The urgent need for a better therapy for advanced ovarian cancer, and our previous work demonstrating the effective delivery of DNA encoding diphtheria toxin (DT-A) to prostate tumor cells in mice following intratumoral injection of nanoparticles formulated with the poly(β-amino ester) C32 (2
), led us to explore the therapeutic efficacy of this nanotherapy for the treatment of ovarian cancer. We reasoned that intraperitoneal delivery of DT-A nanoparticles would avoid biodistribution complications associated with intravenous injection of nanoparticles and would therefore allow their access to metastatic tumors in the peritoneal cavity where most metastatic ovarian tumors are located. Intraperitoneal administration of paclitaxel nanoparticles has been shown to inhibit the progression of ovarian carcinoma in rats (20
). It is noteworthy that a recent report indicating that intraperitoneal chemotherapy for advanced ovarian cancer improves overall and disease-free survival (21
) was the basis for a rare Clinical Announcement posted by the National Cancer Institute in January 2006 recommending that physicians use this mode of drug administration in treating ovarian cancer patients.
We investigated the therapeutic efficacy of using polymeric nanoparticles to deliver DNA encoding DT-A to ovarian cancer cells in three mouse models – a xenograft model, a transgenic mouse model, and a cell implantation model. Direct injection of nanoparticles into xenografts, as well as incubation of nanoparticles with primary ascites cells collected from patients, demonstrated the ability of polymeric nanoparticles to deliver DNA to ovarian cancer cells of human origin. Expression of nanoparticle-delivered DT-A DNA suppressed the growth of xenografts in immunosuppressed mice. TUNEL assays done on DT-A nanoparticle-injected tumors in immunocompetent MISIIR/TAg mice showed that tumor cells undergo apoptosis following DT-A expression.
MISIIR/TAg transgenic mice and the ID8-Fluc cell implant model were used to compare the therapeutic efficacy of DT-A nanoparticles with that of chemotherapy (cisplatin + paclitaxel). The growth of ovarian tumors in MISIIR/TAg mice treated with DT-A nanoparticles was significantly less than in mice treated with the drugs. In fact, 42% (3/7) of tumors in the DT-A treated mice shrunk in size over the 3-week course of treatment, whereas all of the tumors in the drug-treated mice grew in size. An additional study using the ID8-Fluc cell implant model confirms that DT-A nanotherapy is as effective, if not better, than the combined drug therapy in reducing tumor load. While histological analyses of multiple non-tumorous tissues from drug-treated and from DT-A treated mice showed no significant non-specific toxicity, drug-treatment of the ID8-Fluc implant mice did result in significant weight loss, while DT-A treated mice showed no weight loss.
The promoters of two genes, HE4
, are preferentially active in ovarian cancer cells. We used the MSLN promoter to regulate DT-A expression in the nano vs chemotherapy studies as well as in a lifespan study. This promoter appears to target DT-A expression quite effectively because, although we have shown that DNA is delivered to multiple organs following intraperitoneal injection of C32-117 nanoparticles (6
), very little non-specific toxicity was observed upon histological analysis of multiple tissues collected from mice that had received i.p.
injections of DT-A nanoparticles for several months. Nevertheless, in further development of this therapy, we plan to make modifications to the nanoparticle formulations that will enhance targeted delivery of DNA to tumor cells. A combination of targeting delivery as well as targeting expression through the use of specific promoters will help ensure the maintenance of non-cancerous, healthy tissues.
The therapeutic response of tumors in mice treated with DT-A nanoparticles, in the absence of significant non-specific toxicity, establishes the utility of a DNA-based nanotherapy for the treatment of ovarian cancer. The C32-117 poly(β-amino ester) polymer effectively delivers DNA to tumor cells following intraperitoneal delivery, while the use of specific promoter sequences effectively targets expression of the therapeutic DNA to the tumor cells. The acquired resistance of tumor cells to chemotherapeutics is responsible, in large part, for the poor prognosis of patients with advanced ovarian cancer. Given the rapid shutdown of protein synthesis following uptake and expression of DT-A encoding DNA, it is likely that cells will not exhibit resistance to the nanotherapy. Our study suggests that it should be possible to administer DT-A nanotherapy over an extended period of time to suppress tumor growth, and perhaps even reduce tumor burden.
Currently, ovarian cancer patients are treated with chemotherapeutics following surgical debulking. Further studies using the MISIIR/TAg model will be aimed at assessing the therapeutic efficacy of DT-A nanotherapy as an adjuvant therapy to surgical debulking. DT-A nanotherapy is also likely to be an effective therapy for tumors that develop in other organs in the peritoneum, e.g., pancreatic cancer (15
). There is no clinical precedent for the use of C32-117 poly(β-amino ester) polymer as a delivery vehicle. The NCI recently announced that it will sponsor further preclinical development of this therapy for the treatment of solid tumors at the Nanotechnology Characterization Laboratory.