title: Biosynthesis, characterization, and efficacy in retinal degenerative diseases of lens epithelium-derived growth factor fragment (LEDGF1-326), a novel therapeutic protein.
abstract: For vision-threatening retinitis pigmentosa and dry age-related macular degeneration, there are no United States Food and Drug Administration (FDA)-approved treatments. We identified, biosynthesized, purified, and characterized lens epithelium-derived growth factor fragment (LEDGF1-326) as a novel protein therapeutic. LEDGF1-326 was produced at about 20 mg/liter of culture when expressed in the Escherichia coli system, with about 95% purity and aggregate-free homogeneous population with a mean hydrodynamic diameter of 9 ± 1 nm. The free energy of unfolding of LEDGF1-326 was 3.3 ± 0.5 kcal mol(-1), and melting temperature was 44.8 ± 0.2 °C. LEDGF1-326 increased human retinal pigment epithelial cell viability from 48.3 ± 5.6 to 119.3 ± 21.1% in the presence of P23H mutant rhodopsin-mediated aggregation stress. LEDGF1-326 also increased retinal pigment epithelial cell FluoSphere uptake to 140 ± 10%. Eight weeks after single intravitreal injection in Royal College of Surgeons (RCS) rats, LEDGF1-326 increased the b-wave amplitude significantly from 9.4 ± 4.6 to 57.6 ± 8.8 μV for scotopic electroretinogram and from 10.9 ± 5.6 to 45.8 ± 15.2 μV for photopic electroretinogram. LEDGF1-326 significantly increased the retinal outer nuclear layer thickness from 6.34 ± 1.6 to 11.7 ± 0.7 μm. LEDGF1-326 is a potential new therapeutic agent for treating retinal degenerative diseases.
authors: Skaggs School of Pharmacy and Pharmaceutical Sciences, University of Colorado Anschutz Medical Campus, Aurora, Colorado 80045, USA.
authors: Rinku
authors: R
authors: Baid
authors: Arun K
authors: AK
authors: Upadhyay
authors: Toshimichi
authors: T
authors: Shinohara
authors: Uday B
authors: UB
authors: Kompella
mesh_terms: D000818:Animals; D002460:Cell Line; D048430:Cell Shape / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002851:Chromatography, High Pressure Liquid; D002942:Circular Dichroism; D003001:Cloning, Molecular; D004596:Electroretinography; D057092:Geographic Atrophy / Q000188:drug therapy* / Q000473:pathology; D006801:Humans; D056965:Injections, Intraocular; D036341:Intercellular Signaling Peptides and Proteins / Q000008:administration & dosage* / Q000096:biosynthesis / Q000302:isolation & purification; D008027:Light; D008970:Molecular Weight; D010316:Particle Size; D010446:Peptide Fragments / Q000008:administration & dosage* / Q000096:biosynthesis / Q000302:isolation & purification; D010587:Phagocytosis / Q000187:drug effects; D055550:Protein Stability; D017433:Protein Structure, Secondary; D051381:Rats; D017948:Retinal Rod Photoreceptor Cells / Q000187:drug effects / Q000502:physiology; D012174:Retinitis Pigmentosa / Q000188:drug therapy* / Q000473:pathology; D012542:Scattering, Radiation; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013312:Stress, Physiological
chemical_list: D036341:Intercellular Signaling Peptides and Proteins; C582957:LEDGF1-326; D010446:Peptide Fragments
keywords: Dry Age-related Macular Degeneration; Growth Factors; Lens Epithelium-derived Growth Factor; Protein Expression; Protein purification; Retinal Degeneration; Retinitis Pigmentosa; Therapeutic Protein; Transcription Factors
references: BMC Ophthalmol. 2007 Feb 26;7:3
references: 17324285
references: Arch Ophthalmol. 1991 Jan;109(1):92-101
references: 1987956
references: Am J Ophthalmol. 2004 Mar;137(3):486-95
references: 15013873
references: Proc Natl Acad Sci U S A. 1998 Sep 29;95(20):11933-8
references: 9751768
references: J Biol Chem. 2003 Jan 3;278(1):372-81
references: 12407101
references: Mol Med. 2010 Nov-Dec;16(11-12):535-42
references: 20683548
references: Nat Rev Drug Discov. 2005 Apr;4(4):298-306
references: 15803194
references: JAMA. 2005 Apr 20;293(15):1841-5
references: 15840844
references: Mol Ther. 2012 May;20(5):908-17
references: 22334021
references: Lancet. 2006 Nov 18;368(9549):1795-809
references: 17113430
references: Hum Mol Genet. 2008 Oct 1;17(19):3043-54
references: 18635576
references: PLoS One. 2011;6(9):e24616
references: 21915354
references: Invest Ophthalmol Vis Sci. 1989 Nov;30(11):2341-7
references: 2807791
references: Biol Cell. 2004 May;96(4):261-9
references: 15145530
references: Exp Eye Res. 1998 Nov;67(5):509-15
references: 9878212
references: J Biol Chem. 2003 Aug 29;278(35):33528-39
references: 12796494
references: Pharmacol Rep. 2006 May-Jun;58(3):353-63
references: 16845209
references: Invest Ophthalmol Vis Sci. 2000 Apr;41(5):1168-75
references: 10752956
references: Arch Ophthalmol. 1993 Jun;111(6):761-72
references: 8512476
references: J Biol Chem. 2002 Sep 13;277(37):34150-60
references: 12091393
references: J Mol Biol. 2006 Jan 20;355(3):379-94
references: 16318853
references: Postepy Hig Med Dosw (Online). 2007;61:83-94
references: 17369776
references: J Natl Cancer Inst. 1996 Nov 6;88(21):1560-70
references: 8901854
references: Am J Clin Nutr. 1999 Apr;69(4):656-63
references: 10197566
references: Arch Ophthalmol. 2004 Apr;122(4):477-85
references: 15078664
references: J Biol Chem. 2002 May 10;277(19):17016-22
references: 11861639
references: Nucleic Acids Res. 2003 Jul 1;31(13):3784-8
references: 12824418
references: Arch Ophthalmol. 2004 Apr;122(4):564-72
references: 15078675
references: Invest Ophthalmol Vis Sci. 2000 Aug;41(9):2648-57
references: 10937578
references: N Engl J Med. 2000 Feb 17;342(7):483-92
references: 10675430
references: Adv Exp Med Biol. 2010;664:403-9
references: 20238041
references: Nippon Ganka Gakkai Zasshi. 2002 Aug;106(8):461-73
references: 12229196
references: J Hered. 1975 Jul-Aug;66(4):242-4
references: 1172515
references: Methods Enzymol. 2009;466:549-65
references: 21609876
references: Nature. 1990 Jan 25;343(6256):364-6
references: 2137202
references: Invest Ophthalmol Vis Sci. 2001 Apr;42(5):1087-95
references: 11274090
references: Invest Ophthalmol Vis Sci. 1999 Jun;40(7):1444-51
references: 10359326
references: Invest Ophthalmol Vis Sci. 2001 Nov;42(12):2935-41
references: 11687539
references: Arch Ophthalmol. 1993 Nov;111(11):1456-9
references: 8240091
references: Trends Biotechnol. 1990 Apr;8(4):93-8
references: 1367432
references: Photochem Photobiol. 1973 Oct;18(4):263-79
references: 4583619
references: J Am Chem Soc. 2005 Jul 13;127(27):9700-1
references: 15998070