Programming peptide-oligonucleotide nano-assembly for engineering of neoantigen vaccine with potent immunogenicity.

Background: Neoantigen nanovaccine has been recognized as a promising treatment modality for personalized cancer immunotherapy. However, most current nanovaccines are carrier-dependent and the manufacturing process is complicated, resulting in potential safety concerns and suboptimal codelivery of neoantigens and adjuvants to antigen-presenting cells (APCs). Methods: Here we report a facile and general methodology for nanoassembly of peptide and oligonucleotide by programming neoantigen peptide with a short cationic module at N-terminus to prepare nanovaccine. The programmed peptide can co-assemble with CpG oligonucleotide (TLR9 agonist) into monodispersed nanostructures without the introduction of artificial carrier. Results: We demonstrate that the engineered nanovaccine promoted the codelivery of neoantigen peptides and adjuvants to lymph node-residing APCs and instigated potent neoantigen-specific T-cell responses, eliciting neoantigen-specific antitumor immune responses with negligible systemic toxicity. Furthermore, the antitumor T-cell immunity is profoundly potentiated when combined with anti-PD-1 therapy, leading to significant inhibition or even complete regression of established melanoma and MC-38 colon tumors. Conclusions: Collectively, this work demonstrates the feasibility and effectiveness of personalized cancer nanovaccine preparation with high immunogenicity and good biosafety by programming neoantigen peptide for nanoassembly with oligonucleotides without the aid of artificial carrier.

Immunotherapy in leukaemia.

Leukaemia is the common name for a group of malignant diseases of the haematopoietic system with complex classifications and characteristics. Remarkable progress has been made in basic research and preclinical studies for acute leukaemia compared to that of the many other types/subtypes of leukaemia, especially the exploration of the biological basis and application of immunotherapy in acute myeloid leukaemia (AML) and B-cell acute lymphoblastic leukaemia (B-ALL). In this review, we summarize the basic approaches to immunotherapy for leukaemia and focus on the research progress made in immunotherapy development for AML and ALL. Importantly, despite the advances made to date, big challenges still exist in the effectiveness of leukaemia immunotherapy, especially in AML. Therefore, we use AML as an example and summarize the mechanisms of tumour cell immune evasion, describe recently reported data and known therapeutic targets, and discuss the obstacles in finding suitable treatment targets and the results obtained in recent clinical trials for several types of single and combination immunotherapies, such as bispecific antibodies, cell therapies (CAR-T-cell treatment), and checkpoint blockade. Finally, we summarize novel immunotherapy strategies for treating lymphocytic leukaemia and clinical trial results.

Safety and tumour-specific immunological responses of combined dendritic cell vaccination and anti-CD40 agonistic antibody treatment for patients with metastatic pancreatic cancer: protocol for a phase I, open-label, single-arm, dose-escalation study (REACtiVe-2 trial).

INTRODUCTION: The prognosis of patients with advanced pancreatic ductal adenocarcinoma (PDAC) is dismal and conventional chemotherapy treatment delivers limited survival improvement. Immunotherapy may complement our current treatment strategies. We previously demonstrated that the combination of an allogeneic tumour-lysate dendritic cell (DC) vaccine with an anti-CD40 agonistic antibody resulted in robust antitumour responses with survival benefit in a murine PDAC model. In the Rotterdam PancrEAtic Cancer Vaccination-2 trial, we aim to translate our findings into patients. This study will determine the safety of DC/anti-CD40 agonistic antibody combination treatment, and treatment-induced tumour-specific immunological responses. METHODS AND ANALYSIS: In this open-label, single-centre (Erasmus Univsersity Medical Center, Rotterdam, Netherlands), single-arm, phase I dose finding study, adult patients with metastatic pancreatic cancer with progressive disease after FOLFIRINOX chemotherapy will receive monocyte-derived DCs loaded with an allogeneic tumour lysate in conjunction with a CD40 agonistic antibody. This combination-immunotherapy regimen will be administered three times every 2 weeks, and booster treatments will be given after 3 and 6 months following the third injection. A minimum of 12 and a maximum of 18 patients will be included. The primary endpoint is safety and tolerability of the combination immunotherapy. To determine the maximum tolerated dose, DCs will be given at a fixed dosage and anti-CD40 agonist in a traditional 3+3 dose-escalation design. Secondary endpoints include radiographic response according to the RECIST (V.1.1) and iRECIST criteria, and the detection of antitumour specific immune responses. ETHICS AND DISSEMINATION: The Central Committee on Research Involving Human Subjects (CCMO; NL76592.000.21) and the Medical Ethics Committee (METC; MEC-2021-0566) of the Erasmus M.C. University Medical Center Rotterdam approved the conduct of the trial. Written informed consent will be required for all participants. The results of the trial will be submitted for publication in a peer-reviewed scientific journal. TRIAL REGISTRATION NUMBER: NL9723.

In vitro evidence of oncofetal antigen and TLR-9 agonist co-delivery by alginate nanovaccines for liver cancer immunotherapy.

Liver cancer is the most common malignant tumor and liver cancer immunotherapy has been one of the research hotspots. To induce antigen-specific antitumor immune responses against liver cancer, we developed antigen and adjuvant co-delivery nanovaccines (APPCs). Polyanionic alginate (ALG) and polycationic polyethyleneimine (PEI) were utilized to co-deliver a glypican-3 peptide antigen and an unmethylated cytosine-phosphate-guanine (CpG) adjuvant by electrostatic interactions. A cellular uptake study confirmed that APPC could promote antigen and adjuvant uptake by dendritic cells (DCs). Importantly, APPC facilitated the endosomal escape of the peptide for antigen delivery into the cytoplasm. In addition, APPC showed significant stimulation of DC maturation in vitro. APPC could also efficiently prime DCs and induce cytotoxic T lymphocyte responses in vivo. The in vitro cell viability assay and the in vivo histocompatibility showed that APPC was non-toxic within the tested concentration. This study demonstrates that the peptide antigen and the CpG adjuvant co-delivery nanovaccine have potential applications in liver cancer immunotherapy.

Efficient targeting of NY-ESO-1 tumor antigen to human cDC1s by lymphotactin results in cross-presentation and antigen-specific T cell expansion.

BACKGROUND: Type 1 conventional dendritic cells (cDC1s) are characterized by their ability to induce potent CD8+ T cell responses. In efforts to generate novel vaccination strategies, notably against cancer, human cDC1s emerge as an ideal target to deliver antigens. cDC1s uniquely express XCR1, a seven transmembrane G protein-coupled receptor. Due to its restricted expression and endocytic nature, XCR1 represents an attractive receptor to mediate antigen-delivery to human cDC1s. METHODS: To explore tumor antigen delivery to human cDC1s, we used an engineered version of XCR1-binding lymphotactin (XCL1), XCL1(CC3). Site-specific sortase-mediated transpeptidation was performed to conjugate XCL1(CC3) to an analog of the HLA-A*02:01 epitope of the cancer testis antigen New York Esophageal Squamous Cell Carcinoma-1 (NY-ESO-1). While poor epitope solubility prevented isolation of stable XCL1-antigen conjugates, incorporation of a single polyethylene glycol (PEG) chain upstream of the epitope-containing peptide enabled generation of soluble XCL1(CC3)-antigen fusion constructs. Binding and chemotactic characteristics of the XCL1-antigen conjugate, as well as its ability to induce antigen-specific CD8+ T cell activation by cDC1s, was assessed. RESULTS: PEGylated XCL1(CC3)-antigen conjugates retained binding to XCR1, and induced cDC1 chemoattraction in vitro. The model epitope was efficiently cross-presented by human cDC1s to activate NY-ESO-1-specific CD8+ T cells. Importantly, vaccine activity was increased by targeting XCR1 at the surface of cDC1s. CONCLUSION: Our results present a novel strategy for the generation of targeted vaccines fused to insoluble antigens. Moreover, our data emphasize the potential of targeting XCR1 at the surface of primary human cDC1s to induce potent CD8+ T cell responses.

In Situ Tumor Vaccination with Calcium-Linked Degradable Coacervate Nanocomplex Co-Delivering Photosensitizer and TLR7/8 Agonist to Trigger Effective Anti-Tumor Immune Responses.

In situ anti-tumor vaccination is an attractive type of cancer immunotherapy which relies on the effectiveness of dendritic cells (DCs) to engulf tumor antigens, become activated, and present antigens to T cells in lymphoid tissue. Here, a multifunctional nanocomplex based on calcium crosslinked polyaspartic acid conjugated to either a toll-like receptor (TLR)7/8 agonist or a photosensitizer is reported. Intratumoral administration of the nanocomplex followed by laser irradiation induces cell killing and hence generation of a pool of tumor-associated antigens, with concomitant promotion of DCs maturation and expansion of T cells in tumor-draining lymph nodes. Suppression of tumor growth is observed both at the primary site and at the distal site, thereby hinting at successful induction of an adaptive anti-tumor response. This strategy holds promise for therapeutic application in a pre-operative and post-operative setting to leverage to mutanome of the patient's own tumor to mount immunological memory to clear residual tumor cells and metastasis.

In situ vaccination using unique TLR9 ligand K3-SPG induces long-lasting systemic immune response and synergizes with systemic and local immunotherapy.

Although checkpoint inhibitors (CPIs) have changed the paradigm of cancer therapy, low response rates and serious systemic adverse events remain challenging. In situ vaccine (ISV), intratumoral injection of immunomodulators that stimulate innate immunity at the tumor site, allows for the development of vaccines in patients themselves. K3-SPG, a second-generation nanoparticulate Toll-like receptor 9 (TLR9) ligand consisting of K-type CpG oligodeoxynucleotide (ODN) wrapped with SPG (schizophyllan), integrates the best of conventional CpG ODNs, making it an ideal cancer immunotherapy adjuvant. Focusing on clinical feasibility for pancreaticobiliary and gastrointestinal cancers, we investigated the antitumor activity of K3-SPG-ISV in preclinical models of pancreatic ductal adenocarcinoma (PDAC) and colorectal cancer (CRC). K3-SPG-ISV suppressed tumor growth more potently than K3-ISV or K3-SPG intravenous injections, prolonged survival, and enhanced the antitumor effect of CPIs. Notably, in PDAC model, K3-SPG-ISV alone induced systemic antitumor effect and immunological memory. ISV combination of K3-SPG and agonistic CD40 antibody further enhanced the antitumor effect. Our results imply that K3-SPG-based ISV can be applied as monotherapy or combined with CPIs to improve their response rate or, conversely, with CPI-free local immunotherapy to avoid CPI-related adverse events. In either strategy, the potency of K3-SPG-based ISV would provide the rationale for its clinical application to puncturable pancreaticobiliary and gastrointestinal malignancies.

Combination Cancer Immunotherapy with Dendritic Cell Vaccine and Nanoparticles Loaded with Interleukin-15 and Anti-beta-catenin siRNA Significantly Inhibits Cancer Growth and Induces Anti-Tumor Immune Response.

PURPOSE: The invention and application of new immunotherapeutic methods can compensate for the inefficiency of conventional cancer treatment approaches, partly due to the inhibitory microenvironment of the tumor. In this study, we tried to inhibit the growth of cancer cells and induce anti-tumor immune responses by silencing the expression of the ß-catenin in the tumor microenvironment and transmitting interleukin (IL)-15 cytokine to provide optimal conditions for the dendritic cell (DC) vaccine. METHODS: For this purpose, we used folic acid (FA)-conjugated SPION-carboxymethyl dextran (CMD) chitosan (C) nanoparticles (NPs) to deliver anti-ß-catenin siRNA and IL-15 to cancer cells. RESULTS: The results showed that the codelivery of ß-catenin siRNA and IL-15 significantly reduced the growth of cancer cells and increased the immune response. The treatment also considerably stimulated the performance of the DC vaccine in triggering anti-tumor immunity, which inhibited tumor development and increased survival in mice in two different cancer models. CONCLUSIONS: These findings suggest that the use of new nanocarriers such as SPION-C-CMD-FA could be an effective way to use as a novel combination therapy consisting of ß-catenin siRNA, IL-15, and DC vaccine to treat cancer.

Peptides for Vaccine Development.

This review discusses peptide epitopes used as antigens in the development of vaccines in clinical trials as well as future vaccine candidates. It covers peptides used in potential immunotherapies for infectious diseases including SARS-CoV-2, influenza, hepatitis B and C, HIV, malaria, and others. In addition, peptides for cancer vaccines that target examples of overexpressed proteins are summarized, including human epidermal growth factor receptor 2 (HER-2), mucin 1 (MUC1), folate receptor, and others. The uses of peptides to target cancers caused by infective agents, for example, cervical cancer caused by human papilloma virus (HPV), are also discussed. This review also provides an overview of model peptide epitopes used to stimulate non-specific immune responses, and of self-adjuvanting peptides, as well as the influence of other adjuvants on peptide formulations. As highlighted in this review, several peptide immunotherapies are in advanced clinical trials as vaccines, and there is great potential for future therapies due the specificity of the response that can be achieved using peptide epitopes.

Nanovaccines with cell-derived components for cancer immunotherapy.

Cancer nanovaccines as one of immunotherapeutic approaches are able to attack tumors by stimulating tumor-specific immunological responses. However, there still exist multiple challenges to be tackled for cancer nanovaccines to evoke potent antitumor immunity. Particularly, the administration of exogenous materials may cause the off-target immunotherapy responses. In recent years, biomimetic nanovaccines by using cell lysates, cell-derived nanovesicles, or extracted cell membranes as the functional components have received extensive attention. Such nanovaccines based on cell-derived components would show many unique advantages including inherent biocompatibility and the ability to trigger immune responses against a range of tumor-associated antigens. In this review article, we will introduce the recent research progresses of those cell-derived biomimetic nanovaccines for cancer immunotherapy, and discuss the perspectives and challenges associated with the future clinical translation of these emerging vaccine platforms.

Harnessing self-assembling peptide nanofibers toprime robust tumor-specific CD8 T cell responses in mice.

Induction of tumor-specific CD8 + T cell responses is known as a major challenge for cancer vaccine development; here we presented a strategy to improve peptide nanofibers-mounted antitumor immune responses. To this end, peptide nanofibers bearing class I (Kb)-restricted epitope (Epi-Nano) were formulated with polyethylene imine backbone (Epi-Nano-PEI), and characterized using morphological and physicochemicalcharacterizationtechniques. Nanofibers were studied in terms of their uptake by antigen-presenting cells (APCs), antigen cross-presentation capacity, and cytotoxic activity. Furthermore, nanofibers were assessed by their potency to induce NLRP3 inflammasome-related cytokines and factors. Finally, the ability of nanofibers to induce tumor-specific CD8 T cells and tumor protection were investigated in tumor-bearing mice. The formulation of Epi-Nano with PEI led to the formation of short strand nanofibers with a positive surface charge, a low critical aggregation concentration (CAC), and an increased resistancetoproteolytic degradation. Epi-Nano-PEI was significantly taken up more efficiently by antigen-presenting cells (APCs), and was more potent in cross-presentation when compared to Epi-Nano. Moreover, Epi-Nano-PEI, in comparison to Epi-Nano, efficiently up-regulated the expression of NLRP3, caspase-1, IL-1b, IL18 and IL-6. Cell viability analysis showed that formulation of PEI with Epi-Nano not only abolished its cytotoxic activity, but surprisingly induced macrophage proliferation. Furthermore, it demonstrated that Epi-Nano-PEI triggered robust antigen-specific CD8+ T cell responses, and induced maximum antitumor response (tumor growth inhibition and prolonged survival) in tumor-bearing mice that were significantly higher compared to Epi-Nano. Taken together, the formulation of Epi-Nano with PEI is suggested as a promising strategy to improve nanofibers-mounted antitumor immune response.

Cancer immunotherapy using artificial adjuvant vector cells to deliver NY-ESO-1 antigen to dendritic cells in situ.

NY-ESO-1 is a cancer/testis antigen expressed in various cancer types. However, the induction of NY-ESO-1-specific CTLs through vaccines is somewhat difficult. Thus, we developed a new type of artificial adjuvant vector cell (aAVC-NY-ESO-1) expressing a CD1d-NKT cell ligand complex and a tumor-associated antigen, NY-ESO-1. First, we determined the activation of invariant natural killer T (iNKT) and natural killer (NK) cell responses by aAVC-NY-ESO-1. We then showed that the NY-ESO-1-specific CTL response was successfully elicited through aAVC-NY-ESO-1 therapy. After injection of aAVC-NY-ESO-1, we found that dendritic cells (DCs) in situ expressed high levels of costimulatory molecules and produced interleukn-12 (IL-12), indicating that DCs undergo maturation in vivo. Furthermore, the NY-ESO-1 antigen from aAVC-NY-ESO-1 was delivered to the DCs in vivo, and it was presented on MHC class I molecules. The cross-presentation of the NY-ESO-1 antigen was absent in conventional DC-deficient mice, suggesting a host DC-mediated CTL response. Thus, this strategy helps generate sufficient CD8+ NY-ESO-1-specific CTLs along with iNKT and NK cell activation, resulting in a strong antitumor effect. Furthermore, we established a human DC-transferred NOD/Shi-scid/IL-2γcnull immunodeficient mouse model and showed that the NY-ESO-1 antigen from aAVC-NY-ESO-1 was cross-presented to antigen-specific CTLs through human DCs. Taken together, these data suggest that aAVC-NY-ESO-1 has potential for harnessing innate and adaptive immunity against NY-ESO-1-expressing malignancies.

Randomized trial of neoadjuvant vaccination with tumor-cell lysate induces T cell response in low-grade gliomas.

BACKGROUNDLong-term prognosis of WHO grade II low-grade gliomas (LGGs) is poor, with a high risk of recurrence and malignant transformation into high-grade gliomas. Given the relatively intact immune system of patients with LGGs and the slow tumor growth rate, vaccines are an attractive treatment strategy.METHODSWe conducted a pilot study to evaluate the safety and immunological effects of vaccination with GBM6-AD, lysate of an allogeneic glioblastoma stem cell line, with poly-ICLC in patients with LGGs. Patients were randomized to receive the vaccines before surgery (arm 1) or not (arm 2) and all patients received adjuvant vaccines. Coprimary outcomes were to evaluate safety and immune response in the tumor.RESULTSA total of 17 eligible patients were enrolled - 9 in arm 1 and 8 in arm 2. This regimen was well tolerated with no regimen-limiting toxicity. Neoadjuvant vaccination induced upregulation of type-1 cytokines and chemokines and increased activated CD8+ T cells in peripheral blood. Single-cell RNA/T cell receptor sequencing detected CD8+ T cell clones that expanded with effector phenotype and migrated into the tumor microenvironment (TME) in response to neoadjuvant vaccination. Mass cytometric analyses detected increased tissue resident-like CD8+ T cells with effector memory phenotype in the TME after the neoadjuvant vaccination.CONCLUSIONThe regimen induced effector CD8+ T cell response in peripheral blood and enabled vaccine-reactive CD8+ T cells to migrate into the TME. Further refinements of the regimen may have to be integrated into future strategies.TRIAL REGISTRATIONClinicalTrials.gov NCT02549833.FUNDINGNIH (1R35NS105068, 1R21CA233856), Dabbiere Foundation, Parker Institute for Cancer Immunotherapy, and Daiichi Sankyo Foundation of Life Science.

Tumor associated macrophage and microbe: The potential targets of tumor vaccine delivery.

The occurrence and development of tumors depend on the tumor microenvironment (TME), which is made of various immune cells, activated fibroblasts, basement membrane, capillaries, and extracellular matrix. Tumor associated macrophages (TAMs) and microbes are important components in TME. Tumor cells can recruit and educate TAMs and microbes, and the hijacked TAMs and microbes can promote the progression of tumor reciprocally. Tumor vaccine delivery remodeling TME by targeting TAM and microbes can not only enhance the specificity and immunogenicity of antigens, but also contribute to the regulation of TME. Tumor vaccine design benefits from nanotechnology which is a suitable platform for antigen and adjuvant delivery to catalyze new candidate vaccines applying to clinical therapy at unparalleled speed. In view of the characteristics and mechanisms of TME development, vaccine delivery targeting and breaking the malignant interactions among tumor cells, TAMs, and microbes may serve as a novel strategy for tumor therapy.

Long-term first-in-man Phase I/II study of an adjuvant dendritic cell vaccine in patients with high-risk prostate cancer after radical prostatectomy.

BACKGROUND: Patients with high-risk prostate cancer (PC) can experience biochemical relapse (BCR), despite surgery, and develop noncurative disease. The present study aimed to reduce the risk of BCR with a personalized dendritic cell (DC) vaccine, given as adjuvant therapy, after robot-assisted laparoscopic prostatectomy (RALP). METHODS: Twelve weeks after RALP, 20 patients with high-risk PC and undetectable PSA received DC vaccinations for 3 years or until BCR. The primary endpoint was the time to BCR. The immune response was assessed 7 weeks after surgery (baseline) and at one-time point during the vaccination period. RESULTS: Among 20 patients, 11 were BCR-free over a median of 96 months (range: 84-99). The median time from the end of vaccinations to the last follow-up was 57 months (range: 45-60). Nine patients developed BCR, either during (n = 4) or after (n = 5) the vaccination period. Among five patients diagnosed with intraductal carcinoma, three experienced early BCR during the vaccination period. All patients that developed BCR remained in stable disease within a median of 99 months (range: 74-99). The baseline immune response was significantly associated with the immune response during the vaccination period (p = 0.015). For patients diagnosed with extraprostatic extension (EPE), time to BCR was longer in vaccine responders than in non-responders (p = 0.09). Among 12 patients with the International Society of Urological Pathology (ISUP) grade 5 PC, five achieved remission after 84 months, and all mounted immune responses. CONCLUSION: Patients diagnosed with EPE and ISUP grade 5 PC were at particularly high risk of developing postsurgical BCR. In this subgroup, the vaccine response was related to a reduced BCR incidence. The vaccine was safe, without side effects. This adjuvant first-in-man Phase I/II DC vaccine study showed promising results. DC vaccines after curative surgery should be investigated further in a larger cohort of patients with high-risk PC.

A randomized phase 2 trial of idiotype vaccination and adoptive autologous T-cell transfer in patients with multiple myeloma.

We hypothesized that combining adoptively transferred autologous T cells with a cancer vaccine strategy would enhance therapeutic efficacy by adding antimyeloma idiotype (Id)-keyhole limpet hemocyanin (KLH) vaccine to vaccine-specific costimulated T cells. In this randomized phase 2 trial, patients received either control (KLH only) or Id-KLH vaccine, autologous transplantation, vaccine-specific costimulated T cells expanded ex vivo, and 2 booster doses of assigned vaccine. In 36 patients (KLH, n = 20; Id-KLH, n = 16), no dose-limiting toxicity was seen. At last evaluation, 6 (30%) and 8 patients (50%) had achieved complete remission in KLH-only and Id-KLH arms, respectively (P = .22), and no difference in 3-year progression-free survival was observed (59% and 56%, respectively; P = .32). In a 594 Nanostring nCounter gene panel analyzed for immune reconstitution (IR), compared with patients receiving KLH only, there was a greater change in IR genes in T cells in those receiving Id-KLH relative to baseline. Specifically, upregulation of genes associated with activation, effector function induction, and memory CD8+ T-cell generation after Id-KLH but not after KLH control vaccination was observed. Similarly, in responding patients across both arms, upregulation of genes associated with T-cell activation was seen. At baseline, all patients had greater expression of CD8+ T-cell exhaustion markers. These changes were associated with functional Id-specific immune responses in a subset of patients receiving Id-KLH. In conclusion, in this combination immunotherapy approach, we observed significantly more robust IR in CD4+ and CD8+ T cells in the Id-KLH arm, supporting further investigation of vaccine and adoptive immunotherapy strategies. This trial was registered at www.clinicaltrials.gov as #NCT01426828.

A phase 1/2 trial of an immune-modulatory vaccine against IDO/PD-L1 in combination with nivolumab in metastatic melanoma.

Anti-programmed death (PD)-1 (aPD1) therapy is an effective treatment for metastatic melanoma (MM); however, over 50% of patients progress due to resistance. We tested a first-in-class immune-modulatory vaccine (IO102/IO103) against indoleamine 2,3-dioxygenase (IDO) and PD ligand 1 (PD-L1), targeting immunosuppressive cells and tumor cells expressing IDO and/or PD-L1 (IDO/PD-L1), combined with nivolumab. Thirty aPD1 therapy-naive patients with MM were treated in a phase 1/2 study ( https://clinicaltrials.gov/ , NCT03047928). The primary endpoint was feasibility and safety; the systemic toxicity profile was comparable to that of nivolumab monotherapy. Secondary endpoints were efficacy and immunogenicity; an objective response rate (ORR) of 80% (confidence interval (CI), 62.7-90.5%) was reached, with 43% (CI, 27.4-60.8%) complete responses. After a median follow-up of 22.9 months, the median progression-free survival (PFS) was 26 months (CI, 15.4-69 months). Median overall survival (OS) was not reached. Vaccine-specific responses assessed in vitro were detected in the blood of >93% of patients during vaccination. Vaccine-reactive T cells comprised CD4+ and CD8+ T cells with activity against IDO- and PD-L1-expressing cancer and immune cells. T cell influx of peripherally expanded T cells into tumor sites was observed in responding patients, and general enrichment of IDO- and PD-L1-specific clones after treatment was documented. These clinical efficacy and favorable safety data support further validation in a larger randomized trial to confirm the clinical potential of this immunomodulating approach.

Immunization with short peptide particles reveals a functional CD8+ T-cell neoepitope in a murine renal carcinoma model.

BACKGROUND: Induction of CD8+ T cells that recognize immunogenic, mutated protein fragments in the context of major histocompatibility class I (MHC-I) is a pressing challenge for cancer vaccine development. METHODS: Using the commonly used murine renal adenocarcinoma RENCA cancer model, MHC-I restricted neoepitopes are predicted following next-generation sequencing. Candidate neoepitopes are screened in mice using a potent cancer vaccine adjuvant system that converts short peptides into immunogenic nanoparticles. An identified functional neoepitope vaccine is then tested in various therapeutic experimental tumor settings. RESULTS: Conversion of 20 short MHC-I restricted neoepitope candidates into immunogenic nanoparticles results in antitumor responses with multivalent vaccination. Only a single neoepitope candidate, Nesprin-2 L4492R (Nes2LR), induced functional responses but still did so when included within 20-plex or 60-plex particles. Immunization with the short Nes2LR neoepitope with the immunogenic particle-inducing vaccine adjuvant prevented tumor growth at doses multiple orders of magnitude less than with other vaccine adjuvants, which were ineffective. Nes2LR vaccination inhibited or eradicated disease in subcutaneous, experimental lung metastasis and orthotopic tumor models, synergizing with immune checkpoint blockade. CONCLUSION: These findings establish the feasibility of using short, MHC-I-restricted neoepitopes for straightforward immunization with multivalent or validated neoepitopes to induce cytotoxic CD8+ T cells. Furthermore, the Nes2LR neoepitope could be useful for preclinical studies involving renal cell carcinoma immunotherapy.

MHC-Optimized Peptide Scaffold for Improved Antigen Presentation and Anti-Tumor Response.

Tumor Associated Antigens (TAAs) may suffer from an immunological tolerance due to expression on normal cells. In order to potentiate their immunogenicity, heteroclitic peptides (htcPep) were designed according to prediction algorithms. In particular, specific modifications were introduced in peptide residues facing to TCR. Moreover, a MHC-optimized scaffold was designed for improved antigen presentation to TCR by H-2Db allele. The efficacy of such htcPep was assessed in C57BL/6 mice injected with syngeneic melanoma B16F10 or lung TC1 tumor cell lines, in combination with metronomic chemotherapy and immune checkpoint inhibitors. The immunogenicity of htcPep was significantly stronger than the corresponding wt peptide and the modification involving both MHC and TCR binding residues scored the strongest. In particular, the H-2Db-specific scaffold significantly potentiated the peptides' immunogenicity and control of tumor growth was comparable to wt peptide in a therapeutic setting. Overall, we demonstrated that modified TAAs show higher immunogenicity compared to wt peptide. In particular, the MHC-optimized scaffold can present different antigen sequences to TCR, retaining the conformational characteristics of the corresponding wt. Cross-reacting CD8+ T cells are elicited and efficiently kill tumor cells presenting the wild-type antigen. This novel approach can be of high clinical relevance in cancer vaccine development.

Idiotype vaccines produced with a non-cytopathic alphavirus self-amplifying RNA vector induce antitumor responses in a murine model of B-cell lymphoma.

A promising therapy for patients with B-cell lymphoma is based on vaccination with idiotype monoclonal antibodies (mAbs). Since idiotypes are different in each tumor, a personalized vaccine has to be produced for each patient. Expression of immunoglobulins with appropriate post-translational modifications for human use often requires the use of stable mammalian cells that can be scaled-up to reach the desired level of production. We have used a noncytopathic self-amplifying RNA vector derived from Semliki Forest virus (ncSFV) to generate BHK cell lines expressing murine follicular lymphoma-derived idiotype A20 mAb. ncSFV/BHK cell lines expressed approximately 2 mg/L/24 h of A20 mAb with proper quaternary structure and a glycosylation pattern similar to that of A20 mAb produced by hybridoma cells. A20 mAb purified from the supernatant of a ncSFV cell line, or from the hybridoma, was conjugated to keyhole limpet hemocyanin and used to immunize Balb/c mice by administration of four weekly doses of 25 µg of mAb. Both idiotype mAbs were able to induce a similar antitumor protection and longer survival compared to non-immunized mice. These results indicate that the ncSFV RNA vector could represent a quick and efficient system to produce patient-specific idiotypes with potential application as lymphoma vaccines.