A novel 'social contract' - An attempt to harmonize a sponsor's exploratory research with a clinical study participant's data rights.

BACKGROUND: Pharmaceutical drug development rarely addresses a study participant's control of their genomic data, how to return individual incidental findings, and how to make use of genomic data more efficiently for exploratory research purposes. Mutually beneficial solutions to these issues are needed, as whole genome sequencing (WGS) is increasingly adopted in human research and as access to such information could provide impactful health-related information for a participant. METHODS: In this paper, we offer a novel framework to align a trial sponsor's need for broad exploratory research of the human genome with the study participant's right to data access and access control. The Exploratory Genetic Research Project (EGRP) aims to gather WGS on all participants of a sponsor's clinical trials. It is set up as a separate umbrella protocol to facilitate the consenting process, as well as the delineation between clinical trial endpoints versus exploratory future research. CONCLUSION: This concept establishes a participant's autonomy regarding access to genomic data and the disclosure of actionable incidental findings. The feasibility of EGRP will be tested and reassessed as it is deployed over the next few years.

Lipid Conjugates Enhance Endosomal Release of Antisense Oligonucleotides Into Cells.

Antisense oligonucleotides modified with phosphorothioate linkages (PS-ASOs) can enter cells via endocytic pathways and must escape from membraned organelles to reach target RNAs. We recently found that membrane destabilization induced by different lipid species contributes to PS-ASO release from late endosomes (LEs). In this study, we characterized intracellular uptake, trafficking, and activities of PS-ASOs conjugated with different lipid species. We found that palmitic acid-, tocopherol-, and cholesterol-conjugated PS-ASOs have increased protein binding and enhanced intracellular uptake compared to unconjugated PS-ASOs. Similar to the parental PS-ASO, the lipid-conjugated PS-ASOs traffic from early to LEs without incorporation into lipid droplets. Unlike parental PS-ASOs, the lipid-conjugated PS-ASOs tend to remain associated with plasma or endosomal membranes, and this appears to influence their release from endosomes. The lipid-conjugated PS-ASOs were released more rapidly than parental PS-ASO. These results suggest that lipid conjugation enhances the interactions of PS-ASOs with proteins or membranes, in turn facilitating intracellular trafficking and endosomal release.

Membrane Destabilization Induced by Lipid Species Increases Activity of Phosphorothioate-Antisense Oligonucleotides.

Chemically modified antisense oligonucleotides with phosphorothioate linkages (PS-ASOs) mediate site-specific cleavage of RNA by RNase H1 and are broadly used as research and therapeutic tools. PS-ASOs can enter cells via endocytic pathways and escape from membrane-enclosed endocytic organelles to reach target RNAs. We recently found that lysobisphosphatidic acid is required for release of PS-ASOs from late endosomes. Here, we evaluated the effects of other lipids on PS-ASO intracellular trafficking and activities. We show that free fatty acids, ceramide, and cholesterol increase PS-ASO activities. Free fatty acids induced formation of lipid droplets without changing the intracellular localization of PS-ASOs in early or late endosomes. Ceramide and cholesterol did not obviously induce the formation of lipid droplets, but cholesterol caused enlargement of endosome size and volume. Although none of those lipids appeared to influence PS-ASO internalization or intracellular trafficking processes, all led to an increase in leakiness of late endosomes. Thus, the membrane destabilization induced by these lipids likely contributes to PS-ASO release from late endosomes, which, in turn, increases PS-ASO activity.

COPII vesicles can affect the activity of antisense oligonucleotides by facilitating the release of oligonucleotides from endocytic pathways.

RNase H1-dependent, phosphorothioate-modified antisense oligonucleotides (PS-ASOs) can enter cells through endocytic pathways and need to be released from the membrane-enclosed organelles, a limiting step for antisense activity. Accumulating evidence has suggested that productive PS-ASO release mainly occurs from late endosomes (LEs). However, how PS-ASOs escape from LEs is not well understood. Here, we report that upon PS-ASO incubation, COPII vesicles, normally involved in ER-Golgi transport, can re-locate to PS-ASO-containing LEs. Reduction of COPII coat proteins significantly decreased PS-ASO activity, without affecting the levels of PS-ASO uptake and early-to-late endosome transport, but caused slower PS-ASO release from LEs. COPII co-localization with PS-ASOs at LEs does not require de novo assembly of COPII at ER. Interestingly, reduction of STX5 and P115, proteins involved in tethering and fusion of COPII vesicles with Golgi membranes, impaired COPII re-localization to LEs and decreased PS-ASO activity. STX5 can re-locate to LEs upon PS-ASO incubation, can bind PS-ASOs, and the binding appears to be required for this pathway. Our study reveals a novel release pathway in which PS-ASO incubation causes LE re-localization of STX5, which mediates the recruitment of COPII vesicles to LEs to facilitate endosomal PS-ASO release, and identifies another key PS-ASO binding protein.

Cellular uptake mediated by epidermal growth factor receptor facilitates the intracellular activity of phosphorothioate-modified antisense oligonucleotides.

Chemically modified antisense oligonucleotides (ASOs) with phosphorothioate (PS) linkages have been extensively studied as research and therapeutic agents. PS-ASOs can enter the cell and trigger cleavage of complementary RNA by RNase H1 even in the absence of transfection reagent. A number of cell surface proteins have been identified that bind PS-ASOs and mediate their cellular uptake; however, the mechanisms that lead to productive internalization of PS-ASOs are not well understood. Here, we characterized the interaction between PS-ASOs and epidermal growth factor receptor (EGFR). We found that PS-ASOs trafficked together with EGF and EGFR into clathrin-coated pit structures. Their co-localization was also observed at early endosomes and inside enlarged late endosomes. Reduction of EGFR decreased PS-ASO activity without affecting EGF-mediated signaling pathways and overexpression of EGFR increased PS-ASO activity in cells. Furthermore, reduction of EGFR delays PS-ASO trafficking from early to late endosomes. Thus, EGFR binds to PS-ASOs at the cell surface and mediates essential steps for active (productive) cellular uptake of PS-ASOs through its cargo-dependent trafficking processes which migrate PS-ASOs from early to late endosomes. This EGFR-mediated process can also serve as an additional model to better understand the mechanism of intracellular uptake and endosomal release of PS-ASOs.

Associations between food insecurity and the severity of psychological distress among African-Americans.

BACKGROUND: Little research exists on the association between food insecurity and mild to moderate psychological distress (MPD) among Black/African-Americans. In this study, we assess the relationship between food insecurity with and without hunger to that of both MPD and serious psychological distress (SPD) among this population. METHODS: 2009 and 2011/2012 adult public-use data from African-American respondents of the California Health Interview Survey were utilized for this study (n = 4003). Descriptive statistics were utilized to identify prevalence of psychological distress among sociodemographic and mental-health associated variables. Bivariate analyses were conducted between these variables and psychological distress using survey-weighted chi-square analyses. To evaluate the association between psychological distress, our primary exposure variable of food security, and other variables, we utilized survey-weighted multinomial logistic regression. RESULTS: Prevalence of mild to MPD was higher among those reporting food insecurity while SPD was highest for those with food insecurity and hunger. Results of multinomial logistic regression analysis demonstrate that while MPD was significantly associated with food insecurity, Black/African-Americans with food insecurity and hunger displayed over sixfold odds of higher serious psychological distress, as compared to those living at or above 200% federal poverty level. CONCLUSION: Our findings add to this growing segment of the literature on psychological distress and food insecurity. Further focus should be placed on improving the efficacy and reach of both formal and informal food support networks to improve the collective health and well-being of poor Black/African-American communities.

Food insecurity and low self-efficacy are associated with increased healthcare utilization among adults with type II diabetes mellitus.

OBJECTIVE: Food insecurity has been shown to negatively impact health outcomes, disease management and hospitalizations. Despite the increasing burden of type II diabetes mellitus (T2DM) in the United States, little research exists on the role of food insecurity and its association to T2DM-related healthcare utilization. The purpose of our study was to address such a gap in the literature by evaluating the role of food insecurity and T2DM-related past 12-month hospitalization or emergency department (ED) admission among adults with healthcare professional diagnosed T2DM. METHODS: We utilized the California Health Interview Survey (CHIS), 2009, 2011/2012 data to select CHIS participants who were aged 18 or older and reported doctor diagnosis of T2DM; resulting in a total of 8252 participants. Food insecurity was defined as: living at or above 200% federal poverty level (FPL), living below 200% FPL but food secure, living below 200% FPL and food insecure. A secondary exposure variable of interest was self-efficacy, based on the CHIS-provided variable of confidence to manage T2DM. All analyses were survey weighted with alpha less than .05 noting significance. RESULTS: Those with low food security (12.96%) and low-self efficacy (15.14%), reported significantly higher prevalence of T2DM-related healthcare utilization, as compared to their counterparts. Both living with low food security and having low self-efficacy were also associated with over two-fold increase in healthcare utilization. CONCLUSION: Our results demonstrate the cumulative need for community-based resources to improve the continuum of care and ensure that such at-risk populations have adequate resources for disease management.