Arthritis & Rheumatism, Volume 62,
November 2010 Abstract Supplement
Abstracts of the American College of
Rheumatology/Association of Rheumatology Health Professionals
Annual Scientific Meeting
Atlanta, Georgia November 6-11, 2010.
Towards Personalized Arthritis Gene Therapy: Screening of Computationally-Designed Promoters in an In Vivo Mouse Model and in Human Synovial Fibroblasts.
van de Loo, Fons A. J., Vermeij, Eline, J. Arntz, Onno, B. Bennink, Miranda, Geurts, Jeroen, van den Berg, Wim B.
Chronic synovial inflammation is a hallmark of the autoimmune disease rheumatoid arthritis (RA). The synovial membrane is regarded as the primary target tissue for local gene therapeutic treatment of arthritic joints. RA is a truly heterogeneous disease, which is supported by gene expression analyses of RA tissues that have revealed molecularly distinct inflammatory subtypes. Therefore, personalized gene therapy using disease inducible promoters might be a good approach to fine-tune supply of biologicals in an offer-meets-demand fashion, increasing safety and efficacy.
Previously, we developed a computational approach for selecting suitable promoters from endogenous genes differentially regulated in the inflamed synovium of collagen-induced arthritis mice. Based on this approach, we constructed lentiviral luciferase reporters containing proximal promoter regions that were predicted to be predominantly regulated by the transcription factors NFkB (Cxcl1, Cxcl5, Il1b), AP-1 (Mmp3, Mmp13, Timp1) or C/EBPb (Saa3, Chi3l1, Has1).
Next, we investigated kinetics and strength of these promoters as compared to a strong constitutive promoter (PGK) in experimental arthritis with two relapsing flares of inflammation. Surprisingly, only Saa3 and Cxcl1 were strongly upregulated (1050 fold) during the first challenge, while a second challenge induced strong luciferase expression for all constructs (10200 fold). The Saa3 promoter activity showed a clear correlation with joint edema as measured by 99mTc uptake. Interestingly, in the absence of a second flare the Cxcl1 promoter activity still increased whereas the Saa3 promoter activity returned to basal levels. Promoter analysis suggested that hypoxia-response elements in the Cxcl1 promoter are likely causative for this difference in expression profile with Saa3. Finally, we characterized the response of computationally-defined promoters in RA synovial tissue of varying inflammatory subtypes. Saa3, Cxcl1, Cxcl5 and Il1b promoters could be induced by stimulation of RA synovial fibroblasts with pro-inflammatory stimuli. Uniquely, relative Saa3 promoter responses to cytokines and a TLR4 agonist were significantly higher in fibroblasts with an inflammatory genetic imprint both as a group as in individual samples. Relative Cxcl1 promoter responses did not discriminate between inflammatory synovial fibroblast subtypes.
These data demonstrate that computational design of promoters is of great value for the development of disease-regulated gene therapy, Towards personalized arthritis gene therapy, the Saa3 promoter appears an excellent candidate for sensing the extent of joint inflammation and responding differentially in human arthritic synovium according to their inflammatory phenotype.
To cite this abstract, please use the following information:
van de Loo, Fons A. J., Vermeij, Eline, J. Arntz, Onno, B. Bennink, Miranda, Geurts, Jeroen, van den Berg, Wim B.; Towards Personalized Arthritis Gene Therapy: Screening of Computationally-Designed Promoters in an In Vivo Mouse Model and in Human Synovial Fibroblasts. [abstract]. Arthritis Rheum 2010;62 Suppl 10 :1733