Vivo Efficacy Research Articles

Methylene Blue Has Strong Extracellular Virucidal Activity Against a SARS-CoV-2-Related Pangolin Coronavirus with No Intracellular or In Vivo Efficacy

Studies have demonstrated that methylene blue exhibits significant antiviral activity against SARS-CoV-2 or related coronaviruses at the cellular level, suggesting its potential as an anti-SARS-CoV-2 drug. Herein, we report that methylene blue does not exhibit noticeable antiviral activity in a lethal model involving SARS-CoV-2-related pangolin coronavirus GX_P2V (short_3UTR) infection in CAG-hACE2 transgenic mice. We employed plaque reduction assays and cell infection experiments to compare the extracellular virucidal activity of the compound and its ability to inhibit viral replication in cells to those of nirmatrelvir. Methylene blue demonstrated strong virucidal activity but did not inhibit viral replication in cells. The control compound nirmatrelvir lacked virucidal activity but exhibited strong abilities to inhibit viral replication. The virucidal activity of methylene blue was further tested in mouse plasma. Incubation in mouse plasma increased the virucidal EC50 value of methylene blue, indicating that mouse plasma can affect the stability of the compound, although mouse plasma has some extent of natural virucidal activity. These findings elucidate why methylene blue lacks antiviral efficacy in vivo and provide insights for the development of antiviral drugs.

Evaluation of the Rostral Inferior Alveolar Nerve Block via the Mental Foramen in Equids: In Vivo Efficacy Testing.

The use of proper sedative and regional anesthetic protocols is essential when performing equine dental surgical procedures under standing sedation. The efficacy of the rostral inferior alveolar nerve block via the mental foramen has not been previously studied. Aims of this study included determining the efficacy of the block, investigating whether any region (labial mucosa, alveolar mucosa, or teeth) was more reliably anesthetized, and if differences in efficacy existed between bilateral and unilateral blocks. In this blinded trial, 10 horses each were randomly assigned to receive a left unilateral block, right unilateral block or bilateral blocks and 5 horses were bilateral controls. For unilateral groups, the contralateral side of the horse acted as its own unilateral control. Mechanical nociceptive stimulus methods were used to determine response to stimulus at time points pre-sedation, post-sedation, and post-injection at 10, 30, 60, and 90 min. Results were evaluated in 4 groups: unilateral blocks, unilateral controls, bilateral blocks and bilateral controls. Overall, all groups were significantly less likely to respond to stimulus at time points post-sedation, T10, and T30, whereas only blocked sites were less likely to respond at T60 and T90 compared to pre-sedation. There was no significant difference in response to stimuli of blocked sites between the 3 regions at all time points. Overall, at T60 and T90, blocks produced regional anesthesia in 73% and 55% of sites, respectively. This study demonstrated the equine mental foramen regional nerve block has varied efficacy, producing partial or incomplete regional anesthesia in some cases.

Development and Optimization of Transferosomal Gel for Efficient Topical Delivery of Berberine Hydrochloride

Background: Berberine is an isoquinoline alkaloid with potent anti-inflammatory effects. However, its therapeutic efficacy is often restricted by its poor solubility, absorption, and permeability, especially in topical applications. Transferosomes are elastic vesicular carriers with high skin permeability values and retention, making them suitable for encapsulating hydrophilic and lipophilic actives. Objective: The objective of this research was to develop a transferosome-based topical gel formulation of Berberine hydrochloride (BER) to improve its skin permeability and anti-inflammatory efficacy. Method: The thin film hydration method was used to formulate the BER transferosomes. The effects of independent variables, amount of BER in lipid phase (X1), and lipid (Phospholipon 90G) to surfactant ratio (X2) on BER entrapment and vesicle size were studied using face-centered central composite design. The characterization was performed using differential scanning calorimetry, transmission electron microscopy, and X-ray diffraction. The optimized batch (F5) was incorporated in Carbopol gel and further investigated for viscosity, in vitro and ex-vivo diffusion, skin retention by tape stripping, and in-vivo anti-inflammatory efficiency. Results: The formulation optimized with 50 mg of drug and a 5:1 lipid-to-surfactant ratio (F5) demonstrated higher drug entrapment efficiency (72.11%) and lower vesicle size (77.9 nm). TEM validated the spherical vesicle morphology, whereas DSC and XRD analysis confirmed the molecular entrapment of BER within the phospholipid vesicles. The transferosomal gel demonstrated improved BER diffusion (0.63 mg/cm2) confirmed by in vitro and ex-vivo diffusion experiments that revealed a 6-fold increase in flux and permeability coefficient (0.1053 mg. cm-2.h-1). The drug release from transferosome gel was non-Fickian in nature (n = 0.6575), indicating an integration of diffusion and erosion processes. Furthermore, BER transferosomal gel displayed substantial anti- inflammatory activity in rats (p < 0.001). result: The independent variables exerted a substantial impact on drug entrapment and transferosome vesicle size. The optimized batch (F5) exhibited 72.11 % of BER entrapment and 77.9 nm vesicle size. DSC studies indicated molecular dispersion of BER in phospholipon G. The change in BER crystalline state was indicated by the XRD study. In vitro, diffusion study of Transferosome gel formulation indicated the sustained release by a non-fickian mechanism. Ex vivo skin permeation revealed significant improvement in BER permeation (p< 0.05). Further, the tape-stripping method revealed higher BER skin permeability. In-vivo anti-inflammatory efficacy of BER transferosome gel was significantly higher as compared to plain BER gel. Conclusion: The findings demonstrated the potential of transferosomal gel as a promising approach for efficient drug delivery and therapeutic efficacy.

Platelet membrane coated r-SAK to improve thrombolytic efficacy by targeting thrombus

Abstract Background Thrombolytic therapy is one of the most effective treatments to achieve rapid thrombus clearance and recanalization of blocked vessels in thrombotic diseases. However, the application of thrombolytic strategy has been limited due to unsatisfied thrombolytic efficacy, relatively higher bleeding complications, and consequently restricted indications.[1] With the development of nanotechnology and biotechnology, platelet membrane nanoparticles have been developed to enable the targeted delivery of thrombolytic agents to local thrombus sites.[2] Recombinant staphylokinase (r-SAK) is a third-generation thrombolytic agent produced by genetic engineering technology. In this study, platelet membrane coated r-SAK (PLT-r-SAK) was constructed, and its thrombolytic efficacy was evaluated in in-vitro and in-vivo experiments. Purpose We aimed to construct a thrombus-targeting agent of PLT-r-SAK and investigate its thrombolytic efficacy. Methods PLT-r-SAK was prepared by encapsulating r-SAK into platelet membrane vesicles using a membrane extrusion method. An in-vitro thrombolytic experiment was performed to examine the thrombolytic efficacy of PLT-r-SAK, and a totally occluded femoral artery thrombosis model of rabbit was developed to evaluate the in-vivo thrombolytic efficacy of PLT-r-SAK. The changes in femoral artery pressure were adopted to assess the vessel opening status. Besides, the effect of the antiplatelet drugs on the thrombolytic efficacy of PLT-r-SAK was explored. Finally, near-infrared fluorescence imaging system was used to verify the thrombus-targeting effect of PLT-r-SAK. Results PLT-r-SAK was firstly constructed, which exhibited a uniformly dispersed spherical morphology with a diameter of 185.8 ± 1.5 nm and a zeta potential of -26.1 ± 0.8 mV. In the in-vitro animal experiment, the thrombolysis rate of PLT-r-SAK was significantly higher than that of the same (low) dose and 4-fold (high) dose r-SAK (Figure 1a, 1b). In the in-vivo experiment, the initial recanalization time of PLT-r-SAK was significantly shorter than that of the low dose and high dose r-SAK, and the duration of the reperfusion time of PLT-r-SAK was prolonged compared to low dose and high dose r-SAK (Figure 1c). Another in-vitro experiment showed that the increased thrombolysis rate of PLT-r-SAK could be inhibited by platelet poor plasma (PPP) from aspirin- and ticagrelor-treated patients. Finally, both the in-vitro and in-vivo near-infrared fluorescence imaging showed significantly higher radiant efficiency in the PLT-r-SAK group as compared to the NS group (Figure 2). Conclusions Platelet membrane coating significantly improves the thrombolytic efficacy of r-SAK, which is related to the thrombus-targeting activity of the platelet membrane and can be inhibited by aspirin- and ticagrelor-treated plasma.Thrombolytic efficacy of PLT-r-SAKThrombus-targeting evidence of PLT-r-SAK

The Use of a Novel Surgical Irrigant May Be Associated with Decreased Incidence of Surgical Site Infections

Background Periprosthetic joint infection (PJI) represents a formidable challenge to patients undergoing elective total hip arthroplasty (THA) and total knee arthroplasty (TKA). This investigation evaluated a novel antimicrobial surgical irrigation solution, XPerience (XP) (Next Science LLC, Jacksonville, FL), with proven high & persistent in-vivo efficacy against planktonic bacteria and biofilm. The primary objective of this investigation was to compare the incidence of PJI following TKA performed with use of XP versus a standard control rinse (povidone-iodine). The secondary outcome was an all-cause return to operating room (OR) rate. Methods A retrospective cohort study was conducted from 824 consecutive patients undergoing primary THA/TKA from 12/1/2019 - 11/30/2021 treated using povidone-iodine solution and 471 consecutive patients undergoing primary THA/TKA from 12/1/2021 - 12/30/2022 treated with XP at a single institution by the senior author. All surgeries in both groups followed largely identical preoperative, perioperative, and postoperative protocols. A total of 1,295 cases were analyzed. The groups shared largely analogous background characteristics pertaining to age, sex, BMI, and American Society of Anesthesiologists (ASA) class. Results The overall SSI rate was 0% (0/471) in the XP group and 0.49% (4/824) in the control group (P = 0.3). The overall return to OR rate was 0.21% (1/471) in the XP group and 0.85% (7/824) in the control group (P = 0.3). Control group cases returned to the OR for hematoma debridements, periprosthetic fracture fixation, and PJI revisions. Conclusion This investigation is the first to remark on the clinical efficacy of a novel surgical irrigant. There was no statistically significant difference in rates of PJI or return to OR with the irrigant versus without, though a trend towards lower rates was observed in the context of a low baseline infection rate. Limitations pertaining to a single-center experience and sample size may be addressed by further studies incorporating broader enrollments.

Efficacy of Intravitreal Multi-Characteristic Opsin (MCO-010) Optogenetic Gene Therapy in a Mouse Model of Leber Congenital Amaurosis.

Purpose: Leber congenital amaurosis (LCA) is a sight-threatening inherited retinal disorder (IRD) caused by numerous genetic mutations. Multi-characteristic opsin (MCO)-based optogenetic therapy allows the recruitment of residual cells of the retina in LCA for alternative vision transduction while being mutation-agnostic. Using rd12 mice, we investigated the invivo efficacy of an adeno-associated virus2 (AAV2)-transduced ambient light-activatable MCO (MCO-010) containing a metabotropic glutamate receptor-6 bipolar cell-specific promoter/enhancer. Methods: Mice requiring > 40 s to reach and board a dimly lit hidden platform in a water-maze were selected and randomly divided into 2 cohorts. These mice were intravitreally (IVT) injected with either 1.7E9 gene copies/eye of MCO-010 or control AAV2 and re-tested in the water-maze. Spectral-domain optical coherence tomography (SD-OCT), hematoxylin and eosin staining of retinas, and electroretinographic (ERG) studies were also conducted. Results: Safety of MCO-010 in rd12 mice was confirmed by the lack of significant detrimental changes in the mouse behavior, b-wave amplitudes and in retinal thickness. rd12 control mice performed relatively poorly in the water-maze test requiring ≥ 30-60 s to find and board the platform. MCO-010-treated rd12 mice reached the platform much faster than the AAV2-treated rd12 mice, with some mice only requiring < 5 s to achieve this goal (P < 0.01-0.0024). Conclusions: IVT MCO-010 treatment was well tolerated by rd12 mice, and it prevented the decrease in retinal thickness, and preserved ERG parameters. It also significantly improved the vision in rd12 mice relative to control AAV2-injected mice. MCO-010 therefore represents a novel and efficacious optogenetic therapeutic to treat LCA and other IRDs irrespective of the genetic defect(s).

Paroxetine Loaded Nanostructured Lipid Carriers Based In-situ Gel for Brain Delivery via Nasal Route for Enhanced Anti-Depressant Effect: In Vitro Prospect and In Vivo Efficacy.

This study focused on developing a thermosensitive gel with nanostructured lipid carriers (NLCs) loaded with paroxetine (PAR) to enhance the treatment and management of depression via nasal administration. Micro emulsion technique was utilized for the PAR-NLCs preparation. The acetyl alcohol and oleic acid were used in the ratio of 76:24. In the NLCs Tween 40, Span40 and Myrj 52 were used as a surfactant. The NLCs were then added into Poloxamer mixture to get thermosensitive NLCs based gel. Characterization, in vitro and in vivo studies were performed to check the efficiency of formulation in drug delivery. The entrapment efficiency of optimized PAR-NLCs was about 90%. The particle size, zeta potential and PDI were 155 ± 1.4nm, -25.9 ± 0.5mV, and 0.12 ± 0.01 respectively. The optimized gel showed a gelling temperature of 31.50 ± 0.50°C and a gelling time of 1 ± 0.12s with a pH of 6, suitable for nasal administration. The in vitro release assay of PAR-NLC-gel showed a cumulative release of about 59% in the first 6h after comparison with PAR-NLCs which showed almost 100%release. In vivo studies included forced swim test and tail suspension tests showed significant potential for treating depression when compared to PAR-NLCs. PAR-NLCs and NLCs based gel enhanced the tissue architecture and suppressed the expression of TNF-α in brain cortex from histological and immunohistochemical analysis. PAR- NLCs gel-based delivery system can prove to be an effective delivery system for brain targeting through nose for the better management of depression.

Immortelle Essential-Oil-Enriched Hydrogel for Diabetic Wound Repair: Development, Characterization, and In Vivo Efficacy Assessment

Background: Alarming data revealed that 19% to 34% of adults with diabetes mellitus develop chronic wounds, which are characterized by impaired healing and a higher risk of infections. Inspired by the traditional use of immortelle for wound healing and the lack of scientific evidence regarding how it thoroughly influences tissue regeneration, we aimed to formulate a hydrogel loaded with immortelle essential oil and assess its effectiveness on diabetic excision wounds. Methods: The rheological properties of the hydrogel, an in vivo safety test, as well as wound healing capacity, were determined in rats with induced diabetes and excision wounds. Diabetic rats were divided into four groups: untreated, treated with 1% silver sulfadiazine ointment, treated with a gel base, and treated with the immortelle essential oil-based hydrogel. Results: It was revealed that the hydrogel exerts pseudoplastic behavior and has no potential to act as an irritant, thus highlighting its suitability for skin application. Moreover, analysis of macroscopic, biochemical, and histopathological data revealed that the immortelle essential oil-based hydrogel significantly improves wound repair. Superior re-epithelialization, scar maturation, and increased collagen fiber density were achieved after immortelle essential oil-based gel application. Conclusions: These findings suggest that the immortelle essential oil-based hydrogel could be a natural, safe, and effective wound-healing dressing.

Pyrotinib and trastuzumab combination treatment synergistically overcomes HER2 dependency in HER2-positive breast cancer: insights from the PHILA trial

The PHILA study suggests that pyrotinib, trastuzumab, and docetaxel significantly improved progression-free survival (PFS) compared with placebo, trastuzumab, and docetaxel in patients with untreated HER2-positive metastatic breast cancer. In this study, we aimed to investigate the synergistic mechanisms of pyrotinib plus trastuzumab and provide further insights for the PHILA trial. The invitro activity of combination treatments was assessed through cell biological and biochemical experiments. The invivo efficacy was evaluated in cell-derived xenografts, a TUBO tumour model, and one clinical case. Next-generation sequencing was performed on circulating tumour DNA (ctDNA) from patients in the PHILA trial. The combination of pyrotinib and trastuzumab more effectively inhibited cell growth than pyrotinib or trastuzumab alone in models of HER2-dependent breast cancer. It potentiated membrane HER2 ubiquitination and downregulation, which resulted in a comprehensive blockade of the HER2 signalling pathway. The pyrotinib-altered membrane HER2 levels had no significant effect on trastuzumab-mediated antibody-dependent cell-mediated cytotoxicity (ADCC). We further validated the synergistic mechanisms in TUBO tumours and one clinical case, rather than models of HCC1954cells harbouring the PIK3CA H1047R mutation. Similarly, in our centre cohort of the PHILA study, patients with genetic alterations in the HER2 signalling cascade had significantly shorter median PFS than individuals with the wild-type pathway. Our findings underscore the robust synergy between pyrotinib and trastuzumab in overcoming HER2 dependency and provide a rationale for pyrotinib, trastuzumab, and docetaxel as one of the optimal choices for patients with untreated HER2-positive metastatic breast cancer, who are dependent on the HER2 signalling cascade. This work was supported by the National Key Research and Development Program of China (2021YFF1201300), the National Natural Science Foundation of China (82172875), the CAMS Innovation Fund for Medical Sciences (CIFMS) (2022-I2M-2-001), and the Joint Innovative Fund of Beijing Natural Science Foundation and Changping District (L234004).

Design, Synthesis, and Characterization of HBP-Vectorized Methotrexate Prodrug Molecule 1102-39: Evaluation of In Vitro Cytotoxicity Activity in Cell Culture Models, Preliminary In Vivo Safety and Efficacy Results in Rodents.

A novel bone-targeted prodrug, 1102-39, is discussed with the aim of enhancing the therapeutic effects of methotrexate (MTX) within bone tissues while minimizing systemic toxicity. Within the 1102-39 molecule, the central linker part forms a cleavable ester group, with MTX being also linked by a stable imine bond to the specially designed hydroxybisphosphonic (HBP) vector. Synthesized through a convergent approach starting from MTX, this prodrug advantageously modulates MTX's activity by selective esterification of its α-carboxyl group. In vitro tests revealed a 10-fold reduction in cytotoxicity compared to standard MTX, in alignment with prodrug behavior and correlated with gradual MTX release. In vivo in rodents, 1102-39 displayed preliminary encouraging antitumor effects on orthotopic osteosarcoma. Furthermore, various aspects of designing molecules for selective therapy in bone tissue based on bisphosphonate molecules as vectors for delivering active compounds to the bone are discussed. The 1102-39 molecule exhibits strong affinity for hydroxyapatite and a progressive release of MTX in aqueous environments, enhancing the safety and efficacy of bone-specific treatments and enabling sustained activity within bone and bone joints in the therapy of tumor and inflammation.

Targeting HLA-E-overexpressing cancers with a NKG2A/C switch receptor

Human leukocyte antigen (HLA)-E is overexpressed by a large proportion of solid tumors, including malignant glioblastoma, and acts as a major checkpoint for NKG2A+ CD8+ Tcells and natural killer (NK) cells in the tumor microenvironment and circulation. This axis operates alongside PD-L1 to inhibit effector responses by T and NK cells. We engineered a chimeric A/C switch receptor, combining the high HLA-E binding affinity of the NKG2A receptor ectodomain with the activating signaling of the NKG2C receptor endodomain. The cytotoxic function of A/C switch-transduced NK and Tcells was evaluated against tumor cells with varying levels of HLA-E expression. Invivo efficacy was assessed using a xenograft model of glioblastoma. A/C switch-transduced NK and Tcells exhibited superior and specific cytotoxicity against tumor cells with medium to high HLA-E expression. A/C switch-expressing human Tcells demonstrated enhanced anti-tumor function in a glioblastoma xenograft model. The activity of the modified Tcells was governed by an equilibrium between A/C switch levels and HLA-E expression, creating a therapeutic window to minimize on-target, off-tumor toxicities. Normal cells remained insensitive to A/C switch Tcells, even after interferon (IFN)-γ pretreatment to induce HLA-E expression. The A/C switch receptor effectively targets tumor cells expressing high levels of HLA-E, either alone or in combination with other engineered specificities, to overcome the suppressive NKG2A/HLA-E checkpoint. This approach offers a promising therapeutic strategy with a favorable safety profile for targeting HLA-E-overexpressing tumors. This work was funded by The Research Council of Norway, the Norwegian Cancer Society, and the National Cancer Institute.

Macrophage targeted nanocarriers containing methotrexate against arthritis

The present study aimed to develop macrophage-targeted nanocarriers containing methotrexate (MTX) against arthritis. To achieve this goal, mannosylated thiolated chitosan (MTCS) was synthesized and conjugation was confirmed by FTIR analysis. The synthesized MTCS was then used to develop nanocarriers by ionic gelation method utilizing chondroitin sulfate (ChS) as an oppositely charged polymer. The prepared nanocarriers were characterized in terms of size, zeta potential, SEM, and FTIR analysis. In vitro drug release, permeation enhancement, and macrophage targeting potential were investigated along with in-vivo efficacy studies in the arthritic rat model. The size of the macrophage targeted formulation chondroitin sulfate-methotrexate-mannosylated thiolated chitosan nanoparticles (ChS-MTX-MTCS-NPs) was found to be 379 ± 97.74 nm, with a zeta potential of 13 ± 8.76 mV. ChS-MTX-MTCS-NPs exhibited the entrapment efficacy of 78.60 ± 2.3 % with a cumulative drug release of 52 % after 60 h at pH 7.4. Florescent microscopy studies indicated the enhanced potential of ChS-MTX-MTCS-NPs to accumulate inside macrophages compared to non-targeted nanoformulations ChS-MTX-CS-NPs and ChS-MTX-TCS-NPs. Ex vivo intestinal permeation values for ChS- ChS-MTX-MTCS-NPs were measured to be 1.8 × 10-6 cm2/s. In vivo, studies in the arthritic rat model showed that the ChS-MTX-MTCS-NPs nanocarriers significantly reduced the diameter of the arthritic paw (P < 0.05), compared with MTX, ChS-MTX-CS-NPs and disease control. The results suggest that macrophage-targeted MTX-loaded nanocarriers based on MTCS are a promising approach for the treatment of arthritis.

Ultrapotent class I neutralizing antibodies post Omicron breakthrough infection overcome broad SARS-CoV-2 escape variants

The spread of emerging SARS-CoV-2 immune escape sublineages, especially JN.1 and KP.2, has resulted in new waves of COVID-19 globally. The evolving memory B cell responses elicited by the parental Omicron variants to subvariants with substantial antigenic drift remain incompletely investigated. Using the single B cell antibody cloning technology, we isolated single memory B cells, delineated the B cell receptor repertoire and conducted the pseudovirus-based assay for recovered neutralizing antibodies (NAb) screening. We analyzed the cryo-EM structures of top broadly NAbs (bnAbs) and evaluated their invivo efficacy (golden Syrian hamster model). By investigating the evolution of human B cell immunity, we discovered a new panel of bnAbs arising from vaccinees after Omicron BA.2/BA.5 breakthrough infections. Two lead bnAbs neutralized major Omicron subvariants including JN.1 and KP.2 with IC50 values less than 10ng/mL, representing ultrapotent receptor binding domain (RBD)-specific class I bnAbs. They belonged to the IGHV3-53/3-66 clonotypes instead of evolving from the pre-existing vaccine-induced IGHV1-58/IGKV3-20 bnAb ZCB11. Despite sequence diversity, they targeted previously unrecognized, highly conserved conformational epitopes in the receptor binding motif (RBM) for ultrapotent ACE2 blockade. The lead bnAb ZCP3B4 not only protected the lungs of hamsters intranasally challenged with BA.5.2, BQ.1.1 and XBB.1.5 but also prevented their contact transmission. Our findings demonstrated that class I bnAbs have evolved an ultrapotent mode of action protecting against highly transmissible and broad Omicron escape variants, and their epitopes are potential targets for novel bnAbs and vaccine development. A full list of funding bodies that contributed to this study can be found in the Acknowledgements section.

Anti-tissue factor antibody conjugated with monomethyl auristatin E or deruxtecan in pancreatic cancer models.

Antibody-drug conjugates (ADCs) have been recognized as a promising class of cancer therapeutics. Tissue factor (TF), an initiator of the blood coagulation pathway, has been investigated regarding its relationship with cancer, and several preclinical and clinical studies have presented data on anti-TF ADCs, including tisotumab vedotin, which was approved in 2021. However, the feasibility of other payloads in the design of anti-TF ADCs is still unclear because no reports have compared payloads with different cytotoxic mechanisms. For ADCs targeting other antigens, such as Her2, optimizing the payload is also an important issue in order to improve invivo efficacy. In this study, we prepared humanized anti-TF Ab (clone.1084) conjugated with monomethyl auristatin E (MMAE) or deruxtecan (DXd), and evaluated the efficacy in several cell line- and patient-derived xenograft models of pancreatic cancer. As a result, optimizing the drug / Ab ratio was necessary for each payload in order to prevent pharmacokinetic deterioration and maximize delivery efficiency. In addition, MMAE-conjugated anti-TF ADC showed higher antitumor effects in tumors with strong and homogeneous TF expression, while DXd-conjugated anti-TF ADC was more effective in tumors with weak and heterogeneous TF expression. Analysis of a pancreatic cancer tissue array showed weak and heterogeneous TF expression in most TF-positive specimens, indicating that the response rate to pancreatic cancer might be higher for DXd- than MMAE-conjugated anti-TF ADC. Nevertheless, our findings indicated that optimizing the ADC payloads individually in each patient could maximize the potential of ADC therapeutics.

STING Agonist Induced Innate Immune Responses Drive Anti-Respiratory Virus Activity In Vitro with Limited Antiviral Efficacy In Vivo.

The emergence of SARS-CoV-2 and seasonal outbreaks of other respiratory viruses highlight the urgent need for broad-spectrum antivirals to treat respiratory tract infections. Stimulator of interferon genes (STING) is a key component of innate immune signaling and plays a critical role in protection of the host against viral infections. Previously the STING agonist diABZI-4, a diamidobenzimidazole-based compound, demonstrated protection against SARS-CoV-2 both in vitro and in vivo. However, its broad-spectrum antiviral activity against other respiratory viruses in human airway epithelial cells, which are the primary targets of these infections, is not well established. In this study, we demonstrated that diABZI-4 stimulated robust innate immune responses protecting lung cells against a wide range of respiratory viruses, including influenza A virus (IAV), common cold coronaviruses, SARS-CoV-2, human rhinovirus (HRV), and human parainfluenza virus. diABZI-4 was highly active in physiologically relevant human airway epithelial tissues grown at the air-liquid interface, blocking replication of IAV, SARS-CoV-2, and HRV in these tissues. Furthermore, treatment of macrophages with diABZI-4 resulted in the secretion of cytokines that protected the primary airway epithelial cells from IAV infection. Despite the promising in vitro pan-antiviral activity, intranasal administration of diABZI-4 in mice provided early, but not sustained, inhibition of IAV replication in the lungs. These data highlight the complexities of the relationship between timing of STING agonist-driven inflammatory responses and viral replication dynamics, emphasizing the development challenge posed by STING agonists as potential therapeutics against respiratory viruses.

Tanomastat exerts multi-targeted inhibitory effects on viral capsid dissociation and RNA replication in human enteroviruses

Global cyclical outbreaks of human enterovirus infections has positioned human enterovirus A71 (EV-A71) as a neurotropic virus of clinical importance. However, there remains a scarcity of internationally approved antivirals and vaccines. In pursuit of repurposing drugs for combating human enteroviruses, we employed a comprehensive pharmacophore- and molecular docking-based virtual screen targeting EV-A71 capsid protein VP1-4, 3C protease, and 3D polymerase proteins. Among 15 shortlisted ligand candidates, we dissected the inhibitory mechanism of Tanomastat in cell-based studies and evaluated its invivo efficacy in an EV-A71-infected murine model. We demonstrated that Tanomastat exerts dose-dependent inhibition on EV-A71 replication, with comparable efficacy profiles in enterovirus species A, B, C, and D invitro. Time-course studies suggested that Tanomastat predominantly disrupts early process(es) of the EV-A71 replication cycle. Mechanistically, live virus particle tracking and docking predictions revealed that Tanomastat specifically impedes viral capsid dissociation, potentially via VP1 hydrophobic pocket binding. Bypassing its inhibition on entry stages, we utilized EV-A71 replication-competent, 3Dpol replication-defective, and bicistronic IRES reporter replicons to show that Tanomastat also inhibits viral RNA replication, but not viral IRES translation. We further showed that orally administered Tanomastat achieved 85% protective therapeutic effect and alleviated clinical symptoms in EV-A71-infected neonatal mice. Our study establishes Tanomastat as a broad-spectrum anti-enterovirus candidate with promising pre-clinical efficacy, warranting further testing for potential therapeutic application. MOE Tier 2 grants (MOE-T2EP30221-0005, R571-000-068-592, R571-000-076-515, R571-000-074-733) and A∗STARBiomedical Research Council (BMRC).

50717 Hydrolyzed Collagen and Cacay Oil Increase Collagen, Elastin, and Cell Turnover: In-Vivo and Ex-Vivo Efficacy of a Novel Retinol-Alternative Body Cream

50717 Hydrolyzed Collagen and Cacay Oil Increase Collagen, Elastin, and Cell Turnover: In-Vivo and Ex-Vivo Efficacy of a Novel Retinol-Alternative Body Cream

In Vivo Efficacy of Rezafungin, Anidulafungin, Caspofungin, and Micafungin against Four Candida auris Clades in a Neutropenic Mouse Bloodstream Infection Model.

Rezafungin is the first new drug approved to treat candidaemia and invasive candidiasis in more than 10 years. However, data are scant on the in vivo efficacy of rezafungin and the other three approved echinocandins against different Candida auris clades. This study involved 10 isolates representing 4 C. auris clades: South Asian (n = 2), East Asian (n = 2), South African (n = 2), and South American (n = 4, including 2 environmental isolates). In the lethality experiment and fungal tissue burden experiment (kidney, heart, and brain), cyclophosphamide-treated BALB/c male mice were intravenously infected (107 and 8 × 106 colony-forming units [CFU]/mouse, respectively). A 20 mg/kg dose of rezafungin was administered on days 1, 3, and 6. Alternatively, beginning 24 h post-infection, mice received 3 mg/kg of caspofungin, 5 mg/kg of micafungin, or 5 mg/kg of anidulafungin once daily for 6 days. Regardless of isolate and clade, all echinocandin regimens improved survival after 21 days (p = 0.0041 to p < 0.0001). All echinocandins frequently produced >3-log mean CFU/g decreases in the fungal kidney and heart burdens, although some of these decreases were not statistically significant. Rezafungin, regardless of clade, produced 3-5 and 2-4 log CFU/g decreases in the kidney and heart burdens, respectively. Echinocandins did not inhibit fungal growth in the brain. Histopathological examination performed on day 7 showed no fungal cells in the heart and kidneys of rezafungin-treated mice and to a lesser extent, caspofungin-treated mice, regardless of the clinical isolate. All echinocandin-treated mice showed medium and/or large foci of fungal cells in their cerebrum or cerebellum. Regardless of the C. auris clade, rezafungin activity in vivo was comparable to or improved over that of the three previously approved echinocandins.

Exploring nalbuphine loaded chitosan nanoparticles for effective pain management through intranasal administration: a comparative study

Background Intranasal drug delivery shows potential for brain access via olfactory and trigeminal routes. Purpose This work aimed to ensure brain availability of nalbuphine via the nasal route. Method Chitosan based nanoparticles loaded with nalbuphine were successfully prepared using ionic gelation method and characterised. Result SEM results revealed that the nanoparticles were spherical in shape, with an average size of 192.4 ± 11.6 nm. Zeta potential and entrapment efficiency was found 32.8 mV and 88.43 ± 7.75%, respectively. The X-ray diffractometry and DSC results unravel a profound understanding on the physical and thermal characteristics. The in-vitro release of nalbuphine from the nanoparticles was biphasic, with an initial burst release followed by a slow-release profile. In-vitro cell study on HEK-293 cells and microscopic images of brain tissue confirmed the safety profile of formulation. In-vivo efficacy studies on animal confirmed the effectiveness of developed intranasal formulation as compared to the standard therapy. The in-vivo pharmacokinetic studies showed that the prepared nanoparticles were able to efficiently deliver nalbuphine to the brain in comparison to the other body organs. Gamma scintigraphy images showed retention of the drug in the brain. Furthermore, the efficacy studies confirmed that the nanoparticles were found significantly more effective than the marketed formulation in pain management.

Orthoflaviviral Inhibitors in Clinical Trials, Preclinical In Vivo Efficacy Targeting NS2B-NS3 and Cellular Antiviral Activity via Competitive Protease Inhibition.

Orthoflaviviruses, including zika (ZIKV), West Nile (WNV), and dengue (DENV) virus, induce severely debilitating infections and contribute significantly to the global disease burden, yet no clinically approved antiviral treatments exist. This review offers a comprehensive analysis of small-molecule drug development targeting orthoflaviviral infections, with a focus on NS2B-NS3 inhibition. We systematically examined clinical trials, preclinical efficacy studies, and modes of action for various viral replication inhibitors, emphasizing allosteric and orthosteric drugs inhibiting NS2B-NS3 protease with in vivo efficacy and in vitro-tested competitive NS2B-NS3 inhibitors with cellular efficacy. Our findings revealed that several compounds with in vivo preclinical efficacy failed to show clinical antiviral efficacy. NS3-NS4B inhibitors, such as JNJ-64281802 and EYU688, show promise, recently entering clinical trials, underscoring the importance of developing novel viral replication inhibitors targeting viral machinery. To date, the only NS2B-NS3 inhibitor that has undergone clinical trials is doxycycline, however, its mechanism of action and clinical efficacy as viral growth inhibitor require additional investigation. SYC-1307, an allosteric inhibitor, exhibits high in vivo efficacy, while temoporfin and methylene blue represent promising orthosteric non-competitive inhibitors. Compound 71, a competitive NS2B-NS3 inhibitor, emerges as a leading preclinical candidate due to its high cellular antiviral efficacy, minimal cytotoxicity, and favorable in vitro pharmacokinetic parameters. Challenges remain in developing competitive NS2B-NS3 inhibitors, including appropriate biochemical inhibition assays as well as the selectivity and conformational flexibility of the protease, complicating effective antiviral treatment design.