RD Cell Lines Research Articles

Synthesis and Characterization of TiO2 -CQDs Nanocomposites and Testing of Their Anticancer Potential

This work used a green approach to produce carbon quantum dots (CQDs). TiO2 nanoparticles (NPs) and TiO2–CQDs nanocomposites with different weight ratios of CQDs were organized through a facile sol-gel method. XRD showed that the anatase and rutile phases of TiO2 were polycrystalline with a tetragonal structure and that the CQDs exhibited a broad peak at (002) and a hexagonal structure. High resolution-transmission-electron microscopy revealed that the TiO2 NPs agglomerated in mostly spherical shapes and that the anatase and rutile phases of TiO2 had sizes of less than 15 and 25 nm, respectively. The CQDs had a relatively uniform diameter, a spherical shape with a highly crystalline structure, and a size below 2 nm. UV–visible spectroscopy revealed that the absorbance of the TiO2- CQDs nanocomposites increased with the increase in CQD ratio. The energy band gaps of the anatase and rutile phases were 3.07 and 2.7 eV, respectively, whereas that of CQDs was 3.14 eV. Meanwhile, the energy band gaps of the TiO2–CQDs nanocomposites decreased with the increase in CQDs ratio. The growth inhibition rates of the liver cancer HepG2 cell line and the normal cell line RD were measured after 24 hours of experience to the two TiO2 phases and TiO2–CQDs nanocomposites. The cytotoxicity test presented that the tested substances were extremely harmful to cells in cancer. Inhibition rates increased with the increase in CQDs ratio. The inhibition rate of growth in cancer cells, including the liver cancer HepG2 cell line and the normal cell line (RD), was measured for 24 hours after they were exposed to TiO2 (two phases) and TiO2-CQDs nanocomposites. The samples showed a slight effect on the normal line (RD) compared to HepG2 cancer. The highest inhibition rate was 12.11% for the CQD 0.7 sample.

Comparative study of the propagation and plaque titration conditions for human coronavirus OC43 as a surrogate for SARS-CoV-2.

The ongoing COVID-19 pandemic is threatening human health globally. The development of effective drugs and vaccines against SARS-CoV-2 is hindered by the limited access to high-biosafety-level facilities. Although human coronavirus (HCoV) OC43, a low-pathogenic endemic human coronavirus, has been used as a surrogate virus for SARS-CoV-2 research, a standard technique for HCoV-OC43 culture and plaque titration has not been established. Our objective was to establish optimized culture and titration protocols for HCoV-OC43. The growth kinetics and permissibility to HCoV-OC43 infection of seven different cell lines were examined concurrently at two different temperatures, 33°C and 37°C. Cell lines exhibiting a cytopathic effect (CPE) were selected for plaque titration. No significant difference in the rate of cell growth was observed at the two temperatures tested. Interestingly, HCoV-OC43 was found not to be a high-temperature-sensitive virus, since it grew well at 37°C. Although RD, LLC-MK2, MRC-5, and HCT-8 cell lines supported virus growth with an obvious cytopathic effect and a high yield of virus after two days of infection, only RD cells were suitable for producing countable plaques. The incubation of the cells with 1.2% low-viscosity Avicel as an overlay medium at 37°C for 4 days appeared to promote clearer and sharper plaque morphology. However, further optimization of the plaque titration protocol is still required due to the continued observation of plaque size variation and hazy zones. We propose a cost-effective protocol for HCoV-OC43 culture and plaque titration that can be implemented at a standard conventional temperature without the need for additional special equipment.

Phospholipase Cδ-4 (PLCδ4) Acts as a Nuclear Player to Influence Cyclin B Expression in the Embryonal Rhabdomyosarcoma Cell Lines RD and A204.

Rhabdomyosarcoma (RMS), the most common form of sarcoma typical of pediatric age, arises from the malignant transformation of the mesenchymal precursors that fail to differentiate into skeletal muscle cells. Here, we investigated whether the protein phospholipase C δ4 (PLCδ4), a member of the PLC family involved in proliferation and senescence mechanisms of mesenchymal stromal stem cells, may play a role in RMS. Our molecular and morpho-functional data reveal that PLCδ4 is highly expressed in the fusion-negative, p53-positive, SMARCB1 heterozygous mutated embryonal RMS (ERMS) cell line A204, while it is poorly expressed in the ERMS cell lines RD (fusion-negative, MYC amplification, N-RAS (Q61H), homozygous mutated p53) and Hs729 (homozygous mutated p53) and the alveolar rhabdosarcoma (ARMS) cell line SJCRH30 (RH30; fusion positive, heterozygous mutated RARA, polyheterozygous mutated p53). To characterize the role of PLCδ4, the RD cell line was stably transfected with wild-type PLCδ4 (RD/PLCδ4). Overexpressed PLCδ4 mainly localized to the nucleus in RD cells and contributed to the phosphorylation of PRAS40 (T246), Chk2(T68), WNK1(T60), and Akt 1/273 (S473), as revealed by proteome profiler array analysis. Overexpression of PLCδ4 in RD cells enhanced cyclin B1 expression and resulted in G2/M-phase cell cycle arrest. In contrast, siRNA-mediated knockdown of PLCδ4 in A204 cells resulted in reduced cyclin B1 expression. Our study identifies a novel role for nuclear PLCδ4 as a regulator of cyclin B1 via Akt-dependent phosphorylation. The modulation of PLCδ4 expression and its downstream targets could represent a crucial signaling pathway to block embryonal RMS cell proliferation.

Emergence of Recombinant Subclade D3/Y in Coxsackievirus A6 Strains in Hand-Foot-and-Mouth Disease (HFMD) Outbreak in India, 2022.

Coxsackievirus-A6 (CV-A6) is responsible for more severe dermatological manifestations compared to other enteroviruses such as CV-A10, CV-A16, and EV-A71, causing HFMD in children and adults. Between 2005 and 2007, the recombinant subclade D3/RF-A started to expand globally, and a CV-A6 pandemic started. The study aimed to conduct whole-genome sequencing (WGS) of an isolated CV-A6 strain from currently circulating HFMD cases from India in 2022. Gene-specific RT-PCR and sequencing were used to perform molecular characterization of the isolated virus. Confirmation of these isolates was also performed by transmission electron microscopy and WGS. Among eleven positive clinical enterovirus specimens, eight CV-A6 strains were successfully isolated in the RD cell line. Isolates confirmed the presence of the CV-A6 strain based on VP1 and VP2 gene-specific RT-PCR. Sequences of isolates were clustered and identified as the novel CV-A6 strain of the D3/Y sub-genotype in India. The studies revealed that the D3/Y sub-genotype is being introduced into Indian circulation. The predicted putative functional loops found in VP1 of CV-A6 showed that the nucleotide sequences of the amino acid were a remarkably conserved loop prediction compatible with neutralizing linear epitopes. Therefore, this strain represents a potential candidate for vaccine development and antiviral studies.

Comprehensive Elucidation of the Role of L and 2A Security Proteins on Cell Death during EMCV Infection.

The EMCV L and 2A proteins are virulence factors that counteract host cell defense mechanisms. Both L and 2A exhibit antiapoptotic properties, but the available data were obtained in different cell lines and under incomparable conditions. This study is aimed at checking the role of these proteins in the choice of cell death type in three different cell lines using three mutants of EMCV lacking functional L, 2A, and both proteins together. We have found that both L and 2A are non-essential for viral replication in HeLa, BHK, and RD cell lines, as evidenced by the viability of the virus in the absence of both functional proteins. L-deficient infection led to the apoptotic death of HeLa and RD cells, and the necrotic death of BHK cells. 2A-deficient infection induced apoptosis in BHK and RD cells. Infection of HeLa cells with the 2A-deficient mutant was finalized with exclusive caspase-dependent death with membrane permeabilization, morphologically similar to pyroptosis. We also demonstrated that inactivation of both proteins, along with caspase inhibition, delayed cell death progression. The results obtained demonstrate that proteins L and 2A play a critical role in choosing the path of cell death during infection, but the result of their influence depends on the properties of the host cells.

Chemical Constituents and Anticancer Activities of the Extracts from Phlomis × commixta Rech. f. (P. cretica × P. lanata).

The present work is the first report on the ingredients of the P. × commixta hybrid, a plant of the genus Phlomis. So far, thirty substances have been isolated by various chromatographic techniques and identified by spectroscopic methods, such as UV/Vis, NMR, GC-MS and LC-MS. The compounds are classified as flavonoids: naringenin, eriodyctiol, eriodyctiol-7-O-β-D-glucoside, luteolin, luteolin-7-O-β-D-glucoside, apigenin, apigenin-7-O-β-D-glucoside, diosmetin-7-O-β-D-glucoside, quercetin, hesperetin and quercetin-3-O-β-D-glucoside; phenylpropanoids: martynoside, verbascoside, forsythoside B, echinacoside and allysonoside; chromene: 5,7-dihydroxychromone; phenolic acids: caffeic acid, p-hydroxybenzoic acid, chlorogenic acid, chlorogenic acid methyl ester, gallic acid, p-coumaric acid and vanillic acid; aliphatic hydrocarbon: docos-1-ene; steroids: brassicasterol and stigmasterol; a glucoside of allylic alcohol, 3-O-β-D-apiofuranosyl-(1→6)-O-β-D-glucopyranosyl-oct-1-ene-3-ol, was fully characterized as a natural product for the first time. Two tyrosol esters were also isolated: tyrosol lignocerate and tyrosol methyl ether palmitate, the latter one being isolated as a natural product for the first time. Moreover, the biological activities of the extracts from the different polarities of the roots, leaves and flowers were estimated for their cytotoxic potency. All root extracts tested showed a high cytotoxic activity against the Hep2c and RD cell lines.

Indole Family and Neomycin Sulfate: Inductors of Differentiation in C2C12 and RD cell Lines

Rhabdomyosarcoma (RMS) is a highly aggressive tumor primarily affecting the pediatric population, that generally originates from a failure in the embryonic differentiation of myogenic precursor cells. Standard treatment involves surgery, chemotherapy and radiation therapy, with a poor prognosis, especially when it spreads to other parts of the body, highlighting the need for new treatment approaches. In this study, cellular differentiation was investigated as a potential therapeutic approach for an embryonal subtype of RMS (eRMS) employing a human eRMS cell line, RD; and comparing it with a normal murine myoblast cell line, C2C12. We observed that not only a serum free or a low serum concentration in the cultured medium induce morphological changes towards differentiations, but also staurosporine, bisindolylmaleimide, and neomycin sulfate (direct or indirect protein kinase inhibitors) induces differentiation in RD and C2C12 cell lines, suggesting a crucial role of PKC, PLC and its signaling pathways in a proliferation/differentiation switch. These findings underscore the importance of exploring cellular differentiation as a therapeutic strategy against RMS and highlight the need for further research into the underlying mechanisms of kinase inhibitor-induced cellular differentiation as a potential cancer treatment.

Epidemiology and molecular detection of human adenovirus and non-polio enterovirus in fecal samples of children with acute gastroenteritis: A five-year surveillance in northern Brazil.

Acute gastroenteritis (AGE) is a common pediatric infection that remains a significant cause of childhood morbidity and mortality worldwide, especially in low-income regions. Thus, the objective of this study was to detect human adenovirus (HAdV) and non-polio enterovirus (NPEV) in fecal samples from the Gastroenteritis Surveillance Network, and to identify circulating strains by nucleotide sequencing. A total of 801 fecal samples were tested using qPCR/RT-qPCR, and 657 (82.0%) were inoculated into HEp-2C and RD cell lines. The HAdV and NPEV positivity rates obtained using qPCR/RT-qPCR were 31.7% (254/801) and 10.5% (84/801), respectively, with 5.4% (43/801) co-detection. Cytopathic effect was observed in 9.6% (63/657) of patients, 2.7% (18/657) associated with HAdV, and 6.2% (41/657) associated with NPEV after testing by ICC-PCR. A comparison of the two methodologies demonstrated an agreement of 93.5% for EVNP and 64.4% for HAdV. These two viruses were detected throughout the study period, with HAdV positivity rates ranging from 41% in Amapá to 18% in Pará. The NEPV varied from 18% in Pará/Rondônia to 3% in Acre. The most affected age group was over 60 months for both HAdV and NPEV. Samples previously positive for rotavirus and norovirus, which did not show a major difference in the presence or absence of diarrhea, fever, and vomiting, were excluded from the clinical analyses of these two viruses. These viruses circulated over five years, with a few months of absence, mainly during the months corresponding to the waves of SARS-CoV-2 infection in Brazil. Five HAdV species were identified (A, B, C, D, and F), with a greater predominance of HAdV-F41 (56.5%) followed by HAdV-C (15.2%). Three NPEV species (A, B, and C) were detected, with serotypes E14 (19.3%) and CVA-24 (16.1%) being the most prevalent. The present study revealed a high diversity of NPEV and HAdV types circulating in children with AGE symptoms in the northern region of Brazil.

Native Medicinal Plants (Moringa oleifera Lam, Brucea javanica (L.) Merr., Eclipta prostrata (L.), Callisia fragrans (Lindl.) Woodson, and Zingiber zerumbet (L.) Smith) in An Giang, Vietnam: A Preliminary Investigation for Rhabdomyosarcoma Treatments using in-vitro RD cell cytotoxicity test

Cancer, one of the deadliest diseases worldwide, is projected to affect 30.2 million people by 2040. Among the various cancer types, rhabdomyosarcoma (RMS) is a unique tumor primarily impacting the muscular system of children. The current treatment for RMS has limited efficacy and numerous side effects, emphasizing the need for novel therapeutic approaches. This study investigates the potential treatment of the RMS cell line RD using extracts from five folklore-based medicinal plants in An Giang, Vietnam. The plants—Moringa oleifera Lam, Brucea javanica (L.) Merr., Eclipta prostrata (L.), Callisia fragrans (Lindl.) Woodson, and Zingiber zerumbet (L.) Smith—were extracted and fractionated using three solvents: ether, ethanol, and water. These fractions underwent phytochemical screening and cytotoxicity testing on the in-vitro RMS cell line RD. The results indicate that the ether fraction of Eclipta prostrata (L.) and the ether and ethanol fractions of Zingiber zerumbet (L.) Smith exhibit moderate cytotoxic effects on RD cell lines, with IC50 values of 37.08 ± 1.23 µg/mL, 23.15 ± 1.17 µg/mL, and 45.63 ± 2.39 µg/mL, respectively. These findings provide preliminary data for further in-depth research into the anticancer properties of these plants, which are widely grown in the South of Vietnam.

Exometabolites of streptomycetes isolated from the Odesa bay exhibit a toxic effect against human cancer cell lines

The aim of the work was to determine the cytotoxic activity of marine actinobacteria exometabolites on the human larynx adenocarcinoma Hep-2 and rhabdomyosarcoma RD cell lines. Exometabolites of 6 strains of Streptomyces sp. were extracted with ethyl acetate and their composition was determined with UltiMate™ 3000 BioRS UPLC System coupled to mass spectrometer. Cytotoxic activity of exometabolites against cancer cells was determined after 24 hours of incubation by microscopic examination of cell morphological changes and the degree of cell monolayer degeneration. Exometabolites from Streptomyces sp. Myt7b strain showed the highest cytotoxic activity and at concentration of 500.0 μg/ml caused up to 90.0% death of RD and Hep-2 cells. The presence of 53 identified compounds with potential cytotoxic activity, including enterocin, 6-prenyltryptophol, medelamine A and turbinaric acid was detected in Streptomyces sp. Myt7b exometabolome. It is concluded that this strain is a promising producer of compounds with anticancer activity. Keywords: cytotoxic activity, exometabolites, marine streptomycetes, RD and Hep-2 cancer cells

Capparis cartilaginea Decne. as a natural source of antioxidant, anti-inflammatory, and anti-cancer herbal drug

BackgroundCapparis cartilaginea Decne.(C. cartilaginea Decne.) a medicinally important plant that belongs to the family Capparaceae. Traditionally the fruit, leaf, and root have been believed to show anti-inflammatory effects and can also be used as agents. PurposeThe objective of this study is to investigate the chemical compositions, in-vitro antioxidant, anti-cancer and anti-inflammatory activities of the ethanolic extract of Capparis cartilaginea Decne. leaves. Study methodsThe chemical compounds were investigated by Gas Chromatography–Mass Spectrometry (GC–MS) analysis. Ferric-bipyridine reducing capacity of total antioxidants (FBRC), Ferric reducing antioxidant power (FRAP) were utilized to determine antioxidant activity. The cytotoxic activity was evaluated using a 3-[4,5-dimethylthiazole- 2-yl]-2,5-diphenyl tetrazolium bromide (MTT) assay. Additionally, in vitro anti-inflammatory activity of the ethanolic extract of C. cartilaginea Decne. leaves using COX1 and protein denaturation methods. ResultsThe major twenty- four bioactive compounds from fifty compounds were identified including alkaloids, flavonoids, phenols, fatty acids, and carotenes. Pleiocarpamine was the major constituent of the extract (73.07 %). Additionally, in vitro anti-inflammatory activity of the ethanolic extract of C. cartilaginea Decne. leaves using COX1 and protein denaturation methods showed a strong effect with IC50 values of 60.23 and 17.67 µg/mL respectively, compared to Indomethacin and Diclofenac as a standard anti-inflammatory drug.; while FBRC and FRAP were utilized to determine antioxidant activity slowed high antioxidants capacity with values 0.056 ± 0.001and 0.03 ± 0.00196 Ascorbic acid equivalent/g of extract (AAE) and DPPH radical scavenging method show IC50 value 10 ± 1 %. Evaluation of the cytotoxic activity against three human cancer cell lines colon carcinoma cells HCT-116, Breast carcinoma cells MCF-7, and human muscle Rhabdomyosarcoma RD cell line using MTT assay showed a significant effect with IC50 values 59.58 ± 1.5, 39.14 ± 3.09, and 102 ± 8.75 µg/mL, compared to Vinblastine Sulfate as a standard cytotoxic drug. Yemeni C. cartilaginea Decne. showed a higher effect on MCF-7 compared to Iranian C. cartilaginea Decne. as well as established a higher antioxidant activity than that extracts obtained from Jordan. ConclusionEthanolic leaves extract of C. cartilaginea Decne. have the potential for development as therapeutic agents for cytotoxicity on certain cancer cells following further investigation. C. cartilaginea Decne. leaves that could have medicinal values and application in industries, were used to design the drug for the treatment of various types of inflammatory diseases.

Antibacterial and Cytotoxic Activity of Mentha arvensis L. Leaves Methanolic Extract In vitro

Treatment with herbal drugs is considered one of the oldest and safest methods for treating various diseases. Mentha arvensis is a medicinal plant used worldwide in the pharmaceutical industry for its antibacterial, anticancer, antiallergic, and anti-inflammatory properties. In the present study, the antibacterial activity of the methanolic extract of M. arvensis leaves was tested at different concentrations (50, 100, 150, and 200 µg/ml) against bacteria isolated from human burn wound infections, including Pseudomonas aeruginosa, Klebsiella pneumoniae and Staphylococcus aureus. The results demonstrated that at concentrations of 100, 150, and 200 µg/ml, the methanolic extract exhibited strong antibacterial activity against S. aureus and P. aeruginosa, while K. pneumoniae isolates were resistant to the extract at all tested concentrations. Further, the cytotoxic activity of the methanolic extract was assessed at various concentrations (100, 140, 160, 180, 220, and 250 µg/ml) against two cell lines: the RD tumor cell line and Vero normal cell line. The higher concentrations of the extract (220 and 250 µg/ml) showed potent cytotoxic effects on the RD cell line after 72 hours in a dose- and time-dependent manner. In contrast, the extract exhibited lower cytotoxic effects on Vero cells at the same concentrations after 72 h.

Discovery of Dual Lysine Methyltransferase G9a and EZH2 Inhibitors with In Vivo Efficacy against Malignant Rhabdoid Tumor.

In recent years, it has been proposed that G9a/EZH2 dual inhibition is a promising cancer treatment strategy. Herein, we present the discovery of G9a/EZH2 dual inhibitors that merge the pharmacophores of G9a and EZH2 inhibitors. Among them, the most promising compound 15h displayed potent inhibitory activities against G9a (IC50 = 2.90 ± 0.05 nM) and EZH2 (IC50 = 4.35 ± 0.02 nM), superior antiproliferative profiles against RD (CC50 = 19.63 ± 0.18 μM) and SW982 (CC50 = 19.91 ± 0.50 μM) cell lines. In vivo, 15h achieved significant antitumor efficacy in a xenograft mouse model of human rhabdoid tumor with a tumor growth inhibitory rate of 86.6% without causing observable toxic effects. The on-target activity assays illustrated that compound 15h can inhibit tumor growth by specifically inhibiting EZH2 and G9a. Therefore, 15h is a potential anticancer drug candidate for the treatment of malignant rhabdoid tumor.

Abstract 3537: A novel orthotopic xenograft model of spontaneous metastasis for rhabdomyosarcoma using lingual intramuscular injection

Abstract Rhabdomyosarcoma (RMS), a cancer characterized by the defective differentiation of skeletal muscle precursor cells, is the most common pediatric soft tissue sarcoma. Despite significant efforts to characterize the transcriptomic and genetic abnormalities of the disease, the prognosis for patients with advanced or refractory disease has remained unchanged for more than four decades. Preclinically, studies have been hampered by insufficient animal models. For example, the hindlimb orthotopic model, an injection into the gastrocnemius muscle of immunocompromised mice, produces highly encapsulated, non-invasive, non-metastatic tumors. While this model has shown a reasonable recapitulation of tumor proliferation response, investigators using this model are unable to assess the effect of their investigational treatment on local invasion or distant metastasis. Moreover, transgenic mouse models of RMS, while commonly metastatic, diverge significantly from the transcriptomic signature of human tumors. With these considerations, we hypothesized that an alternative orthotopic site, the tongue, may provide a more representative xenograft model of RMS. In a proof of principal experiment, the fusion negative RMS (FN-RMS) cell line RD was injected into the skeletal muscle of the tongue, a lingual intramuscular (LIM) injection. Tongue tumors establish in approximately 60-80% of mice injected within approximately 40 days. By histopathology, the primary tumors show evidence of local invasion. Interestingly, approximately 60% of these tumors also spread to local cervical lymph nodes, a process not observed in hindlimb orthotopic models or subcutaneous models. We expanded our investigation to include additional FN-RMS cell lines: RMS-YM, SMS-CTR, and a MEK inhibitor resistant cell line CTR-557. Mice injected with RMS-YM cells established tumors in at least 60% of mice by 75 days post injection. Additionally, enlarged local and distant lymph nodes are observed in 50% of these mice. Intriguingly, mice injected with SMS-CTR or its MEK inhibitor resistant derivative, CTR-557, frequently have tumors in both local and distant lymph nodes without evidence of a primary tongue tumor. One mouse in each group also had pulmonary metastases. Taken together, this model not only demonstrates the first orthotopic xenograft model of spontaneous metastasis for FN-RMS, it also mimics the spread of disease that is observed clinically in RMS originating in the head and neck. Ongoing and future studies include 1) generating highly metastatic human FN-RMS cell lines; 2) using spatial transcriptomics to compare the tumor microenvironment of non-metastatic hindlimb tumors and invasive and/or metastatic lingual tumors; and 3) identifying drivers of FN-RMS dissemination through transcriptomic, genome accessibility, and phospho-proteomic methods. Citation Format: Katie E. Hebron, Kristine Isanogle, Amy James, Simone Difilippiantonio, Marielle E. Yohe. A novel orthotopic xenograft model of spontaneous metastasis for rhabdomyosarcoma using lingual intramuscular injection. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3537.

Enterovirus detection in different regions of Madagascar reveals a higher abundance of enteroviruses of species C in areas where several outbreaks of vaccine-derived polioviruses occurred

BackgroundPoliomyelitis outbreaks due to pathogenic vaccine-derived polioviruses (VDPVs) are threatening and complicating the global polio eradication initiative. Most of these VDPVs are genetic recombinants with non-polio enteroviruses (NPEVs) of species C. Little is known about factors favoring this genetic macroevolution process. Since 2001, Madagascar has experienced several outbreaks of poliomyelitis due to VDPVs, and most of VDPVs were isolated in the south of the island. The current study explored some of the viral factors that can promote and explain the emergence of recombinant VDPVs in Madagascar.MethodsBetween May to August 2011, we collected stools from healthy children living in two southern and two northern regions of Madagascar. Virus isolation was done in RD, HEp-2c, and L20B cell lines, and enteroviruses were detected using a wide-spectrum 5ʹ-untranslated region RT-PCR assay. NPEVs were then sequenced for the VP1 gene used for viral genotyping.ResultsOverall, we collected 1309 stools, of which 351 NPEVs (26.8%) were identified. Sequencing revealed 33 types of viruses belonging to three different species: Enterovirus A (8.5%), Enterovirus B (EV-B, 40.2%), and Enterovirus C (EV-C, 51.3%). EV-C species included coxsackievirus A13, A17, and A20 previously described as putative recombination partners for poliovirus vaccine strains. Interestingly, the isolation rate was higher among stools originating from the South (30.3% vs. 23.6%, p-value = 0.009). EV-C were predominant in southern sites (65.7%) while EV-B predominated in northern sites (54.9%). The factors that explain the relative abundance of EV-C in the South are still unknown.ConclusionsWhatever its causes, the relative abundance of EV-C in the South of Madagascar may have promoted the infections of children by EV-C, including the PV vaccine strains, and have favored the recombination events between PVs and NPEVs in co-infected children, thus leading to the recurrent emergence of recombinant VDPVs in this region of Madagascar.

Biological Evaluation of a Series of Amine-Containing Mixed-Ligand Copper(II) Coordination Compounds with 2-(2-hydroxybenzylidene)-N-(prop-2-en-1-yl)hydrazinecarbothioamide.

Five compounds 2-(2-hydroxybenzylidene)-N-(prop-2-en-1-yl)hydrazinecarbothioamide (H2L), bis[μ2-2-({2-[(prop-2-en-1-yl)carbamothioyl]hydrazinylidene}methyl)phenolato-S,N,O:O]diaquadicopper(II) nitrate (1), bis[μ2-2-({2-[(prop-2-en-1-yl)carbamothioyl]hydrazinylidene}methyl)phenolato-S,N,O:O]diimidazoldicopper(II) nitrate (2), bis[μ2-2-({2-[(prop-2-en-1-yl)carbamothioyl]-hydrazinylidene}methyl)phenolato-S,N,O:O]bis-(3,5-dibromopyridine)dicopper(II) nitrate (3), bis[μ2-2-({2-[(prop-2-en-1-yl)carbamothioyl]-hydrazinylidene}methyl)phenolato-S,N,O:O]bis(4-methylpyridine)dicopper(II) nitrate hexahydrate (4) were synthesized. The antiproliferative properties of these compounds toward cancer cell lines RD, HeLa, and normal cell line MDCK have been investigated. The tested complexes surpass Doxorubicin (DOXO) in the efficiency of anticancer activity as their IC50 values toward cancer cells are lower than the corresponding values of DOXO and the selectivity indexes exceed the corresponding SI value of DOXO. The tested compounds demonstrated a high antioxidant effect against ABTS•+ radical cations as well as low toxicity on Daphnia magna.

Environmental Surveillance of Poliovirus in Four Districts within Two Regions of Ghana

The eradication of poliovirus is at its last phase through the efforts and strategies of Global Polio Eradication Initiation (GPEI). There are very few countries that are still endemic with Wild Poliovirus (WPV) and others with circulating Vaccine Derived Poliovirus (cVDPV). The aim of the study was to detect silent circulation of WPV and VDPV in four districts within the Eastern and Volta region of Ghana. A systematic longitudinal design was used for the study. The convenient sampling technique was used to collect the samples every four weeks from two open and close sewage systems. The open sewage systems were located in New Juabeng and Ho districts while the close sewage systems were located in Asuogyaman and Ketu South districts. A total of 35 sewage samples were collected from September 2018 to May 2019. L20B and RD cell lines were used for the purification of Poliovirus (PV) while real-time Reverse Transcriptase Polymerase Chain Reaction (rRT-PCR) was use to characterize the serotypes of the PVs. The findings of the study showed that the prevalence of Non-Polio Enterovirus (NPEV) and Sabin were 65.71% and 14.29% respectively. The characterized Sabins were serotype 1 and serotype 3 which were circulating in the two districts within the Eastern Region. The study did not detect any WPV and VDPV but isolated Sabin strains of the poliovirus. This necessitates the need for continuous environmental surveillance for poliovirus nationwide.

MET Inhibition Sensitizes Rhabdomyosarcoma Cells to NOTCH Signaling Suppression.

Rhabdomyosarcoma (RMS) is a pediatric myogenic soft tissue sarcoma. The Fusion-Positive (FP) subtype expresses the chimeric protein PAX3-FOXO1 (P3F) while the Fusion-Negative (FN) is devoid of any gene translocation. FP-RMS and metastatic FN-RMS are often unresponsive to conventional therapy. Therefore, novel therapeutic approaches are needed to halt tumor progression. NOTCH signaling has oncogenic functions in RMS and its pharmacologic inhibition through γ-secretase inhibitors blocks tumor growth in vitro and in vivo. Here, we show that NOTCH signaling blockade resulted in the up-regulation and phosphorylation of the MET oncogene in both RH30 (FP-RMS) and RD (FN-RMS) cell lines. Pharmacologic inhibition of either NOTCH or MET signaling slowed proliferation and restrained cell survival compared to control cells partly by increasing Annexin V and CASP3/7 activation. Co-treatment with NOTCH and MET inhibitors significantly amplified these effects and enhanced PARP1 cleavage in both cell lines. Moreover, it severely hampered cell migration, colony formation, and anchorage-independent growth compared to single-agent treatments in both cell lines and significantly prevented the growth of FN-RMS cells grown as spheroids. Collectively, our results unveil the overexpression of the MET oncogene by NOTCH signaling targeting in RMS cells and show that MET pathway blockade sensitizes them to NOTCH inhibition.

Combined Treatment with PI3K Inhibitors BYL-719 and CAL-101 Is a Promising Antiproliferative Strategy in Human Rhabdomyosarcoma Cells.

Rhabdomyosarcoma (RMS) is a highly malignant and metastatic pediatric cancer arising from skeletal muscle myogenic progenitors. Recent studies have shown an important role for AKT signaling in RMS progression. Aberrant activation of the PI3K/AKT axis is one of the most frequent events occurring in human cancers and serves to disconnect the control of cell growth, survival, and metabolism from exogenous growth stimuli. In the study reported here, a panel of five compounds targeting the catalytic subunits of the four class I PI3K isoforms (p110α, BYL-719 inhibitor; p110β, TGX-221 inhibitor; p110γ, CZC24832; p110δ, CAL-101 inhibitor) and the dual p110α/p110δ, AZD8835 inhibitor, were tested on the RMS cell lines RD, A204, and SJCRH30. Cytotoxicity, cell cycle, apoptosis, and the activation of downstream targets were analyzed. Of the individual inhibitors, BYL-719 demonstrated the most anti-tumorgenic properties. BYL-719 treatment resulted in G1/G0 phase cell cycle arrest and apoptosis. When combined with CAL-101, BYL-719 decreased cell viability and induced apoptosis in a synergistic manner, equaling or surpassing results achieved with AZD8835. In conclusion, our findings indicate that BYL-719, either alone or in combination with the p110δ inhibitor, CAL-101, could represent an efficient treatment for human rhabdomyosarcoma presenting with aberrant upregulation of the PI3K signaling pathway.

D,L-Methadone enhances the cytotoxic activity of standard chemotherapeutic agents on pediatric rhabdomyosarcoma

PurposeIn advanced tumor stages, pediatric rhabdomyosarcoma (RMS) shows an intrinsic resistance to standard chemotherapy, which is associated with a dismal prognosis. Alternative therapeutic approaches and optimization of already existent treatment protocols are urgently needed in these conditions. The µ-opioid receptor (OPRM1) agonist, D,L-methadone is frequently used for analgesia in oncological patients. Recent evidence has shown that D,L-methadone in combination with chemotherapeutic agents may enhance their cytotoxic effect against cancer cells. There are no related data in pediatric rhabdomyosarcoma (RMS).MethodsAntitumor effects of combined D,L-methadone and doxorubicin, carboplatin, and vincristine on RMS cell lines RD and RH30 were analyzed using following outcome data: expression of the OPRM1 receptor (Western blot), cell growth inhibition (MTT assay), cell migration (wound-healing assay), apoptosis induction (caspase-3/7 assay), and reactive oxygen species (ROS) production (flow cytometry).ResultsIn both cell lines, OPRM1 expression was significantly increased after combined treatment of D,L-methadone with all three cytotoxic drugs tested, which resulted in suppression of tumor cell growth and increase of apoptosis rates. These effects were mediated by increased ROS production and up-regulation of caspase-3/7 activity. Doxorubicin combined with D,L-methadone significantly reduced cell migration in both cell lines. Carboplatin or vincristine in combination with D,L-methadone had only an impact on cell migration in RH30 cells.ConclusionsThis new therapeutic approach in RMS provides strong antitumor effects in vitro. The combination of standard chemotherapy and D,L-methadone requires further investigation. Especially advanced tumors with a limited effectiveness of conventional treatment regimens seem a potential target of this approach.