title: In vivo administration of BL-3050: highly stable engineered PON1-HDL complexes.
abstract: BACKGROUND
  Serum paraoxonase (PON1) is a high density lipoprotein (HDL)-associated enzyme involved in organophosphate (OP) degradation and prevention of atherosclerosis. PON1 comprises a potential candidate for in vivo therapeutics, as an anti-atherogenic agent, and for detoxification of pesticides and nerve agents. Because human PON1 exhibits limited stability, engineered, recombinant PON1 (rePON1) variants that were designed for higher reactivity, solubility, stability, and bacterial expression, are candidates for treatment. This work addresses the feasibility of in vivo administration of rePON1, and its HDL complex, as a potentially therapeutic agent dubbed BL-3050.
  
  
  METHODS
  For stability studies we applied different challenges related to the in vivo disfunctionalization of HDL and PON1 and tested for inactivation of PON1's activity. We applied acute, repetitive administrations of BL-3050 in mice to assess its toxicity and adverse immune responses. The in vivo efficacy of recombinant PON1 and BL-3050 were tested with an animal model of chlorpyrifos-oxon poisoning.
  
  
  RESULTS
  Inactivation studies show significantly improved in vitro lifespan of the engineered rePON1 relative to human PON1. Significant sequence changes relative to human PON1 might hamper the in vivo applicability of BL-3050 due to adverse immune responses. However, we observed no toxic effects in mice subjected to repetitive administration of BL-3050, suggesting that BL-3050 could be safely used. To further evaluate the activity of BL-3050 in vivo, we applied an animal model that mimics human organophosphate poisoning. In these studies, a significant advantages of rePON1 and BL-3050 (>87.5% survival versus <37.5% in the control groups) was observed. Furthermore, BL-3050 and rePON1 were superior to the conventional treatment of atropine-2-PAM as a prophylactic treatment for OP poisoning.
  
  
  CONCLUSIONS
  In vitro and in vivo data described here demonstrate the potential advantages of rePON1 and BL-3050 for treatment of OP toxicity and chronic cardiovascular diseases like atherosclerosis. The in vivo data also suggest that rePON1 and BL-3050 are stable and safe, and could be used for acute, and possibly repeated treatments, with no adverse effects.
authors.affiliation: Department of Biological Chemistry, Weizmann Institute of Science, Rehovot, Israel. leongai@mit.edu
authors.forename: Leonid
authors.initials: L
authors.lastname: Gaidukov
authors.forename: Dganit
authors.initials: D
authors.lastname: Bar
authors.forename: Shiri
authors.initials: S
authors.lastname: Yacobson
authors.forename: Esmira
authors.initials: E
authors.lastname: Naftali
authors.forename: Olga
authors.initials: O
authors.lastname: Kaufman
authors.forename: Rinat
authors.initials: R
authors.lastname: Tabakman
authors.forename: Dan S
authors.initials: DS
authors.lastname: Tawfik
authors.forename: Etgar
authors.initials: E
authors.lastname: Levy-Nissenbaum
mesh_terms: D000818:Animals; D043303:Aryldialkylphosphatase / Q000008:administration & dosage* / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000235:genetics; D004390:Chlorpyrifos / Q000008:administration & dosage / Q000031:analogs & derivatives; D004195:Disease Models, Animal; D004795:Enzyme Stability / Q000187:drug effects / Q000235:genetics; D005260:Female; D005978:Glutathione / Q000008:administration & dosage; D006801:Humans; D007275:Injections, Intravenous; D008075:Lipoproteins, HDL / Q000008:administration & dosage* / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000502:physiology; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D010755:Organophosphates / Q000037:antagonists & inhibitors / Q000633:toxicity; D010713:Phosphatidylcholines / Q000008:administration & dosage; D015202:Protein Engineering / Q000379:methods*; D011994:Recombinant Proteins / Q000008:administration & dosage / Q000037:antagonists & inhibitors / Q000737:chemistry
chemical_list: D008075:Lipoproteins, HDL; D010755:Organophosphates; D010713:Phosphatidylcholines; D011994:Recombinant Proteins; C009618:O,O-diethyl O-3,5,6-trichloro-2-pyridyl phosphate; D043303:Aryldialkylphosphatase; C468406:PON1 protein, human; D005978:Glutathione; D004390:Chlorpyrifos; C028694:1-palmitoyl-2-oleoylphosphatidylcholine