Plasma Membrane Lipid Therapy: Disruption of Oncogenic Ras Driven Phenotypes by Membrane Targeted Dietary Bioactives (Mtdb)

Approximately 30-50% of colorectal cancers contain KRas mutations, which confer resistance to standard therapy. A diet high in long-chain n-3 polyunsaturated fatty acids (n-3 PUFA) is generally considered chemoprotective in regard to colon cancer. However, the molecular mechanisms by which n-3 PUFA suppresses tumorigenesis remain to be determined. Therefore, our overall goal is to define the role of n-3 PUFA in the modulation of oncogenic KRas-driven colon cancer. We hypothesized that n-3 PUFA would modify the biophysical properties of the plasma membrane though its incorporation into plasma membrane phospholipids. Utilizing quantitative fluorescence microscopy, we determined that n-3 PUFA reduce the rigidity of the plasma membrane of young adult mouse colonocytes (YAMC), CD3/CD28 activated CD4⁺ T cells, and colonic crypts derived from transgenic Fat-1 mice, which synthesize n-3 PUFA de novo. Interestingly, n-3 PUFA increased the rigidity of cytoskeletal free giant plasma membrane vesicles (GPMVs), derived from the aforementioned samples. Oncogenic Ras signaling is dependent on the formation of specific plasma membrane localized proteolipid complexes. Thus, it is noteworthy in YAMC cells and Drosophila midguts we documented for the first time that n-3 PUFA generates mix clusters of H- and K-Ras isoforms, which signal through ERK less efficiently. This resulted in a reduction of Ras driven colonic phenotypes in mice and Drosophila models. We then assessed the ability of n-3 PUFA to disrupt macropinocytosis, a cellular process in which extra cellular proteins are internalized through dynamic changes in plasma membrane lipids and cytoskeletal proteins. This process provides energy substrates that support the unique metabolic needs of Ras expressing cells. We determined that Ras expression mediates an increase in plasma membrane free cholesterol, which rigidifies the plasma membrane. Attenuating this rigidification by incorporating n-3 PUFA into the plasma membrane disrupted Ras driven macropinocytosis. Overall, we have demonstrated that n-3 PUFA ameliorate oncogenic Ras driven phenotypes through modification of plasma membrane biophysical properties leading to a disruption of Ras signaling. This reduces the ability of tumor cells to acquire energy substrate necessary to maintain its unique metabolic needs. These data contribute to the mechanistic understanding of how n-3 PUFA protect against colon cancer

Long-Chain n-3 Fatty Acids Attenuate Oncogenic KRas-Driven Proliferation by Altering Plasma Membrane Nanoscale Proteolipid Composition

Ras signaling originates from transient nanoscale compartmentalized regions of the plasma membrane composed of specific proteins and lipids. The highly specific lipid composition of these nanodomains, termed nanoclusters, facilitates effector recruitment and therefore influences signal transduction. This suggests that Ras nanocluster proteolipid composition could represent a novel target for future chemoprevention interventions. There is evidence that consumption of fish oil containing long-chain n-3 polyunsaturated fatty acids (n-3 PUFA) such as eicosapentaenoic acid (EPA, 20:5Δ5,8,11,14,17) and docosahexaenoic acid (DHA, 22:6Δ4,7,10,13,16,19) may reduce colon cancer risk in humans, yet the mechanism underlying this effect is unknown. Here, we demonstrate that dietary n-3 PUFA reduce the lateral segregation of cholesterol-dependent and -independent nanoclusters, suppressing phosphatidic acid-dependent oncogenic KRas effector interactions, via their physical incorporation into plasma membrane phospholipids. This results in attenuation of oncogenic Ras-driven colonic hyperproliferation in both Drosophila and murine models. These findings demonstrate the unique properties of dietary n-3 PUFA in the shaping of Ras nanoscale proteolipid complexes and support the emerging role of plasma membrane-targeted therapies. Significance: The influence of dietary long chain n-3 polyunsaturated fatty acids on plasma membrane protein nanoscale organization and KRas signaling supports development of plasma membrane-targeted therapies in colon cancer

Physical exercise, fitness and dietary pattern and their relationship with circadian blood pressure pattern, augmentation index and endothelial dysfunction biological markers: EVIDENT study protocol

Background: Healthy lifestyles may help to delay arterial aging. The purpose of this study is to analyze the relationship of physical activity and dietary pattern to the circadian pattern of blood pressure, central and peripheral blood pressure, pulse wave velocity, carotid intima-media thickness and biological markers of endothelial dysfunction in active and sedentary individuals without arteriosclerotic disease. Methods/Design Design: A cross-sectional multicenter study with six research groups. Subjects: From subjects of the PEPAF project cohort, in which 1,163 who were sedentary became active, 1,942 were sedentary and 2,346 were active. By stratified random sampling, 1,500 subjects will be included, 250 in each group. Primary measurements: We will evaluate height, weight, abdominal circumference, clinical and ambulatory blood pressure with the Radial Pulse Wave Acquisition Device (BPro), central blood pressure and augmentation index with Pulse Wave Application Software (A-Pulse) and SphymgoCor System Px (Pulse Wave Analysis), pulse wave velocity (PWV) with SphymgoCor System Px (Pulse Wave Velocity), nutritional pattern with a food intake frequency questionnaire, physical activity with the 7-day PAR questionnaire and accelerometer (Actigraph GT3X), physical fitness with the cycle ergometer (PWC-170), carotid intima-media thickness by ultrasound (Micromax), and endothelial dysfunction biological markers (endoglin and osteoprotegerin). Discussion: Determining that sustained physical activity and the change from sedentary to active as well as a healthy diet improve circadian pattern, arterial elasticity and carotid intima-media thickness may help to propose lifestyle intervention programs. These interventions could improve the cardiovascular risk profile in some parameters not routinely assessed with traditional risk scales. From the results of this study, interventional approaches could be obtained to delay vascular aging that combine physical exercise and diet