Significance Tumor-associated macrophages (TAMs) are cells of our innate immune system that have been associated with poor prognosis in many types of cancers. When polarized toward the anti-inflammatory state, TAMs promote immune evasion and angiogenesis, thereby driving tumor growth. Using a peptide library selection strategy, we identified a sequence, called M2pep, that preferentially binds to anti-inflammatory murine macrophages. We then used M2pep to carry a proapoptotic peptide to TAMs by i.v. delivery and demonstrated that selective reduction of TAMs resulted in improved survival in tumor-bearing mice. These results suggest that a molecular-targeting approach for TAM depletion is a promising adjunct strategy to add to the arsenal of anticancer therapies.

We have used a subtractive method to clone novel messages enriched in the heart. Here we show that one such message, bves (blood vessel/epicardial substance) is a novel protein that is highly conserved between chicken and mouse. The bves message is detected at high levels in early chick hearts. Using anti-Bves antibodies, we show expression in cells of the proepicardial organ, migrating epicardium, epicardial-derived mesenchyme, and smooth muscle of the developing intracardiac arterial system, including the coronary arteries. Our data suggest that Bves is an early marker of developing vascular smooth muscle cells. In addition, the expression pattern of Bves protein reveals the patterning of intracardiac vascular smooth muscle and possible insights into the cellular regulation of smooth muscle differentiation during vasculogenesis.

Precise genetic manipulation of human pluripotent stem cells will be required to realize their scientific and therapeutic potential. Here, we show that adeno-associated virus (AAV) gene targeting vectors can be used to genetically engineer human embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs). Different types of sequence-specific changes, including the creation and correction of mutations, were introduced into the human HPRT1 and HMGA1 genes (HPRT1 mutations being responsible for Lesch-Nyhan syndrome). Gene targeting occurred at high frequencies in both ESCs and iPSCs, with over 1% of all colony-forming units (CFUs) undergoing targeting in some experiments. AAV vectors could also be used to target genes in human fibroblasts that were subsequently used to derive iPSCs. Accurate and efficient targeting took place with minimal or no cytotoxicity, and most of the gene-targeted stem cells produced were euploid and pluripotent.

RNA interference (RNAi) offers a promising therapeutic approach for dominant genetic disorders that involve gain-of-function mechanisms. One candidate disease for RNAi therapy application is myotonic dystrophy type 1 (DM1), which results from toxicity of a mutant mRNA. DM1 is caused by expansion of a CTG repeat in the 3' UTR of the DMPK gene. The expression of DMPK mRNA containing an expanded CUG repeat (CUG(exp)) leads to defects in RNA biogenesis and turnover. We designed miRNA-based RNAi hairpins to target the CUG(exp) mRNA in the human α-skeletal muscle actin long-repeat (HSA(LR)) mouse model of DM1. RNAi expression cassettes were delivered to HSA(LR) mice using recombinant adeno-associated viral (rAAV) vectors injected intravenously as a route to systemic gene therapy. Vector delivery significantly reduced disease pathology in muscles of the HSA(LR) mice, including a reduction in the CUG(exp) mRNA, a reduction in myotonic discharges, a shift toward adult pre-mRNA splicing patterns, reduced myofiber hypertrophy and a decrease in myonuclear foci containing the CUG(exp) mRNA. Significant reversal of hallmarks of DM1 in the rAAV RNAi-treated HSA(LR) mice indicate that defects characteristic of DM1 can be mitigated with a systemic RNAi approach targeting the nuclei of terminally differentiated myofibers. Efficient rAAV-mediated delivery of RNAi has the potential to provide a long-term therapy for DM1 and other dominant muscular dystrophies.