Poly(beta-amino ester)s are cationic degradable polymers that have significant potential as gene delivery vectors. Here we present a generalized method to modify poly(beta-amino ester)s at the chain ends to improve their delivery performance. End-chain coupling reactions were developed so that polymers could be synthesized and tested in a high-throughput manner, without the need for purification. In this way, many structural variations at the polymer terminus could be rapidly evaluated. End-modification of the terminal amine structure of a previously optimized poly(beta-amino ester), C32, significantly enhanced its in vitro transfection efficiency. In vivo, intraperitoneal (IP) gene delivery using end-modified C32 polymers resulted in expression levels over one order of magnitude higher than unmodified C32 and jet-polyethylenimine (jet-PEI) levels in several abdominal organs. The rapid end-modification strategy presented here has led to the discovery of many effective polymers for gene delivery and may be a useful method to develop and optimize cationic polymers for gene therapy.

Claudin-3 (CLDN3) is a tight junction protein that is overexpressed in 90% of ovarian tumors. Previous in vitro studies have indicated that CLDN3 overexpression promotes the migration, invasion, and survival of ovarian cancer cells. Here, we investigated the efficacy of lipidoid-formulated CLDN3 siRNA in 3 different ovarian cancer models. Intratumoral injection of lipidoid/CLDN3 siRNA into OVCAR-3 xenografts resulted in dramatic silencing of CLDN3, significant reduction in cell proliferation, reduction in tumor growth, and a significant increase in the number of apoptotic cells. Intraperitoneal injection of lipidoid-formulated CLDN3 siRNA resulted in a substantial reduction in tumor burden in MISIIR/TAg transgenic mice and mice bearing tumors derived from mouse ovarian surface epithelial cells. Ascites development was reduced in CLDN3 siRNA-treated mice, suggesting the treatment effectively suppressed metastasis. Toxicity was not observed after multiple i.p. injections. Importantly, treatment of mice with nonimmunostimulatory 2 -OMe modified CLDN3 siRNA was as effective in suppressing tumor growth as unmodifed siRNA. These results suggest that lipidoid-formulated CLDN3 siRNA has potential as a therapeutic for ovarian cancer.lipidoid ͉ ovarian cancer ͉ cancer therapy O varian cancer has the highest mortality rate among gynecologic malignancies, and ranks 4th as the most common cancer in women in the United States (1). Treatment of early stage ovarian carcinoma improves the survival rate up to 90%. However, most women have advanced stage metastatic cancer at the time of diagnosis due to the asymptomatic nature of early stages of the disease and the lack of effective screening modalities. The standard treatment for patients with advanced stage epithelial ovarian cancer is surgical debulking followed by chemotherapy with paclitaxel plus a platinum-based therapy (cisplatin or carboplatin). Although Ϸ80% of patients receiving this therapeutic regimen have an initial favorable response, recurrent disease will occur in a majority of cases. New effective therapies are urgently needed for those patients with advanced-stage ovarian cancer who either do not respond to initial therapy or develop recurrent disease.Claudins are integral membrane proteins associated with tight junctions. Two members of the claudin protein family, claudin-3 (CLDN3) and CLDN4, are overexpressed in epithelial ovarian tumors relative to normal ovarian tissue (2-5). In fact, they are among the most highly expressed proteins in ovarian tumors. High amounts of CLDN3 and CLDN4 are associated with increased cellular motility and survival of ovarian tumor cells, and an increase in matrix metalloproteinase type 2 (MMP-2) (4). These observations implicate a role for CLDN3 and CLDN4 in ovarian tumorigenesis and metastasis, and suggest their importance as target proteins for development of new diagnostic and therapeutic reagents. CLDN3 and CLDN4 have also been identified as receptors for cytotoxic Clostridium perfringens enterotoxin (CPE). Binding of CPE to ...

Growing evidence shows that cancer cells use mRNA-binding proteins and miRNAs to posttranscriptionally regulate signaling pathways to adapt to harsh tumor microenvironments. In ovarian cancer, cytoplasmic accumulation of mRNA-binding protein HuR (ELAVL1) is associated with poor prognosis. In this study, we observed high HuR expression in ovarian cancer cells compared with ovarian primary cells, providing a rationale for targeting HuR. RNAi-mediated silencing of HuR in ovarian cancer cells significantly decreased cell proliferation and anchorage-independent growth, and impaired migration and invasion. In addition, HuR-depleted human ovarian xenografts were smaller than control tumors. A biodistribution study showed effective tumortargeting by a novel Cy3-labeled folic acid (FA)-derivatized DNA dendrimer nanocarrier (3DNA). We combined siRNAs against HuR with FA-3DNA and found that systemic administration of the resultant FA-3DNA-siHuR conjugates to ovarian tumorbearing mice suppressed tumor growth and ascites development, significantly prolonging lifespan. NanoString gene expression analysis identified multiple HuR-regulated genes that function in many essential cellular and molecular pathways, an attractive feature of candidate therapeutic targets. Taken together, these results are the first to demonstrate the versatility of the 3DNA nanocarrier for in vivo-targeted delivery of a cancer therapeutic and support further preclinical investigation of this system adapted to siHuR-targeted therapy for ovarian cancer.

Poly(beta-amino ester)s are cationic degradable polymers that have significant potential as gene delivery vectors. Here we present a generalized method to modify poly(beta-amino ester)s at the chain ends to improve their delivery performance. End-chain coupling reactions were developed so that polymers could be synthesized and tested in a high-throughput manner, without the need for purification. In this way, many structural variations at the polymer terminus could be rapidly evaluated. End-modification of the terminal amine structure of a previously optimized poly(beta-amino ester), C32, significantly enhanced its in vitro transfection efficiency. In vivo, intraperitoneal (IP) gene delivery using end-modified C32 polymers resulted in expression levels over one order of magnitude higher than unmodified C32 and jet-polyethylenimine (jet-PEI) levels in several abdominal organs. The rapid end-modification strategy presented here has led to the discovery of many effective polymers for gene delivery and may be a useful method to develop and optimize cationic polymers for gene therapy.