Omecamtiv mecarbil in chronic heart failure with reduced ejection fraction, GALACTIC‐HF: baseline characteristics and comparison with contemporary clinical trials

Aims: The safety and efficacy of the novel selective cardiac myosin activator, omecamtiv mecarbil, in patients with heart failure with reduced ejection fraction (HFrEF) is tested in the Global Approach to Lowering Adverse Cardiac outcomes Through Improving Contractility in Heart Failure (GALACTIC‐HF) trial. Here we describe the baseline characteristics of participants in GALACTIC‐HF and how these compare with other contemporary trials. Methods and Results: Adults with established HFrEF, New York Heart Association functional class (NYHA) ≥ II, EF ≤35%, elevated natriuretic peptides and either current hospitalization for HF or history of hospitalization/ emergency department visit for HF within a year were randomized to either placebo or omecamtiv mecarbil (pharmacokinetic‐guided dosing: 25, 37.5 or 50 mg bid). 8256 patients [male (79%), non‐white (22%), mean age 65 years] were enrolled with a mean EF 27%, ischemic etiology in 54%, NYHA II 53% and III/IV 47%, and median NT‐proBNP 1971 pg/mL. HF therapies at baseline were among the most effectively employed in contemporary HF trials. GALACTIC‐HF randomized patients representative of recent HF registries and trials with substantial numbers of patients also having characteristics understudied in previous trials including more from North America (n = 1386), enrolled as inpatients (n = 2084), systolic blood pressure < 100 mmHg (n = 1127), estimated glomerular filtration rate < 30 mL/min/1.73 m2 (n = 528), and treated with sacubitril‐valsartan at baseline (n = 1594). Conclusions: GALACTIC‐HF enrolled a well‐treated, high‐risk population from both inpatient and outpatient settings, which will provide a definitive evaluation of the efficacy and safety of this novel therapy, as well as informing its potential future implementation

Therapeutic Potential of Piperlongumine for Pancreatic Ductal Adenocarcinoma

Pancreatic ductal adenocarcinoma (PDAC) is among the most lethal malignancies because it is often diagnosed at a late disease stage and has a poor response rate to currently available treatments. Therefore, it is critical to develop new therapeutic approaches that will enhance the efficacy and reduce the toxicity of currently used therapies. Here we aimed to evaluate the therapeutic potential and mechanisms of action for piperlongumine (PL), an alkaloid from long pepper, in PDAC models. We postulated that PL causes PDAC cell death through oxidative stress and complements the therapeutic efficacy of chemotherapeutic agents in PDAC cells. First, we determined that PL is one of the most abundant alkaloids with antitumor properties in the long pepper plant. We also showed PL in combination with gemcitabine, a chemotherapy agent used to treat advanced pancreatic cancer, reduced tumor weight and volume compared to vehicle-control and individual treatments. Further, biochemical analysis, including RNA sequencing and immunohistochemistry, suggested that the antitumor activity of PL was associated with decreased cell proliferation, induction of cell cycle arrest, and oxidative stress-induced cell death. Moreover, we identified that c-Jun N-terminal kinase (JNK) inhibition blocks PL-induced cell death, translocation of Nrf2, and transcriptional activation of HMOX1 in PDAC. Finally, high-throughput drug and CRISPR screenings identified potential targets that could be used in combination with PL to treat PDAC cells. Collectively, our data suggests that cell cycle regulators in combination with PL might be an effective approach to combat pancreatic cancer.NI

Abstract A86: Piperlongumine enhances the efficacy of gemcitabine in pancreatic cancer cells in vitro and in vivo

Abstract Reactive oxygen species (ROS)-inducing agents have been identified as possible treatments for various cancers, including pancreatic cancer. Cancer cells have high levels of ROS compared to healthy cells due to excessive metabolic activity. This phenomenon makes ROS a viable target for cancer treatment. We investigated the effect of a known ROS inducer, piperlongumine (PL), a bioactive agent found in long peppers, alone and in combination with a chemotherapeutic agent (gemcitabine; Gem) on pancreatic cancer cells in vitro and in vivo. The pancreatic cancer cell line (MIA PaCa-2) was treated with PL, gemcitabine, or a combination of the agents to evaluate the effect of PL on enhancing the efficacy of gemcitabine. Cell survival was assessed by 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) and propidium iodide (PI) staining, and Matrigel, and wound healing assays were performed. Our in vitro results show (MTT assay); 1uM of PL causes a 90% reduction in cell viability, 1nM of gemcitabine causes a 86% reduction, and the combination of PL (1uM) + gemcitabine (1nM) causes a 66.7% reduction compared to control (normalized to 100%). The PI staining (cell membrane integrity) control was 81% viable, 2.5uM PL treatment was 39.4% viable, 100nM gemcitabine treatment was 69.4% viable, and the combination PL (2.5uM) + gemcitabine (100nM) treatment was 36% viable. For the Matrigel and wound healing assays we observed a significant reduction number of cells and wound distance for wells treated with PL+ gemcitabine compared to the control and individual treatments. We evaluated the therapeutic advantage of combining PL and gemcitabine in nude mice bearing orthotopically implanted MIA PaCa-2 cells. Doses of 5 mg/kg of PL and 25 mg/kg gemcitabine were selected for intraperitoneal administration three times a week. The treatment efficacy was determined by considering the mean pancreatic tumor weight and volume immediately following euthanization. Administration of PL caused a 36% and 67% reduction in tumor weight and volume, respectively. Gemcitabine alone caused a 50% reduction in tumor weight and a 64% reduction in tumor volume. However, the combination of PL and gemcitabine showed a significant decrease (p&amp;lt;0.01) in tumor weight and volume relative to DMSO alone, PL alone, and gemcitabine-alone treated mice. No macroscopic evidence of spreading to other visceral organs was evident for any experimental groups. Our results (in vitro and in vivo) suggest that PL has a potential to be used in combination with gemcitabine to more effectively treat pancreatic cancer. Citation Format: Jiyan Mohammad, Harsharan Dhillon, Katie Reindl.{Authors}. Piperlongumine enhances the efficacy of gemcitabine in pancreatic cancer cells in vitro and in vivo. [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer: Advances in Science and Clinical Care; 2016 May 12-15; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2016;76(24 Suppl):Abstract nr A86.</jats:p

Development of a multiphase perfusion model for biomimetic reduced-order dense tumors

AbstractDense fibrous extracellular constitution of solid tumors exerts high resistance to diffusive transport into it; additionally, the scarcity of blood and lymphatic flows hinders convection. The complexity of fluidic transport mechanisms in such tumor environments still presents open questions with translational end goals. For example, clinical diagnosis and targeted drug delivery platforms for such dense tumors can ideally benefit from a quantitative framework on plasma uptake into the tumor. In this study, we present a computational model for physical parameters that may influence blood percolation and penetration into simple biomimetic solid tumor geometry. The model implements three-phase viscous-laminar transient simulation to mimic the transport physics inside a tumor-adhering blood vessel and measures the constituent volume fractions of the three considered phases, viz. plasma, RBCs (red blood cells, also known as “erythrocytes”), and WBCs (white blood cells, also known as “leukocytes”) at three different flow times, while simultaneously recording the plasma pressure and velocity at the entry point to the tumor’s extracellular space. Subsequently, to quantify plasma perfusion within the tumor zone, we proposed a reduced-order two-dimensional transport model for the tumor entry zone and its extracellular space for three different fenestra diameters: 0.1, 0.3, and 0.5 µm; the simulations were two-phase viscous-laminar transient. The findings support the hypothesis that plasma percolation into the tumor is proportional to the leakiness modulated by the size of fenestra openings, and the rate of percolation decays with the diffusion distance.</jats:p

Computational multiphase characterization of perfusion trends inside biomimetic reduced-order dense tumors

AbstractDense fibrous extracellular constitution of solid tumors exerts high resistance to diffusive transport into it; additionally, the scarcity of blood and lymphatic flows hinders convection. The complexity of fluidic transport mechanisms in such tumor environments still presents open questions with translational end goals. For example, clinical diagnosis and targeted drug delivery platforms for such dense tumors can ideally benefit from a quantitative framework on plasma uptake into the tumor. In this study, we present a computational model for physical parameters that may influence blood percolation and penetration into a simple biomimetic solid tumor geometry. The model implements 3-phase viscous laminar transient simulation to mimic the transport physics inside a tumor-adhering blood vessel and measures the constituent volume fractions of the three considered phases, viz. plasma, RBCs (Red Blood Cells, also known as “erythrocytes”), and WBCs (White Blood Cells, also known as “leukocytes”) at three different flow times, while simultaneously recording the plasma pressure and velocity at the entry point to the tumor’s extracellular space. Subsequently, to quantify plasma perfusion within the tumor zone, we have proposed a reduced-order 2D transport model for the tumor entry zone and its extracellular space for three different fenestra diameters: 0.1, 0.3, and 0.5 μm; the simulations were 2-phase viscous laminar transient. The findings support the hypothesis that plasma percolation into the tumor is proportional to the leakiness modulated by the fenestra openings, quantifiable through the opening sizes.</jats:p

Experimental and Computational Multiphase Flow

Dense fibrous extracellular constitution of solid tumors exerts high resistance to diffusive transport into it; additionally, the scarcity of blood and lymphatic flows hinders convection. The complexity of fluidic transport mechanisms in such tumor environments still presents open questions with translational end goals. For example, clinical diagnosis and targeted drug delivery platforms for such dense tumors can ideally benefit from a quantitative framework on plasma uptake into the tumor. In this study, we present a computational model for physical parameters that may influence blood percolation and penetration into simple biomimetic solid tumor geometry. The model implements three-phase viscous-laminar transient simulation to mimic the transport physics inside a tumor-adhering blood vessel and measures the constituent volume fractions of the three considered phases, viz. plasma, RBCs (red blood cells, also known as "erythrocytes "), and WBCs (white blood cells, also known as "leukocytes ") at three different flow times, while simultaneously recording the plasma pressure and velocity at the entry point to the tumor's extracellular space. Subsequently, to quantify plasma perfusion within the tumor zone, we proposed a reduced-order two-dimensional transport model for the tumor entry zone and its extracellular space for three different fenestra diameters: 0.1, 0.3, and 0.5 mu m; the simulations were two-phase viscous-laminar transient. The findings support the hypothesis that plasma percolation into the tumor is proportional to the leakiness modulated by the size of fenestra openings, and the rate of percolation decays with the diffusion distance.National Institutes of Health (NIH) Center of Biomedical Research Excellence (COBRE); North Dakota State University Center for Diagnostic and Therapeutic Strategies in Pancreatic Cancer [5P20GM109024]Published versionThe reported work has been supported by a National Institutes of Health (NIH) Center of Biomedical Research Excellence (COBRE) Pilot Grant from the North Dakota State University Center for Diagnostic and Therapeutic Strategies in Pancreatic Cancer (Project No. 5P20GM109024). Any opinions, findings, and conclusions or recommendations expressed here are, however, those of the authors, and do not necessarily reflect views of the NIH

Glutathione S-Transferase pi-1 Knockdown Reduces Pancreatic Ductal Adenocarcinoma Growth by Activating Oxidative Stress Response Pathways

Glutathione S-transferase pi-1 (GSTP1) plays an important role in regulating oxidative stress by conjugating glutathione to electrophiles. GSTP1 is overexpressed in breast, colon, lung, and prostate tumors, where it contributes to tumor progression and drug resistance; however, the role of GSTP1 in pancreatic ductal adenocarcinoma (PDAC) is not well understood. Using shRNA, we knocked down GSTP1 expression in three different PDAC cell lines and determined the effect on cell proliferation, cell cycle progression, and reactive oxygen species (ROS) levels. Our results show GSTP1 knockdown reduces PDAC cell growth, prolongs the G0/G1 phase, and elevates ROS in PDAC cells. Furthermore, GSTP1 knockdown results in the increased phosphorylation of c-Jun N-terminal kinase (JNK) and c-Jun and the decreased phosphorylation of extracellular signal-regulated kinase (ERK), p65, the reduced expression of specificity protein 1 (Sp1), and the increased expression of apoptosis-promoting genes. The addition of the antioxidant glutathione restored cell viability and returned protein expression levels to those found in control cells. Collectively, these data support the working hypothesis that the loss of GSTP1 elevates oxidative stress, which alters mitogen-activated protein (MAP) kinases and NF-&kappa;B signaling, and induces apoptosis. In support of these in vitro data, nude mice bearing orthotopically implanted GSTP1-knockdown PDAC cells showed an impressive reduction in the size and weight of tumors compared to the controls. Additionally, we observed reduced levels of Ki-67 and increased expression of cleaved caspase-3 in GSTP1-knockdown tumors, suggesting GSTP1 knockdown impedes proliferation and upregulates apoptosis in PDAC cells. Together, these results indicate that GSTP1 plays a significant role in PDAC cell growth and provides support for the pursuit of GSTP1 inhibitors as therapeutic agents for PDAC

Glutathione S-Transferase pi-1 Knockdown Reduces Pancreatic Ductal Adenocarcinoma Growth by Activating Oxidative Stress Response Pathways

Glutathione S-transferase pi-1 (GSTP1) plays an important role in regulating oxidative stress by conjugating glutathione to electrophiles. GSTP1 is overexpressed in breast, colon, lung, and prostate tumors, where it contributes to tumor progression and drug resistance; however, the role of GSTP1 in pancreatic ductal adenocarcinoma (PDAC) is not well understood. Using shRNA, we knocked down GSTP1 expression in three different PDAC cell lines and determined the effect on cell proliferation, cell cycle progression, and reactive oxygen species (ROS) levels. Our results show GSTP1 knockdown reduces PDAC cell growth, prolongs the G0/G1 phase, and elevates ROS in PDAC cells. Furthermore, GSTP1 knockdown results in the increased phosphorylation of c-Jun N-terminal kinase (JNK) and c-Jun and the decreased phosphorylation of extracellular signal-regulated kinase (ERK), p65, the reduced expression of specificity protein 1 (Sp1), and the increased expression of apoptosis-promoting genes. The addition of the antioxidant glutathione restored cell viability and returned protein expression levels to those found in control cells. Collectively, these data support the working hypothesis that the loss of GSTP1 elevates oxidative stress, which alters mitogen-activated protein (MAP) kinases and NF-κB signaling, and induces apoptosis. In support of these in vitro data, nude mice bearing orthotopically implanted GSTP1-knockdown PDAC cells showed an impressive reduction in the size and weight of tumors compared to the controls. Additionally, we observed reduced levels of Ki-67 and increased expression of cleaved caspase-3 in GSTP1-knockdown tumors, suggesting GSTP1 knockdown impedes proliferation and upregulates apoptosis in PDAC cells. Together, these results indicate that GSTP1 plays a significant role in PDAC cell growth and provides support for the pursuit of GSTP1 inhibitors as therapeutic agents for PDAC.</jats:p