Strengthening laboratories in response to outbreaks in humanitarian emergencies and conflict settings: Results, challenges and lessons from expanding PCR diagnostic capacities for COVID-19 testing in Yemen.

BACKGROUND: When the COVID-19 pandemic was declared, Yemen, a country facing years of conflict had only one laboratory with PCR testing capacity. In this article, we describe the outcome of the implementation of molecular based diagnostics platform in Yemen and highlight the key milestones the country went through to increase access to testing for its populations residing in a geographically vast and politically divided country. METHODS: A retrospective assessment of COVID-19 laboratory response activities was done detailing the needs assessment process, timelines, geographical coverage, and outcomes of the activities. Laboratory data was analyzed to construct the geographical locations of COVID-19 testing laboratories and the numbers of tests performed in each facility to highlight the demands of testing for travelers. Finally, we discuss the impact these activities had in enabling the movement of people across international borders for economic gains and in delivery of critical humanitarian aid. OUTCOME: PCR testing capacities in Yemen significantly improved, from one laboratory in Sanaa in April 2020 to 18 facilities across the country by June 2022. In addition, the number of functional Real-Time PCR thermocyclers increased from one to 32, the PCR tests output per day improved from 192 to 6144 tests per day. Results from analysis of laboratory data showed there were four peaks of COVID-19 in Yemen as October 2022. The majority of laboratory tests were performed for travelers than for medical or public health reasons. Demand for laboratory testing in Yemen was generally low and waned over time as the perceived risk of COVID-19 declined, in parallel with rollout of the COVID-19 vaccines. DISCUSSION/CONCLUSION: The successful expansion of laboratory testing capacity was instrumental in the control and management of COVID-19 cases and critical in the implementation of public response strategies, including restrictions on gathering. Laboratory testing also facilitated the movement of humanitarian agencies and delivery of aid and enabled hundreds of thousands of Yemeni nationals to travel internationally. By virtue of these outcomes, the impact of laboratory strengthening activities was thus felt in the health sector and beyond.

Immunoinformatics, molecular docking and dynamics simulation approaches unveil a multi epitope-based potent peptide vaccine candidate against avian leukosis virus.

Lymphoid leukosis is a poultry neoplastic disease caused by avian leukosis virus (ALV) and is characterized by high morbidity and variable mortality rates in chicks. Currently, no effective treatment and vaccination is the only means to control it. This study exploited the immunoinformatics approaches to construct multi-epitope vaccine against ALV. ABCpred and IEDB servers were used to predict B and T lymphocytes epitopes from the viral proteins, respectively. Antigenicity, allergenicity and toxicity of the epitopes were assessed and used to construct the vaccine with suitable adjuvant and linkers. Secondary and tertiary structures of the vaccine were predicted, refined and validated. Structural errors, solubility, stability, immune simulation, dynamic simulation, docking and in silico cloning were also evaluated.The constructed vaccine was hydrophilic, antigenic and non-allergenic. Ramchandran plot showed most of the residues in the favored and additional allowed regions. ProsA server showed no errors in the vaccine structure. Immune simulation showed significant immunoglobulins and cytokines levels. Stability was enhanced by disulfide engineering and molecular dynamic simulation. Docking of the vaccine with chicken's TLR7 revealed competent binding energies.The vaccine was cloned in pET-30a(+) vector and efficiently expressed in Escherichia coli. This study provided a potent peptide vaccine that could assist in tailoring a rapid and cost-effective vaccine that helps to combat ALV. However, experimental validation is required to assess the vaccine efficiency.

Responding to COVID-19 pandemic at the local level: lessons from Yemen.

Background: The health authorities in Hadhramaut Valley and Desert responded to the COVID-19 pandemic differently from other areas in Yemen. Aims: To document the response of the local authority and Ministry of Health in Hadhramaut to COVID-19. Methods: The local authority in Syoun (Hadhramaut Valley) convened a meeting of all key players from the health and related sectors in February 2020 where a decision was made to establish a committee to evaluate the health situation and assess the needs. Based on the results of these assessments, a plan was designed to respond to the pandemic. We reviewed available documents on the COVID-19 response in Hadhramaut, interviewed the main stakeholders, and conducted site visits to the COVID-19 response centres. Results: There was evidence of the crucial role played by the local authority in response to COVID-19. They established 3 well-equipped isolation centres with a total of 142 beds, a stock of 2250 oxygen cylinders, 2 new polymerase chain reaction units, a simplified referral system, and an effective patient follow-up and oxygen home therapy strategy. Conclusion: Political commitment at the local level is crucial to bridge the gap between policy and implementation, especially during infectious disease outbreaks. It is important to train public health leaders on how to effectively assess local health needs and develop effective and efficient response strategy. Lessons from this study in Hadhramaut provide evidence on how local authorities can coordinate response to emerging health needs and update their strategies.

Ensuring access to novel COVID-19 therapeutics in Pacific island countries and areas.

Problem: As of November 2022, over 417 397 confirmed cases and 2631 deaths related to coronavirus disease (COVID-19) were reported in Pacific island countries and areas (PICs). Most PICs have faced challenges accessing therapeutics recommended for the treatment of COVID-19 due to their high demand worldwide and supply chain constraints. Context: The World Health Organization (WHO) coordinates and provides tailored technical and operational support to 21 PICs. Since the start of the pandemic, WHO has worked with partners to establish a mechanism to ensure equitable access to three novel COVID-19 therapeutics (tocilizumab, molnupiravir and nirmatrelvir/ritonavir) for lower-income countries, including 11 eligible PICs. Action: WHO coordinated the requests, procurement and distribution of the three novel therapeutics. In addition, WHO supported PICs by providing trainings in clinical management of COVID-19, developing critical supply needs estimates, and facilitating regulatory approval of clinical therapeutics, including emergency use authorization. Lessons learned: The main barriers to procurement of novel COVID-19 therapeutics were identified as prolonged negotiations with licence holders, sourcing funding, the high cost of therapeutics and limited capacity to provide safety monitoring. Discussion: Uninterrupted supply and availability of essential medicines in the Pacific region is dependent on external and local sourcing. To overcome procurement barriers and ensure access to novel COVID-19 therapeutics in PICs, WHO's pandemic support to Member States focused on strengthening regulatory requirements, safety monitoring and supply chain activities.

A drug repurposing approach for individualized cancer therapy based on transcriptome sequencing and virtual drug screening.

RNA-sequencing has been proposed as a valuable technique to develop individualized therapy concepts for cancer patients based on their tumor-specific mutational profiles. Here, we aimed to identify drugs and inhibitors in an individualized therapy-based drug repurposing approach focusing on missense mutations for 35 biopsies of cancer patients. The missense mutations belonged to 9 categories (ABC transporter, apoptosis, angiogenesis, cell cycle, DNA damage, kinase, protease, transcription factor, tumor suppressor). The highest percentages of missense mutations were observed in transcription factor genes. The mutational profiles of all 35 tumors were subjected to hierarchical heatmap clustering. All 7 leukemia biopsies clustered together and were separated from solid tumors. Based on these individual mutation profiles, two strategies for the identification of possible drug candidates were applied: Firstly, virtual screening of FDA-approved drugs based on the protein structures carrying particular missense mutations. Secondly, we mined the Drug Gene Interaction (DGI) database (https://www.dgidb.org/) to identify approved or experimental inhibitors for missense mutated proteins in our dataset of 35 tumors. In conclusion, our approach based on virtual drug screening of FDA-approved drugs and DGI-based inhibitor selection may provide new, individual treatment options for patients with otherwise refractory tumors that do not respond anymore to standard chemotherapy.

A novel strategy for developing vaccine candidate against Jaagsiekte sheep retrovirus from the envelope and gag proteins: an in-silico approach.

BACKGROUND: Sheep pulmonary adenocarcinoma (OPA) is a contagious lung cancer of sheep caused by the Jaagsiekte retrovirus (JSRV). OPA typically has a serious economic impact worldwide. A vaccine has yet to be developed, even though the disease has been globally spread, along with its complications. This study aimed to construct an effective multi-epitopes vaccine against JSRV eliciting B and T lymphocytes using immunoinformatics tools. RESULTS: The designed vaccine was composed of 499 amino acids. Before the vaccine was computationally validated, all critical parameters were taken into consideration; including antigenicity, allergenicity, toxicity, and stability. The physiochemical properties of the vaccine displayed an isoelectric point of 9.88. According to the Instability Index (II), the vaccine was stable at 28.28. The vaccine scored 56.51 on the aliphatic index and -0.731 on the GRAVY, indicating that the vaccine was hydrophilic. The RaptorX server was used to predict the vaccine's tertiary structure, the GalaxyWEB server refined the structure, and the Ramachandran plot and the ProSA-web server validated the vaccine's tertiary structure. Protein-sol and the SOLPro servers showed the solubility of the vaccine. Moreover, the high mobile regions in the vaccine's structure were reduced and the vaccine's stability was improved by disulfide engineering. Also, the vaccine construct was docked with an ovine MHC-1 allele and showed efficient binding energy. Immune simulation remarkably showed high levels of immunoglobulins, T lymphocytes, and INF-γ secretions. The molecular dynamic simulation provided the stability of the constructed vaccine. Finally, the vaccine was back-transcribed into a DNA sequence and cloned into a pET-30a ( +) vector to affirm the potency of translation and microbial expression. CONCLUSION: A novel multi-epitopes vaccine construct against JSRV, was formed from B and T lymphocytes epitopes, and was produced with potential protection. This study might help in controlling and eradicating OPA.

Naphthoquinone derivatives as P-glycoprotein inducers in inflammatory bowel disease: 2D monolayers, 3D spheroids, and in vivo models.

Inflammatory bowel disease (IBD) represents a chronic inflammation of the gastrointestinal tract characterized by an overreaction of immune responses and damage at the intestinal mucosal barrier. P-glycoprotein (P-gp) plays a key role to protect the intestinal barrier from xenobiotic accumulation and suppressing excessive immune responses. Therefore, induction/activation of P-gp function could serve as a novel therapeutic target to treat IBD. This study aimed to evaluate the potential therapeutic values of naphthoquinone derivatives (NQ-1 - NQ-8) as P-gp modulators to counterbalance intestinal inflammation. The data indicate that NQ-2, NQ-3, and NQ-4 act as P-gp inducers/activators and are recognized as substrates for P-gp. The three derivatives possess anti-inflammatory effects mediated by suppression of NF-κB and HDAC6 activity in Caco2 monolayer cells. Besides, they reversed LPS-induced intestinal barrier dysfunction by enhancing the expression of P-gp and ZO-1 tight junction proteins in a Caco-2 spheroid model. NQ-2, NQ-3, and NQ-4 showed a robust inhibitory effect on IL-1ß maturation in LPS-primed THP-1 cells. This effect may contribute to alleviate the inflammatory cascades associated with IBD. Distinctively, NQ-2 and NQ-3 exerted anti-NLRP3 inflammasome activity evidenced by the inhibition of CASP-1 activity and the promotion of autophagy. Both compounds induced disruptions of the microtubule network in transfected U2OS-GFP-α-tubulin cells. Treatment with NQ-2 remarkably attenuated dextran sulfate sodium (DSS)-induced colitis in rats by suppressing changes in colon length, colon mass index, and intestinal histopathology scores. Thus, 1,4-naphthoquinone derivatives such as NQ-2 may provide potential therapeutic anti-inflammatory effects for IBD patients and for other NLRP3-associated inflammatory diseases.

Nimbolide inhibits 2D and 3D prostate cancer cells migration, affects microtubules and angiogenesis and suppresses B-RAF/p.ERK-mediated in vivo tumor growth.

BACKGROUND: Prostate cancer (PCa) is the most prominent malignancy among men worldwide. PCa cells have a high tendency to metastasize to various distant organs, and this activity is the main cause of PCa mortality. Nimbolide is a promising phytochemical constituent of neem Azadirachta indica (Meliaceae). Previous studies showed that nimbolide exhibited potent anticancer activity however, its role against PCa tumorigenesis has not been fully elucidated. PURPOSE: Our work aims to explore the role of nimbolide in regulating the essential tumor-associated processes involved in the metastatic cascade in PCa cells. STUDY DESIGN: Cytotoxicity assay, wound healing and spheroid invasion assays, western blotting, immunofluorescence, tube-formation assay, in vivo and immunohistochemistry. METHODS: The cytotoxicity of nimbolide towards PCa cell lines was assessed by resazurin assays. The cell mobility and migration of nimbolide-treated DU145 cells were determined by wound healing and spheroid invasion assays. Tubulin network was visualized using U2OS cells and DU145 cells. The effect of nimbolide on E-cadherin, ß-catenin, acetylated α-tubulin and HDAC6 protein expressions levels were measured by Western blot. The potentiality of nimbolide to inhibit angiogenesis was revealed by HUVEC tube-formation assay. Nimbolide antitumor effect was studied in a syngeneic model of murine prostate cancer. RESULTS: The current study indicated that nimbolide negatively affected the migratory and invasive capacity of DU145 prostate cancer cells in 2D and three-dimensional (3D) spheroid cultures. Interestingly, nimbolide induced downregulation of E-cadherin without any influence on the expression level of ß-catenin. Additionally, we demonstrated that nimbolide influenced the microtubule network which was supported by the upregulation of acetylated α-tubulin and the reduction in HDAC6 protein. Moreover, the inhibitory effect of nimbolide on angiogenesis was clearly observed in HUVEC tube formation assay. In vivo experiments revealed the significant suppression of PCa growth and targeting of the B-RAF/p.ERK signaling pathway by nimbolide. CONCLUSION: Our results showed that nimbolide inhibited 2D and 3D prostate cancer cells migration and downregulated E-cadherin protein expression, a marker for metastatic chemoresistance and tumor recurrence. Nimbolide stabilized the microtubules, combated angiogenesis and suppressed B.RAF/ERK-mediated in vivo tumor growth. Nimbolide may be considered as potential therapeutic agent for metastatic and advanced PCa patients and merits further investigations.

SARS-CoV-2 seroprevalence in Aden, Yemen: a population-based study.

BACKGROUND: In Yemen, initial surveillance of coronavirus disease 2019 (COVID-19) focused primarily on patients with symptoms or severe disease. The full spectrum of the disease remains unclear. To the best of the authors' knowledge, this is the first seroprevalence study performed in Yemen. METHODS: This cross-sectional investigation included 2001 participants from all age groups from four districts in Aden, southern Yemen. A multi-stage sampling method was used. Data were collected using a well-structured questionnaire, and blood samples were taken. Healgen COVID-19 IgG/IgM Rapid Diagnostic Test (RDT) Cassettes were used in all participants. All positive RDTs and 14% of negative RDTs underwent enzyme-linked immunosorbent assay (ELISA) testing (WANTAI SARS-CoV-2 Ab ELISA Kit) for confirmation. RESULTS: In total, 549 of 2001 participants were RDT positive and confirmed by ELISA, giving a prevalence of COVID-19 of 27.4%. The prevalence of immunoglobulin G was 25%. The prevalence of asymptomatic COVID-19 in the entire study group was 7.9%. The highest prevalence was observed in Al-Mansurah district (33.4%). Regarding sociodemographic factors, the prevalence of COVID-19 was significantly higher among females, housewives and subjects with a history of contact with a COVID-19 patient: 32%, 31% and 39%, respectively. CONCLUSION: This study found high prevalence of COVID-19 in the study population. Household transmission was common.

Community-based surveillance in internally displaced people's camps and urban settings during a complex emergency in Yemen in 2020.

BACKGROUND: The need for early identification of coronavirus disease (COVID-19) cases in communities was high in Yemen during the first wave of the COVID-19 epidemic because most cases presenting to health facilities were severe. Early detection of cases would allow early interventions to interrupt the transmission chains. This study aimed to describe the implementation of community-based surveillance (CBS) in in internally displaced people (IDP) camps and urban settings in Yemen from 15 April 2020 to 30 September 2020. METHODS: Following the Centers for Disease Control and Prevention guidance for evaluation of surveillance systems, we assessed the usefulness and acceptability of CBS. For acceptability, we calculated the proportion of trained volunteers who reported disease alerts. To assess the usefulness, we compared the alerts reported through the electronic diseases early warning system (eDEWS) with the alerts reported through CBS and described the response activities implemented. RESULTS: In Al-Mukalla City, 18% (14/78) of the volunteers reported at least one alert. In IDP camps, 58% (18/31) of volunteers reported at least one alert. In Al-Mukalla City, CBS detected 49 alerts of influenza-like illness, whereas health facilities detected 561 cases of COVID-19. In IDP camps, CBS detected 91 alerts of influenza-like illness, compared to 10 alerts detected through eDEWS. In IDP camps, CBS detected three other syndromes besides influenza-like illness (febrile illness outbreak suspicion, acute diarrhoea, and skin disease). In IDP camps, public health actions were implemented for each disease detected and no further cases were reported. CONCLUSIONS: In Yemen, CBS was useful for detecting suspected outbreaks in IDP camps. CBS implementation did not yield expected results in general communities in urban areas in the early stage of the COVID-19 pandemic when little was known about the disease. In the urban setting, the system failed to detect suspected COVID-19 cases and other diseases despite the ongoing outbreaks reported through eDEWS. In Yemen, as in other countries, feasibility and acceptability studies should be conducted few months before CBS expansion in urban communities. The project should be expanded in IDP camps, by creating COVID-19 and other disease outbreak reporting sites.

Identification of novel drug resistance mechanisms by genomic and transcriptomic profiling of glioblastoma cells with mutation-activated EGFR.

AIMS: Epidermal growth factor receptor (EGFR) is not only involved in carcinogenesis, but also in chemoresistance. We characterized U87.MGΔEGFR glioblastoma cells with constitutively active EGFR due to deletion at the ligand binding domain in terms of gene expression profiling and chromosomal aberrations. Wild-type U87.MG cells served as control. MATERIALS AND METHODS: RNA sequencing and network analyses (Ingenuity Pathway Analysis) were performed to identify novel drug resistance mechanisms related to expression of mutation activated EGFR. Chromosomal aberrations were characterized by multicolor fluorescence in situ hybridization (mFISH) and array comparative genomic hybridization (aCGH). KEY FINDINGS: U87.MGΔEGFR cells presented much more chromosomal aberrations, amplifications and deletions than wild-type U87.MG cells. Both cell lines were near-triploid. Numerous genes were overexpressed in U87.MGΔEGFR cells, some of which have been already linked to drug resistance. PXDN, which is associated with epithelial mesenchymal transition, was the most upregulated gene (901.8-fold). TENM1 was 331.6-fold upregulated, and it was previously reported to modulate neural development. EGFR-AS1 (161.2-fold upregulated) has been reported to increase the EGFR mRNA stability and its expression - in accordance with that of EGFR - was upregulated (85.5-fold). In addition to well-known resistance genes, numerous novel genes and genomic aberrations were identified. ANGPT2 upregulation and CPM downregulation were validated by Western blotting. SIGNIFICANCE: Transcriptomics and genomics analyses in U87.MGΔEGFR cells unraveled a range of novel drug resistance mechanisms including apoptosis, DNA repair, ferroptosis, glutathione related gene activities, heat shock, oxidative stress, transcription factor activities, which may have important implications for future treatment strategies.

Identification of potential novel drug resistance mechanisms by genomic and transcriptomic profiling of colon cancer cells with p53 deletion.

TP53 (p53) is a pivotal player in tumor suppression with fifty percent of all invasive tumors displaying mutations in the TP53 gene. In the present study, we characterized colon cancer cells (HCT116 p53 -/-) with TP53 deletion, a sub-line derived from HCT116-p53 +/+ cells. RNA sequencing and network analyses were performed to identify novel drug resistance mechanisms. Chromosomal aberrations were identified by multicolor fluorescence in situ hybridization (mFISH) and array comparative genomic hybridization (aCGH). Numerous genes were overexpressed in HCT116 p53 -/- cells: RND3/RhoE (235.6-fold up-regulated), DCLK1 (60.2-fold up-regulated), LBH (31.9-fold up-regulated), MYB (28.9-fold up-regulated), TACSTD2 (110.1-fold down-regulated), NRIP1 (81.5-fold down-regulated) and HLA-DMB (69.7-fold down-regulated) are among the identified genes with potential influence on multidrug resistance (MDR) and they are associated with cancer progression and tumorigenesis, according to previously published studies. Probably due to TP53 deletion, disturbances in DNA repair and apoptosis are leading to aberrancies in cellular and organismal organization, ultimately increasing tumorigenesis and cancer progression potential. With NFκB, PI3K and HSP70, being at the center of merged protein network, and TH1-2 pathways, being among the influenced pathways, it can be speculated that the inflammatory pathway contributes to a resistance phenotype together with cell cycle regulation and heat-shock response. HCT116-p53 -/- cells have more chromosomal aberrations, gains and losses in copy numbers than HCT116-p53 +/+ cells. In conclusion, numerous genomic aberrations, which might be associated with yet unknown drug resistance mechanisms, were identified. This may have important implications for future treatment strategies.

Identification of metastasis-related genes by genomic and transcriptomic studies in murine melanoma.

AIMS: We systematically characterized metastatic murine B16-F10 melanoma, a sub-line derived from murine melanoma B16-F1 cells. MATERIALS AND METHODS: RNA-sequencing and network analyses (Ingenuity Pathway Analysis) were performed to identify novel potential metastasis mechanisms. Chromosomal aberrations were identified by multicolor fluorescence in situ hybridization (mFISH) using all 21 murine whole chromosome painting probes. KEY FINDINGS: Numerous genes were overexpressed in B16-F10 cells, some of which have been already described as being metastasis-linked. Nr5a1/sf1, a known prognostic marker for adrenal tumors, was 177-fold upregulated in B16-F10 cells compared to B16-F1 cells. Hoxb8 was 75-fold upregulated, which was previously associated with gastric cancer progression and metastasis. Ptk7, which is linked with tumorigenesis and metastasis of esophageal squamous carcinoma, was 67-fold upregulated. B16-F10 cells acquired additional chromosomal aberrations compared to B16-F1 cells, including dic(4)(pter->qter:qter->pter), +dic(6;15), +der(10)t(10;?1;16). SIGNIFICANCE: In addition to well-known metastatic genes, numerous novel genes and genomic aberrations were identified, which may serve as targets for treatment in the future. Transcriptomic and genetic analyses in B16-F10 cells unraveled a range of novel metastasis mechanisms, which may also have important implications for future treatment strategies.

2α-Hydroxyalantolactone from Pulicaria undulata: activity against multidrug-resistant tumor cells and modes of action.

BACKGROUND: Sesquiterpene lactones having α-methylene-γ-lactone moiety are promising natural metabolites showing various biological activity. One of the major metabolites isolated from Pulicaria undulata, 2α-hydroxyalantolactone (PU-1), has not been investigated in detail yet. Multidrug resistance (MDR) represents a major obstacle for cancer chemotherapy and the capability of novel natural products to overcoming MDR is of great interest. PURPOSE: Exploring the molecular modes of action for potent natural product metabolites. METHODS: The resazurin reduction assay was employed to evaluate the cytotoxicity of PU-1 on sensitive and their corresponding drug-resistant cell lines (overexpressing P-glycoprotein, BCRP, ABCB5, ΔEGFR, or TP53 knockout). Gene expression profiling was performed by transcriptome-wide mRNA microarray in the human CCRF-CEM leukemic cells after treatment with PU-1. The top significantly up- or down-regulated genes were identified by Chipster program and analyzed using Ingenuity Pathway Analysis (IPA) software. Finally, flow cytometry and Western blotting were performed for cell cycle analyses and apoptosis detection. RESULTS: The sesquiterpene lactone, PU-1, showed potent cytotoxicity towards the drug-sensitive and -resistant cell lines. Transcriptome-wide mRNA expression profiling and pathway analysis pointed to genes involved in DNA damage response and G2/M cell cycle arrest. G2/M arrest was verified by flow cytometry and further confirmed by the upregulation of p21 and downregulation of p-CDC25C expression in Western blotting. Moreover, the suggested DNA damage checkpoint regulation was confirmed by immunofluorescence and Western blotting by upregulation of pS345 Chk1, p-H3 and γ-H2AX. Furthermore, PU-1 inhibited PI3K/AKT pathway, which is involved in signaling DNA damage and G2/M arrest. Cells ultimately induced apoptosis upon PU-1 treatment. CONCLUSIONS: PU-1 is a potent natural product inhibiting otherwise drug-resistant human tumor cell growth through DNA damage, G2/M cell cycle arrest and apoptosis.

Cytotoxicity of apigenin toward multiple myeloma cell lines and suppression of iNOS and COX-2 expression in STAT1-transfected HEK293 cells.

BACKGROUND: Apigenin is one of the most abundant dietary flavonoids that possesses multiple bio-functions. PURPOSE: This study was designed to determine the influence of apigenin on gene expressions, cancer cells, as well as STAT1/COX-2/iNOS pathway mediated inflammation and tumorigenesis in HEK293-STAT1 cells. Furthermore, the cytotoxic activity toward multiple myeloma (MM) cell lines was investigated. METHODS: Bioinformatic analyses were used to predict the sensitivity and resistance of tumor cells toward apigenin and to determine cellular pathways influenced by this compound. The cytotoxic and ferroptotic activity of apigenin was examined by the resazurin reduction assay. Additionally, we evaluated apoptosis, and cell cycle distribution, induction of reactive oxygen species (ROS) and loss of integrity of mitochondrial membrane (MMP) by using the flow cytometry analysis. DAPI staining was used to detect characteristic apoptotic features. Furthermore, we verified its anti-inflammatory and additional mechanism of cell death by western blotting. RESULTS: COMPARE and hierarchical cluster analyses exhibited that 29 of 55 tumor cell lines were sensitive against apigenin (p < 0.001). The Ingenuity Pathway Analysis data showed that important bio-functions affected by apigenin were: gene expression, cancer, hematological system development and function, inflammatory response, and cell cycle. The STAT1 transcription factor was chosen as target protein on the basis of gene promoter binding motif analyses. Apigenin blocked cell proliferation of wild-type HEK293 and STAT1 reporter cells (HEK293-STAT1), promoted STAT1 suppression and subsequent COX-2 and iNOS inhibition. Apigenin also exhibited synergistic activity in combination with doxorubicin toward HEK293-STAT1 cells. Apigenin exerted excellent growth-inhibitory activity against MM cells in a concentration-dependent manner with the greatest activity toward NCI-H929 (IC50 value: 10.73 ± 3.21 µM). Apigenin induced apoptosis, cell cycle arrest, ferroptosis and autophagy in NCI-H929 cells. CONCLUSION: Apigenin may be a suitable candidate for MM treatment. The inhibition of the STAT1/COX-2/iNOS signaling pathway by apigenin is an important mechanism not only in the suppression of inflammation but also in induction of apoptosis.

The first 2 months of the SARS-CoV-2 epidemic in Yemen: Analysis of the surveillance data.

INTRODUCTION: Yemen was one of the last countries in the world to declare the first case of the pandemic, on 10 April 2020. Fear and concerns of catastrophic outcomes of the epidemic in Yemen were immediately raised, as the country is facing a complex humanitarian crisis. The purpose of this report is to describe the epidemiological situation in Yemen during the first 2 months of the SARS-CoV-2 epidemic. METHODS: We analyzed the epidemiological data from 18 February to 05 June 2020, including the 2 months before the confirmation of the first case. We included in our analysis the data from 10 out of 23 governorates of Yemen, located in southern and eastern part of the country. RESULTS: A total of 469 laboratory confirmed, 552 probable and 55 suspected cases with onset of symptoms between 18 February and 5 June 2020 were reported through the surveillance system. The median age among confirmed cases was 46 years (range: 1-90 years), and 75% of the confirmed cases were male. A total of 111 deaths were reported among those with confirmed infection. The mean age among those who died was 53 years (range: 14-88 years), with 63% of deaths (n = 70) occurring in individuals under the age 60 years. A total of 268 individuals with confirmed SARS-CoV-2 infection were hospitalized (57%), among whom there were 95 in-hospital deaths. CONCLUSIONS: The surveillance strategy implemented in the first 2 months of the SARS CoV 2 in the southern and eastern governorates of Yemen, captured mainly severe cases. The mild and moderate cases were not self-reported to the health facilities and surveillance system due to limited resources, stigma, and other barriers. The mortality appeared to be higher in individuals aged under 60 years, and most fatalities occurred in individuals who were in critical condition when they reached the health facilities. It is unclear whether the presence of other acute comorbidities contributed to the high death rate among SARS-CoV-2 cases. The findings only include the southern and eastern part of the country, which is home to 31% of the total population of Yemen, as the data from the northern part of the country was inaccessible for analysis. This makes our results not generalizable to the rest of the country.

Putative molecular determinants mediating sensitivity or resistance towards carnosic acid tumor cell responses.

BACKGROUND: Carnosic acid (CA) is one of the main constituents in rosemary extract. It possesses valuable pharmacological properties, including anti-oxidant, anti-inflammatory, anti-microbial and anti-cancer activities. Numerous in vitro and in vivo studies investigated the anticancer profile of CA and emphasized its potentiality for cancer treatment. Nevertheless, the role of multidrug-resistance (MDR) related mechanisms for CA's anticancer effect is not yet known. PURPOSE: We investigated the cytotoxicity of CA against known mechanisms of anticancer drug resistance (P-gp, ABCB5, BCRP, EGFR and p53) and determined novel putative molecular factors associated with cellular response towards CA. STUDY DESIGN: Cytotoxicity assays, bioinformatic analysis, flow cytometry and western blotting were performed to identify the mode of action of CA towards cancer cells. METHODS: The cytotoxicity to CA was assessed using the resazurin assays in cell lines expressing the mentioned resistance mechanisms. A pharmacogenomic characterization of the NCI 60 cell line panel was applied via COMPARE, hierarchical cluster and network analyses. Flow cytometry was used to detect cellular mode of death and ROS generation. Changes in proteins-related to apoptosis were determined by Western blotting. RESULTS: Cell lines expressing ABC transporters (P-gp, BCRP or ABCB5), mutant EGFR or p53 were not cross-resistant to CA compared to their parental counterparts. By pharmacogenomic approaches, we identified genes that belong to different functional groups (e.g. signal transduction, regulation of cytoskeleton and developmental regulatory system). These genes were predicted as molecular determinants that mediate CA tumor cellular responses. The top affected biofunctions included cellular development, cellular proliferation and cellular death and survival. The effect of CA-mediated apoptosis in leukemia cells, which were recognized as the most sensitive tumor type, was confirmed via flow cytometry and western blot analysis. CONCLUSION: CA may provide a novel treatment option to target refractory tumors and to effectively cooperate with established chemotherapy. Using pharmacogenomic approaches and network pharmacology, the relationship between cancer complexity and multi-target potentials of CA was analyzed and many putative molecular determinants were identified. They could serve as novel targets for CA and further studies are needed to translate the possible implications to clinical cancer treatment.

Lessons learned from detecting and responding to recurrent measles outbreak in Liberia post Ebola-Epidemic 2016-2017.

INTRODUCTION: Measles is an acute viral disease that remains endemic in much of sub-Sahara Africa, including Liberia. The 2014 Ebola epidemic disrupted an already fragile health system contributing to low uptake of immunization services, population immunity remained low thus facilitating recurrent outbreaks of measles in Liberia. We describe lessons learnt from detecting and responding to recurrent outbreaks of measles two years post the 2014 Ebola epidemic in Liberia. METHODS: We conducted a descriptive study using the findings from Integrated Diseases Surveillance and Response (IDSR) 15 counties, National Public Health Institute of Liberia (NPHIL), National Public Health Reference Laboratory (NPHRL) and District Health Information Software (DIHS2) data conducted from October to December, 2017. We perused the outbreaks line lists and other key documents submitted by the counties to the national level from January 2016 to December 2017. RESULTS: From January 2016 to December 2017, 2,954 suspected cases of measles were reported through IDSR. Four hundred sixty-seven (467) were laboratory confirmed (IgM-positive), 776 epidemiologically linked, 574 clinically confirmed, and 1,137 discarded (IgM-negative). Nine deaths out of 1817 cases were reported, a case fatality rate of 0.5%; 49% were children below the age of 5 years. Twenty-two percent (405/1817) of the confirmed cases were vaccinated while the vaccination status of 55% (994/1817) was unknown. CONCLUSION: Revitalization of IDSR contributed to increased detection and reporting of suspected cases of measles thus facilitating early identification and response to outbreaks. Priority needs to be given to increasing the uptake of routine immunization services, introducing a second dose of measles vaccine in the routine immunization program and conducting a high-quality supplementary measles immunization campaign for age group 1 to 10 years to provide protection for a huge cohort of susceptible.

The implementation of integrated disease surveillance and response in Liberia after Ebola virus disease outbreak 2015-2017.

INTRODUCTION: Although Liberia adapted the integrated diseases surveillance and response (IDSR) in 2004 as a platform for implementation of International Health Regulation (IHR (2005)), IDSR was not actively implemented until 2015. Some innovations and best practices were observed during the implementation of IDSR in Liberia after Ebola virus disease outbreak. This paper describes the different approaches used for implementation of IDSR in Liberia from 2015 to 2017. METHODS: We conducted a cross-sectional study using the findings from IDSR supervisions conducted from September to November 2017 and perused the outbreaks linelists submitted by the counties to the national level from January to December 2017 and key documents available at the national level. RESULTS: In 2017, the country piloted the use of mobile phones application to store and send data from the health facilities to the national level. In addition, an electronic platform for acute flaccid paralysis (AFP) surveillance called Auto-Visual AFP Detection and Reporting (AVADAR) was piloted in Montserrado County during the first semester of 2017. The timeliness and completeness of reports submitted from the counties to national level were above the target of 80% stable despite the challenges like insufficient resources, including skilled staff. CONCLUSION: IDSR is being actively implemented in Liberia since 2015. Although the country is facing the same challenges as other countries during the early stages of implementation of IDSR, the several innovations were implemented in a short time. The surveillance system reveled to be resilient, despite the challenges.

Enhancing laboratory capacity during Ebola virus disease (EVD) heightened surveillance in Liberia: lessons learned and recommendations.

INTRODUCTION: Following a declaration by the World Health Organization that Liberia had successfully interrupted Ebola virus transmission on May 9th, 2015; the country entered a period of enhanced surveillance. The number of cases had significantly reduced prior to the declaration, leading to closure of eight out of eleven Ebola testing laboratories. Enhanced surveillance led to an abrupt increase in demand for laboratory services. We report interventions, achievements, lessons learned and recommendations drawn from enhancing laboratory capacity. METHODS: Using archived data, we reported before and after interventions that aimed at increasing laboratory capacity. Laboratory capacity was defined by number of laboratories with Ebola Virus Disease (EVD) testing capacity, number of competent staff, number of specimens tested, specimen backlog, daily and surge testing capacity, and turnaround time. Using Stata 14 (Stata Corporation, College Station, TX, USA), medians and trends were reported for all continuous variables. RESULTS: Between May and December 2015, interventions including recruitment and training of eight staff, establishment of one EVD laboratory facility, implementation of ten Ebola GeneXpert diagnostic platforms, and establishment of working shifts yielded an 8-fold increase in number of specimens tested, a reduction in specimens backlog to zero, and restoration of turn-around time to 24 hours. This enabled a more efficient surveillance system that facilitated timely detection and containment of two EVD clusters observed thereafter. CONCLUSION: Effective enhancement of laboratory services during high demand periods requires a combination of context-specific interventions. Building and ensuring sustainability of local capacity is an integral part of effective surveillance and disease outbreak response efforts.