaluated by western blot. <i>In vivo</i> experiments were performed on SCID mice xenografted with cells with different sensitivity to hormonal treatment. In hormone-sensitive LNCaP and LNCaP-AR cells, the latter expressing high androgen receptor levels, (<i>R</i>)-<b>9</b> and (<i>S</i>)-<b>11</b> exhibited a higher cytotoxic effect compared to that of the reference compound ((<i>R</i>)-bicalutamide), also in the presence of the synthetic androgen R1881. Furthermore, the cytotoxic effect produced by (<i>R</i>)-<b>9</b> was higher than that of (<i>S</i>)-<b>11</b> in the two hormone-resistant LNCaP-AR and VCaP cells. A significant reduction in PSA levels was observed after exposure to both molecules. Moreover, (<i>S</i>)-<b>11</b> and (<i>R</i>)-<b>9</b> inhibited DNA synthesis by blocking the androgen-induced increase in cyclin D1 protein levels. <i>In vivo</i> studies on the toxicological profile of (<i>R</i>)-<b>9</b> did not reveal the presence of adverse events. Furthermore, (<i>R</i>)-<b>9</b> inhibited tumor growth in various <i>in vivo</i> models, especially LNCaP-Rbic xenografts, representative of recurrent disease. Our <i>in vitro</i> results highlight the antitumor activity of the two novel molecules (<i>R</i>)-<b>9</b> and (<i>S</i>)-<b>11</b>, making them a potentially attractive option for the treatment of CRPC.</p></div

(<i>S</i>)-11- and (<i>R</i>)-9-induced inhibition of G1/S progression in androgen-treated LNCaP cells.

<p>In <b>A and B</b>, quiescent LNCaP cells on coverslips were left untreated (control) or treated for 18 hours with the synthetic androgen R1881 (10 nM) in the absence or presence of the indicated antagonists (used at 10 or 20 µM). After <i>in vivo</i> pulsing with 100 µM BrdU, BrdU incorporation was analyzed by immunofluorescence and expressed as % of total nuclei. In <b>A</b> and <b>B</b>, the numbers at the top of each bar represent the mean of three independent experiments (<i>n = 3</i>), with standard deviation (SD) <1. The statistical significance of results in <b>A</b> and <b>B</b> was assessed with the paired <i>t</i> test. <i>P</i> values were <0.005 for cells stimulated with 10 nM R1881. No significance was attributed to the difference in BrdU incorporation between control cells and cells stimulated with 10 nM R1881 in the presence of Casodex®, (<i>S</i>)-<b>11</b> (<b>A</b>) or (<i>R</i>)-<b>9</b> (<b>B</b>). In <b>C</b> and <b>D,</b> quiescent LNCaP cells were left untreated or were treated for the indicated times with 10 nM R1881, in the absence or presence of 10 µM of the indicated antagonists (Casodex®, Cx; (<i>S</i>)-<b>11</b> or (<i>R</i>)-<b>9</b>). Lysate proteins were analyzed by Western blot using antibodies directed against cyclin D1 (Cyc D1) in <b>C,</b> or p27 in <b>D</b>. Filters were stripped and re-probed using the rabbit polyclonal anti CDK-4 antibody as a loading control. Western blots in <b>C</b> and <b>D</b> are representative of two different experiments. In <b>C</b>, a 3.3-fold increase in the levels of 4-hr hormone-induced Cyc D1 expression was detected using the NIH ImageJ program. Casodex, (<i>S</i>)-<b>11</b> and (<i>R</i>)-<b>9</b> inhibited this increase (16% 80% and 66% for Casodex®, for S-11 and R-9, respectively) to varying degrees.</p

Cytotoxic activity <i>in vitro</i>.

<p>Cytotoxic activity of (<i>R</i>)-bicalutamide, (<i>S</i>)-<b>11</b>and (<i>R</i>)-<b>9</b> in human prostate cancer cell lines LNCaP, LNCaP-Rbic, LNCaP-AR and VCaP after a 144-hour exposure, measured by SRB assay (average of three independent experiments). The concentrations (µM) of (<i>R</i>)-bicalutamide, (<i>S</i>)-<b>11</b> and (<i>R</i>)-<b>9</b> causing 50% decrease in cell survival (IC₅₀) are shown to the right of the curves.</p