Accumulation In Normal Tissues Research Articles

Formulation of injectable glycyrrhizic acid-hydroxycamptothecin micelles as new generation of DNA topoisomerase I inhibitor for enhanced antitumor activity.

To develop a new drug delivery system is one of the useful approaches to break through the limitation of hydroxycamptothecin (HCPT), a typical DNA topoisomerase I (Topo I) inhibitor in clinical appliance. Injectable glycyrrhizic acid-hydroxycamptothecin (GL-HCPT) micelles that were able to dramatically improve the solubility and stability of HCPT were prepared through self-assembly process and evaluated both in vitro and in vivo. With a mean particle size (PS) of 105.7 ± 9.7 nm and a drug loading (DL) of 9.0 ± 1.5%, GL-HCPT micelles were rapidly internalized by HepG2 cells after 1 h, significantly increasing the intracellular accumulation of HCPT. Compared with the current used HCPT injection and HCPT/GL physical mixture, GL-HCPT micelles showed enhanced antitumor activity against liver cancer cells (HepG2 and Huh7) as well as a superior suppression on the tumor growth of HepG2 tumor bearing mice. Interestingly, GL-HCPT micelles gathered in liver and simultaneously reduced the drug accumulation in normal tissues, thereby exhibiting minimal cytotoxicity to human normal liver cells (LO2). Therefore, we offered a convenient and cost-effective strategy to construct an intravenous drug delivery system (GL-HCPT micelles) as new generation of DNA Topo I inhibitor for enhanced cancer chemotherapy.

Renal Clearable Luminescent Gold Nanoparticles: From the Bench to the Clinic.

With more and more engineered nanoparticles (NPs) being translated to the clinic, the United States Food and Drug Administration (FDA) has recently issued the latest draft guidance on nanomaterial-containing drug products with an emphasis on understanding their in vivo transport and nano-bio interactions. Following these guidelines, NPs can be designed to target and treat diseases more efficiently than small molecules, have minimum accumulation in normal tissues, and induce minimum toxicity. In this Minireview, we integrate this guidance with our ten-year studies on developing renal clearable luminescent gold NPs. These gold NPs resist serum protein adsorption, escape liver uptake, target cancerous tissues, and report kidney dysfunction at early stages. At the same time, off-target gold NPs can be eliminated by the kidneys with minimum accumulation in the body. Additionally, we identify challenges to the translation of renal clearable gold NPs from the bench to the clinic.

Renal Clearable Luminescent Gold Nanoparticles: From the Bench to the Clinic

AbstractWith more and more engineered nanoparticles (NPs) being translated to the clinic, the United States Food and Drug Administration (FDA) has recently issued the latest draft guidance on nanomaterial‐containing drug products with an emphasis on understanding their in vivo transport and nano–bio interactions. Following these guidelines, NPs can be designed to target and treat diseases more efficiently than small molecules, have minimum accumulation in normal tissues, and induce minimum toxicity. In this Minireview, we integrate this guidance with our ten‐year studies on developing renal clearable luminescent gold NPs. These gold NPs resist serum protein adsorption, escape liver uptake, target cancerous tissues, and report kidney dysfunction at early stages. At the same time, off‐target gold NPs can be eliminated by the kidneys with minimum accumulation in the body. Additionally, we identify challenges to the translation of renal clearable gold NPs from the bench to the clinic.

Surface Modifier Effects on Gold Nanoprobe for the Assay of Matrix Metalloproteinases

AbstractTo directly compare the surface modifier effects on the property regulation of nanoparticle (NP)‐based systems, a fluorescence quenched nanoprobe (GpF) responsive to matrix metalloproteinases (MMPs) is employed as a demonstration. Polyethylene glycol (PEG) and bovine serum albumin (BSA) are modified on the surface of GpF via different modes, the SAu covalent bond and nonspecific interaction, respectively. PEG and BSA display similarly protective capability to GpF and increase their biocompatibility that are demonstrated in stability, cytotoxicity, and hemolysis studies. Although PEGylated NPs show better in vivo long circulation property than BSA‐decorated ones, the merit of BSA in controlling the system size in a relatively small range improves the penetration of GpF‐BSA in tumor tissues and reduces the accumulation in normal tissues. Through an MMP‐dominated catalysis and then a glutathione‐assisted replacement process, the fluorescence recovery of GpF‐BSA shows higher efficiency than PEG‐modified NPs. It can be explained by the low steric hindrance and exchangeable manner of BSA on the surface of GpF. The results in this work demonstrate that an endogenous protein having dysopsonization function will be a promising material for the construction and surface modification of nanosystems for biomedical applications.

Implementation of a proton pump inhibitor step-down program within a community-owned health system

Sigma-1 receptor is a target for tumor imaging. In a previous study, we synthesized a vesamicol analog, (+)-2-[4-(4-bromophenyl)piperidino]cyclohexanol [(+)-pBrV], with a high affinity for sigma-1 receptor, and synthesized radiobrominated (+)-pBrV. This radiobrominated (+)-pBrV showed high tumor uptake in tumor-bearing mice; however, radioactivity accumulation in normal tissues, such as the liver, was high. We assumed that the accumulation of (+)-pBrV in the non-target tissues was partially derived from its high lipophilicity; therefore, we synthesized and evaluated (+)-4-[1-(2-hydroxycyclohexyl)piperidine-4-yl]-2-bromophenol [(+)-BrV-OH], which is a more hydrophilic compound. Although we aimed to develop a PET tracer using 76Br, in these initial studies, we used 77Br because of its longer half-life.(+)-[77Br]BrV-OH was synthesized using the chloramine-T method with a radiochemical purity of 95%. Lipophilicity and affinity for sigma-1 receptor of (+)-[77Br]BrV-OH were determined, and biodistribution experiments were performed. We also performed an in vivo blocking study by co-injecting excess amounts of the sigma-1 receptor ligand, SA4503, into mice.The lipophilicity and affinity for sigma-1 receptor of (+)-[77Br]BrV-OH were lower than those of (+)-[77Br]pBrV. (+)-[77Br]BrV-OH also showed high tumor uptake in biodistribution experiments in DU-145 tumor-bearing mice,. Although (+)-[77Br]pBrV was retained in most tissues, (+)-[77Br]BrV-OH was cleared from these tissues. In blocking studies, the co-injection of SA4503 significantly decreased the tumor uptake of (+)-[77Br]BrV-OH.These results indicate that (+)-[76Br]BrV-OH has potential as a PET probe for sigma-1 receptor imaging.

Development of a novel radiobromine-labeled sigma-1 receptor imaging probe

IntroductionSigma-1 receptor is a target for tumor imaging. In a previous study, we synthesized a vesamicol analog, (+)-2-[4-(4-bromophenyl)piperidino]cyclohexanol [(+)-pBrV], with a high affinity for sigma-1 receptor, and synthesized radiobrominated (+)-pBrV. This radiobrominated (+)-pBrV showed high tumor uptake in tumor-bearing mice; however, radioactivity accumulation in normal tissues, such as the liver, was high. We assumed that the accumulation of (+)-pBrV in the non-target tissues was partially derived from its high lipophilicity; therefore, we synthesized and evaluated (+)-4-[1-(2-hydroxycyclohexyl)piperidine-4-yl]-2-bromophenol [(+)-BrV-OH], which is a more hydrophilic compound. Although we aimed to develop a PET tracer using 76Br, in these initial studies, we used 77Br because of its longer half-life. Methods(+)-[77Br]BrV-OH was synthesized using the chloramine-T method with a radiochemical purity of 95%. Lipophilicity and affinity for sigma-1 receptor of (+)-[77Br]BrV-OH were determined, and biodistribution experiments were performed. We also performed an in vivo blocking study by co-injecting excess amounts of the sigma-1 receptor ligand, SA4503, into mice. ResultsThe lipophilicity and affinity for sigma-1 receptor of (+)-[77Br]BrV-OH were lower than those of (+)-[77Br]pBrV. (+)-[77Br]BrV-OH also showed high tumor uptake in biodistribution experiments in DU-145 tumor-bearing mice,. Although (+)-[77Br]pBrV was retained in most tissues, (+)-[77Br]BrV-OH was cleared from these tissues. In blocking studies, the co-injection of SA4503 significantly decreased the tumor uptake of (+)-[77Br]BrV-OH. ConclusionThese results indicate that (+)-[76Br]BrV-OH has potential as a PET probe for sigma-1 receptor imaging.

Interaction of the Plasma Membrane Monoamine Transporter and Organic Cation Transporters with Meta‐iodobenzylguanidine (mIBG)

Meta‐iodobenzylguanidine (mIBG) is a radiopharmaceutical used as both a diagnostic imaging agent (123I‐mIBG) and a targeted frontline radiotherapy (131I‐mIBG) for neuroblastoma. Scintigraphic imaging of mIBG in human and biodistribution studies in xenografted animal models have shown that alongside its uptake into neuroblastoma tumor tissues, mIBG is also significantly transported into normal organs and tissues including the liver, salivary glands, and kidney. 123I‐or 131I‐mIBG accumulation in normal tissues can interfere with its tumor imaging quality and contribute to peripheral toxicities. While mIBG uptake into the neuroblastoma tumors has been shown to be facilitated predominantly by the human norepinephrine transporter (hNET), the molecular mechanisms involved in its uptake into the peripheral tissues have been poorly understood. We hypothesized that the organic cation transporters 1–3 (hOCT1‐3) and the human plasma membrane monoamine transporter (hPMAT) are involved in mIBG transport and accumulation in normal tissues. Thus, the purpose of this project is to characterize the interaction and transport kinetics of mIBG with these transporters and compare them with hNET. This was done by executing uptake assays in human embryonic kidney cells stably expressing hPMAT, hNET, or hOCT1‐3, followed with mIBG quantification by liquid chromatography‐tandem mass spectrometry. Time‐dependent studies showed that mIBG is an excellent substrate of hPMAT, hNET, and hOCT1‐3. Concentration‐dependent studies were fitted to the Michaelis‐Menten equation, resulting in Km values for hPMAT, hNET, and hOCT2 of 42.0 ± 10.7, 35.7 ± 8.91, and 42.9 ± 12.5 μM, respectively, suggesting that mIBG has a similar apparent affinity towards these transporters. Our data demonstrated that mIBG is a novel substrate for hPMAT and hOCTs, providing a molecular basis for these transporters as potential facilitators for mIBG uptake into normal tissues.Support or Funding InformationThis work is supported by NIH Grants GM066233 and T32‐GM007750.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Genetic and epigenetic alterations in normal tissues have differential impacts on cancer risk among tissues

Genetic and epigenetic alterations are both involved in carcinogenesis, and their low-level accumulation in normal tissues constitutes cancer risk. However, their relative importance has never been examined, as measurement of low-level mutations has been difficult. Here, we measured low-level accumulations of genetic and epigenetic alterations in normal tissues with low, intermediate, and high cancer risk and analyzed their relative effects on cancer risk in the esophagus and stomach. Accumulation of genetic alterations, estimated as a frequency of rare base substitution mutations, significantly increased according to cancer risk in esophageal mucosae, but not in gastric mucosae. The mutation patterns reflected the exposure to lifestyle risk factors. In contrast, the accumulation of epigenetic alterations, measured as DNA methylation levels of marker genes, significantly increased according to cancer risk in both tissues. Patients with cancer (high-risk individuals) were precisely discriminated from healthy individuals with exposure to risk factors (intermediate-risk individuals) by a combination of alterations in the esophagus (odds ratio, 18.2; 95% confidence interval, 3.69-89.9) and by only epigenetic alterations in the stomach (odds ratio, 7.67; 95% confidence interval, 2.52-23.3). The relative importance of epigenetic alterations upon genetic alterations was 1.04 in the esophagus and 2.31 in the stomach. The differential impacts among tissues will be critically important for effective cancer prevention and precision cancer risk diagnosis.

Ultrasound-mediated delivery and distribution of polymeric nanoparticles in the normal brain parenchyma of a metastatic brain tumour model.

The treatment of brain diseases is hindered by the blood-brain barrier (BBB) preventing most drugs from entering the brain. Focused ultrasound (FUS) with microbubbles can open the BBB safely and reversibly. Systemic drug injection might induce toxicity, but encapsulation into nanoparticles reduces accumulation in normal tissue. Here we used a novel platform based on poly(2-ethyl-butyl cyanoacrylate) nanoparticle-stabilized microbubbles to permeabilize the BBB in a melanoma brain metastasis model. With a dual-frequency ultrasound transducer generating FUS at 1.1 MHz and 7.8 MHz, we opened the BBB using nanoparticle-microbubbles and low-frequency FUS, and applied high-frequency FUS to generate acoustic radiation force and push nanoparticles through the extracellular matrix. Using confocal microscopy and image analysis, we quantified nanoparticle extravasation and distribution in the brain parenchyma. We also evaluated haemorrhage, as well as the expression of P-glycoprotein, a key BBB component. FUS and microbubbles distributed nanoparticles in the brain parenchyma, and the distribution depended on the extent of BBB opening. The results from acoustic radiation force were not conclusive, but in a few animals some effect could be detected. P-glycoprotein was not significantly altered immediately after sonication. In summary, FUS with our nanoparticle-stabilized microbubbles can achieve accumulation and displacement of nanoparticles in the brain parenchyma.

Cowpox Virus: A New and Armed Oncolytic Poxvirus

Oncolytic virus therapy has recently been recognized as a promising new therapeutic approach for cancer treatment. In this study, we are proposing for the first time to evaluate the in vitro and in vivo oncolytic capacities of the Cowpox virus (CPXV). To improve the tumor selectivity and oncolytic activity, we developed a thymidine kinase (TK)-deleted CPXV expressing the suicide gene FCU1, which converts the non-toxic prodrug 5-fluorocytosine (5-FC) into cytotoxic 5-fluorouracil (5-FU) and 5-fluorouridine-5′-monophosphate (5-FUMP). This TK-deleted virus replicated efficiently in human tumor cell lines; however, it was notably attenuated in normal primary cells, thus displaying a good therapeutic index. Furthermore, this new recombinant poxvirus rendered cells sensitive to 5-FC. In vivo, after systemic injection in mice, the TK-deleted variant caused significantly less mortality than the wild-type strain. A biodistribution study demonstrated high tumor selectivity and low accumulation in normal tissues. In human xenograft models of solid tumors, the recombinant CPXV also displayed high replication, inducing relevant tumor growth inhibition. This anti-tumor effect was improved by 5-FC co-administration. These results demonstrated that CPXV is a promising oncolytic vector capable of expressing functional therapeutic transgenes.

Shear-responsive injectable supramolecular hydrogel releasing doxorubicin loaded micelles with pH-sensitivity for local tumor chemotherapy

In this study, glycol chitosan-Pluronic F127 conjugate (GC-PF127), produced by an amidation reaction between terminal-carboxylated PF127 and glycol chitosan (GC), was used to prepare doxorubicin (DOX)-loaded micelles. The DOX/GC-PF127 micelles produced at optimal conditions had sizes of about 150nm and pH-sensitive surface charges. DOX/GC-PF127 hydrogel formed after addition of α-cyclodextrin into DOX/GC-PF127 micelle solution. The hydrogel had good shear-responsive, injectable and rapid recovery properties. In vitro release experiment confirmed that the hydrogel could sustainedly release DOX/GC-PF127 micelles via the dissociation of the hydrogel. After peritumoral injection into H22 tumor-bearing mice, the hydrogel could greatly increase DOX accumulation in tumor tissue and synchronously avoid DOX accumulation in normal tissues including heart. At similar total DOX dose administrated, the tumors of free DOX treatment group grew slowly after thrice intravenous injections, the tumors of the micelle group did not grow after twice intravenous injections, and the tumors of the hydrogel group disappeared almost after once peritumoral injection. This study demonstrates that injectable DOX/GC-PF127 hydrogel, which can sustainedly release DOX-loaded micelles with tumor-targeting function, is a promising system for local tumor chemotherapy.

MnO2 Gatekeeper: An Intelligent and O2‐Evolving Shell for Preventing Premature Release of High Cargo Payload Core, Overcoming Tumor Hypoxia, and Acidic H2O2‐Sensitive MRI

Premature leakage of photosensitizer (PS) from nanocarriers significantly reduces the accumulation of PS within a tumor, thereby enhancing nonspecific accumulation in normal tissues, which inevitably leads to a limited efficacy for photodynamic therapy (PDT) and the enhanced systematic phototoxicity. Moreover, local hypoxia of the tumor tissue also seriously hinders the PDT. To overcome these limitations, an acidic H2O2‐responsive and O2‐evolving core–shell PDT nanoplatform is developed by using MnO2 shell as a switchable shield to prevent the premature release of loaded PS in core and elevate the O2 concentration within tumor tissue. The inner core SiO2‐methylene blue obtained by co‐condensation has a high PS payload and the outer MnO2 shell shields PS from leaking into blood after intravenous injection until reaching tumor tissue. Moreover, the shell MnO2 simultaneously endows the theranostic nanocomposite with redox activity toward H2O2 in the acidic microenvironment of tumor tissue to generate O2 and thus overcomes the hypoxia of cancer cells. More importantly, the Mn(ΙΙ) ion reduced from Mn(ΙV) is capable of in vivo magnetic resonance imaging selectively in response to overexpressed acidic H2O2. The facile incorporation of the switchable MnO2 shell into one multifunctional diagnostic and therapeutic nanoplatform has great potential for future clinical application.

191 - Redox-Active Mn Porphyrins, MnTE-2-PyP5+ and MnTnBuOE-2-PyP5+ But Not Redox-Inert MnTBAP3-Suppress Tumor Growth in an Environment Where H2O2 Is Produced

Cationic Mn(III) N-substituted pyridylporphyrins (MnPs), such as MnTnBuOE-2-PyP5+ and MnTE-2-PyP5+, initially optimized as powerful mimics of superoxide dismutase, also impact cellular redox-based signaling pathways, most so redox sensitive thiol bearing proteins such as NF-κB, whereby affecting cellular apoptotic and proliferative pathways. Such actions lead to S-glutathionylation of proteins and their subsequent inactivation. In addition to MnP catalyst, both H2O2 and glutathione are required for such process, which includes initial oxidation of Mn porphyrin by H2O2, followed by oxidation of protein thiols by high-valent MnP in a GPx-like fashion. While tumors already have higher H2O2 levels, they are sensitive to any further increase in H2O2 which strategy is used to induce tumor killing. Mn porphyrins do not differentiate tumor from normal tissue and thereby interact at identical rate constants with small reactive species and proteins in both types of tissues. Yet, the yield of reactions depends upon the MnPs levels and levels of reactive species. In the case of NF-kB, the higher are the levels of reactants (MnPs and reactive species) the larger would be the yield of its oxidative damage. Consequently, the larger would be the inhibition of NF-κB activation and in turn the suppression of anti-apoptotic pathways (for involvement of NF-κB pathways in MnP/Asc anticancer effects see Tovmasyan et al abstract on Redox Proteomics). Using LC-MS/MS we showed the preferential accumulation of 3 different MnPs in tumor over normal tissue in a 4T1 flank mouse tumor model. In such model, the already high tumor H2O2 levels were increased with either radiation (RT) or ascorbate or both. The higher tumor accumulation of MnTE-2-PyP5+ relative to MnTnBuOE-2-PyP5+ resulted in larger tumor suppression. Regardless of much higher tumor vs normal tissue accumulation, the redox-inert MnTBAP3- sensitized tumor neither to ascorbate nor to radiation. As anticipated the tumor GSSG levels were increased only when redox-active MnTE-2-PyP5+, but not MnTBAP3-, was combined with RT/Ascorbate treatment. Grant support NIH 1R03-NS082704-01, 5-P30-CA14236-29, CNPq and CAPES from Brazil.

Zinc finger-inspired nanohydrogels with glutathione/pH triggered degradation based on coordination substitution for highly efficient delivery of anti-cancer drugs

Biodegradable materials used for drug delivery are of great demand due to their ability to degrade into low molecular weight species and further excrete from the body by metabolism. Herein, we report a new kind of zinc(II) crosslinked poly(methacrylic acid) nanohydrogels (ZCLNs) inspired by zinc finger proteins with dual stimuli-triggered degradation (glutathione and pH) for the first time. Compared with the disulfide bond crosslinked nanohydrogels, this new kind of ZCLNs is beneficial to the degradation of a wide range of cells, including normal cells. Ex vivo fluorescence images showed that the DOX-loaded folate-PEG conjugated zinc(II)-crosslinked polymeric nanohydrogels (FPZCLNs-15) preferentially accumulated in tumor tissue and the accumulation in normal tissues was much less compared with DOX-loaded PZCLNs-15 (non-targeted nanohydrogels) and free DOX, the FPZCLNs-15 (targeting system) delivered DOX to the tumor site with approximately 3.6- and 1.6-fold higher than free DOX and PZCLNs-15, respectively. Meanwhile, the PZCLNs-15 and FPZCLNs-15 reduced the concentration of DOX in the heart by 3.2- and 5.0-fold respectively, as compared to the free DOX. Moreover, a superior tumor growth inhibition and negligible damage to normal organs like the heart and kidney, which is reported to be vulnerable to DOX-associated side effects, are further demonstrated.

Monoamine oxidase A inhibitor-near-infrared dye conjugate reduces prostate tumor growth.

Development of anti-cancer agents with high tumor-targeting specificity and efficacy is critical for modern multidisciplinary cancer research. Monoamine oxidase A (MAOA), a mitochondria-bound enzyme, degrades monoamine neurotransmitters and dietary monoamines. Recent evidence suggests a correlation between increased MAOA expression and prostate cancer (PCa) progression with poor outcomes for patients. MAOA induces epithelial-mesenchymal transition (EMT) and augments hypoxic effects by producing excess reactive oxygen species. Thus, development of MAOA inhibitors which selectively target tumors becomes an important goal in cancer pharmacology. Here we describe the design, synthesis, and in vitro and in vivo evaluation of NMI, a conjugate that combines a near-infrared dye for tumor targeting with the moiety derived from the MAOA inhibitor clorgyline. NMI inhibits MAOA with low micromolar IC50, suppresses PCa cell proliferation and colony formation, and reduces migration and invasion. In mouse PCa xenografts, NMI targets tumors with no detectable accumulation in normal tissues, providing effective reduction of the tumor burden. Analysis of tumor specimens shows reduction in Ki-67(+) and CD31(+) cells, suggesting a decrease of cell proliferation and angiogenesis and an increase in M30(+) cells, indicating increased apoptosis. Gene expression profiles of tumors treated with NMI demonstrate reduced expression of oncogenes FOS, JUN, NFKB, and MYC and cell cycle regulators CCND1, CCNE1, and CDK4/6, along with increases in the levels of tumor suppressor gene TP53, cell cycle inhibitors CDKN1A and CDKN2A, and MAOA-downstream genes that promote EMT, tumor hypoxia, cancer cell migration, and invasion. These data suggest that NMI exerts its effect through tumor-targeted delivery of a MAOA-inactivating group, making NMI a valuable anti-tumor agent.

In vivo radioimaging of bradykinin receptor b1, a widely overexpressed molecule in human cancer.

The bradykinin receptor B1R is overexpressed in many human cancers where it might be used as a general target for cancer imaging. In this study, we evaluated the feasibility of using radiolabeled kallidin derivatives to visualize B1R expression in a preclinical model of B1R-positive tumors. Three synthetic derivatives were evaluated in vitro and in vivo for receptor binding and their ability to visualize tumors by PET. Enalaprilat and phosphoramidon were used to evaluate the impact of peptidases on tumor visualization. While we found that radiolabeled peptides based on the native kallidin sequence were ineffective at visualizing B1R-positive tumors, peptidase inhibition with phosphoramidon greatly enhanced B1R visualization in vivo. Two stabilized derivatives incorporating unnatural amino acids ((68)Ga-SH01078 and (68)Ga-P03034) maintained receptor-binding affinities that were effective, allowing excellent tumor visualization, minimal accumulation in normal tissues, and rapid renal clearance. Tumor uptake was blocked in the presence of excess competitor, confirming that the specificity of tumor accumulation was receptor mediated. Our results offer a preclinical proof of concept for noninvasive B1R detection by PET imaging as a general tool to visualize many human cancers.

Preliminary radioimmunoimaging and biodistribution of 131iodinelabeled single-chain antibody fragment against progastrin-releasing peptide(31–98) in small cell lung cancer xenografts

Background Monoclonal antibodies (mAbs) such as DD3, raised against progastrin-releasing peptide(31–98) (ProGRP(31–98)) antigen, have been used to target small cell lung cancer (SCLC). However, as an intact mAb, DD3 is cleared slowly from the body, with an optimal radioimmunoimaging time of 72 hours. More recently, a single-chain antibody fragment has demonstrated reduced excretion time in blood and normal tissues and is increasingly used in diagnostic cancer research. Thereby, it potentially increases the radioimmunoimaging efficacy. However, there have been few studies with this antibody fragment. The aim of this study was to characterize the preliminary radioimmunoimaging and biodistribution of 131I-anti-ProGRP(31–98) scFv in nude mice bearing SCLC xenografts. Methods Anti-ProGRP(31–98) scFv was used to detect ProGRP expression by flow cytometry analysis and immunohistochemistry. 131I-anti-ProGRP(31–98) scFv was injected intravenously into healthy Kunming mice and the percentage injected dose per gram (%ID/g) in various organs was calculated. Similarly, the %ID/g and tumor/non-tumor ratio in xenograft-bearing mice was calculated. After injection of 131I-anti-ProGRP(31–98) scFv, treated mice were imaged at 1, 24, and 30 hours. Then the tumor/base ratios were calculated. Results ProGRP was highly expressed in NCI-H446 cells and xenograft tissue. The metabolism of 131I-anti-ProGRP(31–98) scFv in healthy mice was consistent with a first-order and two-compartment model; T1/2α and T1/2β were 10.2 minutes and 5 hours 18 minutes, respectively. The %ID/g of 131I-anti-ProGRP(31–98) scFv in xenografts was much higher than in healthy tissues at 12 hours after injection, reaching a maximum of (5.38±0.92) %ID/g at 24 hours. Successful imaging of xenograft tissue was achieved as early as 1 hour post-injection and persisted until 30 hours, with 24 hours proving optimal. Conclusion 131I-anti-ProGRP(31–98) scFv shows highly selective tumor uptake with low accumulation in normal tissues and rapid blood clearance, indicating that it could be a promising agent for SCLC radioimmunoimaging.

A Rationally Designed A34R Mutant Oncolytic Poxvirus: Improved Efficacy in Peritoneal Carcinomatosis

Oncolytic poxviruses have demonstrated initial promising results in patients with cancer in clinical trials, yet further improvements are needed. It has been shown that a single point mutation in the A34R gene resulted in the production of more total progeny virus and more extracellular enveloped virus (EEV), a form that can be immune-evasive and with enhanced spread. We have genetically engineered a new oncolytic poxvirus (designated vA34R) by incorporating this mutated A34R gene into a viral backbone (vvDD) which was designed for tumor-selective replication. This rationally designed virus can evade neutralization from antipoxvirus antibodies and is highly cytotoxic to cancer cells. It demonstrates improved spread and increased replication within the peritoneal cavity resulting in improved antitumor effects in a peritoneal carcinomatosis (PC) model of MC38 colon cancer. Impressively, after carrier cell-mediated delivery in the preimmunized host, vA34R displayed high replication in tumor nodules yet low accumulation in normal tissues thus enhancing the therapeutic index leading to 70% long-term cures. These results demonstrate that vA34R gains an enhanced therapeutic index for PC via immune evasion, increased spread, and production of more progeny virus. Thus, vA34R may be a potent oncolytic virus (OV) for patients with PC, even after prior exposure to vaccinia virus (VV).

Pharmacological Characterization of Chemically Synthesized Monomeric phi29 pRNA Nanoparticles for Systemic Delivery

Previous studies have shown that the packaging RNA (pRNA) of bacteriophage phi29 DNA packaging motor folds into a compact structure, constituting a RNA nanoparticle that can be modularized with functional groups as a nanodelivery system. pRNA nanoparticles can also be self-assembled by the bipartite approach without altering folding property. The present study demonstrated that 2'-F-modified pRNA nanoparticles were readily manufactured through this scalable bipartite strategy, featuring total chemical synthesis and permitting diverse functional modularizations. The RNA nanoparticles were chemically and metabolically stable and demonstrated a favorable pharmacokinetic (PK) profile in mice (half-life (T(1/2)): 5-10 hours, clearance (Cl): <0.13 l/kg/hour, volume of distribution (V(d)): 1.2 l/kg). It did not induce an interferon (IFN) response nor did it induce cytokine production in mice. Repeat intravenous administrations in mice up to 30 mg/kg did not result in any toxicity. Fluorescent folate-pRNA nanoparticles efficiently and specifically bound and internalized to folate receptor (FR)-bearing cancer cells in vitro. It also specifically and dose-dependently targeted to FR(+) xenograft tumor in mice with minimal accumulation in normal tissues. This first comprehensive pharmacological study suggests that the pRNA nanoparticle had all the preferred pharmacological features to serve as an efficient nanodelivery platform for broad medical applications.

Abstract 369: Redox nanoparticle for tumor imaging and anticancer drug enhancer

Abstract Recently, a non-invasive in vivo imaging of low pH circumstances has attracted attention as new means of detecting disease and response to clinical treatment. Low pH circumstances, which are an aberration of normal pH homeostasis, are caused by ischemia, infection or inflammation, extracellular acidosis in tumors, wound healing, and so forth. However, there is currently no clinical tool available to capture an image of low pH circumstances in vivo. We have been focusing on preparation of core-shell type nanoparticles by self-assembling of amphiphilic block copolymers. Especially, we have succeeded to prepare pH-sensitive nanoparticles, which disintegrate under acidic environment. Thus, using these nanoparticles, ON-OFF regulation of signals in responce to the acidic environment could be monitored. Since nitroxyl radical is susceptible to both electron spin and nuclear magnetic resonances (ESR and NMR), it is anticipated to utilize as imaging compound. Under in vivo conditions, however, low-molecular-weight nitroxyl radicals pose several problems, such as nonspecific accumulation in normal tissues, preferential renal clearance, and rapid reduction of the nitroxyl radical. To solve these difficulties, we have confined nitroxyl radicals in the core of pH-sensitive nanoparticle (RNP) using a self-assembling amphiphilic block copolymer composed of a hydrophilic poly(ethylene glycol) segment and a hydrophobic polystyrene segment carrying nitroxyl radical in each repeating unit as side chain. The cumulant average diameter of the RNP under neutral pH was about 40 nm, and the RNP collapses under the acidic conditions. At the same time, the RNP emitted an intense ESR and NMR signals. Due to the confinement of nitroxyl radical in the core of the RNP, the blood circulation was extended. The nitroxyl radicals in the nanoparticles showed reduction resistance even in the presence of ascorbic acid. In vitro and in vivo toxicity were also reduced significantly. Thus, we can utilize it as a bioimaging probe, which observe acid environment in vivo. An excessive reactive oxygen species (ROS) causes severe side effects in versatile diseases. For example, ischemia-reperfusion injury is one of the most famous damages by ROS, which are produced after a long ischemic period. Therefore, the protection of organs affected by ROS must be considered in order to minimize the area of damage associated with ischemia-reperfusion. Since nitroxyl radical can react catalytically with ROS, RNP was anticipated to scavenge ROS after reperfusion. The RNP was confirmed to show protective effect to several types of ischemia-reperfusion injuries such as brain, heart and kidney. It is also interested to note that RNP improved efficiency of anticancer drug doxorubicin (DOX) when RNP is administered before DOX administration. Thus, RNP is promising as new nanomedicine for clinical applications for versatile therapy. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 369. doi:10.1158/1538-7445.AM2011-369