Intracellular delivery of antisense oligonucleotides by tri-branched cyclic disulfide units.

Therapeutic oligonucleotides, such as antisense DNA, show promise in treating previously untreatable diseases. However, their applications are still hindered by the poor membrane permeability of naked oligonucleotides. Therefore, it is necessary to develop efficient methods for intracellular oligonucleotide delivery. Previously, our group successfully developed disulfide-based Membrane Permeable Oligonucleotides (MPON), which achieved enhanced cellular uptake and gene silencing effects through an endocytosis-free uptake mechanism.  Herein, we report a new molecular design for the next generation of MPON, called trimer MPON. The trimer MPON consists of a tri-branched backbone, three α-lipoic acid units, and a spacer linker between the oligonucleotides and tri-branched cyclic disulfide unit. We describe the design, synthesis, and functional evaluation of the trimer MPON, offering new insights into the molecular design for efficient oligonucleotide delivery.

Continued cancer drug approvals in Japan and Europe after market withdrawal in the United States: A comparative study of accelerated approvals.

Regulatory authorities must balance ensuring evidence of efficacy and safety of new drugs. Various regulatory pathways, such as the accelerated approval program in the United States (US), allow authorities to quickly approve drugs for severely ill patients by granting market authorization based on surrogate end points and pending confirmatory trials. In this cross-sectional study, we considered 23 indications of cancer drugs that received accelerated approval by the US Food and Drug Administration (FDA) but were subsequently withdrawn as of April 2023. Our investigation extended to assessing the regulatory status of these accelerated approvals in the European Union (EU) and Japan, examining relevant regulatory documents and identifying factors contributing to the withdrawal in the United States. Comparing regions, we found that for 52% (12/23) and 30% (7/23) of withdrawn accelerated approvals in the United States, sponsors had also sought marketing authorization from the European Medicines Agency (EMA) and Japan's Pharmaceuticals and Medical Devices Agency (PMDA), respectively. As of the April 30, 2023 study cutoff date, 83% (10/12) of drug-indication pairs remained approved by the EMA, while the PMDA retained 100% (7/7). For these indications, the time from FDA withdrawal until the study cutoff date ranged from 0.23 years to 11.45 years for EMA approvals (median: 1.28 years) and 1.10 years to 11.45 years for PMDA approvals (median: 3.22 years). These findings highlight substantial regulatory discrepancies concerning cancer drugs with unconfirmed benefits. Addressing these discrepancies may involve requiring pharmaceutical companies to confirm clinical benefits using more robust end points and fostering international harmonization in regulators' assessment.

Illicit Trade of Prescription Medications Through X (Formerly Twitter) in Japan: Cross-Sectional Study.

BACKGROUND: Nonmedical use of prescription drugs can cause overdose; this represents a serious public health crisis globally. In this digital era, social networking services serve as viable platforms for illegal acquisition of excessive amounts of medications, including prescription medications. In Japan, such illegal drug transactions have been conducted through popular flea market applications, social media, and auction websites, with most of the trades being over-the-counter (OTC) medications. Recently, an emerging unique black market, where individuals trade prescription medications-predominantly nervous system drugs-using a specific keyword ("Okusuri Mogu Mogu"), has emerged on X (formerly Twitter). Hence, these dynamic methods of illicit trading should routinely be monitored to encourage the appropriate use of medications. OBJECTIVE: This study aimed to specify the characteristics of medications traded on X using the search term "Okusuri Mogu Mogu" and analyze individual behaviors associated with X posts, including the types of medications traded and hashtag usage. METHODS: We conducted a cross-sectional study with publicly available posts on X between September 18 and October 1, 2022. Posts that included the term "Okusuri Mogu Mogu" during this period were scrutinized. Posts were categorized on the basis of their contents: buying, selling, self-administration, heads-up, and others. Among posts categorized as buying, selling, and self-administration, medication names were systematically enumerated and categorized using the Anatomical Therapeutic Chemical (ATC) classification. Additionally, hashtags in all the analyzed posts were counted and classified into 6 categories: medication name, mental disorder, self-harm, buying and selling, community formation, and others. RESULTS: Out of 961 identified posts, 549 were included for analysis. Of these posts, 119 (21.7%) referenced self-administration, and 237 (43.2%; buying: n=67, 12.2%; selling: n=170, 31.0%) referenced transactions. Among these 237 posts, 1041 medication names were mentioned, exhibiting a >5-fold increase from the study in March 2021. Categorization based on the ATC classification predominantly revealed nervous system drugs, representing 82.1% (n=855) of the mentioned medications, consistent with the previous survey. Of note, the diversity of medications has expanded to include medications that have not been approved by the Japanese government. Interestingly, OTC medications were frequently mentioned in self-administration posts (odds ratio 23.6, 95% CI 6.93-80.15). Analysis of hashtags (n=866) revealed efforts to foster community connections among users. CONCLUSIONS: This study highlighted the escalating complexity of trading of illegal prescription medication facilitated by X posts. Regulatory measures to enhance public awareness should be considered to prevent illegal transactions, which may ultimately lead to misuse or abuse such as overdose. Along with such pharmacovigilance measures, social approaches that could direct individuals to appropriate medical or psychiatric resources would also be beneficial as our hashtag analysis shed light on the formation of a cohesive or closed community among users.

The "Okusuri Charm" movement in Japan: Prescription drug accessories emerging on X (Twitter).

Background: In this digital age, social networks may offer an avenue for individuals to obtain drugs illicitly beyond the prescribed amount. Users on X (Twitter)® have ingeniously fabricated fashionable accessories that employ prescription drug sheets, termed "Okusuri Charm". Methods: This cross-sectional study scrutinized the emerging "Okusuri Charm" trend, by searching the term in Japanese on X (Twitter)® and analyzing related posts. Results: Alongside illegal prescription drug trading, individuals crafted accessories from drug sheets, particularly prescribed psychiatric drugs, and dealt with other users, leading to a growing trend this year. Conclusions: A positive outlook toward this trend is the emergence of a new artistic movement, but a pessimistic viewpoint is the creators' misuse of prescription drugs, potentially fostering illegal drug dealings.

[A Survey of Illegal Medication Trading through Twitter in Japan].

Non-medical use of prescription medications is a serious public health crisis. The black market for prescription medications should be routinely surveyed to encourage their appropriate use. Herein, we focused on Twitter to investigate the possibility of illicit drug trading in Japan. From March 1 to 8, 2021, we examined the characteristics of Twitter posts, identified using the search term "Okusuri Mogu Mogu", a Japanese argot used for trading of medications. The captured data included the date of the posts, whether with a hashtag was used, an indication of the trades type (buy, sell, self-administration, and unknown), and the name of the mentioned pharmaceutical products. The number of named medications in the posts was counted and further categorized according to the Anatomical Therapeutic Chemical (ATC) classification. Two hundred and thirty-eight posts were identified with the searching term "Okusuri Mogu Mogu", of which 154 (64.7%) named specific medications. Of note, 73 posts (30.7%) were associated with buying or selling medications. We examined the 73 posts. These posts included 118 medications (26 types), of which 107 (88.4%) were classified as nervous system drugs. Hypnotics and sedatives were the most frequently mentioned medications. The present study sheds light on pharmaceutical medication trading via Twitter. Reinforcing the surveillance practices or cracking down on traders by authorities may be insufficient. We consider the possible effectiveness of socially supportive approaches to help those who lack support to access the appropriate psychiatric care.

RNA-based cooperative protein labeling that permits direct monitoring of the intracellular concentration change of an endogenous protein.

Imaging the dynamics of proteins in living cells is a powerful means for understanding cellular functions at a deeper level. Here, we report a versatile method for spatiotemporal imaging of specific endogenous proteins in living mammalian cells. The method employs a bifunctional aptamer capable of selective protein recognition and fluorescent probe-binding, which is induced only when the aptamer specifically binds to its target protein. An aptamer for ß-actin protein preferentially recognizes its monomer forms over filamentous forms, resulting in selective G-actin staining in both fixed and living cells. Through actin-drug treatment, the method permitted direct monitoring of the intracellular concentration change of endogenous G-actin. This protein-labeling method, which is highly selective and non-covalent, provides rich insights into the study of spatiotemporal protein dynamics in living cells.

Non-genetic cell-surface modification with a self-assembling molecular glue.

This report describes the development of a non-genetic cell-surface modification method, in which a self-assembling small molecule is combined with Halo-tag proteins. Cell-surface functionalization with cancer-linked extracellular proteins led to enhanced cell motility, angiogenesis, and immune shielding of the cells, paving the way for translational opportunities for cell therapy.

A prospective compound screening contest identified broader inhibitors for Sirtuin 1.

Potential inhibitors of a target biomolecule, NAD-dependent deacetylase Sirtuin 1, were identified by a contest-based approach, in which participants were asked to propose a prioritized list of 400 compounds from a designated compound library containing 2.5 million compounds using in silico methods and scoring. Our aim was to identify target enzyme inhibitors and to benchmark computer-aided drug discovery methods under the same experimental conditions. Collecting compound lists derived from various methods is advantageous for aggregating compounds with structurally diversified properties compared with the use of a single method. The inhibitory action on Sirtuin 1 of approximately half of the proposed compounds was experimentally accessed. Ultimately, seven structurally diverse compounds were identified.

Multifunctionalization of Cells with a Self-Assembling Molecule to Enhance Cell Engraftment.

Cell-based therapy is a promising approach to restoring lost functions to compromised organs. However, the issue of inefficient cell engraftment remains to be resolved. Herein, we take a chemical approach to facilitate cell engraftment by using self-assembling molecules which modify two cellular traits: cell survival and invasiveness. In this system, the self-assembling molecule induces syndecan-4 clusters on the cellular surface, leading to enhanced cell viability. Further integration with Halo-tag technology provided this self-assembly structure with matrix metalloproteinase-2 to functionalize cells with cell-invasion activity. In vivo experiments showed that the pretreated cells were able to survive injection and then penetrate and engraft into the host tissue, demonstrating that the system enhances cell engraftment. Therefore, cell-surface modification via an alliance between self-assembling molecules and ligation technologies may prove to be a promising method for cell engraftment.

Fabrication of a Multiplexed Artificial Cellular MicroEnvironment Array.

Cellular microenvironments consist of a variety of cues, such as growth factors, extracellular matrices, and intercellular interactions. These cues are well orchestrated and are crucial in regulating cell functions in a living system. Although a number of researchers have attempted to investigate the correlation between environmental factors and desired cellular functions, much remains unknown. This is largely due to the lack of a proper methodology to mimic such environmental cues in vitro, and simultaneously test different environmental cues on cells. Here, we report an integrated platform of microfluidic channels and a nanofiber array, followed by high-content single-cell analysis, to examine stem cell phenotypes altered by distinct environmental factors. To demonstrate the application of this platform, this study focuses on the phenotypes of self-renewing human pluripotent stem cells (hPSCs). Here, we present the preparation procedures for a nanofiber array and the microfluidic structure in the fabrication of a Multiplexed Artificial Cellular MicroEnvironment (MACME) array. Moreover, overall steps of the single-cell profiling, cell staining with multiple fluorescent markers, multiple fluorescence imaging, and statistical analyses, are described.