Antitumor Effects Of Newcastle Disease Virus Hemagglutinin-Neuraminidase Used As A Molecular Adjuvant

Although many cancer gene therapy strategies aim at eradicating tumor cellswith the final goal to restrain tumor growth and confer longer survival time,augmentation of anti-tumor immune responses and induction of long-term immunesurveillance are coordinative important. Innate immunity not only plays an importantrole in the induction of adaptive immune responses but contributes to anticanceractivities. Therefore, novel therapeutic strategies including triggering of the innatearm of the immune system may be successful in increasing anti-tumor activity.Molecular adjuvants are kinds of molecules that can synergic stimulate adaptiveand innate immune responses. Furthermore, these adjuvants can not only activate NKbut stimulate innate immune system to produce cytokines. Therefore the studies aboutthese molecules most focus on vaccine development. Although there are only twoadjuvants were currently permitted for clinical use, other molecular adjuvants,including cytokines, bacterial product, heat shock protein, virus like particles andimmunostimulatory complexes, are extensively researched. However, many of themolecules have side effects which make them fail in clinical using. In this study, tofulfill the highlighted demand for non-toxic adjuvants, we investigated the use ofNDV HN as a molecular immune adjuvant.In our previous studies, we generated an oncolytic adenovirus Ad-Apoptin basedon a cancer cell selective apoptosis-inducing gene. It was shown that Ad-Apoptincould perform strong antitumor and apoptosis-inducing effects in vitro. It also couldconfer significant anticancer activity toward solid tumors in vivo. In addition,Ad-Apoptin was shown to be capable of suppressing metastatic nodules. Recently, Ad-Apoptin was shown to directly restrain tumor growth via system delivering.Because of these therapeutic properties of Ad-Apoptin, we sought to evaluate the useof the recombinant adenovirus as a candidate of anti-tumor gene therapy. However,the recombinant was absence of immune stimulating capacities, which made itinadequate to obtain prolonged anti-tumor activity.NDV is a paramyxovirus with a negative single-stranded RNA genome whichcauses inflammation of the respiratory and gastrointestinal tract in a wide variety ofpoultry species. HN, a74kDa multifunctional membrane class II glycoprotein, is oneof the most important structural proteins of NDV. The protein can not only mediatesrecognition of sialic acid containing receptors but also possesses NA which cancleave the sialic acid on those receptors. It was demonstrated that HN could stimulatea strong interferon-α response in peripheral blood mononuclear cells (PBMCs). It wasshown that HN protein could confer T cell co-stimulatory function and inducinginnate immune activities. Furthermore, HN had also the type I interferons inducingcapacities. Additionally, NDV HN was shown to directly activate human NK cells viatriggering the NKp44and NKp46receptors.Antitumor immunity can be either innate or adaptive. Although adaptiveantitumor immune activity can induce reactivity against defined antigens, induce longlasting immune memory, and give low toxicity to normal tissues, innate approachesmay also be of benefit. In fact, the most effective immunological antitumor responseshave been seen with active innate approaches. NK, a key component of the innateimmune system, is characterized by their rapid response and strong cytolytic activitywithout pre-sensitization and involving of major histocompatibility molecules. NKcan not only carry out cytotoxicity through the expression of TNF-related apoptosisinducing ligand (TRAIL) or Fas ligand (FasL) on the NK cell surface or TNF-αsecretion by NK cell, but regulate immune responses through produce IFN-γ,interleukins and growth factors. Furthermore, NK cells produce many chemokinesthat impact dendritic cells (DCs), macrophages and neutrophils during an immuneresponse. NDV HN used in present study can activate NK, DCs, monocytes, and stabilize activated T cells. This highlights the potential adjuvant activity of the HNgene in tumor therapy.In this study, we approached the use of HN as a potential molecular immuneadjuvant in tumor therapy. We demonstrated that Ad-HN was capable of inducinglymphokine effector cytokines of PBMCs in vitro. In addition, Ad-HN adjuvant led toreduction of tumor growth and provided survival benefits when compared to thecontrol adjuvants. Therefore, it is plausible that HN is a suitable immune adjuvant forcancer gene therapy.We demonstrated here that HN successfully expressed in SGC7901cellsperform both NA and HA. We further shown that SGC7901cells carrying HNinduced PBMCs to secrete increased amounts of lymphokine effector cytokinesTNF-α, IFN-α and IFN-γ. It was known that TNF-α could contribute to the antitumorcytotoxicity of activated macrophages, and IFN-γ released by NK could promote Th1cell polarization and subsequent cytotoxic-T-lymphocyte induction. Additionally,IFN-α could exhibit anti-tumor effects as well as had multiple activities on immunecells. All these might be contribute to variety of direct and indirect antitumor effectorfunctions. Subsequently, we showed that the incubation of NK cells withAd-HN-infected SGC7901cells was able to enhance the cytotoxic activity of NKagainst uninfected target cells, which might be contribute to the in vivo oncolyticproperties.We also investigated in settings of therapeutic antitumor model whether HNadjuvant co-application with Ad-Apoptin could improve effectivity. Data showed areduction of tumor burden in tumor bearing mice upon therapeutic treatment withAd-Apoptin and subsequently Ad-HN as adjuvant in comparison to Ad-Apoptintreatment alone. We further observed remarkable survival benefits in the group ofapplication of NDV HN as an immune adjuvant. All these might be due to thesynergistic reaction between Apoptin and HN. Since Apoptin could obstruct theprompt growth of the tumors, meanwhile HN established the immunological defenseagainst the infiltration, the diffusion and the metastasis of the tumors. In conclusion, the results from in vitro experiments and in vivo tumor modellack of adaptive immune system suggested that innate immune response, most likelyinvolving NK cells, plays a key role in the suppression of syngeneic graft andimprovement of the mean survival directed by adjuvant property of HN. The resultsfurther reinforced the potential adjuvant activity of the HN gene in tumor therapy.