Srilalitha Belur, PhD

T32 Support:  2001 – 2004

Education:     1985 – 1988; B.S., University of Bombay, Bombay, India
                      1988 – 1990; M.Sc., University of Bombay, Bombay, India 

Project:          Sleeping Beauty-Mediated Gene Expression and Sustained Expression in Lung

Our goal was to apply the Sleeping Beauty transposon system in a non-viral gene transfer strategy for the treatment of hyperoxia-induced lung injury.  We constructed a series of transposons that contained the rat heme oxygenase-1 (HO-1) gene to be used for gene transfer and expression in an animal model of hyperoxia.  In these constructs, the HO-1 gene is under transcriptional regulation of several different promoters: RSV-LTR, early CMV, CAGS, or Ubiquitin.  We also engineered a bicistronic transposon containing the HO-1 transgene under transcriptional regulation of the CAGS promoter, plus a luciferase transgene downstream of an IRES.  All of these constructs have been tested for expression of HO-1 enzyme activity after transfection into HEK 293 cells.  Based on these in vitro results, the construct expressing the highest level of HO-1 activity (bicistronic construct) was injected systemically into C57BL/6 mice, resulting in both HO-1 and luciferase expression in the lung.  Following this demonstration of in vivo gene transfer and expression, C57BL/6 mice will be exposed to >99% oxygen and then injected with the bicistronic transposon with or without Sleeping Beauty transposase.  Experiments to elucidate the protective effects of the HO-1 transposon and Sleeping Beauty transposase are currently in progress by Dr. Belur in conjunction with Dr. McIvor.

Publications:

Belur LR. Boelk-Galvan D. Diers MD. McIvor RS. Zimmerman CL. Methotrexate accumulates to similar levels in animals transplanted with normal versus drug-resistant transgenic marrow. Cancer Research. 2001; 61(4):1522-6.

Belur L, Frandsen J, Dupuy A, Ingbar DH, Largaespada D, Hackett PB, McIvor RS.  Gene insertion and long-term expression in lung mediated by the Sleeping Beauty transposon system.  Molecular Therapy    2003; 8(3):501-7.

Converse A, Belur L, Gori G, Liu F, Amaya E, Aguilar-Cordova P, Hackett B, McIvor RS.  Counterselection and co-delivery of transposon and transposase functions for Sleeping Beauty-mediated transposition in cultured mammalian cells.  Somatic Cell and Molecular Genetics 2003.

Belur L, McIvor RS.  Polyethyleneimine-mediated non-viral gene delivery to the lung.  Methods in Molecular Biology 2004; 245:137-44.

Belur LR. James RI. May C. Diers MD. Swanson D. Gunther R. McIvor RS. Methotrexate preconditioning allows sufficient engraftment to confer drug resistance in mice transplanted with marrow expressing drug-resistant dihydrofolate reductase activity. Journal of Pharmacology & Experimental Therapeutics.   2005; 314(2):668-74.

Score PR. Belur LR. Frandsen JL. Geurts JL. Yamaguchi T. Somia NV. Hackett PB. Largaespada DA. McIvor RS. Sleeping Beauty-mediated transposition and long-term expression in vivo: use of the LoxP/Cre recombinase system to distinguish transposition-specific expression Molecular Therapy: the Journal of the American Society of Gene Therapy. 2006; 13(3):617-24.

Aronovich EL. Bell JB. Belur LR. Gunther R. Koniar B. Erickson DC. Schachern PA. Matise I. McIvor RS. Whitley CB. Hackett PB. Prolonged expression of a lysosomal enzyme in mouse liver after Sleeping Beauty transposon-mediated gene delivery: implications for non-viral gene therapy of mucopolysaccharidoses. Journal of Gene Medicine  2007; 9(5):403-15.

Wilber A. Linehan JL. Tian X. Woll PS. Morris JK. Belur LR. McIvor RS. Kaufman DS. Efficient and stable transgene expression in human embryonic stem cells using transposon-mediated gene transfer. Stem Cells 2007; 25(11):2919-27.

Belur LR, Podetz-Pedersen, Frandsen J, McIvor RS.  Lung-directed gene therapy in mice using the nonviral Sleeping Beauty transposon system.  Nature Protocols 2007; 2(12):3146-3152.

Bell JB. Podetz-Pedersen KM. Aronovich EL. Belur LR. McIvor RS. Hackett PB.Preferential delivery of the Sleeping Beauty transposon system to livers of mice by hydrodynamic injection.  Nature Protocols 2007; 2(12):3153-3165.

Updated 3-25-2008