scholarly journals A versatile genetic control system in mammalian cells and mice responsive to clinically licensed sodium ferulate

2020 ◽  
Vol 6 (32) ◽  
pp. eabb9484
Author(s):  
Yidan Wang ◽  
Shuyong Liao ◽  
Ningzi Guan ◽  
Yuanxiao Liu ◽  
Kaili Dong ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Transgene Expression ◽  
Expression Profiles ◽  
Therapeutic Effects ◽  
Clinical Use ◽  
Sodium Ferulate ◽  
Wide Range ◽  
Transcriptional Switch ◽  
Logic Operations ◽  
Dose Dependent

Dynamically adjustable gene- and cell-based therapies are recognized as next-generation medicine. However, the translation of precision therapies into clinics is limited by lack of specific switches controlled by inducers that are safe and ready for clinical use. Ferulic acid (FA) is a phytochemical with a wide range of therapeutic effects, and its salt sodium ferulate (SF) is used as an antithrombotic drug in clinics. Here, we describe an FA/SF-adjustable transcriptional switch controlled by the clinically licensed drug SF. We demonstrated that SF-responsive switches can be engineered to control CRISPR-Cas9 systems for on-command genome/epigenome engineering. In addition, we integrated FA-controlled switches into programmable biocomputers to process logic operations. We further demonstrated the dose-dependent SF-inducible transgene expression in mice by oral administration of SF tablets. Engineered switches responsive to small-molecule clinically licensed drugs to achieve adjustable transgene expression profiles provide new opportunities for dynamic interventions in gene- and cell-based precision medicine.

Evaluation of the nucleopolyhedrovirus of Anticarsia gemmatalis as a vector for gene therapy in mammals

2020 ◽  
Vol 20 ◽  
Author(s):  
Cintia N. Parsza ◽  
Diego L. Mengual Gómez ◽  
Jorge Alejandro Simonin ◽  
Mariano Nicolás Belaich ◽  
Pablo Daniel Ghiringhelli
Keyword(s):  
Gene Therapy ◽  
Mammalian Cells ◽  
Insect Cells ◽  
Autographa Californica ◽  
Anticarsia Gemmatalis ◽  
Agricultural Pests ◽  
Mammalian Cell Lines ◽  
Delivery Vector ◽  
Wide Range ◽  
Insect Pathogens

Background: Baculoviruses are insect pathogens with important biotechnological applications that transcend their use as biological controllers of agricultural pests. One species, Autographa californica multiple nucleopolhyedrovirus (AcMNPV) has been extensively exploited as a molecular platform to produce recombinant proteins and as a delivery vector for genes in mammals, because it can transduce a wide range of mammalian cells and tissues without replicating or producing progeny. Objective/Method: To investigate if the budded virions of Anticarsia gemmatalis multiple nucleopolhyedrovirus (AgMNPV) species has the same ability, the viral genome was modified by homologous recombination into susceptible insect cells to integrate reporter genes and then it was evaluated on mammalian cell lines in comparative form with respect to equivalent viruses derived from AcMNPV. Besides, the replicative capacity of AgMNPV´s virions in mammals was determined. Results: The experiments carried out showed that the recombinant variant of AgMNPV transduces and support the expression of delivered genes but not replicates in mammalian cells. Conclusion: Consequently, this insect pathogen is proposed as an alternative of non-infectious viruses in humans to explore new approaches in gene therapy and other applications based on the use of mammalian cells.

Synthesis and Antibacterial / Anticancer Activities of Compounds Containing Pyrazole Ring Linked to Piperazines

2020 ◽  
Vol 16 (4) ◽  
pp. 419-431
Author(s):  
Kishore K. Valluri ◽  
Tejeswara R. Allaka ◽  
IV Kasi Viswanath ◽  
Nagaraju PVVS
Keyword(s):  
Cell Line ◽  
Cancer Cell ◽  
Anticancer Agents ◽  
Cancer Cell Line ◽  
Pyrazole Ring ◽  
Therapeutic Effects ◽  
Anticancer Activities ◽  
Human Colon Cancer ◽  
Parent Molecule ◽  
Wide Range

Background: Many pyrazole piperazine derivatives are known to exhibit a wide range, thus being attractive for the drug design and synthesis of interesting class of widely studied heterocyclic compounds. It is therefore necessary to devote continuing effort for the identification and development of New Chemical Entities (NCEs) as potential antibacterial and anticancer agents to address serious health problems. Methods: A series of new compounds containing pyrazole ring linked to a piperazine hydrochloride moiety were synthesized and screened for their antibacterial activity, cytotoxicity of novel scaffolds are described by variation in therapeutic effects of parent molecule. The structure variants were characterized by using a blend of spectroscopic 1H NMR, 13C NMR, IR, Mass and chromatographic techniques. Results: When tested for in vitro antibacterial and anticancer activities, several of these compounds showed good activities. The target compounds 9b, 9a and 9e exhibited a high degree of anticancer activity against human colon cancer cell line Caco-2 and human breast cancer cell line MDAMB231. Further, 9a, 9b, 9d, and 9h showed better activity towards four medically relevant organisms; Staphylococcus aureus, Bacillus subtilis, Escherichia coli and Klebsiella Species compared to CPF. In the present investigation, cheminfomatics tools Molinspiration, 2003 and MolSoft, 2007 for the prediction of insilico molecular properties and drug likeness for the target compounds 9a-h was evaluated and positive results were observed. Conclusion: Our study revealed that the molecular framework presented here could be a useful template for the identification of novel small molecules as promising antibacterial/ anticancer agents.

Cucurbitacins as Anticancer Agents: A Patent Review

2019 ◽  
Vol 14 (2) ◽  
pp. 133-143 ◽  
Author(s):  
Hidayat Hussain ◽  
Ivan R. Green ◽  
Muhammad Saleem ◽  
Khanzadi F. Khattak ◽  
Muhammad Irshad ◽  
...  
Keyword(s):  
Anticancer Agents ◽  
Wide Spectrum ◽  
Biological Effects ◽  
Therapeutic Effects ◽  
Cytotoxic Effects ◽  
Natural Sources ◽  
Oh Groups ◽  
Drug Candidates ◽  
The Past ◽  
Wide Range

Background: Cucurbitacins belong to a group of tetracyclic triterpenoids that display a wide range of biological effects. In the past, numerous cucurbitacins have been isolated from natural sources and many active compounds have been synthesized using the privileged scaffold in order to enhance its cytotoxic effects. Objective: his review covers patents on the therapeutic effects of natural cucurbitacins and their synthetic analogs published during the past decade. By far, the majority of patents published are related to cancer and Structure-Activity Relationships (SAR) of these compounds are included to lend gravitas to this important class of natural products. Methods: The date about the published patents was downloaded via online open access patent databases. Results: Cucurbitacins display significant cytotoxic properties, in particular cucurbitacins B and D which possess very potent effects towards a number of cancer cells. Numerous cucurbitacins isolated from natural sources have been derivatized through chemical modification at the C(2)-OH and C(25)- OH groups. Most importantly, an acyl ester of the C(25)-OH and, iso-propyl, n-propyl and ethyl ether groups of the C(2)-OH demonstrated the most increased cytotoxic activity. Conclusion: The significant cytotoxic effects of natural and semi-synthetic cucurbitacins make them attractive as new drug candidates. Moreover, cucurbitacins have the capability to form conjugates with other anticancer drugs which will synergistically enhance their anticancer effects. The authors believe that in order to get lead compounds, there should be a greater focus on the synthesis of homodimers, heterodimers, and halo derivatives of cucurbitacins. In the opinion of the authors the analysis of the published patents on the cucurbitacins indicates that these compounds can be developed into a regimen to treat a wide spectrum of cancers.

scholarly journals Autophagy Deficiency by Atg4B Loss Leads to Metabolomic Alterations in Mice

Metabolites ◽  
2021 ◽  
Vol 11 (8) ◽  
pp. 481
Author(s):  
Gemma G. Martínez-García ◽  
Raúl F. Pérez ◽  
Álvaro F. Fernández ◽  
Sylvere Durand ◽  
Guido Kroemer ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Amino Acids ◽  
Mammalian Cells ◽  
Tissue Homeostasis ◽  
In Vivo Studies ◽  
Protective Mechanism ◽  
Cardiac Damage ◽  
Wide Range ◽  
First Time

Autophagy is an essential protective mechanism that allows mammalian cells to cope with a variety of stressors and contributes to maintaining cellular and tissue homeostasis. Due to these crucial roles and also to the fact that autophagy malfunction has been described in a wide range of pathologies, an increasing number of in vivo studies involving animal models targeting autophagy genes have been developed. In mammals, total autophagy inactivation is lethal, and constitutive knockout models lacking effectors of this route are not viable, which has hindered so far the analysis of the consequences of a systemic autophagy decline. Here, we take advantage of atg4b−/− mice, an autophagy-deficient model with only partial disruption of the process, to assess the effects of systemic reduction of autophagy on the metabolome. We describe for the first time the metabolic footprint of systemic autophagy decline, showing that impaired autophagy results in highly tissue-dependent alterations that are more accentuated in the skeletal muscle and plasma. These changes, which include changes in the levels of amino-acids, lipids, or nucleosides, sometimes resemble those that are frequently described in conditions like aging, obesity, or cardiac damage. We also discuss different hypotheses on how impaired autophagy may affect the metabolism of several tissues in mammals.

scholarly journals Antifungal activity of dendritic cell lysosomal proteins against Cryptococcus neoformans

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Benjamin N. Nelson ◽  
Savannah G. Beakley ◽  
Sierra Posey ◽  
Brittney Conn ◽  
Emma Maritz ◽  
...  
Keyword(s):  
Antifungal Activity ◽  
Cryptococcus Neoformans ◽  
Mammalian Cells ◽  
Cysteine Proteases ◽  
Dependent Manner ◽  
Life Threatening ◽  
Opportunistic Fungal Pathogen ◽  
Dose Dependent ◽  
Lysosomal Proteins

AbstractCryptococcal meningitis is a life-threatening disease among immune compromised individuals that is caused by the opportunistic fungal pathogen Cryptococcus neoformans. Previous studies have shown that the fungus is phagocytosed by dendritic cells (DCs) and trafficked to the lysosome where it is killed by both oxidative and non-oxidative mechanisms. While certain molecules from the lysosome are known to kill or inhibit the growth of C. neoformans, the lysosome is an organelle containing many different proteins and enzymes that are designed to degrade phagocytosed material. We hypothesized that multiple lysosomal components, including cysteine proteases and antimicrobial peptides, could inhibit the growth of C. neoformans. Our study identified the contents of the DC lysosome and examined the anti-cryptococcal properties of different proteins found within the lysosome. Results showed several DC lysosomal proteins affected the growth of C. neoformans in vitro. The proteins that killed or inhibited the fungus did so in a dose-dependent manner. Furthermore, the concentration of protein needed for cryptococcal inhibition was found to be non-cytotoxic to mammalian cells. These data show that many DC lysosomal proteins have antifungal activity and have potential as immune-based therapeutics.

scholarly journals N-Acetyl Cysteine Overdose Inducing Hepatic Steatosis and Systemic Inflammation in Both Propacetamol-Induced Hepatotoxic and Normal Mice

Antioxidants ◽  
2021 ◽  
Vol 10 (3) ◽  
pp. 442
Author(s):  
Gunn-Guang Liou ◽  
Cheng-Chi Hsieh ◽  
Yi-Ju Lee ◽  
Pin-Hung Li ◽  
Ming-Shiun Tsai ◽  
...  
Keyword(s):  
Inbred Mouse ◽  
Serum Triglyceride ◽  
Mouse Strains ◽  
High Dose ◽  
Therapeutic Effects ◽  
Protein Nitration ◽  
Cholesterol Levels ◽  
Antioxidative Enzyme Activities ◽  
Acetyl Cysteine ◽  
Dose Dependent

Acetaminophen (APAP) overdose induces acute liver damage and even death. The standard therapeutic dose of N-acetyl cysteine (NAC) cannot be applied to every patient, especially those with high-dose APAP poisoning. There is insufficient evidence to prove that increasing NAC dose can treat patients who failed in standard treatment. This study explores the toxicity of NAC overdose in both APAP poisoning and normal mice. Two inbred mouse strains with different sensitivities to propacetamol-induced hepatotoxicity (PIH) were treated with different NAC doses. NAC therapy decreased PIH by reducing lipid oxidation, protein nitration and inflammation, and increasing glutathione (GSH) levels and antioxidative enzyme activities. However, the therapeutic effects of NAC on PIH were dose-dependent from 125 (N125) to 275 mg/kg (N275). Elevated doses of NAC (400 and 800 mg/kg, N400 and N800) caused additional deaths in both propacetamol-treated and normal mice. N800 treatments significantly decreased hepatic GSH levels and induced inflammatory cytokines and hepatic microvesicular steatosis in both propacetamol-treated and normal mice. Furthermore, both N275 and N400 treatments decreased serum triglyceride (TG) and induced hepatic TG, whereas N800 treatment significantly increased interleukin-6, hepatic TG, and total cholesterol levels. In conclusion, NAC overdose induces hepatic and systemic inflammations and interferes with fatty acid metabolism.

scholarly journals Mechanisms underlying the antifibrotic properties of noninhibitory PAI-1 (PAI-1R) in experimental nephritis

2009 ◽  
Vol 297 (4) ◽  
pp. F1045-F1054 ◽  
Author(s):  
Yufeng Huang ◽  
Wayne A. Border ◽  
Daniel A. Lawrence ◽  
Nancy A. Noble
Keyword(s):  
Half Life ◽  
Stable Form ◽  
Therapeutic Effects ◽  
Dependent Manner ◽  
Therapeutic Action ◽  
Protease Inhibition ◽  
Ecm Degradation ◽  
Dose Dependent ◽  
Pai 1

Administration of a mutant, noninhibitory PAI-1 (PAI-1R), reduces disease in experimental glomerulonephritis. Here we investigated the importance of vitronectin (Vn) binding, PAI-1 stability and protease binding in this therapeutic effect using a panel of PAI-1 mutants differing in half-life, protease binding, and Vn binding. PAI-1R binds Vn normally but does not inhibit proteases. PAI-1AK has a complete defect in Vn binding but retains full inhibitory activity, with a short half-life similar to wild-type (wt)-PAI-1. Mutant 14-lb is identical to wt-PAI-1 but with a longer half-life. PAI-1K has defective Vn binding, inhibits proteases normally, and has a long half-life. In vitro wt-PAI-1 dramatically inhibited degradation of mesangial cell ECM while the AK mutant had much less effect. Mutants 14-1b and PAI-1K, like wt-PAI-1, inhibited matrix degradation but PAI-1R failed to reverse this inhibition although PAI-1R reversed the wt-PAI-1-induced inhibition of ECM degradation in a plasmin-, time-, and dose-dependent manner. Thus the ability of PAI-1 to inhibit ECM degradation is dependent both on its antiproteinase activity and on maintaining an active conformation achieved either by Vn binding or mutation to a stable form. Administration of these PAI-1 mutants to nephritic rats confirmed the in vitro data; only PAI-1R showed therapeutic effects. PAI-1K did not bind to nephritic kidney, indicating that Vn binding is essential to the therapeutic action of PAI-1R. The ability of PAI-1R to remain bound to Vn even in a high-protease environment is very likely the key to its therapeutic efficacy. Furthermore, because both PAI-1R and 14-1b bound to the nephritic kidney in the same pattern and differ only in their ability to bind proteases, lack of protease inhibition is also keyed to PAI-1R's therapeutic action.

scholarly journals A novel human Smad4 mutation is involved in papillary thyroid carcinoma progression

2011 ◽  
Vol 19 (1) ◽  
pp. 39-55 ◽  
Author(s):  
Sonia D'Inzeo ◽  
Arianna Nicolussi ◽  
Caterina Francesca Donini ◽  
Massimo Zani ◽  
Patrizia Mancini ◽  
...  
Keyword(s):  
Papillary Thyroid Carcinoma ◽  
Thyroid Carcinoma ◽  
Mammalian Cells ◽  
Transforming Growth Factor ◽  
Human Cancer ◽  
Papillary Thyroid ◽  
Tgfβ Signaling ◽  
Strong Reduction ◽  
Mesenchymal Transition ◽  
Wide Range

Smad proteins are the key effectors of the transforming growth factor β (TGFβ) signaling pathway in mammalian cells. Smad4 plays an important role in human physiology, and its mutations were found with high frequency in wide range of human cancer. In this study, we have functionally characterized Smad4 C324Y mutation, isolated from a nodal metastasis of papillary thyroid carcinoma. We demonstrated that the stable expression of Smad4 C324Y in FRTL-5 cells caused a significant activation of TGFβ signaling, responsible for the acquisition of transformed phenotype and invasive behavior. The coexpression of Smad4 C324Y with Smad4 wild-type determined an increase of homo-oligomerization of Smad4 with receptor-regulated Smads and a lengthening of nuclear localization. FRTL-5 clones overexpressing Smad4 C324Y showed a strong reduction of response to antiproliferative action of TGFβ1, acquired the ability to grow in anchorage-independent conditions, showed a fibroblast-like appearance and a strong reduction of the level of E-cadherin, one crucial event of the epithelial–mesenchymal transition process. The acquisition of a mesenchymal phenotype gave the characteristics of increased cellular motility and a significant reduction in adhesion to substrates such as fibronectin and laminin. Overall, our results demonstrate that the Smad4 C324Y mutation plays an important role in thyroid carcinogenesis and can be considered as a new prognostic and therapeutic target for thyroid cancer.

scholarly journals Position Effects Are Influenced by the Orientation of a Transgene with Respect to Flanking Chromatin

2001 ◽  
Vol 21 (1) ◽  
pp. 298-309 ◽  
Author(s):  
Yong-Qing Feng ◽  
Matthew C. Lorincz ◽  
Steve Fiering ◽  
John M. Greally ◽  
Eric E. Bouhassira
Keyword(s):  
Mammalian Cells ◽  
Transgene Expression ◽  
Methylation Status ◽  
Regulatory Elements ◽  
Cell Populations ◽  
Methylation Analysis ◽  
Position Effects ◽  
Methylation State ◽  
Cassette Exchange

ABSTRACT We have inserted two expression cassettes at tagged reference chromosomal sites by using recombinase-mediated cassette exchange in mammalian cells. The three sites of integration displayed either stable or silencing position effects that were dominant over the different enhancers present in the cassettes. These position effects were strongly dependent on the orientation of the construct within the locus, with one orientation being permissive for expression and the other being nonpermissive. Orientation-specific silencing, which was observed at two of the three site tested, was associated with hypermethylation but not with changes in chromatin structure, as judged by DNase I hypersensitivity assays. Using CRE recombinase, we were able to switch in vivo the orientation of the transgenes from the permissive to the nonpermissive orientation and vice versa. Switching from the permissive to the nonpermissive orientation led to silencing, but switching from the nonpermissive to the permissive orientation did not lead to reactivation of the transgene. Instead, transgene expression occurred dynamically by transcriptional oscillations, with 10 to 20% of the cells expressing at any given time. This result suggested that the cassette had been imprinted (epigenetically tagged) while it was in the nonpermissive orientation. Methylation analysis revealed that the methylation state of the inverted cassettes resembled that of silenced cassettes except that the enhancer had selectively lost some of its methylation. Sorting of the expressing and nonexpressing cell populations provided evidence that the transcriptional oscillations of the epigenetically tagged cassette are associated with changes in the methylation status of regulatory elements in the transgene. This suggests that transgene methylation is more dynamic than was previously assumed.

scholarly journals Insights into the Mode of Action of Chitosan as an Antibacterial Compound

2008 ◽  
Vol 74 (12) ◽  
pp. 3764-3773 ◽  
Author(s):  
Dina Raafat ◽  
Kristine von Bargen ◽  
Albert Haas ◽  
Hans-Georg Sahl
Keyword(s):  
Antimicrobial Activity ◽  
Cell Wall ◽  
Cell Membrane ◽  
Mode Of Action ◽  
Membrane Lipids ◽  
Expression Profiles ◽  
Transcriptional Response ◽  
Response Analysis ◽  
Sequence Of Events ◽  
Wide Range

ABSTRACT Chitosan is a polysaccharide biopolymer that combines a unique set of versatile physicochemical and biological characteristics which allow for a wide range of applications. Although its antimicrobial activity is well documented, its mode of action has hitherto remained only vaguely defined. In this work we investigated the antimicrobial mode of action of chitosan using a combination of approaches, including in vitro assays, killing kinetics, cellular leakage measurements, membrane potential estimations, and electron microscopy, in addition to transcriptional response analysis. Chitosan, whose antimicrobial activity was influenced by several factors, exhibited a dose-dependent growth-inhibitory effect. A simultaneous permeabilization of the cell membrane to small cellular components, coupled to a significant membrane depolarization, was detected. A concomitant interference with cell wall biosynthesis was not observed. Chitosan treatment of Staphylococcus simulans 22 cells did not give rise to cell wall lysis; the cell membrane also remained intact. Analysis of transcriptional response data revealed that chitosan treatment leads to multiple changes in the expression profiles of Staphylococcus aureus SG511 genes involved in the regulation of stress and autolysis, as well as genes associated with energy metabolism. Finally, a possible mechanism for chitosan's activity is postulated. Although we contend that there might not be a single classical target that would explain chitosan's antimicrobial action, we speculate that binding of chitosan to teichoic acids, coupled with a potential extraction of membrane lipids (predominantly lipoteichoic acid) results in a sequence of events, ultimately leading to bacterial death.

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