Phage Vaccine Research Articles

Comparative Ability of Various Immunosuppressants as Adjuvants on the Activity of T1D Vaccine

Background: Type 1 diabetes (T1D) is an autoimmune disorder characterised by the destruction of insulin-producing beta cells in the pancreatic islets, resulting from a breakdown in immunological tolerance. Currently, T1D treatment primarily relies on insulin replacement or immunosuppressive therapies. However, these approaches often have significant drawbacks, including adverse effects, high costs, and limited long-term efficacy. Consequently, there is a pressing need for innovative immunotherapeutic strategies capable of inducing antigen-specific tolerance and protecting beta cells from autoimmune destruction. Among the various antigens, β-cell antigens like 65 kDa glutamic acid decarboxylase (GAD65) have been explored as vaccine candidates for T1D. Despite their potential, their effectiveness in humans remains modest, necessitating the use of appropriate adjuvants to enhance the vaccine’s protective effects. Methods: In this study, we evaluated the therapeutic potential of kynurenine (KYN), dexamethasone (DXMS), tacrolimus (FK506), and aluminium hydroxide (Alum) in combination with the GAD65 phage vaccine as adjuvants. Results: Our findings demonstrate that KYN, when used in conjunction with the GAD65 vaccine, significantly enhances the vaccine’s immunosuppressive effects. Compared to dexamethasone, FK506, and Alum adjuvants, KYN more effectively reduced the incidence and delayed the onset of T1D, preserved β-cell function, and promoted the induction of regulatory T cells and antigen-specific tolerance. These results suggest that KYN combined with vaccines could offer superior preventive and therapeutic benefits for T1D compared to existing treatments. Additionally, we investigated the dose-dependent effects of the GAD65 vaccine by including a low-dose group in our study. The results indicated that reducing the vaccine dose below 1010 plaque-forming units (pfu) did not confer any protective advantage or therapeutic benefit in combination with KYN. This finding underscores that 1010 pfu is the minimum effective dose for the GAD65 vaccine in achieving a protective response. In conclusion, KYN shows considerable promise as an adjuvant for the GAD65 vaccine in T1D therapy, potentially offering a more effective and durable treatment option than current immunosuppressive strategies.

Adjuvanted Vaccine Induces Functional Antibodies against Pseudomonas aeruginosa Filamentous Bacteriophages.

Pseudomonas aeruginosa (Pa), a WHO priority 1 pathogen, resulted in approximately 559,000 deaths globally in 2019. Pa has a multitude of host-immune evasion strategies that enhance Pa virulence. Most clinical isolates of Pa are infected by a phage called Pf that has the ability to misdirect the host-immune response and provide structural integrity to biofilms. Previous studies demonstrate that vaccination against the coat protein (CoaB) of Pf4 virions can assist in the clearance of Pa from the dorsal wound model in mice. Here, a consensus peptide was derived from CoaB and conjugated to cross-reacting material 197 (CRM197). This conjugate was adjuvanted with a novel synthetic Toll-like receptor agonist (TLR) 4 agonist, INI-2002, and used to vaccinate mice. Mice vaccinated with CoaB-CRM conjugate and INI-2002 developed high anti-CoaB peptide-specific IgG antibody titers. Direct binding of the peptide-specific antibodies to whole-phage virus particles was demonstrated by ELISA. Furthermore, a functional assay demonstrated that antibodies generated from vaccinated mice disrupted the replicative cycle of Pf phages. The use of an adjuvanted phage vaccine targeting Pa is an innovative vaccine strategy with the potential to become a new tool targeting multi-drug-resistant Pa infections in high-risk populations.

A Recombinant RBD-Based Phage Vaccine Report: A Solution to the Prevention of New Diseases?

The safety, inherent immunogenicity, stability, and low-cost production of bacteriophages make them an ideal platform for vaccine development. Most vaccination strategies against COVID-19 have targeted the spike protein of SARS-CoV-2 to generate neutralizing antibodies. P1, a truncated RBD-derived spike protein, has been shown to induce virus-neutralizing antibodies in preclinical studies. In this study, we first investigated whether recombinant phages displaying P1 on the M13 major protein could immunize mice against COVID-19, and second, whether inoculation with 50 µg of purified P1 in addition to the recombinant phages would stimulate the immune systems of the animals. The results showed that the mice that received recombinant phages were immunized against the phage particles, but did not have anti-P1 IgG. In contrast, compared with the negative control, the group that received a combination of P1 protein and recombinant phage was immunized against the P1 protein. In both groups, CD4+ and CD8+ T cells appeared in the lung tissue. These results suggest that the number of antigens on the phage body plays a crucial role in stimulating the immune system against the bacteriophage, although it is immunogenic enough to function as a phage vaccine.

Aspects of Phage-Based Vaccines for Protein and Epitope Immunization.

Because vaccine development is a difficult process, this study reviews aspects of phages as vaccine delivery vehicles through a literature search. The results demonstrated that because phages have adjuvant properties and are safe for humans and animals, they are an excellent vaccine tool for protein and epitope immunization. The phage genome can easily be manipulated to display antigens or create DNA vaccines. Additionally, they are easy to produce on a large scale, which lowers their manufacturing costs. They are stable under various conditions, which can facilitate their transport and storage. However, no medicine regulatory agency has yet authorized phage-based vaccines despite the considerable preclinical data confirming their benefits. The skeptical perspective of phages should be overcome because humans encounter bacteriophages in their environment all the time without suffering adverse effects. The lack of clinical trials, endotoxin contamination, phage composition, and long-term negative effects are some obstacles preventing the development of phage vaccines. However, their prospects should be promising because phages are safe in clinical trials; they have been authorized as a food additive to avoid food contamination and approved for emergency use in phage therapy against difficult-to-treat antibiotic-resistant bacteria. Therefore, this encourages the use of phages in vaccines.

The Breadth of Bacteriophages Contributing to the Development of the Phage-Based Vaccines for COVID-19: An Ideal Platform to Design the Multiplex Vaccine.

Phages are highly ubiquitous biological agents, which means they are ideal tools for molecular biology and recombinant DNA technology. The development of a phage display technology was a turning point in the design of phage-based vaccines. Phages are now recognized as universal adjuvant-free nanovaccine platforms. Phages are well-suited for vaccine design owing to their high stability in harsh conditions and simple and inexpensive large-scale production. The aim of this review is to summarize the overall breadth of the antiviral therapeutic perspective of phages contributing to the development of phage-based vaccines for COVID-19. We show that phage vaccines induce a strong and specific humoral response by targeted phage particles carrying the epitopes of SARS-CoV-2. Further, the engineering of the T4 bacteriophage by CRISPR (clustered regularly interspaced short palindromic repeats) presents phage vaccines as a valuable platform with potential capabilities of genetic plasticity, intrinsic immunogenicity, and stability.

Adjuvanted multivalent vaccine targeting clinically relevant Pf bacteriophage creates cross-reactive antibodies

Abstract Pseudomonas aeruginosa (Pa) is a bacterial pathogen associated with chronic diabetic wounds as well as potentially deadly lung infections in cystic fibrosis patients. Pa is listed by the WHO as “Priority 1” in the Priority Pathogens list and results in hundreds of deaths each year. Pa diversity presents a challenge in developing a vaccine or therapeutic capable of resolving infections caused by different Pa clinical isolates. Filamentous Pf phages (a virus) infect the majority of Pa strains. Additionally, the presence Pf phage is a key virulence factor contributing to Pa infections. Pf is a filamentous-lysogenic phage that is primarily comprised of repeats of coat protein (CoaB). A consensus peptide derived from the coat protein was identified and used as the antigen for our phage vaccine targeted vaccine. This conjugate vaccine targeting Pf phage induced protection against Pa wound challenges in mice. Additionally, our team has implemented the use of a novel TLR4 agonist to further enhance antibody response against Pf coat protein intact Pf phage virions. Based on their amino acid sequence homology, clinical isolates of Pa and associated Pf phage were divided into two major clades. We prepared CRM-197 conjugates using the consensus peptides from both clades. Vaccination with these multivalent conjugates in combination with a novel synthetic TLR4 agonist (adjuvant) boosts antibody response against the peptides and promotes antibody cross-recognition between peptides. Creating a safe and effective vaccine that can mount a robust immune response against both clades of Pf phage is an important advancement towards the development and clinical testing of phage based vaccine targeting deadly Pa infections. Supported by R01 AI138981-01

Adaptive antitumor immune response stimulated by bio-nanoparticle based vaccine and checkpoint blockade

BackgroundInteractions between tumor and microenvironment determine individual response to immunotherapy. Triple negative breast cancer (TNBC) and hepatocellular carcinoma (HCC) have exhibited suboptimal responses to immune checkpoint inhibitors (ICIs). Aspartate β-hydroxylase (ASPH), an oncofetal protein and tumor associated antigen (TAA), is a potential target for immunotherapy.MethodsSubcutaneous HCC and orthotopic TNBC murine models were established in immunocompetent BALB/c mice with injection of BNL-T3 and 4 T1 cells, respectively. Immunohistochemistry, immunofluorescence, H&E, flow cytometry, ELISA and in vitro cytotoxicity assays were performed.ResultsThe ASPH-MYC signaling cascade upregulates PD-L1 expression on breast and liver tumor cells. A bio-nanoparticle based λ phage vaccine targeting ASPH was administrated to mice harboring syngeneic HCC or TNBC tumors, either alone or in combination with PD-1 blockade. In control, autocrine chemokine ligand 13 (CXCL13)-C-X-C chemokine receptor type 5 (CXCR5) axis promoted tumor development and progression in HCC and TNBC. Interactions between PD-L1+ cancer cells and PD-1+ T cells resulted in T cell exhaustion and apoptosis, causing immune evasion of cancer cells. In contrast, combination therapy (Vaccine+PD-1 inhibitor) significantly suppressed primary hepatic or mammary tumor growth (with distant pulmonary metastases in TNBC). Adaptive immune responses were attributed to expansion of activated CD4+ T helper type 1 (Th1)/CD8+ cytotoxic T cells (CTLs) that displayed enhanced effector functions, and maturation of plasma cells that secreted high titers of ASPH-specific antibody. Combination therapy significantly reduced tumor infiltration of immunosuppressive CD4+/CD25+/FOXP3+ Tregs. When the PD-1/PD-L1 signal was inhibited, CXCL13 produced by ASPH+ cancer cells recruited CXCR5+/CD8+ T lymphocytes to tertiary lymphoid structures (TLSs), comprising effector and memory CTLs, T follicular helper cells, B cell germinal center, and follicular dendritic cells. TLSs facilitate activation and maturation of DCs and actively recruit immune subsets to tumor microenvironment. These CTLs secreted CXCL13 to recruit more CXCR5+ immune cells and to lyse CXCR5+ cancer cells. Upon combination treatment, formation of TLSs predicts sensitivity to ICI blockade. Combination therapy substantially prolonged overall survival of mice with HCC or TNBC.ConclusionsSynergistic antitumor efficacy attributable to a λ phage vaccine specifically targeting ASPH, an ideal TAA, combined with ICIs, inhibits tumor growth and progression of TNBC and HCC.

The Effect of Immunosuppressive Adjuvant Kynurenine on Type 1 Diabetes Vaccine

Inducing antigen-specific tolerance is a promising treatment for preventing or reversing Type 1 diabetes (T1D). In contrast to a vaccine that induces immune responses against pathogens, a tolerogenic vaccine can suppress immunity against antigens causing diseases by administrating a mixture of self-antigens with an adjuvant that decreases the strength of antigen-specific response. Kynurenine (Kyn) is an endogenous substance that can inhibit the natural killer cell and T cell proliferation and promote the differentiation of naïve T cells into regulatory T cells (Tregs). In this study, we evaluated the efficacy of Kyn as a novel suppressive adjuvant. Kyn was co-immunized with GAD65 phage vaccine to induce Treg cells and tolerogenic responses for the prevention of T1D in NOD mouse model. Mice were subcutaneously immunized two times with 1011 Pfu (100μL,1012 Pfu/ml) GAD65 phage vaccine doses mixed with 200 μg of Kyn. Serum antibodies and cytokines were detected by ELISA and electrochemiluminescence, respectively. Flow cytometry assay was used to analyze DC and Treg. MTS was used for the analysis of spleen lymphocyte proliferation. RNA sequencing was used to investigate mRNA and miRNA expression profiles in spleen lymphocytes. Compared to GAD65 phage vaccine alone, co-immunization of Kyn and GAD65 phage vaccine resulted in the prevention of hyperglycemia in 60% of mice for at least one month. Further, Kyn enhances GAD65-specific Th2-mediated immune responses; regulates the Th1/Th2 imbalance and increases the secretion of Th2 cytokines and the number of CD4+CD25+Foxp3+T cells; suppresses DC maturation and GAD65-specific T lymphocyte proliferation. Moreover, we integrated Kyn related miRNA and mRNA expression profiles obtained from the spleen lymphocyte RNA-sequencing which was stimulated by Kyn in vitro. These data provide an important basis for understanding the mechanisms underlying Kyn as an immunosuppressive adjuvant which regulated the immune response. These findings suggest that Kyn can serve as an effective suppressive adjuvant candidate for Type 1 diabetes vaccines.

Adjuvanted bacteriophage vaccine targeting Pseudomonas aeruginosa

Abstract Pseudomonas aeruginosa (Pa) is a Gram-negative bacterium which is associated with chronic diabetic wounds as well as potentially deadly lung infections in cystic fibrosis patients. Approximately 51,000 cases of Pa infections occur annually; of these cases, 13% are resistant to antibiotics which results in the deaths of ~400 individuals every year. Diversity between Pa strains presents a challenge in developing a vaccine or therapeutic capable of resolving infections caused by different strains. Pf phage (a filamentous bacteriophage) has been found in Pa from chronic diabetic wound isolates and has been show to increase virulence of Pa. Recent findings by our team indicate that a Pf phage-targeted vaccine or monoclonal antibody provide protection from the establishment of Pa infection in mice. To further improve this innovative new vaccine targeting Pa, adjuvant systems were used to enhance humoral response to the vaccine. A consensus peptide from Pf Phage coat protein was conjugated to the carrier protein CRM197 and combined with novel adjuvants and delivery systems. Selected adjuvants significantly enhanced humoral immunity to the Pf Phage peptide in a dose-dependent manner. Combination adjuvant systems were also used to further enhance antigen-specific immunity to Pf Phage and cell-mediated immunity to the carrier protein. The development of an adjuvanted Pf phage vaccine to protect against Pa infections is a new and innovative strategy with the potential to protect patients at risk for opportunistic bacterial infections.

Bacteriophages as Potential Tools for Use in Antimicrobial Therapy and Vaccine Development.

The constantly growing number of people suffering from bacterial, viral, or fungal infections, parasitic diseases, and cancers prompts the search for innovative methods of disease prevention and treatment, especially based on vaccines and targeted therapy. An additional problem is the global threat to humanity resulting from the increasing resistance of bacteria to commonly used antibiotics. Conventional vaccines based on bacteria or viruses are common and are generally effective in preventing and controlling various infectious diseases in humans. However, there are problems with the stability of these vaccines, their transport, targeted delivery, safe use, and side effects. In this context, experimental phage therapy based on viruses replicating in bacterial cells currently offers a chance for a breakthrough in the treatment of bacterial infections. Phages are not infectious and pathogenic to eukaryotic cells and do not cause diseases in human body. Furthermore, bacterial viruses are sufficient immuno-stimulators with potential adjuvant abilities, easy to transport, and store. They can also be produced on a large scale with cost reduction. In recent years, they have also provided an ideal platform for the design and production of phage-based vaccines to induce protective host immune responses. The most promising in this group are phage-displayed vaccines, allowing for the display of immunogenic peptides or proteins on the phage surfaces, or phage DNA vaccines responsible for expression of target genes (encoding protective antigens) incorporated into the phage genome. Phage vaccines inducing the production of specific antibodies may in the future protect us against infectious diseases and constitute an effective immune tool to fight cancer. Moreover, personalized phage therapy can represent the greatest medical achievement that saves lives. This review demonstrates the latest advances and developments in the use of phage vaccines to prevent human infectious diseases; phage-based therapy, including clinical trials; and personalized treatment adapted to the patient’s needs and the type of bacterial infection. It highlights the advantages and disadvantages of experimental phage therapy and, at the same time, indicates its great potential in the treatment of various diseases, especially those resistant to commonly used antibiotics. All the analyses performed look at the rich history and development of phage therapy over the past 100 years.

Yersinia Outer Membrane Vesicles as Potential Vaccine Candidates in Protecting against Plague.

Despite the relatively low incidence of plague, its etiological agent, Yersinia pestis, is an exceptional epidemic danger due to the high infectivity and mortality of this infectious disease. Reports on the isolation of drug-resistant Y. pestis strains indicate the advisability of using asymmetric responses, such as phage therapy and vaccine prophylaxis in the fight against this problem. The current relatively effective live plague vaccine is not approved for use in most countries because of its ability to cause heavy local and system reactions and even a generalized infectious process in people with a repressed immune status or metabolic disorders, as well as lethal infection in some species of nonhuman primates. Therefore, developing alternative vaccines is of high priority and importance. However, until now, work on the development of plague vaccines has mainly focused on screening for the potential immunogens. Several investigators have identified the protective potency of bacterial outer membrane vesicles (OMVs) as a promising basis for bacterial vaccine candidates. This review is aimed at presenting these candidates of plague vaccine and the results of their analysis in animal models.

Diverse molecular functions of aspartate β‑hydroxylase in cancer (Review)

Aspartate/asparagine β-hydroxylase (AspH) is a type II transmembrane protein that catalyzes the post-translational hydroxylation of definite aspartyl and asparaginyl residues in epidermal growth factor-like domains of substrates. In the last few decades, accumulating evidence has indicated that AspH expression is upregulated in numerous types of human malignant cancer and is associated with poor survival and prognosis. The AspH protein aggregates on the surface of tumor cells, which contributes to inducing tumor cell migration, infiltration and metastasis. However, small-molecule inhibitors targeting hydroxylase activity can markedly block these processes, both in vitro and in vivo. Immunization of tumor-bearing mice with a phage vaccine fused with the AspH protein can substantially delay tumor growth and progression. Additionally, AspH antigen-specific CD4+ and CD8+ T cells were identified in the spleen of tumor-bearing mice. Therefore, these agents may be used as novel strategies for cancer treatment. The present review summarizes the current progress on the underlying mechanisms of AspH expression in cancer development.

Adjuvanted bacteriophage vaccine targeting Pseudomonas aeruginosa

Abstract Pseudomonas aeruginosa (Pa) is a Gram-negative bacterium which is associated with chronic diabetic wounds as well as potentially deadly lung infections in cystic fibrosis patients. Approximately 51,000 cases of Pa infections occur annually; of these cases, 13% are resistant to antibiotics which results in the deaths of ~400 individuals every year. Diversity between Pa strains presents a challenge in developing a vaccine or therapeutic capable of resolving infections caused by different strains. Pf phage (a filamentous bacteriophage) has been found in Pa from chronic diabetic wound isolates and has been show to increase virulence of Pa. Recent findings by our team indicate that a Pf phage-targeted vaccine or monoclonal antibody provide protection from the establishment of Pa infection in mice. To further improve this innovative new vaccine targeting Pa, adjuvant systems were used to enhance humoral response to the vaccine. A consensus peptide from Pf Phage coat protein was conjugated to the carrier protein CRM197 and combined with novel adjuvants and delivery systems. Selected adjuvants significantly enhanced humoral immunity to the Pf Phage peptide in a dose-dependent manner. Combination adjuvant systems were also used to further enhance antigen-specific immunity to Pf Phage and cell-mediated immunity to the carrier protein. The development of an adjuvanted Pf phage vaccine to protect against Pa infections is a new and innovative strategy with the potential to protect patients at risk for opportunistic bacterial infections.

Analysis of the Consolidation Phase of Immunological Memory within the IgG Response to a B Cell Epitope Displayed on a Filamentous Bacteriophage.

Immunological memory can be defined as the ability to mount a response of greater magnitude and with faster kinetics upon re-encounter of the same antigen. We have previously reported that a booster dose of a protein antigen given 15 days after the first dose interferes with the development of memory, i.e., with the ability to mount an epitope-specific IgG response of greater magnitude upon re-encounter of the same antigen. We named the time-window during which memory is vulnerable to disruption a “consolidation phase in immunological memory”, by analogy with the memory consolidation processes that occur in the nervous system to stabilize memory traces. In this study, we set out to establish if a similar memory consolidation phase occurs in the IgG response to a B cell epitope displayed on a filamentous bacteriophage. To this end, we have analyzed the time-course of anti-β-amyloid IgG titers in mice immunized with prototype Alzheimer’s Disease vaccine fdAD(2-6), which consists of a fd phage that displays the B epitope AEFRH of β -amyloid at the N-terminus of the Major Capsid Protein. A booster dose of phage fdAD(2-6) given 15 days after priming significantly reduced the ratio between the magnitude of the secondary and primary IgG response to β-amyloid. This analysis confirms, in a phage vaccine, a consolidation phase in immunological memory, occurring two weeks after priming.

Compensation for cures: Why we should pay a premium for participation in 'challenge studies'.

Antibiotic resistance is one of the most pressing public health problems humanity faces. Research into new classes of antibiotics and new kinds of treatments – including risky experimental treatments such as phage therapy and vaccines – is an important part of improving our ability to treat infectious diseases. In order to aid this research, we will argue that we should permit researchers to pay people any amount of money to compensate for the risks of participating in clinical trials, including ‘challenge studies’ that involve deliberately infecting patients. We think that standard worries about paying for participation in risky research are reducible to concerns that can be addressed with the right screening mechanisms.

Phage vaccines displaying YGKDVKDLFDYAQE epitope induce protection against systemic candidiasis in mouse model

Candida albicans is a common commensal and opportunistic fungal pathogen in human, which poses threat to human health, especially in immunocompromised patients. Unfortunately, few effective prophylactic and therapeutic strategies were applied to clinic practice. Recently, the peptide YGKDVKDLFDYAQE from Fructose-bisphosphate aldolase 1 (Fba1), as a vaccine, was reported to induce protection effects against systemic candidiasis. Here, we displayed this epitope peptide on the coat proteins (pIII or pVIII) of filamentous phage, and investigated their protective effects against C. albicans infections. Mice were immunized with recombinant phages (designated as phage-3F and phage-8F) or protein (rFba1), then challenged with C. albicans yeast cells via lateral tail vein. Results demonstrated that the recombinant phages as well as rFba1 apparently induced humoral and cellular immune responses, reduced fungal burden and relieved kidney damage in infected mice and significantly improved their survival rates. Briefly, all these findings indicated that the recombinant phages displaying the epitope YGKDVKDLFDYAQE have the potential to be developed into a new vaccine against C. albicans infections.

Immunization with tumor neoantigens displayed on T7 phage nanoparticles elicits plasma antibody and vaccine-draining lymph node B cell responses

The aim of this preclinical study was to evaluate T7 bacteriophage as a nanoparticle platform for expression of neoantigens that could allow rapid generation of vaccines for potential studies in human cancer patients. We have generated recombinant T7 phage vaccines carrying neoepitopes derived from mutated proteins of B16-F10 melanoma tumor cells. With the single mutated amino acid (AA) centered, peptides were expressed on the outer coat of T7 phage. All peptides with 11 and 34 AAs were successfully expressed. Trimers of the 11-AA peptides were successfully expressed in only 3 of 8 peptides. The 11-AA peptide was better in stimulating antibodies selective for the mutated region than the longer 34-AA peptide. We observed a dose response for vaccines which provides an initial framework of the minimum phage required for vaccination. A single injection with phage-peptide vaccines in both monomer and trimer formats produced significant immune responses in mice on day 21, as assessed by lymph node cell counts, next generation sequencing (NGS), and plasma titers against T7 phage and vaccine peptides. A trimer provided no additional serum response to the monomer format. Immunization of mice with a mixture of 8 different peptide vaccines resulted in antibodies to most of the peptides. It was encouraging that induced antibodies had higher binding to the mutated peptides compared to the corresponding normal peptides. The NGS of lymph node cells demonstrated a low B cell receptor diversity and clonal hyperpolarization in vaccine-draining lymph nodes in comparison to those in unvaccinated mice nodes. The NGS data also revealed phenomenal increase in IgG and other class-switched antibodies following vaccination. These results agree with the higher plasma titers of IgG antibodies against T7 phage and vaccine peptides. Antibodies bound whole B16-F10 cells, lysates and multiple bands on Western blot. This indicates that these vaccine peptides successfully induced antibodies that bind full proteins from which the vaccine peptides were derived. We demonstrate a preclinical platform for rapid production of vaccines that can deliver mutated peptides and stimulate an appropriate B cell response. We anticipate further research in utilizing the cells from a tumor or vaccine draining lymph node as a resource for therapeutic anticancer reagents.

Lambda phage-based vaccine induces antitumor immunity in hepatocellular carcinoma

Background and aimsHepatocellular carcinoma (HCC) is a difficult to treat tumor with a poor prognosis. Aspartate β-hydroxylase (ASPH) is a highly conserved enzyme overexpressed on the cell surface of both murine and human HCC cells.MethodsWe evaluated therapeutic effects of nanoparticle lambda (λ) phage vaccine constructs against ASPH expressing murine liver tumors. Mice were immunized before and after subcutaneous implantation of a syngeneic BNL HCC cell line. Antitumor actively was assessed by generation of antigen specific cellular immune responses and the identification of tumor infiltrating lymphocytes.ResultsProphylactic and therapeutic immunization significantly delayed HCC growth and progression. ASPH-antigen specific CD4+ and CD8+ lymphocytes were identified in the spleen of tumor bearing mice and cytotoxicity was directed against ASPH expressing BNL HCC cells. Furthermore, vaccination generated antigen specific Th1 and Th2 cytokine secretion by immune cells. There was widespread necrosis with infiltration of CD3+ and CD8+ T cells in HCC tumors of λ phage vaccinated mice compared to controls. Moreover, further confirmation of anti-tumor effects on ASPH expressing tumor cell growth were obtained in another murine syngeneic vaccine model with pulmonary metastases.ConclusionsThese observations suggest that ASPH may serve as a highly antigenic target for immunotherapy.

Cancer immunotherapy by a recombinant phage vaccine displaying EGFR mimotope: an in vivo study

To date, several small molecule inhibitors and monoclonal-antibodies (like ICR-62) have been used to treat tumors over-expressing epidermal growth factor receptor (EGFR). However, the limitations associated with these conventional applications accentuate the necessity of alternative approaches. Mimotopes as compelling molecular tools could rationally be employed to circumvent these drawbacks. In the present study, an M13 phage displaying ICR-62 binding peptide mimotope is exploited as a vaccine candidate. It exhibited high affinity towards ICR62 and polyclonal anti-P-BSA antibodies. Following the mice immunization, phage-based mimotope vaccine induced humoral immunity. Elicited anti-EGFR mimotope antibodies were detected using ELISA method. Moreover, the phage vaccine was tested on the Lewis lung carcinoma mice model to investigate the prophylactic and therapeutic effects. The tumor volume was measured and recorded in different animal groups to evaluate the anti-tumor effects of the vaccine. Our data indicate that the reported phage-based mimotope could potentially elicit specific antibodies resulting in low titers of EGFR-specific antibodies and reduced tumor growth. However, in vivo experiments of prophylactic or therapeutic vaccination showed no specific advantage. Furthermore, phage-mimotope vaccine might be a promising approach in the field of cancer immunotherapy.

Triple Tandem Mimotope Peptide of Epidermal Growth Factor ReceptorDisplaying on the Surface of M13 Phage Induces Anti-Tumor Response in MiceTumor Model

Introduction: Epidermal growth factor receptor (EGFR) has been shown to play a critical role in tumor cell growth and its overexpression has been observed in many epithelial tumors. In the field of cancer vaccine research, displaying the peptide mimotope on the surface of phage particles has shown promising results. Methods: In this study using m13-PVIII phage display system, two constructs were prepared: triple tandem repeat of EGFR mimotpe displaying particles (3M) and single EGFR mimotope displaying phage particle (1M). To investigate the anti-tumor properties of phage vaccine, C57BL/6 mice Lewis lung carcinoma xenograft model was established and treated with 3M phage vaccine, 1M phage vaccine and control agents. Results: Immunization of mice with these phage-based vaccines showed strong immune response against phage-mimotope. 3M phage vaccine showed more potency against tumor in comparison with control groups. Also the survival time was extended in phage vaccine treated tumor-bearing mice compared with untreated mice. Conclusion: Our findings suggest that mimotope-displaying phage vaccine can induce specific antibodies with antitumoral activity, which its potential as a candidate vaccine for EGFR-specific cancer immunotherapy needs to be more investigated in future studies.