Adolescents' Perspective Regarding a Community-Wide Mass Drug Administration Program for Soil-Transmitted Helminths in India.

This study was undertaken to understand the perspective of adolescents in endemic communities of India regarding soil-transmitted helminth (STH) infections and community-wide mass drug administration (cMDA). A multicountry community-based cluster-randomized trial, the Deworm3 trial, tested the feasibility of interrupting STH transmission with cMDA, where all individuals aged 1-99 are treated empirically with albendazole. Using a guideline based on the Consolidated Framework for Implementation Research, eight focus group discussions were conducted among 57 adolescents from the trial site in India and analyzed on ATLAS.ti 8.0 software using an a priori thematic codebook. Adolescents believed that adults could be a source of STH infection because they were not routinely dewormed like the children through the national deworming program. Perceived benefits of cMDA for all were better health and increased work efficiency. Perceived barriers to adults' participation in cMDA was their mistrust about the program, fear of side effects, perceived low risk of infection, and absence during drug distribution. To encourage adult participation in cMDAs, adolescents suggested community outreach activities, engaging village influencers and health workers, and tailoring drug distribution to when adults would be available. Adolescents were confident in their ability to be change agents within their households for treatment compliance. Adolescents provided insights into potential barriers and solutions to improve adult participation in cMDA, identified best practices of cMDA delivery, and suggested that they have unique roles as change agents to increase their household participation in cMDA.

Somatic hypermutation mechanisms during lymphomagenesis and transformation.

B cells undergoing physiologically programmed or aberrant genomic alterations provide an opportune system to study the causes and consequences of genome mutagenesis. Activated B cells in germinal centers express activation-induced cytidine deaminase (AID) to accomplish physiological somatic hypermutation (SHM) of their antibody-encoding genes. In attempting to diversify their immunoglobulin (Ig) heavy- and light-chain genes, several B-cell clones successfully optimize their antigen-binding affinities. However, SHM can sometimes occur at non-Ig loci, causing genetic alternations that lay the foundation for lymphomagenesis, particularly diffuse large B-cell lymphoma. Thus, SHM acts as a double-edged sword, bestowing superb humoral immunity at the potential risk of initiating disease. We refer to off-target, non-Ig AID mutations - that are often but not always associated with disease - as aberrant SHM (aSHM). A key challenge in understanding SHM and aSHM is determining how AID targets and mutates specific DNA sequences in the Ig loci to generate antibody diversity and non-Ig genes to initiate lymphomagenesis. Herein, we discuss some current advances regarding the regulation of AID's DNA mutagenesis activity in B cells.

"Our desire is to make this village intestinal worm free": Identifying determinants of high coverage of community-wide mass drug administration for soil transmitted helminths in Benin, India, and Malawi.

BACKGROUND: Soil-transmitted helminth infections (STH) are associated with substantial morbidity in low-and-middle-income countries, accounting for 2.7 million disability-adjusted life years annually. Current World Health Organization guidelines recommend controlling STH-associated morbidity through periodic deworming of at-risk populations, including children and women of reproductive age (15-49 years). However, there is increasing interest in community-wide mass drug administration (cMDA) which includes deworming adults who serve as infection reservoirs as a method to improve coverage and possibly to interrupt STH transmission. We investigated determinants of cMDA coverage by comparing high-coverage clusters (HCCs) and low-coverage clusters (LCCs) receiving STH cMDA in three countries. METHODS: A convergent mixed-methods design was used to analyze data from HCCs and LCCs in DeWorm3 trial sites in Benin, India, and Malawi following three rounds of cMDA. Qualitative data were collected via 48 community-level focus group discussions. Quantitative data were collected via routine activities nested within the DeWorm3 trial, including annual censuses and coverage surveys. The Consolidated Framework for Implementation Research (CFIR) guided coding, theme development and a rating process to determine the influence of each CFIR construct on cMDA coverage. RESULTS: Of 23 CFIR constructs evaluated, we identified 11 constructs that differentiated between HCCs and LCCs, indicating they are potential drivers of coverage. Determinants differentiating HCC and LCC include participant experiences with previous community-wide programs, communities' perceptions of directly observed therapy (DOT), perceptions about the treatment uptake behaviors of neighbors, and women's agency to make household-level treatment decisions. CONCLUSION: The convergent mixed-methods study identified barriers and facilitators that may be useful to NTD programs to improve cMDA implementation for STH, increase treatment coverage, and contribute to the successful control or elimination of STH. TRIAL REGISTRATION: The parent trial was registered at clinicaltrials.gov (NCT03014167).

Ethanolysis of nitrogen mustards: A novel strategy for nitrogen mustard identification in environmental matrices by liquid chromatography-electrospray ionization-tandem mass spectrometry.

RATIONALE: Nitrogen mustards (NMs) are blistering chemical warfare agents. The ability to detect NMs in environmental samples is very important for obtaining forensic evidence. The most common analytical techniques for NM detection are gas chromatography-mass spectrometry, which detects NMs in their intact form but is disadvantaged by high limits of detection (LODs), and liquid chromatography-electrospray ionization-tandem mass spectrometry (LC/ESI-MS/MS) of their hydrolysis products, which do not provide robust evidence to support NM use. METHODS: We developed a novel approach to detect and identify NMs using LC/ESI-MS/MS after chemical derivatization. The method is based on ethoxide-promoted ethanolysis prior to analysis. The effects of reaction time, temperature, ethoxide concentration and chromatography behavior were studied and optimized. In the developed procedure, 0.1% (v/v) sodium ethoxide solution is added to the NMs in ethanol and agitated for 2 h at 50°C, followed by LC/ESI-MS/MS, without any other pretreatment. RESULTS: The ethanolysis reaction efficiencies were evaluated in ethanolic extracts from soil, asphalt, and ethanol contaminated with 0.5% (v/v) diesel fortified with NMs at a five-point calibration curve. The calibration curves showed good linearity in the range of 0.05-1 ng/mL, with an R2 value of 0.99, and were similar to those of LC/MS-grade ethanol, with almost no observable matrix effects. The derivatization products were stable at room temperature, with LODs of 10 pg/mL, in all investigated extracts. CONCLUSIONS: Through this newly developed strategy, the derivatization of active NMs by ethanolysis was achieved for the first time, and these derivatization products can serve as specific indicators for the use and presence of NMs. The methodology can also verify trace levels of NM chemical warfare agents collected in war or terror scenarios in forensic investigations.

Diagnosis and treatment of diabetic foot ulcer complicated with lower extremity vasculopathy: Consensus recommendation from the Chinese Medical Association (CMA), Chinese Medical Doctor Association (CMDA).

Diabetic foot ulcer complicated with lower extremity vasculopathy is highly prevalent, slow healing and have a poor prognosis. The final progression leads to amputation, or may even be life-threatening, seriously affecting patients' quality of life. The treatment of lower extremity vasculopathy is the focus of clinical practice and is vital to improving the healing process of diabetic foot ulcers. Recently, a number of clinical trials on diabetic foot ulcers with lower extremity vasculopathy have been reported. A joint group of Chinese Medical Association (CMA) and Chinese Medical Doctor Association (CMDA) expert representatives reviewed and reached a consensus on the guidelines for the clinical diagnosis and treatment of this kind of disease. These guidelines are based on evidence from the literature and cover the pathogenesis of diabetic foot ulcers complicated with lower extremity vasculopathy and the application of new treatment approaches. These guidelines have been put forward to guide practitioners on the best approaches for screening, diagnosing and treating diabetic foot ulcers with lower extremity vasculopathy, with the aim of providing optimal, evidence-based management for medical personnel working with diabetic foot wound repair and treatment.

Identifying opportunities to optimize mass drug administration for soil-transmitted helminths: A visualization and descriptive analysis using process mapping.

BACKGROUND: The control of soil-transmitted helminths (STH) is achieved through mass drug administration (MDA) with deworming medications targeting children and other high-risk groups. Recent evidence suggests that it may be possible to interrupt STH transmission by deworming individuals of all ages via community-wide MDA (cMDA). However, a change in delivery platforms will require altering implementation processes. METHODS: We used process mapping, an operational research methodology, to describe the activities required for effective implementation of school-based and cMDA in 18 heterogenous areas and over three years in Benin, India, and Malawi. Planned activities were identified during workshops prior to initiation of a large cMDA trial (the DeWorm3 trial). The process maps were updated annually post-implementation, including adding or removing activities (e.g., adaptations) and determining whether activities occurred according to plan. Descriptive analyses were performed to quantify differences and similarities at baseline and over three implementation years. Comparative analyses were also conducted between study sites and areas implementing school-based vs. cMDA. Digitized process maps were developed to provide a visualization of MDA processes and inspected to identify implementation bottlenecks and inefficient activity flows. RESULTS: Across three years and all clusters, implementation of cMDA required an average of 13 additional distinct activities and was adapted more often (5.2 adaptations per year) than school-based MDA. An average of 41% of activities across both MDA platforms did not occur according to planned timelines; however, deviations were often purposeful to improve implementation efficiency or effectiveness. Visualized process maps demonstrated that receipt of drugs at the local level may be an implementation bottleneck. Many activities rely on the effective setting of MDA dates and estimating quantity of drugs, suggesting that the timing of these activities is important to meet planned programmatic outcomes. CONCLUSION: Implementation processes were heterogenous across settings, suggesting that MDA is highly context and resource dependent and that there are many viable ways to implement MDA. Process mapping could be deployed to support a transition from a school-based control program to community-wide STH transmission interruption program and potentially to enable integration with other community-based campaigns. TRIAL REGISTRATION: NCT03014167.

H2 O2 -Inducible DNA Cross-linking Agents Capable of Releasing Multiple DNA Alkylators as Anticancer Prodrugs.

Three compounds with arylboronate esters conjugated with two equivalent nitrogen mustards [bis(2-chloroethyl)methylamine, HN2] have been synthesized and characterized. These inactive small molecules selectively react with H2 O2 to produce multiple DNA cross-linkers, such as two HN2 molecules alongside a bisquinone methide (bisQM), leading to efficient DNA ICL formation. In comparison to other amine functional groups, using HN2 as a leaving group greatly improves the DNA cross-linking efficiency of these arylboronate esters as well as cellular activity. The introduction of HN2 in these arylboronate ester analogues favored the generation of bisQM that can directly cross-link DNA. Two equivalents of HN2 are also generated from these compounds upon treatment with H2 O2 , which directly produces DNA ICL products. The cumulative effects of HN2 and bisQM on DNA cross-linking makes these molecules highly effective H2 O2 -inducible DNA ICL agents. The three compounds with HN2 as a leaving group showed greatly enhanced cytotoxicity towards cancer cells in comparison to those containing trimethyl amine as a leaving group. This provides an effective strategy for further design of novel potential ROS-activated anticancer prodrugs.

Design, synthesis and anticancer evaluation of novel oncolytic peptide-chlorambucil conjugates.

Nitrogen mustards (NMs) are an important class of chemotherapeutic drugs and have been widely employed for the treatment of various cancers. However, due to the high reactivity of nitrogen mustard, most NMs react with proteins and phospholipids within the cell membrane. Therefore, only a very small fraction of NMs can reach the reach nucleus, alkylating and cross-linking DNA. To efficiently penetrate the cell membrane barrier, the hybridization of NMs with a membranolytic agent may be an effective strategy. Herein, the chlorambucil (CLB, a kind of NM) hybrids were first designed by conjugation with membranolytic peptide LTX-315. However, although LTX-315 could help large amounts of CLB penetrate the cytomembrane and enter the cytoplasm, CLB still did not readily reach the nucleus. Our previous work demonstrated that the hybrid peptide NTP-385 obtained by covalent conjugation of rhodamine B with LTX-315 could accumulate in the nucleus. Hence, the NTP-385-CLB conjugate, named FXY-3, was then designed and systematically evaluated both in vitro and in vivo. FXY-3 displayed prominent localization in the cancer cell nucleus and induced severe DNA double-strand breaks (DSBs) to trigger cell apoptosis. Especially, compared with CLB and LTX-315, FXY-3 exhibited significantly increased in vitro cytotoxicity against a panel of cancer cell lines. Moreover, FXY-3 showed superior in vivo anticancer efficiency in the mouse cancer model. Collectively, this study established an effective strategy to increase the anticancer activity and the nuclear accumulation of NMs, which will provide a valuable reference for future nucleus-targeting modification of nitrogen mustards.

The dinitrobenzamide mustard prodrugs, PR-104A and SN27686, for use in a novel MNDEPT cancer prodrug therapy approach.

Directed enzyme prodrug therapy is a highly promising anti-cancer strategy. However, the current technology is limited by inefficient prodrug activation and the dose-limiting toxicity associated with the prodrugs being tested; to overcome these limitations, the dinitrobenzamide mustard prodrugs, PR-104A and SN27686, have been developed. The present study will assess both of these prodrugs for their potential uses in a novel magnetic-nanoparticle directed enzyme prodrug therapy strategy by determining their kinetic parameters, assessing the products formed during enzymatic reduction using HPLC and finally their ability to cause cell death in the ovarian cancer cell line, SK-OV-3. It was shown for the first time that the dinitrobenzamide mustard prodrugs are able to be reduced by the genetically modified nitroreductases, NfnB-cys and YfkO-cys, and that these enzyme/prodrug combinations can induce a significant cell death in the SK-OV-3 cell line, highlighting the potential for both enzyme/prodrug combinations for use in magnetic-nanoparticle directed enzyme prodrug therapy.

Glutathione conjugation of nitrogen mustards: In vitro study.

RATIONALE: This paper describes an in vitro study designed to identify metabolic biomarkers resulting from the conjugation of nitrogen mustards (NMs) with glutathione (GSH). The method developed is essential in providing evidence in the event of NM exposure in biomedical samples. METHODS: The mass spectral characterization of the proposed NMs-GSH conjugates was performed with liquid chromatography high-resolution tandem mass spectrometry (LC-HRMS/MS). The final reaction mixtures were analysed in positive electrospray ionisation (ESI) at different incubation times. RESULTS: This study identified three types of conjugates in addition to ethanolamines, the hydrolysis products of NMs. Monoglutathionyl, diglutathionyl and phosphorylated conjugates were produced for each of the NMs, bis(2-chloroethyl)ethylamine (HN1), bis(2-chloroethyl)methylamine (HN2) and tris(2-chloroethyl)amine (HN3). The monoglutathionyl conjugates consisted of HN1-GSH, HN2-GSH and HN3-GSH. The spontaneous and primary conjugates of diglutathionyl were HN1-GSH2, HN2-GSH2 and HN3-GSH2. These included phosphorylated conjugates, namely HN1-GSH-PO4 , HN2-GSH-PO4 and HN3-GSH-PO4 , as might have formed due to hydrolysis in phosphate buffer. CONCLUSIONS: The mass spectral data of all conjugates formed in the presence of all NMs and GSH are reported in this study. These GSH metabolites can be used to confirm NMs toxicity in biological samples such as urine.

Activation of the human TRPA1 channel by different alkylating sulfur and nitrogen mustards and structurally related chemotherapeutic drugs.

An important target in toxicology is the ion channel known as human transient receptor potential ankyrin 1 (hTRPA1). It is triggered by a variety of chemicals, including the alkylating chemical warfare agent sulfur mustard (SM). The activation potentials of structural analogs including O- and sesquimustard, nitrogen mustards (HN1, HN2, and HN3), and related chemotherapeutic drugs (bendamustine, cycylophosphamide, and ifosfamide) were examined in the current study. The aequorin assay was used to measure changes in intracellular calcium levels in human hTRPA1 overexpressing HEK293 cells. The XTT assay was used to determine cytotoxicity. The data presented here highlight that all investigated alkylating substances, with the exception of cyclophosphamide and ifosfamide, cause the activation of hTRPA1. Cytotoxicity and activation of hTRPA1 were found to be related. Compounds with high reactivity had higher cytotoxicity and vice versa. However, inhibiting hTRPA1 with the specific inhibitor AP18 could not reduce the cytotoxicity induced by alkylating agents. As a result, hTRPA1 does not play a significant role in the cytotoxicity of alkylating agents.

The Nitrogen Mustards.

The nitrogen mustards are powerful cytotoxic and lymphoablative agents and have been used for more than 60 years. They are employed in the treatment of cancers, sarcomas, and hematologic malignancies. Cyclophosphamide, the most versatile of the nitrogen mustards, also has a place in stem cell transplantation and the therapy of autoimmune diseases. Adverse effects caused by the nitrogen mustards on the central nervous system, kidney, heart, bladder, and gonads remain important issues. Advances in analytical techniques have facilitated the investigation of the pharmacokinetics of the nitrogen mustards, especially the oxazaphosphorines, which are prodrugs requiring metabolic activation. Enzymes involved in the metabolism of cyclophosphamide and ifosfamide are very polymorphic, but a greater understanding of the pharmacogenomic influences on their activity has not yet translated into a personalized medicine approach. In addition to damaging DNA, the nitrogen mustards can act through other mechanisms, such as antiangiogenesis and immunomodulation. The immunomodulatory properties of cyclophosphamide are an area of current exploration. In particular, cyclophosphamide decreases the number and activity of regulatory T cells, and the interaction between cyclophosphamide and the intestinal microbiome is now recognized as an important factor. New derivatives of the nitrogen mustards continue to be assessed. Oxazaphosphorine analogs have been synthesized in attempts to both improve efficacy and reduce toxicity, with varying degrees of success. Combinations of the nitrogen mustards with monoclonal antibodies and small-molecule targeted agents are being evaluated. SIGNIFICANCE STATEMENT: The nitrogen mustards are important, well-established therapeutic agents that are used to treat a variety of diseases. Their role is continuing to evolve.

Bendamustine: A review of pharmacology, clinical use and immunological effects (Review).

Bendamustine is an alkylating agent classified into the group of nitrogen mustard analogues, synthesized almost sixty years ago. It was registered in former East Germany in 1971 and approved by the US Food and Drug Administration in 2008 for treatment of chronic lymphocytic leukemia and indolent B­cell non­Hodgkin lymphoma. Considering its beneficial properties in the therapy of relapsed or refractory hematological malignancies, synergistic effects with other antineoplastic agents and increasing recent reports on its immunomodulatory effects, bendamustine has once again gained its justified attention. The uniqueness of bendamustine­mediated effects should be observed keeping in mind its distinctive structure with structural similarities to both alkylating agents and purine analogs. In the present review, the current knowledge on the use of bendamustine in oncology, its pharmacokinetics, mechanism of action and toxicity was summarized. In addition, its immune­modulating effects that have not been fully elucidated so far are emphasized, hoping to encourage further investigations of this unique drug.

Robust inactivation of Plasmodium falciparum in red blood cell concentrates using amustaline and glutathione pathogen reduction.

BACKGROUND: Plasmodium falciparum is the parasite responsible for most malaria cases globally. The risk of transfusion-transmitted malaria (TTM) is mitigated by donor deferrals and blood screening strategies, which adversely impact blood availability. Previous studies showed robust inactivation of P. falciparum using nucleic acid-targeting pathogen reduction technologies (PRT) for the treatment of plasma and platelet components or whole blood (WB). The efficacy of the amustaline-glutathione (GSH) PRT to inactivate P. falciparum is here evaluated in red blood cells (RBC), as well the impact of PRT on parasite loads, stages, and strains. STUDY DESIGN AND METHODS: RBC units resuspended in AS-1 or AS-5 additive solutions were spiked with ring stage-infected RBC and treated with the amustaline-GSH PRT. Parasite loads and viability were measured in samples at the time of contamination, and after treatment, using serial 10-fold dilutions of the samples in RBC cultures maintained for up to 4 weeks. RESULTS: P. falciparum viability assays allow for the detection of very low levels of parasite. Initial parasite titer was >5.2 log10 /ml in AS-1/5 RBC. No infectious parasites were detected in amustaline-GSH-treated samples after 4 weeks of culture. Amustaline-GSH inactivated high parasite loads regardless of parasite stages and strains. Amustaline readily penetrates the parasite, irreversibly blocks development, and leads to parasite death and expulsion from RBC. DISCUSSION: Amustaline-GSH PRT demonstrated robust efficacy to inactivate malaria parasites in RBC concentrates. This study completes the portfolio of studies demonstrating the efficacy of nucleic acid-targeting PRTs to mitigate TTM risks as previously reported for platelet concentrates, plasma, and WB.

Serum immunoglobulin G as a discriminator of infection in follicular lymphoma patients undergoing chemotherapy with bendamustine in combination with rituximab.

OBJECTIVES: Chemotherapy, including bendamustine, usually causes lymphocytopaenia and hypogammaglobulinaemia as side effects in patients with haematological malignancies. Therefore, the possibility has been considered that these immunological adverse events induced by bendamustine may lead to infectious diseases. However, lymphocytopaenia and/or hypogammaglobulinaemia have not yet been shown to have a statistically significant association with infection in cancer patients who receive bendamustine. METHODS: We retrospectively studied 27 patients with relapsed or refractory indolent follicular lymphoma who were treated with bendamustine and rituximab (BR). In order to elucidate relationships between immune-related laboratory parameters (i.e. peripheral blood leukocyte, neutrophil, lymphocyte and immunoglobulin G [IgG]) and infectious events, receiver operating characteristic (ROC) curve and multivariate logistic regression analyses were performed. RESULTS: Infectious diseases occurred in 11 patients (11/27, 41%), including 3 (3/27, 11%) with severe diseases. The area under the ROC curve (AUC) showed that the lowest IgG level during and after BR discriminated infectious events (cut-off value, 603 mg/dL) with 81.8% sensitivity and 68.8% specificity (AUC, 0.76; 95% CI, 0.52-0.90). Furthermore, a multivariate regression analysis revealed that the minimal serum IgG value during and after BR therapy was the only variable that was significantly associated with infection (odds ratio, 8.29; 95% CI, 1.19-57.62; p value, 0.03). CONCLUSION: Serum IgG ≤603 mg/dL during and after BR therapy was independently associated with an increased risk of infection. The monitoring of serum IgG during chemotherapy may help to predict the development of infection in blood cancer patients undergoing chemotherapy with bendamustine in combination with rituximab.

In Vitro Evaluation of DNA Damage Effect Markers toward Five Nitrogen Mustards Based on Liquid Chromatography-Tandem Mass Spectrometry.

Endogenous DNA lesions frequently occur due to internal effects such as oxidative stress, inflammation, endogenous alkylation, and epigenetic modifications. However, exposure to chemical toxicants from the environment, diet, or drugs can also induce significant endogenous DNA damage. The quantification of endogenous DNA damage effect markers might reflect the actual DNA damage level of chemical toxicants. Herein, we report a liquid chromatography-triple quadrupole tandem mass spectrometry (LC-QqQ MS/MS) method for simultaneous determination of eight representative endogenous DNA damage biomarkers, including five endogenous DNA damage effect markers (oxidative damage, 8-oxo-dG; lipid peroxidation, εdA and N2-Et-dG; inflammation, 5-Cl-dC; and endogenous alkylation, O6-Me-dG), and three epigenetic modifications (5-m-dC, 5-hm-dC, and N6-Me-dA). The method validation was performed, and the linear range was 0.05 pg to 2 ng (on-column), the limit of detection was 0.02 pg (on-column), and the precision, accuracy, matrix effect, and recovery were all between 85 and 115%. We then applied this method to evaluate endogenous DNA damage to human embryonic lung fibroblast cells exposed to five nitrogen mustards [NMs, i.e., HN1, HN2, HN3, chlorambucil (CB), and cyclophosphamide (CTX)], where curcumin exposure was used as a control due to its inability to induce the formation of endogenous DNA adducts. The amounts of eight DNA adducts in the low-, middle-, and high-concentration exposure groups of five NMs were almost all significantly different from those in the blank group (P < 0.05). We obtained a positive correlation between the contents of eight DNA damage biomarkers and the inhibition dose of five NMs, except for N2-Et-dG and 5-Cl-dC. Via further principal component analysis and partial least squares discriminant analysis, we clustered all NMs into three units with different cytotoxicity levels, that is, HN2 and HN1 (highly toxic), HN3 and CB (moderately toxic), and CTX (less toxic). Moreover, for the same concentration of HN1/2/3 exposure groups, as the cytotoxicity increased according to the order of HN3 < HN1 < HN2, the contents of 8-oxo-dG, 5-m-dC, 5-hm-dC, and N6-Me-dA increased, whereas the content of O6-Me-dG decreased. Therefore, the contents of these DNA damage effect markers were somewhat related to the cytotoxicity and concentration of NMs. We hope that this method will provide an alternative evaluation approach for the toxicological effects of NMs and the safety of the medication.

Vitamin D3 protects against nitrogen mustard-induced apoptosis of the bronchial epithelial cells via activating the VDR/Nrf2/Sirt3 pathway.

Respiratory system injury is the main cause of mortality for nitrogen mustard (NM)-induced damage. Previous studies indicate that reactive oxygen species (ROS) participates in NM-mediated respiratory injuries, but the detailed mechanism is not quite clear. Human bronchial epithelial cell lines 16HBE and BEAS-2B were treated with HN2, a type of NM. In detail, it was shown that HN2 treatment induced impaired cell viability, excessive mitochondrial ROS production and enhanced cellular apoptosis in bronchial epithelial cells. Moreover, impaired Sirt3/SOD2 axis was observed upon HN2 treatment, with decreased Sirt3 and increased acetylated SOD2 expression levels. Sirt3 overexpression partially ameliorated HN2-induced cell injury. Meanwhile, vitamin D3 treatment partially attenuated HN2-induced apoptosis and improved the mitochondrial functions upon HN2 intervention. In addition, HN2 exposure decreased VDR expression, thus inhibiting the Nrf2 phosphorylation and Sirt3 activation. Inhibition of Nrf2 or Sirt3 could decrease the protective effects of vitamin D3 and enhance mitochondrial ROS production via modulating mitochondrial redox balance. In conclusion, impaired VDR/Nrf2/Sirt3 axis contributed to NM-induced apoptosis, while vitamin D3 supplementation provides protective effects via the activation of VDR and the improvement of mitochondrial functions. This study provides novel mechanism and strategy for NM exposure-induced pulmonary injuries.

Bioreductive prodrug PR-104 improves the tumour distribution and titre of the nitroreductase-armed oncolytic adenovirus ONYX-411NTR leading to therapeutic benefit.

Advances in the field of cancer immunotherapy have stimulated renewed interest in adenoviruses as oncolytic agents. Clinical experience has shown that oncolytic adenoviruses are safe and well tolerated but possess modest single-agent activity. One approach to improve the potency of oncolytic viruses is to utilise their tumour selectivity to deliver genes encoding prodrug-activating enzymes. These enzymes can convert prodrugs into cytotoxic species within the tumour; however, these cytotoxins can interfere with viral replication and limit utility. In this work, we evaluated the activity of a nitroreductase (NTR)-armed oncolytic adenovirus ONYX-411NTR in combination with the clinically tested bioreductive prodrug PR-104. Both NTR-expressing cells in vitro and xenografts containing a minor population of NTR-expressing cells were highly sensitive to PR-104. Pharmacologically relevant prodrug exposures did not interfere with ONYX-411NTR replication in vitro. In vivo, prodrug administration increased virus titre and improved virus distribution within tumour xenografts. Colonisation of tumours with high ONYX-411NTR titre resulted in NTR expression and prodrug activation. The combination of ONYX-411NTR with PR-104 was efficacious against HCT116 xenografts, whilst neither prodrug nor virus were active as single agents. This work highlights the potential for future clinical development of NTR-armed oncolytic viruses in combination with bioreductive prodrugs.

Widely targeted metabolomics analysis of enriched secondary metabolites and determination of their corresponding antioxidant activities in Elaeagnus angustifolia var. orientalis (L.)Kuntze fruit juice enhanced by Bifidobacterium animalis subsp. Lactis HN-3 fermentation.

Elaeagnus angustifolia var. orientalis (L.)Kuntze fruit contains a large number of naturally occurring molecules present as glycoside, methylated, and methyl ester conjugates, which should be hydolysed or transformed to become bioactive forms. For this purpose, Bifidobacterium animalis subsp. lactis HN-3 was selected to ferment Elaeagnus angustifolia var. orientalis (L.)Kuntze fruit juice (EOJ). After fermentation, the total phenolic content (TPC) and antioxidant capacity of the EOJ increased significantly compared to the non-fermented EOJ. Using widely-targeted metabolomics analysis, polyphenolic compounds involved in the flavonoid biosynthetic pathway were determined to be up-regulated in the fermented EOJ. In addition, the metabolites generated by 8 deglycosidation, 5 demethylation, 5 hydrogenation, and 28 other reactions were detected in higher concentrations in the fermented EOJ compared to the non-fermented EOJ. Interestingly, these up-regulated metabolites have higher antioxidant and other biological activities than their metabolic precursors, which provide a theoretical basis for the development of Bifidobacterium-fermented plant products with stronger functional activities.