Search

An analytical solution for long waves propagating over a submerged atoll is established. The atolls involved in this study are annular coral reefs with large lagoons in the middle, and the expression of the cross section is a trinomial function of the radial distance, i.e., h=ar2s−brs+h0, where s is the positive rational number. This analytical solution extends the theory by Wang et al. (2018) as s is no longer limited to s=2/m, where m is the positive integer. In addition, by adjusting the terrain parameters properly, the analytic solution can be degenerated to describe the wave propagation over topography with a hump or pit. According to the relationship between wave rays and wave energy, the distribution characteristics and formation mechanism of energy over the topography are expounded. When the lagoon is non-existent, all wave rays converge at the x-axis, which results in an abrupt amplification of the wave amplitude around the convergence point. When a lagoon is mounted on the top of the atoll, the rays are scattered due to the refraction of the lagoon, and only some rays converge at the symmetrical axis and the ridges on both sides, which results in the amplification of wave amplitudes in these areas.

Drug delivery systems based on genetically engineered oncolytic bacteria have properties that cannot be achieved by traditional therapeutic interventions. Thus, they have attracted considerable attention in cancer therapies. Attenuated bacteria can specifically target and actively penetrate tumor tissues and play an important role in cancer suppression as the "factories" of diverse anticancer drugs. Over the past decades, several bacterial strains including Salmonella and Clostridium have been shown to effectively retard tumor growth and metastasis, and thus improve survival in preclinical models or clinical cases. In this review, we summarize the unique properties of oncolytic bacteria and their anticancer mechanisms and highlight the particular advantages compared with traditional strategies. With the current research progress, we demonstrate the potential value of oncolytic bacteria-based drug delivery systems for clinical applications. In addition, we discuss novel strategies of cancer therapies integrating oncolytic bacteria, which will provide hope to further improve and standardize the current regimens in the near future.

Salmonella Typhimurium defective in guanosine 5′-diphosphate-3′-diphosphate (ppGpp) synthesis (ΔppGpp) is an attenuated strain with good biosafety and excellent anticancer efficacy. It has been widely applied in preclinical studies of anticancer therapy for various types of solid cancer. VNP20009 is another genetically modified auxotrophic strain with 108-kb deletion, purI−, msbB−, and many single nucleotide polymorphisms (SNPs); it has shown promising therapeutic efficacy in various preclinical tumor models and entered phase I clinical trials. Here, the invasion activities and virulence of ΔppGpp were obviously lower than those of the VNP20009 strain when tested with cancer cells in vitro. In addition, the MC38 tumor-bearing mice showed comparable cancer suppression when treated with ΔppGpp or VNP20009 intravenously. However, the ΔppGpp-treated mice showed 16.7% of complete cancer eradication and prolonged survival in mice, whereas VNP20009 showed higher toxicity to animals, even with equal tumor size individually. Moreover, we found decreased levels of inflammatory cytokines in circulation but strengthened immune boost in tumor microenvironments of ΔppGpp-treated mice. Therefore, the engineered ΔppGpp has high potential for cancer therapeutics, and it is a promising option for future clinical cancer therapy.

Genetically engineered Salmonella typhimurium can specifically colonize tumor tissues and drastically inhibit tumor growth. Vibrio vulnificus flagellin B (FlaB), a natural ligand of Toll-like receptor 5 (TLR5) that can activate robust host immune system, is an excellent adjuvant for cancer immunotherapy with high binding affinity to TLR5. Here, we constructed attenuated S. typhimurium that expresses flagellin B (FlaB) with a controlled expression system to enhance targeted cancer immunotherapy with increased good safety profiles. Visualized therapy can also be achieved with bioluminescence imaging by introducing the lux operon into the attenuated Salmonella.

Genetically engineered bacterial cancer therapy presents several advantages over conventional therapies. However, the anticancer effects of bacterium-based therapies remain insufficient, and serious side effects may be incurred with the increase in therapeutic dosages. Photodynamic therapy (PDT) suppresses tumor growth by producing reactive oxygen species (ROS) through specific laser-activated photosensitizers (PSs). Tumor-specific PS delivery and activatable ROS generation are two critical aspects for PDT advancement. Here, we propose PDT-enhanced oncolytic bacterial immunotherapy (OBI) by using genetically engineered avirulent Salmonella expressing a fluorogen-activating protein (FAP). Upon binding to a fluorogen, FAP could be activated and generate fluorescence and ROS. The instant activation of persistent fluorescence was detected in tumors through bacterium-based imaging. In a colon cancer model, PDT-OBI showed an enhanced tumor inhibition effect and prolonged animal survival. Mechanically, PDT generated ROS, resulting in the killing of cancer cells and over-accumulated bacteria. The pathogen-associated molecular patterns and damage-associated molecular patterns released from the destroyed bacteria and cancer cells recruited and activated immune cells (macrophages, neutrophils, and dendritic cells), which released additional proinflammatory cytokines (TNF-α and IL-1β); reduced anti-inflammatory cytokines (IL-10); and further enhanced immune cell infiltration in a positive-feedback manner, thus reducing bacterium-induced side effects and improving anticancer activities. This synergistic therapy has promising potential for cancer immunotherapy.

This study aimed at investigating the feasibility of bioluminescence imaging (BLI) with engineered Salmonella typhimurium (ΔppGpp S. typhimurium) for visualizing acute hypoxic/ischemic bowels. At the start of 12- or 24-h reperfusion, ΔppGpp S. typhimurium was injected into the lateral tail veins of rats in which three segments of the small intestine were respectively subjected to 2, 3, and 4 h of ischemia. BLI and magnetic resonance imaging were performed at each reperfusion time point. Bioluminescence was exclusively detected in the hypoxic/ischemic segment of the intestine, showing the ability of ΔppGpp S. typhimurium to specifically target and proliferate in a hypoxic/ischemic area. Serial monitoring of these rat models revealed a progressive increase in bacterial bioluminescence in the ischemic intestines in conjunction with viable bacterial counts. The viable bacterial counts were positively correlated with lactate dehydrogenase levels after 24 h of reperfusion following 3 or 4 h of ischemia as well as interleukin-6 levels after 24 h of reperfusion following 4 h of ischemia. Our findings demonstrated that BLI was able to detect the acute hypoxic/ischemic bowel via monitoring of the distribution, internalization, and activity of administered ΔppGpp S. typhimurium. These findings may be useful for the early diagnosis of ischemic bowel disease., Graphical Abstract, Genetically engineered bacteria, ΔppGpp S. typhimurium, showed the ability to specifically target and proliferate in acute hypoxic/ischemic bowel, which could be detected via monitoring of the distribution, internalization, and activity of administered ΔppGpp S. typhimurium using bioluminescence imaging. These findings may be useful for early diagnosis of acute mesenteric ischemia.

This paper presents a semi-Lagrangian meshless framework based on the localized radial basis function collocation method and the second-order explicit Runge–Kutta method (RK2-LRBFCM) for analyzing the sloshing phenomenon in a 2D tank. In the present method, the semi-Lagrangian approach is adopted to constrain the lateral movements of inner collocation nodes. And the LRBFCM is used to obtain the velocity potentials at each time instant governed by Laplace equation, then second-order explicit Runge–Kutta method is introduced to calculate the free-surface elevation and velocity potential at the next time instant. In comparison with the reference results, the efficiency and accuracy of the proposed RK2-LRBFCM are demonstrated in the solutions of several benchmark examples including horizontally excited tanks, vertically excited tanks, horizontally and vertically excited tanks. Moreover, the effect of the protrusion (shape and size) of the bottom wall on the sloshing phenomenon in horizontally excited tank is investigated.

Pancreatic cancer (PC) is a cancer of the digestive system, and pancreatic ductal adenocarcinoma (PDAC) accounts for approximately 90% of all PC cases. Exosomes derived from PDAC (PDAC-exosomes) promote PDAC development and metastasis. Exosomes are nanoscale vesicles secreted by most cells, which can carry biologically active molecules and mediate communication and cargo transportation among cells. Recent studies have focused on transforming exosomes into good drug delivery systems (DDSs) to improve the clinical treatment of PDAC. This review considers PDAC as the main research object to introduce the role of PDAC-exosomes in PDAC development and metastasis. This review focuses on the following two themes: (a) the great potential of PDAC-exosomes as new diagnostic markers for PDAC, and (b) the transformation of exosomes into potential DDSs.

Pancreatic cancer is resistant to conventional therapeutic interventions, mainly due to abundant cancer stromal cells and poor immune cell infiltration. Here, we used a targeted cancer therapy approach based on attenuated Salmonella typhimurium engineered to express cytolysin A (ClyA) to target cancer stromal cells and cancer cells and treat pancreatic cancer in mice. Nude mice bearing subcutaneous or orthotopic human pancreatic cancers were treated with engineered S. typhimurium expressing ClyA. The tumor microenvironment was monitored to analyze stromal cell numbers, stromal cell marker expression, and immune cell infiltration. The attenuated bacteria accumulated and proliferated specifically in tumor tissues after intravenous injection. The bacteria secreted ClyA into the tumor microenvironment. A single dose of ClyA-expressing Salmonella markedly inhibited growth of pancreatic cancer both in subcutaneous xenograft- and orthotopic tumor-bearing nude mice. Histological analysis revealed a marked decrease in expression of stromal cell markers and increased immune cell (neutrophils and macrophages) infiltration into tumors after colonization by ClyA-expressing bacteria. ClyA-expressing S. typhimurium destroyed cancer stromal cells and cancer cells in mouse models of human pancreatic cancer. This approach provides a novel strategy for combining anticancer and anti-stromal therapy to treat pancreatic cancer.

Cancer is the second leading cause of death globally. Abnormity in gene expression regulation characterizes the trajectory of tumor development and progression. RNA-binding proteins (RBPs) are widely dysregulated, and thus implicated, in numerous human cancers. RBPs mainly regulate gene expression post-transcriptionally, but emerging studies suggest that many RBPs can impact transcription by acting on chromatin as transcription factors (TFs) or cofactors. Here, we review the evidence that RBM38, an intensively studied RBP, frequently plays a tumor-suppressive role in multiple human cancer types. Genetic studies in mice deficient in RBM38 on different p53 status also establish RBM38 as a tumor suppressor (TS). By uncovering a spectrum of transcripts bound by RBM38, we discuss the diversity in its mechanisms of action in distinct biological contexts. Examination of the genomic features and expression pattern of RBM38 in human tissues reveals that it is generally lost but rarely mutated, in cancers. By assessing future trends in the study of RBM38 in cancer, we signify the possibility of targeting RBM38 and its related pathways as therapeutic strategies against cancer.

Glioma is one of the most devastating and refractory cancers. The main factors underlying therapeutic failure include extremely invasive characteristics and lack of effective methods for drug delivery. Attenuated Salmonella strains presented a high concentration of tumor targets in various types of cancer models, suggesting a role as potential vectors for drug delivery. In this study, we genetically engineered an attenuated strain of Salmonella as an anti-invasive vector for the targeted delivery and expression of tissue inhibitor of metalloproteinases 2 (TIMP-2) in an orthotopic nude mouse model of glioma. The bioluminescence signals related to tumor size significantly declined in the TIMP-2-expressing Salmonella (SLpTIMP-2)-treated group compared with the control group. Compared with the control group with a survival rate of an average of 33 days, the SLpTIMP-2 group showed an extended survival rate by nearly 60% and lasted an average period of 53 days with TIMP-2 induction. These results indicated the promising therapeutic potential of S. typhimurium for targeted delivery and secretion of TIMP-2 in glioma.

Conventional chemotherapies have some limitations, including the lack of selectivity, high toxicity to normal tissues, multidrug resistance, and tumor relapse. Recently, great progress was made in immunotherapies for anticancer research, with bacteria-mediated cancer therapy one of the most promising approaches among them. Attenuated Salmonella have very specific targeting to various solid cancers, making them ideal vectors for the delivery and expression of immunostimulators. They have native bacterial immunogenicity and induce strong anticancer immunity in vivo. In this review, the recent advances in Salmonella-mediated cancer immunotherapies and the related mechanisms of Salmonella-based cancer therapies are summarized.

Although strains of attenuated Salmonella typhimurium and wild-type Escherichia coli show similar tumor-targeting capacities, only S. typhimurium significantly suppresses tumor growth in mice. The aim of the present study was to examine bacteria-mediated immune responses by conducting comparative analyses of the cytokine profiles and immune cell populations within tumor tissues colonized by E. coli or attenuated Salmonellae. CT26 tumor-bearing mice were treated with two different bacterial strains: S. typhimurium defective in ppGpp synthesis (Delta ppGpp Salmonellae) or wild-type E. coli MG1655. Cytokine profiles and immune cell populations in tumor tissue colonized by these two bacterial strains were examined at two time points based on the pattern of tumor growth after Delta ppGpp Salmonellae treatment: 1) when tumor growth was suppressed ('suppression stage') and 2) when they began to re-grow ('re-growing stage'). The levels of IL-1 beta and TNF-alpha were markedly increased in tumors colonized by Delta ppGpp Salmonellae. This increase was associated with tumor regression; the levels of both IL-1 beta and TNF-alpha returned to normal level when the tumors started to re-grow. To identify the immune cells primarily responsible for Salmonellae-mediated tumor suppression, we examined the major cell types that produce IL-1 beta and TNF-alpha. We found that macrophages and dendritic cells were the main producers of TNF-alpha and IL-1 beta. Inhibiting IL-1 beta production in Salmonellae-treated mice restored tumor growth, whereas tumor growth was suppressed for longer by local administration of recombinant IL-1 beta or TNF-alpha in conjunction with Salmonella therapy. These findings suggested that IL-1 beta and TNF-alpha play important roles in Salmonella-mediated cancer therapy. A better understanding of host immune responses in Salmonella therapy may increase the success of a given drug, particularly when various strategies are combined with bacteriotherapy.

Accompanied by the substantive construction of domestic ports, the negative skin friction on pile becomes a common problem. In order to provide references for the related experiment research in the future, the designs of model experiments reported in the literatures were emphatically analyzed and compared. Compared to conventional pile model test, the model test on negative skin friction on pile needs to apply load on soil surface and it is difficult to simulate large surcharge by conventional test methods. An experimental scheme which could produce large surcharge load by conventional test conditions is given in this paper. Pile stress, displacement of pile top and layered settlement of soil was tested under different surcharge level. The depth of neutral point and the group effect of NSF are discussed.

Several optical imaging techniques have been used to monitor bacterial tropisms for cancer. Most such techniques require genetic engineering of the bacteria to express optical reporter genes. This study investigated a novel tumor-targeting strain of bacteria, Rhodobacter sphaeroides 2.4.1 (R. sphaeroides), which naturally emits near-infrared fluorescence, thereby facilitating the visualization of bacterial tropisms for cancer. To determine the penetration depth of bacterial fluorescence, various numbers of cells (from 10(8) to 10(10) CFU) of R. sphaeroides and two types of Escherichia coli, which stably express green fluorescent protein (GFP) or red fluorescent protein (RFP), were injected s.c. or i.m. into mice. Bacterial tropism for cancer was determined after i.v. injection of R. sphaeroides (10(8) CFU) into mice implanted s.c. with eight types of tumors. The intensity of the fluorescence signal in deep tissue (muscle) from R. sphaeroides was much stronger than from E. coli-expressing GFP or RFP. The near-infrared fluorescence signal from R. sphaeroides was visualized clearly in all types of human or murine tumors via accumulation of bacteria. Analyses of C-reactive protein and procalcitonin concentrations and body weights indicated that i.v. injection of R. sphaeroides does not induce serious systemic immune reactions. This study suggests that R. sphaeroides could be used as a tumor-targeting microorganism for the selective delivery of drugs to tumor tissues without eliciting a systemic immune reaction and for visualizing tumors.

We report a method of cancer immunotherapy using an attenuated Salmonella typhimurium strain engineered to secrete Vibrio vulnificus flagellin B (FlaB) in tumor tissues. Engineered FlaB-secreting bacteria effectively suppressed tumor growth and metastasis in mouse models and prolonged survival. By using Toll-like receptor 5 (TLR5)-negative colon cancer cell lines, we provided evidence that the FlaB-mediated tumor suppression upon bacterial colonization is associated with TLR5-mediated host reactions in the tumor microenvironment. These therapeutic effects were completely abrogated in TLR4 and MyD88 knockout mice, and partly in TLR5 knockout mice, indicating that TLR4 signaling is a requisite for tumor suppression mediated by FlaB-secreting bacteria, whereas TLR5 signaling augmented tumor-suppressive host reactions. Tumor microenvironment colonization by engineered Salmonella appeared to induce the infiltration of abundant immune cells such as monocytes/macrophages and neutrophils via TLR4 signaling. Subsequent secretion of FlaB from colonizing Salmonella resulted in phenotypic and functional activation of intratumoral macrophages with M1 phenotypes and a reciprocal reduction in M2-like suppressive activities. Together, these findings provide evidence that nonvirulent tumor-targeting bacteria releasing multiple TLR ligands can be used as cancer immunotherapeutics.

We report a method of cancer immunotherapy using an attenuated

Dinh An Estuary is one of Bassac River’s estuaries as a part of the Mekong delta. The navigation channel entrance, however, has a limited nautical depth due to the presence of a highly dynamic offshore bar at the estuary. Ship’s size sailing on the Bassac River is therefore limited to 5,000 DWT in optimal conditions. It needs to dredge twice a year and a lot of money paid for keeping the channel. Sometimes, the channel was deposited intermediately after dredging. In this paper, according to the specific morphodynamic conditions at the estuary, the advantages and drawbacks for the proposed regulation schemes are established. Four regulation schemes are numerically investigated and analyzed with respect to the behavior of each scheme to improve the water depth in the navigation channel.

Bacteria-based anticancer therapies aim to overcome the limitations of current cancer therapy by actively targeting and efficiently removing cancer. To achieve this goal, new approaches that target and maintain bacterial drugs at sufficient concentrations during the therapeutic window are essential. Here, we examined the tumor tropism of attenuated Salmonella typhimurium displaying the RGD peptide sequence (ACDCRGDCFCG) on the external loop of outer membrane protein A (OmpA). RGD-displaying Salmonella strongly bound to cancer cells overexpressing alpha v beta 3, but weakly bound to alpha v beta 3-negative cancer cells, suggesting the feasibility of displaying a preferential homing peptide on the bacterial surface. In vivo studies revealed that RGD-displaying Salmonellae showed strong targeting efficiency, resulting in the regression in alpha v beta 3-overexpressing cancer xenografts, and prolonged survival of mouse models of human breast cancer (MDA-MB-231) and human melanoma (MDA-MB-435). Thus, surface engineering of Salmonellae to display RGD peptides increases both their targeting efficiency and therapeutic effect.

Despite substantial advances in the diagnosis of cardiovascular disease, there is a need for 18F-labeled myocardial perfusion agents for the diagnosis of ischemic heart disease because current PET tracers for myocardial perfusion imaging have a short half-life that limits their widespread clinical use in PET. Thus, 18F-labeled fluoroalkylphosphonium derivatives (18F-FATPs), including (5-18F-fluoropentyl)triphenylphosphonium cation (18F-FPTP), (6-18F-fluorohexyl)triphenylphosphonium cation (18F-FHTP), and (2-(2-18F-fluoroethoxy)ethyl)triphenylphosphonium cation (18F-FETP), were synthesized. The myocardial extraction and image quality of the 18F-FATPs were compared with those of 13N-NH3 in rat models.The first-pass extraction fraction (EF) values of the 18F-FATPs (18F-FPTP, 18F-FHTP, 18F-FETP) and 13N-NH3 were measured in isolated rat hearts perfused with the Langendorff method (flow velocities, 0.5, 4.0, 8.0, and 16.0 mL/min). Normal and myocardial infarction rats were imaged with small-animal PET after intravenous injection of 37 MBq of 18F-FATPs and 13N-NH3. To determine pharmacokinetics, a region of interest was drawn around the heart, and time-activity curves of the 18F-FATPs and 13N-NH3 were generated to obtain the counts per pixel per second. Defect size was analyzed on the basis of polar map images of 18F-FATPs and 13N-NH3.The EF values of 18F-FATPs and 13N-NH3 were comparable at low flow velocity (0.5 mL/min), whereas at higher flows EF values of 18F-FATPs were significantly higher than those of 13N-NH3 (4.0, 8.0, and 16.0 mL/min, P0.05). Myocardium-to-liver ratios of 18F-FPTP, 18F-FHTP, 18F-FETP, and 13N-NH3 were 2.10±0.30, 4.36±0.20, 3.88±1.03, and 0.70±0.09, respectively, 10 min after injection, whereas myocardium-to-lung ratios were 5.00±0.25, 4.33±0.20, 7.98±1.23, and 2.26±0.14, respectively. Although 18F-FATPs and 13N-NH3 sharply delineated myocardial perfusion defects, defect size on the 13N-NH3 images was significantly smaller than on the 18F-FATP images soon after tracer injection (0-10 min, P=0.027).18F-FATPs exhibit higher EF values and more rapid clearance from the liver and lung than 13N-NH3 in normal rats, which led to excellent image quality in a rat model of coronary occlusion. Therefore, 18F-FATPs are promising new PET radiopharmaceuticals for myocardial perfusion imaging.

Salmonella typhimurium is a facultative anaerobic bacteria and can selectively grow in tumors following systemic administration. Different strategies have been used to deliver payloads to tumor tissues using engineered Salmonellae. We report a novel method of cancer immunotherapy using an attenuated S. typhimurium strain engineered to secrete Vibrio vulnificus flagellin B (FlaB) in tumor tissues.Previously, we developed an attenuated strain of S. typhimurium, which is defective in ppGpp synthesis (ΔppGpp S. typhimurium), and manifested significantly increased LD50 by 100,000 to 1,000,000-fold. To generate an inducible vector system for bacterial expression of the therapeutic gene, we cloned the flaB gene into the pBAD plasmid vector in which the pelB leader sequence was cloned upstream of flaB to guide extracellular secretion; gene expression from the plasmid (pFlaB) was induced only in the presence of L-arabinose. To evaluate the antitumor activity of engineered S. typhimurium, C57BL/6 mice transplanted with MC38 tumors were injected intravenously (i.v.) with PBS, ΔppGpp S. typhimurium carrying an empty vector, or ΔppGpp S. typhimurium carrying pFlaB (+/− L-arabinose induction). To test whether the FlaB-secreting ΔppGpp S. typhimurium has inhibitory effects on human metastatic cancer, we next implanted HCT116-luc2 tumors into the intestinal wall of BALB/c athymic nu-/nu- mice using a surgical orthotopic implantation (SOI) procedure. Engineered FlaB-secreting Salmonellae significantly suppressed tumor growth and metastasis in mouse models and prolonged survival. By using TLR5-negative colon cancer cell lines (MC38 and HCT116), we have proved that the FlaB-mediated tumor suppression upon bacterial colonization should be associated with TLR5-mediated host reactions in the tumor microenvironment. These therapeutic effects were completely abrogated in TLR4 and MyD88 knockout mice, and partly in TLR5 knockout mice, strongly suggesting that TLR4 signaling is a requisite for the FlaB-secreting bacteria-mediated tumor suppression where TLR5 signaling augmented tumor suppressive host reactions. Tumor colonization by engineered Salmonellae appeared to induce the infiltration of abundant immune cells such as monocytes/macrophages and neutrophils via the TLR4 signaling. Subsequent secretion of FlaB from colonizing Salmonellae resulted in phenotypic and functional activation of intratumoral macrophages with M1 phenotypes and a reciprocal reduction in M2-like suppressive activities. Taken together, these findings provide evidence that non-virulent tumor targeting bacteria liberating multiple TLR ligands can be used as novel cancer immunotherapeutics. Citation Format: Jung-Joon Min, Jin Hai Zheng, Yeongjin Hong, Hyon E. Choy, Joon Haeng Rhee. Targeted cancer immunotherapy with engineered Salmonella typhimurium secreting heterologous bacterial flagellin [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1610. doi:10.1158/1538-7445.AM2017-1610

Several optical imaging techniques have been used to monitor bacterial tropisms for cancer. Most such techniques require genetic engineering of the bacteria to express optical reporter genes. This study investigated a novel tumor-targeting strain of bacteria, Rhodobacter sphaeroides 2.4.1 (R. sphaeroides), which naturally emits near-infrared fluorescence, thereby facilitating the visualization of bacterial tropisms for cancer. To determine the penetration depth of bacterial fluorescence, various numbers of cells (from 10(8) to 10(10) CFU) of R. sphaeroides and two types of Escherichia coli, which stably express green fluorescent protein (GFP) or red fluorescent protein (RFP), were injected s.c. or i.m. into mice. Bacterial tropism for cancer was determined after i.v. injection of R. sphaeroides (10(8) CFU) into mice implanted s.c. with eight types of tumors. The intensity of the fluorescence signal in deep tissue (muscle) from R. sphaeroides was much stronger than from E. coli-expressing GFP or RFP. The near-infrared fluorescence signal from R. sphaeroides was visualized clearly in all types of human or murine tumors via accumulation of bacteria. Analyses of C-reactive protein and procalcitonin concentrations and body weights indicated that i.v. injection of R. sphaeroides does not induce serious systemic immune reactions. This study suggests that R. sphaeroides could be used as a tumor-targeting microorganism for the selective delivery of drugs to tumor tissues without eliciting a systemic immune reaction and for visualizing tumors.

Anticancer therapy using engineered bacteria aims to overcome limitations of current cancer therapy by actively targeting and efficiently removing cancer. In order to achieve this goal, new approaches are essential to target therapeutically resistant regions of tumors, to maintain enough number of bacteria in tumor and to deliver drugs at sufficient concentrations during the period of therapeutic process. Therefore, it is necessary to enhance the bacterial targeting efficiency through bacterial surface engineering. Here, we demonstrate that Salmonella tumor tropism can be strengthened significantly via surface displaying of RGD peptide sequence (ACDCRGDCFCG; RGD sequence) in the external loop of outer membrane protein A (OmpA) of attenuated Salmonella typhimurium. RGD-displaying Salmonella strongly bound to αvβ3 over-expressing cancer cells while showed weak binding to non-expressing cancer cells, indicating the feasibility of surface display with preferential homing peptide. In vivo bioluminescence imaging showed strong targeting efficiency of RGD-displayed Salmonella in αvβ3 over-expressing cancer xenografts (MDA-MB-231, MDA-MB-435, M21, and U87MG). The surface engineered bacteria significantly suppressed both human breast tumor (MDA-MB-231) and human melanoma (MDA-MB-435), and prolonged survival in mice. In conclusion, engineered bacteria displaying RGD peptides on the surface could advance both targeting efficiency and therapeutic effects. Citation Format: Seung-Hwan Park, Jin Hai Zheng, Hee-Seung Yun, In-Kyu Park, Yeongjin Hong, Hyun E. Choy, Jung-Joon Min. Surface-displayed RGD enhanced the targeting and therapeutic efficacy of attenuated Salmonella typhimurium. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4219.

Repebody is a binding scaffold based on variable lymphocyte receptors which are nonimmunoglobulin antibodies composed of leucine-rich repeat modules in jawless vertebrates. Repebody can be developed against variety of epitopes by module engineering. In this study, EGFR-specific repebody (RBEGFR) was developed to visualize the status of receptor expression and to prevent ligand binding that may inhibit autophosphorylation and downstream intracellular signaling. We developed RBEGFR by phage display. H1650, HCC827, A549 (human non-small cell lung cancer) and HT29 (human colon cancer) were selected as EGFR expressing cell lines. MDA-MB-435 (human melanoma) and SW620 (human colon cancer) was selected as a negative control. Specific binding of RBEGFR to cells and cancer tissue was determined by immunofluorescence (IF) staining and/or FACS analysis. In vivo imaging was done by i.v. injection of FNR-675 labeled RBEGFR (30 μg/mouse) or 64Cu-NOTA- RBEGFR (7.4 MBq/mouse) in H1650, HCC827 and HT29-bearing mouse models using cooled CCD camera or microPET, respectively. Orthotopic colon cancer mice were generated by i.p. injection of Azoxymethane (AOM; 10 mg/kg) and oral administration of 2% dextran sulfate sodium (DSS). In vivo imaging was done by i.v. injection FNR-675 labeled RBEGFR (30 μg/mouse) in AOM/DSS mouse models using cooled CCD camera or fluorescence microendoscopy. In vitro and in vivo IF staining demonstrated that strong binding of RBEGFR to H1650, HCC827 and HT29, but not to MDA-MB-435 and SW620. In vivo near infrared (NIR) imaging demonstrated specific targeting of FNR-675-RBEGFR to grafted H1650, HCC827 and HT29 tumor in mice. The 64Cu-NOTA- RBEGFR was detected at the implanted tumor from 1 h (SUVmax: 1.34±0.12) after the injection, peaked at 6 h (1.75±0.18), maintained to 24 h (1.33±0.17). The radioactivity significantly decreased by blocking with cold form of 50 μM naïve RBEGFR 1 day before injection of 64Cu-NOTA- RBEGFR, indicating specific binding of RBEGFR to EGFR in vivo. Optical NIR imaging after i.v. injection of FNR-675-RBEGFR showed specific signal in the abdomen of AOM/DSS mice, but not in control mice. Correlation with surgical/necropsy imaging, fluorescence endoscopy and pathology revealed strong accumulation of FNR-675-RBEGFR in malignant dysplasia, but weak or no accumulation in low grade tumor or benign lesion. In conclusion, the RBEGFR could be developed for specific targeting of cancer overexpressing EGFR. The fluorescence-labeled RBEGFR could be developed for imaging agent for detecting colonic dysplasia and assessing EGFR status. In particular, this agent may have a potential as an imaging companion diagnostics to predict therapeutic outcome of targeted therapy with monoclonal antibody for EGFR through pre-therapeutic visualization of EGFR status. Our work provides a basis to develop potential strategy of targeted immune-detection of cancers which may replace monoclonal antibodies. Citation Format: Misun Yun, Dong-Yeon Kim, Hyeon-Sik Kim, Jin Hai Zheng, AYoung Pyo, Jung-Joon Min. Molecular imaging of EGFR-expressing tumors with novel targeted protein scaffold, anti-EGFR repebody. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4212.

It is generally accepted that there are transverse oscillation, which are concentrated and confined to the backwall and decay asymptotically offshore, existed in the harbor of constant slope, however, whether these oscillations can be induced by the normally incident waves is not clear. This numerical investigation aims at providing the subharmonic generations of transverse oscillations within the harbor of a plane slope by waves normally impacting on. For the harbor of perfectly plane slopes, the subharmonic transverse oscillations are small on the mild and moderate slopes but evident on the steep slope. This instability can take place only if the incident wave amplitude exceeds a threshold value, and transverse oscillations can even grow up to a larger value than that of longitudinal oscillations. The magnitudes of transverse oscillations are approximately the same, only their growth rates are affected by the incident wave amplitude.

Obligate or facultative anaerobic bacteria such as Bifidobacterium, Clostridium, Salmonella, or Escherichia coli specifically colonize and proliferate inside tumor tissues and inhibit tumor growth. Among them, attenuated Salmonella typhimurium (S. typhimurium) has been widely studied in animal cancer models and Phase I clinical trials in human patients. S. typhimurium genes are easily manipulated; thus diverse attenuated strains of S. typhimurium have been designed and engineered as tumor-targeting therapeutics or drug delivery vehicles that show both an excellent safety profile and therapeutic efficacy in mouse models. An attenuated strain of S. typhimurium, VNP20009, successfully targeted human metastatic melanoma and squamous cell carcinoma in Phase I clinical trials; however, the efficacy requires further refinement. Along with the characteristics of self-targeting, proliferation, and deep tissue penetration, the ease of genetic manipulation allows for the production of more attenuated strains with greater safety profiles and vector systems that deliver designable cargo molecules for cancer diagnosis and/or therapy. Here, we discuss recent progress in the field of Salmonellae-mediated cancer therapy.

The Dinh An Estuary is one of the Nine Dragon estuaries of the Mekong River. An international navigation channel was built in the estuary for vessels traveling from the South China Sea to the southwestern area of Vietnam and then to Phnom Penh, Cambodia. The morphological evolution of the navigation channel is complicated and unstable. The back siltation intensity in the navigation channel has largely increased and been concentrated in the curvature segments of the channel since 1980. In this study, based on simulation results and measured data, five key factors that influence the back siltation in the navigation channel were systematically analyzed. These factors included the increasing elevation gap between the channel and the nearby seabed, the disadvantageous hydrodynamic conditions, sediment transport, mixing of saltwater and freshwater, and wave effects in the navigation channel. It is shown that the back siltation to a large extent results from the low current velocity of the secondary ocean circulation, which often occurs in the curvature segments of the channel. Suspended sediment also settles in the channel due to the decrease of the current velocity and the sediment transport capacity when flow passes through the channel. The changes of hydrodynamic conditions are responsible for the majority of the severe siltation in the curvature segments of the navigation channel.