Injectable, Drug-Eluting Nanocrystals Prevent Fibrosis and Stricture Formation In Vivo.

BACKGROUND & AIMS: Tissue fibrosis results from uncontrolled healing responses leading to excessive mesenchymal cell activation and collagen and other extracellular matrix deposition. In the gastrointestinal tract, fibrosis leads to narrowing of the lumen and stricture formation. A drug treatment to prevent fibrosis and strictures in the gastrointestinal tract would be transformational for patient care. We aimed to develop a stricture treatment with the following characteristics and components: a small molecule with strong antifibrotic effects that is delivered locally at the site of the stricture to ensure correct lesional targeting while protecting the systemic circulation, and that is formulated with sustained-release properties to act throughout the wound healing processes. METHODS: A high-throughput drug screening was performed to identify small molecules with antifibrotic properties. Next, we formulated an antifibrotic small molecule for sustained release and tested its antifibrotic potential in 3 animal models of fibrosis. RESULTS: Sulconazole, a US Food and Drug Administration-approved drug for fungal infections, was found to have strong antifibrotic properties. Sulconazole was formulated as sulconazole nanocrystals for sustained release. We found that sulconazole nanocrystals provided superior or equivalent fibrosis prevention with less frequent dosing in mouse models of skin and intestinal tissue fibrosis. In a patient-like swine model of bowel stricture, a single injection of sulconazole nanocrystals prevented stricture formation. CONCLUSIONS: The current data lay the foundation for further studies to improve the management of a range of diseases and conditions characterized by tissue fibrosis.

Nanomedicine for Maternal and Fetal Health.

Conception, pregnancy, and childbirth are complex processes that affect both mother and fetus. Thus, it is perhaps not surprising that in the United States alone, roughly 11% of women struggle with infertility and 16% of pregnancies involve some sort of complication. This presents a clear need to develop safe and effective treatment options, though the development of therapeutics for use in women's health and particularly in pregnancy is relatively limited. Physiological and biological changes during the menstrual cycle and pregnancy impact biodistribution, pharmacokinetics, and efficacy, further complicating the process of administration and delivery of therapeutics. In addition to the complex pharmacodynamics, there is also the challenge of overcoming physiological barriers that impact various routes of local and systemic administration, including the blood-follicle barrier and the placenta. Nanomedicine presents a unique opportunity to target and sustain drug delivery to the reproductive tract and other relevant organs in the mother and fetus, as well as improve the safety profile and minimize side effects. Nanomedicine-based approaches have the potential to improve the management and treatment of infertility, obstetric complications, and fetal conditions.

The cervicovaginal mucus barrier to HIV-1 is diminished in bacterial vaginosis.

Bacterial vaginosis (BV), a condition in which the vaginal microbiota consists of community of obligate and facultative anaerobes rather than dominated by a single species of Lactobacillus, affects ~30% of women in the US. Women with BV are at 60% increased risk for HIV acquisition and are 3-times more likely to transmit HIV to an uninfected partner. As cervicovaginal mucus (CVM) is the first line of defense against mucosal pathogens and the home of the resident vaginal microbiota, we hypothesized the barrier function of CVM to HIV may be diminished in BV. Here, we characterized CVM properties including pH, lactic acid content, and Nugent score to correlate with the microbiota community composition, which was confirmed by 16S rDNA sequencing on a subset of samples. We then quantified the mobility of fluorescently-labeled HIV virions and nanoparticles to characterize the structural and adhesive barrier properties of CVM. Our analyses included women with Nugent scores categorized as intermediate (4-6) and BV (7-10), women that were either symptomatic or asymptomatic, and a small group of women before and after antibiotic treatment for symptomatic BV. Overall, we found that HIV virions had significantly increased mobility in CVM from women with BV compared to CVM from women with Lactobacillus crispatus-dominant microbiota, regardless of whether symptoms were present. We confirmed using nanoparticles and scanning electron microscopy that the impaired barrier function was due to reduced adhesive barrier properties without an obvious degradation of the physical CVM pore structure. We further confirmed a similar increase in HIV mobility in CVM from women with Lactobacillus iners-dominant microbiota, the species most associated with transitions to BV and that persists after antibiotic treatment for BV. Our findings advance the understanding of the protective role of mucus and highlight the interplay between vaginal microbiota and the innate barrier function mucus.

Characterization of an Adapted Murine Model of Intrauterine Inflammation-Induced Preterm Birth.

Preterm birth (PTB) affects nearly 15 million infants each year. Of these PTBs, >25% are a result of inflammation or infection. Animal models have improved our understanding of the mechanisms leading to PTB. Prior work has described induction of intrauterine inflammation in mice with a single injection of lipopolysaccharide (LPS). Herein, we have improved the reproducibility and potency of LPS in the model using two injections distal to the cervix. An in vivo imaging system revealed more uniform distribution of Evans Blue Dye using a double distal injection (DDI) approach compared with a single proximal injection (SPI). Endotoxin concentrations in vaginal lavage fluid from SPI dams were significantly higher than from DDI dams. At equivalent LPS doses, DDI consistently induced more PTB than SPI, and DDI showed a linear dose-response, whereas SPI did not. Gene expression in myometrial tissue revealed increased levels of inflammatory markers in dams that received LPS DDI compared with LPS SPI. The SPI group showed more significant overexpression in cervical remodeling genes, likely due to the leakage of LPS from the uterine horns through the cervix. The more reliable PTB induction and uniform uterine exposure provided by this new model will be useful for further studying fetal outcomes and potential therapeutics for the prevention of inflammation-induced PTB.

Controlled release of dexamethasone sodium phosphate with biodegradable nanoparticles for preventing experimental corneal neovascularization.

Corneal neovascularization (CNV) leads to the loss of corneal transparency and vision impairment, and can ultimately cause blindness. Topical corticosteroids are the first line treatment for suppressing CNV, but poor ocular bioavailability and rapid clearance of eye drops makes frequent administration necessary. Patient compliance with frequent eye drop application regimens is poor. We developed biodegradable nanoparticles (NP) loaded with dexamethasone sodium phosphate (DSP) using zinc ion bridging, DSP-Zn-NP, with dense coatings of poly(ethylene glycol) (PEG). DSP-Zn-NP were safe and capable of providing sustained delivery of DSP to the front of the eye following subconjunctival (SCT) administration in rats. We reported that a single SCT administration of DSP-Zn-NP prevented suture-induced CNV in rats for two weeks. In contrast, the eyes receiving SCT administration of either saline or DSP solution developed extensive CNV in less than 1 week. SCT administration of DSP-Zn-NP could be an effective strategy in preventing and treating CNV.

Hypo-osmolar Formulation of Tenofovir (TFV) Enema Promotes Uptake and Metabolism of TFV in Tissues, Leading to Prevention of SHIV/SIV Infection.

Oral preexposure prophylaxis (PrEP) has been approved for prophylaxis of HIV-1 transmission but is associated with high costs and issues of adherence. Protection from anal transmission of HIV using topical microbicides and methods congruent with sexual behavior offers the promise of improved adherence. We compared the pharmacokinetics (PK) and ex vivo efficacy of iso-osmolar (IOsm) and hypo-osmolar (HOsm) rectal enema formulations of tenofovir (TFV) in rhesus macaques. Single-dose PK of IOsm or HOsm high-dose (5.28 mg/ml) and low-dose (1.76 mg/ml) formulations of TFV enemas were evaluated for systemic uptake in blood, colorectal biopsy specimens, and rectal CD4+ T cells. Markedly higher TFV concentrations were observed in plasma and tissues after administration of the HOsm high-dose formulation than with all other formulations tested. TFV and TFV diphosphate (TFV-DP) concentrations in tissue correlated for the HOsm high-dose formulation, demonstrating rapid uptake and transformation of TFV to TFV-DP in tissues. TFV-DP amounts in tissues collected at 1 and 24 h were 7 times and 5 times higher, respectively (P < 0.01), than the ones collected in tissues with the IOsm formulation. The HOsm high-dose formulation prevented infection in ex vivo challenges of rectal tissues collected at 1, 24, and 72 h after the intrarectal dosing, whereas the same TFV dose formulated as an IOsm enema was less effective.

Modulating Vibrio cholerae quorum-sensing-controlled communication using autoinducer-loaded nanoparticles.

The rise of bacterial antibiotic resistance has created a demand for alternatives to traditional antibiotics. Attractive possibilities include pro- and anti-quorum sensing therapies that function by modulating bacterial chemical communication circuits. We report the use of Flash NanoPrecipitation to deliver the Vibrio cholerae quorum-sensing signal CAI-1 ((S)-3-hydroxytridecan-4-one) in a water dispersible form as nanoparticles. The particles activate V. cholerae quorum-sensing responses 5 orders of magnitude higher than does the identically administered free CAI-1 and are diffusive across in vivo delivery barriers such as intestinal mucus. This work highlights the promise of combining quorum-sensing strategies with drug delivery approaches for the development of next-generation medicines.

Liposome-based mucus-penetrating particles (MPP) for mucosal theranostics: demonstration of diamagnetic chemical exchange saturation transfer (diaCEST) magnetic resonance imaging (MRI).

Mucus barriers lining mucosal epithelia reduce the effectiveness of nanocarrier-based mucosal drug delivery and imaging ("theranostics"). Here, we describe liposome-based mucus-penetrating particles (MPP) capable of loading hydrophilic agents, e.g., the diaCEST MRI contrast agent barbituric acid (BA). We observed that polyethylene glycol (PEG)-coated liposomes containing ≥7 mol% PEG diffused only ~10-fold slower in human cervicovaginal mucus (CVM) compared to their theoretical speeds in water. 7 mol%-PEG liposomes contained sufficient BA loading for diaCEST contrast, and provided improved vaginal distribution compared to 0 and 3mol%-PEG liposomes. However, increasing PEG content to ~12 mol% compromised BA loading and vaginal distribution, suggesting that PEG content must be optimized to maintain drug loading and stability. Non-invasive diaCEST MRI illustrated uniform vaginal coverage and longer retention of BA-loaded 7 mol%-PEG liposomes compared to unencapsulated BA. Liposomal MPP with optimized PEG content hold promise for drug delivery and imaging at mucosal surfaces. FROM THE CLINICAL EDITOR: This team of authors characterized liposome-based mucus-penetrating particles (MPP) capable of loading hydrophilic agents, such as barbituric acid (a diaCEST MRI contrast agent) and concluded that liposomal MPP with optimized PEG coating enables drug delivery and imaging at mucosal surfaces.

Pretreatment of human cervicovaginal mucus with pluronic F127 enhances nanoparticle penetration without compromising mucus barrier properties to herpes simplex virus.

Mucosal drug delivery nanotechnologies are limited by the mucus barrier that protects nearly all epithelial surfaces not covered with skin. Most polymeric nanoparticles, including polystyrene nanoparticles (PS), strongly adhere to mucus, thereby limiting penetration and facilitating rapid clearance from the body. Here, we demonstrate that PS rapidly penetrate human cervicovaginal mucus (CVM), if the CVM has been pretreated with sufficient concentrations of Pluronic F127. Importantly, the diffusion rate of large polyethylene glycol (PEG)-coated, nonmucoadhesive nanoparticles (PS-PEG) did not change in F127-pretreated CVM, implying that F127 did not significantly alter the native pore structure of CVM. Additionally, herpes simplex virus type 1 (HSV-1) remains adherent in F127-pretreated CVM, indicating that the presence of F127 did not reduce adhesive interactions between CVM and the virions. In contrast to treatment with a surfactant that has been approved for vaginal use as a spermicide (nonoxynol-9 or N9), there was no increase in inflammatory cytokine release in the vaginal tract of mice after daily application of 1% F127 for 1 week. Pluronic F127 pretreatment holds potential as a method to safely improve the distribution, retention, and efficacy of nanoparticle formulations without compromising CVM barrier properties to pathogens.

Hypotonic, gel-forming delivery system for vaginal drug administration.

Vaginal drug delivery is often preferred over systemic delivery to reduce side effects and increase efficacy in treating diseases and conditions of the female reproductive tract (FRT). Current vaginal products have drawbacks, including spontaneous ejection of drug-eluting rings and unpleasant discharge from vaginal creams. Here, we describe the development and characterization of a hypotonic, gel-forming, Pluronic-based delivery system for vaginal drug administration. The rheological properties were characterized with and without common hydrogel polymers to demonstrate the versatility. Both qualitative and quantitative approaches were used to determine the Pluronic F127 concentration below the critical gel concentration (CGC) that was sufficient to achieve gelation when formulated to be hypotonic to the mouse vagina. The hypotonic, gel-forming formulation was found to form a thin, uniform gel layer along the vaginal epithelium in mice, in contrast to the rapidly forming conventional gelling formulation containing polymer above the CGC. When the hypotonic, gel-forming vehicle was formulated in combination with a progesterone nanosuspension (ProGel), equivalent efficacy was observed in the prevention of chemically-induced preterm birth (PTB) compared to commercial Crinone® vaginal cream. Further, ProGel showed marked benefits in reducing unpleasant discharge, reducing product-related toxicity, and improving compatibility with vaginal bacteria in vitro. A hypotonic, gel-forming delivery system may be a viable option for therapeutic delivery to the FRT.

Next generation therapeutics for retinal neurodegenerative diseases.

Neurodegenerative diseases affecting the visual system encompass glaucoma, macular degeneration, retinopathies, and inherited genetic disorders such as retinitis pigmentosa. These ocular pathologies pose a serious burden of visual impairment and blindness worldwide. Current treatment modalities include small molecule drugs, biologics, or gene therapies, most of which are administered topically as eye drops or as injectables. However, the topical route of administration faces challenges in effectively reaching the posterior segment and achieving desired concentrations at the target site, while injections and implants risk severe complications, such as retinal detachment and endophthalmitis. This necessitates the development of innovative therapeutic strategies that can prolong drug release, deliver effective concentrations to the back of the eye with minimal systemic exposure, and improve patient compliance and safety. In this review, we introduce retinal degenerative diseases, followed by a discussion of the existing clinical standard of care. We then delve into detail about drug and gene delivery systems currently in preclinical and clinical development, including formulation and delivery advantages/drawbacks, with a special emphasis on potential for clinical translation.

Locally administered nanosuspension increases delivery of estradiol for the treatment of vaginal atrophy in mice.

Vaginal atrophy affects up to 57% of post-menopausal women, with symptoms ranging from vaginal burning to dysuria. Estradiol hormone replacement therapy may be prescribed to alleviate these symptoms, though many vaginal products have drawbacks including increased discharge and local tissue toxicity due to their hypertonic nature. Here, we describe the development and characterization of a Pluronic F127-coated estradiol nanosuspension (NS) formulation for improved vaginal estradiol delivery. We compare the pharmacokinetics to the clinical comparator vaginal cream (Estrace) and demonstrate increased delivery of estradiol to the vaginal tissue. We utilized ovariectomized (OVX) mice as a murine model of post-menopausal vaginal atrophy and demonstrated equivalent efficacy in vaginal re-epithelialization when dosed with either the estradiol NS or Estrace cream. Further, we demonstrate compatibility of the estradiol NS with vaginal bacteria in vitro. We demonstrate that a Pluronic F127-coated estradiol NS may be a viable option for the treatment of post-menopausal vaginal atrophy.

Investigating inhibitors of 1-deoxy-d-xylulose 5-phosphate synthase in a mouse model of UTI.

The rising rate of antimicrobial resistance continues to threaten global public health. Further hastening antimicrobial resistance is the lack of new antibiotics against new targets. The bacterial enzyme, 1-deoxy-d-xylulose 5-phosphate synthase (DXPS), is thought to play important roles in central metabolism, including processes required for pathogen adaptation to fluctuating host environments. Thus, impairing DXPS function represents a possible new antibacterial strategy. We previously investigated a DXPS-dependent metabolic adaptation as a potential target in uropathogenic Escherichia coli (UPEC) associated with urinary tract infection (UTI), using the DXPS-selective inhibitor butyl acetylphosphonate (BAP). However, investigations of DXPS inhibitors in vivo have not been conducted. The goal of the present study is to advance DXPS inhibitors as in vivo probes and assess the potential of inhibiting DXPS as a strategy to prevent UTI in vivo. We show that BAP was well-tolerated at high doses in mice and displayed a favorable pharmacokinetic profile for studies in a mouse model of UTI. Further, an alkyl acetylphosphonate prodrug (homopropargyl acetylphosphonate, pro-hpAP) was significantly more potent against UPEC in urine culture and exhibited good exposure in the urinary tract after systemic dosing. Prophylactic treatment with either BAP or pro-hpAP led to a partial protective effect against UTI, with the prodrug displaying improved efficacy compared to BAP. Overall, our results highlight the potential for DXPS inhibitors as in vivo probes and establish preliminary evidence that inhibiting DXPS impairs UPEC colonization in a mouse model of UTI.IMPORTANCENew antibiotics against new targets are needed to prevent an antimicrobial resistance crisis. Unfortunately, antibiotic discovery has slowed, and many newly FDA-approved antibiotics do not inhibit new targets. Alkyl acetylphosphonates (alkyl APs), which inhibit the enzyme 1-deoxy-d-xylulose 5-phosphate synthase (DXPS), represent a new possible class of compounds as there are no FDA-approved DXPS inhibitors. To our knowledge, this is the first study demonstrating the in vivo safety, pharmacokinetics, and efficacy of alkyl APs in a urinary tract infection mouse model.

Ex vivo characterization of particle transport in mucus secretions coating freshly excised mucosal tissues.

Sustained drug delivery to mucosal surfaces has the potential to improve the effectiveness of prophylactic and therapeutic treatments for numerous diseases and conditions, including inflammatory bowel disease, sexually transmitted diseases, cystic fibrosis, glaucoma, dry eye, and various cancers. Sustained delivery systems such as nanoparticles can be useful for mucosal delivery, but recent work suggests they must penetrate the rapidly cleared mucus barrier that overlies all mucosal epithelia to achieve uniform distribution on epithelial surfaces and enhanced residence time. Thus, it is important to evaluate the mucus-penetrating ability of nanosized delivery systems in preclinical animal studies, and for administration to humans. We describe a simple ex vivo method to visualize and quantify nanoparticle transport in mucus on fresh mucosal tissues. Using this method in murine models, we observed variations in the mucus mesh at different anatomical locations, as well as cyclical variations that may have implications for mucosal delivery.

The microstructure and bulk rheology of human cervicovaginal mucus are remarkably resistant to changes in pH.

The protective barrier, lubricant, and clearance functions of mucus are intimately coupled to its microstructure and bulk rheology. Mucus gels consist of a network of mucin biopolymers along with lipids, salts, and other proteins and exhibit similar biochemical and physical properties across diverse mucosal surfaces. Nevertheless, mucus is exposed to a broad range of pH values throughout the human body. Protein functions are typically sensitive to small changes in pH, and prior investigations using reconstituted, purified mucin gels suggested mucus undergoes a transition from a low-viscosity liquid at neutral pH to a highly viscoelastic solid at low pH. We sought to determine whether those observations hold for fresh, minimally perturbed human mucus ex vivo by using different-sized muco-inert nanoparticles to probe microstructure and cone-and-plate rheometry to measure bulk rheology. We demonstrate that both the microstructure and bulk rheology of fresh, undiluted, and minimally perturbed cervicovaginal mucus exhibit relatively minor changes from pH 1-2 to 8-9, in marked contrast with the pH sensitivity of purified mucin gels. Our work also suggests additional components in mucus secretions, typically eliminated during mucin purification and reconstitution, may play an important role in maintaining the protective properties of mucus.

Reduced inspired oxygen decreases retinal superoxide radicals and promotes cone function and survival in a model of retinitis pigmentosa.

Retinitis pigmentosa (RP) is caused by many different mutations that promote the degeneration of rod photoreceptors and have no direct effect on cones. After the majority of rods have died cone photoreceptors begin to slowly degenerate. Oxidative damage contributes to cone cell death and it has been hypothesized that tissue hyperoxia due to reduced oxygen consumption from the loss of rods is what initiates oxidative stress. Herein, we demonstrate in animal models of RP that reduction of retinal hyperoxia by reducing inspired oxygen to continuous breathing of 11% O2 reduced the generation of superoxide radicals in the retina and preserved cone structure and function. These data indicate that retinal hyperoxia is the initiating event that promotes oxidative damage, loss of cone function, and cone degeneration in the RP retina.

Nanofiber-coated, tacrolimus-eluting sutures inhibit post-operative neointimal hyperplasia in rats.

Post-operative complications of vascular anastomosis procedures remain a significant clinical challenge and health burden globally. Each year, millions of anastomosis procedures connect arteries and/or veins in vascular bypass, vascular access, organ transplant, and reconstructive surgeries, generally via suturing. Dysfunction of these anastomoses, primarily due to neointimal hyperplasia and the resulting narrowing of the vessel lumen, results in failure rates of up to 50% and billions of dollars in costs to the healthcare system. Non-absorbable sutures are the gold standard for vessel anastomosis; however, damage from the surgical procedure and closure itself causes an inflammatory cascade that leads to neointimal hyperplasia at the anastomosis site. Here, we demonstrate the development of a novel, scalable manufacturing system for fabrication of high strength sutures with nanofiber-based coatings composed of generally regarded as safe (GRAS) polymers and either sirolimus, tacrolimus, everolimus, or pimecrolimus. These sutures provided sufficient tensile strength for maintenance of the vascular anastomosis and sustained drug delivery at the site of the anastomosis. Tacrolimus-eluting sutures provided a significant reduction in neointimal hyperplasia in rats over a period of 14 days with similar vessel endothelialization in comparison to conventional nylon sutures. In contrast, systemically delivered tacrolimus caused significant weight loss and mortality due to toxicity. Thus, drug-eluting sutures provide a promising platform to improve the outcomes of vascular interventions without modifying the clinical workflow and without the risks associated with systemic drug delivery.

Cervicovaginal mucus barrier properties during pregnancy are impacted by the vaginal microbiome.

Introduction: Mucus in the female reproductive tract acts as a barrier that traps and eliminates pathogens and foreign particles via steric and adhesive interactions. During pregnancy, mucus protects the uterine environment from ascension of pathogens and bacteria from the vagina into the uterus, a potential contributor to intrauterine inflammation and preterm birth. As recent work has demonstrated the benefit of vaginal drug delivery in treating women's health indications, we sought to define the barrier properties of human cervicovaginal mucus (CVM) during pregnancy to inform the design of vaginally delivered therapeutics during pregnancy. Methods: CVM samples were self-collected by pregnant participants over the course of pregnancy, and barrier properties were quantified using multiple particle tracking. 16S rRNA gene sequencing was performed to analyze the composition of the vaginal microbiome. Results: Participant demographics differed between term delivery and preterm delivery cohorts, with Black or African American participants being significantly more likely to delivery prematurely. We observed that vaginal microbiota is most predictive of CVM barrier properties and of timing of parturition. Lactobacillus crispatus dominated CVM samples showed increased barrier properties compared to polymicrobial CVM samples. Discussion: This work informs our understanding of how infections occur during pregnancy, and directs the engineering of targeted drug treatments for indications during pregnancy.

Antibiotic-free vaginal microbiota transplant with donor engraftment, dysbiosis resolution and live birth after recurrent pregnancy loss: a proof of concept case study.

Background: Vaginal dysbiosis covers imbalances in the vaginal microbiota, defined by altered composition of bacteria, viruses, and fungi and is associated with euploid pregnancy losses, premature birth, infertility, or bacterial vaginosis. A large proportion of women who have vaginal dysbiosis do not experience any symptoms. Antibiotics are the traditional treatment, recently combined with local probiotics in some cases. Vaginal Microbiota Transplantation (VMT) with eubiotic vaginal bacterial microbiota after antibiotic eradication of pathogens has successfully been performed in a case study with five patients, but no VMT has been performed without the use of antibiotics. Methods: This is a proof of concept case study. The patient was found to have vaginal dysbiosis at the RPL clinic at Copenhagen University Hospital Hvidovre, Denmark on the 23rd of June 2021. She was offered and accepted to receive experimental treatment in the form of a VMT as a compassionate use case. VMT is the transfer of cervicovaginal secretions (CVS) from a healthy donor with a Lactobacillus-dominant vaginal microbiome to a recipient with a dysbiotic vaginal microbiome. CVS is a mixture of e.g., mucus, bacteria, metabolites present in the vaginal canal. Potential donors were thoroughly screened for the absence of STIs, and the most suitable donor sample for the specific patient in this study was determined via an in vitro microbiome competition assay. Findings: A 30-year-old patient with one livebirth and a complicated pregnancy history of two stillbirths and 1 s trimester pregnancy loss in gestational weeks 27 (2019), 17 (2020) and 23 (2020) respectively with complaints of vaginal irritation and discharge that had aggravated in all her pregnancies. Her vaginal microbiome composition showed a 90% dominance of Gardnerella spp. After one VMT there was a complete shift in microbiome composition to 81.2% L. crispatus and 9% L. jensenii with a concurrent resolvement of vaginal symptoms. Single nucleotide polymorphism-analysis confirmed her microbiome to be of donor origin and it remain stable now 1.5 years after the VMT. Five months after the VMT she became pregnant and has successfully delivered a healthy baby at term. Interpretation: Here we report a successful VMT with confirmed donor strain engraftment followed by a successful pregnancy and delivery after a series of late pregnancy losses/stillbirths. Findings suggest that VMT is a potential treatment for severe vaginal dysbiosis. Further, larger studies are required. Funding: The study was partially funded (i.e., analysis costs) by Freya Biosciences Aps, Fruebjergvej, 2100 Copenhagen, Denmark.

Six-month effective treatment of corneal graft rejection.

Topical corticosteroid eye drop is the mainstay for preventing and treating corneal graft rejection. While the frequent topical corticosteroid use is associated with risk of intraocular pressure (IOP) elevation and poor patient compliance that leads to graft failure and the requirement for a repeated, high-risk corneal transplantation. Here, we developed dexamethasone sodium phosphate (DSP)-loaded dicarboxyl-terminated poly(lactic acid) nanoparticle (PLA DSP-NP) formulations with relatively high drug loading (8 to 10 weight %) and 6 months of sustained intraocular DSP delivery in rats with a single dosing. PLA DSP-NP successfully reversed early signs of corneal rejection, leading to rat corneal graft survival for at least 6 months. Efficacious PLA DSP-NP doses did not affect IOP and showed no signs of ocular toxicity in rats for up to 6 months. Subconjunctival injection of DSP-NP is a promising approach for safely preventing and treating corneal graft rejection with the potential for improved patient adherence.