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Ultrasound nanodroplets (NDs) have been reported as a promising nanocarrier for siRNA delivery depending on its unique strengths of sonoporation. Presently, common means for NDs-mediated siRNA delivery is through electrostatic interaction, but challenges like cationic toxicity still exist. In this study, we demonstrated a novel strategy to construct negatively charged and ultrasound (US)-responsive O-carboxymethyl chitosan (O-CMS) NDs as a siRNA targeted delivery system through three-way junction of bacteriophage phi29 DNA packaging motor (3WJ-pRNA) nanotechnology. 39nt A10-3.2 aptamer targeting prostate specific membrane antigen (PSMA) and 21nt siRNA against cationic amino acid transporter 1 (siCAT-1) were annealed to 3WJ-pRNA scaffold via complementation with an extended sequence. The cholesterol molecule attached to one branch facilitates the 3WJ-pRNA nanoparticles anchoring onto NDs. The desired O-CMS NDs with siRNA-loading and RNA-aptamer modification (A10-3.2/siCAT-1/3WJ-NDs) were successfully prepared, which were with spherical shapes, core–shell structures and uniform in sizes (198 nm with PDI 0.3). As a main proportion of shell, O-CMC showed a certain anti-tumor effects. In vitro studies demonstrated that A10-3.2/siCAT-1/3WJ-NDs exhibited good contrast-enhanced US imaging, buffering capacity and high bio-safety, were able to deliver siCAT-1 to PSMA-overexpressed prostate cancer cells under US irradiation, thus silence the CAT-1 expression, and consequently suppressing 22RV1 cell proliferation and migration. Taken overall, our findings provide a promising strategy to develop negatively charged and US-responsive NDs for tumor-targeted siRNA delivery.

Encapsulation of water-soluble bioactive compounds for enabling specific accumulation in tumor locations, while avoiding premature clearance and/or degradation in the bloodstream, is one of the main hallmarks in nanomedicine, especially that of NIR fluorescent probes for cancer theragnosis. The herein reported technology furnishes water-dispersible double-walled polyurethane-polyurea hybrid nanocapsules (NCs) loaded with indocyanine green (ICG-NCs), using a versatile and highly efficient one-pot and industrially scalable synthetic process based on the use of two different prepolymers to set up the NCs walls. Flow cytometry and confocal microscopy confirmed that both ICG-loaded NCs internalized in monocyte-derived dendritic cells (moDCs). The in vivo analysis of xenograft A375 mouse melanoma model revealed that amphoteric functionalization of NCs’ surface promotes the selective accumulation of ICG-NCs in tumor tissues, making them promising agents for a less-invasive theragnosis of cancer.

Oncogenic microRNAs are essential components in regulating the gene expression of cancer cells. Especially miR21, which is a major player involved of tumor initiation, progression, invasion and metastasis in several cancers. The delivery of anti-miR21 sequences has significant potential for cancer treatment. Nevertheless, since anti-miR21 sequences are extremely unstable and they need to obtain certain concentration to function, it is intensely difficult to build an effective delivery system for them. The purpose of this work is to construct a self-assembled glutathione (GSH)-responsive system with tumor accumulation capacity for effective anti-miR21 delivery and cancer therapy. A novel drug delivery nanosphere carrying millions of anti-miR21 sequences was developed through the rolling circle transcription (RCT) method. GSH-responsive cationic polymer polyethyleneimine (pOEI) was synthesized to protect the nanosphere from degradation by Dicer or other RNase in normal cells and optimize the pompon-like nanoparticle to suitable size. Dehydroascorbic acid (DHA), a targeting molecule, which is a substrate of glucose transporter 1 (GLUT 1) and highly expressed on malignant tumor cells, was connected to pOEI through PEG, and then the polymer was used for contracting a RNA nanospheres into nanopompons. The anti-miR21 nanopompons showed its potential for effective cancer therapy., Graphical abstract Preparation process, GLUT1-mediated transport and GSH-responsive drug release of DHA-targeting anti-miRNA21 nanopompons were reported. Nanopomponswere first obtained by rolling transformation transcription (RCT) and condensed by tumor-targteing pOEI-PEG-DHA.fx1

Type I interferon (IFN) has been approved as an anticancer agent to treat some malignancies. However, IFNs have a short in vivo half-life, systemic toxicity, and poor biophysical properties, which prevent it from being widely used for cancer therapy. This study aimed to construct recombinant IFN-β-1a mutein immunocytokines that comprise a human epidermal growth factor receptor 2 (HER2)-targeting antibody and IFN-β muteins with an additional glycosylation, which can overcome the limitation of the cytokine itself. Hence, the molecular design aims to 1) enhance productivity and biophysical properties by adding secondary glycosylation in IFN-β, 2) increase the therapeutic index of IFN-β therapy by preferential retention at the tumor by possessing high affinity for HER2-expressing cancer cells, and 3) improve the pharmacokinetics and, thus, the convenience of IFN-β administration. The yield of trastuzumab-IFN-β mutein was higher than that of trastuzumab-wild-type IFN-β in the mammalian cell culture system. Trastuzumab-IFN-β mutein showed similar IFN activity and HER2-targeting ability equivalent to that of IFN-β mutein and trastuzumab, respectively. Trastuzumab-IFN-β mutein directly inhibited the growth of HER2-positive gastric cancer cell lines and was more effective than trastuzumab or IFN-β mutein alone. Trastuzumab-IFN-β mutein and IFN-β mutein displayed enhanced immune cell-mediated cytotoxicity. Collectively, trastuzumab-IFN-β mutein may have indirect immune cell-mediated antitumor effects and direct cell growth inhibitory effects. Tumor-targeting effect of trastuzumab-IFN-β mutein was analyzed using in vivo fluorescence imaging. The accumulation of trastuzumab-IFN-β mutein was observed in HER2-positive tumors rather than other tissues except the liver. To evaluate the both direct tumor growth inhibition effect and indirect immune cell-mediated antitumor effect, we tested the effect of trastuzumab-IFN-β mutein in HER2-positive cancer xenograft models using nude mice or humanized mice. Trastuzumab-IFN-β mutein could significantly enhance tumor regression when compared with trastuzumab or IFN-β mutein. In addition, an increase in tumor-infiltrating lymphocytes was observed in the trastuzumab-IFN-β mutein-treated group, implying that the tumor-targeting IFN-β may have an enhanced antitumor effect through increased immune response. Therefore, targeting IFN-β with an anti-HER2 monoclonal antibody makes the immunocytokine more potent than either agent alone. These novel findings suggest that trastuzumab-IFN-β mutein merits clinical evaluation as a new candidate of anticancer therapeutics.

Diagnostic imaging and radionuclide therapy of prostate (PC) and breast cancer (BC) using radiolabeled gastrin-releasing peptide receptor (GRPR)-antagonists represents a promising approach. We herein propose the GRPR-antagonist based radiotracer [99mTc]Tc-DB15 ([99mTc]Tc-N4-AMA-DGA-DPhe6,Sar11,LeuNHEt13]BBN(6-13); N4: 6-carboxy-1,4,8,11-tetraazaundecane, AMA: aminomethyl-aniline, DGA: diglycolic acid) as a new diagnostic tool for GRPR-positive tumors applying SPECT/CT. The uptake of [99mTc]Tc-DB15 was tested in vitro in mammary (T-47D) and prostate cancer (PC-3) cells and in vivo in T-47D or PC-3 xenograft-bearing mice as well as in BC patients. DB15 showed high GRPR-affinity (IC50 = 0.37 ± 0.03 nM) and [99mTc]Tc-DB15 strongly bound to the cell-membrane of T-47D and PC-3 cells, according to a radiolabeled antagonist profile. In mice, the radiotracer showed high and prolonged GRPR-specific uptake in PC-3 (e.g., 25.56 ± 2.78 %IA/g vs. 0.72 ± 0.12 %IA/g in block; 4 h pi) and T-47D (e.g., 15.82 ± 3.20 %IA/g vs. 3.82 ± 0.30 %IA/g in block; 4 h pi) tumors, while rapidly clearing from background. In patients with advanced BC, the tracer could reveal several bone and soft tissue metastases on SPECT/CT. The attractive pharmacokinetic profile of [99mTc]DB15 in mice and its capability to target GRPR-positive BC lesions in patients highlight its prospects for a broader clinical use, an option currently being explored by ongoing clinical studies.

Jinying Liang,1–3 Xinxin Zhang,2 Yunqiu Miao,2 Juan Li,1 Yong Gan2 1Department of Pharmaceutics, China Pharmaceutical University, Nanjing, People’s Republic of China; 2Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai, People’s Republic of China; 3School of Pharmacy, Xinxiang Medical University, Xinxiang, People’s Republic of China Abstract: The development of noninvasive imaging techniques for the accurate diagnosis of progressive hepatocellular carcinoma (HCC) is of great clinical significance and has always been desired. Herein, a hepatocellular carcinoma cell-targeting fluorescent magnetic nanoparticle (NP) was obtained by conjugating near-infrared fluorescence to the surface of Fe3O4 (NIRF-Fe3O4) NPs, followed by coating the lipids consisting of tumoral hepatocytes-targeting polymer (Gal-P123). This magnetic NP (GPC@NIRF-Fe3O4) with superparamagnetic behavior showed high stability and safety in physiological conditions. In addition, GPC@NIRF-Fe3O4 achieved more specific uptake of human liver cancer cells than free Fe3O4 NPs. Importantly, with superparamagnetic iron oxide and strong NIR absorbance, GPC@NIRF-Fe3O4 NPs demonstrate prominent tumor-contrasted imaging performance both on fluorescent and T2-weighted magnetic resonance (MR) imaging modalities in a living body. The relative MR signal enhancement of GPC@NIRF-Fe3O4 NPs achieved 5.4-fold improvement compared with NIR-Fe3O4 NPs. Therefore, GPC@NIRF-Fe3O4 NPs may be potentially used as a candidate for dual-modal imaging of tumors with information covalidated and directly compared by combining fluorescence and MR imaging. Keywords: dual-imaging, magnetic resonance imaging, hepatocellular carcinoma, tumor-targeting

Conventional cancer chemotherapy presents notable drug side effects due to non-selective action of the chemotherapeutics to normal cells. Nanoparticles decorated with receptor-specific ligands on the surface have shown an important role in improving site-selective binding, retention, and drug delivery to the cancer cells. This review summarizes the recent reported achievements using nanostructured coordination polymers (NCPs) with active targeting properties for cancer treatment in vitro and in vivo. Despite the controversy surrounding the effectivity of active targeting nanoparticles, several studies suggest that active targeting nanoparticles notably increase the selectivity and the cytotoxic effect in tumoral cells over the conventional anticancer drugs and non-targeted nanoparticle platform, which enhances drug efficacy and safety. In most cases, the nanocarriers have been endowed with remarkable capabilities such as stimuli-responsive properties, targeting abilities, or the possibility to be monitored by imaging techniques. Unfortunately, the lack of preclinical studies impedes the evaluation of these unique and promising findings for the translation of NCPs into clinical trials.

Oncolytic viruses represent a promising therapeutic modality, but they have yet to live up to their therapeutic potential. Safety and efficacy concerns impel us to identify least toxic oncolytic agents that would generate durable and multifaceted anti-tumor immune responses to disrupt the tumors. Here we describe a rational engineered oncolytic herpes virus (OVH) that is a selective killer for targeting tumors, has strong safety records, induces complete regression of tumors in multiple tumor models, and elicits potent antitumor immunity. By far, the potential of OVs in promoting the tumor antigen-specific humoral immune responses remains obscure. In this study, we found that effective treatment by OVH induced immunogenic cell death, which facilitates to elicit humoral immune responses. Depletion experiments revealed that B cells were required for maximal antitumor efficacy of oncolytic immunotherapy. Both serum transfer and antibody treatment experiments revealed that endogenous oncolysis-induced antigen-targeting therapeutic antibodies can lead to systemic tumor regression. Our data demonstrate that tumor-targeting immune modulatory properties confer oncolytic OVH virotherapy as potent immunotherapeutic cancer vaccines that can generate specific and efficacious antitumor humoral responses by eliciting endogenous tumor antigen-targeting therapeutic antibodies in situ, resulting in an efficacious and tumor-specific therapeutic effect.

We developed tumor-targeting Salmonella typhimurium (S. typhimurium) A1-R, a facultative anaerobe that is an auxotroph of leucine and arginine. The tumor-targeting efficacy of S. typhimurium A1-R was demonstrated in vivo and vitro using several malignant cell lines including melanoma, sarcoma, glioma, breast, pancreatic, colon, cervical, prostate, and ovarian cancers. Our laboratory also developed a patient-derived orthotopic xenograft (PDOX) model by implanting patient-derived malignant tumor fragments into orthotopic sites in mice. We reviewed studies of S. typhimurium A1-R against recalcitrant cancers. S. typhimurium A1-R was effective against all PDOX tumor models tested and showed stronger efficacies than chemotherapy or molecular-targeting therapy against some tumors. Furthermore, the synergistic efficacy of S. typhimurium A1-R when combined with chemotherapeutic agents, molecular-targeting agents, or recombinant methioninase was also demonstrated. We suggest potential clinical uses of this S. typhimurium A1-R treatment.

// Xiaoxin Zhang 1 , Qiaoqiao Xu 1 , Lirun Yang 1 , Yueyang Lai 1 , Zhuangzhuang Zhang 1 , Chao Han 1 , Chizhou Jiang 1 , Jiahuang Li 1 , Yixin Shi 2 , Zi-Chun Hua 1, 3, 4, 5 1 The State Key Laboratory of Pharmaceutical Biotechnology, School of Life Science and School of Stomatology, Affiliated Stomatological Hospital, Nanjing University, Nanjing, 210023, Jiangsu, China 2 The School of Life Sciences and the Center for Infectious Diseases and Vaccinology at the Biodesign Institute, Arizona State University, Tempe, Arizona, 85287-4501, USA 3 Changzhou High-Tech Research Institute of Nanjing University and Jiangsu TargetPharma Laboratories Inc., Changzhou, 213164, Jiangsu, China 4 The State Key Lab of Natural Medicines, China Pharmaceutical University, Nanjing 210017, China 5 The State Key Laboratory of Bioelectronics, Southeast University, Nanjing 210008, China Correspondence to: Zi-Chun Hua, email: zchua@nju.edu.cn Yixin Shi, email: yixin.shi@asu.edu Keywords: VNP20009, cancer, bacteria, tumor-targeting, immune response Received: April 29, 2016 Accepted: October 14, 2016 Published: November 08, 2016 ABSTRACT VNP20009 is a very effective anti-cancer agent and can specifically target tumors and inhibit tumor growth. It was assumed that the tumor targeting ability of VNP20009 correlated to its anticancer capacity. However, our observation contradicted to this assumption. Three VNP20009 mutant strains ( ΔslyA , ΔSTM3120 and ΔhtrA ) with reduced fitness in normal tissues and unchanged fitness in tumors partially or completely lost their anti-cancer capacities. The genes slyA , STM3120 and htrA were required for survival within macrophages and were indispensable for tumor microenvironment remodeling by VNP20009. The infiltration of immune cells occurred less in the tumors of mice infected with the mutant strains. In addition, the mRNA levels of TNF-α and IL-1β were significantly decreased in the tumors of mice treated with the mutant strains. Our results indicate that the immune responses elicited by bacteria rather than the bacterial titer in tumors play a “decisive” role in VNP20009-mediated bacterial cancer therapy, which provides a novel perspective for the underlying mechanism of bacterial cancer therapy.

Background: Peptide radioligands may serve as radionuclide carriers to tumor sites overexpressing their cognate receptor for diagnostic or therapeutic purposes. Treatment of mice with the neprilysin (NEP)-inhibitor phosphoramidon was previously shown to improve the metabolic stability and tumor uptake of biodegradable radiopeptides. Aiming to clinical translation of this methodology, we herein investigated the impact of the approved pill Entresto, releasing the potent NEP-inhibitor LBQ657 in vivo, on the stability and tumor uptake of two radiopeptides. Methods: The metabolic stability of [99mTc]Tc-DB4 (DB4, N4-Pro-Gln-Arg-Tyr-Gly-Asn-Gln-Trp-Ala-Val-Gly-His-Leu-Nle-NH2) and [111In]In-SG4 (SG4, DOTA-DGlu-Ala-Tyr-Gly-Trp-Nle-Asp-Phe-NH2) was tested in LBQ657/Entresto-treated mice vs. untreated controls. The uptake in gastrin-releasing peptide receptor (GRPR)-, or cholecystokinin subtype 2 receptor (CCK2R)-positive tumors respectively, was compared between LBQ657/Entresto-treated mice and untreated controls. Results: LBQ657/Entresto treatment induced marked stabilization of [99mTc] Tc-DB4 and [111In]In-SG4 in peripheral mice blood, resulting in equally enhanced tumor uptake at 4 h post-injection. Accordingly, the [99mTc]Tc-DB4 uptake of 7.13 &plusmn, 1.76%IA/g in PC-3 tumors increased to 16.17 &plusmn, 0.71/17.50 &plusmn, 3.70%IA/g (LBQ657/Entresto) and the [111In]In-SG4 uptake of 3.07 &plusmn, 0.87%IA/g in A431-CCK2R(+) tumors to 8.11 &plusmn, 1.45/9.61 &plusmn, 1.70%IA/g. Findings were visualized by SPECT/CT. Conclusions: This study has shown the efficacy of Entresto to notably improve the profile of [99mTc]Tc-DB4 and [111In]In-SG4 in mice, paving the way for clinical translation of this approach.

Background Improving the water solubility of hydrophobic photosensitizer and increasing its accumulation in tumor tissue are essential for in vivo photodynamic therapy (PDT). Considering commercialization or clinical application in future, it will be promising to achieve these purposes by developing new agents with simple and non-toxic structure. Results We conjugated multiple chlorin e6 (Ce6) molecules to gelatin polymer, synthesizing two types of gelatin–Ce6 conjugates with different amounts of Ce6: gelatin–Ce6-2 and gelatin–Ce6-8. The resulting conjugates remained soluble in aqueous solutions for a longer time than hydrophobic Ce6. The conjugates could generate singlet oxygen and kill tumor cells upon laser irradiation. After intravenous injection into SCC-7 tumor-bearing mice, gelatin–Ce6-2 showed prolonged blood circulation and highly increased accumulation in tumor tissue as observed in real-time imaging in vivo. After laser irradiation, gelatin–Ce6-2 suppressed tumor growth completely and enabled improved PDT compared to free Ce6 and gelatin–Ce6-8. Conclusions This work demonstrates that a simple structure based on photosensitizer and gelatin can highly improve water solubility and stability. Superior tumor tissue accumulation and increased therapeutic efficacy of gelatin–Ce6 during in vivo PDT showed its high potential for clinical application. Electronic supplementary material The online version of this article (10.1186/s12951-019-0475-1) contains supplementary material, which is available to authorized users.

The present study investigated the feasibility of improving the tumor‑targeting efficacy and decreasing the toxicity of liposomal cabazitaxel (Cab) with aptamer modification. The process involved preparing aptamer (TLS1c)‑modified liposomes and studying the behavior of the liposomes in vitro and in vivo. TLS1c as an aptamer, which has high specificity for BNL 1ME A.7R.1 (MEAR) cells, was conjugated with Cab liposomes (Cab/lipo) to enhance MEAR tumor tissue targeting. Confocal laser scanning microscopy and flow cytometry analyses demonstrated that the fluorescence of the liposomes modified with the aptamer was notably stronger compared with that of the unmodified liposomes. Furthermore, the biodistribution data of the modified liposomes tested in tumor‑bearing mice revealed high specificity of biotinylated TLS1c‑modified Cab/lipo (BioTL‑Cab/lipo) for tumor tissues. Furthermore, the modified liposomes demonstrated decreased cytotoxicity and simultaneously retained potent inhibition against tumor growth. It is likely that the specific binding of the aptamer (TLS1c) to the targeted cells (MEAR) facilitates the binding of the liposomes to the targeted cells. Therefore, BioTL‑Cab/lipo may be considered as a promising delivery system to improve cell targeting and reduce drug toxicity in the treatment of cancer. Published version

// Dom-Gene Tu 1,2,3 , Wen-Wei Chang 4,5 , Song-Tao Lin 6 , Chun-Yu Kuo 7 , Yu-Tzu Tsao 8 and Che-Hsin Lee 6,7,9 1 Department of Nuclear Medicine, Ditmanson Medical Foundation, Chia-Yi Christian Hospital, Chia-Yi, Taiwan 2 Department of Food Science and Technology, Chia Nan University of Pharmacy and Science, Tainan, Taiwan 3 College of Health Sciences, Chang Jung Christian University, Tainan, Taiwan 4 Department of Biomedical Sciences, College of Medical Science and Technology, Chung Shan Medical University, Taichung, Taiwan 5 Department of Medical Research, Chung Shan Medical University Hospital, Taichung, Taiwan 6 Department of Microbiology, School of Medicine, China Medical University, Taichung, Taiwan 7 Graduate Institute of Basic Medical Science, School of Medicine, China Medical University, Taichung, Taiwan 8 Department of Medicine, Taoyuan General Hospital, Taoyuan, Taiwan 9 Department of Biological Sciences, National Sun Yat-sen University, Kaohsiung, Taiwan Correspondence to: Che-Hsin Lee, email: // Keywords : Salmonella, tumor-targeting, angiogenesis, hypoxia-inducible factor-1α, vascular endothelial growth factor Received : August 10, 2015 Accepted : January 19, 2016 Published : January 27, 2016 Abstract Salmonella is a Gram-negative, facultative anaerobe that is a common cause of host intestinal infections. Salmonella grows under aerobic and anaerobic conditions, and it has been proven capable of inhibiting tumor growth. However, the molecular mechanism by which Salmonella inhibits tumor growth is still unclear. Angiogenesis plays an important role in the development and progression of tumors. We investigated the antitumor effect of Salmonella in a syngeneic murine tumor model. Hypoxia-inducible factor-1 (HIF-1)α plays a significant role in tumor angiogenesis. We examined the molecular mechanism by which Salmonella regulated vascular endothelial growth factor (VEGF), which is an important angiogenic factor. The expression of VEGF in tumor cells was decreased by treatment with Salmonella . The conditioned medium from Salmonella -treated cells inhibited the proliferation of endothelial cells. Salmonella inhibited the expression of HIF-1α as well as downregulated its upstream signal mediator protein kinase B (AKT). Salmonella significantly inhibited tumor growth in vivo , and immunohistochemical studies of the tumors revealed decreased intratumoral microvessel density. These results suggest that Salmonella therapy, which exerts anti-angiogenic activities, represents a promising strategy for the treatment of tumors.

Over the past decades, Salmonella has been proven capable of inhibiting tumor growth. It can specifically target tumors and due to its facultative anaerobic property, can be more penetrative than other drug therapies. However, the molecular mechanism by which Salmonella inhibits tumor growth is still incompletely known. The antitumor therapeutic effect mediated by Salmonella is associated with an inflammatory immune response at the tumor site and a T cell-dependent immune response. Many tumors have been proven to have a high expression of indoleamine 2, 3-dioxygenase 1 (IDO), which is a rate-limiting enzyme that catalyzes tryptophan to kynurenine, thus causing immune tolerance within the tumor microenvironment. With decreased expression of IDO, increased immune response can be observed, which might be helpful when developing cancer immunotherapy. The expression of IDO was decreased after tumor cells were infected with Salmonella. In addition, Western blot analysis showed that the expression levels of phospho-protein kinase B (P-AKT), phospho-mammalian targets of rapamycin (P-mTOR), and phospho-p70 ribosomal s6 kinase (P-p70s6K) in tumor cells were decreased after Salmonella infection. In conclusion, our results indicate that Salmonella inhibits IDO expression and plays a crucial role in anti-tumor therapy, which might be a promising strategy combined with other cancer treatments.

Attenuated Salmonella enterica serovar Typhimurium (S. Typhimurium) has been developed as a vector to deliver therapeutic agents to tumors. The potential of S. Typhimurium in cancer therapy is largely due to its reported propensity to accumulate at greater than 1,000-fold higher concentrations in tumors relative to healthy tissues. In this study, we compared bacterial colonization of tumors in a subcutaneous transplantation model with a more clinically relevant autochthonous tumor model. Following intravenous administration of attenuated S. Typhimurium strain SL3261, we observed approximately 10,000-fold less bacteria in autochthonous tumors that sporadically develop in transgenic BALB-neuT mice compared to tumors developed from subcutaneous transplantation of 4T1 murine breast cancer cells in BALB/c mice. Treatment of BALB-neuT mice with a vasculature-disrupting agent (VDA) prior to bacterial treatment caused necrosis of tumor tissue and significantly increased the bacterial targeting of autochthonous tumors by approximately 1,000-fold. These observations emphasize the importance of appropriate model selection in developing bacteria-based cancer therapies and demonstrate the potential of combining VDA pre-treatment with bacteria to facilitate targeting of clinically relevant tumors.

The use of preferentially tumor-targeting bacteria as vectors is one of the most innovative approaches for the treatment of cancer. This method is based on the observation that some obligate or facultative anaerobic bacteria are capable of selectively multiplying in tumors and inhibiting their growth. Previously, we found that the tumor-targeting efficiency of Salmonella could be modulated by modifying the immune response to these bacteria by coating them with poly(allylamine hydrochloride) (PAH), and these organisms are designated PAH-S.C. (S. choleraesuis). PAH can provide a useful platform for the chemical modification of Salmonella, perhaps by allowing a therapeutic gene to bind to tumor-targeting Salmonella. This study aimed to investigate the benefits of the use of PAH-S.C. for gene delivery. To evaluate this modulation, the invasion activity and gene transfer of DNA-PAH-S.C. were measured in vitro and in vivo. Treatment with PAH-S.C. carrying a tumor suppressor gene (connexin 43) resulted in inhibition of tumor growth, which suggested that tumor-targeted gene therapy using PAH-S.C. carrying a therapeutic gene could exert antitumor activities. This technique represents a promising strategy for the treatment of tumors.

In this study, we examined the effect of cationic lipid type in folate (FA)-polyethylene glycol (PEG)-modified cationic liposomes on gene-silencing effects in tumor cells using cationic liposomes/siRNA complexes (siRNA lipoplexes). We used three types of cationic cholesterol derivatives, cholesteryl (3-((2-hydroxyethyl)amino)propyl)carbamate hydroiodide (HAPC-Chol), N-(2-(2-hydroxyethylamino)ethyl)cholesteryl-3-carboxamide (OH-Chol), and cholesteryl (2-((2-hydroxyethyl)amino)ethyl)carbamate (OH-C-Chol), and we prepared three types of FA-PEG-modified siRNA lipoplexes. The modification of cationic liposomes with 1&ndash, 2 mol % PEG-lipid abolished the gene-silencing effect in human nasopharyngeal tumor KB cells, which overexpress the FA receptor (FR). In contrast, FA-PEG-modification of cationic liposomes restored gene-silencing activity regardless of the cationic lipid type in cationic liposomes. However, the optimal amount of PEG-lipid and FA-PEG-lipid in cationic liposomes for selective gene silencing and cellular uptake were different among the three types of cationic liposomes. Furthermore, in vitro transfection of polo-like kinase 1 (PLK1) siRNA by FA-PEG-modified liposomes exhibited strong cytotoxicity in KB cells, compared with PEG-modified liposomes, however, in in vivo therapy, intratumoral injection of PEG-modified PLK1 siRNA lipoplexes inhibited tumor growth of KB xenografts, as well as that of FA-PEG-modified PLK1 siRNA lipoplexes. From these results, the optimal formulation of PEG- and FA-PEG-modified liposomes for FR-selective gene silencing might be different between in vitro and in vivo transfection.

// Takashi Murakami 1, 2, 3 , Kentaro Igarashi 1 , Kei Kawaguchi 1 , Tasuku Kiyuna 1 , Yong Zhang 1 , Ming Zhao 1 , Yukihiko Hiroshima 3 , Scott D. Nelson 5 , Sarah M. Dry 5 , Yunfeng Li 5 , Jane Yanagawa 6 , Tara Russell 6 , Noah Federman 7 , Arun Singh 4 , Irmina Elliott 6 , Ryusei Matsuyama 3 , Takashi Chishima 3 , Kuniya Tanaka 3 , Itaru Endo 3 , Fritz C. Eilber 6 , Robert M. Hoffman 1, 2 1 AntiCancer, Inc., San Diego, California, USA 2 Department of Surgery, University of California, San Diego, California, USA 3 Department of Gastroenterological Surgery, Graduate School of Medicine, Yokohama City University, Yokohama, Japan 4 Division of Hematology-Oncology, University of California, Los Angeles, California, USA 5 Department of Pathology, University of California Los Angeles, California, USA 6 Division of Surgical Oncology, University of California, Los Angeles, California, USA 7 Department of Pediatrics and Department of Orthopaedics, David Geffen School of Medicine, Mattel Children’s Hospital, UCLA’s Jonsson Comprehensive Cancer Center, University of California, Los Angeles, California, USA Correspondence to: Robert M. Hoffman, email: all@anticancer.com Fritz C. Eilber, email: fceilber@mednet.ucla.edu Keywords: osteosarcoma, nude mouse, patient-derived xenograft, Salmonella typhimurium A1-R, tumor-targeting Received: October 20, 2016 Accepted: November 16, 2016 Published: December 20, 2016 ABSTRACT Osteosarcoma occurs mostly in children and young adults, who are treated with multiple agents in combination with limb-salvage surgery. However, the overall 5-year survival rate for patients with recurrent or metastatic osteosarcoma is 20-30% which has not improved significantly over 30 years. Refractory patients would benefit from precise individualized therapy. We report here that a patient-derived osteosarcoma growing in a subcutaneous nude-mouse model was regressed by tumor-targeting Salmonella typhimurium A1-R ( S. typhimurium A1-R, p

Uniform gold nanostars (Au NS) were conjugated with cyclic RGD (cRGD) and near infrared (NIR) fluorescence probe (MPA) or anti-cancer drug (DOX) to obtain multi-functional nanoconstructs, Au-cRGD-MPA and Au-cRGD-DOX respectively. The NIR contrast agent Au-cRGD-MPA was shown to have low cytotoxicity. Using tumor cells and tumor bearing mice, these imaging nanoparticles demonstrated favorable tumor-targeting capability mediated by RGD peptide binding to its over-expressed receptor on the tumor cells. The multi-therapeutic analogue, Au-cRGD-DOX, integrates targeting tumor, chemotherapy and photo-thermotherapy into a single system. The synergistic effect of photo-thermal therapy and chemotherapy was demonstrated in different tumor cell lines and in vivo using S180 tumor-bearing mouse models. The viability of MDA-MB-231 cells was only 40 % after incubation with Au-cRGD-DOX and irradiation with NIR light. Both tail vein and intratumoral injections showed Au-cRGD-DOX treated mice exhibiting the slowest tumor increase. These results indicate that the multifunctional nanoconstruct is a promising combined therapeutic agent for tumor-targeting treatment, with the potential to enhance the anti-cancer treatment outcomes.

Bladder cancer is the second most common urological malignancy in the world. In 70% of cases it is initially diagnosed as non-muscle-invasive bladder cancer (NMIBC) and it is amenable to local treatments, with intravesical (IVES) Bacillus-Calmette-Guerin (BCG) immunotherapy being routinely used after transurethral resection of the lesion. However, this treatment is associated with significant side-effects and treatment failures, highlighting the necessity of novel strategies. One potent approach is the suicide-gene mediated therapy/prodrug combination, provided tumor-specificity can be ensured and anti-tumor immune responses induced. Using the mouse syngeneic orthotopic MB49-bladder tumor model, here we show that IVES human papillomavirus non-replicative pseudovirions (PsV) can pseudoinfect tumors with a ten-fold higher efficacy than normal bladders. In addition, PsV carrying the suicide-gene herpes-simplex virus thymidine kinase (PsV-TK) combined to Ganciclovir (GCV) led to immunogenic cell-death of tumor cells in vitro and to MB49-specific CD8 T-cells in vivo. This was associated with reduction in bladder-tumor growth and increased mice survival. Altogether, our data show that IVES PsV-TK/GCV may be a promising alternative or combinatory treatment for NMIBC.

Human platelets were identified in tumors by Trousseau in 1865, although their roles in tumor microenvironments have only recently attracted the attention of cancer researchers. In this study we exploit and enhance platelet interactions in tumor microenvironments by introducing tumor-targeting and imaging functions. The first step in repurposing human platelets as vehicles for tumor-targeting was to inhibit platelet-aggregation by cytoplasmic-loading of kabiramide (KabC), a potent inhibitor of actin polymerization and membrane protrusion. KabC-Platelets can accumulate high levels of other membrane-permeable cytoxins and probes, including epidoxorubicin, carboxyfluorescein di-ester and chlorin-e6. Finally, mild reaction conditions were developed to couple tumor-targeting proteins and antibodies to KabC-platelets. Fluorescence microscopy studies showed KabC-platelets, surface-coupled with transferrin and Cy5, bind specifically to RPMI8226 and K562 cells, both of which over-express the transferrin receptor. Repurposed platelets circulate for upto 9-days a feature that increases their chance of interacting with target cells. KabC-platelets, surface-coupled with transferrin and Cy7, or chlorin-e6, and injected in immuno-compromised mice were shown to accumulate specifically in sub-cutaneous and intra-cranial myeloma xenotransplants. The high-contrast, in vivo fluorescence images recorded from repurposed platelets within early-stage myeloma is a consequence in part of their large size (φ~2µm), which allows them to transport 100 to 1000-times more targeting-protein and probe molecules respectively. Human platelets can be configured with a plurality of therapeutic and targeting antibodies to help stage tumor environments for an immunotherapy, or with combinations of therapeutic antibodies and therapeutic agents to target and treat cardiovascular and neurologic diseases.

A patient with high grade undifferentiated pleomorphic soft-tissue sarcoma from a striated muscle was grown orthotopically in the right biceps femoris muscle of mice to establish a patient-derived orthotopic xenograft (PDOX) model. Twenty PDOX mice were divided into 4 groups: G1, control without treatment; G2, Salmonella typhimurium (S. typhimurium)A1-R administered by intratumoral (i.t.) injection once a week for 4 weeks; G3, doxorubicin (DOX) administered by intraperitoneal (i.p.) injection once a week for 4 weeks; G4, S. typhimurium A1-R (i.t.) administered once a week for 2 weeks followed by i.p. doxorubicin once a week for 2 weeks. On day 25 from the initiation of treatment, tumor volume in G2, G3, and G4 was significantly lower than G1. Mice found without gross tumor included one mouse (20%) in G2; one mouse (20%) in G3; and 3 mice (60%) in G4. Body weight loss did not significantly differ between the 3 treated groups or from the untreated control. Histological examination revealed eradication of tumor only in G4 where mice were treated with S. typhimurium A1-R followed by DOX. Our present study indicates future clinical potential of combining S. typhimurium A1-R with chemotherapy such as DOX for soft tissue sarcoma patients.

The roles of Matrix MetalloProteinases (MMPs), such as MMP-9, in tumor metastasis are well studied, and this in turns stimulates the development of MMP inhibitors as antitumor agents. Previously, Salmonella accumulation was observed in the metastatic nodules of the lungs after systemic administration. Salmonella significantly enhanced the survival of the pulmonary metastatic tumor-bearing mice. Based on our previous observation, we hypothesized that Salmonella could affect metastasis-related protein expression. The treatment of Salmonella clearly reduced the expression of MMP-9. Meanwhile, the MMP-9 related signaling pathways, including Phosph-Protein Kinase B (P-AKT) and Phosph-mammalian Targets Of Rapamycin (P-mTOR) were decreased after a Salmonella treatment. The Salmonella inhibited tumor cell migration by wound-healing and Transwell assay. The anti-metastatic effects of Salmonella were evaluated in mice bearing experimental metastasis tumor models. Consequently, Salmonella inhibited the expression of MMP-9 by reducing the AKT/mTOR pathway and metastatic nodules in vivo.

Lack of specificity of the therapeutic agent is a primary limitation in the treatment of a tumor. The use of preferentially replicating bacteria as therapeutic agents is an innovative approach to tumor treatment. This is based on the observation that certain obligate or facultative anaerobic bacteria are capable of multiplying selectively in tumors and inhibiting their growth. Bacteria have been employed as antitumor agents that are capable of preferentially amplifying within tumors and inhibiting their growth. Moreover, bacteria-derived factors have an immune-stimulation effect. Therefore, bacteria are able to transfer therapeutic genes into the tumor cells using their infective ability. Herein, we introduce the application of bacteria for tumor therapy and focus on Salmonella, which have been widely used for tumor therapy. Salmonella have mainly been applied as gene-delivery vectors, antitumor immune activators and tumor cell death inducers. This study will not only evaluate the therapeutic efficacy of Salmonella for the treatment of tumor but will also elucidate the mechanisms underlying the antitumor activities mediated by Salmonella, which involve host immune responses and cellular molecular responses.

Hui Lv,1,* Qing Zhu,1,* Kewu Liu,2 Manman Zhu,1 Wenfang Zhao,1 Yuan Mao,1 Kehai Liu1 1Department of Biopharmaceutics, College of Food Science and Technology, Shanghai Ocean University, Shanghai, People's Republic of China; 2Heilongjiang Forest By-Product and Speciality Institute, Mudanjiang, People's Republic of China *These authors contributed equally to this work Background: A degradable polyethylenimine (PEI) derivative coupled to a bifunctional peptide R13 was developed to solve the transfection efficiency versus cytotoxicity and tumor-targeting problems of PEI when used as a gene vector. Methods: We crossed-linked low molecular weight PEI with N-octyl-N-quaternary chitosan (OTMCS) to synthesize a degradable PEI derivative (OTMCS-PEI), and then used a bifunctional peptide, RGDC-Tat (49–57) called R13 to modify OTMCS-PEI so as to prepare a new gene vector, OTMCS-PEI-R13. This new gene vector was characterized by various physicochemical methods. Its cytotoxicity and gene transfection efficiency were also determined both in vitro and in vivo. Results: The vector showed controlled degradation and excellent buffering capacity. The particle size of the OTMCS-PEI-R13/DNA complexes was around 150–250 nm and the zeta potential ranged from 10 mV to 30 mV. The polymer could protect plasmid DNA from being digested by DNase I at a concentration of 23.5 U DNase I/µg DNA. Further, the polymer was resistant to dissociation induced by 50% fetal bovine serum and 400 µg/mL sodium heparin. Compared with PEI 25 kDa, the OTMCS-PEI-R13/DNA complexes showed higher transfection efficiency both in vitro and in vivo. Further, compared with OTMCS-PEI, distribution of OTMCS-PEI-R13 at tumor sites was markedly enhanced, indicating the tumor-targeting specificity of R13. Conclusion: OTMCS-PEI-R13 could be a potential candidate as a safe and efficient gene delivery carrier for gene therapy. Keywords: nonviral gene vector, polyethylenimine, R13, transfection efficiency, tumor-targeting

Jianfeng Liu, Jinjian Liu, Hongyan Xu, Yumin Zhang, Liping Chu, Qingfen Liu, Naling Song, Cuihong YangTianjin Key Laboratory of Molecular Nuclear Medicine, Institute of Radiation Medicine, Chinese Academy of Medical Science and Peking Union Medical College, People's Republic of ChinaAbstract: The poor aqueous solubility and low bioavailability of curcumin restrict its clinical application for cancer treatment. In this study, a novel tumor-targeting nanofiber carrier was developed to improve the solubility and tumor-targeting ability of curcumin using a self-assembled Nap-GFFYG-RGD peptide. The morphologies of the peptide nanofiber and the curcumin-encapsulated nanofiber were visualized by transmission electron microscopy. The tumor-targeting activity of the curcumin-encapsulated Nap-GFFYG-RGD peptide nanofiber (f-RGD-Cur) was studied in vitro and in vivo, using Nap-GFFYG-RGE peptide nanofiber (f-RGE-Cur) as the control. Curcumin was encapsulated into the peptide nanofiber, which had a diameter of approximately 10–20 nm. Curcumin showed sustained-release behavior from the nanofibers in vitro. f-RGD-Cur showed much higher cellular uptake in αvβ3 integrin-positive HepG2 liver carcinoma cells than did non-targeted f-RGE-Cur, thereby leading to significantly higher cytotoxicity. Ex vivo studies further demonstrated that curcumin could accumulate markedly in mouse tumors after administration of f-RGD-Cur via the tail vein. These results indicate that Nap-GFFYG-RGD peptide self-assembled nanofibers are a promising hydrophobic drug delivery system for targeted treatment of cancer.Keywords: nanofiber, tumor-targeting, self-assembling, curcumin, drug delivery