Glycol Chitosan-Poly(lactic acid) Conjugate Nanoparticles Encapsulating Ciprofloxacin: A Mucoadhesive, Antiquorum-Sensing, and Biofilm-Disrupting Treatment Modality for Bacterial Keratitis.

Bacterial keratitis (BK) causes visual morbidity/blindness if not treated effectively. Here, ciprofloxacin (CIP)-loaded nanoparticles (NPs) using glycol chitosan (GC) and poly(lactic acid) (PLA) conjugate at three different ratios (CIP@GC(PLA) NPs (1:1,5,15)) were fabricated. CIP@GC(PLA) NPs (1:1) were more effective than other tested ratios, indicating the importance of optimal hydrophobic/hydrophilic balance for corneal penetration and preventing bacterial invasion. The CIP@GC(PLA) (NPs) (1:1) realized the highest association with human corneal epithelial cells, which were nonirritant to the hen's egg-chorioallantoic membrane test (HET-CAM test) and demonstrated significant antibacterial response in the in vitro minimum inhibitory, bactericidal, live-dead cells, zone of inhibition, and biofilm inhibition assays against the keratitis-inducing pathogen Pseudomonas aeruginosa. The antiquorum sensing activity of GC has been explored for the first time. The NPs disrupted the bacterial quorum sensing by inhibiting the production of virulence factors, including acyl homoserine lactones, pyocyanin, and motility, and caused significant downregulation of quorum sensing associated genes. In the in vivo studies, CIP@GC(PLA) NPs (1:1) displayed ocular retention in vivo (∼6 h) and decreased the opacity and the bacterial load effectively. Overall, the CIP@GC(PLA) NP (1:1) is a biofilm-disrupting antiquorum sensing treatment regimen with clinical translation potential in BK.

Safety and efficacy of McCarey-Kaufman medium supplemented with colistin (polymyxin E) and amphotericin B in inhibiting the multidrug-resistant Pseudomonas aeruginosa using an ex vivo donor corneal infection model.

PURPOSE: This study aimed to evaluate the efficacy and safety of McCarey-Kaufman (MK) medium supplemented with colistin and amphotericin B in inhibiting the growth of multidrug-resistant Pseudomonas (P.) aeruginosa , using an ex vivo experimental model with human donor corneas. METHODS: Cadaveric human corneas deemed unsuitable for corneal transplantation were obtained, and MK media were supplemented with colistin and amphotericin B. Multidrug-resistant P. aeruginosa was cultured and used to infect the human donor corneas ex vivo . Infected corneas were placed in the MK media with additional antibiotics (colistin and amphotericin B) and the standard MK media, which served as the control arm for comparison. Corneal opacity due to infiltration and quantitative analysis of colony-forming units (CFUs) were assessed. The viability of the corneal endothelium was assessed using trypan blue staining. RESULTS: Corneas incubated in MK media supplemented with additional antibiotics showed less corneal opacification compared with those in standard MK media at both 48- and 96-hour (hr) time points. Quantitative analysis revealed a lower bacterial load and a significant reduction in CFU in the corneas incubated in MK media with additional antibiotics compared with the control group. At 48 hrs, there was 84% ( P value = 0.024) reduction in bacterial load, and at 96 hr, a 53% ( P value = 0.016) reduction was observed in comparison with those placed in standard MK media. The trypan blue staining tests revealed that the extent of endothelial cell loss in corneas incubated in supplemented MK media was comparable to the ones in standard MK media. CONCLUSION: The addition of colistin and amphotericin B to MK media demonstrated efficacy in inhibiting the growth of multidrug-resistant P. aeruginosa in an ex vivo cornea infection model. The supplemented media had no detrimental effect on the corneal endothelium. The findings suggest that supplementing the MK media with these broad-spectrum antimicrobial agents may help mitigate the risk of postoperative donor-related infection in the recipients by reducing and containing the load of microbial contamination in donor corneas.

Host antimicrobial peptide S100A12 disrupts the fungal membrane by direct binding and inhibits growth and biofilm formation of Fusarium species.

Fungal keratitis is the foremost cause of corneal infections worldwide, of which Fusariumspp. is the common etiological agent that causes loss of vision and warrants surgical intervention. An increase in resistance to the available drugs along with severe side effects of the existing antifungals demands for new effective antimycotics. Here, we demonstrate that antimicrobial peptide S100A12 directly binds to the phospholipids of the fungal membrane, disrupts the structural integrity, and induces generation of reactive oxygen species in fungus. In addition, it inhibits biofilm formation by Fusariumspp. and exhibits antifungal property against Fusariumspp. both in vitro and in vivo. Taken together, our results delve into specific effect of S100A12 against Fusariumspp. with an aim to investigate new antifungal compounds to combat fungal keratitis.

3D-Printed Inherently Antibacterial Contact Lens-Like Patches Carrying Antimicrobial Peptide Payload for Treating Bacterial Keratitis.

Delivery of therapeutic agents through contact lenses-like patches is a promising strategy to achieve significant bioavailability with negligible eye drainage. The present study investigates the preparation and 3D printing of mucoadhesive gelatin methacryloyl (GelMA)/chitosan methacryloyl (ChiMA) hydrogels to fabricate them as contact lens-like patches (CLP) loaded with antimicrobial peptide, S100A12 (AMP) for treating bacterial keratitis (BK). Extrusion technology is used to print the patches layer by layer to form a hemispherical scaffold suitable for eyewear, and 3D-printed CLP is crosslinked using Irgacure 2959 under UV light. The results from the in vivo experiment conducted on Pseudomonas aeruginosa-infected BK rabbit model after the treatment with AMP-loaded CLP have shown a significant decrease in bacterial load when plated for CFU. The newly developed delivery system containing AMP has great potential to overcome the treatment challenges of multidrug resistance (MDR) in bacteria and eliminate the frequent dosing associated with eye drops. The presence of chitosan in the formulation provides a synergetic effect on the AMP in disrupting bacterial biofilms. The ease of using 3D printing will open new avenues for optimizing the dosage depending on the severity of the BK in the patients, which can be used as personalized medicine.

Anti-fungal efficacy of combination of amphotericin B with colistin and gentamicin in McCarey-Kaufman corneal preservation medium.

Purpose: To curtail the potential of donor corneal tissue disseminating fungi to the recipient's eye, we evaluated the addition of amphotericin B to McCarey-Kaufman (M-K)-corneal storage medium supplemented with colistin. Methods: Amphotericin B was examined for its ability to inhibit the growth of Candida albicans and Aspergillus flavus using a microbroth dilution test and checkerboard assay in combination with only gentamicin and a combination of colistin, gentamicin, and amphotericin B. The safety on epithelium and endothelium was evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Results: The minimal inhibitory concentration of gentamicin was found to be >256 µg/ml against both C. albicans and A. flavus, whereas that of amphotericin B was found to be in a range of 0.25-0.5 and 1-2 µg/ml for C. albicans and A. flavus, respectively. According to the checkerboard assay, 80% (4/5) of C. albicans isolates and 100% (5/5) of A. flavus isolates responded synergistically to the combination of amphotericin B and gentamicin, but only 20% (1/5) of C. albicans isolates showed an additive effect. None of the tested isolates displayed antagonism. The combined effect of the three drugs also did not display any antagonistic effect. Additionally, the MTT assay reveals no toxic effect of the antimicrobials used on corneal epithelial and endothelial cells. Conclusion: In vitro experiments demonstrate that amphotericin B is not toxic to either epithelium or endothelium and is a promising additive to the M-K medium supplemented with colistin.

Successful Management of Extensively Drug Resistant Pseudomonas aeruginosa-Infectious Scleritis after Pterygium Surgery.

INTRODUCTION AND PURPOSE: We report a case of extensively drug-resistant (XDR) Pseudomonas aeruginosa (PA) scleritis after pterygium surgery. METHODS: Case report. RESULTS: A 58-year-old farmer presented with a 40-day history of severe pain, swelling, and blurred vision after a pterygium excision was performed at another institute. The patient was on multiple medications with no relief. The examination showed a nasally located scleral thinning in his right eye, with ulceration and infiltrates. Microbiology revealed Pseudomonas aeruginosa, which showed intermediate sensitivity to colistin only. The patient was administered topical (0.19%) and intravenous colistin and dexamethasone. There was a rapid reduction in symptoms, and the lesions healed over the next 2 months. CONCLUSIONS: To the best of our knowledge, this is the first case report of XDR-PA scleritis. We suggest the possibility of evolving drug resistance caused by the iatrogenic use of antibiotics during the early stages of the disease course.

Tetraspanin CD9-derived peptides inhibit Pseudomonas aeruginosa corneal infection and aid in wound healing of corneal epithelial cells.

Pseudomonas aeruginosa is a leading cause of corneal infection both within India and globally, often causing a loss of vision. Increasing antimicrobial resistance among the bacteria is making its treatment more difficult. Preventing initial bacterial adherence to the host membrane has been explored here to reduce infection of the cornea. Synthetic peptides derived from human tetraspanin CD9 have been shown to reduce infection in corneal cells both in vitro, ex vivo and in vivo. We found constitutive expression of CD9 in immortalized human corneal epithelial cells by flow cytometry and immunocytochemistry. The synthetic peptides derived from CD9 significantly reduced bacterial adherence to cultured corneal epithelial cells and ex vivo human cadaveric corneas as determined by colony forming units. The peptides also significantly reduced bacterial burden in a murine model of Pseudomonas keratitis and lowered the cellular infiltration in the corneal stroma. Additionally, the peptides aided corneal wound healing in uninfected C57BL/6 mice compared to control mice. These potential therapeutics had no effect on cell viability or proliferation of corneal epithelial cells and have the potential to be developed as an alternative therapeutic intervention.

Chitosan-poly(lactide-co-glycolide)/poloxamer mixed micelles as a mucoadhesive thermo-responsive moxifloxacin eye drop to improve treatment efficacy in bacterial keratitis.

A mucoadhesive self-assembling polymeric system was developed to carry moxifloxacin (M) for treating bacterial keratitis (BK). Chitosan-PLGA (C) conjugate was synthesized, and poloxamers (F68/127) were mixed in different proportions (1: 5/10) to prepare moxifloxacin (M)-encapsulated mixed micelles (M@CF68/127(5/10)Ms), including M@CF68(5)Ms, M@CF68(10)Ms, M@CF127(5)Ms, and M@CF127(10)Ms. The corneal penetration and mucoadhesiveness were determined biochemically, in vitro using human corneal epithelial (HCE) cells in monolayers and spheroids, ex vivo using goat cornea, and in vivo via live-animal imaging. The antibacterial efficacy was studied on planktonic biofilms of P. aeruginosa and S. aureus (in vitro) and Bk-induced mice (in vivo). Both M@CF68(10)Ms and M@CF127(10)Ms demonstrated high cellular uptake, corneal retention, muco-adhesiveness, and antibacterial effect, with M@CF127(10)Ms exhibiting superior therapeutic effects in P. aeruginosa and S. aureus-infected BK mouse model by reducing the corneal bacterial load and preventing corneal damage. Therefore, the newly developed nanomedicine is promising for clinical translation in treating BK.

Updates on congenital hereditary endothelial dystrophy.

Congenital hereditary endothelial dystrophy (CHED) is a rare genetic corneal disorder causing progressive cornea clouding and significant visual impairment. CHED remains a leading indication for pediatric corneal transplantation despite its infrequency, particularly in regions with high consanguinity rates like Southeast Asia. Identifying the Solute Carrier Family 4 Member 11 (SLC4A11) gene as the genetic basis of CHED has led to the discovery of it's various genetic variations. However, a comprehensive understanding of its clinical-genetic correlation, pathophysiology, and optimal management is ongoing. This review aims to consolidate current knowledge about CHED, covering its genetic origins, pathophysiological mechanisms, clinical presentation, and management strategies. Surgical intervention, such as penetrating keratoplasty (PK), Descemet stripping automated endothelial keratoplasty (DSAEK), and Descemet membrane endothelial keratoplasty (DMEK), remains the primary treatment. DSAEK and DMEK offer advantages over PK, including quicker visual recovery, reduced complications, and longer graft survival, especially in the pediatric age group. The timing of surgical interventions depends on disease severity, age at presentation, comorbidities, and visual potential. Elevated oxidative stress in CHED corneal tissue suggests potential benefits from anti-inflammatory drugs to rescue mutated endothelial cells. Considering the limitations of corneal graft surgeries, exploring novel gene-based molecular therapies are essential for future management. Early diagnosis, appropriate surgical interventions, amblyopia control, and genetic counseling for predictive analysis are pivotal for optimizing CHED management. A multidisciplinary approach involving ophthalmologists, researchers, and genetic counselors is essential for precise diagnosis and optimal care for CHED patients.

Purification, characterization of an entomopathogenic fungal lectin from Purpureocillium lilacinum and its involvement in pathogenesis leading to mycotic keratitis.

A lectin PCL, from Purpureocillium lilacinum a saprophytic, filamentous fungus was purified from the crude extract of the mycelia using 70% ammonium sulphate precipitation followed by affinity chromatography on mucin-Sepharose 4 B column. PCL is a monomer with an apparent molecular mass of 18.5 kDa as revealed by SDS-PAGE under both reducing and non-reducing conditions. PCL is a blood group non-specific lectin and has highest affinity towards chitin, mucin, asialomucin, fetuin with a MIC of 0.15 µg/mL and also recognizes L-fucose, galactose, lactose, N-acetyl galactosamine, hyaluronic acid. PCL is stable up to 60 °C and within the pH range 4-8. To understand its role in pathogenesis, effect of PCL was evaluated on human corneal epithelial cells (HCECs). PCL showed strong glycan mediated binding to HCECs and PCL showed proinflammatory response at lower concentrations by stimulating secretion of IL-6, 8. In contrast PCL at higher concentrations revealed opposite effect of HCECs growth inhibition. All these results collectively support the involvement of PCL in mediating host pathogen interactions possibly leading to pathogenesis. In addition, considering the entomopathogenic effect of Purpureocillium lilacinum, PCL may be attributed for this beneficiary effect, which needs to be explored.

Oil-free eye drops containing Cyclosporine A/cyclodextrin/PVA supramolecular complex as a treatment modality for dry eye disease.

According to the global mapping of dry eye disease (DED), nearly 5 to 50 % of people suffer from DED, and this number is on the rise. The drug of choice Cyclosporine A (CsA) exhibits poor ocular bioavailability due to high molecular weight and lipophilicity. Moreover, formulations of CsA currently available are in the form of oil-based emulsions that are known to cause ocular irritation and pain. In this study, sulfobutylether-ß-cyclodextrin (SBE-ß-CD) based binary and ternary supramolecular complexes of CsA were developed as completely oil-free, and particle-free eye drops to treat DED. The physicochemical characterizations were supplemented with relevant in silico studies, to ascertain the findings. Further, the efficacy of the complexes was evaluated in the scopolamine-induced mouse model of DED. The complexation improved the CsA solubility by ~21-fold, with ~4-fold improvement in dissolution and transcorneal permeation. The non-irritancy and non-toxicity were confirmed by hen's egg chorioallantoic membrane assay and cytotoxicity assay using human corneal epithelial cells, respectively. The in vivo treatment with the ternary CD complex demonstrated better management of the dry eye supported by the tear volume assessment, corneal fluorescein staining, and histopathological studies of the cornea, lacrimal gland, and harderian gland. The study demonstrates the potential of the supramolecular complex as an alternative to the oil-based formulation of eye drops for drugs that show low solubility and poor corneal permeation.

An experimental study to evaluate the effect of polymixin E (Colistin) alone or in combination with gentamicin in McCarey-Kaufman corneal preservation medium on various drug resistant bacterial and fungal isolates.

Purpose: To assess the efficacy of the addition of polymyxin E (colistin) in the McCarey-Kaufman (MK) corneal storage solution against multi-drug resistant strains of Enterobacteriaceae, Staphylococcus aureus, and Candida spp. Methods: A standard micro broth dilution test and a checkerboard assay were performed for five multi-drug resistant (MDR) clinical strains of P. aeruginosa and five clinical strains of methicillin-resistant S. aureus (MRSA) and C. albicans against colistin and gentamicin alone and in combination. The minimum inhibitory concentration (MIC) and the fractional inhibitory concentration index (FICI) were calculated to assess the efficacy of each combination. Results: The MIC of colistin was in the range of 1-2 µg/mL for P. aeruginosa, whereas it was 256-1024 µg/mL against S. aureus. In comparison, the MIC of gentamicin was found to be 0.5-512 µg/mL and 0.5-8 µg/mL against P. aeruginosa and S. aureus, respectively. All five isolates of C. albicans did not exhibit any susceptibility to either colistin or gentamicin even at a concentration of ≥ 512 µg/mL each. The checkerboard assay was performed to evaluate the nature of the interaction of the combination of colistin and gentamicin. Based on the FICI, it was observed that the colistin and gentamicin combination has a maximum synergistic effect (FIC <0.5) in 80% (4/5) for S. aureus isolates, whereas the maximum additive effect (FIC >0.5-4) was 100% (5/5) for P. aeruginosa and the minimum additive effect was 20% (1/5) for S. aureus isolates. Antagonism (FIC ≥ 4) was not observed in any combination between the strains used in the study. Both colistin and gentamicin alone or in combination were, however, ineffective against Candida spp. Conclusion: The addition of colistin has an inhibitory effect on bacterial contamination that could be possibly caused by MDR strains and could potentially be considered as an additional additive in corneal storage media.

Structural insights into the interaction of antifungal peptides and ergosterol containing fungal membrane.

The treatment of invasive drug-resistant and potentially life-threatening fungal infections is limited to few therapeutic options that are usually associated with severe side effects. The development of new effective antimycotics with a more tolerable side effect profile is therefore of utmost clinical importance. Here, we used a combination of complementary in vitro assays and structural analytical methods to analyze the interaction of the de novo antimicrobial peptide VG16KRKP with the sterol moieties of biological cell membranes. We demonstrate that VG16KRKP disturbs the structural integrity of fungal membranes both invitro and in model membrane system containing ergosterol along with phosphatidylethanolamine lipid and exhibits broad-spectrum antifungal activity. As revealed by systematic structure-function analysis of mutated VG16KRKP analogs, a specific pattern of basic and hydrophobic amino acid side chains in the primary peptide sequence determines the selectivity of VG16KRKP for fungal specific membranes.

Antimicrobial peptide S100A12 (calgranulin C) inhibits growth, biofilm formation, pyoverdine secretion and suppresses type VI secretion system in Pseudomonas aeruginosa.

Pseudomonas aeruginosa is an opportunistic pathogen and is the major cause of corneal infections in India and worldwide. The increase in antimicrobial resistance among Pseudomonas has prompted rise in significant research to develop alternative therapeutics. Antimicrobial peptides (AMPs) are considered as potent alternatives to combat bacterial infections. In this study, we investigated the role of S100A12, a host defense peptide, against PAO1 and an ocular clinical isolate. Increased expression of S100A12 was observed in corneal tissues obtained from Pseudomonas keratitis patients by immunohistochemistry. S100A12 significantly inhibited growth of Pseudomonas in vitro as determined from colony forming units. Furthermore, recombinant S100A12 reduced the corneal opacity and the bacterial load in a mouse model of Pseudomonas keratitis. Transcriptome changes in PAO1 in response to S100A12 was investigated using RNA sequencing. The pathway analysis of transcriptome data revealed that S100A12 inhibits expression of genes involved in pyoverdine synthesis and biofilm formation. It also impedes several important pathways like redox, pyocyanin synthesis and type 6 secretion system (T6SS). The transcriptome data was further validated by checking the expression of several affected genes by quantitative PCR. Our study sheds light on how S100A12 impacts Pseudomonas and that it might have the potential to be used as therapeutic intervention in addition to antibiotics to combat infection in future.

A rationally designed synthetic antimicrobial peptide against Pseudomonas-associated corneal keratitis: Structure-function correlation.

Contact lens wearers are at an increased risk of developing Pseudomonas-associated corneal keratitis, which can lead to a host of serious ocular complications. Despite the use of topical antibiotics, ocular infections remain a major clinical problem, and a strategy to avoid Pseudomonas-associated microbial keratitis is urgently required. The hybrid peptide VR18 (VARGWGRKCPLFGKNKSR) was designed to have enhanced antimicrobial properties in the fight against Pseudomonas-induced microbial keratitis, including contact lens-related keratitis. In this paper, VR18's modes of action against Pseudomonas membranes were shown by live cell Raman spectroscopy, live cell NMR, live-cell fluorescence microscopy and measures taken using sparsely tethered bilayer lipid membrane bacterial models to be via a bacterial-specific membrane disruption mechanism. The high affinity and selectivity of the peptide were then demonstrated using in vivo, in vitro and ex vivo models of Pseudomonas infection. The extensive data presented in this work suggests that topical employment of the VR18 peptide would be a potent therapeutic agent for the prevention or remedy of Pseudomonas-associated microbial keratitis.

Streptococcus pneumoniae exerts oxidative stress, subverts antioxidant signaling and autophagy in human corneal epithelial cells that is alleviated by tert-Butylhydroquinone.

Streptococcus pneumoniae is one of the leading causes of bacterial keratitis in the developing world and globally. In the current study, we have determined oxidative stress as pathogenesis of S. pneumoniae infection in corneal tissues and human corneal epithelial cells (HCEC) and explored host immune response of HCEC towards S. pneumoniae. We also determined whether treatment with tert-Butylhydroquinone (tBHQ), a Nrf2 inducer, could alleviate oxidative stress and reduce bacterial cytotoxicity in these cells. Oxidative stress was determined in corneal tissues of patients and HCEC by immunohistochemistry and immunofluorescence analysis, respectively. The expression of antioxidant genes, cytokines and antimicrobial peptides was determined by quantitative PCR. Infection of HCEC by S. pneumoniae was determined by colony-forming units. The autophagy and cell death were determined by fluorescence microscopy. The phosphorylation of signaling proteins was evaluated by immunoblot analysis. S. pneumoniae induced oxidative stress during corneal infections and inhibited antioxidant signaling pathways and immune responses like autophagy. tBHQ aided in restoring Nrf2 activation, reduced reactive oxygen species generation and prevented cytotoxicity and cell death in S. pneumoniae-infected HCEC. tBHQ also induced autophagy in a Nrf2-dependent manner and reduced bacterial survival in HCEC. Increased expression of antimicrobial peptides by tBHQ might have contributed to a reduction of bacterial load and cytotoxicity, as exemplified in LL-37 depleted corneal epithelial cells exposed to S. pneumoniae compared to control siRNA-transfected cells. tBHQ mediates alleviation of oxidative stress induced by S. pneumoniae by activating Nrf2-mediated antioxidant signaling in corneal epithelial cells. tBHQ also enhances expression of antimicrobial peptides in corneal cells and aids in inhibition of bacterial survival and cytotoxicity of HCEC.

Voriconazole-Cyclodextrin Supramolecular Ternary Complex-Loaded Ocular Films for Management of Fungal Keratitis.

Fungal keratitis is one of the leading causes of ophthalmic mycosis affecting the vision due to corneal scarring. Voriconazole (VRC) is the most preferred azole antifungal agent for treating ocular mycotic infections. Ocular drug delivery is challenging due to the shorter corneal residence time of the formulation requiring frequent administration, leading to poor patient compliance. The present study aimed at improving the solubility, transcorneal permeation, and efficacy of voriconazole via the formation of cyclodextrin-based ternary complexes and incorporation of the complex into mucoadhesive films. A phase solubility study suggested a ∼14-fold improvement in VRC solubility, whereas physicochemical characterization confirmed the inclusion of VRC in the cyclodextrin inner cavity. In silico docking studies were performed to predict the docking conformation and stability of the inclusion complex. Complex-loaded films showed sustained release of voriconazole from the films and improved transcorneal permeation by ∼4-fold with an improved flux of 8.36 µg/(cm2 h) for ternary complex-loaded films compared to 1.86 µg/(cm2 h) for the pure VRC film. The 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) and hen's egg-chorioallantoic membrane test (HET-CAM) assays confirmed that the complexes and ocular films were nonirritant and safe for ocular administration. The antifungal study performed using Aspergillus fumigatus and Fusarium oxysporum suggested improved antifungal activity compared to the pure drug film. In conclusion, the supramolecular cyclodextrin ternary complex proved to be a promising strategy for enhancing the solubility and permeability and augmenting the antifungal activity of voriconazole in the management of fungal keratitis.

A supramolecular thermosensitive gel of ketoconazole for ocular applications: In silico, in vitro, and ex vivo studies.

The incidence of corneal fungal infections continues to be a growing concern worldwide. Ocular delivery of anti-fungal drugs is challenging due to the anatomical and physiological barriers of the eye. The ocular bioavailability of ketoconazole (KTZ), a widely prescribed antifungal agent, is hampered by its limited aqueous solubility and permeation. In the study, the physicochemical properties of KTZ were improved by complexation with sulfobutylether-ß-cyclodextrin (SBE-ß-CD).KTZ-SBE-ß-CD complex was studied in silico with docking and dynamics simulations, followed by wet-lab experiments.The optimized KTZ-SBE-ß-CD complex was loaded into a thermosensitivein situ gel to increase corneal bioavailability. The supramolecular complex increased the solubility of KTZ by 5-folds and exhibited a 10-fold increment in drug release compared to the pure KTZ. Owing to the diffusion, thein situ gel exhibited a more sustained drug release profile. Theex vivocorneal permeation studies showed higher permeation from KTZ-SBE-ß-CD in situ gel (flux of ∼19.11 µg/cm2/h) than KTZin situ gel (flux of ∼1.17 µg/cm2/h). The cytotoxicity assays and the hen's egg chorioallantoic membrane assay (HET-CAM) confirmed the formulations' safety and non-irritancy. In silico guided design of KTZ-SBE-ß-CD inclusion complexes successfully modified the physicochemical properties of KTZ. In addition, the loading of the KTZ-SBE-ß-CD complex into an in situ gel significantly increased the precorneal retention and permeation of KTZ, indicating that the developed formulation is a viable modality to treat fungal keratitis.

Hydroxypropyl methacrylamide-based copolymeric nanoparticles loaded with moxifloxacin as a mucoadhesive, cornea-penetrating nanomedicine eye drop with enhanced therapeutic benefits in bacterial keratitis.

Bacterial keratitis (BK) is a leading cause of visual impairment. The fluoroquinolone antibiotic moxifloxacin (Mox), being highly water-soluble, suffers from poor corneal penetration leading to unsatisfactory therapeutic outcomes in BK. Here, we prepared Mox-loaded co-polymeric nanoparticles (NPs) by entrapping the drug in co-polymeric NPs constituted by the self-assembly of a water-soluble copolymer, poly(ethylene glycol)-b-p(hydroxypropyl) methacrylamide (mPH). The polymer (mPH) was prepared using a radical polymerization technique at different mPEG: HPMA ratios of 1:70/100/150. The polymer/nanoparticles were characterized by GPC, CAC, DLS, SEM, XRD, DSC, FTIR, % DL, % EE, and release studies. The ex vivo muco-adhesiveness and corneal permeation ability were judged using a texture analyzer and Franz Diffusion Cells. In vitro cellular uptake, cytotoxicity, and safety assessment were performed using HCE cells in monolayers, spheroids, and multilayers in transwells. The DOE-optimized colloidal solution of Mox-mPH NPs (1:150) displayed a particle size of ~116 nm, superior drug loading (8.3%), entrapment (83.2%), robust mucoadhesion ex vivo, and ocular retention in vivo (~6 h) (judged by in vivo image analysis). The non-irritant formulation, Mox-mPH NPs (1:150) (proven by HET-CAM test) exhibited intense antimicrobial activity against P. aeruginosa, S. pneumoniae, and S. aureus in vitro analyzed by live-dead cells assay, zone of inhibition studies, and by determining the minimum inhibitory and bactericidal concentrations. The polymeric nanoparticles, mPH (1:150), decreased the opacity and the bacterial load compared to the other treatment groups. The studies warrant the safe and effective topical application of the Mox-mPH NPs solution in bacterial keratitis.