Journal articles on the topic '[SDV:BIO] Life Sciences/Biotechnology'

Revolutionary progress in the life sciences and in biotechnology presents us with completely new ethical questions concerning the acceptability of individual and generic eugenics, and of the production of ‘monsters’ of different kinds (including hybrids). Cultural developments related to what is called post or late modernity create a context in which biotechnology is freely used for the realization of all sorts of extravagant desires. Bioethics which is supposed to deal with new problems in this predicament has lost contact with concrete, lived morality and seems to have become the handmaiden of progress. It is argued here that a new kind of bioethics is required which is not divorced from ordinary, common sense morality and which is guided by fundamental philosophical reflections on morality and its place in culture.

THIS BOOK IS AN INSIGHTFUL and theoretically ambitious anthropological study of the genomics and biotech industries in the United States and India. These and related science-based sectors form part of the bio-economy, a larger complex of manufacturing, service, and research and development (R&D) activities, grounded increasingly in advancements in the biological sciences. In his groundbreaking study, Kaushik Sunder Rajan seeks to explain the intersection between biological knowledge ? the new knowledge of life itself ? and the economic accumulation process in which large pharmaceutical firms are dominant actors. The most striking manifestation of the bio-economy is the emergence of thousands of small biotechnology and other science-intensive start-up firms. These populate areas close to major universities in the developed parts of the world ? in California, Massachusetts, the Cambridge region in the United Kingdom, and elsewhere ? but have emerged also in some centres in developing countries. This is described in Sunder Rajan?s empirical analysis which investigates the global evolution of the bio-economy, with a particular focus on India and California. He explains the interdependencies between giant pharmaceutical companies and small dedicated biotechnology firms, which operate in conjunction with a myriad of intermediaries, such as venture capitalist firms that provide funding for promising science and facilitate interaction between different bio-economic actors.

The purpose of this paper is to present a holistic and practicable methodology to address new ethical issues emerging from biotechnology. With the advent of the age of life sciences, related technologies have extended life beyond the purpose of disease treatment without limitation and have even played a role in selecting excellent genes. With the acquisition of this technology by life scientists, mankind has come to encounter ethical problems that are completely different from the past. Ethicists point out that the logic of the age of life sciences has limitations in solving ethical problems. It is pointed out that human beings are in danger of discussing ethical issues that will emerge in the present and future due to bio-science technology. The human community had to call for a new ethical approach. This method is a predictable philosophical reflection on the environment and future of society and the human community beyond the ethical issues of science and technology. Therefore, this research method is not only to approach the ethical issues of biotechnology itself, but also to ensure the possibility of predicting a transparent future

The development of national and international biotechnology law in the context of new constitutional priorities, threats and challenges to life, public health and safety, is one of the current and yet under-researched topics in the scientific legal literature. Unfortunately, pro-gress in the life sciences is proceeding faster than the legal thinking that should accompany them. Breakthroughs in biology and medicine dictate the corresponding development of law, which today unfortunately lags chronically behind, resulting in gaps and contradictions between existing legal norms. As contemporary researchers have noted, and as is also evident in judicial practice, the issues of regulating the objects of bio-law - the body, life, procreation, self-identification, and ecology - mirror the contradictions of our society. The need to legally regulate the application of biotechnology has led to the formation of bioclaw as a new, integrated set of laws and, in the long term, as a branch of law of a new generation. As a response to the rapid introduction of advances in biotechnology into human life, we believe that bio-law must take into account the benefits and challenges associated with the impact of new technologies on the human body that may fundamentally alter the human condition as a physical individual. In this case, ethical issues, i. e. the requirements of bioethics, lead to a reflection on the content of bioethics in contemporary democracies and legal states. In particular, the introduction of technologies associated with gene editing, cloning, surrogacy, transplantation medicine, sex reassignment surgeries and other interventions into human nature, the consequences of which are not fully understood and threaten the physical and mental health of individuals, can cause irreparable harm to the spiritual, social, moral and ethical foundations of individuals, families, society, states and humanity as a whole. In our opinion, in the next few decades, bio-law will emerge as a global, integrated branch of law that simultaneously incorporates the norms of international law and national law. The main purpose of bio-law as a new branch of law is to counteract the global threat to humanity which consists in the emergence of a trend toward mass, large-scale changes to human nature and its unique natural qualities through the use of biotechnological advances. Therefore, the object of regulation and, accordingly, protection by this branch of law is not only the individual, his or her physical and spiritual health, but humanity as a whole, current and future generations. The uniqueness of bio-law as a new branch of law also lies in the fact that the individual, humanity, present and future generations are both the objects and subjects of law. The integrated nature of bioprinciples is manifested in the close connection of public and private interests with the obvious predominance of the public significance of their legal norms.

Biotechnology is a broad area of biology, involving the use of living systems and organisms to develop products. Depending on the tools and applications, it often overlaps with related scientific fields. In the late 20th and early 21st centuries, biotechnology has expanded to include new and diverse sciences, such as genomics, recombinant gene techniques, applied immunology, and development of pharmaceutical therapies and diagnostic tests. Biotechnology has also led to the development of antibiotics. Biotechnology has applications in four major industrial areas, including health care (medical), crop production and agriculture, non-food (industrial) uses of crops and other products and environmental uses. In medicine, modern biotechnology has many applications in areas such as pharmaceutical drug discoveries and production, pharmacogenomics, and genetic testing. Pharmaceutical biotechnology is a relatively new and growing field in which the principles of biotechnology are applied to the development of drugs. A majority of therapeutic drugs in the current market are bio formulations, such as antibodies, nucleic acid products and vaccines. Such bio formulations are developed through several stages that include: understanding the principles underlying health and disease the fundamental molecular mechanisms governing the function of related biomolecules synthesis and purification of the molecules determining the product shelf life, stability, toxicity and immunogenicity drug delivery systems patenting and clinical trials. This review article describes the purpose of biotechnology in pharmaceutical industry, particularly pharmaceutical biotechnology along with its prospects and challenges.

The paper aims to explain two basic standpoints regarding the enhancement of human beings through genetic engineering. Savulescu seems to be starting from a technoprogressive, (neo)liberal orientation, while Fukuyama's position implies a step back - to (bio)conservatism, returning to the natural human rights. These opposing general attitudes reflect the same aspiration - towards greater control and monitoring by the state for the benefit of individuals and (or) humankind. While Fukuyama justifies the narrow use of biotechnology for the purpose of therapy and prevention of disease, so far Savulescu goes a step further, understanding 'enhancement' to include increasing the length and quality of life, and focuses exclusively on genetic intervention for this purpose. Discussing the same problems Fukuyama lists several reasons why we should limit the use of biotechnology - reasons of religious, utilitarian and philosophical nature. In this paper, arguments for and against genetic intervention are discussed, while cited examples are commented ad hoc.

The term "biotechnology" is widely used and encompasses many different technologies. Consulting firms provide common definitions of modern biotechnology. The term "modern biotechnology" refers to all innovative methods, processes or products, including the use of living organisms or their cellular compartments, and the use of biochemistry, molecular biology, immunology, virology, microbiology, cell biology or environmental sciences and engineering. Biotechnology and entrepreneurship are intrinsically linked together, and are studied biotechnology at the regional, firm, and individual level of analysis The concept of "bioentrepreneurship", is described as a wealth created by applying the life sciences to a business environment. Bioentrepreneurs seek business value in the technologies they use to conduct biotechnology research. Some well-known bio startups are based on multiple companies. Biotechnology and entrepreneurship are essentially linked. In recent years, a large number of articles in the business literature have studied biotechnology at the level of analysis of regions, companies and individuals. This review article will encourage stakeholders to address the research space which have been recognized and will help more progress in this captivating area of ​​interest in the field of biotechnology and entrepreneurship.

The life sciences present a politically and ethically sensitive area of technology development. NBIC convergence—the convergence of nanotechnology, biotechnology, and information and cognitive technology—presents an increased interaction between the biological and physical sciences. As a result the bio-debate is no longer dominated by biotechnology, but driven by NBIC convergence. NBIC convergence enables two bioengineering megatrends: “biology becoming technology” and “technology becoming biology.” The notion of living technologies captures the latter megatrend. Accordingly, living technology presents a politically and ethically sensitive area. This implies that governments sooner or later are faced with the challenge of both promoting and regulating the development of living technology. This article describes four current political models to deal with innovation promotion and risk regulation. Based on two specific developments in the field of living technologies—(psycho)physiological computing and synthetic biology—we reflect on appropriate governance strategies for living technologies. We conclude that recent pleas for anticipatory and deliberative governance tend to neglect the need for anticipatory regulation as a key factor in guiding the development of the life sciences from a societal perspective. In particular, when it is expected that a certain living technology will radically challenge current regulatory systems, one should opt for just such a more active biopolitical approach.

AbstractGelatin is a protein obtained from the hydrolysis of collagen. Gelatin is an attractive biodegradable material for use in nano-biotechnology and nano-pharmaceutics. Gelatin nanoparticles (NPs) have been widely used as drug and gene carrier to targeted sick tissues including cancer, tuberculosis, HIV infection along with the treatment of vasospasm and restenosis, due to its biocompatibility and biodegradability. For instance, coating with gelatin lowers the cytotoxicity of quantum dots. Moreover, gelatin NPs have the ability to cross the blood-brain barrier, hence proven as a promising candidate to target brain disorders. Macrophage targeting with gelatin NPs for remedy of different diseases is repeatedly reported in previous years. In tissue engineering gelatin is actively utilized for construction of biological and life-long 3D scaffolds for bio-artificial tissues and organ production. Gelatins have a wide range of potential applications which needs to be unraveled in more detail. This review is mainly focused on the applications of gelatin NPs in biomedical sciences.

The grand challenge facing the chemical and allied industries in the twenty-first century is the transition to greener, more sustainable manufacturing processes that efficiently use raw materials, eliminate waste and avoid the use of toxic and hazardous materials. It requires a paradigm shift from traditional concepts of process efficiency, focusing on chemical yield, to one that assigns economic value to replacing fossil resources with renewable raw materials, eliminating waste and avoiding the use of toxic and/or hazardous substances. The need for a greening of chemicals manufacture is readily apparent from a consideration of the amounts of waste generated per kilogram of product (the E factors) in various segments of the chemical industry. A primary source of this waste is the use of antiquated ‘stoichiometric’ technologies and a major challenge is to develop green, catalytic alternatives. Another grand challenge for the twenty-first century, driven by the pressing need for climate change mitigation, is the transition from an unsustainable economy based on fossil resources—oil, coal and natural gas—to a sustainable one based on renewable biomass. In this context, the valorization of waste biomass, which is currently incinerated or goes to landfill, is particularly attractive. The bio-based economy involves cross-disciplinary research at the interface of biotechnology and chemical engineering, focusing on the development of green, chemo- and biocatalytic technologies for waste biomass conversion to biofuels, chemicals and bio-based materials. Biocatalysis has many benefits to offer in this respect. The catalyst is derived from renewable biomass and is biodegradable. Processes are performed under mild conditions and generally produce less waste and are more energy efficient than conventional ones. Thanks to modern advances in biotechnology ‘tailor-made’ enzymes can be economically produced on a large scale. However, for economic viability it is generally necessary to recover and re-use the enzyme and this can be achieved by immobilization, e.g. as solid cross-linked enzyme aggregates (CLEAs), enabling separation by filtration or centrifugation. A recent advance is the use of ‘smart’, magnetic CLEAs, which can be separated magnetically from reaction mixtures containing suspensions of solids; truly an example of cross-disciplinary research at the interface of physical and life sciences, which is particularly relevant to biomass conversion processes.

Recent research on biological materials and bioartificial systems has created one of the most dynamic field at the confluence of physical sciences, molecular engineering, cell biology, materials sciences, biotechnology and (nano) medicine. This field concerns better understanding of living systems, design of bio-inspired materials, synthesis of bioartificial technologies with new properties depending on their multi-scale architectures. Biological and man-made systems show the first level of organization at the nanoscale, where the fundamental properties and functions are settled (e.g., proteome and genome). The nanoscale properties reflect on larger scales: mesoscale, microscale, and continuum. Mechanisms by which phenomena at the different length and time scales are coupled and influence each other is the central issue in linking properties to functionalities, with a dramatic impact in designing and engineering biosystems. To get insights into the progressive trough-scales cascade effects-from molecular to macroscale level and from nanoseconds to life expectancy duration-multiscale/multiphysics models are required, dealing with inorganic, biological and hybrid matter. Thus, bioartificial systems technology depends upon our ability in assembling molecules into objects, hierarchically along several length scales, and in disassembling objects into molecules, in a tailored manner. As a peculiar feature, in bioartificial systems, the definition of the interactions between artificial and biological components needs to incorporate the “time” variable, in order to reproduce the evolution of the overall system, and to simulate complex phenomena as biodegradation and tissue remodeling. Herein, a number of paradigmatic multiscale models that attend the investigation of biological systems and the engineering of bioartificial systems is reviewed and discussed.

Sweet potato (Ipomoea batatas) is one of the most important crops in eastern Asia, including Korea. Consumption of sweet potato is increasing gradually because of its growing reputation as a health food. Recently, outbreaks of viruses infecting sweet potatoes have increased all over the world, probably because sweet potatoes are produced via vegetative propagation (1,2). In Korea, most sweet potatoes in fields have been infected by a begomovirus, Sweet potato leaf curl virus (SPLCV), and other viruses such as Sweet potato feathery mottle virus, Sweet potato virus G, and Sweet potato latent virus (3). Many countries have monitored sweet potato virus infections in fields as well as in germplasm collections to select virus-free stocks. In 2013, 20 sweet potato plants showing leaf roll symptoms in Muan, South Korea, were collected and analyzed. Total DNA was isolated from sweet potato leaves (Viral Gene-spin Viral DNA/RNA Extraction Kit, iNtRON Biotechnology, Seongnam, Korea) and viral DNA was amplified by rolling circle amplification (RCA, TempliPhi Amplification Kit, GE Healthcare Life Sciences, Uppsala, Sweden) following the manufacturer's instructions. Amplicons were digested by restriction enzyme SacI (TaKaRa Bio, Shiga, Japan) and products were run on a 1.5% agarose gel. A 2.8-kb DNA fragment was purified from a gel, ligated into a pGEM-T easy vector (Promega, Madison, WI), and sequenced (Macrogen, Seoul, Korea). Based on a BLAST search, most of the sequences (36/38) were identified as SPLCV, but two independent clones 2,824 nt in length from sweet potato cv. Sincheonmi were similar to Sweet potato golden vein associated virus (SPGVaV) isolate US:MS:1B-3 (94.38%, GenBank Accession No. HQ333143). The complete genome sequence of the SPGVaV-Korea isolate contained six ORFs, as expected for a typical monopartite begomovirus. The sequence was deposited in GenBank under accession number KF803170. SPGVaV is a whitefly (Bemisia tabaci)-transmitted virus (genus Begomovirus, family Geminiviridae). A phylogenetic analysis that included other begomoviruses that infect sweet potato showed SPGVaV-Korea to segregate with other SPGVaV isolates. SPGVaV has previously only been reported in Brazil and the United States (1). This is the first report of SPGVaV in sweet potato outside of the Americas. References: (1) L. C. Albuquerque et al. Virol. J. 9:241, 2012. (2) E. Choi et al. Acta Virol. 56:187, 2012. (3) H. R. Kwak et al. Plant Pathol. J. 22:239, 2006.

Eustoma (Eustoma grandiflorum), also called lisianthus, belongs to the family Gentianaceae and is cultivated for flower production globally (1), including in Korea. At least 10 viruses can infect eustoma, including Cucumber mosaic virus (genus Cucumovirus), Tobacco mosaic virus (genus Tobamovirus), Tomato spotted wilt virus (genus Tospovirus), and Tomato yellow leaf curl virus (TYLCV, genus Begomovirus) (1,2). In December 2012, disease symptoms such as leaf curling and stunting were observed on eustoma plants grown in Gumi, Korea, where TYLCV outbreak was reported on tomato farms. In a eustoma greenhouse, about 5% of eustoma plants showed the leaf curling and stunting symptoms. Total DNA was isolated from 15 symptomatic eustoma plants with a Viral Gene-spin Viral DNA/RNA Extraction Kit (iNtRON Biotechnology, Seongnam, Korea) and viral DNA was amplified by rolling circle amplification (TempliPhi Amplification Kit, GE Healthcare Life Sciences, Uppsala, Sweden) following the manufacturer's instructions. All amplicons were digested with the restriction enzyme SacI (TaKaRa Bio, Shiga, Japan) and 2.8-kb DNA fragments were verified on an agarose gel. Fifteen digested DNA fragments were purified from the gel, ligated into pGEM-T easy vector (Promega, Madison, WI), and sequenced (Macrogen, Seoul, Korea, GenBank Accession No. KF225312.1). A BLAST search exhibited a 99% identity to TYLCV previously reported in Korea (GenBank HM856911.1). This is the first report of TYLCV in eustoma plants in Korea. To identify the movement and replication of TYLCV in infected eustoma plants, PCR and Southern hybridization analysis were performed with samples from four organs (flower, leaf, stem, and root) of three individual TYLCV-infected plants. TYLCV TYL DNA from each organ sample was amplified using 2× Taq PCR MasterMix (Bioneer, Daejeon, Korea) with TYLCV-specific primers (TYLCV-F: 5′-ATATTACCGGATGGCCGCGCCT-3′, CV-R: 5′-TCCACGGGGAACATCAGGGCTT-3′). Single-stranded as well as double-stranded TYLCV DNA were identified from all organs of symptomatic eustoma, indicating TYLCV can replicate and move systemically in eustoma plants. Whitefly (Bemisia tabaci)-mediated plant-to-plant viral transmission was performed with one TYLCV-infected eustoma plant and five healthy eustoma plants and revealed that 80% (4 of 5) of the eustoma plants were infected by whitefly-mediated transmission. These results indicate that TYLCV-infected eustoma plants could act as virus reservoirs to healthy eustoma plants as well as other potential TYLCV hosts, such as tomatoes. In Korea, TYLCV has been the most notorious plant virus since 2008 (3), but, until now, TYLCV infection in eustoma plants has not been reported in Korea. References: (1) C. C. Chen et al. Plant Dis. 84:506, 2000. (2) A. Kritzman et al. Plant Dis. 84:1185, 2000. (3) H. Lee et al. Mol. Cells 30:467, 2010.

Background: Relapsed and refractory myeloid malignancies including myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML) remain a clinical challenge due to high morbidity and mortality as well as a paucity of effective therapeutic agents. PLX2853 is an orally available, non-benzodiazepine bromodomain and extraterminal domain (BET) inhibitor that exhibits low nanomolar potency and a modest preference for binding to the second bromodomain (BD2) of the BET proteins. By regulating genes (e.g., MYC and BCL2) critical to leukemic cell growth and survival, PLX2853 has demonstrated broad anti-leukemic activity both as a single agent and in combination with other therapeutic agents in preclinical models. The pharmacokinetic (PK) profiles in patients with AML or MDS revealed high peak plasma concentrations, a short terminal half-life (T1/2 < 3 hour), and nearly complete elimination from the plasma by 9 hour post dose. This PK profile is hypothesized to enable improved tolerability by allowing transient target engagement followed by time for recovery after daily dosing. Methods: This is an open-label, Phase 1b (Ph1b) study of PLX2853 as a single oral agent administered daily for 21 day cycles in adult patients with relapsed or refractory AML or high risk MDS. A modified continuous reassessment model with escalation with overdose control is employed to determine the recommended phase 2 dose (RP2D). Primary objectives include safety and PK. Secondary objectives include measures of preliminary efficacy, and exploratory objectives include pharmacodynamics (PD) biomarker assessments. Enrollment through Cohort 5 (140 mg QD) is ongoing as of July 2020. Results: 9 subjects with relapsed or refractory AML and 7 subjects with high risk MDS (median age 65.7 years, range 47-77 years old; median number of prior therapies 2.3, range 1-5 prior therapies) have received PLX2853 in escalating doses from 20 to 140 mg QD. Through the data cut-off of 19 Jul 2020 (n=16), the most common treatment emergent adverse events (AEs) regardless of causality occurring in ≥20% of patients (n≥4) are: decreased appetite (n=9), (n=8), fatigue (N=8), hyperbilirubinemia (N=8), nausea (n=7), constipation (n=6), hypokalemia (n=6), leukopenia (n=6), vomiting (n=5), international normalised ratio increased (n=5), proteinuria (n=5), and dyspnea (n=5). Most were grade (G) 1-2. Treatment emergent AEs > G2 in > 1 patients are: anemia (n=7), leukopenia (n=5) hyperbilirubinemia (n=4), febrile neutropenia (n=4), sepsis (n=3), thrombocytopenia (n=3), hypertension (n=2), lymphocyte count decreased (n=3), and neutrophil count decreased (n=3). No dose limiting toxicity (DLT) has been observed. Following a daily dose of PLX2853 the median time to reach maximal plasma concentration (Tmax) is 1 hour, and no accumulation observed at steady state, which is consistent with the short T1/2 (< 3 hours). Dose-dependent increases in exposures were observed across the dose range tested (20-80 mg daily). Fifteen of 16 subjects have completed at least 1 cycle of treatment. The following best overall responses have been observed: 1 subject with a confirmed marrow complete remission, 1 subject with a partial remission (myeloid sarcoma), 10 subjects with stable disease, 1 subject with progressive disease, and 3 subjects not evaluable. Conclusions: In an ongoing Ph1b study of PLX2853, four cohorts have been completed with continuous, once daily dosing in patients with relapsed or refractory AML or high risk MDS, and no DLTs have been observed yet. As dose escalation continues, PK, PD, preliminary safety, and efficacy data will be reviewed further to determine the clinical significance of this BET inhibitor and identify the RP2D. This clinical trial is registered at clinicaltrials.gov: NCT03787498. Disclosures Mims: Agios: Consultancy; Kura Oncology: Membership on an entity's Board of Directors or advisory committees; Leukemia and Lymphoma Society: Other: Senior Medical Director for Beat AML Study; Novartis: Speakers Bureau; Abbvie: Membership on an entity's Board of Directors or advisory committees; Syndax Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees; Jazz Pharmaceuticals: Other: Data Safety Monitoring Board. Borate:Pfizer: Membership on an entity's Board of Directors or advisory committees, Research Funding; Novartis: Membership on an entity's Board of Directors or advisory committees, Research Funding; Jazz Pharmaceuticals: Research Funding; AbbVie: Other: Investigator in AbbVie-funded clinical trials; Takeda: Membership on an entity's Board of Directors or advisory committees, Research Funding; Daiichi Sankyo: Membership on an entity's Board of Directors or advisory committees; Genentech: Membership on an entity's Board of Directors or advisory committees. Pemmaraju:Daiichi Sankyo: Research Funding; Novartis: Honoraria, Research Funding; Pacylex Pharmaceuticals: Consultancy; Cellectis: Research Funding; SagerStrong Foundation: Other: Grant Support; Celgene: Honoraria; Blueprint Medicines: Honoraria; MustangBio: Honoraria; DAVA Oncology: Honoraria; Roche Diagnostics: Honoraria; Samus Therapeutics: Research Funding; Affymetrix: Other: Grant Support, Research Funding; Incyte Corporation: Honoraria; Stemline Therapeutics: Honoraria, Research Funding; AbbVie: Honoraria, Research Funding; Plexxikon: Research Funding; LFB Biotechnologies: Honoraria. Borthakur:BioTherix: Consultancy; BioLine Rx: Consultancy; Argenx: Consultancy; FTC Therapeutics: Consultancy; Curio Science LLC: Consultancy; Oncoceutics: Research Funding; Xbiotech USA: Research Funding; Polaris: Research Funding; BMS: Research Funding; Jannsen: Research Funding; BioLine Rx: Research Funding; Cyclacel: Research Funding; GSK: Research Funding; Abbvie: Research Funding; Novartis: Research Funding; Incyte: Research Funding; PTC Therapeutics: Research Funding; Nkarta Therapeutics: Consultancy; Treadwell Therapeutics: Consultancy; PTC Therapeutics: Consultancy; AstraZeneca: Research Funding. Roboz:Agios: Consultancy; Amphivena: Consultancy; MEI Pharma: Consultancy; Helsinn: Consultancy; Epizyme: Consultancy; Jasper Therapeutics: Consultancy; Cellectis: Research Funding; Trovagene: Consultancy; Takeda: Consultancy; Otsuka: Consultancy; Orsenix: Consultancy; AstraZeneca: Consultancy; Daiichi Sankyo: Consultancy; Astellas: Consultancy; Argenx: Consultancy; Actinium: Consultancy; Sandoz: Consultancy; Astex: Consultancy; Celgene: Consultancy; Janssen: Consultancy; Novartis: Consultancy; Pfizer: Consultancy; Abbvie: Consultancy; Array BioPharma: Consultancy; Bayer: Consultancy; Celltrion: Consultancy; Eisai: Consultancy; Jazz: Consultancy; Roche/Genentech: Consultancy. Powell:Plexxikon Inc.: Current Employment. Severson:Plexxikon Inc.: Current Employment. Matusow:Plexxikon Inc.: Current Employment. Halladay:Plexxikon Inc.: Current Employment. Hsu:Daiichi Sankyo Inc.: Current Employment. Watkins:Plexxikon Inc.: Current Employment. Zhang:Plexxikon Inc.: Current Employment. Walling:Plexxikon Inc.: Consultancy; Aduro Biotech: Consultancy; Arch Oncology: Consultancy; CytomX Therapeutics: Consultancy; Harpoon Therapeutics: Consultancy; ImmuNext: Consultancy; Myovant Sciences: Consultancy; Nurix Therapeutics: Consultancy; Que Oncology: Consultancy; Sesen Bio: Consultancy; Amgen: Consultancy; Aminex: Consultancy; Crown Bio: Consultancy; Leap Therapeutics: Consultancy; Prothena Corporation: Consultancy; Puma Biotechnology: Consultancy; Rhizen Pharmaceuticals: Consultancy; Shanghai Pharmaceuticals: Consultancy; Stealth Biotherapeutics: Consultancy; Sunesis Pharmaceuticals: Consultancy; Upsher-Smith Laboratories: Consultancy; Flag Therapeutics: Consultancy; January Therapeutics: Consultancy. Tsiatis:Plexxikon Inc.: Current Employment. DeZern:Celgene: Consultancy, Honoraria; Astex: Research Funding; Abbvie: Consultancy; MEI: Consultancy.

Abstract Introduction: Multiple myeloma (MM) is a chronic hematologic malignancy with a high symptom burden that can have a substantial negative impact on patients' (pts) health-related quality of life (HRQoL). The introduction of novel triplet regimens for newly diagnosed MM (NDMM) has extended progression-free survival (PFS) and overall survival (OS); however, adverse events and demanding administration and monitoring schedules have a further negative effect on HRQoL, especially among pts who are transplant ineligible (TIE) due to older age and/or frailty. Optimizing initial treatment is particularly important in older pts, many of whom receive only 1 line of therapy. The phase 3 MAIA trial compared daratumumab, lenalidomide, and dexamethasone (D-Rd) vs lenalidomide and dexamethasone (Rd) in TIE pts with NDMM. At a median follow-up of 28 months, D-Rd significantly prolonged PFS and was associated with faster and sustained clinically meaningful improvements in patient-reported outcomes (PROs) vs Rd. Results of an updated analysis with longer follow-up recently confirmed a significant benefit in OS with D-Rd vs Rd as well as a continued significant PFS benefit and higher rates of complete response or better and very good partial response or better. Here we present an update of the HRQoL analysis with additional follow-up. Methods: MAIA (NCT02252172) is a randomized, open-label, active controlled, multicenter, phase 3 study of TIE pts with NDMM who were randomly assigned 1:1 to receive D-Rd or Rd until disease progression (PD) or unacceptable toxicity. PROs were recorded using the European Organization for Research and Treatment of Cancer Quality of Life Questionnaire Core 30-item (EORTC QLQ-C30) and the EQ-5D-5L visual analog scale. EORTC QLQ-C30 has 30 items comprising 5 functional scales (physical, role, emotional, cognitive, and social functioning), 1 global health status (GHS) scale, 3 symptom scales (fatigue, nausea and vomiting, and pain) and 6 single items (dyspnea, insomnia, appetite loss, constipation, diarrhea, and financial difficulties). Questionnaires were completed at baseline, on day 1 of cycles 3, 6, 9, and 12 for year 1, and every 6 months thereafter until PD. Analyses were conducted on all pts with a baseline and ≥1 post-baseline PRO assessment. Pts were censored at PD or discontinuation of study treatment. Thresholds for meaningful improvement and worsening were defined a priori based on published literature (≥10-point change). Treatment effect was analyzed using a mixed-effects model for repeated measurements including baseline value, visit, treatment, visit by treatment interaction, and randomization stratification factors as fixed effects and individual subject as random effect. Results: At a median follow-up of 56.2 months, discontinuation rates were lower with D-Rd than Rd (56.8% vs 80.8%). For GHS, physical functioning, fatigue, pain, and dyspnea, PROs that are particularly relevant to pts with MM, a numerically greater proportion of pts achieved a meaningful improvement with D-Rd vs Rd (Table 1). Differences were significant for physical functioning, fatigue, and dyspnea. The proportion of pts achieving a meaningful worsening on therapy was similar in both treatment groups (Table 1). The median time to improvement was numerically shorter with D-Rd vs Rd for physical functioning and pain and with Rd vs D-Rd for GHS and fatigue; differences were not significant (Table 2). The median time to worsening of fatigue was similar between groups, numerically longer for D-Rd vs Rd for GHS, and significantly longer with D-Rd than Rd for physical functioning, pain, and dyspnea (Table 2). Median time to worsening of pain with D-Rd vs Rd was 39.43 vs 17.97 months, reflective of an additional ~21 months without worsening pain among pts treated with D-Rd. Between-group differences for least squares mean change from baseline for these 5 PROs favored D-Rd vs Rd at all assessment time points except cycle 3 for physical functioning and cycle 6 for fatigue; differences were significant at ≥1 timepoint for each scale. Conclusions: These updated PRO analyses from the MAIA study demonstrate sustained and clinically meaningful improvements in HRQoL with D-Rd vs Rd with almost 5 years of follow-up. These results are consistent with the clinical benefits of superior PFS, OS, and deep responses observed with D-Rd compared with Rd and support the use of D-Rd in older pts. Figure 1 Figure 1. Disclosures Perrot: Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees; GSK: Honoraria, Membership on an entity's Board of Directors or advisory committees; Celgene/BMS: Honoraria, Membership on an entity's Board of Directors or advisory committees; Abbvie: Honoraria; Amgen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Sanofi: Honoraria; Takeda: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding. Kumar: Roche-Genentech: Consultancy, Research Funding; Novartis: Research Funding; Celgene: Membership on an entity's Board of Directors or advisory committees, Research Funding; Janssen: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Sanofi: Research Funding; Bluebird Bio: Consultancy; Tenebio: Research Funding; Adaptive: Membership on an entity's Board of Directors or advisory committees, Research Funding; Astra-Zeneca: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Carsgen: Research Funding; KITE: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Merck: Research Funding; Antengene: Consultancy, Honoraria; Oncopeptides: Consultancy; Beigene: Consultancy; BMS: Consultancy, Research Funding; Abbvie: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Amgen: Consultancy, Research Funding; Takeda: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding. Plesner: Takeda: Research Funding; Oncopeptides: Other: Advisor, Research Funding; Genentech: Other: Advisor, Research Funding; CSL Behring: Other: Advisor; AbbVie: Other: Advisor, Research Funding; Celgene: Other: Advisor, Research Funding; Janssen: Other: Advisor, Research Funding; Genmab: Research Funding. Orlowski: Amgen, Inc., BioTheryX, Inc., Bristol-Myers Squibb, Celgene, EcoR1 Capital LLC, Genzyme, GSK Biologicals, Janssen Biotech, Karyopharm Therapeutics, Inc., Neoleukin Corporation, Oncopeptides AB, Regeneron Pharmaceuticals, Inc., Sanofi-Aventis, and Takeda P: Consultancy, Honoraria; CARsgen Therapeutics, Celgene, Exelixis, Janssen Biotech, Sanofi-Aventis, Takeda Pharmaceuticals North America, Inc.: Other: Clinical research funding; Asylia Therapeutics, Inc., BioTheryX, Inc., and Heidelberg Pharma, AG.: Other: Laboratory research funding; Asylia Therapeutics, Inc.: Current holder of individual stocks in a privately-held company, Patents & Royalties; Amgen, Inc., BioTheryX, Inc., Bristol-Myers Squibb, Celgene, Forma Therapeutics, Genzyme, GSK Biologicals, Janssen Biotech, Juno Therapeutics, Karyopharm Therapeutics, Inc., Kite Pharma, Neoleukin Corporation, Oncopeptides AB, Regeneron Pharmaceuticals, I: Membership on an entity's Board of Directors or advisory committees. Moreau: Oncopeptides: Honoraria; Sanofi: Honoraria; Janssen: Honoraria; Celgene BMS: Honoraria; Amgen: Honoraria; Abbvie: Honoraria. Bahlis: BMS/Celgene: Consultancy, Honoraria; Janssen: Consultancy, Honoraria; Pfizer: Consultancy, Honoraria; Sanofi: Consultancy, Honoraria; Genentech: Consultancy; GlaxoSmithKline: Consultancy, Honoraria; Amgen: Consultancy, Honoraria; Abbvie: Consultancy, Honoraria; Takeda: Consultancy, Honoraria; Karyopharm: Consultancy, Honoraria. Nahi: XNK Therapeutics AB: Consultancy. Hulin: Takeda: Honoraria; Sanofi: Honoraria; Celgene/BMS: Honoraria; Janssen: Honoraria; abbvie: Honoraria. Quach: CSL: Consultancy, Membership on an entity's Board of Directors or advisory committees; Antengene: Consultancy, Membership on an entity's Board of Directors or advisory committees; GlaxoSmithKline: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Amgen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Bristol Myers Squibb: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Celgene: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Karyopharm: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Sanofi: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Janssen/Cilag: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Takeda: Consultancy, Membership on an entity's Board of Directors or advisory committees. Goldschmidt: Chugai: Honoraria, Other: Grants and/or Provision of Investigational Medicinal Product, Research Funding; BMS: Consultancy, Honoraria, Other: Grants and/or Provision of Investigational Medicinal Product, Research Funding; Incyte: Research Funding; Janssen: Consultancy, Honoraria, Other: Grants and/or Provision of Investigational Medicinal Product, Research Funding; Johns Hopkins University: Other: Grant; Molecular Partners: Research Funding; MSD: Research Funding; Mundipharma: Research Funding; Sanofi: Consultancy, Honoraria, Other: Grants and/or Provision of Investigational Medicinal Product, Research Funding; Takeda: Consultancy, Research Funding; Adaptive Biotechnology: Consultancy; Celgene: Consultancy, Honoraria, Other: Grants and/or Provision of Investigational Medicinal Product, Research Funding; Novartis: Honoraria, Research Funding; Dietmar-Hopp-Foundation: Other: Grant; GSK: Honoraria; Amgen: Consultancy, Honoraria, Other: Grants and/or Provision of Investigational Medicinal Product, Research Funding. O'Dwyer: ONK Therapeutics: Current Employment, Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Janssen: Consultancy; Bristol Myers Squibb: Research Funding. Venner: BMS: Honoraria; Amgen: Research Funding; Celgene: Research Funding; Amgen: Honoraria; Takeda: Honoraria; Janssen: Honoraria; Sanofi: Honoraria; Pfizer: Honoraria. Weisel: Celgene: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Amgen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Sanofi: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Bristol Myers Squibb: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Adaptive Biotechnologies: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; GSK: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Janssen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Roche: Honoraria; Takeda: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Karyopharm: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Oncopeptides: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Abbvie: Consultancy; Novartis: Honoraria; Pfizer: Honoraria. Raje: Celgene, Amgen, Bluebird Bio, Janssen, Caribou, and BMS: Other. Macro: Sanofi: Honoraria; GSK: Honoraria; Takeda: Honoraria, Other: Travel accomodation, Research Funding; Janssen: Honoraria, Other: Travel accomodation, Research Funding; Celgen/BMS: Honoraria. Leleu: Sanofi: Honoraria; Takeda: Honoraria, Other: Non-financial support; Janssen-Cilag: Honoraria; Karyopharm Therapeutics: Honoraria; Merck: Honoraria; Mundipharma: Honoraria; Novartis: Honoraria; Oncopeptides: Honoraria; Gilead Sciences: Honoraria; Celgene: Honoraria; Carsgen Therapeutics Ltd: Honoraria; Bristol-Myers Squibb: Honoraria; Amgen: Honoraria; AbbVie: Honoraria; Pierre Fabre: Honoraria; Roche: Honoraria. Liu: Janssen: Current Employment, Current equity holder in publicly-traded company. Fastenau: Janssen: Current Employment, Current equity holder in publicly-traded company. Gries: Janssen: Current Employment, Current holder of individual stocks in a privately-held company. Ho: DRG Abacus: Consultancy; Janssen: Consultancy; Emalex Biosciences: Consultancy. Mistry: Janssen: Current Employment, Current equity holder in publicly-traded company. Tromp: Janssen: Current Employment, Current equity holder in publicly-traded company. Delioukina: Janssen: Current Employment. Vermeulen: Janssen: Current Employment, Current equity holder in publicly-traded company. Usmani: Abbvie: Consultancy; Array BioPharma: Consultancy, Research Funding; Celgene/BMS: Consultancy, Research Funding, Speakers Bureau; GSK: Consultancy, Research Funding; EdoPharma: Consultancy; Janssen: Consultancy, Research Funding, Speakers Bureau; Sanofi: Consultancy, Research Funding, Speakers Bureau; Merck: Consultancy, Research Funding; Pharmacyclics: Consultancy, Research Funding; Seattle Genetics: Consultancy, Research Funding; SkylineDX: Consultancy, Research Funding; Takeda: Consultancy, Research Funding, Speakers Bureau; Janssen Oncology: Consultancy, Research Funding; Bristol-Myers Squibb: Research Funding; Amgen: Consultancy, Research Funding, Speakers Bureau.

Abstract T cell regenerative medicine represents an emerging immunotherapeutic approach using antigen-specific Induced Pluripotent Stem Cells (iPSC) to rejuvenate CD8 + cytotoxic T lymphocytes (CTL). Here we report on an iPSC-derived therapeutic strategy targeting B-Cell Maturation Antigen (BCMA) against multiple myeloma (MM) via establishment of antigen-specific iPSC, followed by differentiation into highly functional BCMA-specific CD8 + CTL. The reprogrammed BCMA-specific iPSC displayed normal karyotypes and pluripotency potential as evidenced by expression of stem cell markers (SSEA-4, TRA1-60) and alkaline phosphatase, along with differentiation into three germ layers (Ectoderm, Mesoderm, Endoderm). During embryoid body formation, BCMA-specific iPSC further polarized into the mesoderm germ layer, evidenced by the activation of SNAI2, TBX3, PLVAP, HAND1 and CDX2 transcriptional regulators. Next, the BCMA-specific iPSC clones committed to CD8 + T cell differentiation were characterized by analyzing their hematopoietic progenitor cells (HPC; CD34 + CD43 +/CD14 - CD235a -) for specific transcriptional regulation. RNAseq analyses indicated a low variability and similar profiles of gene transcription within the iPSC clones committed to CD8 + CTL compared to increased transcriptional variability within iPSC clones committed to different cell types. The unique transcriptional profiles of the iPSC committed to CD8 + T cells included upregulation of transcriptional regulators controlling CD4/CD8 T cell differentiation ratio, memory CTL formation, NF-kappa-B/JNK pathway activation, and cytokine transporter/cytotoxic mediator development, as well as downregulation of regulators controlling B and T cell interactions, CD4 + Th cells, and inhibitory receptor development. Specifically, a major regulatory shift, indicated by upregulation of specific genes involved in immune function, was detected in HPC from the iPSC committed to CD8 + T cells. BCMA-specific T cells differentiated from the iPSC were characterized as displaying mature CTL phenotypes including high expression of CD3, CD8a, CD8b, TCRab, CD7 along with no CD4 expression (Fig. 1). In addition, the final BCMA iPSC-T cells were predominantly CD45RO + memory cells (central memory and effector memory cells) expressing high level of T cell activation (CD38, CD69) and costimulatory (CD28) molecules. Importantly, these BCMA iPSC-T cells lacked immune checkpoints (CTLA4, PD1, LAG3, Tim3) expression and regulatory T cells induction, distinct from other antigen-stimulated T cells. The rejuvenated BCMA iPSC-T cells demonstrated a high proliferative (1,000 folds increase) during the differentiation process as well as poly-functional anti-tumor activities and Th1 cytokine (IFN-g, IL-2, TNF-a) production triggered in response to MM patients' cells in HLA-A2-restricted manner (Fig. 2). Furthermore, the immune responses induced by these BCMA iPSC-T cells were specific to the parent heteroclitic BCMA 72-80 (YLMFLLRKI) peptide used to reprogram and establish the antigen-specific iPSC. Evaluation of 88 single cell Tetramer + CTL from the BCMA iPSC-T cells revealed a clonotype of unique T cell receptor (TCRa, TCRb) sequence. The BCMA-specific iPSC clones maintained their specific differentiation potential into the antigen-specific CD8 + memory T cells, following multiple subcloning in long-term cultures under feeder-free conditions or post-thaw after long-term (18 months) cryopreservation at -140 oC, which provides additional benefits to treat patients in a continuous manner. Taken together, rejuvenated CD8 + CTL differentiated from BCMA-specific iPSC were highly functional with significant (*p < 0.05) levels of anti-MM activities including proliferation, cytotoxic activity and Th-1 cytokine production. Therefore, the antigen-specific iPSC reprogramming and T cells rejuvenation process can provide an effective and long-term source of antigen-specific memory CTL lacking immune checkpoints and suppressors for clinical application in adoptive immunotherapy to improve patient outcome in MM. Figure 1 Figure 1. Disclosures Munshi: Amgen: Consultancy; Karyopharm: Consultancy; Takeda: Consultancy; Adaptive Biotechnology: Consultancy; Bristol-Myers Squibb: Consultancy; Celgene: Consultancy; Abbvie: Consultancy; Janssen: Consultancy; Legend: Consultancy; Oncopep: Consultancy, Current equity holder in publicly-traded company, Other: scientific founder, Patents & Royalties; Novartis: Consultancy; Pfizer: Consultancy. Ritz: Amgen: Research Funding; Equillium: Research Funding; Kite/Gilead: Research Funding; Avrobio: Membership on an entity's Board of Directors or advisory committees; Akron: Consultancy; Biotech: Consultancy; Blackstone Life Sciences Advisor: Consultancy; Clade Therapeutics, Garuda Therapeutics: Consultancy; Immunitas Therapeutic: Consultancy; LifeVault Bio: Consultancy; Novartis: Consultancy; Rheos Medicines: Consultancy; Talaris Therapeutics: Consultancy; TScan Therapeutics: Consultancy. Anderson: Sanofi-Aventis: Membership on an entity's Board of Directors or advisory committees; Gilead: Membership on an entity's Board of Directors or advisory committees; Millenium-Takeda: Membership on an entity's Board of Directors or advisory committees; Janssen: Membership on an entity's Board of Directors or advisory committees; Bristol Myers Squibb: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees; Pfizer: Membership on an entity's Board of Directors or advisory committees; Scientific Founder of Oncopep and C4 Therapeutics: Current equity holder in publicly-traded company, Current holder of individual stocks in a privately-held company; AstraZeneca: Membership on an entity's Board of Directors or advisory committees; Mana Therapeutics: Membership on an entity's Board of Directors or advisory committees.

Abstract Introduction: In oncology clinical trials, overall survival (OS) is considered the gold standard efficacy endpoint. However, in multiple myeloma, the prolonged survival times and availability of multiple salvage therapies render it difficult to rely on OS as a primary endpoint. Instead, progression-free survival (PFS) or clinical response can be a relevant biomarker. At the same time, some health technology assessment bodies will only consider evidence based on patient-relevant endpoints, such as morbidity, mortality, and health-related quality of life (HRQoL), within their benefit assessments. Patient-reported outcomes (PROs) provide insights into how a treatment affects HRQoL, including symptoms and functioning. In the phase 3 MAIA trial, daratumumab, lenalidomide, and dexamethasone (D-Rd) demonstrated a significantly prolonged PFS and rapid and sustained improvements in PROs compared with lenalidomide and dexamethasone (Rd) alone at a median follow-up of 28 months. Updated results with longer follow-up confirmed a continued PFS benefit, deepening best clinical responses, and a significant OS benefit with D-Rd. Here, we report the results of analyses exploring the relationship between clinical efficacy endpoints and PROs in the MAIA trial. Methods: In MAIA (NCT02252172), TIE patients with NDMM were randomized to D-Rd or Rd until disease progression (PD) or unacceptable side effects. Clinical response and PD were defined per the International Myeloma Working Group uniform response criteria. PROs were assessed at baseline, on day 1 of cycles 3, 6, 9, and 12 for year 1, every 6 months thereafter until PD, or at end of treatment and at 8 and 16 weeks post-progression using the European Organization for Research and Treatment of Cancer Quality of Life Questionnaire Core 30-item (EORTC QLQ-C30). The current analyses were conducted on patients with a baseline PRO assessment and ≥1 post-baseline PRO assessment using a clinical cutoff date of February 19, 2021. Patients from both treatment groups were pooled and stratified by best clinical response. For PROs, the threshold for clinically meaningful change from baseline was defined a priori as ≥10 points based on published literature. Analyses were conducted for the global health status (GHS), fatigue, and pain scales of the EORTC QLQ-C30. The proportions of patients with clinically meaningful improvement in these PROs from baseline at any time while on treatment were calculated for and compared across best clinical response subgroups. For this analysis, patients were censored at the time of PD or discontinuation of therapy. Results were summarized with odds ratios and 95% confidence intervals. Additionally, the proportions of patients with clinically meaningful worsening in PROs from baseline at the time of and post-progression were also calculated for those who had PD. No adjustments were made for multiplicity. Nominal P-values are presented. Results: Best clinical response at a median follow-up of 56.2 months for the 710 patients included in this analysis is shown in the Table. For GHS, fatigue, and pain, the proportion of patients who reported clinically meaningful improvement from baseline increased with increasing depth of best clinical response. Odds ratios comparing best clinical response vs stable disease reflect this relationship and are shown in the Figure. Of the 264 patients who experienced PD and had PRO data at the time of or post-progression, 38.6%, 54.9%, and 39.4% reported meaningful worsening of GHS, pain, and fatigue, respectively, at the time of or post-progression. Conclusions: Patients with deeper best clinical response were more likely to report meaningful improvements in PROs. Patients who experienced PD reported worsening of GHS and symptoms at the time of or following progression. These analyses provide evidence of the association between key clinical efficacy endpoints and PROs and demonstrate the patient relevance of these clinical endpoints. Figure 1 Figure 1. Disclosures Perrot: Abbvie: Honoraria; BMS Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees; Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Amgen: Honoraria, Membership on an entity's Board of Directors or advisory committees; GSK: Honoraria, Membership on an entity's Board of Directors or advisory committees; Sanofi: Honoraria, Research Funding; Takeda: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding. Kumar: Amgen: Consultancy, Research Funding; BMS: Consultancy, Research Funding; KITE: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Novartis: Research Funding; Astra-Zeneca: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Merck: Research Funding; Abbvie: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Roche-Genentech: Consultancy, Research Funding; Bluebird Bio: Consultancy; Janssen: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Carsgen: Research Funding; Antengene: Consultancy, Honoraria; Oncopeptides: Consultancy; Beigene: Consultancy; Tenebio: Research Funding; Takeda: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Celgene: Membership on an entity's Board of Directors or advisory committees, Research Funding; Adaptive: Membership on an entity's Board of Directors or advisory committees, Research Funding; Sanofi: Research Funding. Plesner: Takeda: Research Funding; Genentech: Other: Advisor, Research Funding; Oncopeptides: Other: Advisor, Research Funding; AbbVie: Other: Advisor, Research Funding; CSL Behring: Other: Advisor; Janssen: Other: Advisor, Research Funding; Celgene: Other: Advisor, Research Funding; Genmab: Research Funding. Orlowski: Asylia Therapeutics, Inc., BioTheryX, Inc., and Heidelberg Pharma, AG.: Other: Laboratory research funding; CARsgen Therapeutics, Celgene, Exelixis, Janssen Biotech, Sanofi-Aventis, Takeda Pharmaceuticals North America, Inc.: Other: Clinical research funding; Amgen, Inc., BioTheryX, Inc., Bristol-Myers Squibb, Celgene, EcoR1 Capital LLC, Genzyme, GSK Biologicals, Janssen Biotech, Karyopharm Therapeutics, Inc., Neoleukin Corporation, Oncopeptides AB, Regeneron Pharmaceuticals, Inc., Sanofi-Aventis, and Takeda P: Consultancy, Honoraria; Asylia Therapeutics, Inc.: Current holder of individual stocks in a privately-held company, Patents & Royalties; Amgen, Inc., BioTheryX, Inc., Bristol-Myers Squibb, Celgene, Forma Therapeutics, Genzyme, GSK Biologicals, Janssen Biotech, Juno Therapeutics, Karyopharm Therapeutics, Inc., Kite Pharma, Neoleukin Corporation, Oncopeptides AB, Regeneron Pharmaceuticals, I: Membership on an entity's Board of Directors or advisory committees. Moreau: Abbvie: Honoraria; Oncopeptides: Honoraria; Celgene BMS: Honoraria; Sanofi: Honoraria; Amgen: Honoraria; Janssen: Honoraria. Bahlis: Amgen: Consultancy, Honoraria; Janssen: Consultancy, Honoraria; Abbvie: Consultancy, Honoraria; Pfizer: Consultancy, Honoraria; Sanofi: Consultancy, Honoraria; BMS/Celgene: Consultancy, Honoraria; Karyopharm: Consultancy, Honoraria; Takeda: Consultancy, Honoraria; Genentech: Consultancy; GlaxoSmithKline: Consultancy, Honoraria. Nahi: XNK Therapeutics AB: Consultancy. Hulin: abbvie: Honoraria; Celgene/BMS: Honoraria; Sanofi: Honoraria; Takeda: Honoraria; Janssen: Honoraria. Quach: Bristol Myers Squibb: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; CSL: Consultancy, Membership on an entity's Board of Directors or advisory committees; Janssen/Cilag: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; GlaxoSmithKline: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Amgen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Antengene: Consultancy, Membership on an entity's Board of Directors or advisory committees; Celgene: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Karyopharm: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Takeda: Consultancy, Membership on an entity's Board of Directors or advisory committees; Sanofi: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding. Goldschmidt: GSK: Honoraria; Incyte: Research Funding; Janssen: Consultancy, Honoraria, Other: Grants and/or Provision of Investigational Medicinal Product, Research Funding; Amgen: Consultancy, Honoraria, Other: Grants and/or Provision of Investigational Medicinal Product, Research Funding; BMS: Consultancy, Honoraria, Other: Grants and/or Provision of Investigational Medicinal Product, Research Funding; Adaptive Biotechnology: Consultancy; Celgene: Consultancy, Honoraria, Other: Grants and/or Provision of Investigational Medicinal Product, Research Funding; Chugai: Honoraria, Other: Grants and/or Provision of Investigational Medicinal Product, Research Funding; Novartis: Honoraria, Research Funding; Dietmar-Hopp-Foundation: Other: Grant; Sanofi: Consultancy, Honoraria, Other: Grants and/or Provision of Investigational Medicinal Product, Research Funding; Takeda: Consultancy, Research Funding; Johns Hopkins University: Other: Grant; Mundipharma: Research Funding; MSD: Research Funding; Molecular Partners: Research Funding. O'Dwyer: Bristol Myers Squibb: Research Funding; ONK Therapeutics: Current Employment, Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Janssen: Consultancy. Venner: Amgen: Research Funding; Celgene: Research Funding; Takeda: Honoraria; Amgen: Honoraria; Janssen: Honoraria; BMS: Honoraria; Sanofi: Honoraria; Pfizer: Honoraria. Weisel: Adaptive Biotechnologies: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Bristol Myers Squibb: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Takeda: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Amgen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Janssen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Roche: Honoraria; Sanofi: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Karyopharm: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; GSK: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Celgene: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Oncopeptides: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Abbvie: Consultancy; Novartis: Honoraria; Pfizer: Honoraria. Raje: Celgene, Amgen, Bluebird Bio, Janssen, Caribou, and BMS: Other. Macro: Takeda: Honoraria, Other: Travel accomodation, Research Funding; Janssen: Honoraria, Other: Travel accomodation, Research Funding; GSK: Honoraria; Celgen/BMS: Honoraria; Sanofi: Honoraria. Leleu: Sanofi: Honoraria; Roche: Honoraria; Pierre Fabre: Honoraria; Oncopeptides: Honoraria; Novartis: Honoraria; Mundipharma: Honoraria; Merck: Honoraria; Karyopharm Therapeutics: Honoraria; Amgen: Honoraria; Bristol-Myers Squibb: Honoraria; Carsgen Therapeutics Ltd: Honoraria; Celgene: Honoraria; Gilead Sciences: Honoraria; Janssen-Cilag: Honoraria; AbbVie: Honoraria; Takeda: Honoraria, Other: Non-financial support. Liu: Janssen: Current Employment, Current equity holder in publicly-traded company. Fastenau: Janssen: Current Employment, Current equity holder in publicly-traded company. Gries: Janssen: Current Employment, Current holder of individual stocks in a privately-held company. Ho: DRG Abacus: Consultancy; Emalex Biosciences: Consultancy; Janssen: Consultancy. Mistry: Janssen: Current Employment, Current equity holder in publicly-traded company. Tromp: Janssen: Current Employment, Current equity holder in publicly-traded company. Delioukina: Janssen: Current Employment. Vermeulen: Janssen: Current Employment, Current equity holder in publicly-traded company. Usmani: Amgen: Consultancy, Research Funding, Speakers Bureau; Abbvie: Consultancy; Array BioPharma: Consultancy, Research Funding; GSK: Consultancy, Research Funding; EdoPharma: Consultancy; SkylineDX: Consultancy, Research Funding; Celgene/BMS: Consultancy, Research Funding, Speakers Bureau; Takeda: Consultancy, Research Funding, Speakers Bureau; Seattle Genetics: Consultancy, Research Funding; Pharmacyclics: Consultancy, Research Funding; Merck: Consultancy, Research Funding; Janssen: Consultancy, Research Funding, Speakers Bureau; Sanofi: Consultancy, Research Funding, Speakers Bureau; Janssen Oncology: Consultancy, Research Funding; Bristol-Myers Squibb: Research Funding.

Abstract Background: Chimeric antigen receptor (CAR)-T cells are highly effective in patients (pts) with multiple myeloma (MM), but duration of response can be limited, and pts with rapidly progressing disease require a fast and reliable CAR-T cell manufacturing process. Here, we report initial clinical data from a Phase I trial assessing PHE885 manufactured using the T-Charge TM process and characterization of in vivo expansion, suggesting a preserved T-cell stemness (T scm) phenotype in pts with relapsed/refractory (r/r) MM (NCT04318327). Methods: PHE885 is a unique and fully human BCMA CAR-T cell product manufactured using the novel T-Charge TM platform, which reduces ex vivo culture time to about 24 hours and takes <2 days to manufacture the final product, thereby relying entirely on in vivo expansion after CAR-T cell infusion. Pts with MM r/r to ≥2 prior lines of treatment (tx), including an immunomodulatory drug, proteasome inhibitor, and an anti-CD38 monoclonal antibody, were eligible. Pts received fludarabine and cyclophosphamide for lymphodepletion prior to a single PHE885 intravenous injection. Primary objectives were safety, including dose-limiting toxicities (DLTs) and adverse events (AEs). Secondary objectives were clinical responses, evaluation of the T-Charge TM process, and pharmacokinetic properties. Results: As of data cut (April 1, 2021), 7 pts were enrolled in the dose escalation portion; 1 pt failed screening (prolonged QTc), and 6 pts were successfully infused with PHE885. All pts were heavily pretreated, penta-refractory, and refractory to the last line of tx. Fixed doses received were 5×10 6 (n=5) and 14.3×10 6 CAR+ T cells (n=1). All 6 pts were eligible for safety and efficacy. Two DLTs were reported: asymptomatic grade 3 transaminitis in the pt infused with 14.3×10 6 CAR+ T cells, and asymptomatic grade 4 lipase increased in 1 pt infused with 5×10 6 CAR+ T cells. Treatment-related grade ≥3 AEs included anemia and neutropenia in all pts; thrombocytopenia (n=4, 67%); and leukopenia, cytokine release syndrome (CRS), ALT and AST increase, and decreased blood fibrinogen (each n=2, 33%). All pts experienced grade ≤3 CRS per Lee 2014 criteria; median times to CRS onset and resolution were 7 d (range, 4-9 d) and 22 d (range, 10-27 d), respectively. All pts received at least 1 dose each of steroids and tocilizumab; 3 pts received anakinra to manage CRS. Two pts experienced grade 2 neurotoxicity related to PHE885. Both events were nonserious and temporally associated with grade 3 CRS. No deaths occurred on study. At 1 mo after tx, all pts had achieved at least a partial response (PR), with complete response (CR) in 1 pt (17%) and very good PR in 2 pts (33%). Of 4 pts evaluable at 3 mo after tx, 2 had stringent CR, 1 had PR, and 1 pt in PR experienced progressive disease, presumed to be due to loss of BCMA. Of 3 pts evaluable for minimal residual disease (MRD) at 1 mo after tx, all were MRD negative: 2 at sensitivity of 10 -6 and 1 at 10 -5. Robust cellular expansion was observed in all pts via qPCR and flow cytometry; maximum expansion (geometric mean C max) was 283000 copies/μg by qPCR and 69.3% of circulating T cells by flow cytometry. Maximum expansion was reached by 30 d, with median T max of 21.1 d by qPCR (16.4 d by flow cytometry). PHE885 was detectable in peripheral blood up to the latest measured sample for each pt (6 mo for the longest followed pt; range of follow-up, 1-6 mo). A naïve-like T-cell phenotype (T naïve+T scm) was preserved during manufacturing of all PHE885 products. Conclusions: Initial data from this Phase I study demonstrate that low doses of BCMA CAR-T cells manufactured by T-Charge TM in <2 days have encouraging clinical activity and a manageable safety profile in pts with r/r MM. PHE885 CAR-T cells expand rapidly in vivo, persist at relatively high levels for prolonged periods, and demonstrate a relatively immature T-cell phenotype. The trial is ongoing and updated data will be presented at the annual meeting. Clinical trial information: NCT04318327 Figure 1 Figure 1. Disclosures Sperling: Adaptive: Consultancy. Nikiforow: Kite/Gilead: Other: ad HOC Advisory Boards; Novartis: Other: ad Hoc Advisory Boards; Iovance: Other: ad Hoc Advisory Boards; Glaxo Smith Kline (GSK): Other: ad Hoc Advisory Boards. Nadeem: Bristol Myer Squibb: Consultancy; GSK: Consultancy; Adaptive: Consultancy; Karyopharm: Consultancy; Takeda: Consultancy. Mo: Eli Lilly: Consultancy; Epizyme: Consultancy; GSK: Consultancy, Membership on an entity's Board of Directors or advisory committees; Janssen: Honoraria; Karyopharm: Honoraria, Membership on an entity's Board of Directors or advisory committees; Sanofi: Honoraria, Membership on an entity's Board of Directors or advisory committees; BMS: Membership on an entity's Board of Directors or advisory committees; AbbVIE: Consultancy. Anderson: Sanofi-Aventis: Membership on an entity's Board of Directors or advisory committees; Pfizer: Membership on an entity's Board of Directors or advisory committees; Janssen: Membership on an entity's Board of Directors or advisory committees; Gilead: Membership on an entity's Board of Directors or advisory committees; Bristol Myers Squibb: Membership on an entity's Board of Directors or advisory committees; Millenium-Takeda: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees; Scientific Founder of Oncopep and C4 Therapeutics: Current equity holder in publicly-traded company, Current holder of individual stocks in a privately-held company; AstraZeneca: Membership on an entity's Board of Directors or advisory committees; Mana Therapeutics: Membership on an entity's Board of Directors or advisory committees. Ikegawa: Bristol Myers Squibb: Honoraria. Shaw: Orchard Therapeutics, Ltd: Current equity holder in publicly-traded company. Ansari: Novartis: Current Employment. Quinn: Novartis: Current Employment, Current equity holder in publicly-traded company. Pearson: Novartis: Current Employment, Current equity holder in publicly-traded company. Hack: Novartis: Current Employment. Treanor: Novartis: Current Employment, Current holder of individual stocks in a privately-held company, Divested equity in a private or publicly-traded company in the past 24 months, Patents & Royalties: no royalties as company-held patents. Bu: Novartis: Current Employment, Patents & Royalties: Co-inventor on patent applications. Mataraza: Novartis: Current Employment, Current holder of stock options in a privately-held company. Rispoli: Novartis: Current Employment. Credi: Novartis: Current Employment, Current equity holder in publicly-traded company, Divested equity in a private or publicly-traded company in the past 24 months. Ritz: Amgen: Research Funding; Equillium: Research Funding; Kite/Gilead: Research Funding; Avrobio: Membership on an entity's Board of Directors or advisory committees; Akron: Consultancy; Biotech: Consultancy; Blackstone Life Sciences Advisor: Consultancy; Clade Therapeutics, Garuda Therapeutics: Consultancy; Immunitas Therapeutic: Consultancy; LifeVault Bio: Consultancy; Novartis: Consultancy; Rheos Medicines: Consultancy; Talaris Therapeutics: Consultancy; TScan Therapeutics: Consultancy. De Vita: Novartis: Current Employment. Munshi: Celgene: Consultancy; Amgen: Consultancy; Takeda: Consultancy; Adaptive Biotechnology: Consultancy; Abbvie: Consultancy; Oncopep: Consultancy, Current equity holder in publicly-traded company, Other: scientific founder, Patents & Royalties; Janssen: Consultancy; Karyopharm: Consultancy; Novartis: Consultancy; Pfizer: Consultancy; Legend: Consultancy; Bristol-Myers Squibb: Consultancy.

Problem statement. The main issues that exist in the management of strains of particularly dangerous microorganisms are primarily related to the weakening of state supervision and control. Surprisingly, nowadays, there is no single official register of owners of pathogenic microorganisms and strains of dangerous and especially dangerous infectious diseases. Some biological objects are found, even in private collections. There are no permitting procedures for handling such facilities and sanctions for violating them. That is why it is not surprising that the Prosecutor General’s Office has announced that the head of the State Research and Control Institute of Biotechnology and Microorganism Strains has identified dangerous virus strains that are known as infectious animal pathogens and can spread rapidly on a large scale across national borders. Dual-use studies should be supervised to address the risks arising from the development of biomedical sciences. Continuous monitoring and verification of scientific and technological advances sensitive from the point of view of security allow to minimize the possibility of the adoption of biological and toxic weapons and other risks. Analysis of recent sources and publications. Theoretical issues of legal regulation of activities in the field of circulation of certain groups of drugs containing strains of microorganisms, in particular in vaccination, attract the attention of many researchers. They were studied, in particular by: R.A. Maidanyk, I.Ya. Senyuta, etc. However, the peculiarities of activities in the field of particularly dangerous microorganisms’ strains treatment have been left without due attention of researchers, including specialists in medical law. Formulating Goals. The aim of the work is to study the state of relations in the field of especially dangerous microorganisms’ strains treatment legal regulation in Ukraine, analysis of the current legislation of Ukraine and the practice of its application in this area. There have been used scientific publications of leading experts and current Ukrainian legislation. The research is based on an organic combination of general scientific and special legal research methods. Presenting main material. The main sources of biological threats are: 1) epidemics and outbreaks of infectious human diseases; 2) epizootics (high incidence among animals); 3) epitophytia (spread of infectious plant disease in large areas); 4) accidents at biologically dangerous objects; 5) natural reservoirs of pathogenic microorganisms; 6) transboundary transfer of pathogenic microorganisms, representatives of flora and fauna, dangerous for ecological systems; 7) sabotage at biologically dangerous objects; 8) biological terrorism; 9) the use of biological weapons by the state. It should be noted that the regulatory framework in the field of particularly dangerous microorganisms’ strains treatment in the context of biosafety in Ukraine is fragmentary. Although the list of laws and other regulations governing biosafety and/or biosecurity in Ukraine is impressive. Nowadays, the use of strains of microorganisms is gaining popularity, in particular in agriculture, because they can be used for the needs of veterinary medicine and for the production of certain foods such as yogurt, kefir etc. Therefore, in addition to the basic law, regulation of the use of strains and protection of rights to them is regulated by other acts, in particular the Law of Ukraine “On protection of rights to inventions and utility models”. Namely, in accordance with Part 2 of Art. 6 strains of microorganisms that have been bred or would be bred shall be considered as the objects of the invention. It follows that the owner who invented the strain must certify the authorship and the right to obtain a patent or declaratory patent. However, this procedure, unlike others, is complicated. After all, to obtain a full patent, you need to conduct an appropriate qualification examination, which would establish whether the strain meets the conditions of patentability. Also, in addition to filing an application for the invention of a utility model and obtaining a patent, in accordance with the Law of Ukraine “On Veterinary Medicine” when registering a domestic veterinary immunobiological agent, the applicant must deposit strains of microorganisms in a special collection – depository. There are currently three national depositories in Ukraine, each of which specializes in a specific type: non-pathogenic strains; pathogenic to humans; pathogenic to animals. The procedure of depositing strains of microorganisms is carried out in accordance with the Instruction on the procedure of depositing strains of microorganisms in Ukraine for the purpose of patent procedure, approved by the order № 106/115 of the State Patent and the National Academy of Sciences of Ukraine of 26.06.1995. Moreover, in accordance with the Regulations on the National Center for Microorganism Strains and the procedure for depositing microorganism strains, approved by the Cabinet of Ministers of Ukraine regulation № 637 of 07.05.1998, deposit of microorganism strains in Ukraine is carried out by the National Center for Microorganism Strains. Its task is to preserve the production and control of strains of microorganisms, maintain biotechnological indicators, control the state of their population, as well as the preparation of new strains. It is necessary to pay attention to the decision of the National Security and Defense Council of Ukraine “On Biosafety of Ukraine”, which states that at the moment there is an increase in the negative impact of various biological factors on the population, which may lead to threats of biological origin. The reasons for such plural regulation are: 1) the lack of a program on biosafety and prevention of biological terrorism, no national system of counteraction to possible biothreats; 2) no automated and integrated data banks on possible threats of biological and chemical origin. Funding and logistics for laboratories also remain unsatisfactory. Also, the state supervision and control in the field of biosafety is weakened, namely when the owner of high-risk facilities changes or when there is a risk of unauthorized access to laboratories due to imperfect protection of pathogenic microorganisms and strains of dangerous and especially dangerous infectious diseases, which in turn can lead to the leakage of pathogenic microorganisms into the environment and cause mass infectious disease. In addition, there is a legal gap in the legislation of Ukraine regarding the location and control of viral and biological laboratories by foreign states. However, in almost all European countries, as well as in the United States, domestic law prohibits the placement of such laboratories in these countries, because they are potentially dangerous to the population. The danger behind viral and biological laboratories, even with the strictest observance of all necessary safety rules, is extremely great, because the pathogenic microorganisms of human and animal origin in them are considered potential agents of biological weapons. Although Ukraine ratified the Convention on the Prohibition of the Development, Production and Stockpiling of Bacteriological (Biological) and Toxin Weapons and on their Destruction of 10 April, 1972, an agreement was signed in 2005 between the US Department of Defense and the Ministry of Health of Ukraine (expired on 31.05.2013) regarding cooperation in the field of prevention of the spread of technologies, pathogens and knowledge that can be used in the development of biological weapons and the US Department of Defense has begun construction of its objects on the territory of our country. At the same time, microbiological laboratories and production facilities are considered to be areas of the highest biological risk. Infecting of persons while working with microorganisms in laboratories is noted throughout the existence of microbiology and is considered as an indisputable confirmation of occupational hazards. In general, there is a wide variety of ways of potential attacks involving bio-toxic weapons and there are several ways to assimilate biotechnologies with their transformation into weapons due to: 1) use of various agents (e.g., bacteria, viruses, fungi, toxins, bioregulators); 2) use against various aims (humans, animals and plants); 3) different scales of application (tactical, strategic); 4) use for various purposes (open or covert war, murder, terrorism or criminal activity). Probably because of this, in 2012 the WHO adopted the Framework Strategy for Laboratory Biorisk Management for 2012-2016, aimed at creating sustainable global, regional and national plans for biological risk management in laboratories. It recognizes that “In accordance with the International Health Regulations (IHR (2005), all participating countries have made a legal commitment to evaluate, develop and maintain their national key oversight, evaluation and response functions”. Conclusions. The current state of legal regulation of relations in the field of strain management requires, given the intersectoral nature of the issue, consideration of the creation of a single intersectoral body for the supervision of hazardous biological objects. Such a body may not only be a licensing body, but also responsible for the introduction of a register of owners of hazardous biological objects that are not economic entities (for example, individual owners of collections of such biological objects). In this paper, it is impractical to consider the circulation of products with GMOs, on the one hand, this issue is perfectly regulated in other special regulations, on the other hand, today there is an issue concerning the treatment of dangerous strains of pathogenic viruses, bacteria and other microorganisms and toxins, as well as poisons of animal and plant origin. For entities engaged in economic activities with pathogenic microorganisms and strains of dangerous infectious diseases, it is advisable to introduce licensing of this type of activity, which requires further development of a bylaw on licensing conditions. For other persons who are the owners of such biological objects and who are not engaged in economic activities, it is necessary to introduce other forms of permit, including special requirements for their handling and storage conditions. The next important bylaw should be the Regulations on the state register of pathogenic microorganisms and strains of dangerous and especially dangerous infectious diseases. Particular attention should be paid to the disposal of hazardous biological objects. Nowadays, for example, there is no effective system for the disposal of vaccines and other immunobiological drugs that have expired (for reference, the shelf life of the flu vaccine is 7-8 months). The cost of recycling is not always profitable for pharmaceutical operators. There are cases when such drugs are falsified by replacing the expiration date, which adversely affect the health of patients. In this context, it can also be mentioned the fact of importing humanitarian aid in Ukraine in the form of a vaccine against measles and rubella (from the Ukrainian diaspora in Canada). The storage temperature of this vaccine was up to minus 48 degrees. After importation into the customs territory of Ukraine, given the shortcomings of customs legislation and the lack of effective control, the temperature regime was not observed. However, the Ministry of Health still carried out compulsory vaccination of children (there were cases of deteriorating health), which is not only negative for the health of children, but also for the very idea of vaccination. In this case, the Ministry of Health, given the interest, was not able to make the right and lawful decision. This is why an independent intersectoral body of state control (supervision) is needed.

Environments containing significant concentration of NaCl such as salt lakes harbor extremophiles microorganisms which have a great biotechnology interest. To explore the diversity of Bacteria in Chott Tinsilt (Algeria), an isolation program was performed. Water samples were collected from the saltern during the pre-salt harvesting phase. This Chott is high in salt (22.47% (w/v). Seven halophiles Bacteria were selected for further characterization. The isolated strains were able to grow optimally in media with 10–25% (w/v) total salts. Molecular identification of the isolates was performed by sequencing the 16S rRNA gene. It showed that these cultured isolates included members belonging to the Halomonas, Staphylococcus, Salinivibrio, Planococcus and Halobacillus genera with less than 98% of similarity with their closest phylogenetic relative. The halophilic bacterial isolates were also characterized for the production of biosurfactant and industrially important enzymes. Most isolates produced hydrolases and biosurfactants at high salt concentration. In fact, this is the first report on bacterial strains (A4 and B4) which were a good biosurfactant and coagulase producer at 20% and 25% ((w/v)) NaCl. In addition, the biosurfactant produced by the strain B4 at high salinity (25%) was also stable at high temperature (30-100°C) and high alkalinity (pH 11).Key word: Salt Lake, Bacteria, biosurfactant, Chott, halophiles, hydrolases, 16S rRNAINTRODUCTIONSaline lakes cover approximately 10% of the Earth’s surface area. The microbial populations of many hypersaline environments have already been studied in different geographical regions such as Great Salt Lake (USA), Dead Sea (Israel), Wadi Natrun Lake (Egypt), Lake Magadi (Kenya), Soda Lake (Antarctica) and Big Soda Lake and Mono Lake (California). Hypersaline regions differ from each other in terms of geographical location, salt concentration and chemical composition, which determine the nature of inhabitant microorganisms (Gupta et al., 2015). Then low taxonomic diversity is common to all these saline environments (Oren et al., 1993). Halophiles are found in nearly all major microbial clades, including prokaryotic (Bacteria and Archaea) and eukaryotic forms (DasSarma and Arora, 2001). They are classified as slight halophiles when they grow optimally at 0.2–0.85 M (2–5%) NaCl, as moderate halophiles when they grow at 0.85–3.4 M (5–20%) NaCl, and as extreme halophiles when they grow at 3.4–5.1 M (20–30%) NaCl. Hyper saline environments are inhabited by extremely halophilic and halotolerant microorganisms such as Halobacillus sp, Halobacterium sp., Haloarcula sp., Salinibacter ruber , Haloferax sp and Bacillus spp. (Solomon and Viswalingam, 2013). There is a tremendous demand for halophilic bacteria due to their biotechnological importance as sources of halophilic enzymes. Enzymes derived from halophiles are endowed with unique structural features and catalytic power to sustain the metabolic and physiological processes under high salt conditions. Some of these enzymes have been reported to be active and stable under more than one extreme condition (Karan and Khare, 2010). Applications are being considered in a range of industries such as food processing, washing, biosynthetic processes and environmental bioremediation. Halophilic proteases are widely used in the detergent and food industries (DasSarma and Arora, 2001). However, esterases and lipases have also been useful in laundry detergents for the removal of oil stains and are widely used as biocatalysts because of their ability to produce pure compounds. Likewise, amylases are used industrially in the first step of the production of high fructose corn syrup (hydrolysis of corn starch). They are also used in the textile industry in the de-sizing process and added to laundry detergents. Furthermore, for the environmental applications, the use of halophiles for bioremediation and biodegradation of various materials from industrial effluents to soil contaminants and accidental spills are being widely explored. In addition to enzymes, halophilic / halotolerants microorganisms living in saline environments, offer another potential applications in various fields of biotechnology like the production of biosurfactant. Biosurfactants are amphiphilic compounds synthesized from plants and microorganisms. They reduce surface tension and interfacial tension between individual molecules at the surface and interface respectively (Akbari et al., 2018). Comparing to the chemical surfactant, biosurfactant are promising alternative molecules due to their low toxicity, high biodegradability, environmental capability, mild production conditions, lower critical micelle concentration, higher selectivity, availability of resources and ability to function in wide ranges of pH, temperature and salinity (Rocha et al., 1992). They are used in various industries which include pharmaceuticals, petroleum, food, detergents, cosmetics, paints, paper products and water treatment (Akbari et al., 2018). The search for biosurfactants in extremophiles is particularly promising since these biomolecules can adapt and be stable in the harsh environments in which they are to be applied in biotechnology.OBJECTIVESEastern Algeria features numerous ecosystems including hypersaline environments, which are an important source of salt for food. The microbial diversity in Chott Tinsilt, a shallow Salt Lake with more than 200g/L salt concentration and a superficies of 2.154 Ha, has never yet been studied. The purpose of this research was to chemically analyse water samples collected from the Chott, isolate novel extremely or moderate halophilic Bacteria, and examine their phenotypic and phylogenetic characteristics with a view to screening for biosurfactants and enzymes of industrial interest.MATERIALS AND METHODSStudy area: The area is at 5 km of the Commune of Souk-Naâmane and 17 km in the South of the town of Aïn-Melila. This area skirts the trunk road 3 serving Constantine and Batna and the railway Constantine-Biskra. It is part the administrative jurisdiction of the Wilaya of Oum El Bouaghi. The Chott belongs to the wetlands of the High Plains of Constantine with a depth varying rather regularly without never exceeding 0.5 meter. Its length extends on 4 km with a width of 2.5 km (figure 1).Water samples and physico-chemical analysis: In February 2013, water samples were collected from various places at the Chott Tinsilt using Global Positioning System (GPS) coordinates of 35°53’14” N lat. and 06°28’44”E long. Samples were collected randomly in sterile polythene bags and transported immediately to the laboratory for isolation of halophilic microorganisms. All samples were treated within 24 h after collection. Temperature, pH and salinity were measured in situ using a multi-parameter probe (Hanna Instruments, Smithfield, RI, USA). The analytical methods used in this study to measure ions concentration (Ca2+, Mg2+, Fe2+, Na+, K+, Cl−, HCO3−, SO42−) were based on 4500-S-2 F standard methods described elsewhere (Association et al., 1920).Isolation of halophilic bacteria from water sample: The media (M1) used in the present study contain (g/L): 2.0 g of KCl, 100.0/200.0 g of NaCl, 1.0 g of MgSO4.7HO2, 3.0 g of Sodium Citrate, 0.36 g of MnCl2, 10.0 g of yeast extract and 15.0 g agar. The pH was adjusted to 8.0. Different dilutions of water samples were added to the above medium and incubated at 30°C during 2–7 days or more depending on growth. Appearance and growth of halophilic bacteria were monitored regularly. The growth was diluted 10 times and plated on complete medium agar (g/L): glucose 10.0; peptone 5.0; yeast extract 5.0; KH2PO4 5.0; agar 30.0; and NaCl 100.0/200.0. Resultant colonies were purified by repeated streaking on complete media agar. The pure cultures were preserved in 20% glycerol vials and stored at −80°C for long-term preservation.Biochemical characterisation of halophilic bacterial isolates: Bacterial isolates were studied for Gram’s reaction, cell morphology and pigmentation. Enzymatic assays (catalase, oxidase, nitrate reductase and urease), and assays for fermentation of lactose and mannitol were done as described by Smibert (1994).Optimization of growth conditions: Temperature, pH, and salt concentration were optimized for the growth of halophilic bacterial isolates. These growth parameters were studied quantitatively by growing the bacterial isolates in M1 medium with shaking at 200 rpm and measuring the cell density at 600 nm after 8 days of incubation. To study the effect of NaCl on the growth, bacterial isolates were inoculated on M1 medium supplemented with different concentration of NaCl: 1%-35% (w/v). The effect of pH on the growth of halophilic bacterial strains was studied by inoculating isolates on above described growth media containing NaCl and adjusted to acidic pH of 5 and 6 by using 1N HCl and alkaline pH of 8, 9, 10, 11 and 12 using 5N NaOH. The effect of temperature was studied by culturing the bacterial isolates in M1 medium at different temperatures of incubation (4°C–55°C).Screening of halophilic bacteria for hydrolytic enzymes: Hydrolase producing bacteria among the isolates were screened by plate assay on starch, tributyrin, gelatin and DNA agar plates respectively for amylase, lipase, protease and DNAse activities. Amylolytic activity of the cultures was screened on starch nutrient agar plates containing g/L: starch 10.0; peptone 5.0; yeast extract 3.0; agar 30.0; NaCl 100.0/250.0. The pH was 7.0. After incubation at 30 ºC for 7 days, the zone of clearance was determined by flooding the plates with iodine solution. The potential amylase producers were selected based on ratio of zone of clearance diameter to colony diameter. Lipase activity of the cultures was screened on tributyrin nutrient agar plates containing 1% (v/v) of tributyrin. Isolates that showed clear zones of tributyrin hydrolysis were identified as lipase producing bacteria. Proteolytic activity of the isolates was similarly screened on gelatin nutrient agar plates containing 10.0 g/L of gelatin. The isolates showing zones of gelatin clearance upon treatment with acidic mercuric chloride were selected and designated as protease producing bacteria. The presence of DNAse activity on plates was determined on DNAse test agar (BBL) containing 10%-25% (w/v) total salt. After incubation for 7days, the plates were flooded with 1N HCl solution. Clear halos around the colonies indicated DNAse activity (Jeffries et al., 1957).Milk clotting activity (coagulase activity) of the isolates was also determined following the procedure described (Berridge, 1952). Skim milk powder was reconstituted in 10 mM aqueous CaCl2 (pH 6.5) to a final concentration of 0.12 kg/L. Enzyme extracts were added at a rate of 0.1 mL per mL of milk. The coagulation point was determined by manual rotating of the test tube periodically, at short time intervals, and checking for visible clot formation.Screening of halophilic bacteria for biosurfactant production. Oil spread Assay: The Petridis base was filled with 50 mL of distilled water. On the water surface, 20μL of diesel and 10μl of culture were added respectively. The culture was introduced at different spots on the diesel, which is coated on the water surface. The occurrence of a clear zone was an indicator of positive result (Morikawa et al., 2000). The diameter of the oil expelling circles was measured by slide caliber (with a degree of accuracy of 0.02 mm).Surface tension and emulsification index (E24): Isolates were cultivated at 30 °C for 7 days on the enrichment medium containing 10-25% NaCl and diesel oil as the sole carbon source. The medium was centrifuged (7000 rpm for 20 min) and the surface tension of the cell-free culture broth was measured with a TS90000 surface tensiometer (Nima, Coventry, England) as a qualitative indicator of biosurfactant production. The culture broth was collected with a Pasteur pipette to remove the non-emulsified hydrocarbons. The emulsifying capacity was evaluated by an emulsification index (E24). The E24 of culture samples was determined by adding 2 mL of diesel oil to the same amount of culture, mixed for 2 min with a vortex, and allowed to stand for 24 h. E24 index is defined as the percentage of height of emulsified layer (mm) divided by the total height of the liquid column (mm).Biosurfactant stability studies : After growth on diesel oil as sole source of carbone, cultures supernatant obtained after centrifugation at 6,000 rpm for 15 min were considered as the source of crude biosurfactant. Its stability was determined by subjecting the culture supernatant to various temperature ranges (30, 40, 50, 60, 70, 80 and 100 °C) for 30 min then cooled to room temperature. Similarly, the effect of different pH (2–11) on the activity of the biosurfactant was tested. The activity of the biosurfactant was investigated by measuring the emulsification index (El-Sersy, 2012).Molecular identification of potential strains. DNA extraction and PCR amplification of 16S rDNA: Total cellular DNA was extracted from strains and purified as described by Sambrook et al. (1989). DNA was purified using Geneclean® Turbo (Q-BIO gene, Carlsbad, CA, USA) before use as a template in polymerase chain reaction (PCR) amplification. For the 16S rDNA gene sequence, the purified DNA was amplified using a universal primer set, forward primer (27f; 5′-AGA GTT TGA TCM TGG CTC AG) and a reverse primer (1492r; 5′-TAC GGY TAC CTT GTT ACG ACT T) (Lane, 1991). Agarose gel electrophoresis confirmed the amplification product as a 1400-bp DNA fragment.16S rDNA sequencing and Phylogenic analysis: Amplicons generated using primer pair 27f-1492r was sequenced using an automatic sequencer system at Macrogene Company (Seoul, Korea). The sequences were compared with those of the NCBI BLAST GenBank nucleotide sequence databases. Phylogenetic trees were constructed by the neighbor-joining method using MEGA version 5.05 software (Tamura et al., 2011). Bootstrap resembling analysis for 1,000 replicates was performed to estimate the confidence of tree topologies.Nucleotide sequence accession numbers: The nucleotide sequences reported in this work have been deposited in the EMBL Nucleotide Sequence Database. The accession numbers are represented in table 5.Statistics: All experiments were conducted in triplicates. Results were evaluated for statistical significance using ANOVA.RESULTSPhysico-chemical parameters of the collected water samples: The physicochemical properties of the collected water samples are reported in table 1. At the time of sampling, the temperature was 10.6°C and pH 7.89. The salinity of the sample, as determined in situ, was 224.70 g/L (22,47% (w/v)). Chemical analysis of water sample indicated that Na +and Cl- were the most abundant ions (table 1). SO4-2 and Mg+2 was present in much smaller amounts compared to Na +and Cl- concentration. Low levels of calcium, potassium and bicarbonate were also detected, often at less than 1 g/L.Characterization of isolates. Morphological and biochemical characteristic feature of halophilic bacterial isolates: Among 52 strains isolated from water of Chott Tinsilt, seven distinct bacteria (A1, A2, A3, A4, B1, B4 and B5) were chosen for further characterization (table 2). The colour of the isolates varied from beige, pale yellow, yellowish and orange. The bacterial isolates A1, A2, A4, B1 and B5 were rod shaped and gram negative (except B5), whereas A3 and B4 were cocci and gram positive. All strains were oxidase and catalase positive except for B1. Nitrate reductase and urease activities were observed in all the bacterial isolates, except B4. All the bacterial isolates were negative for H2S formation. B5 was the only strain positive for mannitol fermentation (table 2).We isolated halophilic bacteria on growth medium with NaCl supplementation at pH 7 and temperature of 30°C. We studied the effect of NaCl, temperature and pH on the growth of bacterial isolates. All the isolates exhibited growth only in the presence of NaCl indicating that these strains are halophilic. The optimum growth of isolates A3 and B1 was observed in the presence of 10% NaCl, whereas it was 15% NaCl for A1, A2 and B5. A4 and B4 showed optimum growth in the presence of 20% and 25% NaCl respectively. A4, B4 and B5 strains can tolerate up to 35% NaCl.The isolate B1 showed growth in medium supplemented with 10% NaCl and pH range of 7–10. The optimum pH for the growth B1 was 9 and they did not show any detectable growth at or below pH 6 (table 2), which indicates the alkaliphilic nature of B1 isolate. The bacterial isolates A1, A2 and A4 exhibited growth in the range of pH 6–10, while A3 and B4 did not show any growth at pH greater than 8. The optimum pH for growth of all strains (except B1) was pH 7.0 (table 2). These results indicate that A1, A2, A3, A4, B4 and B5 are neutrophilic in nature. All the bacterial isolates exhibited optimal growth at 30°C and no detectable growth at 55°C. Also, detectable growth of isolates A1, A2 and A4 was observed at 4°C. However, none of the bacterial strains could grow below 4°C and above 50°C (table 2).Screening of the halophilic enzymes: To characterize the diversity of halophiles able to produce hydrolytic enzymes among the population of microorganisms inhabiting the hypersaline habitats of East Algeria (Chott Tinsilt), a screening was performed. As described in Materials and Methods, samples were plated on solid media containing 10%-25% (w/v) of total salts and different substrates for the detection of amylase, protease, lipase and DNAse activities. However, coagulase activity was determined in liquid medium using milk as substrate (figure 3). Distributions of hydrolytic activity among the isolates are summarized in table 4.From the seven bacterial isolates, four strains A1, A2, A4 and B5 showed combined hydrolytic activities. They were positive for gelatinase, lipase and coagulase. A3 strain showed gelatinase and lipase activities. DNAse activities were detected with A1, A4, B1 and B5 isolates. B4 presented lipase and coagulase activity. Surprisingly, no amylase activity was detected among all the isolates.Screening for biosurfactant producing isolates: Oil spread assay: The results showed that all the strains could produce notable (>4 cm diameter) oil expelling circles (ranging from 4.11 cm to 4.67 cm). The average diameter for strain B5 was 4.67 cm, significantly (P < 0.05) higher than for the other strains.Surface tension and emulsification index (E24): The assimilation of hydrocarbons as the sole sources of carbon by the isolate strains led to the production of biosurfactants indicated by the emulsification index and the lowering of the surface tension of cell-free supernatant. Based on rapid growth on media containing diesel oil as sole carbon source, the seven isolates were tested for biosurfactant production and emulsification activity. The obtained values of the surface tension measurements as well as the emulsification index (E24) are shown in table 3. The highest reduction of surface tension was achieved with B5 and A3 isolates with values of 25.3 mN m−1 and 28.1 mN m−1 respectively. The emulsifying capacity evaluated by the E24 emulsification index was highest in the culture of isolate B4 (78%), B5 (77%) and A3 (76%) as shown in table 3 and figure 2. These emulsions were stable even after 4 months. The bacteria with emulsification indices higher than 50 % and/or reduction in the surface tension (under 30 mN/m) have been defined as potential biosurfactant producers. Based on surface tension and the E24 index results, isolates B5, B4, A3 and A4 are the best candidates for biosurfactant production. It is important to note that, strains B4 and A4 produce biosurfactant in medium containing respectively 25% and 20% (w/v) NaCl.Stability of biosurfactant activities: The applicability of biosurfactants in several biotechnological fields depends on their stability at different environmental conditions (temperatures, pH and NaCl). For this study, the strain B4 appear very interesting (It can produce biosurfactant at 25 % NaCl) and was choosen for futher analysis for biosurfactant stability. The effects of temperature and pH on the biosurfactant production by the strain B4 are shown in figure 4.biosurfactant in medium containing respectively 25% and 20% (w/v) NaCl.Stability of biosurfactant activities: The applicability of biosurfactants in several biotechnological fields depends on their stability at different environmental conditions (temperatures, pH and NaCl). For this study, the strain B4 appear very interesting (It can produce biosurfactant at 25 % NaCl) and was chosen for further analysis for biosurfactant stability. The effects of temperature and pH on the biosurfactant production by the strain B4 are shown in figure 4. The biosurfactant produced by this strain was shown to be thermostable giving an E-24 Index value greater than 78% (figure 4A). Heating of the biosurfactant to 100 °C caused no significant effect on the biosurfactant performance. Therefore, the surface activity of the crude biosurfactant supernatant remained relatively stable to pH changes between pH 6 and 11. At pH 11, the value of E24 showed almost 76% activity, whereas below pH 6 the activity was decreased up to 40% (figure 4A). The decreases of the emulsification activity by decreasing the pH value from basic to an acidic region; may be due to partial precipitation of the biosurfactant. This result indicated that biosurfactant produced by strain B4 show higher stability at alkaline than in acidic conditions.Molecular identification and phylogenies of potential isolates: To identify halophilic bacterial isolates, the 16S rDNA gene was amplified using gene-specific primers. A PCR product of ≈ 1.3 kb was detected in all the seven isolates. The 16S rDNA amplicons of each bacterial isolate was sequenced on both strands using 27F and 1492R primers. The complete nucleotide sequence of 1336,1374, 1377,1313, 1305,1308 and 1273 bp sequences were obtained from A1, A2, A3, A4, B1, B4 and B5 isolates respectively, and subjected to BLAST analysis. The 16S rDNA sequence analysis showed that the isolated strains belong to the genera Halomonas, Staphylococcus, Salinivibrio, Planococcus and Halobacillus as shown in table 5. The halophilic isolates A2 and A4 showed 97% similarity with the Halomonas variabilis strain GSP3 (accession no. AY505527) and the Halomonas sp. M59 (accession no. AM229319), respectively. As for A1, it showed 96% similarity with the Halomonas venusta strain GSP24 (accession no. AY553074). B1 and B4 showed for their part 96% similarity with the Salinivibrio costicola subsp. alcaliphilus strain 18AG DSM4743 (accession no. NR_042255) and the Planococcus citreus (accession no. JX122551), respectively. The bacterial isolate B5 showed 98% sequence similarity with the Halobacillus trueperi (accession no. HG931926), As for A3, it showed only 95% similarity with the Staphylococcus arlettae (accession no. KR047785). The 16S rDNA nucleotide sequences of all the seven halophilic bacterial strains have been submitted to the NCBI GenBank database under the accession number presented in table 5. The phylogenetic association of the isolates is shown in figure 5.DICUSSIONThe physicochemical properties of the collected water samples indicated that this water was relatively neutral (pH 7.89) similar to the Dead Sea and the Great Salt Lake (USA) and in contrast to the more basic lakes such as Lake Wadi Natrun (Egypt) (pH 11) and El Golea Salt Lake (Algeria) (pH 9). The salinity of the sample was 224.70 g/L (22,47% (w/v). This range of salinity (20-30%) for Chott Tinsilt is comparable to a number of well characterized hypersaline ecosystems including both natural and man-made habitats, such as the Great Salt Lake (USA) and solar salterns of Puerto Rico. Thus, Chott Tinsilt is a hypersaline environment, i.e. environments with salt concentrations well above that of seawater. Chemical analysis of water sample indicated that Na +and Cl- were the most abundant ions, as in most hypersaline ecosystems (with some exceptions such as the Dead Sea). These chemical water characteristics were consistent with the previously reported data in other hypersaline ecosystems (DasSarma and Arora, 2001; Oren, 2002; Hacěne et al., 2004). Among 52 strains isolated from this Chott, seven distinct bacteria (A1, A2, A3, A4, B1, B4 and B5) were chosen for phenotypique, genotypique and phylogenetique characterization.The 16S rDNA sequence analysis showed that the isolated strains belong to the genera Halomonas, Staphylococcus, Salinivibrio, Planococcus and Halobacillus. Genera obtained in the present study are commonly occurring in various saline habitats across the globe. Staphylococci have the ability to grow in a wide range of salt concentrations (Graham and Wilkinson, 1992; Morikawa et al., 2009; Roohi et al., 2014). For example, in Pakistan, Staphylococcus strains were isolated from various salt samples during the study conducted by Roohi et al. (2014) and these results agreed with previous reports. Halomonas, halophilic and/or halotolerant Gram-negative bacteria are typically found in saline environments (Kim et al., 2013). The presence of Planococcus and Halobacillus has been reported in studies about hypersaline lakes; like La Sal del Rey (USA) (Phillips et al., 2012) and Great Salt Lake (Spring et al., 1996), respectively. The Salinivibrio costicola was a representative model for studies on osmoregulatory and other physiological mechanisms of moderately halophilic bacteria (Oren, 2006).However, it is interesting to note that all strains shared less than 98.7% identity (the usual species cut-off proposed by Yarza et al. (2014) with their closest phylogenetic relative, suggesting that they could be considered as new species. Phenotypic, genetic and phylogenetic analyses have been suggested for the complete identification of these strains. Theses bacterial strains were tested for the production of industrially important enzymes (Amylase, protease, lipase, DNAse and coagulase). These isolates are good candidates as sources of novel enzymes with biotechnological potential as they can be used in different industrial processes at high salt concentration (up to 25% NaCl for B4). Prominent amylase, lipase, protease and DNAase activities have been reported from different hypersaline environments across the globe; e.g., Spain (Sánchez‐Porro et al., 2003), Iran (Rohban et al., 2009), Tunisia (Baati et al., 2010) and India (Gupta et al., 2016). However, to the best of our knowledge, the coagulase activity has never been detected in extreme halophilic bacteria. Isolation and characterization of crude enzymes (especially coagulase) to investigate their properties and stability are in progress.The finding of novel enzymes with optimal activities at various ranges of salt concentrations is of great importance. Besides being intrinsically stable and active at high salt concentrations, halophilic and halotolerant enzymes offer great opportunities in biotechnological applications, such as environmental bioremediation (marine, oilfiel) and food processing. The bacterial isolates were also characterized for production of biosurfactants by oil-spread assay, measurement of surface tension and emulsification index (E24). There are few reports on biosurfactant producers in hypersaline environments and in recent years, there has been a greater increase in interest and importance in halophilic bacteria for biomolecules (Donio et al., 2013; Sarafin et al., 2014). Halophiles, which have a unique lipid composition, may have an important role to play as surface-active agents. The archae bacterial ether-linked phytanyl membrane lipid of the extremely halophilic bacteria has been shown to have surfactant properties (Post and Collins, 1982). Yakimov et al. (1995) reported the production of biosurfactant by a halotolerant Bacillus licheniformis strain BAS 50 which was able to produce a lipopeptide surfactant when cultured at salinities up to 13% NaCl. From solar salt, Halomonas sp. BS4 and Kocuria marina BS-15 were found to be able to produce biosurfactant when cultured at salinities of 8% and 10% NaCl respectively (Donio et al., 2013; Sarafin et al., 2014). In the present work, strains B4 and A4 produce biosurfactant in medium containing respectively 25% and 20% NaCl. To our knowledge, this is the first report on biosurfactant production by bacteria under such salt concentration. Biosurfactants have a wide variety of industrial and environmental applications (Akbari et al., 2018) but their applicability depends on their stability at different environmental conditions. The strain B4 which can produce biosurfactant at 25% NaCl showed good stability in alkaline pH and at a temperature range of 30°C-100°C. Due to the enormous utilization of biosurfactant in detergent manufacture the choice of alkaline biosurfactant is researched (Elazzazy et al., 2015). On the other hand, the interesting finding was the thermostability of the produced biosurfactant even after heat treatment (100°C for 30 min) which suggests the use of this biosurfactant in industries where heating is of a paramount importance (Khopade et al., 2012). To date, more attention has been focused on biosurfactant producing bacteria under extreme conditions for industrial and commercial usefulness. In fact, the biosurfactant produce by strain B4 have promising usefulness in pharmaceutical, cosmetics and food industries and for bioremediation in marine environment and Microbial enhanced oil recovery (MEOR) where the salinity, temperature and pH are high.CONCLUSIONThis is the first study on the culturable halophilic bacteria community inhabiting Chott Tinsilt in Eastern Algeria. Different genera of halotolerant bacteria with different phylogeneticaly characteristics have been isolated from this Chott. Culturing of bacteria and their molecular analysis provides an opportunity to have a wide range of cultured microorganisms from extreme habitats like hypersaline environments. Enzymes produced by halophilic bacteria show interesting properties like their ability to remain functional in extreme conditions, such as high temperatures, wide range of pH, and high salt concentrations. These enzymes have great economical potential in industrial, agricultural, chemical, pharmaceutical, and biotechnological applications. Thus, the halophiles isolated from Chott Tinsilt offer an important potential for application in microbial and enzyme biotechnology. In addition, these halo bacterial biosurfactants producers isolated from this Chott will help to develop more valuable eco-friendly products to the pharmacological and food industries and will be usefulness for bioremediation in marine environment and petroleum industry.ACKNOWLEDGMENTSOur thanks to Professor Abdelhamid Zoubir for proofreading the English composition of the present paper.CONFLICT OF INTERESTThe authors declare that they have no conflict of interest.Akbari, S., N. H. Abdurahman, R. M. Yunus, F. Fayaz and O. R. Alara, 2018. Biosurfactants—a new frontier for social and environmental safety: A mini review. Biotechnology research innovation, 2(1): 81-90.Association, A. P. H., A. W. W. Association, W. P. C. 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The quantity of municipal solid waste (MSW) generation is escalating at an alarming rate with every passing year alongside the modernization of our economy. Unfortunately, the majority of this waste remains uncollected or ends up in open dumping and followed by uncontrolled burning. Citing the deep-rooted consequences, open dumping should be absolutely abandoned and scientific interventions should be aggressively exercised to reclaim the municipal brownfields. The present research work undertook the judicial task of assessing the comparative feasibility of biomining and scientific capping as a technology selection for reclamation of about a decade old 120 million tons of waste chunk laying at Jawahar Nagar dump yard. Primary dump samples were collected from various locations, considering depth as a variable. While leachate and groundwater samples were collected from Malkaram lake and preinstalled borewells receptively. Additionally, the ambient air quality and noise level also been ascertained within the buffer zone. The blended representative solid sample was segregated using a 70 mm mesh size trommel into organic and inorganic fractions. The organic fraction was composted using a lab-scale aerobic static pile composting (ASPC) while the trommel reject was processed as refuse derived fuel (RDF). Evidently, the compost lagged quality and depicted nutrient deficiency. While the burning of RDF produced siloxane gas, significantly due to elevated silicon level in the primary waste. Furthermore, due to the prolonged leaching tenure and seasonal dilution, the concentration of legacy leachate was relatively weaker. Borewell samples collected from a depth of 20 feet also portrayed minor contamination up to 500 meters horizontal radius. The issue of leachability can solely be resolved with the capping of the existing dump and the end product quality derived from the biomining process is highly questionable. Thus, handling such large quantity capping is a befitting option over biomining for Jawahar Nagar dumpsite. INTRODUCTION Presently, in India due to rapid urbanization and industrialization, the generation of MSW has been increasing tremendously and also expected to continue a similar trend in the future (Scott, 1995; Bhat et al., 2017; Sethurajan et al., 2018; Sharma et al., 2018). Annually, the comprehensive urban MSW generation in India is more than 62 million tons. Metro cities are the mammoth contributor of the entire chunk and waste production had already reached an alarming figure of 50,000 tonnes/day. While the waste generation from the tier 2 cities is also rigorously escalating and presently contribute up to 20,000 tones/day (Sharma et al., 2018). A study conducted by the central pollution control board (CPCB) revealed MSW generation in India is increasing at a distressing rate of 5 % per annum with a sharp escalation in the quantities of domestic hazardous waste (Sharma et al., 2018). With major financial constraints, inefficacy of collection, treatment, and disposal incurs further reasons to worry. So far India has miserably failed to set up wholesome source segregation and collection method. Presently, the country spends more than 60% of its annual waste management budget only in collection. Besides, only 20% or less of the collected materials are scientifically handled and treated. Citing the statistics, it is evident that the majority of the MSW is simply gets dumped on the low laying grounds located somewhere on the outskirts of the cities. The precipitation, infiltration, surface water runoff, bird menace, rodent interference etc. triggers the vulnerability of waste and leads to mal odor, ground and surface water contamination, human and environmental health deterioration (Jayawardhana et al., 2016). Further, the perseverance of the inorganic and inert fractions leads to soil contamination, poses a fire threat, and also may incur carcinogenicity and acute toxicity among the animals (Mir et al., 2021). There are numerous techniques for the reclamation and remediation of the dumpsites, includes processes such as capping and closure, in-situ vitrification, sub-surface cut-off walls, and waste biomining (Chakrabarti and Dubey, 2015; Thakare and Nandi, 2016). Waste biomining is a stable way to get rid of the entire range of problems associated with open dumping and reclaim valuable land (Kaksonen et al., 2017). There are several instances including reclamation of Mumbai Gorai dump yard by IL & FS Environment, 70 – 80 years old 12,00,000 tons of dump clearance by Nagar Nigam Indore within a minute span of 3 years and many more. But the process of biomining is highly sensitive and case-specific. The success of the process solely depends on factors such as characteristics of the waste, efficacy of the effective microorganism culture, acceptability of the processed end product at the local market etc. (Jerez, 2017; Banerjee et al., 2017; Venkiteela, 2020). Contrarily, though the scientific capping is not an end-to-end solution but still advisable in the cases where the quantity of waste is gigantic, land scarcity is prevalent, no nearby industries to consume the end products etc. Mehta et al. (2018) have also supported the above claim based on the assessment of locations specific MSW dump reclamation case studies. While in another Nagpur-based case study conducted by Ashootosh et al. (2020) reported the superiority of the biominingprocess over simple land capping due to the favorability of the local conditions. Capping eliminates the environmental interference and thereby reduces biosphere contamination and leachate generation. Further, it captivates rodent and vector breeding and thereby curtails the spreading of communicable diseases and improves aesthetics. But right consolidation through compaction and execution is utmost necessary in the above case. As non-compaction and faulty sloping will easily lead to heavy settlement and slope failure (Berkun et al., 2005; Al-Ghouti et al., 2021). The present study has been pursued with the primary objective to run a techno-commercial assessment between scientific capping and biomining. While the secondary objective was to ascertain the level of contamination and propose mitigative measures. MATERIALS AND METHODStudy Area Spanning over 350 acres of a precious piece of land at the outskirts of Hyderabad city, Jawahar Nagar dumping yard was brutally utilized by the Greater Hyderabad Municipal Corporation (GHMC) for open dumping for a prolonged tenure of 10 years. It housed nearly 12 lakh metric tons of heterogeneous solid and domestic hazardous waste and continues polluting until 2015, until the Ramky group was offered to cap the legacy dumping and scientifically handle the site. The present study has been facilitated at Hyderabad Municipal Solid Waste Limited, formerly known as Jawahar Nagar dump yard to analyze and assess the feasibility of bio-mining as handling and management alternate to the existing practice of scientific capping. The epicenter of processing and disposal facility is lying approximately on the cross-section of 17°31'24.45"N and 78°35'23.37"E. As per the contract, the comprehensive legacy dumping to be capped in three phases over about 150 acres of area and Ramky has significantly entered the phase two of the operation only within a span of five years by successfully capping more than half of the legacy footprint. Sampling Methodology The waste pile was divided into three layers namely, base, middle, and top. A uniform amount of sample was collected from the successive layers of all five different corners which cover north, south, east, west, and central of the garbage pile. Sampling inspections were performed using a manual auger besides large samples were collected using a JCB excavator. The top six-inch layer of the pile was removed to avoid any contamination while collecting the samples and 5-10 kg of sample was collected from each of the locations. Further, intermediate and bottom layer samples were collected by digging a 500 mm diameter hole through the heap. A composite was prepared by a homogenized blending of all the fifteen grub samples. The blend was distributed into four equal quadrants and the top and bottom quadrants were eliminated diagonally while the left-over quadrants were mixed thoroughly. This process was repeated until a sample of the required bulk of 20 kg is obtained. Surface and subsurface water samples from borewell were collected in and around the facility. Piezometric monitoring borewells located near the landfills were utilized for the subsurface sample collection. While a rainwater pond turned leachate lake named Malkaram was determined as the primary source for leachate collection. Buffer samples were collected from Ambedkar Nagar, the nearby colony exiting at a distance of only 300 meters. Lab-scale Experimentation The representative sample was characterized for composition and further screened through a 70 mm mesh size trommel. The trommel permeate was considered as the organic fraction while the reject was mostly inorganics and inert. The organics were subjected to ASPC. The quantity of the air required is arrived using the method delineated below (Figure 1). MSW Pile size: 2m x 0.5m x 0.5m Volume of pile: 0.5 m3 Average Density of MSW: 620 Kg/m3 Weight of pile: 310 Kg Nitrogen required for matured compost: 9300 mg/kg dry : 9300 X 310 mg : 2.88 x 106 mg : 2.88 Kg Total air required: 2.88 x 100/76 [as Nitrogen in air is 76% by weight] : 3.79 Kg of dry air : 3.79/1.225 m3 [@ 15 deg C density of air 1.225 kg/m3] : 3.1 m3 This air is to be supplied for 100 min / day for 0.5 m pile Air flow rate required: 3.1 x 60/100 = 1.86 m3/h (for practical purpose a flowrate of 2 m3/h was maintained). The maturation period was considered as 28 days and post-maturation, the stabilized material was further cured for 24 hours and screened using 12 mm and 4 mm trommel respectively to obtain the desired product quality and particle size. Whereas, the trommel reject was evenly spreader on the copper trays and dried in an oven at 1050C for 2 hours. The dried material was micronized to the size of 50 mm or below using a scissor and inert such as glass, sand, stone etc. were segregated manually (Mohan and Joseph, 2020). Concurrently, a bench-scale capped landfill prototype was built using the below-mentioned procedure to evaluate the factors such as settlement and slope stability. A 30 mm thick low permeable soil was laid on the top of the waste, followed by a 60 mm layer of compacted clay liner (CCL). Each join between successive liner material was closely monitored. A 1.5 mm thick HDPE liner was placed on the top of the CCL. A 285 GSM geotextile membrane was placed as the successive above layer followed by a 15 mm thick drainage media layer. A further layer of geotextile membrane was placed on top of the drainage media for better stabilization, grip, and strength. The top vegetative soil layer of 45 mm thickness was laid off on top of the geotextile media and St. Augustine grass was rooted (Cortellazzo et al., 2020; Ashford et al., 2000). 2.4 Sample Analysis pH, Electrical Conductivity (EC) and Turbidity of the samples were analyzed using pH, EC-TDS, and Nephelometer of Mettler Toledo. The pH meter was calibrated with the buffer solution of 4.0, 7.0 & 9.12 at a controlled temperature. EC-TDS meter was calibrated with 0.1 M KCL having 12.8 mS/cm of conductivity. Nephelometer was calibrated with Formazine solution of 10 & 100 NTU. Total Dissolved Solids (TDS), (mg/L) was performed using the gravimetric method at 1800C in the oven. Titrimetric parameters such as Total Alkalinity as CaCO3 (mg/L), Total Hardness as CaCO3 (mg/L), Chloride as Cl- (mg/L), Calcium as Ca2+ (mg/L), Residual Free Chlorine (RFC), (mg/L) were analyzed using APHA (American Public Health Associations) method, 23rd Edition, 2017. Total Kjeldahl Nitrogen (mg/L) and Ammonical Nitrogen (mg/L) were performed through distillation followed by titration with H2SO4 as a titrant. Sulphide as S2- was done with the Iodometric method after distillation. Each titrimetric parameter was analyzed in triplicate after standardizing the titrant with required reagents and crossed checked by keeping a check standard. Sodium as Na (mg/L) and Potassium as K (mg/L) were performed using Flame Photometer. The photometer was calibrated with different standards from 10 to 100 (mg/L) standard solutions. The leachate sample was diluted enough to get the value within the standard range and cross-checked with check standards at the same time. Chemical Oxygen Demand (COD), (mg/L) was performed using the open reflux method for 2 hours at 1500C in COD Digestor. Biochemical Oxygen Demand (BOD), (mg/L) was performed using the alkali iodide azide method for 3 days. The samples were kept in a BOD incubator at 270C for 3 days. It was kept in duplicate to have a check on quality control. Sulphate was analyzed by the gravimetric method instead of turbidimetric or through UV-Visible spectrophotometer as its concentration was found more than 40 mg/L. Nitrate as NO3- was analyzed after filtration at 220-275 nm, while Hexavalent Chromium as Cr6+ was analyzed at 540 nm in the UV-Vis. Parameters like Cyanide as CN-, Fluoride as F-, and Phenolic Compounds were gone through a distillation process followed by UV-Vis. The distillation process ensures the removal of interferences presents either positive or negative. For the parameters like Total Iron or Ferric Iron, the samples were digested properly with the required reagents on the hot plate before analyzing in UV-Vis. For the metal analysis the water samples were digested at a temperature of 1000C using aqua regia as a media. The samples were digested to one-fourth of the volume on a hot plate. The recommended wavelengths as per APHA 3120 B were selected for each of the metals. The standard graph was plotted for each of the metals before analysis and crossed checked with the check standard at the same time. Parameters such as bulk density and particle size were performed through the certified beaker and sieve. The percentage of moisture content was estimated using the oven by keeping the compost sample for 2 hours at 1050C. C/N ratio was estimated through CHNS analyzer keeping sulfanilamide as a check standard. The analysis was performed by extracting the desired component in the desired solution prescribed in the method followed by converting the same from mg/L to mg/Kg. RESULTS AND DISCUSSION An exhaustive bench-study has been pursued and real-time samples were collected and analyzed for all possible parameters to determine the pros and cons attributed to both processes. The investigation begins by collecting the samples and concluded by impact assessment studies inclusive of the buffer zone. Both solid, liquid, and gaseous samples were precisely investigated to opt for the best solution. A detailed finding of the investigation is summarized below. Primarily, the representative solid sample was characterized through a manual separation process and the results are portrayed in Figure 1. Compost Characterization ASPC of the organic fraction has resulted in a recovery of 46.7% of the initial load. While 53.3% of the influent mass were inert and barely degradable fraction contributes to reject, the rest 4.1% is miscellaneous process loss. The processed compost was extensively analyzed including for metal contamination and the same is tabulated in Table 1. The value of C/N ratio, OC, TN, K2O, P2O5, and NPK evidently portrays the shortcoming in terms of nutrient availability. Though it is highly enriched in organic carbon and thus the same can be effectively utilized as a soil preconditioner. Ayilara et al. (2020) also reported a similar finding, where the city compost sourced from MSW lagged major plant nutrients. RDF Characterization Processed trommel rejects constitute cloth, rexine, leather, jute, paper, plastics, coir and other inert contributed to RDF. The fraction of inert was as high as 37.2% of the overall RDF mass and it mostly constituted glass and sand. The combined weight of sand and glass fragments contributed 73.5% of the total inert, while the rest was stone and small brickbats. The higher level of silicon associated with the presence of glass and sand yielded siloxane and triggered the possibility of kiln corrosion. A detailed RDF analysis report is enclosed in Table 2. The values explicitly portray the quality of RDF is moderately lower and higher salts concentration is extremely prevalent. With relatively lower NCV and such high salt concentration, the above specimen will certainly pose a corrosion threat to the kiln and shall be either neglected as kiln feed or can be utilized after dilution with Grade III RDF quality. Further, such high ash generation will also induct high transportation and landfill charges. Leachate Characterization The Malkaram leachate lake is the end result of prolonged, slow, and steady mixing of the legacy leachate through the existing fissure cracks in the sheath rock bottom profile. Apparently, the concentration of leachate is significantly lower due to the dilution. Samples were analyzed in triplicates and the mean value is tabulated here in Table 3. The metal concertation and rest of the parameter values are well within the secondary treatment influent range, except for TDS. Thus, a modular aerobic biological treatment unit such as moving bed biofilm bioreactor (MBBR) or membrane bioreactor (MBR) would be a well-suited pick. However, a reverse osmosis (RO) system needs to be installed to get rid of the high TDS content. The permeate of RO can be reused back into the system. Whereas, the reject can be converted into dried powder through forced evaporation mechanisms. The higher concentration of salts in RDF collaterally justifies the elevated TDS level in leachate. In a leachate impact assessment study performed by El-Salam and Abu-Zuid (2015) the reported BOD/COD ratio of 0.69 is greater than double the value of 0.301 reported in Table 3. Though the difference in both the values are quite high, it is relatable and justifiable by the huge age difference of the source waste. The primarily characterized data is of a fresh leachate generated from regular MSW, while the later one is from a decade old waste that barely has any unstabilized organic content. Groundwater Contamination The obvious reason for downward leachate infiltration and osmotic movement facilitates groundwater contamination. Both surface and subsurface water samples were collected within the dump yard and the buffer zone and analyzed using the standard methods. The results are portrayed in Table 4. The slightly alkaline pH of the borewell sample is an indication of the ongoing anaerobic process. The dissolved oxygen value of 3.5 mg/L further validates the correlation. Higher TDS and hardness values are self-indicative of elevated salt concentration in source waste. Eventually, the same interfered with the RDF quality. Positively in the case of all the parameters, a successive decrement in pollution concentration has been spotted from dump ground towards the buffer zone. In a similar study conducted by Singh et al. (2016) at Varanasi, Uttar Pradesh the reported concentration of the parameters is significantly higher than reported in Table 4. The basic reason behind variation is the dissimilarities of the local soil profile. The sandy and clay loam soil profile of Varanasi allows a greater rate of percolation and infiltration. While the bottom sheath rock profile at Jawahar Nagar permits the only a minute to little percolation rate. The difference in percolation rate is directly correlated to the concentration levels in this case. Contrarily, Kurakalva et al. (2016) have reported much-elevated pollutant concertation both in ground and surface water for a study conducted at the same site in 2016. The higher concentration is relatable to the fact of the non-closure of the open dump back then. Capping activity had at Jawahar Nagar gained its pace 2018 onwards and capping for the primary section of 70 acres got concluded only during mid of 2019. Due to the decrement in runoff and percolation, the quality of both surface and subsurface water has improved drastically. Impact Assessment The odor and groundwater contamination are two of the primary issues that triggered a massive public agitation initially. The root causes of both the issues are identified as rainwater percolation and anaerobic digestion respectively. Eventually, the completion of the capping process would resolve both the problems effectively. Other non-tangential impacts include nausea; headache; irritation of the eye, nasal cavity, and throat; diarrhoeal diseases; vector-borne disease, cattle toxicity etc. Scientific capping can easily cater as the wholesome solution for all (Cortellazzo et al., 2020). Yu et al. (2018) had performed an extensive study to comprehend the relativity of respiratory sickness and MSW borne air pollution. The study made a couple of dreadful revelations such as gases released due to the anaerobic digestion of MSW such as methane, hydrogen sulphide, and ammonia incur detrimental impact on Lysozyme and secretory immunoglobulin A (SIgA). While SO2 was reported as the lung capacity and functionality reducer. Further, a gender-specific study executed by the same research group revealed, air pollution impacts more severely on male children than the female and retards immune functions. Presently, the area of 351 acres has been developed as Asia’s one of the largest state of the art municipal solid waste processing and disposal facility by Ramky Enviro Engineers Limited. This ensured zero dumping and no further environmental interventions. As legal compliance, the facility monitors the quality of groundwater and ambient air quality in and around the facility on monthly basis to assure the biosafety. The variation in concentration of various monitoring parameters between 2012 to 2020 is summarized in Figure 2. The concentration of each of the parameters are showcased in ppm and a standard equipment error was settled at 3% for respirable dust sampler and multi-gas analyzer (Taheri et al., 2014). Despite all parameter values have gradually increased except for methane, the facility still managed to maintain them well under the regulatory limits. The decrement in methane concentration is directly correlated to the practice of aerobic composting and aeration-based secondary treatment that prevented the formation of the anaerobic atmosphere and henceforth methane generation. While for the rest of the parameters the increment in values is quite substantial and predictable due to the sudden escalation in MSW generation in the past decade in correlation with Gross domestic product (GDP) enhancement. The observed and interpreted impacts due to the elevated pollutant level are in-line with the georeferenced findings reported by Deshmukh and Aher (2016) based on a study conducted at Sangamner, Maharashtra. CONCLUSION The study critically analyzed and investigated every techno-environmental and socio-economic aspect correlated to open dumping. The bench-scale experimentation revealed the efficiency of the single liner scientific capping is fair enough to eliminate any further rainwater infiltration, however, it has no control over the generation of leachate due to the inherent moisture. Internal moisture related issue was anyhow compensated with pertinent compaction prior to dispose of the waste. Contrarily, both the products derived through the biomining process namely, compost and RDF lagged quality due to scantier nutrient content and higher salt and silicon content respectively. Besides, impact assessment studies concede the pollutant concentration in groundwater in and around the plant has drastically diminished post-July 2019 due to the partial completion of waste capping. It also abetted lowering the dust and odor issues relatively in the surrounding. ACKNOWLEDGMENT The authors would like to sincerely acknowledge GHMC, Hyderabad Integrated Municipal Solid Waste Limited, and Ramky Enviro Engineers Limited for enabling us to pursue the sample collection and other necessary onsite activities. Further, the authors would like to register profound acknowledgment to EPTRI for supporting us with the essential experimental facilities. REFERENCES Sharma, A., Gupta, A.K., Ganguly, R. (2018), Impact of open dumping of municipal solid waste on soil properties in mountainous region. 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Hamsters were exposed to sodium arsenite (173 mg As/L) in drinking water for 6 days. Equal amounts of proteins from urinary bladder or liver extracts of control and arsenic-treated hamsters were labeled with Cy3 and Cy5 dyes, respectively. After differential in gel electrophoresis and analysis by the DeCyder software, several protein spots were found to be down-regulated and several were up regulated. Our experiments indicated that in the bladder tissues of hamsters exposed to arsenite, transgelin was down-regulated and GST-pi was up-regulated. The loss of transgelin expression has been reported to be an important early event in tumor progression and a diagnostic marker for cancer development [29-32]. Down-regulation of transgelin expression may be associated with the carcinogenicity of inorganic arsenic in the urinary bladder. In the liver of arsenite-treated hamsters, ornithine aminotransferase was up-regulated, and senescence marker protein 30 and fatty acid binding protein were down-regulated. The volume ratio changes of these proteins in the bladder and liver of hamsters exposed to arsenite were significantly different than that of control hamsters. Introduction Chronic exposure to inorganic arsenic can cause cancer of the skin, lungs, urinary bladder, kidneys, and liver [1-6]. The molecular mechanisms of the carcinogenicity and toxicity of inorganic arsenic are not well understood [7-9). Humans chronically exposed to inorganic arsenic excrete MMA(V), DMA(V) and the more toxic +3 oxidation state arsenic biotransformants MMA(III) and DMA (III) in their urine [10, 11], which are carcinogen [12]· After injection of mice with sodium arsenate, the highest concentrations of the very toxic MMA(III) and DMA(III) were in the kidneys and urinary bladder tissue, respectively, as shown by experiments of Chowdhury et al [13]. Many mechanisms of arsenic toxicity and carcinogenicity have been suggested [1, 7, 14] including chromosome abnormalities [15], oxidative stress [16, 17], altered growth factors [18], cell proliferation [19], altered DNA repair [20], altered DNA methylation patterns [21], inhibition of several key enzymes [22], gene amplification [23] etc. Some of these mechanisms result in alterations in protein expression. Methods for analyzing multiple proteins have advanced greatly in the last several years. In particularly, mass spectrometry (MS) and tandem MS (MS/MS) are used to analyze peptides following protein isolation using two-dimensional (2-D) gel electrophoresis and proteolytic digestion [24]. In the present study, Differential In Gel Electrophoresis (DIGE) coupled with Mass Spectrometry (MS) has been used to study some of the proteomic changes in the urinary bladder and liver of hamsters exposed to sodium arsenite in their drinking water. Our results indicated that transgelin was down-regulated and GST-pi was up-regulated in the bladder tissues. In the liver tissues ornithine aminotransferase was up-regulated, and senescence marker protein 30, and fatty acid binding protein were down-regulated. Materials and Methods Chemicals Tris, Urea, IPG strips, IPG buffer, CHAPS, Dry Strip Cover Fluid, Bind Silane, lodoacetamide, Cy3 and Cy5 were from GE Healthcare (formally known as Amersham Biosciences, Uppsala, Sweden). Thiourea, glycerol, SDS, DTT, and APS were from Sigma-Aldrich (St. Louis, MO, USA). Glycine was from USB (Cleveland, OH, USA). Acrylamide Bis 40% was from Bio-Rad (Hercules, CA, USA). All other chemicals and biochemicals used were of analytical grade. All solutions were made with Milli-Q water. Animals Male hamsters (Golden Syrian), 4 weeks of age, were purchased from Harlan Sprague Dawley, USA. Upon arrival, hamsters were acclimated in the University of Arizona animal care facility for at least 1 week and maintained in an environmentally controlled animal facility operating on a 12-h dark/12-h light cycle and at 22-24°C. They were provided with Teklad (Indianapolis, IN) 4% Mouse/Rat Diet # 7001 and water, ad libitum, throughout the acclimation and experimentation periods. Sample preparation and labelling Hamsters were exposed to sodium arsenite (173 mg) in drinking water for 6 days and the control hamsters were given tap water. On the 6th day hamsters were decapitated rapidly by guillotine. Urinary bladder tissues and liver were removed, blotted on tissue papers (Kimtech Science, Precision Wipes), and weighed. Hamster urinary bladder or liver tissues were homogenized in lysis buffer (30mMTris, 2M thiourea, 7M urea, and 4% w/w CHAPS adjusted to pH 8.5 with dilute HCI), at 4°C using a glass homogenizer and a Teflon coated steel pestle; transferred to a 5 ml acid-washed polypropylene tube, placed on ice and sonicated 3 times for 15 seconds. The sonicate was centrifuged at 12,000 rpm for 10 minutes at 4°C. Small aliquots of the supernatants were stored at -80°C until use (generally within one week). Protein concentration was determined by the method of Bradford [25] using bovine serum albumin as a standard. Fifty micrograms of lysate protein was labeled with 400 pmol of Cy3 Dye (for control homogenate sample) and Cy5 Dye (for arsenic-treated urinary bladder or liver homogenate sample). The samples containing proteins and dyes were incubated for 30 min on ice in the dark. To stop the labeling reaction, 1uL of 10 mM lysine was added followed by incubation for 10 min on ice in the dark. To each of the appropriate dye-labeled protein samples, an additional 200 ug of urinary bladderor liver unlabeled protein from control hamster sample or arsenic-treated hamster sample was added to the appropriate sample. Differentially labeled samples were combined into a single Microfuge tube (total protein 500 ug); protein was mixed with an equal volume of 2x sample buffer [2M thiourea, 7M urea, pH 3-10 pharmalyte for isoelectric focusing 2% (v/v), DTT 2% (w/v), CHAPS 4% (w/v)]; and was incubated on ice in the dark for 10 min. The combined samples containing 500 ug of total protein were mixed with rehydration buffer [CHAPS 4% (w/v), 8M urea, 13mM DTT, IPG buffer (3-10) 1% (v/v) and trace amount of bromophenol blue]. The 450 ul sample containing rehydration buffer was slowly pipetted into the slot of the ImmobilinedryStripReswelling Tray and any large bubbles were removed. The IPG strip (linear pH 3-10, 24 cm) was placed (gel side down) into the slot, covered with drystrip cover fluid (Fig. 1), and the lid of the Reswelling Tray was closed. The ImmobillineDryStrip was allowed to rehydrate at room temperature for 24 hours. First dimension Isoelectric focusing (IEF) The labeled sample was loaded using the cup loading method on universal strip holder. IEF was then carried out on EttanIPGphor II using multistep protocol (6 hr @ 500 V, 6 hr @ 1000 V, 8 hr @ 8000 V). The focused IPG strip was equilibrated in two steps (reduction and alkylation) by equilibrating the strip for 10 min first in 10 ml of 50mM Tris (pH 8.8), 6M urea, 30% (v/v) glycerol, 2% (w/v) SDS, and 0.5% (w/v) DTT, followed by another 10 min in 10 ml of 50mM Tris (pH 8.8), 6M urea, 30% (v/v) glycerol, 2% (w/v) SDS, and 4.5% (w/v) iodoacetamide to prepare it for the second dimension electrophoresis. Second dimension SDS-PAGE The equilibrated IPG strip was used for protein separation by 2D-gel electrophoresis (DIGE). The strip was sealed at the top of the acrylamide gel for the second dimension (vertical) (12.5% polyacrylamide gel, 20x25 cm x 1.5 mm) with 0.5% (w/v) agarose in SDS running buffer [25 mMTris, 192 mM Glycine, and 0.1% (w/v) SDS]. Electrophoresis was performed in an Ettan DALT six electrophoresis unit (Amersham Biosciences) at 1.5 watts per gel, until the tracking dye reached the anodic end of the gel. Image analysis and post-staining The gel then was imaged directly between glass plates on the Typhoon 9410 variable mode imager (Sunnyvale, CA, USA) using optimal excitation/emission wavelength for each DIGE fluor: Cy3 (532/580 nm) and Cy5 (633/670 nm). The DIGE images were previewed and checked with Image Quant software (GE Healthcare) where all the two separate gel images could be viewed as a single gel image. DeCyde v.5.02 was used to analyze the DIGE images as described in the Ettan DIGE User Manual (GE Healthcare). The appropriate up-/down regulated spots were filtered based on an average volume ratio of ± over 1.2 fold. After image acquisition, the gel was fixed overnight in a solution containing 40% ethanol and 10% acetic acid. The fixed gel was stained with SyproRuby (BioRad) according to the manufacturer protocol (Bio-Rad Labs., 2000 Alfred Nobel Drive, Hercules, CA 94547). Identification of proteins by MS Protein spot picking and digestion Sypro Ruby stained gels were imaged using an Investigator ProPic and HT Analyzer software, both from Genomic Solutions (Ann Arbor, MI). Protein spots of interest that matched those imaged using the DIGE Cy3/Cy5 labels were picked robotically, digested using trypsin as described previously [24] and saved for mass spectrometry identification. Liquid chromatography (LC)- MS/MS analysis LC-MS/MS analyses were carried out using a 3D quadrupole ion trap massspectrometer (ThermoFinnigan LCQ DECA XP PLUS; ThermoFinnigan, San Jose, CA) equipped with a Michrom Paradigm MS4 HPLC (MichromBiosources, Auburn, CA) and a nanospray source, or with a linear quadrupole ion trap mass spectrometer (ThermoFinnigan LTQ), also equipped with a Michrom MS4 HPLC and a nanospray source. Peptides were eluted from a 15 cm pulled tip capillary column (100 um I.D. x 360 um O.D.; 3-5 um tip opening) packed with 7 cm Vydac C18 (Vydac, Hesperia, CA) material (5 µm, 300 Å pore size), using a gradient of 0-65% solvent B (98% methanol/2% water/0.5% formic acid/0.01% triflouroacetic acid) over a 60 min period at a flow rate of 350 nL/min. The ESI positive mode spray voltage was set at 1.6 kV, and the capillary temperature was set at 200°C. Dependent data scanning was performed by the Xcalibur v 1.3 software on the LCQ DECA XP+ or v 1.4 on the LTQ [27], with a default charge of 2, an isolation width of 1.5 amu, an activation amplitude of 35%, activation time of 50 msec, and a minimal signal of 10,000 ion counts (100 ion counts on the LTQ). Global dependent data settings were as follows: reject mass width of 1.5 amu, dynamic exclusion enabled, exclusion mass width of 1.5 amu, repeat count of 1, repeat duration of a min, and exclusion duration of 5 min. Scan event series were included one full scan with mass range of 350-2000 Da, followed by 3 dependent MS/MS scans of the most intense ion. Database searching Tandem MS spectra of peptides were analyzed with Turbo SEQUEST, version 3.1 (ThermoFinnigan), a program that allows the correlation of experimental tandem MS data with theoretical spectra generated from known protein sequences. All spectra were searched against the latest version of the non redundant protein database from the National Center for Biotechnology Information (NCBI 2006; at that time, the database contained 3,783,042 entries). Statistical analysis The means and standard error were calculated. The Student's t-test was used to analyze the significance of the difference between the control and arsenite exposed hamsters. P values less than 0.05 were considered significant. The reproducibility was confirmed in separate experiments. Results Analysis of proteins expression After DIGE (Fig. 1), the gel was scanned by a Typhoon Scanner and the relative amount of protein from sample 1 (treated hamster) as compared to sample 2 (control hamster) was determined (Figs. 2, 3). A green spot indicates that the amount of protein from sodium arsenite-treated hamster sample was less than that of the control sample. A red spot indicates that the amount of protein from the sodium arsenite-treated hamster sample was greater than that of the control sample. A yellow spot indicates sodium arsenite-treated hamster and control hamster each had the same amount of that protein. Several protein spots were up-regulated (red) or down-regulated (green) in the urinary bladder samples of hamsters exposed to sodium arsenite (173 mg As/L) for 6 days as compared with the urinary bladder of controls (Fig. 2). In the case of liver, several protein spots were also over-expressed (red) or under-expressed (green) for hamsters exposed to sodium arsenite (173 mg As/L) in drinking water for 6 days (Fig. 3). The urinary bladder samples were collected from the first and second experiments in which hamsters were exposed to sodium arsenite (173 mg As/L) in drinking water for 6 days and the controls were given tap water. The urinary bladder samples from the 1st and 2nd experiments were run 5 times in DIGE gels on different days. The protein expression is shown in Figure 2 and Table 1. The liver samples from the 1st and 2nd experiments were also run 3 times in DIGE gels on different days. The proteins expression were shown in Figure 3 and Table 2. The volume ratio changed of the protein spots in the urinary bladder and liver of hamsters exposed to arsenite were significantly differences than that of the control hamsters (Table 1 and 2). Protein spots identified by LC-MS/MS Bladder The spots of interest were removed from the gel, digested, and their identities were determined by LC-MS/MS (Fig. 2 and Table 1). The spots 1, 2, & 3 from the gel were analyzed and were repeated for the confirmation of the results (experiments; 173 mg As/L). The proteins for the spots 1, 2, and 3 were identified as transgelin, transgelin, and glutathione S-transferase Pi, respectively (Fig. 2). Liver We also identified some of the proteins in the liver samples of hamsters exposed to sodium arsenite (173 mg As/L) in drinking water for 6 days (Fig. 3). The spots 4, 5, & 6 from the gels were analyzed and were repeated for the confirmation of the results. The proteins for the spots 4, 5, and 6 were identified as ornithine aminotransferase, senescence marker protein 30, and fatty acid binding protein, respectively (Fig. 3) Discussion The identification and functional assignment of proteins is helpful for understanding the molecular events involved in disease. Weexposed hamsters to sodium arsenite in drinking water. Controls were given tap water. DIGE coupled with LC-MS/MS was then used to study the proteomic change in arsenite-exposed hamsters. After electrophoresis DeCyder software indicated that several protein spots were down-regulated (green) and several were up-regulated (red). Our overall results as to changes and functions of the proteins we have studied are summarized in Table 3. Bladder In the case of the urinary bladder tissue of hamsters exposed to sodium arsenite (173 mg As/L) in drinking water for 6 days, transgelin was down-regulated and GST-pi was up-regulated. This is the first evidence that transgelin is down-regulated in the bladders of animals exposed to sodium arsenite. Transgelin, which is identical to SM22 or WS3-10, is an actin cross linking/gelling protein found in fibroblasts and smooth muscle [28, 29]. It has been suggested that the loss of transgelin expression may be an important early event in tumor progression and a diagnostic marker for cancer development [30-33]. It may function as a tumor suppressor via inhibition of ARA54 (co-regulator of androgen receptor)-enhanced AR (androgen receptor) function. Loss of transgelin and its suppressor function in prostate cancer might contribute to the progression of prostate cancer [30]. Down-regulation of transgelin occurs in the urinary bladders of rats having bladder outlet obstruction [32]. Ras-dependent and Ras-independent mechanisms can cause the down regulation of transgelin in human breast and colon carcinoma cell lines and patient-derived tumorsamples [33]. Transgelin plays a role in contractility, possibly by affecting the actin content of filaments [34]. In our experiments loss of transgelin expression may be associated or preliminary to bladder cancer due to arsenic exposure. Arsenite is a carcinogen [1]. In our experiments, LC-MS/MS analysis showed that two spots (1 and 2) represent transgelin (Fig. 2 and Table 1). In human colonic neoplasms there is a loss of transgelin expression and the appearance of transgelin isoforms (31). GST-pi protein was up-regulated in the bladders of the hamsters exposed to sodium arsenite. GSTs are a large family of multifunctional enzymes involved in the phase II detoxification of foreign compounds [35]. The most abundant GSTS are the classes alpha, mu, and pi classes [36]. They participate in protection against oxidative stress [37]. GST-omega has arsenic reductase activity [38]. Over-expression of GST-pi has been found in colon cancer tissues [39]. Strong expression of GST-pi also has been found in gastric cancer [40], malignant melanoma [41], lung cancer [42], breast cancer [43] and a range of other human tumors [44]. GST-pi has been up-regulated in transitional cell carcinoma of human urinary bladder [45]. Up-regulation of glutathione – related genes and enzyme activities has been found in cultured human cells by sub lethal concentration of inorganic arsenic [46]. There is evidence that arsenic induces DNA damage via the production of ROS (reactive oxygen species) [47]. GST-pi may be over-expressed in the urinary bladder to protect cells against arsenic-induced oxidative stress. Liver In the livers of hamsters exposed to sodium arsenite, ornithine amino transferase was over-expressed, senescence marker protein 30 was under-expressed, and fatty acid binding protein was under-expressed. Ornithine amino transferase has been found in the mitochondria of many different mammalian tissues, especially liver, kidney, and small intestine [48]. Ornithine amino transferase knockdown inhuman cervical carcinoma and osteosarcoma cells by RNA interference blocks cell division and causes cell death [49]. It has been suggested that ornithine amino transferase has a role in regulating mitotic cell division and it is required for proper spindle assembly in human cancer cells [49]. Senescence marker protein-30 (SMP30) is a unique enzyme that hydrolyzes diisopropylphosphorofluoridate. SMP30, which is expressed mostly in the liver, protects cells against various injuries by stimulating membrane calcium-pump activity [50]. SMP30 acts to protect cells from apoptosis [51]. In addition it protects the liver from toxic agents [52]. The livers of SMP30 knockout mice accumulate phosphatidylethanolamine, cardiolipin, phosphatidyl-choline, phosphatidylserine, and sphingomyelin [53]. Liver fatty acid binding protein (L-FABP) also was down- regulated. Decreased liver fatty acid-binding capacity and altered liver lipid distribution hasbeen reported in mice lacking the L-FABP gene [54]. High levels of saturated, branched-chain fatty acids are deleterious to cells and animals, resulting in lipid accumulation and cytotoxicity. The expression of fatty acid binding proteins (including L-FABP) protected cells against branched-chain saturated fatty acid toxicity [55]. Limitations: we preferred to study the pronounced spots seen in DIGE gels. Other spots were visible but not as pronounced. Because of limited funds, we did not identify these others protein spots. In conclusion, urinary bladders of hamsters exposed to sodium arsenite had a decrease in the expression of transgelin and an increase in the expression of GST-pi protein. Under-expression of transgelin has been found in various cancer systems and may be associated with arsenic carcinogenicity [30-33). Inorganic arsenic exposure has resulted in bladder cancer as has been reported in the past [1]. Over-expression of GST-pi may protect cells against oxidative stress caused by arsenite. In the liver OAT was up regulated and SMP-30 and FABP were down regulated. These proteomic results may be of help to investigators studying arsenic carcinogenicity. The Superfund Basic Research Program NIEHS Grant Number ES 04940 from the National Institute of Environmental Health Sciences supported this work. Additional support for the mass spectrometry analyses was provided by grants from NIWHS ES06694, NCI CA023074 and the BIOS Institute of the University of Arizona. Acknowledgement The Author wants to dedicate this paper to the memory of his former supervisor Dr. H. VaskenAposhian who passed away in September 6, 2019. He was an emeritus professor of the Department of Molecular and Cellular Biology at the University of Arizona. 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ABSTRACT First Person is a series of interviews with the first authors of a selection of papers published in Biology Open, helping early-career researchers promote themselves alongside their papers. Kun Guo is first author on ‘The thermal dependence and molecular basis of physiological color change in Takydromus septentrionalis (Lacertidae)’, published in BiO. Kun conducted the research described in this article while a PhD student in Xiang Ji's lab at Jiangsu Key Laboratory for Biodiversity and Biotechnology, College of Life Sciences, Nanjing Normal University, Jiangsu, China. They are now research assistant in the lab of Xiang Ji at College of Life and Environmental Sciences, Wenzhou University, Zhejiang, China, and Jiangsu Key Laboratory for Biodiversity and Biotechnology, College of Life Sciences, Nanjing Normal University, Nanjing, Jiangsu, China.

In the policy discourses of the Organization for Economic Cooperation and Development (OECD) and European Commission (EC), modern biotechnology and the life sciences are represented as an emerging „bio-economy“ in which the latent value underpinning biological materials and products offers the opportunity for sustainable economic growth. This articulation of modern biotechnology and economic development is an emerging scholarly field producing numerous „bio-concepts.“ Over the last decade or so, there have been a number of attempts to theorize this relationship between biotechnologies and their capitalization. This article highlights some of the underlying ambiguities in these conceptualizations, especially the fetishism about any „bio“-item We offer an alternative view of the bio-economy by rethinking the theoretical importance of several key economic and financial processes.

ABSTRACT Euglenoids (Euglenida) are unicellular flagellates possessing exceptionally wide geographical and ecological distribution. Euglenoids combine a biotechnological potential with a unique position in the eukaryotic tree of life. In large part these microbes owe this success to diverse genetics including secondary endosymbiosis and likely additional sources of genes. Multiple euglenoid species have translational applications and show great promise in production of biofuels, nutraceuticals, bioremediation, cancer treatments and more exotically as robotics design simulators. An absence of reference genomes currently limits these applications, including development of efficient tools for identification of critical factors in regulation, growth or optimization of metabolic pathways. The Euglena International Network (EIN) seeks to provide a forum to overcome these challenges. EIN has agreed specific goals, mobilized scientists, established a clear roadmap (Grand Challenges), connected academic and industry stakeholders and is currently formulating policy and partnership principles to propel these efforts in a coordinated and efficient manner.

Bio‐inspired approaches in materials science and systems chemistry are yielding a variety of stimuli‐responsive and dynamic materials that are gradually changing our everyday life. However, the ability to chemically program these materials to exhibit macroscopic higher‐order behaviours such as self‐assembly, contractility, swarming, taxis, chemical communication, or predator‐prey dynamics remains an ongoing challenge. While still in its infancy, the successful fabrication of bio‐inspired materials displaying higher‐order behaviours not only will help bridging the gap between living and non‐living matter, but it will also contribute to the development of advanced materials for potential applications ranging from tissue engineering and biotechnology, to soft robotics and regenerative medicine. Our Mini‐Review will systematically discuss the higher‐order behaviours developed thus far in bio‐inspired systems, namely (i) polymer networks (ii) microbots, (iii) protocells, and (iv) prototissues. For each system it will provide key examples and highlight how the emergent behaviour could be chemically programmed.

Increasingly, bio-based products made via sugar-powered microbial cell factories and industrial fermentation are reaching the market and presenting themselves as sustainable alternatives to fossil and animal-based products. The sustainability potential of biotechnology, however, has been shown to come with trade-offs and cannot be taken for granted. Shared environmental impact hotspots have been identified across industrial fermentation-based products, including biomass production, energy consumption, and end-of-life fate. Based on both these patterns and our direct experience in preparing for the commercial-scale production of Brewed Protein™, we outline practical considerations for improving the sustainability performance of bio-based products made via industrial fermentation.

Tremendous advances in the field of synthetic biology have been witnessed in multiple areas including life sciences, industrial development, and environmental bio-remediation. However, due to the limitations of human understanding in the code of life, any possible intended or unintended uses of synthetic biology, and other unknown reasons, the development and application of this technology has raised concerns over biosafety, biosecurity, and even cyberbiosecurity that they may expose public health and the environment to unknown hazards. Over the past decades, some countries in Europe, America, and Asia have enacted laws and regulations to control the application of synthetic biology techniques in basic and applied research and this has resulted in some benefits. The outbreak of the COVID-19 caused by novel coronavirus SARS-CoV-2 and various speculations about the origin of this virus have attracted more attention on bio-risk concerns of synthetic biology because of its potential power and uncertainty in the synthesis and engineering of living organisms. Therefore, it is crucial to scrutinize the control measures put in place to ensure appropriate use, promote the development of synthetic biology, and strengthen the governance of pathogen-related research, although the true origin of coronavirus remains hotly debated and unresolved. This article reviews the recent progress made in the field of synthetic biology and combs laws and regulations in governing bio-risk issues. We emphasize the urgent need for legislative and regulatory constraints and oversight to address the biological risks of synthetic biology.

Simulations Plus Announces Preliminary Success in Malaria Drug Design Project. xCELLigence System Evaluated in EU Project to Replace Animal Experiments in Cosmetics Industry. Progress in $100 Million Biodiesel and Commercial Fish Food Project. QIAGEN and Bio-X Center Open Shanghai Translational Medicine Lab. Index Ventures Launches First 150m Life Sciences Fund. Enzo Biochem Expands Distribution with Japan's Cosmo Bio. Maxwell Biotech Venture Fund Invests in Hepatitis B/D. ScinoPharm and NHRI Announce Jointly Developed Diabetes Drug. Calibr: A New Paradigm for Academic – Industry Cooperation. Biocon and Pfizer End Commercialization Agreement. Plandai Biotech Demonstrates Significance of Flavonoid & Polyphenol Bioavailability. Ceva Uses ProBioGen's AGE1.CR Cell Line for Viral Vaccine Production. Quintiles to Sign Memorandum of Agreement with Malaysia Biotechnology Corporation.

In current year, there has been enhancing in the usage of plastic and disposal of waste coming from daily life. Various type of plastics are using for our daily needs, in order to reduce the impact of petroleum based plastics and other waste to be considered and focused on green plastics. Polyethylene are the polymers which can be produced by repeating the single units. It is one of the bittersweet coinage for human's better life. But this coinage causes a lot problems to the environment. To overcome this problem an alternative and eco-friendly Polyethylene is introduced called bio- polyethylene. Bio-polyethylene are from natural biomass sources. Among various source, starch has chief role in production of bio-polyethylene due to its low cost and nature abundance with plasticizers such as glycerol and vinegar. Starch based bio-polyethylene has good mechanical properties, tensile strength and biodegradability. Cellulose also used as source in development of biodegradable polyethylene. In this review starch and cellulose has led to their great innovative uses in food packaging and to improved biodegradation and mechanical properties.

Advances in bottom-up synthetic biology offer the exciting—albeit contentious—prospect of transitioning bio-science researchers from passive observers of life to potential creators of it. Synthetic cells closely emulate the attributes of their biological counterparts. These rationally designed microsystems exhibit emergent properties and life-like functionalities. They can therefore be used as simplified cell models to decipher the rules of life, and as programmable biologically powered micromachines for application in healthcare and biotechnology more broadly. While there is a consensus that current synthetic cells are not yet ‘living’, the question of what defines ‘aliveness’ is gaining increasing relevance. Exploring this concept necessitates a multidisciplinary approach, where scientists from across domains in the physical, life, engineering and social sciences participate in community-level discussions, together with the acceptance of a set of criteria which defines a living system. Achieving a widely accepted definition of ‘living’ represents a possible mission-oriented endpoint to the synthetic cell endeavour, uniting the community towards a common goal. As the field evolves, researchers must address regulatory, ethical, societal and public perception implications, while fostering collaborative efforts to harness the transformative potential of synthetic cells.

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Acrux Enters Commercial Manufacturing Alliance with Orion Corporation Agilent Technologies and University of Technology, Sydney Establish Unique “Elemental Bio-Imaging” Facility DuPont and Hexima Partner to Develop and Commercialize Fungal Disease Resistance Technology Canopus Biopharma and Leading Chinese Researcher Team Up to Treat Avian Influenza All Asia Licensing Announces Proposed Joint Venture with Bio Research Firm for Bovine Testing, Beef Grading Piramal Healthcare Launches French Dermo-cosmetic Range from Pierre Fabre Avesthagen and ICMR Join Hands to Promote Biomedical Research in India ICRISAT and DBT to Establish New Facility for Agri-biotechnology Research Philips Launches Its First Patient Monitoring System “Designed For Emerging Markets” in India Biocon, Abraxis Launch Abraxane in India to Treat Breast Cancer Carna Signs Reagent Supply Agreement with Caliper Life Sciences American Peptide Company to be Acquired by Otsuka Chemical Company EpiCept and GNI Partner to Develop Anti-Viral Hepatitis Drug in Asia USM Signs Nanotechnology Agreement with BiotechCorp AstraZeneca Partners with Singapore Institutions to Develop Anti-Cancer Compounds Metabasis Therapeutics Collaborates with Roche to Develop Liver-Targeted Compounds for the Treatment of Hepatitis C Genelabs Signs Research Agreement with National Health Research Institutes of Taiwan and Genovate Biotech Taiwan Liposome Inks Deal with Japanese Company

Bacteriophage are a class of virus that exclusively infect bacteria cells, with MS2 a group 1 RNA coliphage (affecting the bacteria Escherichia coli). The viral life cycle is comprised of six stages, the first two involving attachment and injection of the RNA genome directly into the host. Lysis is the final stage; including compromise of host cell membrane integrity and release of progeny phage, it is mediated by the lysis, or “L” protein. Consisting of seventy‐five amino acids,(1) L is one of four proteins coded for by the simple MS2 genome, approximately 3500 nucleotides in length.(2) It produces a larger hole in the host cell membrane compared to other bacteriophage lysis proteins and L retains lysis ability, although less efficient, in truncated form.(1) The virus‐free expression of L and the lysis process it elicits is the focus of this study. Toward this goal, a synthetic L‐gene was cloned and ligated to an inducible arabinose operon and RBS using “Bio‐brick” technology. After induction, lysis will be induced and relativity cell density monitored over time via spectrophotometry. A C‐terminally‐fused L‐GFP construct, to track protein localization within the cell and to provide a “handle” for future pull‐down experiments, will also be constructed. Investigation into the role of bacterial host proteins involved in the lysis process has not been previously reported.Support or Funding InformationThis study was graciously funded by Drs. Allen and Judy Freedman. (Hartwick College Freedman Prize ‐ Physical and Life Sciences)

Research Facility at Macquarie University Joins International Proteomic Alliance. Public Databases have Errors and no Quick Fix. Genetic Viability of Australian Grasslands and Heathlands. China Drafts Biotechnology Accord with Brazil. China Calls for Closer Cooperation with US in Biomedical Research. Opportunities in China's Biomedicine Industry. China Aims to Be Leading Pharmacy Processing Center. Geneticist Tsui Lap Chee Appointed as Vice Chancellor of University of Hong Kong. India to Start Growing Pest-resistant Cotton. Health Ministry Confirms Fourth Mad Cow Case. Importance of Japanese Funding to Rice Genomics. Bio Expo Korea 2002 Scheduled for September. Korea Establishes an Arctic Science Base. The Sciences not Popular among Korean Students. Soya Bean the Only GM Food in the Malaysian Market. Fear Prevents AIDS Patients from Getting Cheaper Treatment. Biotech Industry Receives Big Funding. Genome Institute of Singapore Moves to New Facility. Scientist Lays out his Vision for the Life Sciences in Singapore. Industry Conference on Lab Design and Management. Singapore and University of Washington form Bioengineering Alliance. Taiwan Hsinchu Science Park's March Trade Highest in One Year. New Drug for ED Going on Sale Soon. Traders Urged to Market Higher Quality Rice. Hybrid Rice Meeting in Vietnam.

NASA's Rodent Research (RR) project is playing a critical role in advancing biomedical research on the physiological effects of space environments. Due to the limited resources for conducting biological experiments aboard the International Space Station (ISS), it is imperative to use crew time efficiently while maximizing high‐quality science return. NASA's GeneLab project has as its primary objectives to 1) further increase the value of these experiments using a multi‐omics, systems biology‐based approach, and 2) disseminate these data without restrictions to the scientific community. The current investigation assessed viability of RNA, DNA, and protein extracted from archived RR‐1 tissue samples for epigenomic, transcriptomic, and proteomic assays. During the first RR spaceflight experiment, a variety of tissue types were harvested from subjects, snap‐frozen or RNAlater‐preserved, and then stored at least a year at −80°C after return to Earth. They were then prioritized for this investigation based on likelihood of significant scientific value for spaceflight research. All tissues were made available to GeneLab through the bio‐specimen sharing program managed by the Ames Life Science Data Archive and included mouse adrenal glands, quadriceps, gastrocnemius, tibialis anterior, extensor digitorum longus, soleus, eye, and kidney. We report here protocols for and results of these tissue extractions, and thus, the feasibility and value of these kinds of omics analyses. In addition to providing additional opportunities for investigation of spaceflight effects on the mouse transcriptome and proteome in new kinds of tissues, our results may also be of value to program managers for the prioritization of ISS crew time for rodent research activities.Support or Funding InformationSupport from the NASA Space Life and Physical Sciences Division and the International Space Station Program is gratefully acknowledged

Modern food science is providing ever more information on the functions and mechanisms of specific food components in health promotion and/or disease prevention. In response to demands from increasingly health conscious consumers, the global trend is for food industries to translate nutritional information into consumer reality by developing food products that provide not only superior sensory appeal, but also nutritional and health benefits. Today’s busy life styles are also driving the development of healthy convenience foods. Recent innovations in food sciences have led to the use of many traditional technologies, such as fermentation, extraction, encapsulation, fat replacement, and enzyme technology, to produce new health food ingredients, reduce or remove undesirable food components, add specific nutrient or functional ingredients, modify food compositions, mask undesirable flavours or stabilize ingredients. Modern biotechnology has even revolutionized the way foods are created. Recent discoveries in gene science are making it possible to manipulate the components in natural foods. In combination with bio-fermentation, desirable natural compounds can now be produced in large amounts at a low cost and with little environmental impact.In this paper authors analyzed effect new technology on food safety, food quality and food security.

Insulin‐like growth factor‐II (IGF‐II) mRNA‐binding protein‐2 (IMP2) is one of the three homologues (IMP1‐3) that play an important role in the post‐transcriptional regulation of gene expression during development in several tissues including the nervous system. An alternative splice product of IMP2 lacking exon 10 aberrantly expresses in human hepatocellular carcinoma and is identified as HCC. Several lines of studies have demonstrated that IMP1/ZBP1 (zipcode binding protein) is critical in axon guidance and regeneration by regulating localization and translation of specific mRNAs. However, a role of IMP2 in the neural tissue is largely unknown. Previously, we used a custom made IMP2‐specific antibody to locate the endogenous IMP2 expression in mouse neurons and glial cells at different developmental stages. Our pilot study shows that IMP2 expression is sustained throughout life and it may play a role to facilitate axon regeneration. In this study, we used the synapsin promoter‐driven adeno‐associated viral (AAV) 9 constructs to express YFP‐IMP2 and –HCC specifically in neurons both in vitro and in vivo. We applied AAV9.hSyn.YFP.PolyA, AAV9.hSyn.YFP‐hIMP2.PolyA and AAV9.hSyn.YFP‐hHCC.PolyA in the primary dorsal root ganglion mouse neuron culture as an in vitro model. We also injected the viral vectors into the crushed mouse sciatic nerve as an in vivo model. Our results demonstrate the moderate neuron‐specific transgene expression of IMP2/HCC in both models. The data indicate that the synapsin promoter‐driven AAV9 can be used as an important tool for further study of the role of IMP2 in the nervous system.Support or Funding InformationThis work is supported by the Department of Bio‐Medical Sciences, PCOM and Health Research Grant, Pennsylvania Department of Health.

In both recipient and source economies given the mutually beneficial effects Liberalization in APEC that foreign investment should be given high priority this article proposes that. Asia-Pacific and between the region and the rest of the world Facilitating greater foreign investment flows, High growth in the region and Provides an alternative to quality of life. Risk analysis and specifically the strategic risk framework, for investment in Internet capabilities an alternative approach to prioritization is, it is cross-domain and for an analysis of whole-of-government functions Very suitable. Generate insights for full-force capability analysis Cyber risk for a new purpose this article explains the application of risk analysis. Single-valued complex neutrosophic EDAS (estimation based on distance from mean slope) model established and used in green supplier selection. Also, a single-valued complex Neutrosophic EDAS model established and all computational steps are depicted in detail. To examine the use of supplier selection proposed an extended EDAS method. With an intuitive fuzzy number a new EDAS method was proposed and applied. Alternatives are Technological factors (C1), Economic factors (C2), Political and legislative factors (C3), Total costs of investment (C4), Social (personnel) factors (C5), Suppliers (C6) and Ecological (environmental) factors (C7). Evaluation Parameter is biomedical micro electromechanical systems (Bio MEMS) (A1), Nano technology (A2), Biotechnology (A3) and Biomedical engineering (A4). In this type of analysis, EDAS methods determine for the best solution to the negative Short distance and very long distance to settlement, but Comparison of these distances doesn’t underestimate the importance. As a result, Technological factors (C1) is first rank, and Total costs of investment (C4) is lowest rank.

Bio Knowledge Lab (hereinafter BKL) is a young company from Cordoba in the biotechnology and bioinformatics sector specialized in the implementation of massive sequencing projects and computer analysis of biological data. Its objective is to support research groups and companies that work in areas related to life sciences, biotechnology, biomedicine or biopharmaceuticals. In addition, in parallel, BKL also develops its own lines of research. Among them, the project “Bioprospecting of extreme environments of the Andalusian Atlantic coast for the isolation of new bioactive compounds with antitumor activity”, a public-private project carried out in collaboration with the Universities of Huelva and Granada, and aimed to search for new compounds from extremophilic microorganisms. This project can be framed within the area of ​​Drug Discovery, a process by which new drug candidates are discovered. This constitutes the initial stage of the entire development of a drug, a process that is long and expensive, mainly due to the low ratio of drug candidates that can finally be brought to market. For this reason, there are many attempts to innovate in the drug discovery process, for example, through new approaches or using new technologies. Here, omics and computational approaches, as well as the use of new sources of resources, could provide the innovation that this process needs to overcome the public health challenges it faces. In this line, these are the approaches proposed by BKL and its collaborators, all of them integrated into a novel bioprospecting project. The project "Bioprospecting of extreme environments of the Andalusian Atlantic coast for the isolation of new bioactive compounds with antitumor activity" aims to study the microbiodiversity of extreme environments and identify microorganisms of interest, carry out isolations of new microbial species, especially microalgae and archaea, and obtain from them functional extracts rich in bioactive components to evaluate their antioxidant and antitumor capacity. In this first stage of the process, the studied locations have been points of interest of Marismas del Odiel, the Riotinto Mines and the phospho-gypsum pools, all of them in the province of Huelva and characterized by hosting extreme environments, either due to their high salinity, due to its high content of heavy metals and / or its radioactivity. In our project, the microbial, eukaryotic and prokaryotic populations of these environments have been characterized. In addition, two haloarchaeas and four microalgae have been isolated and functional extracts have been obtained, and their antioxidant and antitumor potential has been evaluated, the latter in colorectal cancer models.