Introduction A major barrier to the advancement of therapeutic nanomedicines has been the non-target toxicity caused by the accumulation of the drug delivery systems in organs associated with the reticuloendothelial system, particularly the liver and spleen. Herein, we report the development of peptide based metabolically active linkers (MALs) that are enzymatically cleaved by cysteine cathepsin B and S, two proteases highly expressed in the liver and spleen. The overall goal of this approach is to utilize the MALs to lower the non-target retention and toxicity of radiolabeled drug delivery systems, thus resulting in higher diagnostic and radiotherapeutic efficacy. Methods In this study three MALs (MAL0, MAL1 and MAL2) were investigated. MAL1 and MAL2 are composed of known substrates of cathepsin B and S, respectively, while MAL0 is a non-cleavable control. Both MAL1 and MAL2 were shown to undergo enzymatic cleavage with the appropriate cathepsin protease. Subsequent to conjugation to the HPMA copolymer and radiolabeling with 177Lu, the peptide-polymer conjugates were renamed 177Lu- metabolically active copolymers (177Lu-MACs) with the corresponding designation 177Lu-MAC0, 177Lu-MAC1 and 177Lu-MAC2. Results In vivo evaluation of the 177Lu-MACs was performed in a HPAC human pancreatic cancer xenograft mouse model. 177Lu-MAC1 and 177Lu-MAC2 demonstrated 3.1 and 2.1 fold lower liver retention, respectively, compared to control (177Lu-MAC0) at 72 h post-injection. With regard to spleen retention, 177Lu-MAC1 and 177Lu-MAC2 each exhibited a nearly fourfold lower retention, relative to control, at the 72 h time point. However, the tumor accumulation of the 177Lu-MAC0 was two to three times greater than 177Lu-MAC1 and 177Lu-MAC2 at the same time point. The MAL approach demonstrated the capability of substantially reducing the non-target retention of the 177Lu-labeled HPMA copolymers. Conclusions While further studies are needed to optimize the pharmacokinetics of the 177Lu-MACs design, the ability of the MAL to significantly decrease non-target retention establishes the potential this avenue of research may have for the improvement of diagnostic and radiotherapeutic drug delivery systems.

Receptor-targeted agents, such as BB2r-targeted peptides, have been investigated extensively in preclinical and clinical studies. In an attempt to increase the effectiveness of diagnostic and/or radiotherapeutic agents, we have begun to explore the incorporation of the hypoxia-selective prodrug 2-nitroimidazole into receptor-targeted peptides. Hypoxia is a well-known characteristic of many solid tumors, including breast, prostate and pancreatic cancers. The aim of this approach is to utilize the hypoxia trapping capability of 2-nitroimidazoles to increase the retention of the agent in hypoxic, BB2r-positive tumors. We have demonstrated that incorporation of one or more 2-nitroimidazoles into the BB2r-targeted peptide significantly increases the in vitro retention of the agent in hypoxic prostate cancer cells. The study described herein represents our first investigation of the in vivo properties of these hypoxia-enhanced BB2r-targeted agents in a PC-3 xenograft mouse model. Method Four 111In-labeled BB2r-targeted conjugates, 111In-1, 111In-2, 111In-3 and 111In-4, composed of 0–3 2-nitroimidazole moieties, respectively, were synthesized, labeled and purified. The BB2r binding affinities, externalization and protein association properties of these radioconjugates were assessed using the BB2r-positive PC-3 human prostate cancer cell line under hypoxic and normoxic environments. The in vivo biodistribution and microSPECT/CT imaging of the 111In-1, 111In-2 and 111In-4 radioconjugates were investigated in PC-3 tumor bearing SCID mice. Results All conjugates and natIn-conjugates demonstrated nanomolar binding affinities. 111In-1-4 demonstrated 41.4, 60.7, 69.1 and 69.4 % retention, correspondingly, of internalized radioactivity under hypoxic conditions relative to 34.8, 35.3, 33.2 and 29.7 % retention under normoxic conditions. Protein-association studies showed significantly higher levels of association under hypoxic conditions for 2-nitroimidazole containing BB2r-targeted radioconjugates compared to control. Based on the initial 1 hour uptake in the PC-3 tumors, 111In-1, 111In-2 and 111In-4 demonstrated tumor retentions of 1.5, 6.7 and 21.0%, respectively, by 72 h post-injection. Micro-SPECT/CT imaging studies of 111In-1, 111In-2 and 111In-4 radioconjugates resulted in clear delineation of the tumors. Conclusion Based on the in vitro and in vivo studies, the BB2r-targeted agents that incorporated 2-nitroimidazole moieties demonstrated improved retention. These results indicate that further exploration into the potential of hypoxia-selective trapping agents for BB2r-targeted agents, as well as other targeted compounds, is warranted.

ABSTRACT:The detrimental effect of cell adhesion on polymer surfaces has been a limiting factor in the medical deployment of many implants. We examined the potential to decrease cell proliferation while simultaneously increasing mechanical performance through Zn-Al layered double hydroxide (LDH) organically modified with ibuprofen dispersed in poly(L-lactic acid) (PLLA). These composites are commonly referred to as nanocomposites. The thermophysical and mechanical properties of the hybrids were studied with wide-angle X-ray diffraction (WAXD), transmission electron microscopy (TEM), differential scanning calorimetry, thermogravimetric analysis, dynamic mechanical analysis, and tensile testing. The WAXD and TEM results indicated that intercalated and exfoliated nanocomposites were obtained. The storage modulus, tensile modulus, and ultimate tensile strength were improved. The LDH affected the cold crystallization and reduced the thermal stability of the neat PLLA. Smooth muscle cells were used for in vitro studies of the nanocomposites. It was found that the hybrids reduced cell proliferation, and the amount of cell reduction was related to ibuprofen release.