Ferritin nanovehicle for targeted delivery of cytochrome C to cancer cells

In this work, we have exploited the unique properties of a chimeric archaeal-human ferritin to encapsulate, deliver and release cytochrome c and induce apoptosis in a myeloid leukemia cell line. The chimeric protein combines the versatility in 24-meric assembly and cargo incorporation capability of Archaeglobus fulgidus ferritin with specific binding of human H ferritin to CD71, the “heavy duty” carrier responsible for transferrin-iron uptake. Delivery of ferritin-encapsulated cytochrome C to the Acute Promyelocytic Leukemia (APL) NB4 cell line, highly resistant to transfection by conventional methods, was successfully achieved in vitro. The effective liberation of cytochrome C within the cytosolic environment, demonstrated by double fluorescent labelling, induced apoptosis in the cancer cells

SEMA6C: a novel adhesion-independent FAK and YAP activator, required for cancer cell viability and growth

Transmembrane semaphorins are signaling molecules, controlling axonal wiring and embryo development, which are increasingly implicated in human diseases. Semaphorin 6C (Sema6C) is a poorly understood family member and its functional role is still unclear. Upon targeting Sema6C expression in a range of cancer cells, we observed dramatic growth suppression, decreased ERK phosphorylation, upregulation of cell cycle inhibitor proteins p21, p27 and p53, and the onset of cell senescence, associated with activation of autophagy. These data are consistent with a fundamental requirement for Sema6C to support viability and growth in cancer cells. Mechanistically, we unveiled a novel signaling pathway elicited by Sema6C, and dependent on its intracellular domain, mediated by tyrosine kinases c-Abl and Focal Adhesion Kinase (FAK). Sema6C was found in complex with c-Abl, and induced its phosphorylation, which in turn led to FAK activation, independent of cell–matrix adhesion. Sema6C-induced FAK activity was furthermore responsible for increased nuclear localization of YAP transcriptional regulator. Moreover, Sema6C conferred YAP signaling-dependent long-term cancer cell survival upon nutrient deprivation. In conclusion, our findings demonstrate that Sema6C elicits a cancer promoting-signaling pathway sustaining cell viability and self-renewal, independent of growth factors and nutrients availability

Self-assembling ferritin-dendrimer nanoparticles for targeted delivery of nucleic acids to myeloid leukemia cells

Background: In recent years, the use of ferritins as nano-vehicles for drug delivery is taking center stage. Compared to other similar nanocarriers, Archaeoglobus fulgidus ferritin is particularly interesting due to its unique ability to assemble-disassemble under very mild conditions. Recently this ferritin was engineered to get a chimeric protein targeted to human CD71 receptor, typically overexpressed in cancer cells. Results: Archaeoglobus fulgidus chimeric ferritin was used to generate a self-assembling hybrid nanoparticle hosting an aminic dendrimer together with a small nucleic acid. The positively charged dendrimer can indeed establish electrostatic interactions with the chimeric ferritin internal surface, allowing the formation of a protein-dendrimer binary system. The 4 large triangular openings on the ferritin shell represent a gate for negatively charged small RNAs, which access the internal cavity attracted by the dense positive charge of the dendrimer. This ternary protein-dendrimer-RNA system is efficiently uptaken by acute myeloid leukemia cells, typically difficult to transfect. As a proof of concept, we used a microRNA whose cellular delivery and induced phenotypic effects can be easily detected. In this article we have demonstrated that this hybrid nanoparticle successfully delivers a pre-miRNA to leukemia cells. Once delivered, the nucleic acid is released into the cytosol and processed to mature miRNA, thus eliciting phenotypic effects and morphological changes similar to the initial stages of granulocyte differentiation. Conclusion: The results here presented pave the way for the design of a new family of protein-based transfecting agents that can specifically target a wide range of diseased cells. Graphic abstract: [Figure not available: see fulltext.]

The Safe Use of Pesticides: A Risk Assessment Procedure for the Enhancement of Occupational Health and Safety (OHS) Management

The attention paid to the use of pesticides has increased notably in recent years as demonstrated by the issue of laws and regulations requiring their safe and environmentally-conscious use (e.g. Directive 2009/128/EC and Regulation (EC) no. 1272/2008). Despite the benefits that can be achieved by pursuing the targets of stricter legislative framework, the difficulties for farmers in complying with it are remarkable, especially for small-sized companies. In fact, in contrast to other occupational health and safety (OHS) contexts, in the case of pesticides even a preliminary analysis on the relationship between pesticide use and the consequent exposure risks for the workers is a complex task. In order to reduce the above-mentioned gap, the present study is focused on the development of an easy-to-use tool for carrying out occupational risk assessment of agricultural activities related to the use of pesticides. The procedure was developed by starting from the Agricultural Health Study (AHS) approach and its improvements, and continuing to the thorough development of a tool for preliminary risk assessment, providing a simplified model for its practical application by farmers. A case study concerning olive cultivation was used for its first verification. The results achieved should be considered as an initial step for the promotion of safer practices when using pesticides, providing a consistent base for their further validation

Retinoic acid synergizes with the unfolded protein response and oxidative stress to induce cell death in FLT3-ITD+ AML.

Acute myeloid leukemia (AML) is often characterized by the expression of fusion or mutant proteins that cause impaired differentiation and enhanced proliferation and survival. The presence of mutant proteins prone to misfolding can render the cells sensitive to endoplasmic reticulum (ER) stress and oxidative stress that could otherwise be overcome. Here, we show that the triple combination of the differentiating agent retinoic acid (RA), the ER stress-inducing drug tunicamycin (Tm), and arsenic trioxide (ATO), able to generate oxidative stress, leads to the death of AML cell lines expressing fusion proteins involving the gene MLL and the internal tandem duplication (ITD) in the FLT3 tyrosine kinase receptor. Importantly, the combination of RA, Tm, and ATO decreased the colony-forming capacity of primary leukemic blasts bearing the FLT-ITD mutation without affecting healthy hematopoietic progenitor cells. We demonstrate in cell lines that combination of these drugs generates ER and oxidative stresses and impairs maturation and causes accumulation of FLT3 protein in the ER. Our data provide a proof of concept that low amounts of drugs that generate ER and oxidative stresses combined with RA could be an effective targeted therapy to hit AML cells characterized by MLL fusion proteins and FLT3-ITD mutation

The Small Satellite-Based, Imaging X-Ray Polarimeter Explorer (IXPE) Mission

The Imaging X-ray Polarimeter Explorer (IXPE) focuses on high energy astrophysics in the 2—8 keV x-ray band. IXPE is designed to explore general relativistic and quantum physics effects of gravity, energy, electric and magnetic fields at extreme limits. IXPE, a NASA Small Explorer (SMEX) Mission, will add new dimensions to on-orbit x-ray science: polarization degree, polarization angle and extended object polarization imaging. Polarization uniquely probes physical anisotropies that are not otherwise measurable—ordered magnetic fields, aspheric matter distributions, or general relativistic coupling to black-hole spin. Detailed imaging enables the specific properties of extended x-ray sources to be differentiated. The IXPE Observatory consists of spacecraft and payload modules built up in parallel to form the Observatory during system integration and test. The payload includes three polarization-sensitive, x-ray detector arrays paired with three x-ray mirror module assemblies (MMA). A deployable boom provides the correct separation (focal length) between the detector units and MMAs. Currently, the boom has been delivered, all four detectors units (DU) are complete, the detectors service unit (DSU) is complete, instrument system testing has been completed (DSU with 3 DUs), three of four MMAs is built and all spacecraft components except the solar array have been delivered along with the spacecraft and payload structures. Payload and spacecraft integration and test (I&T) started in March 2020. This paper overviews the flight segment (the Observatory, payload, and spacecraft implementation concepts) with emphasis on the build status and summarizes the launch segment. Launch is planned to occur on a Falcon 9 launch vehicle during Summer 2021. The paper summarizes the impacts of switching from the ‘design-to baseline’ of Pegasus XL to the selected launch vehicle for flight, Falcon 9. COVID-19 impacts to the Project are also summarized. The paper will close with a summary of the mission development status. The Project is firmly into the build phase for both the spacecraft and payload and rapidly approaching Observatory I&T

Liquid Flyback Booster Pre-Phase A Study Assessment

Mw concept of a flyback booster has been around since early in the Shuttle program. The original two-stage Shuttle concepts used a manned flyback booster. These boosters were eliminated from the program for funding and size reasons. The current Shuttle uses two Redesigned Solid Rocket Motors (RSRMs), which are recovered and refurbished after each flight; this is one of the major cost factors of the program. Replacement options have been studied over the past ten years. The conclusion reached by the most recent study is that the liquid flyback booster (LFBB) is the only competitive option from a life-cycle cost perspective. The purpose of this study was to assess the feasibility and practicality of LFBBs. The study provides an expansion of the recommendations made during the during the aforementioned study. The primary benefits are the potential for enhanced reusability and a reuction of recurring costs. The potential savings in vehicle turnaround could offset the up-front costs. Development of LFBBs requires a commitment to the Shuttle program for 20 to 30 years. LFBBs also offer enhanced safety and abort capabilities. Currently, any failure of an RSRM can be considered catastrophic since them we no intact abort capabilities during the burn of the RSRMS. The performance goal of the LFBBs was to lift a fully loaded Orbiter under optimal conditions, so as not to be the limiting factor of the performance capability of the Shuttle. In addition, a final benefit is the availability of growth paths for applications other than the Shuttle

DNAJA1 controls the fate of misfolded mutant p53 through the mevalonate pathway

Stabilization of mutant p53 (mutp53) in tumours greatly contributes to malignant progression. However, little is known about the underlying mechanisms and therapeutic approaches to destabilize mutp53. Here, through high-throughput screening we identify statins, cholesterol-lowering drugs, as degradation inducers for conformational or misfolded p53 mutants with minimal effects on wild-type p53 (wtp53) and DNA contact mutants. Statins preferentially suppress mutp53-expressing cancer cell growth. Specific reduction of mevalonate-5-phosphate by statins or mevalonate kinase knockdown induces CHIP ubiquitin ligase-mediated nuclear export, ubiquitylation, and degradation of mutp53 by impairing interaction of mutp53 with DNAJA1, a Hsp40 family member. Knockdown of DNAJA1 also induces CHIP-mediated mutp53 degradation, while its overexpression antagonizes statin-induced mutp53 degradation. Our study reveals that DNAJA1 controls the fate of misfolded mutp53, provides insights into potential strategies to deplete mutp53 through the mevalonate pathway–DNAJA1 axis, and highlights the significance of p53 status in impacting statins’ efficacy on cancer therapy

Genetic Dissection of Epidermal Growth Factor Receptor Signaling during Luteinizing Hormone-Induced Oocyte Maturation

Recent evidence that luteinizing hormone (LH) stimulation of ovulatory follicles causes transactivation of the epidermal growth factor receptor (EGFR) has provided insights into the mechanisms of ovulation. However, the complete array of signals that promote oocyte reentry into the meiotic cell cycle in the follicle are still incompletely understood. To elucidate the signaling downstream of EGFR involved in oocyte maturation, we have investigated the LH responses in granulosa cells with targeted ablation of EGFR. Oocyte maturation and ovulation is disrupted when EGFR expression is progressively reduced. In granulosa cells from mice with either global or granulosa cell-specific disruption of EGFR signaling, LH-induced phosphorylation of MAPK3/1, p38MAPK, and connexin-43 is impaired. Although the LH-induced decrease in cGMP is EGFR-dependent in wild type follicles, LH still induces a decrease in cGMP in Egfrdelta/f Cyp19-Cre follicles. Thus compensatory mechanisms appear activated in the mutant. Spatial propagation of the LH signal in the follicle also is dependent on the EGF network, and likely is important for the control of signaling to the oocyte. Thus, multiple signals and redundant pathways contribute to regulating oocyte reentry into the cell cycle

Targeted next generation sequencing approach identifies eighteen new candidate genes in normosmic hypogonadotropic hypogonadism and Kallmann Syndrome

The genetic basis is unknown for ∼60% of normosmic hypogonadotropic hypogonadism (nHH)/Kallmann syndrome (KS). DNAs from (17 male and 31 female) nHH/KS patients were analyzed by targeted next generation sequencing (NGS) of 261 genes involved in hypothalamic, pituitary, and/or olfactory pathways, or suggested by chromosome rearrangements. Selected variants were subjected to Sanger DNA sequencing, the gold standard. The frequency of Sanger-confirmed variants was determined using the ExAC database. Variants were classified as likely pathogenic (frameshift, nonsense, and splice site) or predicted pathogenic (nonsynonymous missense). Two novel FGFR1 mutations were identified, as were 18 new candidate genes including: AMN1, CCKBR, CRY1, CXCR4, FGF13, GAP43, GLI3, JAG1, NOS1, MASTL, NOTCH1, NRP2, PALM2, PDE3A, PLEKHA5, RD3, and TRAPPC9, and TSPAN11. Digenic and trigenic variants were found in 8/48 (16.7%) and 1/48 (2.1%) patients, respectively. NGS with confirmation by Sanger sequencing resulted in the identification of new causative FGFR1 gene mutations and suggested 18 new candidate genes in nHH/KS