Chronic intestinal inflammation significantly contributes to the development of colorectal cancer and remains a pertinent clinical challenge, necessitating novel therapeutic approaches. Indole-based microbial metabolite mimics Felix Kopp Kortagere 6 (FKK6), which is a ligand and agonist of the pregnane X receptor (PXR), was recently demonstrated to have PXR-dependent anti-inflammatory and protective effects in a mouse model of dextran sodium sulfate (DSS)-induced acute colitis. Here, we examined the therapeutic potential of FKK6 in a mouse model (C57BL/6 FVB humanized PXR mice) of colitis-associated colon cancer (CAC) induced by azoxymethane and DSS. FKK6 (2 mg/kg) displayed substantial antitumor activity, as revealed by reduced size and number of colon tumors, improved colon histopathology, and decreased expression of tumor markers (c-MYC, β-catenin, Ki-67, and cyclin D) in the colon. In addition, we carried out a chronic toxicity (30 days) assessment of FKK6 (1 mg/kg and 2 mg/kg) in C57BL/6 mice. Histological examination of tissues, biochemical blood analyses, and immunohistochemical staining for Ki-67 and γ-H2AX showed no difference between FKK6-treated and control mice. Comparative metabolomic analyses in mice exposed for 5 days to DSS and administered with FKK6 (0.4 mg/kg) revealed no significant effects on several classes of metabolites in the mouse fecal metabolome. Ames and micronucleus tests showed no genotoxic and mutagenic potential of FKK6 in vitro. In conclusion, anticancer effects of FKK6 in azoxymethane/DSS-induced CAC, together with FKK6 safety data from in vitro tests and in vivo chronic toxicity study, and comparative metabolomic study, are supportive of the potential therapeutic use of FKK6 in the treatment of CAC. SIGNIFICANCE STATEMENT: Microbial metabolite mimicry proposes that chemical mimics of microbial metabolites that serve to protect hosts against aberrant inflammation in the gut could serve as a new paradigm for the development of drugs targeting inflammatory bowel disease if, like the parent metabolite, is devoid of toxicity but more potent against the microbial metabolite receptor. We identified a chemical mimic of Felix Kopp Kortagere 6, and we propose that Felix Kopp Kortagere 6 is devoid of toxicity yet significantly reduces tumor formation in an azoxymethane-dextran sodium sulfate model of murine colitis-induced colon cancer.
- MeSH
- Azoxymethane toxicity MeSH
- Chronic Disease MeSH
- Indoles pharmacology therapeutic use MeSH
- Colitis drug therapy chemically induced metabolism pathology MeSH
- Colorectal Neoplasms * drug therapy metabolism pathology MeSH
- Disease Models, Animal * MeSH
- Molecular Mimicry MeSH
- Mice, Inbred C57BL * MeSH
- Mice MeSH
- Colitis-Associated Neoplasms pathology drug therapy metabolism MeSH
- Dextran Sulfate toxicity MeSH
- Inflammation drug therapy metabolism MeSH
- Animals MeSH
- Check Tag
- Male MeSH
- Mice MeSH
- Female MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
BACKGROUND: Over the past two decades, the global incidence of gout has markedly increased, affecting people worldwide. Considering the side effects of xanthine oxidase (XO) inhibitor drugs (e.g. allopurinol and febuxostat) used in the treatment of hyperuricemia and gout, the potential application of phytochemicals has been widely studied. In addition, XO also takes part in the elimination of certain drugs, including 6-mercaptopurine. In the current explorative study, we aimed to examine the potential effects of tea catechins, resveratrol, silymarin flavonolignans and some of their conjugated metabolites on XO-catalyzed xanthine and 6-mercaptopurine oxidation, applying in vitro assays and modeling studies. RESULTS: Catechins, resveratrol and resveratrol conjugates exerted no or only weak inhibitory effects on XO. Silybin A, silybin B and isosilybin A were weak, silychristin was a moderate, while 2,3-dehydrosilychristin was a potent inhibitor of the enzyme. Sulfate metabolites of silybin A, silybin B and isosilybin A were considerably stronger inhibitors compared to the parent flavonolignans, and the sulfation of 2,3-dehydrosilychristin slightly increased its inhibitory potency. Silychristin was the sole flavonolignan tested, where sulfate conjugation decreased its inhibitory effect. CONCLUSION: 2,3-Dehydrosilychristin seems to be a promising candidate for examining its in vivo antihyperuricemic effects, because both the parent compound and its sulfate conjugate are highly potent inhibitors of XO. © 2024 The Author(s). Journal of the Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.
- MeSH
- Enzyme Inhibitors * chemistry pharmacology MeSH
- Catalysis MeSH
- Catechin * chemistry analogs & derivatives pharmacology MeSH
- Humans MeSH
- Mercaptopurine * chemistry pharmacology metabolism MeSH
- Oxidation-Reduction * MeSH
- Resveratrol * chemistry pharmacology MeSH
- Silymarin * pharmacology chemistry MeSH
- Xanthine chemistry metabolism pharmacology MeSH
- Xanthine Oxidase * antagonists & inhibitors metabolism chemistry MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
Bacillus is well known for producing a wide range of compounds that inhibit microbial phytopathogens. From this perspective, we were interested in evaluating the biocontrol potential of 5 plant growth-promoting rhizobacteria Bacillus species (PGPR-Bacillus) on 21 microbial pectinolytic plant pathogens isolated from previous studies. Phytopathogenicity and in vivo biocontrol potential of PGPR curative and preventive treatments were investigated from this angle. Overall, the pathogenicity test on healthy tomato, zucchini, and mandarin showed low rot to no symptoms for all PGPR strain culture treatments. Conversely, zucchini pre-treated with PGPR strains B. circulans and B. cereus for 72 h showed no signs of soft rot and remained healthy when in vitro contaminated with phytopathogens (Neisseria cinerea and Pichia anomala). Additionally, the PGPR-Bacillus strains were shown to be effective in mitigating the symptoms of soft rot in tomatoes, zucchini, and oranges using in vivo curative treatment. It is true that the majority of pectinolytic phytopathogenic strains exhibited antibiotic resistance. In vivo tests revealed that PGPR-Bacillus cell culture was effective against plant pathogens. Thus, PGPR-Bacillus can be considered a potential biocontrol agent for pectinolytic plant pathogens.
- MeSH
- Antibiosis * MeSH
- Bacillus * physiology MeSH
- Pest Control, Biological * methods MeSH
- Biological Control Agents * MeSH
- Citrus microbiology growth & development MeSH
- Plant Diseases * microbiology prevention & control MeSH
- Pectins metabolism MeSH
- Soil Microbiology MeSH
- Solanum lycopersicum microbiology growth & development MeSH
- Plant Development MeSH
- Publication type
- Journal Article MeSH
The alveolar-capillary interface is the key functional element of gas exchange in the human lung, and disruptions to this interface can lead to significant medical complications. However, it is currently challenging to adequately model this interface in vitro, as it requires not only the co-culture of human alveolar epithelial and endothelial cells but mainly the preparation of a biocompatible scaffold that mimics the basement membrane. This scaffold should support cell seeding from both sides, and maintain optimal cell adhesion, growth, and differentiation conditions. Our study investigates the use of polycaprolactone (PCL) nanofibers as a versatile substrate for such cell cultures, aiming to model the alveolar-capillary interface more accurately. We optimized nanofiber production parameters, utilized polyamide mesh UHELON as a mechanical support for scaffold handling, and created 3D-printed inserts for specialized co-cultures. Our findings confirm that PCL nanofibrous scaffolds are manageable and support the co-culture of diverse cell types, effectively enabling cell attachment, proliferation, and differentiation. Our research establishes a proof-of-concept model for the alveolar-capillary interface, offering significant potential for enhancing cell-based testing and advancing tissue-engineering applications that require specific nanofibrous matrices.
Personalizace a individualizace terapie a diagnostiky pomocí pokročilých in vitro modelů využívajících tkáň pacienta je budoucností klinické medicíny. Tyto modely zahrnují organoidy, tedy trojrozměrné in vitro modely odvozené z dospělých nebo indukovaných pluripotentních kmenových buněk, modely orgánů na čipu a další kombinované modely. Kmenové buňky pro založení těchto kultur lze získat od pacienta za pomocí minimálně invazivních metod a expandovat in vitro pro následná personalizovaná využití. Tento přehledový článek shrnuje charakteristiky jednotlivých přístupů a jejich potenciální využití zejména v transplantační terapii diabetu mellitu a diabetického onemocnění ledvin. Oproti současným metodám transplantace pankreatu a pankreatických ostrůvků, respektive transplantace ledvin, nabízí tyto přístupy řadu výhod. Na závěr jsou prezentována současná klinická využití těchto modelů v prediktivním testování léčebné odpovědi a transplantační terapii s prvními klinickými studiemi využívajícími tyto modely v kauzální léčbě diabetu mellitu.
Personalization and individualization of therapy and diagnostics using advanced in vitro models utilizing patient tissue is the future of clinical medicine. These models include organoids, i.e., three-dimensional in vitro models derived from adult or induced pluripotent stem cells, organ-on-a-chip models, and other combination models. Stem cells for establishing these cultures can be obtained from the patient using minimally invasive methods and expanded in vitro for subsequent personalized applications. This review article summarizes the characteristics of each approach and their potential applications, particularly in transplantation therapy for diabetes mellitus and diabetic kidney disease. Compared with current approaches to pancreas and pancreatic islet transplantation and kidney transplantation, respectively, these approaches offer a number of potential advantages. In conclusion, the current clinical applications of these models in predictive testing of treatment response and transplantation therapy are presented, with the first clinical trials using these models in the causal treatment of diabetes mellitus.
A Mycobacterium smegmatis transcriptional regulator, MSMEG_5850, and its ortholog in M. tuberculosis, rv0775 were annotated as putative TetR Family Transcriptional Regulators. Our previous study revealed MSMEG_5850 is involved in global transcriptional regulation in M. smegmatis and the presence of gene product supported the survival of bacteria during nutritional starvation. Phylogenetic analysis showed that MSMEG_5850 diverged early in comparison to its counterparts in virulent strains. Therefore, the expression pattern of MSMEG_5850 and its counterpart, rv0775, was compared during various in-vitro growth and stress conditions. Expression of MSMEG_5850 was induced under different environmental stresses while no change in expression was observed under mid-exponential and stationary phases. No expression of rv0775 was observed under any stress condition tested, while the gene was expressed during the mid-exponential phase that declined in the stationary phase. The effect of MSMEG_5850 on the survival of M. smegmatis under stress conditions and growth pattern was studied using wild type, knockout, and supplemented strain. Deletion of MSMEG_5850 resulted in altered colony morphology, biofilm/pellicle formation, and growth pattern of M. smegmatis. The survival rate of wild-type MSMEG_5850 was higher in comparison to knockout under different environmental stresses. Overall, this study suggested the role of MSMEG_5850 in the growth and adaptation/survival of M. smegmatis under stress conditions.
- MeSH
- Bacterial Proteins * genetics metabolism MeSH
- Biofilms growth & development MeSH
- Phylogeny MeSH
- Stress, Physiological * MeSH
- Microbial Viability MeSH
- Mycobacterium smegmatis * genetics growth & development physiology metabolism MeSH
- Gene Expression Regulation, Bacterial MeSH
- Transcription Factors * genetics metabolism MeSH
- Publication type
- Journal Article MeSH
In vitro dissolution testing is commonly performed to ensure that oral solid dosage medicines are of high quality and will achieve their targeted in vivo performance. However, this testing is time and material consuming. Therefore, pharmaceutical companies have been developing predictive dissolution models (PDMs) for drug product release based on fast at- and/or on-line measurements, including real-time release testing of dissolution (RTRT-D). Recently, PDMs have seen acceptance by major regulatory bodies as release tests for the dissolution critical quality attribute. In this paper, several methodologies are described to develop and validate a fit-for-purpose model, then to implement it as a surrogate release test for dissolution. These approaches are further exemplified by real-life case studies, which demonstrate that PDMs for release are not only viable but more sustainable than in vitro dissolution testing and can significantly accelerate drug product release. The rise of continuous manufacturing within the pharmaceutical industry further favors the implementation of real-time release testing. Therefore, a steep uptake of PDMs for release is expected once this methodology is globally accepted. To that end, it is advantageous for global regulators and pharmaceutical innovators to coalesce around a harmonized set of expectations for development, validation, implementation, and lifecycle of PDMs as part of drug product release testing.
BACKGROUND: The treatment of non-small cell lung cancer (NSCLC) patients is correlated with the efficacy of immune checkpoint blockade therapy (ICB) targeting programmed cell death ligand 1 (PD-L1) or its cognate receptor (PD-1) on cancer cells or infiltrating immune cells. Analysis of PD-L1/PD-1 expression in tumor tissue represents a crucial step before PD-L1/PD-1 blocker usage. METHODS: We used directed evolution of protein variants derived from a 13 kDa Myomedin loop-type combinatorial library with 12 randomized amino acid residues to select high-affinity binders of human PD-L1 (hPD-L1). After the ribosome display, individual clones were screened by ELISA. Detailed analysis of binding affinity and kinetics was performed using LigandTracer. The specificity of Myomedins was assessed using fluorescent microscopy on HEK293T-transfected cells and cultured cancer cells in vitro, formalin-fixed paraffin-embedded (FFPE) sections of human tonsils, and FFPE tumor samples of NSCLC patients. RESULTS: Seven identified PD-L1 binders, called MLE, showed positive staining for hPD-L1 on transfected HEK293T cells and cultured MCF-7 cells. MLE031, MLE105, MLE249, and MLE309 exhibited high affinity to both human and mouse PD-L1-transfected HEK293T cells measured with LigandTracer. The diagnostic potential of MLE variants was tested on human tonsillitis tissue and compared with diagnostic anti-PD-L1 antibody DAKO 28-8 and PD-L1 IHC 22C3 pharmDx antibody. MLE249 and MLE309 exhibited an excellent overlap with diagnostic DAKO 28-8 (Pearson ́s coefficient (r) = 0.836 and 0.731, respectively) on human tonsils on which MLE309 exhibited also excellent overlap with diagnostic 22C3 antibody (r = 0.876). Using three NSCLC tissues, MLE249 staining overlaps with 28-8 antibody (r = 0.455-0.883), and MLE309 exhibited overlap with 22C3 antibody (r = 0.534-0.619). Three MLE proteins fused with Fc fragments of rabbit IgG, MLE249-rFc, MLE309-rFc and MLE031-rFc, exhibited very good overlap with anti-PD-L1 antibody 28-8 on tonsil tissue (r = 0.691, 0.610, and 0.667, respectively). Finally, MLE249-rFc, MLE309-rFc and MLE031-rFc exhibited higher sensitivity in comparison to IHC 22C3 antibody using routine immunohistochemistry staining system Ventana, which is one of gold standards for PD-L1 diagnosis. CONCLUSIONS: We demonstrated the development of MLE Myomedins specifically recognizing hPD-L1 that may serve as a refinement tool for clinical PD-L1 detection.
Given the high incidence of diet-related diseases, including type 2 diabetes and cancer, there is a growing need to explore new strategies for their prevention. Although polyphenols are known to reduce starch digestibility and lower the in vitro glycemic index, their antioxidant capacity and cytotoxic properties, when complexed with starches, remain underexplored. Therefore, this study aimed to investigate the antioxidant activity, total polyphenol content, and cytotoxic potential of polyphenol-starch complexes formed using common dietary polyphenols-(+)-catechin, epigallocatechin gallate, hesperidin, naringenin, trans-ferulic acid, p-coumaric acid, quercetin, and kaempferol-and widely consumed starches from wheat, rice, potato, and maize. Antioxidant activity (FRAP and DPPH) together with the total polyphenols content (Folin-Ciocalteu) were tested: (1) before (undigested) enzymatic hydrolysis of the tested sample; (2) after (digested) enzymatic hydrolysis of the tested sample and (3) after hydrolysis of the sample and its centrifugation (supernatant). Cytotoxicity against colon cancer (Caco-2, HT29) and normal colon (CCD 841CoN) cell lines were determined in vitro by the MTT method. In undigested samples, the highest antioxidant activity was obtained with the addition of quercetin to wheat, rice, and maize starch (6735.8 μmol Fe2+/g d.m., 678.8, 539.4 μmol Trolox/g d.m., respectively), and epigallocatechin gallate to wheat, rice, potato, and maize starch (692.1, 538.0, 625.8, 573.6 μmol Trolox/g d.m., respectively). In digested samples, the highest antioxidant activity was obtained with the addition of quercetin to wheat and rice starch (2104.5 μmol Fe2+/g d.m., 742.1 μmol Trolox/g d.m., respectively). In the case of the natant of the digested samples, the highest value was recorded for the addition of (+)-catechin to potato starch and trans-ferulic acid to maize starch (823.7 μmol Fe2+/g d.m., 245.1 μmol Trolox/g d.m., respectively). The addition of quercetin to wheat and rice starch and (+)-catechin to potato starch (0.239, 0.151, 0.085 g gallic acid/g d.m., respectively) resulted in the highest total polyphenol content. Furthermore, quercetin demonstrated the most significant level of cytotoxic activity against the tumor cell line Caco-2 (IC50 = 275.6 μg/mL; potato starch). Overall, quercetin was identified as the most significant or one of the most significant for all parameters evaluated.
- MeSH
- Antioxidants * pharmacology chemistry MeSH
- HT29 Cells MeSH
- Caco-2 Cells MeSH
- Catechin analogs & derivatives MeSH
- Zea mays chemistry MeSH
- Humans MeSH
- Cell Line, Tumor MeSH
- Polyphenols * chemistry pharmacology MeSH
- Oryza chemistry MeSH
- Starch * chemistry pharmacology MeSH
- Cell Survival drug effects MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
Drug resistance is a growing problem for many pathogens, including mycobacteria. Small heterocyclic molecules are among the leading scaffolds for developing potential antimycobacterial agents. Therefore, based on the molecular hybridization approach, we have prepared an extensive series of N-substituted 5-(3,5-dinitrophenyl)-1,3,4-oxadiazol-2-amine derivatives. We also investigated their isosteres and acyclic synthetic precursors. The compounds were evaluated for their in vitro activity against Mycobacterium tuberculosis (Mtb) H37Rv, a panel of multidrug- and extensively drug-resistant Mtb isolates and two nontuberculous mycobacterial strains (NTM; M. avium and M. kansasii). The ability to inhibit mycobacterial growth was quantified using minimum inhibitory concentration (MIC) values. Many compounds achieved MIC values ≤ 0.03 μM for NTM and Mtb, regardless of their resistance profile. The highest activity was associated with oxadiazole and thiadiazole scaffolds with benzylamino or C5-C9 alkylamino substitution. The experimentally confirmed mechanism of action of these compounds consists of disruption of mycobacterial cell wall biosynthesis via inhibition of decaprenylphosphoryl-β-D-ribose 2'-epimerase (DprE1). In vitro toxicity evaluation was performed in a hepatocyte model (HepG2), while in vivo toxicity was evaluated using Danio rerio embryos. These findings identify a promising new chemotype with potent, broad-spectrum and selective antimycobacterial activity, including efficacy against resistant strains, and support its further development as a potential therapeutic candidate.
- MeSH
- Antitubercular Agents * pharmacology chemical synthesis chemistry toxicity MeSH
- Zebrafish MeSH
- Humans MeSH
- Microbial Sensitivity Tests MeSH
- Mycobacterium tuberculosis drug effects MeSH
- Oxadiazoles * pharmacology chemical synthesis chemistry MeSH
- Animals MeSH
- Check Tag
- Humans MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
