The cancer-testis antigen NY-ESO-1 is one of the most promising candidates for generic vaccination of cancer patients. Here we analyzed the CD8 + T cell response to a NY-ESO-1 peptidevaccine composed of the two previously defined peptides 157-165 and 157-167, administered with GM-CSF as a systemic adjuvant. The NY-ESO-1 peptidevaccine elicited a CD8 + T cell response directed against multiple distinct epitopes in the 157-167 region, as revealed by using A2/peptide multimers incorporating overlapping A2 binding peptides in this region. However, only a minor fraction of the elicited CD8 + T cells, namely those recognizing the peptide 157-165 with sufficiently high functional avidity, recognized the naturally processed target on NY-ESO-1 + tumor cells. In contrast, the majority of peptide 157-165–specific CD8 + T cells exhibited lower functional avidity and no tumor reactivity. In addition, vaccine-elicited CD8 + T cells specific for other overlapping epitopes in the 157- 167 region failed to significantly recognize NY-ESO-1–expressing tumor targets. Thus, because of the complexity of the CD8 + T cell repertoire that can be elicited by vaccination with synthetic peptides, a precise definition of the targeted epitope, and hence, of the corresponding peptide to be used as immunogen, is required to ensure a precise tumor targeting.
Transmembrane 4 superfamily member 5 protein (TM4SF5) is a potential therapeutic target for hepatocellular carcinoma (HCC) and colon cancer. In a previous study, we demonstrated the prophylactic and therapeutic effects of a TM4SF5-specific peptidevaccine and monoclonal antibody in HCC and colon cancer in a mouse model. Here, we designed a cyclic peptide targeting TM4SF5. Cyclic peptide-specific antibodies were produced in mice after immunization with a complex of the peptide, CpG-DNA, and liposomes. Intravenous injection of the CT-26 mouse colon cancer cell line into mice induced tumors in the lung. Immunization with the peptidevaccine improved the survival rate and reduced the growth of lung tumors. We established a monoclonal antibody specific to the cyclic TM4SF5-based peptide and humanized the antibody sequence by complementarity determining region-grafting. The humanized antibody was reactive to the cyclic peptide and TM4SF5 protein. Treatment of CT-26 cells with the humanized antibody reduced cell motility in vitro. Furthermore, direct injection of the humanized anti-TM4SF5 antibody in vivo reduced growth of lung tumors in mouse metastasis model. Therefore, we conclude that the immunization with the cyclic peptidevaccine and injection of the TM4SF5-specifc humanized antibody have an anti-metastatic effect against colon cancer in mice. Importantly, the humanized antibody may serve as a starting platform for further development and application in clinical settings.
Alpha-fetoprotein (AFP) is a marker of hepatocellular carcinoma (HCC) and serves as a target for immunotherapy. However, current treatments targeting AFP are not reproducible and do not provide complete protection against cancer. This issue may be solved by developing novel therapeutic vaccines with enhanced immunogenicity that could effectively target AFP-expressing tumors. In this study, we report construction of a therapeutic peptidevaccine by linking heat shock protein 70 (HSP70) functional peptide to the AFP epitope to obtain HSP70-P/AFP-P. This novel peptide was administered into BALB/c mice to observe the effects. Quantification of AFP-specific CD8 + T cells that secrete IFN-γ in these mice via ELISPOT revealed the
Abstract: Human coronavirus (HCoV), a member of Coronaviridae family, is the causative agent of upper respiratory tract infections and “atypical pneumonia”. Despite severe epidemic outbreaks on several occasions and lack of antiviral drug, not much progress has been made with regard to an epitope-based vaccine designed for HCoV. In this study, a computational approach was adopted to identify a multiepitope vaccine candidate against this virus that could be suitable to trigger a significant immune response. Sequences of the spike proteins were collected from a protein database and analyzed with an in silico tool, to identify the most immunogenic protein. Both T cell immunity and B cell immunity were checked for the peptides to ensure that they had the capacity to induce both humoral and cell-mediated immunity. The peptide sequence from 88–94 amino acids and the sequence KSSTGFVYF were found as the most potential B cell and T cell epitopes, respectively. Furthermore, conservancy analysis was also done using in silico tools and showed a conservancy of 64.29% for all epitopes. The peptide sequence could interact with as many as 16 human leukocyte antigens (HLAs) and showed high cumulative population coverage, ranging from 75.68% to 90.73%. The epitope was further tested for binding against the HLA molecules, using in silico docking techniques, to verify the binding cleft epitope interaction. The allergenicity of the epitopes was also evaluated. This computational study of design of an epitope-based peptidevaccine against HCoVs allows us to determine novel pep- tide antigen targets in spike proteins on intuitive grounds, albeit the preliminary results thereof require validation by in vitro and in vivo experiments.
Amino acid sequences (peptide) are the building block for peptidevaccine. Antigen stimulates the immune response by recognizing the tumor cell. Epitope based peptide vaccines are upgraded and effective in the treatment of tumor in comparison to conventional whole organism based vaccines.  They generate adaptive immune response by introduction of tumor-associated antigens directly to patient. The response can be either allogeneic or autologous. Antigen can express themselves at tumor specific site or at different tumor types with tumor-specific post-translational modified proteins.
Abstract: The bacterial species Acinetobacter baumannii is a major cause of hospital acquired infection throughout the world and it is increasing public health concern. Infection caused by multidrug resistant A. baumannii is currently among the most difficult to treat due to propensity to acquire mobile genetic element. To date there is no vaccine or specific drug available for its treatment, this necessitate the need for the identification of therapeutic target enzyme and vaccine. Pharmacogenomic and computational biology represent an attractive alternative approach for the identification of common drug target and peptide-vaccine candidates in the pathogen. Vaccine designing is shifted from entire pathogen or whole antigen to peptide or epitope based-vaccines that are specific, safe and easy to produce. Comparative genomic approach was used to identify conserved protein signatures among five genomes. Three outer membrane proteins conserved among the genomes with high vaxijen scores were used to produce both B-cell and T-cell mediated immunity. Propred and propred1 were used to predict promiscuos helper T-Lymphocytes (HTL), Cytotoxic T-Lymphocyte (CTL) epitopes and MHCPred for their binding affinity.Three T-cell epitopes derived from identified B-cells bind to maximum number of MHC class I and class II alleles and specifically bind to HLA alleles such as DRB1*0101 and DRB1*0401.The epitopes are YEKLAAGPS, FYTSQPEDS and YVTGNPLGL with high potential to induce humoral and cell mediated immune responses. These predicted epitopes (small peptide) might be promising candidates for vaccine design against A. baumannii infection, though experimental validation.
The approach that we used for designing potential peptidevaccine candidates takes a community- level personalized strategy, that we refer to as populationalized vaccinomics, which enables the design of peptides based on the genome sequence of the target virus and the immune characteristics of the target populations, variable at the gene level. Based on the human leukocyte antigen (HLA) gene allele variation, the peptide vaccines can be made more efficient for a given community or individual. The selection of the peptides that could act as vaccines is determined by the binding of the processed viral peptide with the MHC class I and II molecules, and the relevant HLA alleles. We identified multiepitope peptidevaccine candidates against nCov that can potentially trigger both CD4+ and CD8+ T cell immune response.
To identify biomarkers of response and peripheral immunoreactivity patterns, we performed RNA sequencing (RNA-seq) on peripheral blood mononuclear cells (PBMCs) from patients prior to the start of vaccine treatment and at multiple time points during treatment (n = 54 total samples). We found that HLA-V, a class I MHC pseudogene, and GSTM1 expression could differentiate between SD, PR, and CR patients in our trial. Furthermore, T cell activity genes (GZMB and PRF1) were elevated at an early time point in CR patients compared with SD and PR patients, whereas monocytic pathways were elevated at early time points in SD (week 6) and PR (week 15) patients. Significantly, at week 34 following the start of treatment, only CR patients demonstrated strong induction of both IFN signaling and Poly-IC response pathways. Furthermore, genes involved in DC activation of T cells were correlated with ELISPOT respons- es to survivin. Finally, low expression levels of genes encoding immune checkpoint proteins at week 0 and high expression levels of T cell activation markers at week 6 were associated with longer PFS. Overall, our data suggest the presence of unique peripheral immune pathways associated with pediatric LGG patient response to peptidevaccine immunotherapy. Although intriguing, based on the small sample size, our identified gene signatures will ultimately need to be validated in prospective randomized trials. Finally, our findings highlight the utility of RNA-seq as a broadly useful tool for the monitoring of peripheral immune responses in patients receiving immunotherapies.
Abstract: Cell-penetrating peptides (CPPs), a group of small peptides capable of promoting the transport of molecular cargo across the plasma membrane, have become important tools in promoting the cellular uptake of exogenously delivered macromolecules. GV1001, a peptide derived from a reverse-transcriptase subunit of telomerase (hTERT) and developed as a vaccine against various cancers, reportedly has unexpected CPP properties. Unlike typical CPPs, such as the HIV-1 TAT peptide, GV1001 enabled the cytosolic delivery of macromolecules such as proteins, DNA and siRNA via extracellular heat shock protein 90 (eHSP90) and 70 (eHSP70) complexes. The eHSP-GV1001 interaction may have biological effects in addition to its cytosolic delivery function. GV1001 was originally designed as a MHC class II-binding cancer epitope, but its CPP properties may contribute to its strong anti-cancer immune response relative to other telomerase peptide-based vaccines. Cell signaling via eHSP-GV1001 binding may lead to unexpected biological effects, such as direct anticancer or antiviral effects. In this review, we focus on the CPP effects of GV1001 bound to eHSP90 and ehsp70.
Clinical trial. Between August 1996 and August 1998, 19 patients with breast or ovarian cancer were enrolled in a phase I HER-2/neu peptide–based vaccine trial approved by the University of Washington’s Human Subjects Division and the United States Food and Drug Administration. Eligibility was dependent upon subjects (a) being diagnosed with stage III/IV breast or ovarian cancer and having been treated for their pri- mary and metastatic disease according to recommend- ed disease-appropriate standards with surgery, chemotherapy, radiation therapy, or combined modal- ity, (b) having a white blood cell count greater than 3.5 dl/ml, (c) showing HER-2/neu protein overexpression in the primary tumor or metastasis, (d) being off immunosuppressive drugs and chemotherapy for at least 30 days before enrolling, and (e) being HLA-A2 positive. Patients were tested for immune competence responsiveness to a minimum of two of seven recall antigens by skin testing with Multitest CMI (Pasteur Merieux Connaught Labs, Institut Merieux, Lyon, France). All patients signed a protocol-specific consent and received monthly vaccinations with three 15–amino acid (15-aa) HER-2/neu–derived peptides, p369–p384, p688–p703, and p971–p984, containing within each the putative HLA-A2–binding motifs p369–p377 (6), p689–p697 (7), and p971–p979 (18). Five hundred micrograms of each peptide (1.5 µg total peptide dose) were solubilized in 10 mM sodium acetate (pH 4.0) and admixed with 125 µg rhuGM-CSF (kindly supplied by Immunex Corp., Seattle, Washing- ton, USA) as an adjuvant. The vaccine preparation was divided into two intradermal injections administered to the same draining lymph node site monthly for 6 months. Subjects underwent peripheral blood draws or a leukapheresis before and 30 days after each vaccina- tion for immunologic monitoring.
An effective combinatorial vaccine therapy will likely need to address three goals: building a robust antigen- specific CTL response; altering the tumor microenvironment to allow CTL infiltration and reduce migration of regulatory T-cells and myeloid-derived suppressor cells; and counteract- ing CTL inhibitory mechanisms such as immune checkpoints that lead to immunosuppression (Figure 1). An encouraging study using a combination of a peptidevaccine, anti-PD-1 antibody, and low-dose cyclophosphamide in a murine tumor model demonstrated that this combination of drugs syner- gized in increasing survival and reducing tumor burden. 158
(including therapeutic cancer vaccine, immune checkpoint inhibitors) remains optimistic [186–191]. Many of these successful human immunotherapeutics do hold similar promise in veterinary medicine [192–194] however, drug administration to wild Tasmanian devils is very different from the clinical setting of human and companion animals and therefore treatments such as adoptive cell transfer would be difficult to implement. The fact that DFT1 expresses tumour associated antigens (TAA’s) such as ERBB3 invites the application of monoclonal anti- bodies and therapeutic cancer vaccines as prospective treatments. The passive administration of monoclonal antibodies to ERBB3 primarily focused on blocking receptor epitopes are still experimental [57, 151–176] and any humanised anti-ERBB3 would certainly have to be become species specific (devil anti-ERBB3) to prevent adverse immunologic reactions . Very few monoclonal antibodies have been developed in veterinary oncology although two caninised anti- bodies anti-ERBB1  and anti-CD20  show promise. Therapeutic cancer vaccination modalities applicable to wildlife include antigen delivery vaccines that utilise inactivated cancer cells (autologous or allogenic) or peptide vaccines that mimic antigen sequences. Results using an inactivated cancer cell vaccine trial (allogenic DFT1 cell line) are eagerly awaited (http:// www.utas.edu.au/news/2015/10/16/19-world-first-trial-of-tasmanian-devil-vaccine-begins-in- the-wild/). Confidence that immunisation can be successful stems from research showing that Tasmanian devils have a competent immune system [21, 198–200] and can produce cytotoxic antibodies [14, 201]. An alternative antigen presentation modality to cancer cell vaccine is a peptidevaccine, where single or multiple amino acid sequences (long or short) representing a defined antigen is combined with adjuvant to elicit an immune response . Development of just a single ERBB3 peptidevaccine can be found in the literature  however, peptide vac- cines targeting ERBB1 [203, 204], ERBB2 [205–207] or both ERBB1/2  including monoclo- nal antibody against tyrosine related protein 1 (TRP-1) and altered peptide sequence to gp100 for mouse melanoma  all show promise. Overcoming self-tolerance is a major hurdle, one such strategy is the use of Xenoantigens, that is the exact same antigen but from a different species that has considerable sequence homology, differing only by several amino acids which appear to the host as altered epitopes or as “altered self” and therefore tolerance can be broken causing a T-cell response against the endogenous self-antigen . Veterinary xenogeneic vac- cinations include a DNA plasmid vaccine encoding human Tyrosinase (TYR)  the only vet- erinary therapeutic tumour vaccine licensed by the United States department of Agriculture (USDA) for the use of oral and digital melanoma, now marketed as Oncept TM .
The importance of immune surveillance in eradicating ma- lignant cells is well demonstrated, and several observations have motivated the development of therapeutic cancer vaccines. However, except for virus-induced cancers, tumor antigens are mostly self or near-self protein epitopes that are often poorly immunogenic and submitted to central and peripheral tolerance. For therapeutic cancer vaccines to be effective, they must overcome regulatory and immuno- suppressive mechanisms raised by the immune system itself and by the tumor microenvironment [1,2]. It is now well demonstrated that DCs are the central cell population making the decision between immunity or immune toler- ance, depending on the stimuli that they receive [2,3]. For instance, the development of tolerogenic DCs will lead to antigen tolerance in particular through the expansion of T regulatory cells (Tregs). We have recently shown that the vaccine formulation, and in particular the presence of the Toll-like Receptor (TLR) agonist CpG, can significantly promote the maturation of pro-inflammatory DCs, which favors Type I T cell responses while restricting the expansion of Tregs . In addition to TLR ligands, CD1d-restricted invariant NKT (iNKT) cells have been shown to efficiently promote the transactivation of DCs through the CD40L- CD40 interaction upon recognition of the CD1d-glycolipid antigen complex by their semi-invariant TCR. In view of the capacity of iNKT cells to promote DC maturation and NK cell transactivation, several pre-clinical studies have investigated the use of the CD1d/iNKT super agonist α- galactosylceramide (αGalCer), or a related analog as vaccine adjuvant [5-7]. More recently, several studies have demon- strated a cooperative effect on DC maturation between TLR ligands and iNKT cell activation, resulting in highly pro-inflammatory DCs, as seen by enhanced expression of activation markers [8-10]. However, if generating pro- inflammatory DCs should result in a good expansion of antigen-specific T cells, it will not guarantee their efficient homing to the tumor, unless a targeting strategy is used. In this context, we have developed CD1d-antitumor fusion proteins consisting of the soluble part of the CD1d molecule fused to a scFv antibody fragment specific for the tumor an- tigens CEA or HER2. We have previously demonstrated in vitro and in vivo that these fusion proteins were able to redirect iNKT, NK and T cells to the tumor expressing the relevant antigen resulting in a potent antitumor effect [11,12]. In the present study, we aimed to combine a CpG- based peptidevaccine with the activation and tumor target- ing of iNKT cells via the CD1d-anti-HER2 fusion protein.
Abstract: Ebola virus (EBOV) is one of the lethal viruses, causing more than 24 epidemic outbreaks to date. Despite having available molecular knowledge of this virus, no definite vaccine or other remedial agents have been developed yet for the management and avoidance of EBOV infections in humans. Disclosing this, the present study described an epitope-based peptidevaccine against EBOV, using a combination of B-cell and T-cell epitope predictions, followed by molecular docking and molecular dynamics simulation approach. Here, protein sequences of all glycoproteins of EBOV were collected and examined via in silico methods to determine the most immunogenic protein. From the identified antigenic protein, the peptide region ranging from 186 to 220 and the sequence HKEGAFFLY from the positions of 154–162 were considered the most potential B-cell and T-cell epitopes, correspondingly. Moreover, this peptide (HKEGAFFLY) interacted with HLA-A*32:15 with the highest bind- ing energy and stability, and also a good conservancy of 83.85% with maximum population coverage. The results imply that the designed epitopes could manifest vigorous enduring defensive immunity against EBOV.
Our vaccination approach utilized the adjuvant Montanide ISA-51 along with the cytokine GM-CSF included in the emulsion. It is not clear that this is an optimal vaccine adjuvant for peptide vaccines, and fur- ther improvements in the vaccine formulation are con- ceivable. After this trial was initiated, Slingluff and colleagues reported that the inclusion of GM-CSF with Montanide may result in diminished peptide-induced T cell responses in a melanoma vaccine study . The TLR agonists CpG 7909 and polyIC:LC have recently been explored and should continue to be investigated as a possible adjuvant . The MAGE-3 protein-based vaccine from GSK-Bio utilizes a combination of the TLR9 agonist CpG7909 and the TLR4 agonist MPL . Other cytokines with immune-potentiating activities could be considered, including IL-12 [37-39]. Thus, while our study has identified an optimal dose of CEA peptide, further improvements in vaccine potency might be achievable through optimization of the adjuvant component.
Vaccine effectiveness depends on host (immune response) factors and pathogen (strain coverage) factors. Polysaccharide and conjugate vaccine immunogenicity is assessed by the serum bactericidal antibody (SBA) assay, which measures complement- mediated killing for representative strains expressing different serogroups (18–22). Strain coverage for vaccines which target the polysaccharide can be estimated by observing the serogroup frequencies of invasive meningococcal populations. This approach is not practical for subcapsular protein antigens, which are likely to vary both in their levels of expression and in their peptide sequences across the many different endemic disease-associated meningococcal strains (23, 24). MLST-based lineage fre- quencies cannot be used either, due to the dynamic and highly recombining popula- tion structure of meningococcal species (25).
IgA is the major antibody isotype present in mucosal secretions (1) and is important in protective responses against viral infections. However, it remains unclear whether HIV-1-speciﬁc IgA responses are beneﬁcial or detrimental (31). Serum anti-Env IgA titers were directly correlated with acquisition risk in the RV144 clinical trial (6, 7), and the mechanism of this effect has been hypothesized to involve C1-speciﬁc IgA that reduces the ADCC function of Env-speciﬁc IgG (7). However, it has also been suggested that Env-speciﬁc HIV-1 IgA responses may only be markers and not actual mechanisms of risk of infection (32). Moreover, it is unclear whether correlates from an ALVAC/gp120 vaccine would be relevant for an Ad26/gp140 vaccine. We utilized the high-throughput peptide microarray as a tool to study vaccine-elicited IgA antibody responses in greater detail, with the limitation that conformational epitopes were not evaluated in this assay. Peptide microarrays were utilized to assess antibody diversity against HIV-1 linear epitopes (6, 33). We detected consistent systemic and mucosal IgA responses, which generally had lower titers than IgG responses. Puriﬁed IgG and IgA samples from our vaccinated monkeys bound to similar regions within the Env protein, predominantly in the V1 to V3, C4, and C5 regions (Fig. 4B), with minimal to no linear IgG and IgA responses being directed against the C1 linear epitope. The MAb A32 blocks ADCC and binds a conformational C1 epitope (22, 34). IgA did not block A32 binding at concen- trations up to 20 g/ml (Fig. 6).
(S). The plasmid also contains a T H epitope, ovalbumin (OVA) (323–339) (O), which is targeted to the MHC II bind- ing pathway by the dileucine lysosomal targeting motif (LL) (36). The second plasmid, RAOLL, is identical to SAOLL except that it encodes the rat KC chemokine (R) leader (Fig. 1A). Subsequently, DNA was prepared using the EndoFree Giga Prep kit (Qiagen, Valencia, CA). Three- to 5-week-old female BALB/cJ mice (The Jackson Laboratory, Bar Harbor, ME) were injected intradermally (i.d.) using a 26-gauge syringe with 200 g of DNA in 80 l of endotoxin-free phosphate- buffered saline (PBS; Sigma-Aldrich, St. Louis, MO) and were boosted twice at 2-week intervals with the same amount of DNA. Two weeks later, mice were sacrificed by cervical dislo- cation and the spleens were excised from immunized and non- immunized (naive) mice. Splenocytes from individual mice were then in vitro stimulated with 1 M gp120 peptide (317– 326) for 5 days and used in a chromium release assay. Immu- nization of mice with the RAOLL vector elicited statistically significantly higher CTL responses than did immunization with the SAOLL vector (P ⬍ 0.01) (Fig. 1B). On average, we ob- served 20% higher specific lysis in mice immunized with the RAOLL vector than in mice immunized with the SAOLL vec- tor. Thus, the use of the rat KC chemokine leader resulted in improved immune responses against the CTL epitope.