Genetically engineered probiotic E. coli Nissle to consume amino acids associated with orphan metabolic diseases

Orphan metabolic diseases are rare genetic defects that interfere with metabolism due to ineffective or missing enzymes. Two of them, Phenylketonuria (PKU) and Maple Syrup Urine Disease (MSUD) are defined by accumulation of amino acids to toxic levels due to defective metabolism of protein break down products. PKU is caused by a defect in the gene encoding phenylalanine hydroxylase (PAH). MSUD is caused by a defect in a multi-enzyme complex found in mitochondria called branched chain ɑ-ketoacid dehydrogenase “BCKDH”. Without the activity of these enzymes, the amino acid phenylalanine (Phe) in the case of PKU or the branched-chain amino acids leucine (Leu), isoleucine and valine for MSUD build up to neurotoxic levels in the blood and brain, leading to neurological deficits. Current treatment options focus on dietary protein restriction, are insufficient and, unfortunately, can lead to a failure to thrive. Lifelong compliance with a prescription diet is also a concern. We have genetically engineered Nissle, a probiotic strain of E. coli, to reduce serum phenylalanine and leucine levels in patients with PKU or MSUD; preclinical data supporting the activity of these strains are described. Please click Additional Files below to see the full abstract

Engineering and manufacturing of probiotic E. Coli to treat metabolic disorder

The fields of synthetic biology and microbiome research developed greatly over the last decade. The convergence of those two disciplines is now enabling the development of new therapeutic strategies, using engineered microbes that operate from within the gut as living medicines. Inborn errors of metabolism represent candidate diseases for these therapeutics, particularly those disorders where a toxic metabolite causing a syndrome is also present in the intestinal lumen. Phenylketonuria (PKU), a rare inherited disease caused by a defect in phenylalanine hydroxylase (PAH) activity, is one such disease and is characterized by the accumulation of systemic phenylalanine (Phe) that can lead to severe neurological deficits unless patients are placed on a strict low-Phe diet. As an alternative treatment, Escherichia coli Nissle (EcN), a well-characterized probiotic, was genetically modified to efficiently import and degrade Phe (SYNB1618). The coupled expression of a Phe transporter with a Phe ammonia lyase (PAL) allows rapid conversion of Phe into trans-cinnamic acid (TCA) in vitro, which is then further metabolized by the host to hippuric acid (HA) and excreted in the urine. Experiments conducted in the enu2-/- PKU mouse model showed that the oral administration of SYNB1618 is able to significantly reduce blood Phe levels triggered by subcutaneous Phe injection. Decreases in circulating Phe levels were associated with proportional increases in urinary HA, confirming that Phe metabolism was caused by the engineered pathway in SYNB1618. Subsequent studies have shown that SYNB1618 is similarly operative in a non-human primate model, providing a translational link to inform future human clinical studies. Consistent with preclinical studies, recent Phase 1/2a clinical data demonstrate that oral administration of SYNB1618 resulted in significant dose-dependent production of biomarkers specifically associated with SYNB1618 activity, demonstrating proof-of-mechanism of this cell therapy

Engineering of probiotic E.coli to treat metabolic disorders

The fields of synthetic biology and microbiome research developed greatly over the last decade. The convergence of those two disciplines is now enabling the development of new therapeutic strategies, using engineered microbes that operate from within the gut as living medicines. Inborn errors of metabolism represent candidate diseases for these therapeutics, particularly those disorders where a toxic metabolite causing a syndrome is also present in the intestinal lumen. Phenylketonuria (PKU), a rare inherited disease caused by a defect in phenylalanine hydroxylase (PAH) activity, is one such disease and is characterized by the accumulation of systemic phenylalanine (Phe) that can lead to severe neurological deficits unless patients are placed on a strict low-Phe diet. As an alternative treatment, Escherichia coli Nissle (EcN), a well-characterized probiotic, was genetically modified to efficiently import and degrade Phe (SYN-PKU). The coupled expression of a Phe transporter with a Phe ammonia lyase (PAL) allows rapid conversion of Phe into trans-cinnamic acid (TCA) in vitro, which is then further metabolized by the host to hippuric acid (HA) and excreted in the urine. Experiments conducted in the enu2-/- PKU mouse model showed that the oral administration of SYN-PKU is able to significantly reduce blood Phe levels triggered by subcutaneous Phe injection. Decreases in circulating Phe levels were associated with proportional increases in urinary HA, confirming that Phe metabolism was caused by the engineered pathway in SYN-PKU. Subsequent studies have shown that SYN-PKU is similarly operative in a non-human primate model, providing a translational link to inform future human clinical studies. In addition to SYN-PKU, a second EcN strain was genetically engineered to rapidly import and degrade branched-chain amino acids (BCAAs) for the treatment of maple syrup urine disease (SYN-MSUD). MSUD, similar to PKU, is a rare genetic disorder caused by a defect in branched-chain ketoacid dehydrogenase activity leading to the toxic accumulation of BCAAs, particularly leucine, and their ketoacid derivatives. The controlled expression in SYN-MSUD of two BCAA transporters, a leucine dehydrogenase, a ketoacid decarboxylase and an alcohol dehydrogenase, result in the efficient degradation of BCAAs into branched-chain alcohols. In a mouse model of MSUD, the oral delivery of SYN-MSUD suppressed the increase in blood BCAAs level induced by a high-protein challenge and prevented the associated moribund phenotype, as measured by locomotor activity. In conclusion, the therapeutic effects observed with SYN-PKU and SYN-MSUD in pre-clinical studies support the further evaluation of engineered microbes as promising approaches for serious inborn errors of metabolism

Revascularization for coronary artery disease in diabetes mellitus: Angioplasty, stents and coronary artery bypass grafting

Author Manuscript: 2011 April 14Patients with diabetes mellitus (DM) are prone to a diffuse and rapidly progressive form of atherosclerosis, which increases their likelihood of requiring revascularization. However, the unique pathophysiology of atherosclerosis in patients with DM modifies the response to arterial injury, with profound clinical consequences for patients undergoing percutaneous coronary intervention (PCI). Multiple studies have shown that DM is a strong risk factor for restenosis following successful balloon angioplasty or coronary stenting, with greater need for repeat revascularization and inferior clinical outcomes. Early data suggest that drug eluting stents reduce restenosis rates and the need for repeat revascularization irrespective of the diabetic state and with no significant reduction in hard clinical endpoints such as myocardial infarction and mortality. For many patients with 1- or 2-vessel coronary artery disease, there is little prognostic benefit from any intervention over optimal medical therapy. PCI with drug-eluting or bare metal stents is appropriate for patients who remain symptomatic with medical therapy. However, selection of the optimal myocardial revascularization strategy for patients with DM and multivessel coronary artery disease is crucial. Randomized trials comparing multivessel PCI with balloon angioplasty or bare metal stents to coronary artery bypass grafting (CABG) consistently demonstrated the superiority of CABG in patients with treated DM. In the setting of diabetes CABG had greater survival, fewer recurrent infarctions or need for re-intervention. Limited data suggests that CABG is superior to multivessel PCI even when drug-eluting stents are used. Several ongoing randomized trials are evaluating the long-term comparative efficacy of PCI with drug-eluting stents and CABG in patients with DM. Only further study will continue to unravel the mechanisms at play and optimal therapy in the face of the profoundly virulent atherosclerotic potential that accompanies diabetes mellitus.National Institutes of Health (U.S.) (GM 49039

Molecular mechanisms of cell death: recommendations of the Nomenclature Committee on Cell Death 2018.

Over the past decade, the Nomenclature Committee on Cell Death (NCCD) has formulated guidelines for the definition and interpretation of cell death from morphological, biochemical, and functional perspectives. Since the field continues to expand and novel mechanisms that orchestrate multiple cell death pathways are unveiled, we propose an updated classification of cell death subroutines focusing on mechanistic and essential (as opposed to correlative and dispensable) aspects of the process. As we provide molecularly oriented definitions of terms including intrinsic apoptosis, extrinsic apoptosis, mitochondrial permeability transition (MPT)-driven necrosis, necroptosis, ferroptosis, pyroptosis, parthanatos, entotic cell death, NETotic cell death, lysosome-dependent cell death, autophagy-dependent cell death, immunogenic cell death, cellular senescence, and mitotic catastrophe, we discuss the utility of neologisms that refer to highly specialized instances of these processes. The mission of the NCCD is to provide a widely accepted nomenclature on cell death in support of the continued development of the field

A sociocultural conversation analysis of Japanese university students' English turn-taking

EThOS - Electronic Theses Online ServiceGBUnited Kingdo