Anti-CD22 antibody targeting of pH-responsive micelles enhances small interfering RNA delivery and gene silencing in lymphoma cells.

The application of small interfering RNA (siRNA) for cancer treatment is a promising strategy currently being explored in early phase clinical trials. However, efficient systemic delivery limits clinical implementation. We developed and tested a novel delivery system comprised of (i) an internalizing streptavidin-conjugated monoclonal antibody (mAb-SA) directed against CD22 and (ii) a biotinylated diblock copolymer containing both a positively charged siRNA condensing block and a pH-responsive block to facilitate endosome release. The modular design of the carrier facilitates the exchange of different targeting moieties and siRNAs to permit its usage in a variety of tumor types. The polymer was synthesized using the reversible addition fragmentation chain transfer (RAFT) technique and formed micelles capable of binding siRNA and mAb-SA. A hemolysis assay confirmed the predicted membrane destabilizing activity of the polymer under acidic conditions typical of the endosomal compartment. Enhanced siRNA uptake was demonstrated in DoHH2 lymphoma and transduced HeLa-R cells expressing CD22 but not in CD22 negative HeLa-R cells. Gene knockdown was significantly improved with CD22-targeted vs. nontargeted polymeric micelles. Treatment of DoHH2 cells with CD22-targeted polymeric micelles containing 15 nmol/l siRNA produced 70% reduction of gene expression. This CD22-targeted polymer carrier may be useful for siRNA delivery to lymphoma cells.

Identification of gene expression changes associated with the progression of retinal degeneration in the rd1 mouse.

PURPOSE: One approach to gaining insight into the biological pathways contributing to rod and cone photoreceptor death is to identify patterns of gene expression changes. In the present study, a custom retinal microarray was developed to analyze the rd1 mouse, a well-characterized animal model of human retinal degeneration. METHODS: A microarray was constructed containing cDNA fragments corresponding to genes known or postulated to be involved in normal retinal function, development, and disease. Gene expression in rd1 retina was compared with age-matched control retinas at three time points: the peak of rod degeneration (postnatal day [P]14), early in cone degeneration (P35), and during cone degeneration (P50). Selected microarray results were confirmed with real-time PCR. The cellular distribution of one of the differentially expressed genes, dickkopf 3 (Dkk3), was assessed by in situ hybridization. RESULTS: At each stage of degeneration, there was only limited overlap of the genes that showed increased expression, suggesting the involvement of temporally distinct molecular pathways. Genes active in transport mechanisms and in signaling pathways were differentially expressed during rod degeneration, whereas genes with functions in protein modification and cellular metabolism were differentially expressed during cone degeneration. Increased expression of genes involved in cell proliferation pathways and oxidative stress was observed at each time point. CONCLUSIONS: These microarray results provide clues to understanding the molecular pathways underlying photoreceptor degeneration and indicate directions for future studies. In addition, comparisons of normal and degenerated retina identified numerous genes and ESTs that are potentially enriched in rod photoreceptors.