identification-characterization-metastatic-factors-gene-transfer-into

Identification and Characterization of Metastatic Factors by Gene Transfer into the Novel <em>RIP-Tag; RIP-tva</em> Murine Model

Metastatic cancer accounts for 90% of deaths in patients with solid tumors. There is an urgent need to better understand the drivers of cancer metastasis ...

Weill Cornell Medicine

Survivin is highly expressed in most human tumors and fetal tissue, and absent in terminally differentiated cells. It promotes tumor cell proliferation by negatively regulating cell apoptosis and facilitating cell division. Survivin's selective expression pattern suggests that it might be a suitable target for cancer therapy, which would promote death of transformed but not normal cells. This was tested using artificial microRNAs (amiRNAs) targeting survivin. After screening, two effective amiRNAs, which knocked down survivin expression, were identified and cloned into a replication-defective adenoviral vector. Tumor cells infected with the recombinant vector downregulated expression of survivin and underwent apoptotic cell death. Further studies showed that apoptosis was associated with increases in caspase 3 and cleaved Poly (ADP-ribose) polymerase, and activation of the p53 signaling pathway. Furthermore, amiRNA treatment caused blockade of mitosis and cell cycle arrest at the G2/M phase. In vivo, survivin-targeting amiRNAs expressed by adenoviral vectors effectively delayed growth of hepatocellular and cervical carcinomas in mouse xenograft models. These results indicate that silencing of survivin by amiRNA has potential for treatment of cancer.

Survivin-targeting Artificial MicroRNAs Mediated by Adenovirus Suppress Tumor Activity in Cancer Cells and Xenograft Models.

Hand, foot and mouth disease (HFMD) is an important public health problem that has emerged over the past several years. HFMD predominantly infects children under seven years old and occasionally causes severe disease in adults. Among the enteroviruses, enterovirus 71 (EV71) and coxsackievirus 16 (CA16) are the major causative agents of HFMD. In addition, adenovirus cocirculates with enterovirus and has become a possible additional pathogenic factor for HFMD in some cases. Here, we have investigated the neutralizing antibody responses to both enterovirus and adenovirus in adults, with the aim of exploring the prevalence trends of these viruses and the nature of protective immunity in humans to these viral infections. Sera from 391 healthy adults from 21 provinces and cities in China were tested for the presence of antibodies against EV71, CA16, adenovirus human serotype 5 (AdHu5) and chimpanzee adenovirus pan7 (AdC7) using neutralization tests. High seroprevalence rates of EV71, CA16 and AdHu5 were found in the population (85.7%, 58.8% and 74.2%, respectively). The coseropositivity rate of these three viruses was 39.4% (154 of 391), with median neutralizing antibody titers of 80, 40 and 640, respectively, and the neutralizing antibody titer for EV71 was found to be correlated with those of CA16 and AdHu5. AdC7 was found to be a rare adenovirus serotype in the human population, with a seropositivity rate of 11.8%, suggesting that it could be a good choice for a vaccine carrier that could be used in vaccine development.

Neutralizing antibody responses to enterovirus and adenovirus in healthy adults in China.

Hand, foot and mouth disease (HFMD) is a serious public health problem that has emerged over the past several decades. Pathogen detection by the Chinese national HFMD surveillance system has focused mainly on enterovirus 71 (EV71) and coxsackievirus A16 (CA16). Therefore, epidemiological information regarding the other causative enteroviruses is limited. To identify the pandemic enterovirus in Suzhou, Jiangsu province, China, clinical samples from patients with HFMD were collected from 2012 to 2013 and analyzed. The results revealed that CA16 was the most dominant HFMD pathogen in 2012, whereas CA6 and CA10 were the dominant pathogens in 2013. Phylogenetic analysis revealed that the C4a sub-genogroup of EV71 and the B1a and B1b sub-genogroups of CA16 continued to evolve and circulate in Suzhou. The CA6 strains were assigned to six genotypes (A-F) and the CA10 strains were assigned to seven genotypes (A-G), with clear geographical and temporal distributions. All of the CA6 strains in Suzhou belonged to genogroup F, and there were several lineages circulating in Suzhou. All of the CA10 strains in Suzhou belonged to genogroup G, and they had the same genetic origin. Co-infections of EV71/CA16 and CA6/CA10 were found in the samples, and bootscan analysis of 5'-untranslated regions (UTRs) revealed that some CA16 strains in Suzhou had genetic recombination with EV71. This property might allow CA16 to alter its evolvability and circulating ability. This study underscores the need for surveillance of CA6 and CA10 in the Yangtze River Delta and East China.

Phylogenetic analysis of the major causative agents of hand, foot and mouth disease in Suzhou City, Jiangsu province, China, in 2012-2013.

Hand, foot and mouth disease (HFMD) is a major public health concern in Asia; more efficient vaccines against HFMD are urgently required. Adenoviral (Ad) capsids have been used widely for the presentation of foreign antigens to induce specific immune responses in the host. Here, we describe a novel bivalent vaccine for HFMD based on the hexon-modified, E1-deleted chimpanzee adenovirus serotype 68 (AdC68). The novel vaccine candidate was generated by incorporating the neutralising epitope of Coxsackievirus A16 (CA16), PEP71, into hypervariable region 1 (HVR1), and a shortened neutralising epitope of Enterovirus 71 (EV71), sSP70, into HVR2 of the AdC68 hexon. In order to enhance the immunogenicity of EV71, VP1 of EV71 was cloned into the E1-region of the AdC68 vectors. The results demonstrated that these two epitopes were well presented on the virion surface and had high affinity towards specific antibodies, and VP1 of EV71 was also significantly expressed. In pre-clinical mouse models, the hexon-modified AdC68 elicited neutralising antibodies against both CA16 and EV71, which conferred protection to suckling mice against a lethal challenge of CA16 and EV71. In summary, this study demonstrates that the hexon-modified AdC68 may represent a promising bivalent vaccine carrier against EV71 and CA16 and an epitope-display platform for other pathogens.

Hexon-modified recombinant E1-deleted adenoviral vectors as bivalent vaccine carriers for Coxsackievirus A16 and Enterovirus 71.

Acting as inflammatory mediators, tumor oncogenes or suppressors, microRNAs are involved in cell survival, death, epithelial-mesenchymal transition and metastasis, etc. Investigating the communication between microRNAs and tumorigenesis is critical to our understanding of the pathogenesis of multiple disease states.

MicroRNAs in colorectal carcinoma--from pathogenesis to therapy.

Cetuximab is a chimeric monoclonal antibody, approved to treat patients with metastatic colorectal cancer (mCRC), head and neck squamous cell carcinoma (HNSCC), non-small-cell lung cancer (NSCLC) for years. It functions by blocking the epidermal growth factor receptor (EGFR) from receiving signals or interacting with other proteins. Although the demand for cetuximab for the treatment of cancer patients in clinics is increasing, the complicated techniques involved and its high cost limit its wide applications. Here, a new, cheaper form of cetuximab was generated for cancer gene therapy. This was achieved by cloning the full-length cetuximab antibody into two serotypes of adenoviral vectors, termed as AdC68-CTB and Hu5-CTB. In vivo studies showed that a single dose of AdC68-CTB or Hu5-CTB induced sustained cetuximab expression and dramatically suppressed tumor growth in NCI-H508- or DiFi-inoculated nude mice. In conclusion, gene therapy using adenovirus expressing full-length cetuximab could be a novel alternative method for the effective treatment of colorectal cancer.

Gene therapy for colorectal cancer using adenovirus-mediated full-length antibody, cetuximab.

Vaccination is considered to be the most effective method of preventing infectious or other diseases. Adenovirus (Ad) is one the most promising vectors in vaccine research and development. It can induce not only potent humoral but also cellular immune responses, and has therefore been widely applied in basic and translational studies. Chimpanzee Ad is a rare serotype circulating in humans. This circumvents the problem of preexisting immunity to human Ad serotypes, enhancing Chimpanzee Ad prospects in vaccine development. Here we describe experimental procedures used to generate a new generation of rabies vaccine based on a chimpanzee Ad vector, which can be extended in the development of novel vaccines against other infectious diseases.

Development of Novel Vaccines Against Infectious Diseases Based on Chimpanzee Adenoviral Vector.

Among the oncolytic virotherapy, an emerging treatment for tumor, adenoviruses are widely used at present in preclinical and clinical trials. Traditionally, oncolytic adenoviruses were developed based on the human adenovirus serotype 5 (AdHu5). However, AdHu5 has the drawbacks of preexisting anti-AdHu5 immunity in most populations, and extensive sequestration of Adhu5 by the liver through hexon, blood coagulation factor X (FX), and FX receptor interactions. To tackle these problems, we explored a novel oncolytic adenovirus AdC7-SP/E1A-ΔE3 for cancer treatment. AdC7-SP/E1A-ΔE3 was constructed by replacing the E1A promoter with tumor specific promoter survivin promoter and deleting E3 region using direct cloning methods based on simian adenovirus serotype 24 (namely AdC7). We showed that AdC7-SP/E1A-ΔE3 significantly killed tumor cell lines NCI-H508 and Huh7, and inhibited tumor growth in both NCI-H508 and Huh7 xenograft tumor models. Importantly, AdC7-SP/E1A-ΔE3 exhibited the antitumor efficacy via systemic administration. Mechanistically, infected cells were killed by AdC7-SP/E1A-ΔE3 via the p53-independent mitochondrial apoptosis pathway in which phosphorylation of BAD markedly declined and the expresses of Bik significantly went up. Therefore, AdC7-SP/E1A-ΔE3 is a promising candidate for liver and colon tumor treatment.

A novel oncolytic adenovirus based on simian adenovirus serotype 24.

Since 2013, the outbreak or sporadic infection of a new reassortant H7N9 influenza virus in China has resulted in hundreds of deaths and thousands of illnesses. An H7N9 vaccine is urgently needed, as a licensed human vaccine against H7N9 influenza is currently not available. Here, we developed a recombinant adenovirus-based vaccine, AdC68-H7HA, by cloning the H7N9 haemagglutinin (HA) gene into the chimpanzee adenoviral vector AdC68. The efficacy of AdC68-H7HA was evaluated in mice as well as guinea pigs. For comparison, an H7N9 DNA vaccine based on HA was also generated and tested in mice and guinea pigs. The results demonstrated that both AdC68-H7HA and the DNA vaccine prime-adenovirus boost regimen induced potent immune responses in animals and completely protected mice from lethal H7N9 influenza viral challenge. A post-immunization serum transfer experiment showed that antibody responses could completely protect against lethal challenge, while a T cell depletion experiment indicated that HA-specific CD8(+) T cells responses also contributed to protection. Therefore, both HA-specific humoral immunity and cellular immunity play important roles in the protection. These data suggest that the chimpanzee adenovirus expressing HA is a promising vaccine candidate for H7N9 virus or other influenza viral subtypes.

Both haemagglutinin-specific antibody and T cell responses induced by a chimpanzee adenoviral vaccine confer protection against influenza H7N9 viral challenge.

Article	Year
Survivin-targeting Artificial MicroRNAs Mediated by Adenovirus Suppress Tumor Activity in Cancer Cells and Xenograft Models. Molecular therapy. Nucleic acids\| PubMed ID: 25368912	2014
Neutralizing antibody responses to enterovirus and adenovirus in healthy adults in China. Emerging microbes & infections\| PubMed ID: 26038738	2014
Phylogenetic analysis of the major causative agents of hand, foot and mouth disease in Suzhou City, Jiangsu province, China, in 2012-2013. Emerging microbes & infections\| PubMed ID: 26038764	2015
Hexon-modified recombinant E1-deleted adenoviral vectors as bivalent vaccine carriers for Coxsackievirus A16 and Enterovirus 71. Vaccine\| PubMed ID: 26296491	2015
MicroRNAs in colorectal carcinoma--from pathogenesis to therapy. Journal of experimental & clinical cancer research : CR\| PubMed ID: 26964533	2016
Gene therapy for colorectal cancer using adenovirus-mediated full-length antibody, cetuximab. Oncotarget\| PubMed ID: 27058423	2016
Development of Novel Vaccines Against Infectious Diseases Based on Chimpanzee Adenoviral Vector. Methods in molecular biology (Clifton, N.J.)\| PubMed ID: 28374240	2017
A novel oncolytic adenovirus based on simian adenovirus serotype 24. Oncotarget\| PubMed ID: 28460470	2017
Both haemagglutinin-specific antibody and T cell responses induced by a chimpanzee adenoviral vaccine confer protection against influenza H7N9 viral challenge. Scientific reports\| PubMed ID: 28500340	2017

Yudan Chi

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