Advances in Drug Treatments for Companion Animal Obesity.

Companion animal obesity has emerged as a significant veterinary health concern globally, with escalating rates posing challenges for preventive and therapeutic interventions. Obesity not only leads to immediate health problems but also contributes to various comorbidities affecting animal well-being and longevity, with consequent emotional and financial burdens on owners. While past treatment strategies have shown limited success, recent breakthroughs in human medicine present new opportunities for addressing this complex issue in companion animals. Here, we discuss the potential of GLP-1 receptor agonists, specifically semaglutide and tirzepatide, already approved for human use, for addressing companion animal obesity. These drugs, originally developed to treat type 2 diabetes in humans and subsequently repurposed to treat obesity, have demonstrated remarkable weight loss effects in rodents, non-human primates and people. Additionally, newer drug combinations have shown even more promising results in clinical trials. Despite current cost and supply challenges, advancements in oral and/or extended-release formulations and increased production may make these drugs more accessible for veterinary use. Thus, these drugs may have utility in companion animal weight management, and future feasibility studies exploring their efficacy and safety in treating companion animal obesity are warranted.

Prophylactic activity of orally administered dry-heat-sterilized Acremonium egyptiacum against Trypanosoma congolense-induced animal African trypanosomosis.

Animal African trypanosomosis (AAT) is an important global disease of livestock that causes economic losses of up to 4.5 billion US dollars per year. Thus, eliminating AAT in endemic countries will improve agricultural productivity and economic growth. To prevent AAT, vector control and the development of prophylactic drugs are crucial. Ascofuranone (AF) is a bioactive fungal compound with proven in vitro trypanocidal potency and in vivo treatment efficacy. However, the complex stereoselective synthesis of AF has prevented its cost-effective industrial production. Recently, a genetically modified strain of Acremonium egyptiacum fungus that produces a high yield of AF was developed. Therefore, we hypothesized that the oral administration of the AF-producing fungus itself may be effective against AAT. Hence, this study aimed to evaluate the prophylactic activity of orally administered dry-heat-sterilized A. egyptiacum against Trypanosoma congolense IL3000 infection using a mouse model. The survival rate was significantly prolonged (p = 0.009), and parasitemia was suppressed in all AF-fungus-treated groups (Group 1-9) compared with that in the untreated control group (Group 10). Hence, the trypanocidal activity of AF was retained after dry-heat-sterilization of the AF-producing fungus and that its oral administration effectively prevented AAT. Since AAT is endemic to rural areas with underdeveloped veterinary infrastructure, dry-heat-sterilized A. egyptiacum would be the most cost-effective potential treatment for AAT.

Silica nanoparticles enhance interfacial self-adherence of a multi-layered extracellular matrix scaffold for vascular tissue regeneration.

PURPOSE: Based on the clinical need for grafts for vascular tissue regeneration, our group developed a customizable scaffold derived from the human amniotic membrane. Our approach consists of rolling the decellularized amniotic membrane around a mandrel to form a multilayered tubular scaffold with tunable diameter and wall thickness. Herein, we aimed to investigate if silica nanoparticles (SiNP) could enhance the adhesion of the amnion layers within these rolled grafts. METHODS: To test this, we assessed the structural integrity and mechanical properties of SiNP-treated scaffolds. Mechanical tests were repeated after six months to evaluate adhesion stability in aqueous environments. RESULTS: Our results showed that the rolled SiNP-treated scaffolds maintained their tubular shape upon hydration, while non-treated scaffolds collapsed. By scanning electron microscopy, SiNP-treated scaffolds presented more densely packed layers than untreated controls. Mechanical analysis showed that SiNP treatment increased the scaffold's tensile strength up to tenfold in relation to non-treated controls and changed the mechanism of failure from interfacial slipping to single-point fracture. The nanoparticles reinforced the scaffolds both at the interface between two distinct layers and within each layer of the extracellular matrix. Finally, SiNP-treated scaffolds significantly increased the suture pullout force in comparison to untreated controls. CONCLUSION: Our study demonstrated that SiNP prevents the unraveling of a multilayered extracellular matrix graft while improving the scaffolds' overall mechanical properties. In addition to the generation of a robust biomaterial for vascular tissue regeneration, this novel layering technology is a promising strategy for a number of bioengineering applications.

Silica nanoparticles enhance the cyto- and hemocompatibility of a multilayered extracellular matrix scaffold for vascular tissue regeneration.

PURPOSE: The limited availability of autologous vessels for vascular bypass surgeries is a major roadblock to treating severe cardiovascular diseases. Based on this clinical priority, our group has developed a novel engineered vascular graft by rolling human amniotic membranes into multilayered extracellular matrixes (ECM). When treated with silica nanoparticles (SiNP), these rolled scaffolds showed a significant improvement in their structural and mechanical properties, matching those from gold standard autologous grafts. However, it remained to be determined how cells respond to SiNP-treated materials. As a first step toward understanding the biocompatibility of SiNP-dosed biomaterials, we aimed to assess how endothelial cells and blood components interact with SiNP-treated ECM scaffolds. METHODS: To test this, we used established in vitro assays to study SiNP and SiNP-treated scaffolds' cyto and hemocompatibility. RESULTS: Our results showed that SiNP effects on cells were concentration-dependent with no adverse effects observed up to 10 µg/ml of SiNP, with higher concentrations inducing cytotoxic and hemolytic responses. The SiNP also enhanced the scaffold's hydrophobicity state, a feature known to inhibit platelet and immune cell adhesion. Accordingly, SiNP-treated scaffolds were also shown to support endothelial cell growth while preventing platelet and leukocyte adhesion. CONCLUSION: Our findings suggest that the addition of SiNP to human amniotic membrane extracellular matrixes improves the cyto- and hemocompatibility of rolled scaffolds and highlights this strategy as a robust mechanism to stabilize layered collagen scaffolds for vascular tissue regeneration.

RNA polymerase II pausing is essential during spermatogenesis for appropriate gene expression and completion of meiosis.

Male germ cell development requires precise regulation of gene activity in a cell-type and stage-specific manner, with perturbations in gene expression during spermatogenesis associated with infertility. Here, we use steady-state, nascent and single-cell RNA sequencing strategies to comprehensively characterize gene expression across male germ cell populations, to dissect the mechanisms of gene control and provide new insights towards therapy. We discover a requirement for pausing of RNA Polymerase II (Pol II) at the earliest stages of sperm differentiation to establish the landscape of gene activity across development. Accordingly, genetic knockout of the Pol II pause-inducing factor NELF in immature germ cells blocks differentiation to spermatids. Further, we uncover unanticipated roles for Pol II pausing in the regulation of meiosis during spermatogenesis, with the presence of paused Pol II associated with double-strand break (DSB) formation, and disruption of meiotic gene expression and DSB repair in germ cells lacking NELF.

In vitro trypanocidal potency and in vivo treatment efficacy of oligomeric ethylene glycol-tethered nitrofurantoin derivatives.

African trypanosomiasis is a significant vector-borne disease of humans and animals in the tsetse fly belt of Africa, particularly affecting production animals such as cattle, and thus, hindering food security. Trypanosoma congolense (T. congolense), the causative agent of nagana, is livestock's most virulent trypanosome species. There is currently no vaccine against trypanosomiasis; its treatment relies solely on chemotherapy. However, pathogenic resistance has been established against trypanocidal agents in clinical use. This underscores the need to develop new therapeutics to curb trypanosomiasis. Many nitroheterocyclic drugs or compounds, including nitrofurantoin, possess antiparasitic activities in addition to their clinical use as antibiotics. The current study evaluated the in vitro trypanocidal potency and in vivo treatment efficacy of previously synthesized antileishmanial active oligomeric ethylene glycol derivatives of nitrofurantoin. The trypanocidal potency of analogues 2a-o varied among the trypanosome species; however, T. congolense strain IL3000 was more susceptible to these drug candidates than the other human and animal trypanosomes. The arylated analogues 2k (IC50 0.04 µM; SI >6365) and 2l (IC50 0.06 µM; SI 4133) featuring 4-chlorophenoxy and 4-nitrophenoxy moieties, respectively, were revealed as the most promising antitrypanosomal agents of all analogues against T. congolense strain IL3000 trypomastigotes with nanomolar activities. In a preliminary in vivo study involving T. congolense strain IL3000 infected BALB/c mice, the oral administration of 100 mg/kg/day of 2k caused prolonged survival up to 18 days post-infection relative to the infected but untreated control mice which survived 9 days post-infection. However, no cure was achieved due to its poor solubility in the in vivo testing medium, assumably leading to low oral bioavailability. These results confirm the importance of the physicochemical properties lipophilicity and water solubility in attaining not only in vitro trypanocidal potency but also in vivo treatment efficacy. Future work will focus on the chemical optimization of 2k through the investigation of analogues containing solubilizing groups at certain positions on the core structure to improve solubility in the in vivo testing medium which, in the current investigation, is the biggest stumbling block in successfully treating either animal or human Trypanosoma infections.

Role for Nongenomic Estrogen Signaling in Male Fertility.

Estrogen actions are mediated by both nuclear (n) and membrane (m) localized estrogen receptor 1 (ESR1). Male Esr1 knockout (Esr1KO) mice lacking functional Esr1 are infertile, with reproductive tract abnormalities. Male mice expressing nESR1 but lacking mESR1 (nuclear-only estrogen receptor 1 mice) are progressively infertile due to testicular, rete testis, and efferent ductule abnormalities similar to Esr1KO males, indicating a role for mESR1 in male reproduction. The H2NES mouse expresses only mESR1 but lacks nESR1. The goal of this study was to identify the functions of mESR1 alone in mice where nESR1 was absent. Breeding trials showed that H2NES males are fertile, with decreased litter numbers but normal pup numbers/litter. In contrast to Esr1KO mice, H2NES testicular, and epididymal weights were not reduced, and seminiferous tubule abnormalities were less pronounced. However, Esr1KO and H2NES males both had decreased sperm motility and a high incidence of abnormal sperm morphology. Seminiferous tubule and rete testis dilation and decreased efferent ductule epithelial height characteristic of Esr1KO males were reduced in H2NES. Consistent with this, expression of genes involved in fluid transport and ion movement that were reduced in Esr1KO (Aqp1, Car2, Car14, Cftr) were partially or fully restored to wild-type levels in H2NES. In summary, in contrast to Esr1KO males, H2NES males are fertile and have reduced phenotypic and functional abnormalities in the testis and efferent ductules. Thus, mESR1 alone, in the absence of nESR1, can partially regulate male reproductive tract structure and function, emphasizing its importance for overall estrogen action.

Horizon scanning the application of probiotics for wildlife.

The provision of probiotics benefits the health of a wide range of organisms, from humans to animals and plants. Probiotics can enhance stress resilience of endangered organisms, many of which are critically threatened by anthropogenic impacts. The use of so-called 'probiotics for wildlife' is a nascent application, and the field needs to reflect on standards for its development, testing, validation, risk assessment, and deployment. Here, we identify the main challenges of this emerging intervention and provide a roadmap to validate the effectiveness of wildlife probiotics. We cover the essential use of inert negative controls in trials and the investigation of the probiotic mechanisms of action. We also suggest alternative microbial therapies that could be tested in parallel with the probiotic application. Our recommendations align approaches used for humans, aquaculture, and plants to the emerging concept and use of probiotics for wildlife.

Overcoming the clinical challenges of traditional ayahuasca: a first-in-human trial exploring novel routes of administration of N,N-Dimethyltryptamine and harmine.

Recently, the Amazonian plant medicine "ayahuasca"-containing the psychedelic compound N,N-dimethyltryptamine (DMT) and numerous ß-carboline alkaloids, such as harmine-has been suggested to exhibit beneficial effects in patients with affective and other mental health disorders. Although ayahuasca ingestion is considered safe, its pharmacokinetics/pharmacodynamics and tolerability profile pose some challenges and may limit the clinical applicability in vulnerable patient populations. While overdosing and the admixture of intolerable plant constituents may explain some of the common adverse reactions, the peroral route of administration may represent another relevant source of gastro-intestinal intolerabilities and unpredictable pharmacokinetics across users. To overcome these challenges, the present work aimed at creating ayahuasca-analogue formulations with improved pharmacokinetics and tolerability profiles. To this end, we developed peroral formulas and compared them with parenteral formulas specifically designed to circumvent the gastro-intestinal tract. In more detail, peroral administration of a capsule (containing purified DMT and harmine) was tested against a combined administration of an oromucosal harmine tablet and an intranasal DMT spray at two dose levels in an open-label within-subject study in 10 healthy male subjects. Pharmacokinetic and pharmacodynamic profiles were assessed by means of continuous blood sampling, vital sign monitoring, and psychometric assessments. Common side effects induced by traditional herbal ayahuasca such as nausea, vomiting, and diarrhea were significantly attenuated by our DMT/harmine formulations. While all preparations were well tolerated, the combined buccal/intranasal administration of harmine and DMT yielded substantially improved pharmacokinetic profiles, indicated by significantly reduced variations in systemic exposure. In conclusion, the combined buccal/intranasal administration of harmine and DMT is an innovative approach that may pave the way towards a safe, rapid-acting, and patient-oriented administration of DMT/harmine for the treatment of affective disorders. Clinical Trial Registration: clinicaltrials.gov, identifier NCT04716335.

Targeting estrogen signaling and biosynthesis for aged skin repair.

Non-healing skin wounds are disproportionally prevalent in older adults. Current treatments do not account for the particularities of aged skin and result in inadequate outcomes. Overall, healing chronic wounds in the elderly remains a major unmet clinical need. Estrogens play a critical role in reproduction but also have important actions in non-reproductive organs. Estrogen biosynthesis and signaling pathways are locally activated during physiological wound healing, processes that are inhibited in elderly estrogen-deprived skin. Estrogen deprivation has been shown to be a critical mediator of impaired wound healing in both postmenopausal women and aged men, and topical estrogen application reverses age-associated delayed wound healing in both elderly men and women. These data indicate that adequate estrogen biosynthesis and properly regulated estrogen signaling pathways are essential for normal wound healing and can be targeted to optimize tissue repair in the elderly. However, due to fundamental questions regarding how to safely restore estrogen signaling locally in skin wounds, there are currently no therapeutic strategies addressing estrogen deficiency in elderly chronic wounds. This review discusses established and recent literature in this area and proposes the hypothesis that estrogen plays a pleiotropic role in skin aging and that targeting estrogen signaling and biosynthesis could promote skin repair in older adults.

Candida antarctica lipase B performance in organic solvent at varying water activities studied by molecular dynamics simulations.

Applications of lipases in low-water environments are found across a broad range of industries, including the pharmaceutical and oleochemical sectors. This includes condensation reactions in organic solvents where the enzyme activity has been found to depend strongly on both the solvent and the water activity (aw). Despite several experimental and computational studies, knowledge is largely empirical, and a general predictive approach is much needed. To close this gap, we chose native Candida antarctica lipase B (CALB) and two mutants thereof and used molecular dynamics (MD) simulations to gain a molecular understanding of the effect of aw on the specific activity of CALB in hexane. Based on the simulations, we propose four criteria to understand the performance of CALB in organic media, which is supported by enzyme kinetics experiments. First, the lipase must be stable in the organic solvent, which was the case for native CALB and the two mutants studied here. Secondly, water clusters that form and grow close to the active site must not block the path of substrate molecules into the active site. Thirdly, the lipase's lid must not cover the active site. Finally, mutations and changes in aw must not disrupt the geometry of the active site. We show that mutating specific residues close to the active site can hinder water cluster formation and growth, making the lipase resistant to changes in aw. Our computational screening criteria could potentially be used to screen in-silico designed variants, so only promising candidates could be pushed forward to characterisation.

In Vitro and In Vivo Trypanocidal Efficacy of Nitrofuryl- and Nitrothienylazines.

African trypanosomiasis is a vector-borne disease of animals and humans in the tsetse fly belt of Africa. Trypanosoma congolense ("nagana") is the most pathogenic trypanosome in livestock and causes high morbidity and mortality rates among cattle. In the absence of effective preventative vaccines, the management of trypanosomiasis relies on chemoprophylaxis and/or -therapy. However, the trypanocides in clinical use exhibit poor oral bioavailability and toxicity, and therapeutic failures occur because of resistant strains. Because nitrofurantoin displayed, in addition to its clinical use, promising antiparasitic activity, the current study was conducted to evaluate the in vitro trypanocidal activity and preliminary in vivo treatment efficacy of previously synthesized nitrofuranylazines. The trypanocidal activity of these nitrofuran derivatives varied among the evaluated trypanosome species; however, T. congolense strain IL3000 was more susceptible than other animal and human trypanosomes. The nitrofurylazines 4a (IC50 0.04 µM; SI > 7761) and 7a (IC50 0.03 µM; SI > 9542) as well as the nitrothienylazine 8b (IC50 0.04 µM; SI 232), with nanomolar IC50 values, were revealed as early antitrypanosomal leads. Although these derivatives showed strong trypanocidal activity in vitro, no in vivo treatment efficacy was observed in T. congolense IL3000 infected mice after both oral and intraperitoneal administration in a preliminary study. This was attributed to the poor solubility of the test compounds in the in vivo testing media. Indeed, a challenge in drug discovery is finding a balance between the physicochemical properties of a drug candidate, particularly lipophilicity and water solubility, and maintaining adequate potency to provide an effective dose. Hence, future chemical modifications may be required to generate lead-like to lead-like nitrofuranylazines that possess optimal physicochemical and pharmacokinetic properties while retaining in vitro and, ultimately, in vivo trypanocidal efficacy.

Design, synthesis, electrochemistry and anti-trypanosomatid hit/lead identification of nitrofuranylazines.

Chagas disease and leishmaniasis are vector-borne infectious diseases affecting both humans and animals. These neglected tropical diseases can be fatal if not treated. Hundreds to thousands of new Chagas disease and leishmaniasis cases are being reported by the WHO every year, and currently available treatments are insufficient. Severe adverse effects, impractical administrations and increased pathogen resistance against current clinical treatments underscore a serious need for the development of new drugs to curb these ailments. In search for such drugs, we investigated a series of nitrofuran-based azine derivatives. Herein, we report the design, synthesis, electrochemistry, and biological activity of these derivatives against promastigotes and amastigotes of Leishmania major, and L. donovani strains, as well as epimastigotes and trypomastigotes of Trypanosoma cruzi. Two leishmanicidal early leads and one trypanosomacidal hit with submicromolar activity were uncovered and stand for further in vivo investigation in the search for new antitrypanosomatid drugs. Future objective will focus on the identification of involved biological targets with the parasites.

Two Total Syntheses of Trigoxyphins K and L.

Two total syntheses are presented for trigoxyphins K and L, tricyclic terpenoids from Trigonostemon xyphophylloides. The first proceeds via electrophlic cyclization in A/C-ring substrates to close the B ring at C4-C5 and then 1O2-mediated hydroxybutenolide formation to trigoxyphin L, with Luche reduction leading to trigoxyphin K. The second route develops from tetralone ring expansion to a B/C-ring intermediate that, by one-step O-demethylation-lactonization-isomerization, affords trigoxyphin K and then trigoxyphin L following enolate oxygenation.

Effects of Deslorelin on Testosterone Secretion and Testicular Volume in Male Rhesus Macaques (Macaca mulatta).

Sterility in male NHP has long been achieved through surgical castration or vasectomy. However, these techniques are irreversible, require a surgical procedure, and have potential consequences such as sperm granulomas and long recovery time. Deslorelin is a gonadotropin-releasing hormone agonist that temporarily and reversibly suppresses sex hormone secretion. Our goal in this study was to investigate the effects of deslorelin on testosterone secretion and testicular volume in male rhesus macaques (Macaca mulatta). Male macaques (n = 4) each received two, 4.7-mg deslorelin implants subcutaneously in the interscapular region. Serum testosterone and testicular volume were then monitored at specific time points until 10 mo after treatment. Testosterone suppression was defined as testosterone levels lower than 0.6 ng/mL for a sustained period of at least 30 d. After implantation, mean testicular volume was significantly reduced by day 121. Testosterone suppression was observed in all subjects. However, the time from implantation to testosterone suppression and duration of suppression varied. Two macaques were hormonally suppressed by day 26 after implantation and remained suppressed for at least 6 mo. The other 2 macaques were hormonally suppressed by 2 mo after implantation; of these two, one remained suppressed for 70 days while the other was suppressed for at least 245 days. We conclude that deslorelin can safely suppress testosterone secretion in male rhesus macaques, but individual variation in onset and duration of action should be considered when establishing reimplantation time points and potential return to reproductive activity.

Gold-Catalyzed Cyclization of Yndiamides with Isoxazoles via α-Imino Gold Fischer Carbenes.

Gold catalysis is an important method for alkyne functionalization. Here we report the gold-catalyzed formal [3+2] aminative cyclization of yndiamides and isoxazoles in a direct synthesis of polysubstituted diaminopyrroles, which are important motifs in drug discovery. Key to this process is the formation, and subsequent cyclization, of an α-imino gold Fischer carbene, which represents a new type of gold carbene intermediate. The reaction proceeds rapidly under mild conditions, with high regioselectivity being achieved by introducing a subtle steric bias between the nitrogen substituents on the yndiamide. DFT calculations revealed that the key to this regioselectivity was the interconversion of isomeric gold keteniminiun ions via a low-barrier π-complex transition state, which establishes a Curtin-Hammett scenario for isoxazole addition. By using benzisoxazoles as substrates, the reaction outcome could be switched to a formal [5+2] cyclization, leading to 1,4-oxazepines.

On the basis of sex: male vs. female rat adenosine A1/A2A receptor affinity.

OBJECTIVE: To ensure reproducibility in biomedical research, the biological variable sex must be reported; yet a reason for using male (instead of female) rodents is seldom given. In our search for novel adenosine receptor ligands, our research group routinely determines a test compound's binding affinities at male Sprague-Dawley rat (r) adenosine A1 and A2A receptors via in vitro radioligand binding studies. This pilot study compared the binding affinities of four adenosine receptor ligands (frequently used as reference standards) at male and female adenosine rA1 and rA2A receptors. RESULTS: The inhibition constant (Ki) values determined using female rats correspond well to the values obtained using male rats and no markable difference could be observed in affinity and selectivity of reference standards. For example, DPCPX the selective adenosine A1 receptor antagonist: male rA1Ki: 0.5 ± 0.1 nM versus female rA1Ki: 0.5 ± 0.03 nM; male rA2AKi: 149 ± 23 nM versus female rA2AKi: 135 ± 29 nM. From the limited data at hand, we conclude that even when using female rats for in vitro studies without regard for the oestrous cycle, the obtained data did not vary much from their male counterparts.

In vitro trypanocidal activities and structure-activity relationships of ciprofloxacin analogs.

Tropical diseases, such as African trypanosomiasis, by their nature and prevalence lack the necessary urgency regarding drug development, despite the increasing need for novel, structurally diverse antitrypanosomal drugs, using different mechanisms of action that would improve drug efficacy and safety. Traditionally antibacterial agents, the fluoroquinolones, reportedly possess in vitro trypanocidal activities against Trypanosoma brucei organisms. During our research, the fluroquinolone, ciprofloxacin (1), and its analogs (2-24) were tested against bloodstream forms of T. brucei brucei, T. b. gambiense, T. b. rhodesiense, T. evansi, T. equiperdum, and T. congolense and Madin-Darby bovine kidney cells (cytotoxicity). Ciprofloxacin [CPX (1)] demonstrated selective trypanocidal activity against T. congolense (IC50 7.79 µM; SI 39.6), whereas the CPX derivatives (2-10) showed weak selective activity (25 < IC50 < 65 µM; 2 < SI < 4). Selectivity and activity of the CPX and 1,2,3-triazole (TZ) hybrids (11-24) were governed by their chemical functionality at C-3 (carboxylic acid, or 4-methylpiperazinyl amide) and their electronic effect (electron-donating or electron-withdrawing para-benzyl substituent), respectively. Trypanocidal hits in the micromolar range were identified against bloodstream forms of T. congolense [CPX (1); CPX amide derivatives 18: IC50 8.95 µM; SI 16.84; 22: IC50 5.42 µM; SI 25.2] and against T. brucei rhodesiense (CPX acid derivative 13: IC50 4.51 µM; SI 10.2), demonstrating more selectivity toward trypanosomes than mammalian cells. Hence, the trypanocidal hit compound 22 may be optimized by retaining the 4-methylpiperazine amide functional group (C-3) and the TZ moiety at position N-15 and introducing other electron-withdrawing ortho-, meta-, and/or para-substituents on the aryl ring in an effort to improve the pharmacokinetic properties and increase the trypanocidal activity. Structure-activity relationships of ciprofloxacin-1,2,3-triazole hybrids were governed by the chemical functionality at C-3 and electronic effect.