Cargo Delivery Research Articles

Combating Antimicrobial Resistance by Resensitising Bacteria to Antibiotics Using CRISPR: A Narrative Review

Background: Antimicrobial resistance (AMR) remains a formidable global health threat. Conventional strategy of developing new antibiotics is costly and unsustainable. Thus, innovative approaches for resensitising bacteria using clustered regularly inter-spaced short palindromic repeats (CRISPR) technology are sought. Aims and Objectives: The aim of this study was to review existing technologies and approaches with the aim to identify the most feasible and practical approach to solve AMR globally. Methods: It is a narrative review of CRISPR research to identify a practical medical solution for combating AMR. By considering biological, social and pharmaceutical factors, feasible solutions and their limitations were identified. Next, CRISPR solutions were examined, then filtered based on strict medical expectations. A suitable cargo type and delivery mechanism will be established. Finally, the feasibility from a practical and economic perspective is considered. Results: Among the Cas effector modules examined, Cas12k was identified as the most promising candidate. Notably, Cas12k possesses a small protein size and lacks trans-cleavage activity, enhancing its specificity for targeted bacterial strains. Additionally, its naturally inactivated endonuclease domain further contributes to its precision. In line with this investigation, DNA plasmid was identified as the optimal cargo storage, while lipid nanoparticles were identified as the optimal delivery method. Conclusion: This is the first time an end-to-end consideration to AMR was considered for CRISPR-based strategies. This review offers valuable insights into a promising solution to addressing the AMR crisis. Such knowledge is essential to guide the development of novel AMR solutions.

In Silico Design of Novel EpCAM-Binding Aptamers for Targeted Delivery of RNA Therapeutics

Aptamers are short DNA or RNA sequences that adopt 3D structures and can bind to protein targets with high binding affinity and specificity. Aptamers exhibit excellent tissue penetration, are inexpensive to produce, and can be internalized by cells. Therefore, aptamers are attractive targeting ligands to direct the delivery of theranostic agents to the desired cells. Epithelial cell adhesion molecule (EpCAM) is a tumor-associated antigen that is aberrantly overexpressed on many epithelial-derived cancers, including on cholangiocarcinoma (CCA) cells. Its expression on treatment-resistant cancer stem cells, along with its abundance in the CCA tumor microenvironment, highlights the need to develop EpCAM-targeted therapies for CCA. Herein, an in silico approach was used to design and screen DNA aptamers capable of binding to the EpCAM monomer and homodimer. Two aptamers, PLD01 and PLD02, met the selection criteria and were validated in vitro. Both aptamers exhibited high affinity for EpCAM+ CCA cells, with negligible binding to EpCAM- leukemia cells. Modified versions of PLD01 and PLD02 were successfully incorporated into the membranes of milk-derived nanovesicles. PLD01-functionalized nanovesicles enabled EpCAM-targeted delivery of the therapeutic cargo to CCA cells. In summary, these EpCAM-targeting aptamers can be utilized to direct the delivery of theranostic agents to EpCAM-expressing cells.

Neurotrophic extracellular matrix proteins promote neuronal and iPSC astrocyte progenitor cell- and nano-scale process extension for neural repair applications.

The extracellular matrix plays a critical role in modulating cell behaviour in the developing and adult central nervous system influencing neural cell morphology, function and growth. Neurons and astrocytes, play vital roles in neural signalling and support respectively and respond to cues from the surrounding matrix environment. However, a better understanding of the impact of specific individual extracellular matrix proteins on both neurons and astrocytes is critical for advancing the development of matrix-based scaffolds for neural repair applications. This study aimed to provide an in-depth analysis of how different commonly used extracellular matrix proteins- laminin-1, Fn, collagen IV, and collagen I-affect the morphology and growth of trophic induced pluripotent stem cell (iPSC)-derived astrocyte progenitors and mouse motor neuron-like cells. Following a 7-day culture period, morphological assessments revealed that laminin-1, fibronectin, and collagen-IV, but not collagen I, promoted increased process extension and a stellate morphology in astrocytes, with collagen-IV yielding the greatest increases. Subsequent analysis of neurons grown on the different extracellular matrix proteins revealed a similar pattern with laminin-1, fibronectin, and collagen-IV supporting robust neurite outgrowth. fibronectin promoted the greatest increase in neurite extension, while collagen-I did not enhance neurite growth compared to poly-L-lysine controls. Super-resolution microscopy highlighted extracellular matrix-specific nanoscale changes in cytoskeletal organization, with distinct patterns of actin filament distribution where the three basement membrane-associated proteins (laminin-1, fibronectin, and collagen-IV) promoted the extension of fine cellular processes. Overall, this study demonstrates the potent effect of laminin-1, fibronectin and collagen-IV to promote both iPSC-derived astrocyte progenitor and neuronal growth, yielding detailed insights into the effect of extracellular matrix proteins on neural cell morphology at both the whole cell and nanoscale levels. The ability of laminin-1, collagen-IV and fibronectin to elicit strong growth-promoting effects highlight their suitability as optimal extracellular matrix proteins to incorporate into neurotrophic biomaterial scaffolds for the delivery of cell cargoes for neural repair.

An extracellular vesicle delivery platform based on the PTTG1IP protein

Extracellular vesicles (EVs) are promising therapeutic delivery vehicles, although their potential is limited by a lack of efficient engineering strategies to enhance loading and functional cargo delivery. Using an in-house bioinformatics analysis, we identified N-glycosylation as a putative EV-sorting feature. PTTG1IP (a small, N-glycosylated, single-spanning transmembrane protein) was found to be a suitable scaffold for EV loading of therapeutic cargoes, with loading dependent on its N-glycosylation at two arginine residues. Chimeric proteins consisting of PTTG1IP fused with various cargo proteins, and separated by self-cleaving sequences (to promote cargo release), were shown to enable highly efficient functional delivery of Cre protein to recipient cell cultures and mouse xenograft tumors, and delivery of Cas9-sgRNA complexes to recipient reporter cells. The favorable membrane topology of PTTG1IP enabled facile engineering of further variants with improved properties, highlighting its versatility and potential as a platform for EV-based therapeutics.

Artificial Internalizing Receptors: Intracellular Delivery of Cargo Through Bio-Orthogonal Recognition.

Drug targeting is a methodology that helps to overcome the side effects of therapeutic molecules. However, insufficient targeting specificity and the on-target/off-site delivery leave much room for improvement in the targeting endeavors. One approach to enhance the specificity of drug targeting is to engineer artificial receptors with recognition ligands not observed in nature. To this end, artificial internalizing receptors that feature cholesterylamine as the artificial pull-in mechanism, and an anti-fluorescein antibody as the exofacial recognition and capture tool are developed. Fluorescein labeling is among the most routine techniques in biochemistry and can readily provide a way to make cognate derivatives for receptor-mediated endocytosis using these artificial receptors. Herein, the synthesis and the structure-activity relationship for these artificial receptors are detailed, their potency and efficacy in mediating drug delivery for the antibody-drug conjugates are illustrated, and the scope and limitations of targeting the chemically engineered cells via artificial receptors are investigated. Taken together, the presented data explore an innovative approach to drug targeting and contribute to the development of techniques in cell engineering using the tools of chemistry.

Global cellular proteo-lipidomic profiling of diverse lysosomal storage disease mutants using nMOST.

Lysosomal storage diseases (LSDs) comprise ~50 monogenic disorders marked by the buildup of cellular material in lysosomes, yet systematic global molecular phenotyping of proteins and lipids is lacking. We present a nanoflow-based multi-omic single-shot technology (nMOST) workflow that quantifies HeLa cell proteomes and lipidomes from over two dozen LSD mutants. Global cross-correlation analysis between lipids and proteins identified autophagy defects, notably the accumulation of ferritinophagy substrates and receptors, especially in NPC1-/- and NPC2-/- mutants, where lysosomes accumulate cholesterol. Autophagic and endocytic cargo delivery failures correlated with elevated lyso-phosphatidylcholine species and multi-lamellar structures visualized by cryo-electron tomography. Loss of mitochondrial cristae, MICOS-complex components, and OXPHOS components rich in iron-sulfur cluster proteins in NPC2-/- cells was largely alleviated when iron was provided through the transferrin system. This study reveals how lysosomal dysfunction affects mitochondrial homeostasis and underscores nMOST as a valuable discovery tool for identifying molecular phenotypes across LSDs.

Abstract B046: Enhanced immune cell engineering utilizing a novel, effective intracellular delivery method for introduction of diverse cargo types

Abstract Engineered immune cell therapies have great potential to revolutionize cancer therapeutics and progress has been made in developing both engineering and manufacturing methodologies. However, limitations remain in some areas including long manufacturing times, potential for cell exhaustion, and limited access to the disease site as in the case for solid tumors. In addition, some immune cell types such as monocytes, B cells, and gamma delta T cells have remained difficult to engineer.Portal has developed a novel implementation of mechanoporation technology using a silicon membrane with pores which enables deformation of the cell and poration of the cell membrane, allowing for diffusion of any cargo, charged or uncharged, into the cell. Mechanoporation is a gentle intracellular delivery technique which preserves cell viability and function in cells (DiTommaso, et al PNAS 2018) and has the potential to shorten manufacturing time as well as allow for introduction of key therapy enhancing factors via transient modifications shortly before therapy administration. Portal has established methodology for cytoplasmic introduction of many diverse cargo including mRNA, circular RNA, siRNA, proteins, peptides, and CRISPR RNPs to primary immune cells, including both unstimulated and activated T cells, B cells, NK cells, monocytes and iPSCs. The Portal technology is easily scalable, enabling both research scale (1e6 - 5e7 cells) and process scale (over 1e9 cells) deliveries. Editing and RNA expression efficiencies are above 80% with viability maintained at 70% or above. We have demonstrated sustained expression of circular RNA, and have successfully delivered an mRNA CAR. Cargo can easily be multiplexed with equivalent delivery efficiencies across cargo classes. Portal’s methodology is gentle and effective enough to be utilized multiple times within a therapeutic workflow. For example, we have demonstrated high editing efficiencies in unstimulated T cells, followed by rapid expansion for 7 days and subsequent mRNA delivery at clinical scale - yielding over 50% knockout and 90% mRNA expression. Portal’s technology and approach has the potential to further advance immune cell therapy developments, particularly in multiplexed engineering approaches involving simultaneous delivery of multiple cargos. Coupled with Portal’s compatibility with diverse immune cell types and next generation cargos, such as circRNA, we believe this simple approach to intracellular delivery can dramatically reduce manufacturing time and cost, easily integrate into existing clinical equipment and enable unique enhancements that underpin a new generation of therapeutics. Citation Format: Zhihui Song, Eleni Rogers, Sophia Hirsch, Andrew Larocque, Darby Kreienberg, Rachel Conover, Jacquelyn Hanson, Alec Barclay, Mathias Pawlak, Armon Sharei. Enhanced immune cell engineering utilizing a novel, effective intracellular delivery method for introduction of diverse cargo types [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor Immunology and Immunotherapy; 2024 Oct 18-21; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2024;12(10 Suppl):Abstract nr B046.

VitelloTag: a tool for high-throughput cargo delivery into oocytes.

Delivering molecular tools into oocytes is essential for developmental and reproductive biology. Microinjection, the conventional method, is equipment intensive, often technically challenging and has a low yield, and is impractical in species with delicate oocytes or restricted spawning seasons. To overcome these limitations, we developed VitelloTag, a cost-effective, high-throughput system using vitellogenin-derived fusion proteins to enable efficient cargo delivery via receptor-mediated endocytosis. We demonstrate its utility by delivering Cas9/sgRNA complexes in two distantly related species for gene knockout.

Secure Unmanned Aerial Vehicle Communication in Dual-Function Radar Communication System by Exploiting Constructive Interference

In contrast from traditional unmanned aerial vehicle communication via unlicensed spectrum, connecting unmanned aerial vehicles with cellular networks can extend their communication coverage and improve the quality of their service. In addition, the emerging dual-functional radar communication paradigm in cellular systems can better meet the requirements of location-sensitive tasks such as reconnaissance and cargo delivery. Based on the above considerations, in this paper, we study the simultaneous communication and target sensing issue in cellular-connected unmanned aerial vehicle systems. Specifically, we consider a two-cell coordinated system with two base stations, cellular unmanned aerial vehicles, and potential aerial targets. In such systems, the communication security issue of cellular unmanned aerial vehicles regarding eavesdropping on their target is inevitable since the main beam of the transmit waveform needs to point to the direction of the target for achieving a sufficient detection performance. Aiming at protecting the privacy of cellular transmission as well as performing target sensing, we exploit the physical layer security technique with the aid of constructive interference-based precoding. A transmit power minimization problem is formulated with constraints on secure and reliable cellular transmission and a sufficient radar signal-to-interference-plus-noise ratio. By specially designing the transmit beamforming vectors at the base stations, the received signals at the cellular users are located in the decision regions of the transmitted symbols while the targets can only receive wrong symbols. We also compare the performance of the proposed scheme with that of the traditional one without constructive interference. The simulation results show that the proposed constructive interference-based strategy can meet the requirements of simultaneous target sensing and secure communication, and also save transmit power compared with the traditional scheme.

CD81-guided heterologous EVs present heterogeneous interactions with breast cancer cells

BackgroundExtracellular vesicles (EVs) are cell-secreted particles conceived as natural vehicles for intercellular communication. The capacity to entrap heterogeneous molecular cargoes and target specific cell populations through EV functionalization promises advancements in biomedical applications. However, the efficiency of the obtained EVs, the contribution of cell-exposed receptors to EV interactions, and the predictability of functional cargo release with potential sharing of high molecular weight recombinant mRNAs are crucial for advancing heterologous EVs in targeted therapy applications.MethodsIn this work, we selected the popular EV marker CD81 as a transmembrane guide for fusion proteins with a C-terminal GFP reporter encompassing or not Trastuzumab light chains targeting the HER2 receptor. We performed high-content imaging analyses to track EV-cell interactions, including isogenic breast cancer cells with manipulated HER2 expression. We validated the functional cargo delivery of recombinant EVs carrying doxorubicin upon EV-donor cell treatment. Then, we performed an in vivo study using JIMT-1 cells commonly used as HER2-refractory, trastuzumab-resistant model to detect a more than 2000 nt length recombinant mRNA in engrafted tumors.ResultsFusion proteins participated in vesicular trafficking dynamics and accumulated on secreted EVs according to their expression levels in HEK293T cells. Despite the presence of GFP, secreted EV populations retained a HER2 receptor-binding capacity and were used to track EV-cell interactions. In time-frames where the global EV distribution did not change between HER2-positive (SK-BR-3) or -negative (MDA-MB-231) breast cancer cell lines, the HER2 exposure in isogenic cells remarkably affected the tropism of heterologous EVs, demonstrating the specificity of antiHER2 EVs representing about 20% of secreted bulk vesicles. The specific interaction strongly correlated with improved cell-killing activity of doxorubicin-EVs in MDA-MB-231 ectopically expressing HER2 and reduced toxicity in SK-BR-3 with a knocked-out HER2 receptor, overcoming the effects of the free drug. Interestingly, the fusion protein-corresponding transcripts present as full-length mRNAs in recombinant EVs could reach orthotopic breast tumors in JIMT-1-xenografted mice, improving our sensitivity in detecting penetrant cargoes in tissue biopsies.ConclusionsThis study highlights the quantitative aspects underlying the creation of a platform for secreted heterologous EVs and shows the limits of single receptor-ligand interactions behind EV-cell engagement mechanisms, which now become the pivotal step to predict functional tropism and design new generations of EV-based nanovehicles.

Saponin is Essential for the Isolation of Proteins and RNA from Biological Nanoparticles.

Extracellular vesicles (EVs), biomimetics, and other biological nanoparticles (BNs) produced from human cells are gaining increasing attention in the fields of molecular diagnostics and nanomedicine for the delivery of therapeutic cargo. In particular, BNs are considered prospective delivery vehicles for different biologics, including protein and RNA therapeutics. Moreover, EVs are widely used in molecular diagnostics for early detection of disease-associated proteins and RNA. Technical approaches for measuring biologics mostly originated from the field of EVs and were later adopted for other BNs, such as extracellular vesicle-mimetic nanovesicles, membrane nanoparticles (nanoghosts), and hybrid nanoparticles, with minimal modifications. Here, we demonstrate that BNs are highly resistant to protocols that severely underestimate the protein and RNA content of BNs, and provide the relevance of these data both for general BNs characterization and practical applications of CRISPR/Cas-based therapies. We demonstrate that the addition of saponin leads to an ∼2- to 7-fold enhancement in protein isolation and an ∼2- to 242-fold improvement in RNA recovery rates and detection efficiency. Differences in the proteolipid contents of BNs, measured by Raman and surface-enhanced Raman spectroscopy, correlate with their susceptibility to saponin treatment for cargo extraction. Finally, we develop a unified protocol using saponin to efficiently isolate proteins and RNA from the BNs. These data demonstrate that previously utilized protocols underestimate BN cargo contents and offer gold standard protocols that can be broadly adopted into the field of nanobiologics, molecular diagnostics, and analytical chemistry.

Electrotaxis of Self-Propelling Artificial Swimmers in Microchannels.

Biological microswimmers alter their swimming trajectories to follow the direction of an applied electric field, exhibiting electrotaxis. We show that synthetic active droplet microswimmers also autonomously change swimming trajectories in microchannels, even undergoing "U-turns," in response to an electric field, mimicking electrotaxis. We exploit such electrotaxis, in the presence of an external flow, to robustly tune the swimming trajectory of active droplets between wall-adjacent, oscillatory, and channel centerline swimming. A general hydrodynamic model demonstrates that the electrotactic dynamics is governed by the electrical effects due to the swimmer's inherent surface charge, besides its motility, hydrodynamic wall interactions, and relative orientations of the electric field and imposed flow. Our study demonstrates a simple method for controlling active agents in complex geometries for microrobotic applications, like autonomous cargo delivery.

Picoplatin (II)-loaded chitosan nanocomposites as effective drug delivery systems: Preparation, mechanistic investigation of BSA/5-GMP/GSH binding and biological evaluations

The goal of the current study is to improve the characteristics and bioavailability of the drug picoplatin (PPt) by encapsulating it in chitosan nanoparticles (CS NPs) which allows for the targeted delivery of cytotoxic cargo to cancerous tissue, reducing toxic side effects and raising the therapeutic index. When picoplatin was delivered into the CS, it was able to produce an complex with CS (PPt@CS NPs) that had an appropriate particle size of 275 ± 10 nm, a reasonably low PDI of 0.15 ± 0.05, and high stability (ζ = −22.1 ± 0.3 mV). Since almost all pharmaceuticals work by binding to specific proteins or DNA, the in vitro binding mechanism and affinity of bovine serum albumin (BSA), low molecular building units of nucleic acids (5−GMP), and Glutathione (GSH) (considering that cisplatin resistance could be due to a reaction between cisplatin and GSH) to PPt and PPt@CS NPs were examined using stopped-flow and other spectroscopic approaches. Through two reversible processes, a rapid second-order binding followed by a slower first-order isomerization reaction, and a static quenching mechanism, PPt and PPt@CS NPs bind to BSA with relative reactivity of around (PPt)/(PPt@CS NPs) = 1/2.5. The 5−GMP interaction studies demonstrated that, in addition to changing the binding mechanism, PPt's encapsulation in CS increases its rate of reaction through coordination affinity. PPt interacted with 5-GMP via two reversible processes, a rapid second-order binding to phosphate followed by a slower first−order migration to the N7 of pyrimidine moiety. PPt@CS NPs showed weaker binding to GSH compared to PPt and hence PPt@CS NPs exhibits a lower resistance factor. It was also found that the in vitro drug release of PPt@CS NPs in PBS at pH 7.4 was steady, releasing 30% of the PPt in just 5 hours. Nonetheless, 75% of the release in a pH 5.4 solution containing 10 mM GSH—a solution that mimics the tumor microenvironment—shows that the PPt@CS NPs system is sensitive to GSH and specifically targets malignant tissue. The encapsulation of PPt in CS complex maintained its anticancer activity, as shown by an in vitro cell-survival assay on HepG2 cancer cell lines and also cleavage efficiency toward the minor groove of pBR322 DNA via the hydrolytic way. These findings collectively suggested that inclusion PPt in CS would be an effective strategy to formulate a novel picoplatin formulation intended for use as targeted anticancer treatment.

Fluorescent Reporters, Imaging, and Artificial Intelligence Toolkits to Monitor and Quantify Autophagy, Heterophagy, and Lysosomal Trafficking Fluxes.

Lysosomal compartments control the clearance of cell-own material (autophagy) or of material that cells endocytose from the external environment (heterophagy) to warrant supply of nutrients, to eliminate macromolecules or parts of organelles present in excess, aged, or containing toxic material. Inherited or sporadic mutations in lysosomal proteins and enzymes may hamper their folding in the endoplasmic reticulum (ER) and their lysosomal transport via the Golgi compartment, resulting in lysosomal dysfunction and storage disorders. Defective cargo delivery to lysosomal compartments is harmful to cells and organs since it causes accumulation of toxic compounds and defective organellar homeostasis. Assessment of resident proteins and cargo fluxes to the lysosomal compartments is crucial for the mechanistic dissection of intracellular transport and catabolic events. It might be combined with high-throughput screenings to identify cellular, chemical, or pharmacological modulators of these events that may find therapeutic use for autophagy-related and lysosomal storage disorders. Here, discuss qualitative, quantitative and chronologic monitoring of autophagic, heterophagic and lysosomal protein trafficking in fixed and live cells, which relies on fluorescent single and tandem reporters used in combination with biochemical, flow cytometry, light and electron microscopy approaches implemented by artificial intelligence-based technology.

Engineering Functional Particles to Modulate T Cell Responses.

T cells play a critical role in adaptive immune responses. They work with other immune cells such as B cells to protect our bodies when the first line of defense, the innate immune system, is overcome by certain infectious diseases or cancers. Studying and regulating the responses of T cells, such as activation, proliferation, and differentiation, helps us understand not only their behavior in vivo but also their translation and application in the field of immunotherapy, such as adoptive T cell therapy and immune checkpoint therapy, the situations in which T cells cannot fight cancer alone and require external engineering regulation to help them. Nano- to micrometer-sized particulate biomaterials have achieved great progress in the assistance of T cell-based immunomodulation. For example, various types of microparticles decorated with T cell recognition and activation signals to mimic native antigen-presenting cells have shown successful ex vivo expansion of primary T cells and have been approved for clinical use in adoptive T cell therapy. Functional particles can also serve as vehicles for transporting cargos including small molecule drugs, cytokines, and antibodies. Especially for cargos with limited bioavailability and high repeat-dose toxicity, systemic administration in their free form is difficult. By using particle-assisted systems, the delivery can be tailored on demand, of which targeting and controlled release are two typical examples, ultimately aiding in the regulation of T cell responses. Furthermore, when T cells become overactive and behave in ways that contradict our expectations, such as attacking our own cells or innocuous foreign molecules, this can lead to a breakdown of immune tolerance. In such cases, particles to help reprogram those overactive T cells or suppress their activity are appreciated in vivo. The urgent need to introduce immune stimulation into the treatment of cancers, infectious diseases, and autoimmune diseases has driven recent advances in the engineering of functional particulate biomaterials that regulate T cell responses. In this Account, we will first cover a brief overview of the process of T cell-based immunomodulation from principle to development. It then outlines critical points in the design of functional particle platforms, including materials, size, morphology, surface engineering, and delivery of cargos, to modulate the features of T cells, and introduces selected work from our and other research groups with a focus on three major therapeutic applications: adoptive T cell therapy, immune checkpoint therapy, and immune tolerance restoration. Current challenges and future opportunities are also discussed.

Territories of the «northern delivery»: new approaches to categorization

Ensuring the sustainability of settlements in the Far North regions is achievable only through a comprehensive set of measures for the procurement and delivery of essential goods (primarily food and fuel) from other areas, known as the «Northern Delivery» The current supply system is marked by inefficiencies that create high risks of delivery disruptions. Enhancing this system requires developing and implementing optimal measures to influence the economic relationships within the «Northern Delivery» framework, alongside determining effective legal regulatory methods. A critical step in this process is the establishment of precise and strictly defined territorial boundaries for implementing these measures. At present, there is no legal formalization of spatial limits for the array of managerial, logistic, and socio-economic processes aimed at ensuring uninterrupted deliveries of goods for the «Northern Delivery» underscoring the importance of this research. The aim of this study is to define the criteria and their quantitative parameters to categorize the regions of the Far North and equivalent areas with limited cargo delivery periods. By synthesizing the work of both domestic and international scholars on the zoning of northern territories and assessing their transport accessibility, as well as utilizing cartographic research methods and focus groups, an algorithm for categorizing the settlements of the Far North regions has been developed. The identified selection criteria include transport accessibility, the presence of stable supply logistics chains, the duration of heating period, and the level of provision with basic food products. This categorization resulted in identifying of three groups of territories based on the supply organization mechanism: critical, supporting, and basic supply. The proposed categorization can be used to determine the spatial limits for legal regulation of the «Northern Delivery» and to develop a specific set of measures to ensure the uninterrupted delivery of essential goods.

RESEARCH AND EVALUATION OF LOGISTICS SCHEMES EFFICIENCY FOR THE GRAIN LOADS TRANSPORTATION FOR EXPORT UNDER THE CONDITIONS OF THE MILITARY STATE

Purpose. The aim of the work is to study the effectiveness of various logistics schemes for the delivery of grain cargoes for export from production sites to its transshipment points - to sea ports and to border terminals on the border with the countries of the European Union. Methodology. In the research process analysis and synthesis methods were used to study the content and main provisions of scientific publications on the state of the logistic system of transporting Ukrainian grain for export, the theory of operational work of railways for the development of logistics schemes for the delivery of grain cargoes, methods of mathematical statistics for the analysis of tariffs for road transportation, methods of technical economic calculations and determination of transportation tariffs. Results. In order to evaluate the effectiveness of grain delivery logistics schemes, the routes from one of the areas with the largest concentration of grain loading to a large seaport and to a railway and road border checkpoint were considered. At the same time, the following logistics schemes of grain delivery were chosen: automobile transportation by grain trucks and in containers, railway transportation by wagons and route shipments in grain wagons and in containers on fitting platforms. Calculations showed that transportation costs in the direction of ports are 10...15% lower than for transportation to land checkpoints; rail transportation is generally 30...40% cheaper than road transportation, and transportation in containers allows to save 20...25% of the cost of transportation. Scientific novelty. The obtained results make it possible to provide a scientific and economic basis for the development and improvement of the grain cargo transportation system, as well as for the optimization of the parameters of its individual links. Practical significance. Analysis and improvement of grain transportation logistics schemes will increase the efficiency of grain export logistics, reduce the share of the logistics component in the final cost of Ukrainian grain and, accordingly, increase both its competitiveness on foreign markets and foreign currency revenues to the state budget.

Анализ технологических процессов погрузочно-разгрузочных, транспортных, перегрузочных и складских операций на основе контейнерных перевозок

The article discusses key technological operations, including loading and unloading, transportation, transshipment and storage of goods in containers, as well as analyzes modern approaches to managing and optimizing these processes. The main attention is paid to the automation and mechanization of loading and unloading operations, the use of multimodal transportation to improve the efficiency of transport processes, the development of strategies for optimizing routes and logistics schemes, as well as the use of modern information technologies for cargo tracking and inventory management. The authors of the article emphasize the importance of integrating logistics operations and creating a unified information system for coordinating all stages of container transportation. As a result of the analysis, ways to improve technological processes aimed at reducing costs, increasing the speed of cargo delivery and increasing the overall efficiency of logistics chains are proposed. The article contains recommendations on the introduction of innovative solutions and technologies that can help optimize the work of enterprises engaged in container transportation. The research is based on the analysis of real data, including statistics on the operation of ports, transport companies and warehouse complexes, as well as on the study of best practices in this area. The article provides examples of successful implementation of the proposed approaches in practice, which makes it useful for specialists in the field of logistics, supply chain management and transportation.

PH-Triggered, Lymph Node Focused Immunodrug Release by Polymeric 2-Propionic-3-Methyl-maleic Anhydrides with Cholesteryl End Groups.

Gaining spatial control over innate immune activation is of great relevance during vaccine delivery and anticancer therapy, where one aims at activating immune cells at draining lymphoid tissue while avoiding systemic off-target innate immune activation. Lipid-polymer amphiphiles show high tendency to drain to lymphoid tissue upon local administration. Here, pH-sensitive, cholesteryl end group functionalized polymers as stimuli-responsive carriers are introduced for controlled immunoactivation of draining lymph nodes. Methacrylamide-based monomers bearing pendant 2-propionic-3-methylmaleic anhydride groups are polymerized by Reversible Addition-Fragmentation Chain Transfer (RAFT) polymerization using a cholesterol chain-transfer agent (chol-CTA). The amine-reactive anhydrides are conjugated with various amines, however, while primary amines afforded irreversible imides, secondary amines provided pH-responsive conjugates that are released upon acidification. This can be applied to fluorescent dyes for irreversibly carrier labeling or immunostimulatory Toll-like receptor (TLR) 7/8 agonists as cargos for pH-responsive delivery. Hydrophilization of remaining anhydride repeating units with short PEG-chains yielded cholesteryl-polymer amphiphiles that showed efficient cellular uptake and increased drug release at endosomal pH. Moreover, reversibly conjugated TLR 7/8 agonist amphiphiles efficiently drained to lymph nodes and increased the number of effectively maturated antigen-presenting cells after subcutaneous injection in vivo. Consequently, cholesteryl-linked methacrylamide-based polymers with pH-sensitive 2-propionic-3-methylmaleic anhydride side groups provide ideal features for immunodrug delivery.

Analysis and directions of development of transport and logistics schemes for the delivery of grain cargoes with the participation of inland waterway transport in Russia

In this article, the authors analyzed the transport and logistics schemes for transporting grain cargoes with the possible participation of water transport in a new geopolitical situation. After the introduction of sanctions by Western countries against the Russian Federation, the existing grain delivery schemes lost their relevance, therefore, the authors proposed a scheme for transporting grain cargoes from grain-producing regions of central Russia to southern ports on the Caspian Sea. Based on the presented scheme, the authors proposed options for the delivery of grain cargoes from the subjects of the Russian Federation, tending to inland waterways to Iranian ports, within the framework of the developing international transport corridor (ITC) "North-South". The proposed delivery options necessarily imply the operation of a river fleet and river-sea vessels to deliver grain raw materials to the countries of the Middle East and Africa during navigation, as well as during the inter-navigation period.