Alternative splice forms of CTLA-4 induced by antisense mediated splice-switching influences autoimmune diabetes susceptibility in NOD mice.
Activated and regulatory T cells express the negative co-stimulatory molecule cytotoxic T-lymphocyte-associated antigen-4 (CTLA-4) that binds B7 on antigen-presenting cells to mediate cellular responses. Single nucleotide polymorphisms in the CTLA-4 gene have been found to affect alternative splicing and are linked to autoimmune disease susceptibility or resistance. Increased expression of a soluble splice form (sCTLA-4), lacking the transmembrane domain encoded by exon 3, has been shown to accelerate autoimmune pathology. In contrast, an exon 2-deficient form lacking the B7 ligand binding domain (liCTLA-4), expressed by diabetes resistant mouse strains has been shown to be protective when expressed as a transgene in diabetes susceptible non-obese diabetic (NOD) mice. We sought to employ an antisense-targeted splice-switching approach to independently produce these CTLA-4 splice forms in NOD mouse T cells and observe their relative impact on spontaneous autoimmune diabetes susceptibility. In vitro antisense targeting of the splice acceptor site for exon 2 produced liCTLA-4 while targeting exon 3 produced the sCTLA-4 form in NOD T cells. The liCTLA-4 expressing T cells exhibited reduced activation, proliferation and increased adhesion to intercellular adhesion molecule-1 (ICAM-1) similar to treatment with agonist α-CTLA-4. Mice treated to produce liCTLA-4 at the time of elevated blood glucose levels exhibited a significant reduction in the incidence of insulitis and diabetes, whereas a marked increase in the incidence of both was observed in animals treated to produce sCTLA-4. These findings provide further support that alternative splice forms of CTLA-4 affects diabetes susceptibility in NOD mice and demonstrates the therapeutic utility of antisense mediated splice-switching for modulating immune responses.
1 Sarepta Therapeutics , Corvallis, Oregon.
Dan V
DV
Mourich
Shannon K
SK
Oda
Frederick J
FJ
Schnell
Stacy L
SL
Crumley
Laura L
LL
Hauck
Carla A
CA
Moentenich
Nikki B
NB
Marshall
Dave J
DJ
Hinrichs
Patrick L
PL
Iversen
D000069594:Abatacept; D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000818:Animals; D015551:Autoimmunity / Q000235:genetics*; D001483:Base Sequence; D060908:CTLA-4 Antigen / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000276:immunology; D002448:Cell Adhesion; D049109:Cell Proliferation; D003922:Diabetes Mellitus, Type 1 / Q000235:genetics* / Q000276:immunology / Q000473:pathology; D004198:Disease Susceptibility / Q000276:immunology*; D005091:Exons; D005786:Gene Expression Regulation; D018796:Immunoconjugates / Q000494:pharmacology; D018799:Intercellular Adhesion Molecule-1 / Q000235:genetics / Q000276:immunology; D008213:Lymphocyte Activation; D051379:Mice; D016688:Mice, Inbred NOD; D008969:Molecular Sequence Data; D016376:Oligonucleotides, Antisense / Q000235:genetics* / Q000276:immunology; D020033:Protein Isoforms / Q000037:antagonists & inhibitors / Q000235:genetics / Q000276:immunology; D012720:Severity of Illness Index; D050378:T-Lymphocytes, Regulatory / Q000187:drug effects / Q000276:immunology / Q000473:pathology
D060908:CTLA-4 Antigen; D018796:Immunoconjugates; D016376:Oligonucleotides, Antisense; D020033:Protein Isoforms; D018799:Intercellular Adhesion Molecule-1; D000069594:Abatacept