Impact of optimised quasi-block structures on the properties of polymer electrolytes.

A set of ionic quasi-block copolymers were investigated to determine the effects of their composition and structure on their performance in their application as solid-state battery electrolytes. Diffusion and electrochemical tests have shown that these new quasi-block electrolytes have comparable performance to traditional block copolymers reaching ionic conductivities of 3.8 × 10-4 S cm-1 and lithium-ion diffusion of 4.6 × 10-12 m2 s-1 at 80 °C. It was illustrated that the mechanical properties of each quasi-block electrolyte are highly dependent on the order of monomer addition in polymer synthesis while the phase morphology hints at each of the quasi-blocks' unique compositional make up.

Peptide Sequence Variations Govern Hydrogel Stiffness: Insights from a Multi-Scale Structural Analysis of H-FQFQFK-NH2 Peptide Derivatives.

Throughout the past decades, amphipathic peptide-based hydrogels have proven to be promising materials for biomedical applications. Amphipathic peptides are known to adopt ß-sheet configurations that self-assemble into fibers that then interact to form a hydrogel network. A fundamental understanding of how the peptide sequence alters the structural properties of the hydrogels would allow for a more rational design of novel peptides for a variety of biomedical applications in the future. Therefore, the current work investigates how changing the type of amino acid, the amphipathic pattern, and the peptide length affects the secondary structure, fiber characteristics, and stiffness of peptide-based hydrogels. Hereto, seven amphipathic peptides of different sequence and length, four of which have not been previously reported, based on and including the hexapeptide H-Phe-Gln-Phe-Gln-Phe-Lys-NH2, are synthesized and thoroughly characterized by circular dichroism (CD), Fourier Transform Infrared (FTIR) spectroscopy, Wide Angle X-ray Scattering (WAXS), Small Angle X-ray Scattering (SAXS), Transmission Electron Microscopy (TEM), and Thioflavin T (ThT) fibrillization assays. The results show that a high amount of regularly spaced ß-sheets, a high amount of fibers, and fiber bundling contribute to the stiffness of the hydrogel. Furthermore, a study of the time-dependent fibril formation process reveals complex transient dynamics. The peptide strands structure through an intermediate helical state prior to ß-sheet formation, which is found to be concentration- and time-dependent.

A systematic study of the effect of lipid architecture on cytotoxicity and cellular uptake of cationic cubosomes.

HYPOTHESIS: Lipid nanoparticles containing a cationic lipid are increasingly used in drug and gene delivery as they can display improved cellular uptake, enhanced loading for anionic cargo such as siRNA and mRNA or exhibit additional functionality such as cytotoxicity against cancer cells. This research study tests the hypothesis that the molecular structure of the cationic lipid influences the structure of the lipid nanoparticle, the cellular uptake, and the resultant cytotoxicity. EXPERIMENTS: Three potentially cytotoxic cationic lipids, with systematic variations to the hydrophobic moiety, were designed and synthesised. All the three cationic lipids synthesised contain pharmacophores such as the bicyclic coumarin group (CCA12), the tricyclic etodolac moiety (ETD12), or the large pentacyclic triterpenoid "ursolic" group (U12) conjugated to a quaternary ammonium cationic lipid containing twin C12 chains. The cationic lipids were doped into monoolein cubosomes at a range of concentrations from 0.1 mol% to 5 mol% and the effect of the lipid molecular architecture on the cubosome phase behaviour was assessed using a combination of Small Angle X-Ray Scattering (SAXS), Dynamic Light Scattering (DLS), zeta-potential and cryo-Transmission Electron Microscopy (Cryo-TEM). The resulting cytotoxicity of these particles against a range of cancerous and non-cancerous cell-lines was assessed, along with their cellular uptake. FINDINGS: The molecular architecture of the cationic lipid was linked to the internal nanostructure of the resulting cationic cubosomes with a transition to more curved cubic and hexagonal phases generally observed. Cubosomes formed from the cationic lipid CCA12 were found to have improved cellular uptake and significantly higher cytotoxicity than the cationic lipids ETD12 and U12 against the gastric cancer cell-line (AGS) at lipid concentrations ≥ 75 µg/mL. CCA12 cationic cubosomes also displayed reasonable cytotoxicity against the prostate cancer PC-3 cell-line at lipid concentrations ≥ 100 µg/mL. In contrast, 2.5 mol% ETD12 and 2.5 mol% U12 cubosomes were generally non-toxic against both cancerous and non-cancerous cell lines over the entire concentration range tested. The molecular architecture of the cationic lipid was found to influence the cubosome phase behaviour, the cellular uptake and the toxicity although further studies are necessary to determine the exact relationship between structure and cellular uptake across a range of cell lines.

Polysaccharide-Targeting Lipid Nanoparticles to Kill Gram-Negative Bacteria.

The rapid increase and spread of Gram-negative bacteria resistant to many or all existing treatments threaten a return to the preantibiotic era. The presence of bacterial polysaccharides that impede the penetration of many antimicrobials and protect them from the innate immune system contributes to resistance and pathogenicity. No currently approved antibiotics target the polysaccharide regions of microbes. Here, describe monolaurin-based niosomes, the first lipid nanoparticles that can eliminate bacterial polysaccharides from hypervirulent Klebsiella pneumoniae, are described. Their combination with polymyxin B shows no cytotoxicity in vitro and is highly effective in combating K. pneumoniae infection in vivo. Comprehensive mechanistic studies have revealed that antimicrobial activity proceeds via a multimodal mechanism. Initially, lipid nanoparticles disrupt polysaccharides, then outer and inner membranes are destabilized and destroyed by polymyxin B, resulting in synergistic cell lysis. This novel lipidic nanoparticle system shows tremendous promise as a highly effective antimicrobial treatment targeting multidrug-resistant Gram-negative pathogens.

Inverse Cubic and Hexagonal Mesophase Evolution within Ionizable Lipid Nanoparticles Correlates with mRNA Transfection in Macrophages.

mRNA lipid nanoparticle (LNP) technology presents enormous opportunities to prevent and treat various diseases. Here, we developed a novel series of LNPs containing ionizable amino-lipids showing a remarkable array of tunable and pH-sensitive lyotropic liquid crystalline mesophases including the inverse bicontinuous cubic and hexagonal phases characterized by high-throughput synchrotron radiation X-ray scattering. Furthermore, with an interest in developing mRNA therapeutics for lung macrophage targeting, we discovered that there is a strong correlation between the mesophase transition of the LNPs during acidification and the macrophage association/transfection efficiency of mRNAs. The slight molecular structural differences between the SM-102 and ALC-0315 ionizable lipids are linked to the LNP's ability to transform their internal structures from an amorphous state to the inverse micellar, hexagonal, and finally cubic structures during endosomal maturation. SM-102 LNPs showed exceptionally improved transfection efficiency due to their ability to form a cubic structure at a lower pH than the ALC-0315 analogues, which remained within the hexagonal structure, previously attributed to promoting endosomal escape of the ionizable LNPs. Overall, the new knowledge draws our attention to the important role of mesophase transition in endosomal escape, and the novel LNP libraries reported herein have broad prospects for advancing mRNA therapeutics.

Novel bioactive cationic cubosomes enhance the cytotoxic effect of paclitaxel against a paclitaxel resistant prostate cancer cell-line.

Hypothesis The study aimed to use molecular hybridization of a cationic lipid with a known pharmacophore to produce a bifunctional lipid having a cationic charge to enhance fusion with the cancer cell surface and biological activity via the pharmacophoric head group. Experiments The novel cationic lipid DMP12 [N-(2-(3-(3,4-dimethoxyphenyl) propanamido) ethyl)-N-dodecyl-N-methyldodecan-1-aminium iodide] was synthesised by conjugating 3-(3,4-dimethoxyphenyl) propanoic acid (or 3,4-dimethoxyhydrocinnamic acid) to twin 12 carbon chains bearing a quaternary ammonium group [N-(2-aminoethyl)-N-dodecyl-N-methyldodecan-1-aminium iodide]. The physicochemical and biological properties of DMP12 were investigated. Cubosome particles consisting of monoolein (MO) doped with DMP12 and paclitaxel were characterized using Small-angle X-ray Scattering (SAXS), Dynamic Light Scattering (DLS), and Cryo-Transmission Electron Microscopy (Cryo-TEM). Combination therapy using these cubosomes was assessed in vitro against the gastric (AGS) and prostate (DU-145 and PC-3) cancer cell lines using cytotoxicity assay. Findings Monoolein (MO) cubosomes doped with DMP12 were observed to be toxic against the AGS and DU-145 cell-lines at higher cubosome concentrations (≥100 µg/ml) but had limited activity against the PC-3 cell-line. However, combination therapy consisting of 5 mol% DMP12 and 0.5 mol% paclitaxel (PTX) significantly increased the cytotoxicity against the PC-3 cell-line which was resistant to either DMP12 or PTX individually. The results demonstrate that DMP12 has a prospective role as a bioactive excipient in cancer therapy.

Prospective Subunit Nanovaccine against Mycobacterium tuberculosis Infection─Cubosome Lipid Nanocarriers of Cord Factor, Trehalose 6,6' Dimycolate.

An improved vaccine is urgently needed to replace the now more than 100-year-old Bacillus Calmette-Guérin (BCG) vaccine against tuberculosis (TB) disease, which represents a significant burden on global public health. Mycolic acid, or cord factor trehalose 6,6' dimycolate (TDM), a lipid component abundant in the cell wall of the pathogen Mycobacterium tuberculosis (MTB), has been shown to have strong immunostimulatory activity but remains underexplored due to its high toxicity and poor solubility. Herein, we employed a novel strategy to encapsulate TDM within a cubosome lipid nanocarrier as a potential subunit nanovaccine candidate against TB. This strategy not only increased the solubility and reduced the toxicity of TDM but also elicited a protective immune response to control MTB growth in macrophages. Both pre-treatment and concurrent treatment of the TDM encapsulated in lipid monoolein (MO) cubosomes (MO-TDM) (1 mol %) induced a strong proinflammatory cytokine response in MTB-infected macrophages, due to epigenetic changes at the promoters of tumor necrosis factor alpha (TNF-α) and interleukin 6 (IL-6) in comparison to the untreated control. Furthermore, treatment with MO-TDM (1 mol %) cubosomes significantly improved antigen processing and presentation capabilities of MTB-infected macrophages to CD4 T cells. The ability of MO-TDM (1 mol %) cubosomes to induce a robust innate and adaptive response in vitro was further supported by a mathematical modeling study predicting the vaccine efficacy in vivo. Overall, these results indicate a strong immunostimulatory effect of TDM when delivered through the lipid nanocarrier, suggesting its potential as a novel TB vaccine.

High ploidy large cytoplasmic megakaryocytes are hematopoietic stem cells regulators and essential for platelet production.

Megakaryocytes (MK) generate platelets. Recently, we and others, have reported MK also regulate hematopoietic stem cells (HSC). Here we show high ploidy large cytoplasmic megakaryocytes (LCM) are critical negative regulators of HSC and critical for platelet formation. Using a mouse knockout model (Pf4-Srsf3Δ/Δ) with normal MK numbers, but essentially devoid of LCM, we demonstrate a pronounced increase in BM HSC concurrent with endogenous mobilization and extramedullary hematopoiesis. Severe thrombocytopenia is observed in animals with diminished LCM, although there is no change in MK ploidy distribution, uncoupling endoreduplication and platelet production. When HSC isolated from a microenvironment essentially devoid of LCM reconstitute hematopoiesis in lethally irradiated mice, the absence of LCM increases HSC in BM, blood and spleen, and the recapitulation of thrombocytopenia. In contrast, following a competitive transplant using minimal numbers of WT HSC together with HSC from a microenvironment with diminished LCM, sufficient WT HSC-generated LCM regulates a normal HSC pool and prevents thrombocytopenia. Importantly, LCM are conserved in humans.

Cubosome Lipid Nanocarriers As a Drug Delivery Vehicle for Intracellular Mycobacterium tuberculosis Infections.

Mycobacterium tuberculosis (MTB) causes the infectious disease tuberculosis (TB), responsible for more deaths than any other single infectious disease in history. Intracellular MTB are slow growing and difficult to target with traditional antitubercular drugs, leading to the emergence of multidrug resistance in TB infection, which is a major global public health issue. Recent advances in innovative lipid nanotechnologies for drug delivery have demonstrated promising outcomes for chronic infectious diseases but have not yet been tested as potential delivery systems for intracellular infections such as TB. The current study evaluates the potential of monoolein (MO)-based cationic cubosomes for the encapsulation and delivery of the first line antitubercular drug rifampicin (RIF) against an MTB-H37Ra in vitro culture model. In particular, we show that the use of cationic cubosomes as delivery vehicles reduced the minimum inhibitory concentration (MIC) of RIF by 2-fold against actively replicating MTB-H37Ra (compared to that of the free drug) and also shortened the lifecycle duration of axenic MTB-H37Ra from 5 to 3 days. The cubosome-mediated delivery was also found to be effective against intracellular MTB-H37Ra within THP-1 human macrophages, with a 2.8 log reduction in viability of the bacilli after 6 days incubation at the MIC. The killing time was also reduced from 8 to 6 days without distressing the host macrophages. Mechanistic studies on the uptake of RIF-loaded cationic cubosomes using total internal reflection fluorescence microscopy (TIRFM) demonstrated the capacity of these lipid particles to effectively target intracellular bacteria. Overall, these results demonstrate that cationic cubosomes are a potent delivery system for the antitubercular drug RIF for therapeutic management of TB.

The role of glycosylation in amyloid fibril formation of bovine κ-casein.

In order to explore the functions of glycosylation of κ-Casein (κ-CN) in bovine milk, unglycosylated (UG) and twice glycosylated (2G) forms of κ-CN B were purified by selective precipitation followed by anion exchange chromatography from κ-CN BB milk and tested for their amyloid fibril formation and morphology, oligomerisation states and protein structure. The diameter of self-assembled κ-CN B aggregates of both glyco-form were shown for the first time to be in the same 26.0-28.7 nm range for a 1 mg mL-1 solution. The presence of two bound glycans in the protein structure of 2G κ-CN B led to a greater increase in the maximum amyloid fibril formation rate with increasing protein concentration and a difference in both length (82.0 ± 29.9 vs 50.3 ± 13.7 nm) and width (8.6 ± 2.1 vs 13.9 ± 2.5 nm) for fibril morphology compared to UG κ-CN B. The present results suggest that amyloid fibril formation proceeds at a slow but steady rate via the self-assembly of dissociated, monomeric κ-CN B proteins at concentrations of 0.22-0.44 mg mL-1. However amyloid fibril formation proceeds more rapidly via the assembly of either aggregated κ-CN present in a micelle-like form or dissociated monomeric κ-CN, packed into reorganised formational structures above the critical micellar concentration to form fibrils of differing width. The degree of glycosylation has no effect on the polarity of the adjacent environment, nor non-covalent and disulphide interactions between protein molecules when in the native form. Yet glycosylation can influence protein folding patterns of κ-CN B leading to a reduced tryptophan intrinsic fluorescence intensity for 2G compared to UG κ-CN B. These results demonstrate that glycosylation plays an important role in the modulation of aggregation states of κ-CN and contributes to a better understanding of the role of glycosylation in the formation of amyloid fibrils from intrinsically disordered proteins.

A Multifunctional Porous Silicon Nanocarrier for Glioblastoma Treatment.

Clinical treatment of glioblastoma (GBM) remains a major challenge because of the blood-brain barrier, chemotherapeutic resistance, and aggressive tumor metastasis. The development of advanced nanoplatforms that can efficiently deliver drugs and gene therapies across the BBB to the brain tumors is urgently needed. The protein "downregulated in renal cell carcinoma" (DRR) is one of the key drivers of GBM invasion. Here, we engineered porous silicon nanoparticles (pSiNPs) with antisense oligonucleotide (AON) for DRR gene knockdown as a targeted gene and drug delivery platform for GBM treatment. These AON-modified pSiNPs (AON@pSiNPs) were selectively internalized by GBM and human cerebral microvascular endothelial cells (hCMEC/D3) cells expressing Class A scavenger receptors (SR-A). AON was released from AON@pSiNPs, knocked down DRR and inhibited GBM cell migration. Additionally, a penetration study in a microfluidic-based BBB model and a biodistribution study in a glioma mice model showed that AON@pSiNPs could specifically cross the BBB and enter the brain. We further demonstrated that AON@pSiNPs could carry a large payload of the chemotherapy drug temozolomide (TMZ, 1.3 mg of TMZ per mg of NPs) and induce a significant cytotoxicity in GBM cells. On the basis of these results, the nanocarrier and its multifunctional strategy provide a strong potential for clinical treatment of GBM and research for targeted drug and gene delivery.

SARS-CoV-2 Virus Detection Via a Polymeric Nanochannel-Based Electrochemical Biosensor.

The development of simple, cost-effective, rapid, and quantitative diagnostic tools remains critical to monitor infectious COVID-19 disease. Although numerous diagnostic platforms, including rapid antigen tests, are developed and used, they suffer from limited accuracy, especially when tested with asymptomatic patients. Here, a unique approach to fabricate a nanochannel-based electrochemical biosensor that can detect the entire virion instead of virus fragments, is demonstrated. The sensing platform has uniform nanoscale channels created by the convective assembly of polystyrene (PS) beads on gold electrodes. The PS beads are then functionalized with bioreceptors while the gold surface is endowed with anti-fouling properties. When added to the biosensor, SARS-CoV-2 virus particles block the nanochannels by specific binding to the bioreceptors. The nanochannel blockage hinders the diffusion of a redox probe; and thus, allows quantification of the viral load by measuring the changes in the oxidation current before and after virus incubation. The biosensor shows a low limit of detection of ≈1.0 viral particle mL-1 with a wide detection range up to 108 particles mL-1 in cell culture media. Moreover, the biosensor is able to differentiate saliva samples with SARS-CoV-2 from those without, demonstrating the potential of this technology for translation into a point-of-care biosensor product.

Impact of doubling peptide length on in vivo hydrogel stability and sustained drug release.

Peptide-based hydrogels represent promising systems for the sustained release of different types of drugs, ranging from small molecules to biologicals. Aiming at subcutaneous injection, which is a desirable parenteral administration route, especially for biologicals, we herein focus on physically crosslinked systems possessing thixotropic behaviour. The purpose of this study was to evaluate the in vitro and in vivo properties of hydrogels based on the amphipathic hexapeptide H-FQFQFK-NH2, which served as the lead sequence. Upon doubling the length of this peptide, the dodecapeptide H-FQFQFKFQFQFK-NH2 gave a significant improvement in terms of in vivo stability of the hydrogel post-injection, as monitored by nuclear SPECT/CT imaging. This increased hydrogel stability also led to a more prolonged in vivo release of encapsulated peptide cargoes. Even though no direct link with the mechanical properties of the hydrogels before injection could be made, an important effect of the subcutaneous medium was noticed on the rheological properties of the hydrogels in post in vivo injection measurements. The results were validated in vivo for a therapeutically relevant analgesic peptide using the hot-plate test as an acute pain model. It was confirmed that elongation of the hydrogelator sequence induced more extended antinociceptive effects. Altogether, this simple structural modification of the hydrogelating peptide could provide a basis for reaching longer durations of action upon use of these soft biomaterials.

Fabrication of a Reusable Carbon Dot/Gold Nanoparticle/Metal-Organic Framework Film for Fluorescence Detection of Lead Ions in Water.

Solid-state sensing platforms are desirable for the development of reusable sensors to promote public health measures such as testing for drinking water contamination. A bioinspired metal-organic framework (MOF)-based material has been developed by imitating metal-protein interactions in biological systems to attain high sensitivity and selectivity to Pb2+ through fluorescence sensing. A zirconium terephthalate-type framework (also known as NH2-UiO-66) was modified with both gold nanoparticles and thiol-functionalized carbon dots to give HS-C/Au(x)/UiO-66 composites with different Au content (x) and were subsequently adapted into films that show extraordinary sensitivity to Pb2+. The HS-C/Au(1.4)/UiO-66 film that consists of 1.4 wt % Au shows a quenching response with the limit of detection of 80 parts per trillion and sustained performance for five cycles. Moreover, the fluorescence response of the HS-C/Au(x)/UiO-66 film to Pb2+ can be reversed from emission quenching to enrichment of fluorescence by increasing the Au content. The performance of the HS-C/Au(x)/UiO-66 film as a solid-state sensor demonstrates its potential for application in reusable sensing devices to ensure public safety from Pb2+ contamination in drinking water.

Variation in Hydrogel Formation and Network Structure for Telo-, Atelo- and Methacrylated Collagens.

As the most abundant protein in the extracellular matrix, collagen has become widely studied in the fields of tissue engineering and regenerative medicine. Of the various collagen types, collagen type I is the most commonly utilised in laboratory studies. In tissues, collagen type I forms into fibrils that provide an extended fibrillar network. In tissue engineering and regenerative medicine, little emphasis has been placed on the nature of the network that is formed. Various factors could affect the network structure, including the method used to extract collagen from native tissue, since this may remove the telopeptides, and the nature and extent of any chemical modifications and crosslinking moieties. The structure of any fibril network affects cellular proliferation and differentiation, as well as the overall modulus of hydrogels. In this study, the network-forming properties of two distinct forms of collagen (telo- and atelo-collagen) and their methacrylated derivatives were compared. The presence of the telopeptides facilitated fibril formation in the unmodified samples, but this benefit was substantially reduced by subsequent methacrylation, leading to a loss in the native self-assembly potential. Furthermore, the impact of the methacrylation of the collagen, which enables rapid crosslinking and makes it suitable for use in 3D printing, was investigated. The crosslinking of the methacrylated samples (both telo- and atelo-) was seen to improve the fibril-like network compared to the non-crosslinked samples. This contrasted with the samples of methacrylated gelatin, which showed little, if any, fibrillar or ordered network structure, regardless of whether they were crosslinked.

Biomimetic metal-organic frameworks as protective scaffolds for live-virus encapsulation and vaccine stabilization.

The invaluable health, economic and social impacts of vaccination are hard to exaggerate. The ability to stabilize vaccines is urgently required for their equitable distribution without the dependence on the 'cold-chain' logistics. Herein, for the first time we report biomimetic-mineralization of live-viral vaccines using metal-organic frameworks (MOFs) to enhance their storage stability from days to months. Applying ZIF-8 and aluminium fumarate (Alfum), the Newcastle Disease Virus (NDV) V4 strain and Influenza A WSN strain were encapsulated with remarkable retention of their viral titre. The ZIF-8@NDV, ZIF-8@WSN and Alfum@WSN composites were validated for live-virus recovery using a tissue culture infectious dose (TCID50) assay. With the objective of long-term stabilization, we developed a novel, trehalose (T) and skim milk (SM) stabilized, freeze-dried MOF@Vaccine composite, ZIF-8@NDV+T/SM. The thermal stability of this composite was investigated and compared with the control NDV and non-encapsulated, freeze-dried NDV+T/SM composite at 4 °C, RT, and 37 °C over a period of 12 weeks. We demonstrate the fragility of the control NDV vaccine which lost all viability at RT and 37°C by 12 and 4 weeks, respectively. Comparing the freeze-dried counterparts, the MOF encapsulated ZIF-8@NDV+T/SM demonstrated significant enhancement in stability of the NDV+T/SM composite especially at RT and 37 °C upto 12 weeks. STATEMENT OF SIGNIFICANCE: Vaccination is undoubtedly one of the most effective medical interventions, saving millions of lives each year. However, the requirement of 'cold-chain' logistics is a major impediment to widespread immunization. Live viral vaccines (LVVs) are widely used vaccine types with proven efficacy and low cost. Nonetheless, their complex composition increases their susceptability to thermal stress. Several LVV thermostabilization approaches have been investigated, including their complex engineering and the facile addition of stabilizers. Still, the lack of a universal approach urgently requires finding a stabilization technique especially when additives alone may not be sufficient. Herein, we demonstrate MOF biomimetic-mineralization technology to encapsulate LVVs developing an optimised composite which significantly preserves vaccines without refrigeration for extended periods of time.

A polytherapy based approach to combat antimicrobial resistance using cubosomes.

A depleted antimicrobial drug pipeline combined with an increasing prevalence of Gram-negative 'superbugs' has increased interest in nano therapies to treat antibiotic resistance. As cubosomes and polymyxins disrupt the outer membrane of Gram-negative bacteria via different mechanisms, we herein examine the antimicrobial activity of polymyxin-loaded cubosomes and explore an alternative strategy via the polytherapy treatment of pathogens with cubosomes in combination with polymyxin. The polytherapy treatment substantially increases antimicrobial activity compared to polymyxin B-loaded cubosomes or polymyxin and cubosomes alone. Confocal microscopy and neutron reflectometry suggest the superior polytherapy activity is achieved via a two-step process. Firstly, electrostatic interactions between polymyxin and lipid A initially destabilize the outer membrane. Subsequently, an influx of cubosomes results in further membrane disruption via a lipid exchange process. These findings demonstrate that nanoparticle-based polytherapy treatments may potentially serve as improved alternatives to the conventional use of drug-loaded lipid nanoparticles for the treatment of "superbugs".

Zwitterionic Block Copolymer Prodrug Micelles for pH Responsive Drug Delivery and Hypoxia-Specific Chemotherapy.

Tirapazamine (TPZ) and its derivatives (TPZD) have shown their great potential for efficiently killing hypoxic cancer cells. However, unsatisfactory clinical outcomes resulting from the low bioavailability of the low-molecular TPZ and TPZD limited their further applications. Precise delivery and release of these prodrugs via functional nanocarriers can significantly improve the therapeutic effects due to the targeted drug delivery and enhanced permeability and retention (EPR) effect. Herein, zwitterionic block copolymer (BCP) micelles with aldehyde functional groups are prepared from the self-assembly of poly(2-methacryloyloxyethyl phosphorylcholine-b-poly(di(ethylene glycol) methyl ether methacrylate-co-4-formylphenyl methacrylate) [PMPC-b-P(DEGMA-co-FPMA)]. TPZD is then grafted onto PMPC-b-P(DEGMA-co-FPMA) to obtain a polymer-drug conjugate, PMPC-b-P(DEGMA-co-FPMA-g-TPZD) (BCP-TPZ), through the formation of a pH-responsive imine bond, exhibiting a pH-dependent drug release profile owing to the cleavage of the imine bond under acidic conditions. Outstandingly, BCP-TPZ shows around 13.7-fold higher cytotoxicity to hypoxic cancer cells in comparison to normoxic cancer cells evaluated through an in vitro cytotoxicity assay. The pH-responsiveness and hypoxia-specific cytotoxicity confer BCP-TPZ micelles a great potential to achieve precise delivery of TPZD and thus enhance the therapeutic effect toward tumor-hypoxia.

The RNA-binding protein SRSF3 has an essential role in megakaryocyte maturation and platelet production.

RNA processing is increasingly recognized as a critical control point in the regulation of different hematopoietic lineages including megakaryocytes responsible for the production of platelets. Platelets are anucleate cytoplasts that contain a rich repertoire of RNAs encoding proteins with essential platelet functions derived from the parent megakaryocyte. It is largely unknown how RNA binding proteins contribute to the development and functions of megakaryocytes and platelets. We show that serine-arginine-rich splicing factor 3 (SRSF3) is essential for megakaryocyte maturation and generation of functional platelets. Megakaryocyte-specific deletion of Srsf3 in mice led to macrothrombocytopenia characterized by megakaryocyte maturation arrest, dramatically reduced platelet counts, and abnormally large functionally compromised platelets. SRSF3 deficient megakaryocytes failed to reprogram their transcriptome during maturation and to load platelets with RNAs required for normal platelet function. SRSF3 depletion led to nuclear accumulation of megakaryocyte mRNAs, demonstrating that SRSF3 deploys similar RNA regulatory mechanisms in megakaryocytes as in other cell types. Our study further suggests that SRSF3 plays a role in sorting cytoplasmic megakaryocyte RNAs into platelets and demonstrates how SRSF3-mediated RNA processing forms a central part of megakaryocyte gene regulation. Understanding SRSF3 functions in megakaryocytes and platelets provides key insights into normal thrombopoiesis and platelet pathologies as SRSF3 RNA targets in megakaryocytes are associated with platelet diseases.

Rational design of potent ultrashort antimicrobial peptides with programmable assembly into nanostructured hydrogels.

Microbial resistance to common antibiotics is threatening to cause the next pandemic crisis. In this context, antimicrobial peptides (AMPs) are receiving increased attention as an alternative approach to the traditional small molecule antibiotics. Here, we report the bi-functional rational design of Fmoc-peptides as both antimicrobial and hydrogelator substances. The tetrapeptide Fmoc-WWRR-NH2-termed Priscilicidin-was rationally designed for antimicrobial activity and molecular self-assembly into nanostructured hydrogels. Molecular dynamics simulations predicted Priscilicidin to assemble in water into small oligomers and nanofibrils, through a balance of aromatic stacking, amphiphilicity and electrostatic repulsion. Antimicrobial activity prediction databases supported a strong antimicrobial motif via sequence analogy. Experimentally, this ultrashort sequence showed a remarkable hydrogel forming capacity, combined to a potent antibacterial and antifungal activity, including against multidrug resistant strains. Using a set of biophysical and microbiology techniques, the peptide was shown to self-assemble into viscoelastic hydrogels, as a result of assembly into nanostructured hexagonal mesophases. To further test the molecular design approach, the Priscilicidin sequence was modified to include a proline turn-Fmoc-WPWRR-NH2, termed P-Priscilicidin-expected to disrupt the supramolecular assembly into nanofibrils, while predicted to retain antimicrobial activity. Experiments showed P-Priscilicidin self-assembly to be effectively hindered by the presence of a proline turn, resulting in liquid samples of low viscosity. However, assembly into small oligomers and nanofibril precursors were evidenced. Our results augur well for fast, adaptable, and cost-efficient antimicrobial peptide design with programmable physicochemical properties.