Nonspecific Side Effects Research Articles

Bromelain-facilitated GNPs: A strategic innate anticancer delivery system for methotrexate into osteosarcoma cells

Osteosarcoma is the most common malignant bone tumor in children and adolescents during rapid bone growth. Traditional chemotherapy has markedly improved patient outcomes but suffers from nonspecific side effects due to nonspecific drug distribution, highlighting the need for targeted therapies. Methotrexate, although effective, poses toxicity concerns and prompts exploration into advanced drug delivery systems like nanoparticles. Here, we developed a novel method of synthesizing gold nanoparticles using bromelain (G-BNPs) as a reducing and capping agent, and we conjugated G-BNPs with methotrexate (G-B-MNPs) via 1-Ethyl-3-(3-dimethylaminopropyl)carbodiimide/N-hydroxysuccinimide (EDC/NHS) as crosslinker. Our meticulous characterization, which included UV–visible spectroscopy, Transmission electron microscopy (TEM), Dynamic light scattering (DLS), Zeta potential, and Fourier-transform infrared spectroscopy (FTIR) analyses confirmed the successful fabrication and drug conjugation of the G-BNPs. We then evaluated the cytotoxicity of the G-BNPs (sized ∼ 21 nm) and G-B-MNPs (sized ∼ 24 nm) on the osteosarcoma cell lines, Saos-2 and MG-63, as well as the non-cancerous primary osteoblast cells. Our findings revealed that G-B-MNPs significantly increased the cytotoxic impact on osteosarcoma cells compared to methotrexate alone while showing minimal toxicity toward primary osteoblast cells. Importantly, using 4`,6-diamidino-2-phenylindole(DAPI), we demonstrated that G-B-MNPs were delivered directly into the nucleus of the cells without encountering lysosomes. G-B-MNPs can be considered a potential drug delivery method for osteosarcoma treatment by improving effectiveness and reducing adverse effects, thereby enhancing patient prognosis.

The method for assessing the specificity of developing CAR therapies

The effectiveness of antitumor chimeric antigen receptor (CAR) therapy mainly dealt with an elevated sensitivity of CAR cells to target cells. However, CAR therapies are associated with nonspecific side effects: on-target off-tumor toxicity. Sensitivity and specificity of CAR cells are the most important properties of the recognition process of target cells among other cells. Current developments are mainly concentrated on exploring molecular biology methods for designing CAR cells with the highest sensitivity, while the problem of the CAR cell specificity is rarely considered. For the assessment of CAR cell specificity, we suggest that, in addition to an elevated level of CAR-antigen affinity, the ability of CARs for clustering should be taken into account. We assume that the CAR cell cytotoxicity is determined by CAR clustering. The latter is treated within the framework of nucleation theory. The master equation for the probability of CAR cell cytotoxicity is derived. The size of a critical CAR cluster is found to be one of two most essential parameters. The conditions for necessary sensitivity and sufficient specificity are explored. Relevant parametric diagrams are derived. Possible applications of the method for assessing the specificity of developing CAR therapies are discussed.

Osteopontin: A Key Multifaceted Regulator in Tumor Progression and Immunomodulation.

The tumor microenvironment (TME) is composed of various cellular components such as tumor cells, stromal cells including fibroblasts, adipocytes, mast cells, lymphatic vascular cells and infiltrating immune cells, macrophages, dendritic cells and lymphocytes. The intricate interplay between these cells influences tumor growth, metastasis and therapy failure. Significant advancements in breast cancer therapy have resulted in a substantial decrease in mortality. However, existing cancer treatments frequently result in toxicity and nonspecific side effects. Therefore, improving targeted drug delivery and increasing the efficacy of drugs is crucial for enhancing treatment outcome and reducing the burden of toxicity. In this review, we have provided an overview of how tumor and stroma-derived osteopontin (OPN) plays a key role in regulating the oncogenic potential of various cancers including breast. Next, we dissected the signaling network by which OPN regulates tumor progression through interaction with selective integrins and CD44 receptors. This review addresses the latest advancements in the roles of splice variants of OPN in cancer progression and OPN-mediated tumor-stromal interaction, EMT, CSC enhancement, immunomodulation, metastasis, chemoresistance and metabolic reprogramming, and further suggests that OPN might be a potential therapeutic target and prognostic biomarker for the evolving landscape of cancer management.

Programmed cell death-1-modified pig developed using electroporation-mediated gene editing for in vitro fertilized zygotes.

Programmed cell death-1 (PD-1) is an immunoinhibitory receptor required to suppress inappropriate immune responses such as autoimmunity. Immune checkpoint antibodies that augment the PD-1 pathway lead to immune-related adverse events (irAEs), organ non-specific side effects due to autoimmune activation in humans. In this study, we generated a PD-1 mutant pig using electroporation-mediated introduction of the CRISPR/Cas9 system into porcine zygotes to evaluate the PD-1 gene deficiency phenotype. We optimized the efficient guide RNAs (gRNAs) targeting PD-1 in zygotes and transferred electroporated embryos with the optimized gRNAs and Cas9 into recipient gilts. One recipient gilt became pregnant and gave birth to two piglets. Sequencing analysis revealed that both piglets were biallelic mutants. At 18mo of age, one pig showed non-purulent arthritis of the left elbow/knee joint and oligozoospermia, presumably related to PD-1 modification. Although this study has a limitation because of the small number of cases, our phenotypic analysis of PD-1 modification in pigs will provide significant insight into human medicine and PD-1-deficient pigs can be beneficial models for studying human irAEs.

Intratracheal administration of programmable DNA nanostructures combats acute lung injury by targeting microRNA-155

Acute lung injury (ALI) is an acute inflammatory process that can result in life-threatening consequences. Programmable DNA nanostructures have emerged as excellent nanoplatforms for microRNA-based therapeutics, offering potential nanomedicines for ALI treatment. Nonetheless, the traditional systematic administration of nanomedicines is constrained by low delivery efficiency, poor pharmacokinetics, and nonspecific side effects. Here, we identify macrophage microRNA-155 as a novel therapeutic target using the magnetic bead sorting technique. We further construct a DNA nanotubular nucleic acid drug antagonizing microRNA-155 (NT-155) for ALI treatment through intratracheal administration. Flow cytometry results demonstrate that NT-155, when inhaled, is taken up much more effectively by macrophages and dendritic cells in the bronchoalveolar lavage fluid of ALI mice. Furthermore, NT-155 effectively silences the overexpressed microRNA-155 in macrophages and exerts excellent inflammation inhibition effects in vitro and ALI mouse models. Mechanistically, NT-155 suppresses microRNA-155 expression and activates its target gene SOCS1, inhibiting the p-P65 signaling pathway and suppressing proinflammatory cytokine secretion. The current study suggests that deliberately designed nucleic acid drugs are promising nanomedicines for ALI treatment and the local administration may open up new practical applications of DNA in the future.

Electrical Stimulation for Immunomodulation.

The immune system plays a key role in the development and progression of numerous diseases such as chronic wounds, autoimmune diseases, and various forms of cancer. Hence, controlling the behavior of immune cells has emerged as a promising approach for treating these diseases. Current modalities for immunomodulation focus on chemical based approaches, which while effective have the limitations of nonspecific systemic side effects or requiring invasive delivery approaches to reduce the systemic side effects. Recent advances have unraveled the significance of electrical stimulation as an attractive noninvasive approach to modulate immune cell phenotype and activity. This review provides insights on electrical stimulation strategies employed for regulating the behavior of macrophages, T and B cells, and neutrophils. For obtaining a better understanding, two major types of electrical stimulation sources, conventional and self-powered sources, that have been used for immunomodulation are extensively discussed. Next, the strategies of electrical stimulation that may be applied to cells in vitro and in vivo are discussed, with a focus on conventional and stimuli-responsive self-powered sources. A description of how these strategies influence the polarization, phagocytosis, migration, and differentiation of immune cells is also provided. Finally, recent developments in the use of highly localized and efficient platforms for electrical stimulation based immunomodulation are also highlighted.

Abstract A029: Identification of cancer-specific CPP from human telomerase peptide library and its drug delivery potential in anti-cancer strategy

Abstract A reverse-transcriptase-subunit of telomerase (hTERT) derived peptide has been identified as a cell-penetrating peptide (CPP) in our previous study. This hTERT derived CPP was delivered into a variety of cells including various cancer cell lines and primary blood cells. Moreover, the CPP was predominantly located in the cytoplasm of the cells, while a significantly higher proportion of TAT peptide was localized in the nucleus. In order to reduce adverse non-specific toxic side effects of current anticancer drug, we developed cancer-specific anticancer drug delivery system using hTERT derived CPP. The cell penetrating activity was Pep 98, a specific hTERT derived CPP, was evaluated in head and neck cancer cells. The permeability of specific telomerase derived Pep 98-FITC into cancer cells was 20-300 fold higher than control peptide groups in head and neck cancer cells. These result suggest that pep98 can be used to reduce the side effects and increase the efficacy of cancer therapy through the use of specific anticancer drug delivery system.We conjugated Pep 98 with STAT 3 inhibitor (NSC74859) which has been presented as a molecular therapeutic target for head and neck cancer and evaluated concentration of NSC74589 for anti-cancer activity in head and neck cancer cell line. The result showed that the concentration of Pep-98 conjugated NSC74589 could be reduced up to 1/10-20 dose level compared with NSC74589 alone in head and neck cancer cells. Pep 98-NSC74859 conjugates were found to be capable of specific killing head and neck cancer cells without affecting normal fibroblast cells.We have confirmed the possibility of developing a new head neck cancer anticancer drug with improved therapeutic efficacy through the conjugation of the STAT3 inhibitor with hTERT derived peptide. Citation Format: Yoon Seon Lee, Myung Jin Lee, Won Jun Shon. Identification of cancer-specific CPP from human telomerase peptide library and its drug delivery potential in anti-cancer strategy [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr A029.

The developing role of extracellular vesicles in autoimmune diseases: special attention to mesenchymal stem cell-derived extracellular vesicles.

Autoimmune diseases are complex, chronic inflammatory conditions initiated by the loss of immunological tolerance to self-antigens. Nowadays, there is no effective and useful therapy for autoimmune diseases, and the existing medications have some limitations due to their nonspecific targets and side effects. During the last few decades, it has been established that mesenchymal stem cells (MSCs) have immunomodulatory functions. It is proposed that MSCs can exert an important therapeutic effect on autoimmune disorders. In parallel with these findings, several investigations have shown that MSCs alleviate autoimmune diseases. Intriguingly, the results of studies have demonstrated that the effective roles of MSCs in autoimmune diseases do not depend on direct intercellular communication but on their ability to release a wide spectrum of paracrine mediators such as growth factors, cytokines and extracellular vehicles (EVs). EVs that range from 50 to 5,000nm were produced by almost any cell type, and these nanoparticles participate in homeostasis and intercellular communication via the transfer of a broad range of biomolecules such as modulatory proteins, nucleic acids (DNA and RNA), lipids, cytokines, and metabolites. EVs derived from MSCs display the exact properties of MSCs and can be safer and more beneficial than their parent cells. In this review, we will discuss the features of MSCs and their EVs, EVs biogenesis, and their cargos, and then we will highlight the existing discoveries on the impacts of EVs from MSCs on autoimmune diseases such as multiple sclerosis, arthritis rheumatic, inflammatory bowel disease, Type 1 diabetes mellitus, systemic lupus erythematosus, autoimmune liver diseases, Sjögren syndrome, and osteoarthritis, suggesting a potential alternative for autoimmune conditions therapy.

TLR Agonists Delivered by Plant Virus and Bacteriophage Nanoparticles for Cancer Immunotherapy.

Toll-like receptors (TLRs) are promising targets in cancer immunotherapy due to their role in activating the immune system; therefore, various small-molecule TLR agonists have been tested in clinical applications. However, the clinical use of TLR agonists is hindered by their non-specific side effects and poor pharmacokinetics. To overcome these limitations, we used plant virus nanoparticles (VNPs) and bacteriophage virus-like particles (VLPs) as drug delivery systems. We conjugated TLR3 or TLR7 agonists to cowpea mosaic virus (CPMV) VNPs, cowpea chlorotic mottle virus (CCMV) VNPs, and bacteriophage Qβ VLPs. The conjugation of TLR7 agonist, 2-methoxyethoxy-8-oxo-9-(4-carboxybenzyl)adenine (1V209), resulted in the potent activation of immune cells and promoted the production of pro-inflammatory cytokine interleukin 6. We found that 1V209 conjugated to CPMV, CCMV, and Qβ reduced tumor growth in vivo and prolonged the survival of mice compared to those treated with free 1V209 or a simple admixture of 1V209 and viral particles. Nucleic acid-based TLR3 agonist, polyinosinic acid with polycytidylic acid (poly(I:C)), was also delivered by CPMV VNPs, resulting in enhanced mice survival. All our data suggest that coupling and co-delivery are required to enhance the anti-tumor efficacy of TLR agonists and simple mixing of the VLPs with the agonists does not confer a survival benefit. The delivery of 1V209 or poly(I:C) conjugated to VNPs/VLPs probably enhances their efficacy due to the multivalent presentation, prolongation of tumor residence time, and targeting of the innate immune cells mediated by the VNP/VLP carrier.

What Is the Benefit of Adding Placebo Side-Effect Information to Positively Framed Patient Leaflets?

Abstract: Background: Positively framing side-effect risk in patient information leaflets (PILs) can reduce side-effect expectations and resulting nocebo effects (nonspecific medication side effects unrelated to the drug’s pharmacological action). There is scope to educate patients about nocebo effects in PILs to minimize their occurrence further. Aims: To investigate if incorporating information on placebo-reported side effects reduces side-effect expectations compared to a positively framed-only or standard PIL. Methods: Participants ( N = 443) completed an online study and were randomized to read one of three PILs for a hypothetical antibiotic: standard PIL ( n = 140), positively framed PIL ( n = 151), or positively framed PIL with placebo side-effect information ( n = 152). Participants’ side-effect expectations, absolute risk perceptions, and intended adherence were recorded. Results: The standard PIL resulted in significantly higher side-effect expectations compared to the positively framed + placebo side-effect information PIL. Including the placebo side-effect results had no effect on side-effect expectations compared to the positive framing only PIL, however, there was a significant interaction between health literacy and PIL condition on side-effect expectations. Both positively framed PILs produced more accurate risk estimates for the more common side effects. There was no difference in intended adherence between the three PILs. Limitations: Our findings are limited by the highly educated sample and hypothetical context. Conclusions: There was no benefit of adding placebo side-effect information, however alternative ways of explaining nocebo effects in PILs should be explored utilizing clinical contexts and samples with a wider range of participant ages, and health literacy.

Berplatin: A Conjugate of Berberine and Cisplatin Serving as a Potent Antitumor Agent with Distinctive Tumor Cell Uptake and Minimal Side Effects

AbstractDesign of Pt(IV) prodrugs with biologically active axial ligands is an excellent approach to overcome the disadvantages (nonspecific killing, side effects, and drug resistance) of conventional first‐line Pt(II)‐based chemotherapeutic drugs, such as cisplatin, carboplatin and oxaliplatin. The selection of ideal axial ligands is essential for the success of Pt(IV) drugs. Previously, that berberine exhibits great superiority in tumor selectivity is found with uncovered mechanisms. In this study, that berberine is specifically internalized and accumulated in tumors but not normal cells through endocytosis pathways is first disclosed. Therefore a new Pt(IV) prodrug of cisplatin bearing berberine as an axial ligand, named berplatin, is designed and synthesized. Different from cisplatin, berplatin can be effectively and selectively internalized into cancer cells, through macropinocytosis and caveolin‐1 (CAV‐1)‐dependent active endocytosis, and cleaved in intracellular reductive environments to generate cisplatin and berberine, which synergistically induces substantial DNA breaking, apoptosis and cell cycle arrest in both cisplatin‐sensitive and ‐resistant cancer cells. In vivo, berplatin manifests much higher antitumor activity and undetectable side effects compared to cisplatin. This study demonstrates that berplatin can serve as a potent Pt(IV) prodrug for the treatment of cisplatin‐sensitive and ‐resistant cancers with active uptake, excellent specificity, and minimal side effects.

Tumor-Targeting Polymer–Drug Conjugate for Liver Cancer Treatment In Vitro

Bufalin (buf) has poor solubility in aqueous solution, poor tumor targeting, and many non-specific toxic and side effects. The advantages of high-molecular-weight polymer conjugates are that they can improve the water solubility of buf, prolong plasma half-life, and reduce non-specific toxicity. A novel water-soluble polymer-drug conjugate with buf and fluorescein pendants was prepared by the combination of reversible addition-fragmentation transfer (RAFT) polymerization and click chemistry. Its anticancer performance and cellular uptake behavior against liver cancer were investigated in vitro. The polymer-buf conjugates exhibit controlled release and tumor-targeting capabilities, showing promise for clinical applications.

Functionalized PAMAM-based Nanoformulation for Targeted Delivery of 5-Fluorouracil in Hepatocellular Carcinoma.

The efficacy of a traditional anticancer drug is challenged by adverse effects of the drug, including its nonspecific bio-distribution, short half-life, and side effects. Dendrimer-based targeted drug delivery system has been considered a promising strategy to increase targeting ability and reduce adverse effects of anti-cancer drugs. This study analyzed the feasibility of whether the anticancer drug 5-fluorouracil (5-FU) could be delivered by functionalized fifth-poly(amidoamine) (PAMAM) with the peptide WP05 and the acetic anhydride to the liver cancer cells, reducing the toxicity of the PAMAM and improving the targeting property of 5-FU during delivery. The functionalized PAMAM-based nanoformulation (WP05-G5.0NHAC-FUA) was fabricated through an amide condensation reaction to improve the therapeutic efficacy of 5-Fluorouracil (5-FU) in hepatocellular carcinoma (HCC). The physicochemical structure, particle size, zeta potential, stability, and in vitro release characteristics of WP05-G5.0NHAC-FUA were evaluated. In addition, the targeting, biocompatibility, anti-proliferation, and anti-migration of WP05-G5.0NHAC-FUA were investigated. The anti-tumor effect of WP05-G5.0NHAC-FUA in vivo was evaluated by constructing xenograft tumor models of human hepatoma cells (Bel-7402) implanted in nude mice. The resultant WP05-G5.0NHAC-FUA displayed spherical-like nanoparticles with a size of 174.20 ± 3.59 nm. Zeta potential and the drug loading of WP05-G5.0NHAC-FUA were 5.62 ± 0.41mV and 28.67 ± 1.25%, respectively. Notably, the optimized 5-FU-loaded formulation showed greater cytotoxicity with an IC50 of 30.80 ± 4.04 μg/mL than free 5-FU (114.93 ± 1.43 μg/mL) in Bel-7402 cancer liver cells, but a significantly reduced side effect relative to free 5-FU in L02 normal liver cells. In vivo animal study further confirmed efficient tumor accumulation and enhanced therapeutic efficiency. The developed nanoformulation is a promising platform for the targeting delivery of 5-FU and provides a promising solution for improving the efficacy of hepatocellular carcinoma chemotherapy.

Inhalation of tetrandrine liposomes for the treatment of bleomycin induced idiopathic pulmonary fibrosis

BackgroundIdiopathic pulmonary fibrosis (IPF) is a chronic and progressive lung disease of unknown cause, with limited therapeutic options and an urgent need to develop new treatment methods. However, there is still a lack of suitable inhaled drugs for the treatment of IPF due to non-specific side effects, inability to cross the pulmonary barrier, and inadequate management of heterogeneity. In this study, we hypothesized that tetrandrine liposomes (TET-LIPs) delivered by inhalation might have a direct targeting effect on lung tissue, effectively treat IPF and limit its adverse side effects on other organs. MethodsMorphology, particle size, zeta potential, and in vitro dissolution studies of the prepared TET-LIPs were performed. Blood aspartate aminotransferase (AST), alanine aminotransferase (ALT), Urea nitrogen (BUN), creatinine (Cr) were measured to determine the liver and renal function. Bleomycin-induced mice model of IPF was used to evaluate the in vivo pharmacodynamics of TET-LIPs. The levels of hydroxyproline, tumor necrosis factor (TNF-α), and transforming growth factor (TGF-β1) were examined by enzyme-linked immunosorbent assay (ELISA). Finally, alveolar lavage fluid lipid profiles were obtained by using ultra-performance liquid chromatography combined with quadrupole-Exactive Orbitrap mass spectrometry (UPLC-Q-Exactive Orbitrap MS). ResultsInhaled TET-LIPs significantly inhibited the progression of bleomycin-induced acute inflammation and pulmonary fibrosis in lung tissue. Furthermore, delivering the drug locally to the lungs showed little accumulation in the liver and kidneys after long-term inhalation and had a favorable safety profile. Interestingly, we observed a relationship between inhaled TET-LIPs and increased levels of triglycerides (TG), fatty acids (FA) while decreased levels of lysophosphatidylcholine (LPC) in BALF. ConclusionsOverall, as TET-LIPs were safe and well-tolerated in the BLM-induced mice model, tetrandrine encapsulation in liposomes could be an effective method for locally delivering to patients with IPF. Simultaneously, our findings revealed that various lipid metabolic disorders were linked to the early inflammatory phase of the fibrosis process, and the regulation of lipid metabolism may be one of the mechanisms to improve pulmonary fibrosis using TET-LIPs.

Audit Of Psychotropic Prescribing in the Crisis Team at Fieldhead Hospital According to NICE Guidelines

AimsTo ensure that psychotropic prescribing and monitoring in the Crisis Team is compliant with NICE guidelines and to provide excellent patient care and to practice medicine safely.MethodsMedication prescribing should be a collaborative decision by the service user and the prescriber. This allows patients to have autonomy to decide their treatment plan. NICE provides guidelines for prescribing medication which includes baseline investigations, reviews of treatment including side effects, and physical health monitoring.We selected 50 admitted patients for the audit from April 2021 until September 2021, who were prescribed psychotropic medications. We used medication cards and electronic patients’ records (System One). Our exclusion criteria were the 72-hour post-discharge follow-up from the inpatient ward.The audit standards included as follows: age, gender, the indication, the start of medications, dose, within BNF limits, discussion, consent from the patient, comorbidities, physical health monitoring, response to treatment, monitoring of side effects, and other important information.Results100% results for indication, dosage, discussion with the patient, and side effects monitoring.We had promising results for benefits from the treatment (46 out of 50 patients responded to treatment) and 4/50 did not respond to treatment. Unfortunately, one patient died from an overdose of illicit drugs, not with prescribed medication and one was admitted, and one felt worse, and one did not have any response.However, 22 out of 50 patients were prescribed antipsychotic medication.11 out of 22 patients had ECG and blood done by the Crisis Team and 4 done by other parties (hospital and primary care).3 had recent blood tests but no ECG.2 patients did not have physical health monitoring and the reason was not documented.2 patients were started on antipsychotic by the Crisis Team, but the dose was not changed.In terms of side effects, 8 out of 50 reported some side effects.6 of them were prescribed antidepressants. They reported difficulty in sleeping and palpitations with Venlafaxine, nausea with Fluoxetine, nonspecific side effects with Citalopram, and sedation with Trazadone. 2 patients felt dizziness, diarrhoea, and muscle spasms with Mirtazapine. One patient had a metallic taste with Zopiclone. For side effects with antipsychotics, only one patient reported side effects with Olanzapine.Conclusion•The Crisis Team is working at excellent standards on most areas of psychotropic prescribing and monitoring•The Crisis Team needs to improve physical health monitoring of their patients.

Anticancer nanomedicines harnessing tumor microenvironmental components

Introduction Small-molecular drugs are extensively used in cancer therapy, while they have issues of nonspecific distribution and consequent side effects. Nanomedicines that incorporate chemotherapeutic drugs have been developed to enhance the therapeutic efficacy of these drugs and reduce their side effects. One of the promising strategies is to prepare nanomedicines by harnessing the unique tumor microenvironment (TME). Areas covered The TME contains numerous cell types that specifically express specific antibodies on the surface including tumor vascular endothelial cells, tumor-associated adipocytes, tumor-associated fibroblasts, tumor-associated immune cells and cancer stem cells. The physicochemical environment is characterized with a low pH, hypoxia, and a high redox potential resulting from tumor-specific metabolism. The intelligent nanomedicines can be categorized into two groups: the first group which is rapidly responsive to extracellular chemical/biological factors in the TME and the second one which actively and/or specifically targets cellular components in the TME. Expert opinion In this paper, we review recent progress of nanomedicines by harnessing the TME and illustrate the principles and advantages of different strategies for designing nanomedicines, which are of great significance for exploring novel nanomedicines or translating current nanomedicines into clinical practice. We will discuss the challenges and prospects of preparing nanomedicines to utilize or alter the TME for achieving effective, safe anticancer treatment.

LONG-TERM EFFECTS OF SHAM SURGERY ON PHAGOCYTE FUNCTIONS IN RATS

Animal models of inflammatory disorders, including those of the nervous system are commonly used to explore the pathophysiological role of immune cell response in disease triggering and course and to develop biotechnology products for therapeutic use. Modeling some of these disorders, particularly neurodegenerative diseases, implies surgical manipulations for the intracerebral introduction of disease-initiating substances (toxins, amyloids etc.). Design of these experiments involves the use of sham-operated animals as a control of non-specific intrinsic side-effects elicited by surgical manipulations per se, including local and systemic inflammation, where phagocytic cells are key participants. Short-term post-surgical immunomodulatory effects are widely reported. However, no study thus far has examined the long term effects of sham-surgery on phagocyte functions. The purpose of this study was to evaluate the effect of sham-surgery, commonly used for modeling neurodegenerative diseases, on phagocyte functions in the far terms after the surgical manipulations. Materials and Methods. Adult male Wistar rats were used in the study. Sham surgery consisted of stereotactic unilateral injection of saline solution into the median forebrain bundle (sham-operated 1, SO1) or directly into the substantia nigra (sham-operated 2, SO2). Before the placebo surgery, animals were anaesthetized using nembutal and ketamine/xylazine correspondingly. Functional characteristics (phagocytic activity, oxidative metabolism, CD80/86 and CD206 expression) of phagocytes (microglia, peritoneal macrophages, circulating monocytes and granulocytes) were examined by flow cytometry. Differential leukocyte count was conducted using hematological analyzer. Results. Phagocytes from animals underwent of different protocols of placebo surgery, demonstrated various patterns of functional changes on day 29 after the manipulations. In animals from SO1 group, we observed signs of residual neuroinflammation (pro-inflammatory shift of microglia functional profile) along with ongoing resolution of systemic inflammation (anti-inflammatory metabolic shift of circulating phagocytes and peritoneal macrophages). In rats from SO2 group, pro-inflammatory polarized activation of peritoneal phagocytes was registered along with anti-inflammatory shift in microglia and circulating phagocytes. Conclusions. Sham surgery influences functions of phagocytic cells of different locations even in the far terms after the manipulations. These effects can be considered as combined long-term consequences of surgical brain injury and the use of anesthetics. Our observations evidences, that sham associated non-specific immunomodulatory effects should always be taken into consideration in animal models of inflammatory central nervous system diseases.

Systemic L-ornithine supplementation specifically increases ovarian putrescine levels during ovulation in mice†.

In all mammalian species examined thus far, the ovaries produce a burst of ornithine decarboxylase (ODC) and putrescine during ovulation or after application of human chorionic gonadotropin (hCG). Aged mice have significantly reduced levels of this periovulatory ODC and putrescine rise. Putrescine supplementation, in vitro during oocyte maturation or in mouse drinking water during the periovulatory period, reduces egg aneuploidies and embryo resorption, improving fertility of aged mice. These studies suggest that periovulatory putrescine supplementation may be a simple and effective therapy for reproductive aging for women. However, putrescine supplementation is expected to increase widespread tissue putrescine levels, raising concerns of nonspecific and unwanted side effects. Given that ODC is highly expressed in the ovaries during ovulation but otherwise exhibits low activity in most tissues, we hypothesized that periovulatory supplementation of L-ornithine, the substrate of ODC, might be suitable for delivering putrescine specifically to the ovaries. In this study, we have demonstrated that systemic application of L-ornithine via oral gavage or subcutaneous injection increased ovarian putrescine levels; the increase was restricted to animals that had been injected with hCG. Furthermore, L-ornithine specifically increased ovarian putrescine levels without affecting putrescine levels in any other tissues. However, our attempts to improve fertility of aged mice through L-ornithine supplementation in mouse drinking water produced either no effects (1% L-ornithine) or negative impact on fertility (4% ornithine). Our results suggest that it might not be feasible to achieve fertility-enhancing ovarian putrescine levels via L-ornithine supplementation in drinking water without encountering undesired consequences of high dose of exogenous L-ornithine.

Design, Synthesis and Biological Evaluation of New N-Acyl Hydrazones with a Methyl Sulfonyl Moiety as Selective COX-2 Inhibitors.

The mechanism of action of nonsteroidal anti-inflammatory drugs (NSAIDs) is inhibition of specific prostaglandin (PG) synthesis by inhibition of cyclooxygenase (COX) enzymes. The two COX isoenzymes show 60 % similarity. It is known that the nonspecific side effects of conventional NSAIDs are physiologically caused by inhibition of the COX-1 enzyme. Therefore, the use of COX-2 selective inhibitors is seen to be a more beneficial approach in reducing these negative effects. However, some of the existing COX-2 selective inhibitors show cardiovascular side effects. Therefore, studies on the development of new selective COX-2 inhibitors remain necessary. It is important to develop new COX-2 inhibitors in the field of medicinal chemistry. Accordingly, novel N-acyl hydrazone derivatives were synthesized as new COX-2 inhibitors in this study. The hydrazone structure, also known for its COX activity, is important in terms of many biological activities and was preferred as the main structure in the design of these compounds. A methyl sulfonyl pharmacophore was added to the structure in order to increase the affinity for the polar side pocket present in the COX-2 enzyme. It is known that methyl sulfonyl groups are suitable for polar side pockets. The synthesis of the compounds (3a-3j) was characterized by spectroscopic methods. Evaluation of in vitro COX-1/COX-2 enzyme inhibition was performed by fluorometric method. According to the enzyme inhibition results, the obtained compounds displayed the predicted selectivity for COX-2 enzyme inhibition. Compound 3j showed important COX-2 inhibition with a value of IC50 =0.143 uM. Interaction modes between the COX-2 enzyme and compound 3j were investigated by docking studies.

The inhibitory effect of Agrimonia Pilosa methanolic extract on matrix metalloproteinases in HT1080 cells.

The risk of cancer increases with aging due to the accumulation of cellular deterioration that can spread to other organs through the blood and lymphatic vessels. Therefore, the inhibition of metastasis is a major concern for the treatment of cancer. Several synthetic drugs have been developed for the treatment of various cancers. However, these drugs are effective; nonspecific action and side effects on the normal human cells limit their wide acceptance, thus demanding some potential alternative. Hence, the present study emphasizes investigating the effect of a methanolic extract of Agrimonia Pilosa (APLME) on matrix metalloproteinases (MMPs) in human fibroblast sarcoma cells. The action of APLME on MMP-2 and MMP-9 was investigated using gelatin zymography. APLME suppressed the activities of MMP-2 and MMP-9 in PMA (phorbol myristate acetate)-treated HT1080 cells. In addition, western blot analysis and immunofluorescence were performed to investigate the effect of APLME on the expression of the proteins that are the major proteins involved in cell invasion and metastasis. APLME treatment inhibited the expression of MMP-2 and MMP-9 in addition to the activations of JNK, ERK, and AKT-1. Furthermore, APLME was observed to suppress cell invasion related to metastasis using cell invasion assay. Therefore, the above findings indicate that APLME inhibits the expression activity of MMP-2 and MMP-9 via inactivation of ERK and JNK in addition to AKT-1, leading to inhibiting cell invasion. Therefore, these results indicate that APLME may be used as a candidate substance for inhibiting cell invasion. PRACTICAL APPLICATIONS: Cancer increases the cell invasion to other organs through the blood and lymphatic vessels. Cancer cells deplete the nutrients and create new blood vessels that infiltrate and metastasize to other tissues. Therefore, this present study examined the effect of Agrimonia Pilosa on cell invasion. It was found that Agrimonia Pilosa methanolic extract inhibited the invasion of cancer cells through the inactivation of ERK and JNK. In addition, APLME reduced the activation and protein expression of MMP-2 and MMP-9 in addition to AKT-1. Thus, APLME can be utilized as a potential alternative therapeutic agent for inhibiting metastasis.