Microscopic Raman illustrating antitumor enhancement effects by the combination drugs of γ-secretase inhibitor and cisplatin on osteosarcoma cells.

By using Raman microspectroscopy, it aims to elucidate the cellular variations caused by the combination drug of γ-secretase inhibitor (DAPT) and cisplatin in osteosarcoma (OS) cells. Illustrated by the obtained results of spectral analysis, the intracellular composition significantly changed after combined drug actions compared to the solo DAPT treatment, indicating the synergistic effect of DAPT combined with cisplatin on OS cells. Meanwhile, multivariate curve resolution-alternating least squares (MCR-ALS) algorithm was utilized to address the biochemical constitution changes in all investigated groups including the untreated (UT), DAPT (40D) and combined drug (40D + 20C) treated cells. K-means cluster and univariate imaging were both utilized to visualize the changes in subcellular morphology and biochemical distribution. The presented study provides a unique understanding on the cellular responses to DAPT combined with cisplatin from the natural biochemical perspectives, and laids an experimental foundation for exploring the therapeutic strategies of other combined anticancer drugs in cancer cell model.

An Update on the Biologic Effects of Fenbendazole.

Fenbendazole remains the drug of choice to treat pinworm infection in laboratory rodents. When fenbendazole was last reviewed (15 y ago), the literature supported the drug's lack of toxic effects at therapeutic levels, yet various demonstrated physiologic effects have the potential to alter research outcomes. Although more recent reports continue to reflect an overall discordancy of results, several studies support the premise that fenbendazole affects the bone marrow and the immune system. No effects on reproduction were reported in an extensive study that assessed common treatment protocols in mice, and food intake was unchanged in rats. Behavioral studies are sparse, with only a single report of a subtle change in a rotarod performance in mice. Notably, unexpected results in tumor models during facility treatment with fenbendazole have prompted preclinical and clinical studies of the potential roles of benzimidazoles in cancer.

Emerging insights on functions of the anthelmintic flubendazole as a repurposed anticancer agent.

While flubendazole has been used as a macrofilaricide in humans and animals for some 40 years, work in vitro and in preclinical models over the last decade has suggested its potential use as an anticancer agent. This article reviews recent studies in a range of tumor types indicating novel functions for flubendazole in its control of processes associated with tumor growth, spread and renewal including ferroptosis, autophagy, cancer stem-like cell killing and suppression of intratumoral myeloid-derived suppressor cell accumulation and programmed cell death protein 1. Flubendazole's potential use in clinical oncology will require further understanding of its mechanistic roles, range of inhibition of cancer types, capacity for adjunctive therapy and possible reformulation for enhanced solubility, bioavailability and potency.

Effects of management of infection source of echinococcosis in Linzhi, Tibet Autonomous Region of China.

BACKGROUND: Echinococcosis is highly endemic in western and northern China. Tibet Autonomous Region (TAR) is the most serious prevalent area. Linzhi is located in southeastern part of TAR. Dogs are the primary infection source for the transmission of echinococcosis to humans. A control and prevention campaign based on dog management has been implemented in the past three years. This study aims to evaluate the effects of dog management on the infection rate of dogs. METHODS: Data of dog population, registration and de-worming of seven counties/district in Linzhi between 2017 and 2019 were obtained from the annual prevention and control report. Domestic dog fecal samples were collected from each endemic town of seven counties/district in Linzhi in 2019 to determine the infection of domestic dogs using coproantigen enzyme-linked immunosorbent assay (ELISA). Data analysis was processed using SPSS statistics to compare dog infection rate between 2016 and 2019 by chi-square test, and maps were mapped using ArcGIS. RESULTS: In Linzhi, domestic dog population has decreased from 17 407 in 2017 to 12 663 in 2019, while the registration rate has increased from 75.9% in 2017 to 98.6% in 2019. Similarly, stray dog population has decreased from 14 336 in 2017 to 11 837 in 2019, while sheltered rate has increased from 84.6% in 2017 to 96.6% in 2019. Dog de-worming frequency has increased from 4 times per annum in 2017 to 12 times in 2019, indicating that approximately every dog was dewormed monthly. A total of 2715 dog fecal samples were collected for coproantigen ELISA assay. The dog infection rate was 2.8% (77/2715) in 2019, which was significantly lower than 7.3% (45/618) in 2016 (P < 0.05). CONCLUSIONS: Increased dog registration, decreased dog population, and increased dog de-worming frequency contributed to significantly decrease the dog infection rate in Linzhi. Control and prevention campaign based on dog management could significantly decrease dog infection with Echinococcus spp. in echinococcosis endemic areas.

Health assessment of wild speckled dwarf tortoises, CHERSOBIUS SIGNATUS.

BACKGROUND: In free-ranging reptile populations, bacterial, fungal, viral and parasitic pathogens may affect hosts through impairment in movements, thermoregulation, reproduction, survival, and population dynamics. The speckled dwarf tortoise (Chersobius [Homopus] signatus) is a threatened species that is mostly restricted to the Succulent Karoo biome in South Africa, and little information on pathogens of this species is available yet. We derived baseline parameters for five males and five females that were captured to genetically enhance a conservation breeding program in Europe. Upon collection of the tortoises, ticks were removed and identified. Immediately upon arrival in Europe, ocular, nasal, oral and cloacal swabs were taken for viral, bacteriological and mycological examinations. Fecal samples were collected before and 1 month after fenbendazole treatment, and analyzed for parasites. A panel of PCR, aiming to detect herpesviruses, adenoviruses and iridoviruses, was carried out. RESULTS: Samples were negative for viruses, while bacteriological examination yielded detectable growth in 82.5% of the swabs with a mean load of 16 × 107 ± 61 × 108 colony forming units (CFU) per swab, representing 34 bacterial species. Cloacal and oral swabs yielded higher detectable growth loads than nasal and ocular swabs, but no differences between sexes were observed. Fungi and yeasts (mean load 5 × 103 ± 13 × 103 CFU/swab) were detected in 25% of the swabs. All pre-treatment fecal samples were positive for oxyurid eggs, ranging from 200 to 2400 eggs per gram of feces, whereas after the treatment a significantly reduced egg count (90-100% reduction) was found in seven out of 10 individuals. One remaining individual showed 29% reduction, and two others had increased egg counts. In five tortoises, Nycthocterus spp. and coccidian oocysts were also identified. Soft ticks were identified as Ornithodoros savignyi. CONCLUSIONS: Our baseline data from clinically healthy individuals will help future studies to interpret prevalences of microorganisms in speckled dwarf tortoise populations. The study population did not appear immediately threatened by current parasite presence.

Pre-clinical modeling of cutaneous melanoma.

Metastatic melanoma is challenging to manage. Although targeted- and immune therapies have extended survival, most patients experience therapy resistance. The adaptability of melanoma cells in nutrient- and therapeutically-challenged environments distinguishes melanoma as an ideal model for investigating therapy resistance. In this review, we discuss the current available repertoire of melanoma models including two- and three-dimensional tissue cultures, organoids, genetically engineered mice and patient-derived xenograft. In particular, we highlight how each system recapitulates different features of melanoma adaptability and can be used to better understand melanoma development, progression and therapy resistance.

Urinary capillariosis in a cat from Rio de Janeiro, Brazil - Clinical, morphological and phylogenetic characterization.

Capillaria spp. infections of the urinary tract of domestic carnivores are uncommon worldwide. Infections are rarely diagnosed and are typically asymptomatic. This study aimed to evaluate a case of capillariosis in a cat from the state of Rio de Janeiro, Brazil. A seven-year-old female cat with apathy and reduced appetite was presented. Urine analysis revealed C. plica eggs in urine sediment, and cystitis was evidenced by the presence of bacteria, pyuria, proteinuria and hematuria. The subject was treated with 50 mg/kg fenbendazole for five days. Urine samples were frozen for molecular analysis and species confirmation. Polymerase chain reaction for amplification of the 18S rRNA gene followed by sequencing confirmed the occurrence of Capillaria sp. There has been limited phylogenetic study of Capillaria spp. in cats, so further studies are needed to identify the species present in different locations and associated with feline pathogenesis.

Clinical investigations and treatment outcome in a European wildcat (Felis silvestris silvestris) infected by cardio-pulmonary nematodes.

Recently, feline cardio-pulmonary nematodes have attracted high scientific interest, as they are increasingly reported from various areas. Most of these parasites have similar transmission patterns and/or host reservoirs, thus they may affect domestic and wild felids living in sympatry. In the present study, a case of multiple cardio-pulmonary parasitism in co-infection with other parasites in a European wildcat is presented. The animal, found exhausted, was hospitalised for recovery and parasitological, haematological, clinical and imaging examinations were performed. The parasitological examinations revealed 4 cardio-pulmonary nematodes, i.e. Aelurostrongylus abstrusus, Troglostrongylus brevior, Eucoleus aerophilus, Angiostrongylus chabaudi, 3 intestinal parasites, i.e. Toxocara cati, ancylostomatids, Cystoisospora felis, 2 haemoparasites, i.e. Hepatozoon felis and elements morphologically compatible with small Babesia/Cytauxzoon spp., and Ixodes ricinus and Haemaphysalis erinacei ticks. Treatment with a spot-on formulation containing imidacloprid 10% and moxidectin 1% (Advocate® spot-on solution for cats, Bayer) was decided and follow-up faecal examinations were performed until the release of the animal. By the end of the hospitalisation, all metazoan endoparasites were no longer detectable in faecal examinations, with the exception of a low number of A. abstrusus larvae. Thus, the animal was released after a second treatment with the same product. This is the first description of an apparently successful treatment of multiple cardio-respiratory parasitosis in a naturally infected wildcat showing compatible clinical signs. The evidence that Advocate® may be effective against A. chabaudi could be useful for treating infected, hospitalised, wildcats and it is promising in the case A. chabaudi infection will spread to domestic cats in a near future.

Multiple resistance in equine cyathostomins: a case study from military establishments in Rio Grande do Sul, Brazil / Múltipla resistência de ciatostomíneos de equinos: um estudo de caso em estabelecimentos militares no Rio Grande do Sul, Brasil

Abstract Semi-intensive equine breeding system favors gastrointestinal nematode infections. The treatment of these infections is based on the use of anthelmintics. However, the inappropriate use of these drugs has led to parasitic resistance to the available active principles. The objective of this study was to evaluate the efficacy of the main classes of antiparasitic (ATP) used in control in adult and young animals, including: benzimidazoles (fenbendazole), pyrimidines (pyrantel pamoate), macrocyclic lactones (ivermectin and moxidectin), as well as the combination of active ingredients (ivermectin + pyrantel pamoate). The study was carried out in two military establishments, located in Rio Grande do Sul (RS), from January to December, 2018. The intervals between the treatments of the animals were performed from 30 to 90 days. Coproparasitological evaluations were determined by the egg count reduction in the faeces. Cyatostomine larvae were identified in pre and post-treatment cultures. The results demonstrated the multiple parasitic resistance of cyathostomins to fenbendazole, moxidectin in young animals, and to fenbendazole, pyrantel pamoate in adult animals. Thus, it is necessary to define or diagnose parasitic resistance to assist in the creation of prophylactic parasitic control, using suppressive treatment with ATP associated with integrated alternatives. The progress of parasitic resistance can be slowed.

Resumo O sistema semi-intensivo de criação de equinos favorece infecções por nematoides gastrointestinais. O tratamento dessas infecções é baseado no uso de anti-helmínticos. No entanto, o uso inadequado desses medicamentos levou à resistência parasitária aos princípios ativos disponíveis. O objetivo deste estudo foi avaliar a eficácia das principais classes de antiparasitários (ATP), utilizados no controle em animais adultos e jovens, incluindo: benzimidazois (fenbendazol), pirimidinas (pamoato de pirantel), lactonas macrocíclicas (ivermectina e moxidectina), bem como a combinação de ingredientes ativos (ivermectina + pamoato de pirantel). O estudo foi realizado em dois estabelecimentos militares, localizados no Rio Grande do Sul (RS), de janeiro a dezembro de 2018. Os intervalos entre os tratamentos foram realizados de 30 a 90 dias. As avaliações coproparasitológicas foram determinadas pela redução da contagem de ovos nas fezes. Foram identificadas larvas de ciatostomíneos nas coproculturas pré e pós tratamentos. Os resultados demonstraram a resistência parasitária múltipla dos ciatostomíneos ao fenbendazol, moxidectina em animais jovens, febendazole e pamoato de pirantel em animais adultos. Estabelecer o diagnóstico da resistência parasitária auxiliará na elaboração de um controle parasitário profilático, reduzindo o tratamento supressivo com ATP juntamente com alternativas de controle integrado. Dessa forma, o avanço da resistência parasitária poderá ser retardado.

Flubendazole demonstrates valid antitumor effects by inhibiting STAT3 and activating autophagy.

BACKGROUND: Signal transducer and activator of transcription 3 (STAT3) is an oncogene, which upregulates in approximately 70% of human cancers. Autophagy is an evolutionarily conserved process which maintains cellular homeostasis and eliminates damaged cellular components. Moreover, the STAT3 signaling pathway, which may be triggered by cancer cells, has been implicated in the autophagic process. METHODS: In this study, we found that the anthelmintic flubendazole exerts potent antitumor activity in three human colorectal cancer (CRC) cell lines and in the nude mouse model. The inhibition of cell proliferation in vitro by flubendazole was evaluated using a clonogenic assay and the MTT assay. Western blot analysis, flow cytometry analysis, siRNA growth experiment and cytoplasmic and nuclear protein extraction were used to investigate the mechanisms of inhibiting STAT3 signaling and activation of autophagy induced by flubendazole. Additionally, the expression of STAT3 and mTOR was analyzed in paired colorectal cancer and normal tissues collected from clinical patients. RESULTS: Flubendazole blocked the IL6-induced nuclear translocation of STAT3, which led to inhibition of the transcription of STAT3 target genes, such as MCL1, VEGF and BIRC5. In addition, flubendazole also reduced the expression of P-mTOR, P62, BCL2, and upregulated Beclin1 and LC3-I/II, which are major autophagy-related genes. These processes induced potent cell apoptosis in CRC cells. In addition, flubendazole displayed a synergistic effect with the chemotherapeutic agent 5-fluorouracil in the treatment of CRC. CONCLUSIONS: Taken together, these results indicate that flubendazole exerts antitumor activities by blocking STAT3 signaling and inevitably affects the autophagy pathway. Flubendazole maybe a novel anticancer drug and offers a distinctive therapeutic strategy in neoadjuvant chemotherapy of CRC.

The efficacy of an alternative mebendazole formulation in mice infected with Echinococcus multilocularis.

This study was conducted to investigate the efficacy of a new formulation of MBZ oily suspension (MBZ-OS) in experimentally Echinococcus multilocularis-infected mice. MBZ-OS was prepared and administered to mice infected with E. multilocularis at 12.5 and 25 mg/kg for 14 consecutive days. Then, the cysts were collected, weighed and histologically examined. The results showed that the reduction rate of cyst weight induced by MBZ-OS at two doses was 95.23% and 92.67%, which was significantly higher than that of MBZ-1% tragacanth (positive control) at corresponding concentrations (87.41% and 69.47%), indicating that the treatment of alveolar echinococcosis at lower doses could be achieved by the use of MBZ-OS. This finding shows that MBZ-OS is also a promising formulation for alveolar echinococcosis as well as cystic echinococcosis and deserves to be investigated in clinical applications against echinococcosis.

Phomaketide A Inhibits Lymphangiogenesis in Human Lymphatic Endothelial Cells.

Lymphangiogenesis is an important biological process associated with cancer metastasis. The development of new drugs that block lymphangiogenesis represents a promising therapeutic strategy. Marine fungus-derived compound phomaketide A, isolated from the fermented broth of Phoma sp. NTOU4195, has been reported to exhibit anti-angiogenic and anti-inflammatory effects. However, its anti-lymphangiogenic activity has not been clarified to date. In this study, we showed that phomaketide A inhibited cell growth, migration, and tube formation of lymphatic endothelial cells (LECs) without an evidence of cytotoxicity. Mechanistic investigations revealed that phomaketide A reduced LECs-induced lymphangiogenesis via vascular endothelial growth factor receptor-3 (VEGFR-3), protein kinase Cδ (PKCδ), and endothelial nitric oxide synthase (eNOS) signalings. Furthermore, human proteome array analysis indicated that phomaketide A significantly enhanced the protein levels of various protease inhibitors, including cystatin A, serpin B6, tissue factor pathway inhibitor (TFPI), and tissue inhibitor matrix metalloproteinase 1 (TIMP-1). Importantly, phomaketide A impeded tumor growth and lymphangiogenesis by decreasing the expression of LYVE-1, a specific marker for lymphatic vessels, in tumor xenograft animal model. These results suggest that phomaketide A may impair lymphangiogenesis by suppressing VEGFR-3, PKCδ, and eNOS signaling cascades, while simultaneously activating protease inhibitors in human LECs. We document for the first time that phomaketide A inhibits lymphangiogenesis both in vitro and in vivo, which suggests that this natural product could potentially treat cancer metastasis.

Cell-Free DNA Next-Generation Sequencing Prediction of Response and Resistance to Third-Generation EGFR Inhibitor.

INTRODUCTION: The genomic alterations driving resistance to third-generation EGFR tyrosine kinase inhibitors (TKIs) are not well established, and collecting tissue biopsy samples poses potential complications from invasive procedures. Cell-free circulating DNA (cfDNA) testing provides a noninvasive approach to identify potentially targetable mechanisms of resistance. Here we utilized a 70-gene cfDNA next-generation sequencing test to interrogate pretreatment and progression samples from 77 EGFR-mutated non-small cell lung cancer (NSCLC) patients treated with a third-generation EGFR TKI. PATIENTS AND METHODS: Rociletinib was evaluated in advanced or metastatic (second line or higher) disease with EGFR T790M-positive NSCLC in the TIGER-X (NCT01526928) and TIGER-2 (NCT02147990) studies. Plasma samples were collected at baseline and at the time of systemic progression while receiving rociletinib. The critical exons in 70 genes were sequenced in cfDNA isolated from plasma samples to elucidate a comprehensive genomic profile of alterations for each patient. RESULTS: Plasma-based cfDNA analysis identified 93% of the initial EGFR activating and 85% of the EGFR T790M resistance mutations in pretreatment samples with detectable tumor DNA. Profiling of progression samples revealed significant heterogeneity, with different variant types (eg, mutations, amplifications, and fusions) detected in multiple genes (EGFR, MET, RB1) that may be driving resistance in patients. Novel alterations not previously described in association with resistance to third-generation TKIs were also detected, such as an NTRK1 fusion. CONCLUSION: cfDNA next-generation sequencing identified initial EGFR activating and secondary T790M resistance mutations in NSCLC patients with high sensitivity, predicted treatment response equivalent to tissue analysis, and identified multiple novel and established resistance alterations.

A Young Man With Hemoptysis and Cavitary Lung Lesions.

CASE PRESENTATION: A man in his 20s presented with 2 months of mild fatigue and intermittent hemoptysis of less than a tablespoon per episode. He was previously healthy and was on no medications. He denied fevers, night sweats, weight loss, wheezing, dyspnea, musculoskeletal symptoms, and rashes. He had emigrated from a South American country to the United States 3 years earlier. He worked as a groundskeeper but had no exposures to animals, mold, or dusts. He reported rare prior cigarette smoking with no history of alcohol or drug use. He was unsure whether he had received the Bacillus Calmette-Guérin vaccine.

Control of sheep gastrointestinal nematodes using the combination of Duddingtonia flagrans and Levamisole Hydrochloride 5% / Controle de nematódeos gastrintestinais de ovinos utilizando a combinação de Duddingtonia flagrans e Cloridrato de Levamisole 5%

Abstract The objective was to evaluate the action of D. flagrans pellets in association with Levamisole Hydrochloride 5% for controlling sheep gastrointestinal nematodes in the northeastern Brazil. Three groups of six sheep each were formed: group 1 received 3 g of the pellets (0.6 g of D. flagrans mycelium) for each 10 kg b.w., twice a week for six months, and deworming with Levamisole Hydrochloride 5% when EPG ≥ 1500; group 2 received a dosage of Levamisole Hydrochloride 5% when EPG ≥ 1500; and group 3 received 3 g of pellets without fungi for each 10 kg b.w., twice a week for six months. EPG counts, larval cultures, packed cell volume (PCV) and weighing were performed every 15 days; monthly, samples of grass from each paddock were collected. The mean EPG of the groups began to statistically differ from day 30 (p < 0.05). Group 1 required less deworming with Levamisole Hydrochloride 5% and showed superiority of PCV values ​​throughout the experiment (p < 0.05). There was a significant reduction (p < 0.05) in L3 recovery in the group 1 paddock from day 30 onwards. The use of D. flagrans pellets in association with Levamisole Hydrochloride 5% was effective for controlling gastrointestinal nematodes.

Resumo O objetivo foi avaliar a ação de péletes de Duddingtonia flagrans em associação ao Cloridrato de Levamisole 5% no controle de nematódeos gastrintestinais de ovinos no Nordeste do Brasil. Foram formados três grupos de seis animais cada: grupo 1 recebeu 3 g de péletes (0,6 g de micélio de D. flagrans) para cada 10 kg p.v., duaz vezes por semana durante seis meses, e vermifugações com Cloridrato de Levamisole 5% quando OPG > 1500; grupo 2 recebeu uma dosagem de Cloridrato de Levamisole 5% quando OPG ≥ 1500; e grupo 3 recebeu 3 g de péletes sem fungos para cada 10 kg de p.v., duas vezes por semana durante seis meses. Contagens de OPG, coproculturas, de volumes globulares (VG) e pesagens foram realizadas a cada 15 dias. Mensalmente, amostras de pasto de cada piquete eram coletadasa. A média de OPG dos grupos começou a diferir estatisticamente a partir do dia 30 (p < 0,05). O grupo 1 necessitou de menos vermifugações com Cloridrato de Levamisole 5% e demonstrou superioridade nos valores de VG durante todo o experimento (p < 0,05). Houve redução significativa (p < 0,05) nas L3 recuperadas no piquete do grupo 1 a partir do dia 30. Em conclusão, a utilização de péletes de D. flagrans em associação ao Cloridrato de Levamisole 5% foi eficaz no controle de nematódeos gastrintestinais de ovinos.