The molecular determinants of calcium ATPase inhibition by curcuminoids.

The natural product curcumin and some of its analogs are known inhibitors of the transmembrane enzyme sarco/endoplasmic reticulum calcium ATPase (SERCA). Despite their widespread use, the curcuminoids' binding site in SERCA and their relevant interactions with the enzyme remain elusive. This lack of knowledge has prevented the development of curcuminoids into valuable experimental tools or into agents of therapeutic value. We used the crystal structures of SERCA in its E1 conformation in conjunction with computational tools such as docking and surface screens to determine the most likely curcumin binding site, along with key enzyme/inhibitor interactions. Additionally, we determined the inhibitory potencies and binding affinities for a small set of curcumin analogs. The predicted curcumin binding site is a narrow cleft in the transmembrane section of SERCA, close to the transmembrane/cytosol interface. In addition to pronounced complementarity in shape and hydrophobicity profiles between curcumin and the binding pocket, several hydrogen bonds were observed that were spread over the entire curcumin scaffold, involving residues on several transmembrane helices. Docking-predicted interactions were compatible with experimental observations for inhibitory potencies and binding affinities. Based on these findings, we propose an inhibition mechanism that assumes that the presence of a curcuminoid in the binding site arrests the catalytic cycle of SERCA by preventing it from converting from the E1 to the E2 conformation. This blockage of conformational change is accomplished by a combination of steric hinderance and hydrogen-bond-based cross-linking of transmembrane helices that require flexibility throughout the catalytic cycle.

Structural analysis and blood-enriching effects comparison based on biological potency of Angelica sinensis polysaccharides.

Angelica sinensis is a long-standing medicine used by Chinese medical practitioners and well-known for its blood-tonic and blood-activating effects. Ferulic acid, ligustilide, and eugenol in Angelica sinensis activate the blood circulation; however, the material basis of their blood-tonic effects needs to be further investigated. In this study, five homogeneous Angelica sinensis polysaccharides were isolated, and their sugar content, molecular weight, monosaccharide composition, and infrared characteristics determined. Acetylphenylhydrazine (APH) and cyclophosphamide (CTX) were used as inducers to establish a blood deficiency model in mice, and organ indices, haematological and biochemical parameters were measured in mice. Results of in vivo hematopoietic activity showed that Angelica sinensis polysaccharide (APS) could elevate erythropoietin (EPO), granulocyte colony-stimulating factor (G-CSF), and interleukin-3 (IL-3) serum levels, reduce tumor necrosis factor-α (TNF-α) level in mice, and promote hematopoiesis in the body by regulating cytokine levels. Biological potency test results of the in vitro blood supplementation indicated strongest tonic activity for APS-H2O, and APS-0.4 has the weakest haemopoietic activity. The structures of APS-H2O and APS-0.4 were characterized, and the results showed that APS-H2O is an arabinogalactan glycan with a main chain consisting of α-1,3,5-Ara(f), α-1,5- Ara(f), ß-1,4-Gal(p), and ß-1,4-Gal(p)A, and two branched chains of ß-t-Gal(p) and α-t-Glc(p) connected to each other in a (1→3) linkage to α-1,3,5-Ara(f) on the main chain. APS-0.4 is an acidic polysaccharide with galacturonic acid as the main chain, consisting of α-1,4-GalA, α-1,2-GalA, α-1,4-Gal, and ß-1,4-Rha. In conclusion, APS-H2O can be used as a potential drug for blood replenishment in patients with blood deficiency, providing a basis for APS application in clinical treatment and health foods, as well as research and development of new polysaccharide-based drugs.

In vitro CAR-T cell killing: validation of the potency assay.

For advanced therapy medicinal products, the development and validation of potency assays are required, in accordance with international guidelines, to characterise the product and obtain reliable and consistent data. Our purpose was to validate the killing assay for the evaluation of autologous anti-CD19 chimeric antigen receptor (CAR) T potency. We used CD4 + and CD8 + lymphocytes or anti-CD19 CAR-T cells as effector cells and REH (CD19 +) or MOLM-13 (CD19 -) cell lines as target cells. After co-culturing target and effector cells (1:1 ratio) for 24 h, samples were labelled with 7-AAD, anti-CD3 and anti-CD19 antibodies and the frequency of CD19 + dead cells was evaluated by flow cytometry. In order to verify the CAR-T specificity for the CD19 + target, the co-culture between CAR-T and REH or MOLM-13 at different effector-to-target ratios was scheduled. Moreover, not transduced CD4 + and CD8 + lymphocytes were tested in comparison with CAR-T from the same donor to demonstrate the assay specificity. Linearity and accuracy were evaluated, and established acceptance criteria were compiled for both parameters (r2 ≥ 0.97 for linearity and average relative error ≤ 10% for accuracy). Furthermore, the method was considered robust when performed between 23 and 25 h of co-culture, and the intra-assay, inter-assay and inter-day precision was obtained. Finally, in order to verify the inter-analyst precision, the test was executed by three different operators and the intra-class correlation coefficient was > 0.4 in both cases. In conclusion, we consider this CAR-T potency assay as validated and usable in all steps of product development and quality control.

Potency Assay Variability Estimation in Practice.

During the drug development process, testing potency plays an important role in the quality assessment required for the manufacturing and marketing of biologics. Due to multiple operational and biological factors, higher variability is usually observed in bioassays compared with physicochemical methods. In this paper, we discuss different sources of bioassay variability and how this variability can be statistically estimated. In addition, we propose an algorithm to estimate the variability of reportable results associated with different numbers of runs and their corresponding OOS rates under a given specification. Numerical experiments are conducted on multiple assay formats to elucidate the empirical distribution of bioassay variability.

Supercoiled DNA percentage: A key in-process control of linear DNA template for mRNA drug substance manufacturing.

The development of messenger RNA (mRNA) vaccines and therapeutics necessitates the production of high-quality in vitro-transcribed mRNA drug substance with specific critical quality attributes (CQAs), which are closely tied to the uniformity of linear DNA template. The supercoiled plasmid DNA is the precursor to the linear DNA template, and the supercoiled DNA percentage is commonly regarded as a key in-process control (IPC) during the manufacturing of linear DNA template. In this study, we investigate the influence of supercoiled DNA percentage on key mRNA CQAs, including purity, capping efficiency, double-stranded RNA (dsRNA), and distribution of poly(A) tail. Our findings reveal a significant impact of supercoiled DNA percentage on mRNA purity and in vitro transcription yield. Notably, we observe that the impact on mRNA purity can be mitigated through oligo-dT chromatography, alleviating the tight range of DNA supercoiled percentage to some extent. Overall, this study provides valuable insights into IPC strategies for DNA template chemistry, manufacturing, and controls (CMC) and process development for mRNA drug substance.

Characterization of near-field temporal and spatial variations of pesticide residues using honeybee specimens as bio-sensing matrices.

This study was prompted by recent reports of the ubiquity of neonicotinoids (neonics) in environment and the likelihood of exposures and health hazards to non-target organisms. We aimed to quantify neonics levels in time- and location-match pollen and nectar samples foraged by honeybees (Apis mellifera) and characterized the temporal and spatial variations using a relative potency factor method to determine the total neonic levels, expressed as the imidacloprid-adjusted total neonics, IMIRPF (ng/g). Six pairs of pollen and nectar samples, a total of twelve samples, were collected from each of the thirty-two experimental hives during the active foraging months of March, April, and June and analyzed for eight neonics. We found 59% and 64% of pollen and nectar contained at least one neonic, respectively. Among those neonic-detected pollen and nectar samples, 45% and 77% of them contained more than one neonic, respectively. Imidacloprid and acetamiprid in pollen and clothianidin and thiamethoxam in nectar accounted for 60% and 83% detection, respectively. The highest 3-month average of IMIRPF in pollen (6.56 ng/g) and nectar (11.19 ng/g) were detected in a location with the predominant production of citrus fruit. The temporal and spatial variations of IMIRPF levels demonstrated the robustness of using paired pollen and nectar data as the bio-sensing matrices to facilitate the assessment of near-field exposure to total neonics and the delineation of risks.

Analysing the Potency of a Seasonal Influenza Vaccine Using Reference Antisera from Heterologous Strains.

The potency of inactivated seasonal influenza vaccine is harmonised by establishing the haemagglutinin (HA) content using the compendial single radial diffusion (SRD) method. SRD reagents (antigens and antisera) are prepared, calibrated and distributed by regulatory agencies as standards for potency testing, following the biannual World Health Organization (WHO) announcements of the virus strains suitable for inclusion in the vaccine. The generation of a homologous hyperimmune sheep antiserum constrains the time to vaccine release. This study tests the application of heterologous antisera to determine the potency of influenza vaccine compared to that of a standard homologous antiserum. The results indicate that the selected heterologous sheep antisera directed to seasonal H1N1, H3N2 or B Victoria virus strains can be used to determine the accurate potency of inactivated seasonal influenza vaccines. Individually selected antisera could be useful for two to fourteen seasons. A limitation to the heterologous antiserum approach is the diversity of each individual serum, indicating that the empirical determination of a specific serum is required. This application has the potential to enable the earlier availability of a seasonal vaccine and reduce animal usage.

Generation and characterization of mesenchymal stem cells from the affected femoral heads of dogs with Legg Calvé Perthes disease.

Background: Canine Legg Calvé Perthes disease (LCPD) occurs during the growth period, and the cause of ischemic necrosis of the femoral head during growth remains unclear. If LCPD-affected femoral head-derived mesenchymal stem cells (LCPD-MSCs) can be generated, they can be used as a new tool for the pathophysiological analysis of canine LCPD. Aim: To generate affected femoral head-derived mesenchymal stem cells (MSCs) from dogs with LCPD and investigate the mRNA expression levels of angiogenesis-related factors and osteogenic differentiation potency of LCPD-MSCs. Methods: This study was performed using affected femoral heads from dogs diagnosed with LCPD and underwent femoral head and neck ostectomy. The necrotic tissue was harvested from the LCPD-affected femoral head and cultured statically (LCPD group, n = 6). Canine bone marrow-derived MSCs (BM-MSCs) were used as controls (control group, n = 6). First, the morphology of the cultured cells was observed, and the expression of CD29, CD34, CD44, CD45, CD90, and major histocompatibility complex class II was analyzed using flow cytometry. Additionally, the trilineage differentiation potency of the LCPD-affected head-derived adherent cells was examined. Furthermore, the expression levels of HIF1A, VEGFA, VEGFB, and PDGFB mRNAs and the bone differentiation potency of LCPD-affected head-derived adherent cells were investigated. Results: LCPD-affected femoral head-derived adherent cells showed a fibroblast-like morphology, and the expression of cell surface antigens was similar to that of BM-MSCs. In addition, LCPD-affected femoral head-derived adherent cells showed the same trilineage differentiation potency as BM-MSCs and were consistent with MSC characteristics. Furthermore, the mRNA expression levels of angiogenesis-related factors could be objectively measured in LCPD-MSCs and those MSCs had bone differentiation potency. Conclusion: In the present study, canine LCPD-MSCs were successfully generated, suggesting their usefulness as a tool for pathological analysis of LCPD in dogs.

A Hybrid Approach to Hood-Sparing Robotic Prostatectomy to Maximize Functional Outcomes and Maintain Early Oncologic Efficacy.

Background: We detail our approach and experience with a hybrid version of the endopelvic hood-sparing (HS) robot-assisted radical prostatectomy (RARP) using the da Vinci robotic platform. Materials and Methods: We retrospectively reviewed the records of 200 patients who underwent RARP by a single surgeon. Patients were propensity-matched into three cohorts depending on biopsy and prostatectomy Gleason Grade Groups: traditional retropubic (RP) (n = 80), retzius-sparing (RS) (n = 40), and HS (n = 80). Patient characteristics and oncologic and functional outcomes were examined. Zero pads per day defined return of continence. Erections suitable for penetrative intercourse with/without medications defined return of sexual function. Results: Patient characteristics were similar between cohorts excluding prostate-specific antigen levels (p = 0.014), which were significantly lower in the RS cohort (7.1 ± 5.3 ng/mL) compared with RP (9.2 ± 9.3 ng/mL) and HS (8.8 ± 8.9 ng/mL). Clinically significant positive margin rates were significantly higher (p = 0.046) in the RS cohort (32.5%) compared with RP (17.5%) and HS (13.9%). Biochemical recurrence and metastasis rates were similar between all cohorts. Median time to continence was significantly lower for RS and HS-RARP (p < 0.001) compared with RP-RARP at 1.3, 1.6, and 5.4 months, respectively. Median time to return of sexual function was significantly lower for RS and HS-RARP (p < 0.001) compared with RP-RARP at 4.0, 7.7, and 15.1 months, respectively. Conclusions: Our hybrid HS-RARP approach provides functional outcomes similar to RS-RARP with the early oncologic control of traditional RP-RARP.

The Relationship among Internet Addiction, Moral Potency, Mindfulness, and Psychological Capital.

This research aimed to contribute to the literature on internet addiction (IA) and moral development among university students. Moral potency (MP) encompasses the interconnected dimensions of moral courage, moral ownership, and moral efficacy. Studies on the relationships between students' problematic behaviors (e.g., IA) and cognitive processes like MP, mindfulness (MI), and psychological capital (PsyCap) are scarce in educational research. Therefore, this study investigated the relationships among IA, MP, MI, and PsyCap in university students. This study included 868 undergraduate students from a state university in Ethiopia, with 526 male students (60.6%) and 342 female students (39.4%). Participants' ages ranged from 21 to 29 years, with a mean age of 22.31 and a standard deviation of 4.03. The findings indicated that IA was negatively correlated with MI, PsyCap, and MP. Both MI and PsyCap showed positive correlations with MP. Importantly, this study revealed that IA had a direct and negative impact on MI, PsyCap, and MP. Further, MI and PsyCap partially mediated and fully mediated the relationship between IA and MP. These findings suggest that cultivating MI and positive PsyCap among university students could be an important strategy to reduce the risks of IA and enhance their moral development. This study contributes to the limited research on the complex relationships between technology use, psychological resources, and moral functioning in emerging adulthood.

Update on Non-Interchangeability of Botulinum Neurotoxin Products.

The growing use of botulinum neurotoxins (BoNTs) for medical and aesthetic purposes has led to the development and marketing of an increasing number of BoNT products. Given that BoNTs are biological medications, their characteristics are heavily influenced by their manufacturing methods, leading to unique products with distinct clinical characteristics. The manufacturing and formulation processes for each BoNT are proprietary, including the potency determination of reference standards and other features of the assays used to measure unit potency. As a result of these differences, units of BoNT products are not interchangeable or convertible using dose ratios. The intrinsic, product-level differences among BoNTs are compounded by differences in the injected tissues, which are innervated by different nerve fiber types (e.g., motor, sensory, and/or autonomic nerves) and require unique dosing and injection sites that are particularly evident when treating complex therapeutic and aesthetic conditions. It is also difficult to compare across studies due to inherent differences in patient populations and trial methods, necessitating attention to study details underlying each outcome reported. Ultimately, each BoNT possesses a unique clinical profile for which unit doses and injection paradigms must be determined individually for each indication. This practice will help minimize unexpected adverse events and maximize efficacy, duration, and patient satisfaction. With this approach, BoNT is poised to continue as a unique tool for achieving individual goals for an increasing number of medical and aesthetic indications.

Synthesis and larvicidal efficacy of pyrazolopyrimidine derivatives conjugated with selenium nanoparticles against Culex pipiens L. and Musca domestica L. larvae.

The synthesized pyrazolopyrimidine derivatives conjugated with selenium nanoparticles were prepared via a reaction of pyrazolone 1 with aryl-aldehyde and malononitrile or 3-oxo-3-phenylpropanenitrile in the presence ammonium acetate or pipridine using an ultrasonic bath as a modified method in the organic synthesis for such materials. The structure of the synthesized compounds was elucidated through various techniques. All the synthesized pyrazolopyrimidines were used in the synthesis of selenium nanoparticles (SeNPs). These nanoparticles were confirmed using UV-spectra, Dynamic Light scattering and (TEM) techniques. The larvicidal efficiency;of the synthesized;compounds; was investigated against some strains such as Culex pipiens;and Musca domestica larvae. Bioassay test showed pyrazolopyrimide derivatives to exhibit an acceptable larvicidal;bio-efficacy. The derivative (3) exhibited;the highest;efficiency for more than; lab strains of both species. Moreover, C. pipiens larvae were more sensitive towards the examined compounds than M. domestica. The field;strain displayed lower affinity for the 2 folds compounds. Some biochemical changes were tracked through analysis of insect main metabolites (protein, lipid and carbohydrate), in addition to measuring the changes in seven enzymes after treatment. Generally, there was a reduction in the protein, lipids and carbohydrates after treatment with all tested compounds. Moreover, a decrement was noticed for acetylcholine esterase and glutathione;S-transferase; enzymes. There was an increment in the acid;phosphatase; and alkaline phosphatase. In addition, there was elevation in Phenoloxidase level but it noticed the declination in both Cytochrome P450 and Ascorbate peroxidase activity after treatment both flies with derivatives of selenium-nanoparticles in both lab and field strain. Generally, the experiments carried out indicate that antioxidant and detoxification enzymes may play a significant role in mechanism of action of our novel nanocompounds. The cytotoxicity of the synthesized compounds and conjugated with SeNPs showed enhanced compatibility with human normal fibroblast cell line (BJ1) with no toxic effect.

Impact of chitosan extracted from shrimp shells on the shrinkage and mechanical properties of cement-based composites using dendritic fibrous nanosilica.

Dendritic fibrous nanosilica (DFNS) was functionalized using microcrystalline chitosan, derived from shrimp exoskeletons, to act as a robust anchor, resulting in DFNS@Chitosan. In order to prevent the restacking of chitosan sheets, the supramolecular polymerized chitosan not only served as a spacer but was also incorporated into cement-based composites. The physical-chemical characteristics of DFNS@Chitosan were assessed through various analytical techniques such as TEM, SEM, TGA, FTIR, AFM, XPS, and EDX. The potency and auto-induced contraction of Cement-based composite materials fortified with DFNS@Chitosan were probed. The incorporation of DFNS@Chitosan resulted in an increase in both compressive and interfacial stretching potency of the cement-based composites. Furthermore, the presence of DFNS@Chitosan effectively inhibited the occurrence of auto-induced contraction in the cement-based paste. This research endeavor is anticipated to promote an alternative utilization of DFNS and shrimp waste shells in the development of sustainable building materials.

Effect of 3D and 2D cell culture systems on trophoblast extracellular vesicle physico-chemical characteristics and potency.

The growing understanding of the role of extracellular vesicles (EVs) in embryo-maternal communication has sparked considerable interest in their therapeutic potential within assisted reproductive technology, particularly in enhancing implantation success. However, the major obstacle remains the large-scale production of EVs, and there is still a gap in understanding how different culture systems affect the characteristics of the EVs. In the current study, trophoblast analogue human chorionic carcinoma cell line was cultivated in both conventional monolayer culture (2D) and as spheroids in suspension culture (3D) and how the cell growth environment affects the physical, biochemical and cellular signalling properties of EVs produced by them was studied. Interestingly, the 3D system was more active in secreting EVs compared to the 2D system, while no significant differences were observed in terms of morphology, size, and classical EV protein marker expression between EVs derived from the two culture systems. There were substantial differences in the proteomic cargo profile and cellular signalling potency of EVs derived from the two culture systems. Notably, 2D EVs were more potent in inducing a cellular response in endometrial epithelial cells (EECs) compared to 3D EVs. Therefore, it is essential to recognize that the biological activity of EVs depends not only on the cell of origin but also on the cellular microenvironment of the parent cell. In conclusion, caution is warranted when selecting an EV production platform, especially for assessing the functional and therapeutic potential of EVs through in vitro studies.

Isthmus progenitor cells contribute to homeostatic cellular turnover and support regeneration following intestinal injury.

The currently accepted intestinal epithelial cell organization model proposes that Lgr5+ crypt-base columnar (CBC) cells represent the sole intestinal stem cell (ISC) compartment. However, previous studies have indicated that Lgr5+ cells are dispensable for intestinal regeneration, leading to two major hypotheses: one favoring the presence of a quiescent reserve ISC and the other calling for differentiated cell plasticity. To investigate these possibilities, we studied crypt epithelial cells in an unbiased fashion via high-resolution single-cell profiling. These studies, combined with in vivo lineage tracing, show that Lgr5 is not a specific ISC marker and that stemness potential exists beyond the crypt base and resides in the isthmus region, where undifferentiated cells participate in intestinal homeostasis and regeneration following irradiation (IR) injury. Our results provide an alternative model of intestinal epithelial cell organization, suggesting that stemness potential is not restricted to CBC cells, and neither de-differentiation nor reserve ISC are drivers of intestinal regeneration.

Advances in ex vivo expansion of hematopoietic stem and progenitor cells for clinical applications.

As caretakers of the hematopoietic system, hematopoietic stem cells assure a lifelong supply of differentiated populations that are responsible for critical bodily functions, including oxygen transport, immunological protection and coagulation. Due to the far-reaching influence of the hematopoietic system, hematological disorders typically have a significant impact on the lives of individuals, even becoming fatal. Hematopoietic cell transplantation was the first effective therapeutic avenue to treat such hematological diseases. Since then, key use and manipulation of hematopoietic stem cells for treatments has been aspired to fully take advantage of such an important cell population. Limited knowledge on hematopoietic stem cell behavior has motivated in-depth research into their biology. Efforts were able to uncover their native environment and characteristics during development and adult stages. Several signaling pathways at a cellular level have been mapped, providing insight into their machinery. Important dynamics of hematopoietic stem cell maintenance were begun to be understood with improved comprehension of their metabolism and progressive aging. These advances have provided a solid platform for the development of innovative strategies for the manipulation of hematopoietic stem cells. Specifically, expansion of the hematopoietic stem cell pool has triggered immense interest, gaining momentum. A wide range of approaches have sprouted, leading to a variety of expansion systems, from simpler small molecule-based strategies to complex biomimetic scaffolds. The recent approval of Omisirge, the first expanded hematopoietic stem and progenitor cell product, whose expansion platform is one of the earliest, is predictive of further successes that might arise soon. In order to guarantee the quality of these ex vivo manipulated cells, robust assays that measure cell function or potency need to be developed. Whether targeting hematopoietic engraftment, immunological differentiation potential or malignancy clearance, hematopoietic stem cells and their derivatives need efficient scaling of their therapeutic potency. In this review, we comprehensively view hematopoietic stem cells as therapeutic assets, going from fundamental to translational.

Mechanistic insights into complement pathway inhibition by CR1 domain duplication.

Complement receptor 1 (CR1) is a membrane glycoprotein with a highly duplicated domain structure able to bind multiple ligands such as C3b and C4b, the activated fragments of complement components C3 and C4, respectively. We have previously used our knowledge of this domain structure to identify CSL040, a soluble extracellular fragment of CR1 containing the long homologous repeat (LHR) domains A, B, and C. CSL040 retains the ability to bind both C3b and C4b but is also a more potent complement inhibitor than other recombinant CR1-based therapeutics. To generate soluble CR1 variants with increased inhibitory potential across all three complement pathways, or variants with activity skewed to specific pathways, we exploited the domain structure of CR1 further by generating LHR domain duplications. We identified LHR-ABCC, a soluble CR1 variant containing a duplicated C3b-binding C-terminal LHR-C domain that exhibited significantly enhanced alternative pathway inhibitory activity in vitro compared to CSL040. Another variant, LHR-BBCC, containing duplications of both LHR-B and LHR-C with four C3b binding sites, was shown to have reduced classical/lectin pathway inhibitory activity compared to CSL040, but comparable alternative pathway activity. Interestingly, multiplication of the C4b-binding LHR-A domain resulted in only minor increases in classical/lectin pathway inhibitory activity. The CR1 duplication variants characterized in these in vitro potency assays, as well as in affinity in solution C3b and C4b binding assays, not only provides an opportunity to identify new therapeutic molecules but also additional mechanistic insights to the multiple interactions between CR1 and C3b/C4b.

Neurovascular Structure-Adjacent Frozen-Section Examination (NeuroSAFE) Technique of Nerve-Sparing Robot-Assisted Radical Prostatectomy (RARP) in Indian Scenario: Technique, Feasibility, and Early Outcomes.

Potency and urinary continence are adversely affected post-prostatectomy. The primary objective is oncological safety by ensuring negative surgical margins (NSM) and best functional recovery through nerve preservation in appropriate patients. NeuroSAFE technique of intra-operative frozen-section (IFS) analysis was devised for comprehensive assessment of surgical margins adjacent to the neurovascular tissue surface of the prostate. We analyzed our initial experience with this technique. Five NS-RARPs were performed utilizing the NeuroSAFE technique between October 2021 and February 2022. Patient demographics, disease stage, operative console time, post-operative complications, final histopathology, biochemical recurrence (BCR), erectile function, and urinary continence were recorded. The mean age of patients was 59.2 ± 1.3 years. All had clinically organ-confined disease with ISUP grade ≤ 3. The mean operative time of NS-RARP with NeuroSAFE was 240 ± 21 min and average NeuroSAFE time was 45 ± 3.8 min. All patients had NSM on IFS. No patient had Clavien-Dindo grade > 1 complications. Margins were negative on final histopathology. No patient had BCR at 6 and 12 weeks. Three patients were able to have sexual intercourse and only one patient required single precaution pad at 12 weeks. NeuroSAFE is feasible and can ensure intra-operative oncological safety of the NS procedure. Moreover, it gives the opportunity to convert positive surgical margin to prognostically favorable NSM by secondary resection. Our initial experience which is the first in India is encouraging with favorable functional outcomes. Large prospective studies and longer follow-up are required specially to evaluate the oncological benefit.

Macrocyclization strategy for improving candidate profiles in medicinal chemistry.

Macrocycles are defined as cyclic compounds with 12 or more members. In medicinal chemistry, they are categorized based on their core chemistry into cyclic peptides and macrocycles. Macrocycles are advantageous because of their structural diversity and ability to achieve high affinity and selectivity towards challenging targets that are often not addressable by conventional small molecules. The potential of macrocyclization to optimize drug-like properties while maintaining adequate bioavailability and permeability has been emphasized as a key innovation in medicinal chemistry. This review provides a detailed case study of the application of macrocyclization over the past 5 years, starting from the initial analysis of acyclic active compounds to optimization of the resulting macrocycles for improved efficacy and drug-like properties. Additionally, it illustrates the strategic value of macrocyclization in contemporary drug discovery efforts.

Effect of conditioned media on the angiogenic activity of mesenchymal stem cells.

Mesenchymal stem cells (MSCs) are promising candidates for use in novel cell therapies, although such live cell products are highly complex compared with traditional drugs. For example, difficulties such as the control of manufacturing conditions hinder the manufacture of stable cell populations that maintain their therapeutic potency. Here, assuming that medium selection significantly affects cell potency, we focused on the culture media as a critical manufacturing factor influencing the therapeutic efficacy of MSCs. We therefore performed a tube formation assay to quantify the angiogenic activities of conditioned media used to culture human umbilical vein endothelial cells compared with unconditioned media. Comprehensive molecular genetic analysis using microarrays was applied to determine the effects of these media on signal transduction pathways. We found that activation of the vascular endothelial growth factor (VEGF) signaling pathway differed, and that VEGF concentration was dependent on the composition of the conditioned media. These results indicate that the activation level of cell signaling pathways which contribute to therapeutic efficacy may vary depending on the media components affecting MSCs during their cultivation. Moreover, they indicate that therapeutic efficacy will likely depend on how cells are handled during manufacture. These findings will enhance our understanding of the quality control measures required to ensure the efficacy and safety of cell therapy products.