Engagement of the B-cell antigen receptor (BCR) allows efficient transduction of ZAP-70-positive primary B-CLL cells by recombinant adeno-associated virus (rAAV) vectors

KKG Gene Therapy, GSF-National Research Center for Environment and Health, Munich, Germany.
Gene Therapy (Impact Factor: 3.1). 10/2004; 11(18):1416-24. DOI: 10.1038/sj.gt.3302279
Source: PubMed


Engagement of the B-cell antigen receptor (BCR) by crosslinking of the surface immunoglobulin (sIg) homodimer was studied for recombinant adeno-associated virus (rAAV)-mediated gene transfer into B-cell chronic lymphocytic leukaemia (B-CLL) cells. Leukemic cells obtained from 20 patients were stimulated with anti-sIg-directed antibodies and transduced with rAAV vectors coding for enhanced green fluorescent protein (EGFP) (AAV/EGFP) or CD40L (AAV/CD40L). Transduction of B-CLL cells was enhanced after BCR engagement compared to unstimulated controls (P=0.0356). BCR crosslinking induced a significant, dose- and time-dependent upregulation of heparan sulfate proteoglycan (HSPG), the primary receptor for AAV, on B-CLL cells (mean: 38.2 versus 1.7%; P=0.0006). A correlation of HSPG expression after BCR crosslinking with transduction efficiency by AAV/EGFP (P=0.0153) and AAV/CD40L (P=0.0347) was observed. High expression of zeta-associated protein 70 (ZAP-70) in B-CLL cells correlated with a better transduction efficiency by AAV/EGFP (P<0.0001) and AAV/CD40L (P=0.002), respectively: 48 h after transduction of ZAP-70-positive samples, transgene expression was seen in a mean of 33.8% (s.e.m. 3.7%) and 28.9% (s.e.m. 6.7%) of cells, respectively, and could be specifically blocked by heparin, a soluble competitor of HSPG (P<0.0001). In summary, engagement of the BCR on ZAP-70 positive B-CLL cells allows efficient rAAV-mediated gene delivery.

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Available from: Clemens-Martin Wendtner, Mar 10, 2014
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    • "Cells were cultured as described before [20]. BCR stimulation was performed as described by Kofler et al. [21] Anti-IgM-polyacrylamid immunobead (anti-IgM) reagent (Irvine Scientific, Santa Ana, CA, USA) was added to the PBMC cultures at a concentration of 100 µg/mL for 3 or 24 hours. Anti-IgA-immunobeads (anti-IgA, Irvine Scientific) served as a negative control. "
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