Peptide-specific Immune Responses Research Articles

Testing a vaccine candidate against Hepatitis C virus designed by combinatorial optimization

This paper presents a new procedure for vaccine design against highly variable viruses such as Hepatitis C. The procedure uses an optimization algorithm to design vaccines that maximize the coverage of epitopes across different virus variants. Weighted epitopes based on the success ratio of immunological assays are used to prioritize the selection of epitopes for vaccine design. The procedure was successfully applied to design DC vaccines loaded with two HCV peptides, STG and DYP, which were shown to be safe, immunogenic, and able to induce significant levels of anti-viral cytokines, peptide-specific cellular immune responses and IgG antibodies. The procedure could potentially be applied to other highly variable viruses that currently lack effective vaccines.

Experimental cystic echinococcosis as a proof of concept for the development of peptide-based vaccines following a novel rational workflow

Vaccines are among the most important advances in medicine throughout the human history. However, conventional vaccines exhibit several drawbacks in terms of design and production costs. Peptide-based vaccines are attractive alternatives, since they can be designed mainly in silico, can be produced cheaply and safely, and are able to induce immune responses exclusively towards protective epitopes. Yet, a proper peptide design is needed, not only to generate peptide-specific immune responses, but also for them to recognize the native protein in the occurrence of a natural infection. Herein, we propose a rational workflow for developing peptide-based vaccines including novel steps that assure the cross-recognition of native proteins. In this regard, we increased the probability of generating efficient antibodies through the selection of linear B-cell epitopes free of post-translational modifications followed by analyzing the 3D-structure similarity between the peptide in-solution vs. within its parental native protein. As a proof of concept, this workflow was applied to a set of seven previously suggested potential protective antigens against the infection by Echinococcus granulosus sensu lato. Finally, two peptides were obtained showing the capacity to induce specific antibodies able to exert anti-parasite activities in different in vitro settings, as well as to provide significant protection in the murine model of secondary echinococcosis.

Durable and dynamic hTERT immune responses following vaccination with the long-peptide cancer vaccine UV1: long-term follow-up of three phase I clinical trials

BackgroundTherapeutic cancer vaccines represent a promising approach to improve clinical outcomes with immune checkpoint inhibition. UV1 is a second generation telomerase-targeting therapeutic cancer vaccine being investigated across multiple indications. Although...

Phase I trial to determine safety and immunogenicity of amplivant, a synthetic toll-like receptor 2 ligand, conjugated to two HPV16 E6 synthetic long peptides.

2614 Background: Therapeutic vaccines based on synthetic long peptides (SLPs) have a great potential for immunotherapy of cancer patients as these SLPs include both human leukocyte antigen (HLA) class I and II epitopes and no patient selection for HLA types is required. The antigen-induced immune response can be strengthened with immune stimulating additives. Amplivant (AV) is a synthetic Toll-like receptor 2 ligand which can be directly conjugated to tumor peptide antigens. In preclinical studies, AV-conjugation to antigens led to both enhanced antigen presentation by dendritic cells and T-cell priming and caused superior induction of effective anti-tumor responses. Moreover, AV-conjugated SLPs showed a 100 times higher immune response compared to unconjugated SLP. The current study is a first-in-human trial to investigate safety and immunogenicity of AV-conjugated human papillomavirus (HPV)16-SLPs. Methods: A dose escalation phase I trial was performed in 24 patients with HPV16 positive (pre-) malignant lesions. AV was conjugated to two SLPs derived from the most immunodominant regions of the HPV16 E6 oncoprotein. Four dose groups (1, 5, 20 or 50 μg of each peptide) in 6 patients each were studied. The vaccine was injected three times intradermally in DMSO / water with a three-week interval. Adverse events (AE) were collected according to CTCAE v4.0 up to 26 weeks. Peptide-specific T-cell immune responses were determined in blood samples taken before and after vaccination using complementary immunological assays (proliferation assay, IFNγ-ELISPOT and cytokine bead array). Results: Toxicity after three AV-conjugated HPV16-SLP vaccinations was limited to CTCAE grade 1 or 2, with predominantly inflammation at the vaccination site and sometimes flu-like symptoms, which generally resolved within one day. Dose increase resulted from no AE in the lowest dose group to mild/moderate AE in all vaccinated persons in the highest dose group. In the lowest dose group, minor vaccine-induced T-cell responses were observed in three of six vaccinated persons. In the highest dose group, all patients displayed a strong HPV16-specific T-cell response after vaccination. The induced T-cell response against HPV16 lasted until the end of the trial. Conclusions: This first-in-human study showed that AV conjugated to SLPs can safely be used as an intradermal therapeutic vaccine. AV-conjugated HPV16-SLP was able to induce robust HPV16-specific T-cell immunity in patients treated for HPV16 positive (pre-) malignancies without any other vaccine adjuvant or formulation. Increase in dose resulted in both a higher number of mild adverse events as well as stronger T-cell immunity. Clinical trial information: NCT02821494.

A peptide-based subunit candidate vaccine against SARS-CoV-2 delivered by biodegradable mesoporous silica nanoparticles induced high humoral and cellular immunity in mice.

Development of a rapidly scalable vaccine is still an urgent task to halt the spread of COVID-19. We have demonstrated biodegradable mesoporous silica nanoparticles (BMSNs) as a good drug delivery carrier for tumor therapy. In this study, seven linear B cell epitopes and three CD8+ T cell epitopes were screened from the spike (S) glycoprotein of SARS-CoV-2 by computer-based immunoinformatic approaches for vaccine design. A nanoparticle-based candidate vaccine (B/T@BMSNs) against SARS-CoV-2 was rapidly prepared by encapsulating these ten epitope peptides within BMSNs, respectively. BMSNs with potential biodegradability, proved to possess excellent safety in vitro and in vivo, could efficiently deliver epitope peptides into the cytoplasm of RAW264.7 cells. Strong Th1-biased humoral and cellular immunity were induced by B/T@BMSNs in mice and all the 10 selected epitopes were identified as effective antigen epitopes, which could induce robust peptide-specific immune response. The elicited functional antibody could bind to the recombinant S protein and block the binding of the S protein to the ACE-2 receptor. These results demonstrate the potential of a nanoparticles vaccine platform based on BMSNs to rapidly develop peptide-based subunit vaccine candidates against SARS-CoV-2.

Early Phase Clinical Immunogenicity of Valoctocogene Roxaparvovec, an AAV5-Mediated Gene Therapy for Hemophilia A

Evaluation of immune responses to adeno-associated virus (AAV)-mediated gene therapies prior to and following dose administration plays a key role in determining therapeutic safety and efficacy. This report describes up to 3 years of immunogenicity data following administration of valoctocogene roxaparvovec (BMN 270), an AAV5-mediated gene therapy encoding human B domain-deleted FVIII (hFVIII-SQ) in a phase 1/2 clinical study of adult males with severe hemophilia A. Patients with pre-existing humoral immunity to AAV5 or with a history of FVIII inhibitors were excluded from the trial. Blood plasma and peripheral blood mononuclear cell (PBMC) samples were collected at regular intervals following dose administration for assessment of humoral and cellular immune responses to both the AAV5 vector and transgene-expressed hFVIII-SQ. The predominant immune response elicited by BMN 270 administration was largely limited to the development of antibodies against the AAV5 capsid that were cross-reactive with other common AAV serotypes. No FVIII inhibitor responses were observed within 3 years following dose administration. In a context of prophylactic or on-demand corticosteroid immunosuppression given after vector infusion, AAV5 and hFVIII-SQ peptide-specific cellular immune responses were intermittently detected by an interferon (IFN)-γ and tumor necrosis factor (TNF)-α FluoroSpot assay, but they were not clearly associated with detrimental safety events or changes in efficacy measures.

Leishmania-Specific Promiscuous Membrane Protein Tubulin Folding Cofactor D Divulges Th1/Th2 Polarization in the Host via ERK-1/2 and p38 MAPK Signaling Cascade.

Visceral leishmaniasis (VL)-related mortality and morbidity imposes a great deal of health concern across the globe. The existing anti-leishmanial drug regimen generally fails to eliminate newly emerging resistant isolates of this dreadful parasite. In such circumstances, the development of a prophylactic strategy to impart protection against the disease is likely to take center stage. In order to develop a promising prophylactic vaccine, it is desirable to identify an adequately potential vaccine candidate. In silico analysis of Leishmania tubulin folding cofactor D protein predicted its potential to activate both B- and T-cell repertoires. Furthermore, the ELISA employing anti-peptide27 (a segment of tubulin folding cofactor D) antibody revealed its proficiency in VL diagnosis and treatment monitoring. The peptide27 and its cocktail with another Leishmania peptide (peptide23) prompted the up-regulation of pro-inflammatory cytokines, such as IFN-γ, TNF-α, IL-2, IL-17, etc., and the down-regulation of immune-regulatory cytokines, such as IL-10, in the immunized BALB/c mice. Coherent to the consequence of peptide-specific humoral immune response, peptide cocktail-based immunization ensued in the predominant amplification of pathogen-specific IgG2a over the IgG1 isotype, up-regulated proliferation of T lymphocytes, and enhanced production of nitric oxide, reactive oxygen species, etc. We also established that the peptide cocktail modulated host MAPK signaling to favor the amplification of Th1-dominated immune response in the host. The peptide cocktail mediated the activation of the host immune armory, which was eventually translated into a significant decline in parasitic load in the visceral organs of experimental animals challenged with Leishmania donovani.

A phase I study of multi-HLA-binding peptides derived from heat shock protein 70/glypican-3 and a novel combination adjuvant of hLAG-3Ig and Poly-ICLC for patients with metastatic gastrointestinal cancers: YNP01 trial

BackgroundThis phase I study aimed to evaluate the safety, peptide-specific immune responses, and anti-tumor effects of a novel vaccination therapy comprising multi-HLA-binding heat shock protein (HSP) 70/glypican-3 (GPC3) peptides and a novel adjuvant combination of hLAG-3Ig and Poly-ICLC against metastatic gastrointestinal cancers.MethodsHSP70/GPC3 peptides with high binding affinities for three HLA types (A*24:02, A*02:01, and A*02:06) were identified with our peptide prediction system. The peptides were intradermally administered with combined adjuvants on a weekly basis. This study was a phase I dose escalation clinical trial, which was carried out in a three patients’ cohort; in total, 11 patients were enrolled for the recommended dose.ResultsSeventeen patients received this vaccination therapy without dose-limiting toxicity. All treatment-related adverse events were of grades 1 to 2. Peptide-specific CTL induction by HSP70 and GPC3 proteins was observed in 11 (64.7%) and 13 (76.5%) cases, respectively, regardless of the HLA type. Serum tumor marker levels were decreased in 10 cases (58.8%). Immunological analysis using PBMCs indicated that patients receiving dose level 3 presented with significantly reduced T cell immunoglobulin and mucin-domain containing-3 (TIM3)-expressing CD4 + T cells after one course of treatment. PD-1 or TIM3-expressing CD4 + T cells and T cell immunoreceptor with immunoglobulin and ITIM domains (TIGIT)-expressing CD8 + T cells in PBMCs before vaccination were negative predictive factors for survival.ConclusionsThis novel peptide vaccination therapy was safe for patients with metastatic gastrointestinal cancers.

Heterologous peptide display on chromatin nanofibers: A new strategy for peptide vaccines

Chromatin organization starts from a “beads-on-a string” 10 nm fiber, a basic nucleosomal structure consisting of DNA and core histones. Given its regular nucleosome array on DNA backbone where N-terminal tails of each histone are exposed on the surface of chromatin fiber, we hypothesized that chromatin can be utilized as a heterologous peptide carrier to elicit a peptide-specific immune response. The plasmid DNA containing the Widom’s clone 601 sequence and the recombinant chimeric histones containing the peptide derived from ras oncogene (G12V) were used to assemble the chromatin fiber in vitro. The immunogenicity of the assembled chromatin was tested in mice as a single vaccine component or formulated with adjuvants. G12V tagged-chromatin co-administered with adjuvants induced higher antibody responses against the G12V peptide than vaccination with adjuvant alone, while chimeric histones did not generate a significant antibody response. Interestingly, splenocytes from mice vaccinated with the G12V tagged-chromatin vaccine did not generate significant antigen-specific cytokine responses. Our studies suggest that chromatin can be utilized as an effective carrier of antigenic peptides for inducing specific antibody responses.

Application potential of bacterial flagellin in treatment of autoimmune rheumatic diseases with Treg epitope peptides

Immunization with regulatory T cell (Treg) epitope peptides to activate and induce Tregs, by which to suppress pathological autoimmune responses and reconstitute a new homeostasis, is a promising therapeutic regimen for autoimmune rheumatic diseases. However, it is usually hard to induce potent peptide-specific immune responses in vivo with small molecular peptides. Bacterial flagellin is one of the agonists triggering innate immune responses. When used as carrier, it shows strong adjuvant activity to its conjugated antigens. In some particular situations, bacterial flagellin can also activate and induce Tregs. Thus if Treg epitope peptides are covalently conjugated to a bacterial flagellin, the conjugates should be able to effectively enhance the Treg-based immune responses via flagellin itself and the adjuvanticity of flagellin to Treg epitope peptides, and thereby enhance the immunotherapeutic effects on autoimmune rheumatic diseases. Key words: Bacterial flagellin; Treg epitope; Autoimmune rheumatic disease; Conjugate; Vaccine efficacy

EpCAM peptide-primed dendritic cell vaccination confers significant anti-tumor immunity in hepatocellular carcinoma cells.

Cancer stem-like cells (CSCs) may play a key role in tumor initiation, self-renewal, differentiation, and resistance to current treatments. Dendritic cells (DCs) play a vital role in host immune reactions as well as antigen presentation. In this study, we explored the suitability of using CSC peptides as antigen sources for DC vaccination against human breast cancer and hepatocellular carcinoma (HCC) with the aim of achieving CSC targeting and enhancing anti-tumor immunity. CD44 is used as a CSC marker for breast cancer and EpCAM is used as a CSC marker for HCC. We selected CD44 and EpCAM peptides that bind to HLA-A2 molecules on the basis of their binding affinity, as determined by a peptide-T2 binding assay. Our data showed that CSCs express high levels of tumor-associated antigens (TAAs) as well as major histocompatibility complex (MHC) molecules. Pulsing DCs with CD44 and EpCAM peptides resulted in the efficient generation of mature DCs (mDCs), thus enhancing T cell stimulation and generating potent cytotoxic T lymphocytes (CTLs). The activation of CSC peptide-specific immune responses by the DC vaccine in combination with standard chemotherapy may provide better clinical outcomes in advanced carcinomas.

Polyactin A is a novel and potent immunological adjuvant for peptide-based cancer vaccine

Synthetic peptide cancer vaccines are poorly immunogenic sub-unit vaccines and thus essentially need adjuvants in their formulations to increase the efficacy by enhancing the peptide-specific immune response. However, aluminum-based compounds are almost dependent for clinical use at present. Therefore, the increasing use of peptide-based vaccines makes the need for novel and potent adjuvants. Polyactin A (PAA) has been used for the clinical treatment of impaired immunity in China for over 30years. To figure out the adjuvant effects of PAA for E75 peptide breast cancer vaccine (Her2 p369-p377), the generation of mature dendritic cells (DCs) from peripheral blood monocytes (PBMCs) cultured with PAA, IL-4 and TNF-α was assessed by morphologic features and the expressions of special surface markers using flow cytometry. Then the functional features of PBMCs-derived DCs cultured with PAA, IL-4 and TNF-α were investigated by inducing E75-specific cytotoxicity. Finally, C57BL/6-Tg(HLA-A2.1)1Enge/J transgenic mice were immunized with E75 and various amounts of PAA, and splenic lymphocyte proliferation and the IFN-γ level were determined. The results showed that PAA, just like granulocyte-macrophage colony stimulating factor, with IL-4 and TNF-α efficiently induced mature DCs from PBMCs, and these DCs could trigger a potent E75 peptide-specific CD8+ T-cell response in vitro. Immunization with E75 and PAA significantly increased positive rates of CD4+ and CD8+ T lymphocytes, and enhanced splenocytes proliferation and levels of IFN-γ in splenocytes when induced by E75. Our results indicated that PAA could efficiently induce E75-specific immunologic responses in vitro and in vivo. Therefore, PAA possesses potent adjuvant effect on peptide-based cancer vaccine. Our study provides a safe, effective and novel adjuvant for clinical use.

Immunological efficacy of glypican-3 peptide vaccine in patients with advanced hepatocellular carcinoma

ABSTRACTWe have previously conducted a phase I trial to test the efficacy of a glypican-3 (GPC3) peptide vaccine in patients with advanced hepatocellular carcinoma (HCC); however, its immunological mechanism of action remains unclear. Here, we report a pilot study conducted to evaluate the immunological mechanisms of action of this GPC3 peptide vaccine (UMIN-CTR number 000005093). Eleven patients with advanced HCC were vaccinated with the GPC3 peptide in this trial. The primary end point was GPC3 peptide-specific immune response induced by the GPC3 peptide vaccination. The secondary endpoints were clinical and biologic outcomes. We demonstrated that the present vaccine induced GPC3 peptide-specific cytotoxic T lymphocytes (CTLs), which were found to infiltrate into the tumor. Moreover, we established GPC3 peptide-specific CTL clones from a biopsy specimen: these cells exhibited GPC3 peptide-specific cytokine secretion and cell cytotoxicity. The plasma GPC3 level tended to decrease temporarily at least once during the follow-up period. The GPC3-specific CTL frequency after vaccination was correlated with overall survival. The degree of skin reactions at the injection site correlated with the GPC3 peptide-specific CTLs. Furthermore, we sequenced the T cell receptors (TCRs) of tumor-infiltrating lymphocyte (TIL) clones, and confirmed the existence of this TCR repertoire in both tumor tissue and PBMCs. In response to these data, we are developing TCR-engineered T cell therapy using TCR sequences obtained from GPC3 peptide-specific CTL clones for improved efficacy in patients with advanced HCC.

Immunization with antigenic peptides complexed with β-glucan induces potent cytotoxic T-lymphocyte activity in combination with CpG-ODNs

The induction of antigen-specific immune responses requires immunization with not only antigens, but also adjuvants. CpG oligonucleotides (CpG-ODNs) are well-known ligands for Toll-like receptor 9 and a potent adjuvant that induces both Th1-type humoral and cellular immune responses including cytotoxic T-lymphocyte responses. We previously demonstrated that β-glucan schizophyllan (SPG) can form complexes with CpG-ODNs with attached dA40 (CpG-dA/SPG), which can accumulate in macrophages in the draining inguinal lymph nodes and induce strong immune responses by co-administration of antigenic proteins, namely ovalbumin (OVA). Immunization with antigenic peptides, OVA257–264, did not induce these antigen-specific immune responses even in combination with CpG-dA/SPG, indicating that peptides require a carrier to antigen presenting cells. In this study, we prepared conjugates comprising OVA257–264 and dA40, and made complexes with SPG. Immunization with OVA257–264-dA/SPG induced peptide-specific immune responses in combination with CpG-dA regardless of complexation with SPG both in vitro and in vivo. When splenocytes from immunized mice were incubated with E.G7-OVA tumor model cells presenting OVA peptides, the number of cells drastically decreased after 24h. Furthermore, mice pre-immunized with OVA257–264-dA/SPG and CpG-ODNs exhibited a long delay in tumor growth after tumor inoculation. Therefore, these peptide-dA/SPG and CpG-dA/SPG complexes could be used as a potent vaccine for the treatment of cancers and infectious diseases.

Administration of sulfosuccinimidyl-4-[N-maleimidomethyl] cyclohexane-1-carboxylate conjugated GP10025–33 peptide-coupled spleen cells effectively mounts antigen-specific immune response against mouse melanoma

It remains a top research priority to develop immunotherapeutic approaches to induce potent antigen-specific immune responses against tumors. However, in spite of some promising results, most strategies are ineffective because they generate low numbers of tumor-reactive cytotoxic T lymphocytes (CTLs). Here we designed a strategy to enhance antigen-specific immune response via administering sulfosuccinimidyl-4-[N-maleimidomethyl] cyclohexane-1-carboxylate (sulfo-SMCC)-conjugated melanoma tumor antigen GP10025–33 peptide-coupled syngeneic spleen cells in a mouse model of melanoma. We found that infusion of GP10025–33 peptide-coupled spleen cells significantly attenuated the growth of melanoma in prophylactic and therapeutic immunizations. Consistent with these findings, the adoptive transfer of spleen cells from immunized mice to naïve syngeneic mice was able to transfer anti-tumor effect, suggesting that GP10025–33 peptide-specific immune response was induced. Further studies showed that, CD8+ T cell proliferation and the frequency of interferon (IFN)-γ-producing CD8+ T cells upon ex vivo stimulation by GP10025–33 were significantly increased compared to control groups. Tumor antigen, GP10025–23 specific immune response was also confirmed by ELISpot and GP100-tetramer assays. This approach is simple, easy-handled, and efficiently delivering antigens to lymphoid tissues. Our study offers an opportunity for clinically translating this approach into tumor immunotherapy.

MP2-20 FEASIBILITY STUDY OF PERSONALIZED PEPTIDE VACCINATION FOR PLATINUM-BASED CHEMOTHERAPY RESISTANT METASTATIC UPPER TRACT UROTHELIAL CARCINOMA PATIENTS

You have accessJournal of UrologyBladder Cancer: Upper Tract TCC I1 Apr 2015MP2-20 FEASIBILITY STUDY OF PERSONALIZED PEPTIDE VACCINATION FOR PLATINUM-BASED CHEMOTHERAPY RESISTANT METASTATIC UPPER TRACT UROTHELIAL CARCINOMA PATIENTS Shigetaka Suekane, Masanori Noguchi, Kousuke Ueda, Tsukasa Igawa, Tetsuro Sasada, and Kyogo Itoh Shigetaka SuekaneShigetaka Suekane More articles by this author , Masanori NoguchiMasanori Noguchi More articles by this author , Kousuke UedaKousuke Ueda More articles by this author , Tsukasa IgawaTsukasa Igawa More articles by this author , Tetsuro SasadaTetsuro Sasada More articles by this author , and Kyogo ItohKyogo Itoh More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2015.02.143AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES The prognosis of metastatic upper tract urothelial carcinoma ( UTUC ) patients who failed platinum-based chemotherapy remains very poor. We conducted a feasibility study of personalized peptide vaccine ( PPV ) for platinum-based chemotherapy resistant metastatic UTUC patients to explore cancer vaccine as a new treatment modality. METHODS A total of 34 UTUC patients were enrolled to this study, who were platinum-based chemotherapy resistant metastatic 21 pelvic cancer and 13 ureter cancer. A maximum of four HLA-matched peptides showing higher antigen-specific immunoglobulin G ( IgG ) responses were selected based on measurement of IgG against different cytotoxic T lymphocyte ( CTL ) epitope peptides followed by subcutaneous vaccination. This study protocol was approved by the institutional ethical review boards of Kurume University, and conducted in accordance with the Declaration of Helsinki. Written informed consent was obtained from all patients before entering this clinical trial. RESULTS There were no vaccine-related severe adverse events of PPV in all patients. Median overall survival ( OS ) time from the first PPV was 13.0 months in platinum-based chemotherapy resistant metastatic UTUC patients. Increased the anti-peptide IgG titer were revealed seventeen patients ( 50.0% ) of thirty-four patients at post 6th vaccination, and sixteen patients ( 100% ) of sixteen patients at post 12th vaccination. Seventeen patients with 12th more PPV showed significantly longer survival than 17 patients with less than 12th PPV (hazard ratio [HR], 4.775, P=0.0015). Peptide-specific immune response at 12th PPV in platinum-based chemotherapy resistant metastatic UTUC patients was favorable biomarker for OS ( P=0.0002 ), regardless of the response at 6th PPV. CONCLUSIONS PPV as a new treatment modality in patients with platinum-based chemotherapy resistant metastatic UTUC is active and well tolerated, improving survival with immune responses. Further large-scale, randomized trials are needed to confirm these results. © 2015 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 193Issue 4SApril 2015Page: e18 Advertisement Copyright & Permissions© 2015 by American Urological Association Education and Research, Inc.MetricsAuthor Information Shigetaka Suekane More articles by this author Masanori Noguchi More articles by this author Kousuke Ueda More articles by this author Tsukasa Igawa More articles by this author Tetsuro Sasada More articles by this author Kyogo Itoh More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

Immunogenicity of GX301 cancer vaccine: Four (telomerase peptides) are better than one

Peptide540–548, peptide611–626, peptide672–686 and peptide766–780, which are derived from human telomerase, constitute the immunogenic component of the GX301 cancer vaccine. The relative immunogenicity of these peptides is unknown, thus it is unsure whether their combined use offers real advantages over single peptide stimulation. Hence, this study compared the number of specific immune responses and responders to each peptide, as well as to their mixture (meaning the co-presence of the 4 peptides in the same culture well), achieved after ex vivo stimulation of PBMC from 21, HLA-A2+ (n.11) or HLA-A2- (n.10), healthy donors. The study was performed on freshly collected PBMC (T0) and on PBMC stimulated for 10 d with single peptides or their mixture (T1). Peptide-specific immune responses were analyzed by Elispot and cytokine intracellular staining by flow cytometry. The results showed that each peptide induced specific immune responses in some subjects, with different panels of responders among the peptides. Moreover, the numbers of responses and responders to the single peptides or their mixture were comparable. Importantly, the overall number of responders to the 4 peptides was higher than to each single peptide, or to their mixture, both at T0 and T1. These data demonstrate the immunogenicity of each of the 4 GX301 telomerase peptides. Moreover, they show the advantage of multi-peptide over single peptide stimulation, providing a clear support to their combined administration in vaccination protocols. However, the data pose a warning against peptide administration as a mixture due to possible interference phenomena during antigen presentation processes.

Increasing cellular immunogenicity to peptide-based vaccine candidates using a fluorocarbon antigen delivery system

Traditionally, synthetic peptide vaccines for infectious diseases and cancer require adjuvants to achieve optimal immunogenicity. Here we describe a novel method of peptide modification using a fluorocarbon chain which can substantially increase peptide-specific cellular immune responses in the absence of adjuvant. We demonstrate that fluorocarbon-modified peptides (fluoropeptides) derived from HIV, influenza and hepatitis C virus can significantly increase interferon gamma ELISpot responses against cytotoxic and T-helper epitopes compared to unmodified peptides or lipopeptides in mice. Increases in both T-helper1 and T-helper2 cytokines are observed. Fluoropeptides show enhanced ability of the antigen to persist at the site of administration and persistence is associated with a prolonged and elevated immune response. Additionally we demonstrate that fluoropeptides have increased proteolytic resistance thereby potentially supporting their increased half-life in vivo. Fluorocarbon-modification of peptides provides a valuable tool for increasing cellular immunogenicity of vaccines for infectious diseases and cancer without requirement for traditional adjuvants.

Epitope shifting of gp90-specific cellular immune responses in EIAV-infected ponies

Unlike other lentiviruses, EIAV replication can be controlled in most infected horses leading to an inapparent carrier state free of overt clinical signs which lasts for many years. While the resolution of the initial infection is correlated with the appearance of virus specific cellular immune responses, the precise immune mechanisms responsible for control of the infection are not yet identified. Since the virus undergoes rapid mutation following infection, the immune response must also adapt to meet this challenge. We hypothesize that this adaptation involves peptide-specific recognition shifting from immunodominant variable determinants to conserved immunorecessive determinants following EIAV infection. Forty-four peptides, spanning the entire surface unit protein (gp90) of EIAV, were used to monitor peptide-specific T cell responses in vivo over a six-month period following infection. Peptides were injected intradermally and punch biopsies were collected for real-time PCR analysis to monitor the cellular peptide-specific immune responses in vivo. Similar to the CMI response to HIV infection, peptide-specific T cell recognition patterns changed over time. Early post infection (1 month), immune responses were directed to the peptides in the carboxyl-terminus variable region. By six months post infection, the peptide recognition spanned the entire gp90 sequence. These results indicate that peptide recognition broadens during EIAV infection.

A phase II trial evaluating the clinical and immunologic response of HLA-A2+ non-small cell lung cancer patients vaccinated with an hTERT cryptic peptide

ObjectivesThe immunological and clinical responses of patients with NSCLC treated, in the context of an expanded action program, with the cryptic hTERT-targeting Vx-001 vaccine are presented. Materials and methodsForty-six HLA-A*0201-positive patients with advanced NSCLC and residual (n=27) or progressive (n=19) disease following front-line treatment received two subcutaneous injections of the optimized TERT572Y peptide followed by four injections of the native TERT572 peptide, every 3 weeks. Peptide-specific immune responses were monitored by enzyme-linked immunosorbent spot assay at baseline, and after the 2nd and the 6th vaccinations. Thirty-eight HLA-A*0201-positive matched patients were used as historical controls. ResultsTwenty-three patients (50%) completed the vaccination protocol and 87% received at least two administrations. Twelve patients (26%) without disease progression after the 6th vaccination received boost vaccinations. Three (7%) patients achieved a partial response and 13 (28%) disease stabilization. The disease control rate was significantly higher in patients with non-squamous histology compared to those with squamous-cell histology [n=14 (45%) versus n=2 (13%); p=0.03]. The median progression-free survival (PFS) and overall survival (OS) was 3.8 (range, 0.7–99.4) and 19.8 months (range, 0.7–99.4), respectively. Patients who developed immune response had a numerically higher PFS compared to those who failed to mount any (6.7 versus 2.7 months; p=0.090). However, the median OS for the immune-responders was significantly prolonged compared to non-responders (40.0 versus 9.2 months, respectively; p=0.02). Toxicity was <grade 2. ConclusionVx-001 vaccine is well tolerated and induced a TERT-specific immunological response, which was significantly correlated with improved clinical outcome.