Improved I.P. Drug Delivery with Bioadhesive Nanoparticles
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Improved i.p. drug delivery with bioadhesive nanoparticles Yang Denga, Fan Yanga, Emiliano Coccob, Eric Songa, Junwei Zhanga,c, Jiajia Cuia, Muneeb Mohideena, Stefania Belloneb, Alessandro D. Santinb, and W. Mark Saltzmana,c,1 aDepartment of Biomedical Engineering, Yale University, New Haven, CT 06511; bDepartment of Obstetrics, Gynecology & Reproductive Sciences, School of Medicine, Yale University, New Haven, CT 06511; and cDepartment of Chemical & Environmental Engineering, Yale University, New Haven, CT 06511 Edited by Joseph M. DeSimone, University of North Carolina at Chapel Hill and Carbon, Chapel Hill, NC, and approved August 19, 2016 (received for review November 22, 2015) The i.p. administration of chemotherapy in ovarian and uterine (∼10–100 μm) and therefore, should distribute more evenly serous carcinoma patients by biodegradable nanoparticles may throughout the peritoneum. Furthermore, nanoparticles are small represent a highly effective way to suppress peritoneal carcinoma- enough to be internalized by tumor cells (17, 18), allowing them to tosis. However, the efficacy of nanoparticles loaded with chemo- release the encapsulated drugs near the biological target, which is therapeutic agents is currently hampered by their fast clearance by often in the cell nucleus. Although many studies have shown that lymphatic drainage. Here, we show that a unique formulation of nanoparticles are promising vehicles for the delivery of chemother- bioadhesive nanoparticles (BNPs) can interact with mesothelial apeutic agents in preclinical in vivo models, the application of cells in the abdominal cavity and significantly extend the retention nanoparticles for i.p. delivery remains hampered by their fast of the nanoparticles in the peritoneal space. BNPs loaded with a clearance from the peritoneal cavity, mainly as a result of lymphatic potent chemotherapeutic agent [epothilone B (EB)] showed signif- drainage (19). icantly lower systemic toxicity and higher therapeutic efficacy In a recent study (20), we described a novel form of bio- against i.p. chemotherapy-resistant uterine serous carcinoma- adhesive nanoparticles (BNPs) and showed that their adhesive- derived xenografts compared with free EB and non-BNPs loaded ness results in prolonged retention on the skin after topical with EB. delivery. These new materials are based on polylactic acid block– hyperbranched polyglycerol (PLA-HPG) copolymers, which can drug delivery | nanoparticles | ovarian cancer | intraperitoneal | be formed into “stealthy,” nonadhesive nanoparticles (NNPs) chemotherapy that circulate for long times after i.v. injection (21). The BNPs were produced by oxidation of the NNPs, which results in con- igh-grade ovarian and uterine serous carcinoma (USC) are version of vicinal diols on the surface of NNPs into aldehydes on Hbiologically aggressive tumors, which commonly spread into the BNPs (Fig. 1). Aldehydes spontaneously react with proteins the peritoneal cavity, disseminating along the abdominal and to form a variety of bonds, such as Schiff-base bonds; therefore, pelvic peritoneum and resulting in peritoneal metastases. Al- the presence of surface aldehydes on BNPs leads to particle though these tumors are often responsive to initial cytoreductive adhesion to protein-rich materials. For example, we previously surgery and platinum–paclitaxel chemotherapy, most patients showed the rapid and robust adhesion of the BNPs to poly-L- – develop recurrence and eventually die from progression of che- lysine coated surfaces and pig or mouse skin (20). In this work, motherapy-resistant disease (1). we hypothesize that BNPs will interact and adhere strongly with In cases of advanced ovarian cancer, the infusion of chemo- mesothelial cells layering the abdominal cavity after i.p. delivery therapeutic agents directly into the peritoneum (i.e., i.p. chemo- therapy) improves overall and disease-free survival compared with Significance standard i.v. chemotherapy (2). However, despite level 1 evidence from meta-analyses of three large, prospectively randomized, Resistance to platinum-based chemotherapies and paclitaxel is controlled trials, this route of administration is still not widely used common in recurrence of both high-grade ovarian and endo- in the clinic because of its more severe side effects compared with metrial cancers. Paclitaxel resistance has been correlated with i.v. chemotherapy and the need for frequent dosing schedules (2, overexpression of class III β-tubulin, the preferential target of 3). To enhance the retention of chemotherapeutic drugs in the the epothilones, microtubule-stabilizing agents. Epothilone B peritoneal cavity and improve their bioavailability, controlled re- (EB) is manifold more effective than paclitaxel, but clinical use lease drug carriers, such as microparticles and hydrogels, have is limited by side effects. To reduce side effects, we encapsu- recently been tested in preclinical animal models (4–8). Although lated EB into bioadhesive nanoparticles (BNPs), reasoning that these carriers can lead to improvement over conventional delivery bioadhesive nanoparticles loaded with epothilone B (EB/BNPs) methods, both strategies only use the microparticles or hydrogel as would interact with abdominal tissues and gradually release EB reservoirs for drugs. To achieve expected therapeutic efficacy, the in proximity of peritoneal cancer implants, thus maintaining EB concentration of the free drugs in the peritoneal cavity has to be concentration at the site of action and limiting systemic ex- maintained in the effective range. Sustained drug concentrations posure and toxicity. Our experiments show the higher thera- require a balance between the drug release from carriers and clear- peutic activity and limited toxicity of EB/BNPs compared with ance from peritoneal cavity by metabolism. For a specific drug and nonadhesive nanoparticles loaded with EB or carrier-free EB. SCIENCES drug carrier combination, release that is too slow might also com- APPLIED BIOLOGICAL Author contributions: Y.D., F.Y., E.C., E.S., J.Z., J.C., M.M., A.D.S., and W.M.S. designed promise the therapeutic efficacy as observed when limited disso- research; Y.D., F.Y., E.C., E.S., J.Z., J.C., M.M., S.B., A.D.S., and W.M.S. performed research; lution of paclitaxel from a microparticle/hydrogel system impaired Y.D., F.Y., E.C., E.S., J.Z., J.C., M.M., S.B., A.D.S., and W.M.S. analyzed data; and Y.D., E.C., therapeutic outcomes (5). In addition, side effects of some of these A.D.S., and W.M.S. wrote the paper. carriers do exist (i.e., microparticles accumulate in the lower ab- The authors declare no conflict of interest. domen and cause peritoneal adhesion) (9–11). This article is a PNAS Direct Submission. Degradable polymer nanoparticles have several potential advan- 1To whom correspondence should be addressed. Email: [email protected]. – ENGINEERING tages over microparticles and hydrogels in cancer therapy (12 16). This article contains supporting information online at www.pnas.org/lookup/suppl/doi:10. Nanoparticles (∼100 nm) are small compared with microparticles 1073/pnas.1523141113/-/DCSupplemental. www.pnas.org/cgi/doi/10.1073/pnas.1523141113 PNAS | October 11, 2016 | vol. 113 | no. 41 | 11453–11458 Downloaded by guest on September 24, 2021 successfully used in prior work to generate consistent i.p. tumors in mice (28). Results In a previous work, we showed that NNPs had an extended blood circulation because of their resistance to nonspecific interaction to proteins (21). In another recent study, we found that BNPs had an enhanced interaction with protein-rich surfaces caused by their aldehyde groups (20). These results suggest that this class of nanoparticles can be converted from a nonadhesive to a strongly adhesive state and then back again by regulation of the density of aldehydes on the nanoparticle surface. To illustrate this flexi- bility, nanoparticles were converted by a cycle of oxidation and reduction between the hydroxyl-rich NNP state, the aldehyde-rich BNP state, and a reduced nonadhesive nanoparticle (NNP-R) state. We reduced the aldehydes to alcohols by treatment with a common reducing agent (29), NaBH4 (Fig. S1A). The concen- tration of aldehyde groups on BNPs was monitored as a function of duration of NaBH4 treatment; most aldehyde groups were converted after 2 min of treatment (Fig. 2D). It is important to note that NaBH4 treatment cannot reduce the aldehydes to the original vicinal diols, because one carbon is truncated by the initial conversion of vicinal diols to aldehydes (Fig. S1B). BNP, NNP, and NNP-R were identical in shape by transmission EM (TEM) (Fig. 2 A–C) and hydrodynamic size measured by dy- namic light scattering (DLS) (Table 1), suggesting that oxidation and reduction cycles had no detrimental effect to the nano- particles. To quantitate bioadhesion, we tested nanoparticles at various states of aldehyde activation by examining attachment to poly(L-lysine)–coated plates and USC cells (Fig. 2 E and F). To facilitate quantification of their bioadhesive properties, all nanoparticles were loaded with 0.2% 1,1′-dioctadecyl- 3,3,3′,3′-tetramethylindodicarbocyanine, 4-chlorobenzenesulfo- nate salt (DiD) dye. We first evaluated the bioadhesive property of the three nanoparticles on poly(L-lysine)–coated surfaces. After incubation and extensive washing, the BNPs showed much higher retention on the poly(L-lysine)–coated plates (Fig. 2E). The BNPs also showed much higher attachment to USC cells (Fig. 2F). The retention of NNPs or NNP-Rs on both poly(L- lysine) surfaces and USC cell monolayers was negligible (Fig. 2 E Fig. 1. Schematic showing the conversion of BNPs from NNPs and the fate and F). We further tested the nanoparticles for blood circulation of NNPs and BNPs after i.p. delivery. BNPs can be converted from NNPs by times after i.v. injection in mice. The percentage dose of BNPs NaIO4 treatment. After i.p. delivery, (Left) NNPs are cleared by lymphatic dropped to 1.5% after 2 h, whereas significant numbers of NNPs drainage, but (Right) BNPs are retained in the peritoneal cavity because of (21.3%) were still circulating after 24 h (Fig. 2G). NNP-Rs cir- their bioadhesive property.