Antigen-specific Immune Responses Research Articles

TRPV1 nociceptors are required to optimize antigen-specific primary antibody responses to novel antigens

BackgroundKey to the advancement of the field of bioelectronic medicine is the identification of novel pathways of neural regulation of immune function. Sensory neurons (termed nociceptors) recognize harmful stimuli and initiate a protective response by eliciting pain and defensive behavior. Nociceptors also interact with immune cells to regulate host defense and inflammatory responses. However, it is still unclear whether nociceptors participate in regulating primary IgG antibody responses to novel antigens.MethodsTo understand the role of transient receptor potential vanilloid 1 (TRPV1)-expressing neurons in IgG responses, we generated TRPV1-Cre/Rosa-ChannelRhodopsin2 mice for precise optogenetic activation of TRPV1 + neurons and TRPV1-Cre/Lox-diphtheria toxin A mice for targeted ablation of TRPV1-expressing neurons. Antigen-specific antibody responses were longitudinally monitored for 28 days.ResultsHere we show that TRPV1 expressing neurons are required to develop an antigen-specific immune response. We demonstrate that selective optogenetic stimulation of TRPV1+ nociceptors during immunization significantly enhances primary IgG antibody responses to novel antigens. Further, mice rendered deficient in TRPV1- expressing nociceptors fail to develop primary IgG antibody responses to keyhole limpet hemocyanin or haptenated antigen.ConclusionThis functional and genetic evidence indicates a critical role for nociceptor TRPV1 in antigen-specific primary antibody responses to novel antigens. These results also support consideration of potential therapeutic manipulation of nociceptor pathways using bioelectronic devices to enhance immune responses to foreign antigens.

Breaking the Tumor Chronic Inflammation Balance with a Programmable Release and Multi-Stimulation Engineering Scaffold for Potent Immunotherapy.

Tumor-associated chronic inflammation severely restricts the efficacy of immunotherapy in cold tumors. Here, a programmable release hydrogel-based engineering scaffold with multi-stimulation and reactive oxygen species (ROS)-response (PHOENIX) is demonstrated to break the chronic inflammatory balance in cold tumors to induce potent immunity. PHOENIX can undergo programmable release of resiquimod and anti-OX40 under ROS. Resiquimod is first released, leading to antigen-presenting cell maturation and the transformation of myeloid-derived suppressor cells and M2 macrophages into an antitumor immune phenotype. Subsequently, anti-OX40 is transported into the tumor microenvironment, leading to effector T-cell activation and inhibition of Treg function. PHOENIX consequently breaks the chronic inflammation in the tumor microenvironment and leads to a potent immune response. In mice bearing subcutaneous triple-negative breast cancer and metastasis models, PHOENIX effectively inhibited 80% and 60% of tumor growth, respectively. Moreover, PHOENIX protected 100% of the mice against TNBC tumor rechallenge by electing a robust long-term antigen-specific immune response. An excellent inhibition and prolonged survival in PHOENIX-treated mice with colorectal cancer and melanoma is also observed. This work presents a potent therapeutic scaffold to improve immunotherapy efficiency, representing a generalizable and facile regimen for cold tumors.

Single-cell analysis of bronchoalveolar cells in inflammatory and fibrotic post-COVID lung disease

BackgroundPersistent radiological lung abnormalities are evident in many survivors of acute coronavirus disease 2019 (COVID-19). Consolidation and ground glass opacities are interpreted to indicate subacute inflammation whereas reticulation is thought to reflect fibrosis. We sought to identify differences at molecular and cellular level, in the local immunopathology of post-COVID inflammation and fibrosis.MethodsWe compared single-cell transcriptomic profiles and T cell receptor (TCR) repertoires of bronchoalveolar cells obtained from convalescent individuals with each radiological pattern, targeting lung segments affected by the predominant abnormality.ResultsCD4 central memory T cells and CD8 effector memory T cells were significantly more abundant in those with inflammatory radiology. Clustering of similar TCRs from multiple donors was a striking feature of both phenotypes, consistent with tissue localised antigen-specific immune responses. There was no enrichment for known SARS-CoV-2-reactive TCRs, raising the possibility of T cell-mediated immunopathology driven by failure in immune self-tolerance.ConclusionsPost-COVID radiological inflammation and fibrosis show evidence of shared antigen-specific T cell responses, suggesting a role for therapies targeting T cells in limiting post-COVID lung damage.

Organ- and Cell-Selective Delivery of mRNA In Vivo Using Guanidinylated Serinol Charge-Altering Releasable Transporters.

Selective RNA delivery is required for the broad implementation of RNA clinical applications, including prophylactic and therapeutic vaccinations, immunotherapies for cancer, and genome editing. Current polyanion delivery relies heavily on cationic amines, while cationic guanidinium systems have received limited attention due in part to their strong polyanion association, which impedes intracellular polyanion release. Here, we disclose a general solution to this problem in which cationic guanidinium groups are used to form stable RNA complexes upon formulation but at physiological pH undergo a novel charge-neutralization process, resulting in RNA release. This new delivery system consists of guanidinylated serinol moieties incorporated into a charge-altering releasable transporter (GSer-CARTs). Significantly, systematic variations in structure and formulation resulted in GSer-CARTs that exhibit highly selective mRNA delivery to the lung (∼97%) and spleen (∼98%) without targeting ligands. Illustrative of their breadth and translational potential, GSer-CARTs deliver circRNA, providing the basis for a cancer vaccination strategy, which in a murine model resulted in antigen-specific immune responses and effective suppression of established tumors.

Improvement of the Nuclease Resistance and Immunostimulatory Activity of CpG Oligodeoxynucleotides by Conjugation to Sugar-Immobilized Gold Nanoparticles.

Adjuvants are essential substances for vaccines and immunotherapies that enhance antigen-specific immune responses. Single-stranded oligodeoxynucleotides containing an unmethylated CpG motif (CpG ODNs) are agonistic ligands for toll-like receptor 9 that initiate an innate immune response. They represent promising adjuvants for antiviral and antitumor immunotherapies; however, CpG ODNs have some limitations, such as poor nuclease resistance and low cell membrane permeability. Therefore, an effective formulation is needed to improve the nuclease resistance and immunostimulatory effects of CpG ODNs. Previously, we demonstrated the selective delivery of a small molecule toll-like receptor 7 ligand to immune cells through sugar-binding receptors using sugar-immobilized gold nanoparticles (SGNPs), which significantly enhanced the potency of the ligand. In this study, we examined SGNPs as carriers for partially phosphorothioated A-type CpG ODN (D35) and an entirely phosphorothioated B-type CpG ODN (K3) and evaluated the functionality of the sugar moiety on SGNPs immobilized with CpG ODN. SGNPs immobilized with D35 (D35-SGNPs) exhibited improved nuclease resistance and the in vitro and in vivo potency was significantly higher compared with that of unconjugated D35. Furthermore, the sugar structure on the GNPs was a significant factor in enhancing the cell internalization ability, and enhanced intracellular delivery of D35 resulted in improving the potencies of the A-type CpG ODN, D35. SGNPs immobilized with K3 (K3-SGNPs) exhibited significantly higher induction activities for both humoral and cellular immunity compared with unconjugated K3 and D35-SGNPs. On the other hand, sugar structure on K3-SGNPs did not affect the immunostimulatory effects. These results indicate that the sugar moiety on K3-SGNPs primarily functions as a hydrophilic dispersant for GNPs and the formulation of K3 to SGNPs contributes to improving the immunostimulatory activity of K3. Because our CpG ODN-SGNPs have superior induction activities for antigen-specific T-cell mediated immune responses, they may be effective adjuvants for vaccines and immunotherapies.

Dendritic-cell-targeting virus-like particles as potent mRNA vaccine carriers.

Messenger RNA vaccines lack specificity for dendritic cells (DCs)-the most effective cells at antigen presentation. Here we report the design and performance of a DC-targeting virus-like particle pseudotyped with an engineered Sindbis-virus glycoprotein that recognizes a surface protein on DCs, and packaging mRNA encoding for the Spike protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) or for the glycoproteins B and D of herpes simplex virus 1. Injection of the DC-targeting SARS-CoV-2 mRNA vaccine in the footpad of mice led to substantially higher and durable antigen-specific immunoglobulin-G titres and cellular immune responses than untargeted virus-like particles and lipid-nanoparticle formulations. The vaccines also protected the mice from infection with SARS-CoV-2 or with herpes simplex virus 1. Virus-like particles with preferential uptake by DCs may facilitate the development of potent prophylactic and therapeutic vaccines.

Development of an mRNA-based therapeutic vaccine mHTV-03E2 for high-risk HPV-related malignancies

Human Papillomavirus (HPV) 16 and 18 infections are related to many human cancers. Despite several preventative vaccines for high-risk (hr) HPVs, there is still an urgent need to develop therapeutic HPV vaccines for targeting pre-existing hrHPV infections and lesions. In this study, we developed a lipid nanoparticle (LNP)-formulated mRNA-based HPV therapeutic vaccine (mHTV)-03E2, simultaneously targeting the E2/E6/E7 of both HPV16 and HPV18. mHTV-03E2 dramatically induced antigen-specific cellular immune responses, leading to significant CD8+ T cell infiltration and cytotoxicity in TC-1 tumors derived from primary lung epithelial cells of C57BL/6 mice expressing HPV E6/E7 antigens, mediated significant tumor regression, and prolonged animal survival, in a dose-dependent manner. We further demonstrated significant T cell immunity against HPV16/18 E6/E7 antigens for up to 4 months post-vaccination in immunological and distant tumor rechallenging experiments, suggesting robust memory T cell immunity against relapse. Finally, mHTV-03E2 synergized with immune checkpoint blockade to inhibit tumor growth and extend animal survival, indicating the potential in combination therapy. We conclude that mHTV-03E2 is an excellent candidate therapeutic mRNA vaccine for treating malignancies caused by HPV16 or HPV18 infections.

Polymeric Caffeic Acid Acts as an Antigen Delivery Carrier for Mucosal Vaccine Formulation by Forming a Complex with an Antigenic Protein.

The development of mucosal vaccines, which can generate antigen-specific immune responses in both the systemic and mucosal compartments, has been recognized as an effective strategy for combating infectious diseases caused by pathogenic microbes. Our recent research has focused on creating a nasal vaccine system in mice using enzymatically polymerized caffeic acid (pCA). However, we do not yet understand the molecular mechanisms by which pCA stimulates antigen-specific mucosal immune responses. In this study, we hypothesized that pCA might activate mucosal immunity at the site of administration based on our previous findings that pCA possesses immune-activating properties. However, contrary to our initial hypothesis, the intranasal administration of pCA did not enhance the expression of various genes involved in mucosal immune responses, including the enhancement of IgA responses. Therefore, we investigated whether pCA forms a complex with antigenic proteins and enhances antigen delivery to mucosal dendritic cells located in the lamina propria beneath the mucosal epithelial layer. Data from gel filtration chromatography indicated that pCA forms a complex with the antigenic protein ovalbumin (OVA). Furthermore, we examined the promotion of OVA delivery to nasal mucosal dendritic cells (mDCs) after the intranasal administration of pCA in combination with OVA and found that OVA uptake by mDCs was increased. Therefore, the data from gel filtration chromatography and flow cytometry imply that pCA enhances antigen-specific antibody production in both mucosal and systemic compartments by serving as an antigen-delivery vehicle.

Orchestrated codelivery of peptide antigen and adjuvant to antigen-presenting cells by using an engineered chimeric peptide enhances antitumor T-cell immunity.

The intrinsic pharmacokinetic limitations of traditional peptide-based cancer vaccines hamper effective cross-presentation and codelivery of antigens and adjuvants, which are crucial for inducing robust antitumor CD8+ T-cell responses. Here, we report the development of a versatile strategy that simultaneously addresses the different pharmacokinetic challenges of soluble subunit vaccines composed of antigens and CpG to modulate vaccine efficacy via translating an engineered chimeric peptide, eTAT, as an intramolecular adjuvant. Linking antigens to eTAT enhanced cytosolic delivery of the antigens. This, in turn, led to improved activation and lymph node-trafficking of antigen-presenting cells and antigen cross-presentation, thus promoting antigen-specific T-cell immune responses. Simple mixing of eTAT-linked antigens and CpG significantly enhanced codelivery of antigens and CpG to the antigen-presenting cells, and this substantially augmented the adjuvant effect of CpG, maximized vaccine immunogenicity and elicited robust and durable CD8+ T-cell responses. Vaccination with this formulation altered the tumor microenvironment and exhibited potent antitumor effects, with generally further enhanced therapeutic efficacy when used in combination with anti-PD1. Altogether, the engineered chimeric peptide-based orchestrated codelivery of antigen and adjuvant may serve as a promising but simple strategy to improve the efficacy of peptide-based cancer vaccines.

Preclinical evaluation of two phylogenetically distant arenavirus vectors for the development of novel immunotherapeutic combination strategies for cancer treatment

BackgroundEngineered arenavirus vectors have recently been developed to leverage the body’s immune system in the fight against chronic viral infections and cancer. Vectors based on Pichinde virus (artPICV) and lymphocytic...

The adaptive immune response to Trichuris in wild versus laboratory mice: An established model system in context.

Laboratory model organisms have provided a window into how the immune system functions. An increasing body of evidence, however, suggests that the immune responses of naive laboratory animals may differ substantially to those of their wild counterparts. Past exposure, environmental challenges and physiological condition may all impact on immune responsiveness. Chronic infections of soil-transmitted helminths, which we define as establishment of adult, fecund worms, impose significant health burdens on humans, livestock and wildlife, with limited treatment success. In laboratory mice, Th1 versus Th2 immune polarisation is the major determinant of helminth infection outcome. Here we compared antigen-specific immune responses to the soil-transmitted whipworm Trichuris muris between controlled laboratory and wild free-ranging populations of house mice (Mus musculus domesticus). Wild mice harbouring chronic, low-level infections produced lower levels of cytokines in response to Trichuris antigen than laboratory-housed C57BL/6 mice. Wild mouse effector/memory CD4+ T cell phenotype reflected the antigen-specific cytokine response across the Th1/Th2 spectrum. Increasing egg shedding was associated with body condition loss. However, local Trichuris-specific Th1/Th2 balance was positively associated with worm burden only in older wild mice. Thus, although the fundamental relationships between the CD4+ T helper cell response and resistance to T. muris infection are similar in both laboratory and wild M. m. domesticus, there are quantitative differences and age-specific effects that are analogous to human immune responses. These context-dependent immune responses demonstrate the fundamental importance of understanding the differences between model and natural systems for translating mechanistic models to 'real world' immune function.

Injectable Hydrogel Mucosal Vaccine Elicits Protective Immunity against Respiratory Viruses.

Intranasal vaccines, eliciting mucosal immune responses, can prevent early invasion, replication, and transmission of pathogens in the respiratory tract. However, the effective delivery of antigens through the nasal barrier and boosting of a robust systematic and mucosal immune remain challenges in intranasal vaccine development. Here, we describe an intranasally administered self-healing hydrogel vaccine with a reversible strain-dependent sol-gel transition by precisely modulating the self-assembly processes between the natural drug rhein and aluminum ions. The highly bioadhesive hydrogel vaccine enhances antigen stability and prolongs residence time in the nasal cavity and lungs by confining the antigen to the surface of the nasal mucosa, acting as a "mucosal mask". The hydrogel also stimulates superior immunoenhancing properties, including antigen internalization, cross-presentation, and dendritic cell maturation. Furthermore, the formulation recruits immunocytes to the nasal mucosa and nasal-associated lymphoid tissue (NALT) while enhancing antigen-specific humoral, cellular, and mucosal immune responses. Our findings present a promising strategy for preparing intranasal vaccines for infectious diseases or cancer.

Self-Amplifying RNA Generated with the Modified Nucleotides 5-MethylCytidine and 5-MethylUridine Mediate Strong Expression & Immunogenicity in vivo

Abstract When utilized in therapeutic applications, synthetic self-amplifying RNA can lead to higher and more sustained expression than standard messenger RNA. This feature is particularly important for gene replacement therapy applications where prolonged expression could reduce the dose and frequency of treatments. The inclusion of modified nucleotides in synthetic non-amplifying mRNA has been shown to increase RNA stability, reduce immune activation and enhance gene expression. Preclinical and clinical studies with self-amplifying RNA (saRNA) have so far exclusively relied on RNA containing the canonical nucleotides adenosine, cytidine, guanosine and uridine. For the first time, we show that non-canonical nucleotides, such as m5C and m5U, are sufficiently compatible with a replicon derived from Venezuelan equine encephalitis alphavirus mediating protein translation in vitro, while those containing m1ψ in place of uridine show no detectable expression. When administered in vivo, saRNA generated with m5C or m5U mediate sustained gene expression of the luciferase reporter gene with those incorporating m5U appearing to lead to more prolonged expression. Finally, distinct antigen-specific humoral and cellular immune responses were induced by modified saRNA encoding the model antigen ovalbumin. The use of modified nucleotides with saRNA-based platforms could enhance their potential to be used effectively in a variety of applications.

Armed with IL-2 based fusion protein improves CAR-T cell fitness and efficacy against solid tumors

Chimeric antigen receptor T (CAR-T) cell therapy is regarded as a potent immunotherapy and has made significant success in hematologic malignancies by eliciting antigen-specific immune responses. However, response rates of CAR-T cell therapy against solid tumors with immunosuppressive microenvironments remain limited. Co-engineering strategies are advancing methods to overcome immunosuppressive barriers and enhance antitumor responses. Here, we engineered an IL-2 mutein co-engineered CAR-T for the improvement of CAR-T cells against solid tumors and the efficient inhibition of solid tumors. We equipped the CAR-T cells with co-expressing both tumor antigen-targeted CAR and a mutated human interleukin-2 (IL-2m), conferring enhanced CAR-T cells fitness in vitro, reshaped immune-excluded TME, enhanced CAR-T infiltration in solid tumors, and improved tumor control without significant systemic toxicity. Overall, this subject demonstrates the universal CAR-T cells armed strategy for the development and optimization of CAR-T cells against solid tumors.

Abstract CT193: Phase 1 clinical trial evaluating the safety, tolerability, and immunogenicity of AST-021p, a novel cancer vaccine targeting HSP90-derived epitopes in advanced solid tumor patients (CornerStone-002)

Abstract Background: AST-021p is an "off-the-shelf" therapeutic cancer vaccine composed of two peptides, AST-021p4 and AST-021p5, derived from MHC class II-specific epitopes of Hunam HSP90 protein. Both peptides serve as antigenic determinants for HSP90 and act as a tumor-associated antigen (TAA) inducing activation of type 1 helper T cells (Th1). Th1 cell immunologically elicited by AST-021p secrete cytokines, including IFN-γ, and activate local antigen presenting cells (APC) enhancing cross priming at the tumor site. This immunological activity, through the mechanism of cross priming, sequentially leads to the activation of cytotoxic T cells and epitope spreading. In preclinical studies using the MMTV neu gene overexpressing mouse model, the administration of the HSP90 peptide vaccine, AST-021p, significantly inhibited tumor growth. Based on these scientific rationales, The first-in-human (FIH) study was conducted to evaluate the safety, tolerability, and immunogenicity of AST-021p in advanced solid tumor patients have no treatment option. Methods: Phase 1 study (CornerStone-002, NCT No.: 04864418) was conducted on patients with recurrent or metastatic solid tumors for whom applicable standard treatment options were unavailable (patient enrollment period: June 9, 2021, to June 14, 2023). Four-dose cohorts of AST-021p (Cohort 1: 1.2mg, Cohort 2: 2.4mg, Cohort 3: 3.6mg, Cohort 4: 4.8mg) received three vaccinations at 2-week intervals (Priming immunization) followed by boosting immunization with 4-week interval based on PI's discretion. Enrolling one patient initially in the lowest dose cohort, Cohort 1 (1.2mg), dose escalation occurred sequentially after confirming safety and tolerability. Subsequently, each dose cohort (Cohort 2: 2.4mg, Cohort 3: 3.6mg, Cohort 4: 4.8mg) enrolled six patients each for vaccination. Antigen-specific immune response was assessed by IFNγ ELISPOT using PBMCs from the subjects at baseline, 4 weeks, and 8 weeks. Results: During the study period, a total of 22 patients were enrolled (Cohort 1: 1, Cohort 2: 6, Cohort 3: 8, Cohort 4: 7). The median age of the patients was 66, with a gender distribution of 14 males and 8 females. Among the 22 patients, 20 completed the priming immunization with 3 vaccinations, and 7 of them proceeded with the boosting immunization. A total of 75 adverse events were reported in 21 patients (95.5%). Adverse drug reactions (ADRs) occurred in 15 patients (68.2%), accounting for 92 cases. The most common ADR was injection site reaction (ISR), observed in 15 patients (68.2%), with 35 cases and all reactions were graded as 1 or 2. Serious adverse events (SAEs), including gastrointestinal hemorrhage, vomiting, and aspiration, were observed in 3 patients (13.6%), totaling 3 cases. However, these events were unrelated to AST-021p. Conclusions: This study, as the first-in-human (FIH) administration of the AST-021p, demonstrated safety and tolerability across all four dose groups. Plus, optimal immunogenic dose (OID) and recommended phase 2 dose (RP2D) were confirmed based on AST-021p-specific immunogenicity and safety. Citation Format: Eun Joo Kang, Kyoungmin Lee, In Ho Kim, Ju Won Kim, Jwa Hoon Kim, Soohyeon Lee, Jiwon Lee, Kabsoo Shin, Sejun Park, Joori Kim, Hyun Won Shin, Kyong Hwa Park. Phase 1 clinical trial evaluating the safety, tolerability, and immunogenicity of AST-021p, a novel cancer vaccine targeting HSP90-derived epitopes in advanced solid tumor patients (CornerStone-002) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr CT193.

Immunomodulatory effects of Diospyros peregrina fruit preparation (DFP) in non-small cell lung cancer (NSCLC) by utilizing dendritic cell-mediated antigen presentation and T helper (TH) cell differentiation.

Diospyros peregrina is a dioecious plant which is native to India. It belongs to the family of Ebenaceae and is extensively used to treat various ailments, such as leucorrhoea and other uterine-related problems. Though few studies have been on D. peregrina for their anti-tumour response, little is known. Therefore, this intrigued us to understand its immunomodulator capabilities on various types of cancer extensively. Our primary focus is on NSCLC (Non-Small Cell Lung Cancer), which is ranked as the second largest form of cancer in the world, and the treatments demand non-invasive agents to target NSCLC effectively. In an objective to generate an efficient Lung Cancer Associated Antigen (LCA) specific anti-tumour immune response, LCA was presented using dendritic cells (DCs) in the presence of D. peregrina fruit preparation (DFP). Moreover, we also investigated DFP's role in the differentiation of T-helper (TH) cells. Therefore, this study aimed at better LCA presentation mediated by DFP by activating the LCA pulsed DCs and T helper cell differentiation for better immune response. DCs were pulsed with LCA for tumour antigen presentation in vitro, with and without DFP. Differentially pulsed DCs were irradiated to co-culture with autologous and allogeneic lymphocytes. Extracellular supernatants were collected for the estimation of cytokine levels by ELISA. LDH release assay was performed to test Cytotoxic T lymphocytes (CTLs) mediated lung tumour cell cytotoxicity. Thus, DFP may be a potential vaccine to generate anti-LCA immune responses to restrict NSCLC.

Abstract CT052: A phase IB study of mRNA-based active cancer vaccine, BI1361849, combined with durvalumab and tremelimumab immunotherapy in patients with non-small cell lung cancer (NSCLC)

Abstract Introduction. BI1361849 is an RNA-based cancer vaccine, encoding for 6 tumor-associated antigens, including MUC1, survivin, NY-ESO-1, 5T4, MAGE-C2 and MAGE-C1, to balance systemic antigen-specific immune response. This open-label multicenter 2-arm Phase IB study evaluated the safety and preliminary efficacy of the addition of BI1361849 to durvalumab with or without tremelimumab for NSCLC. Methods. Patients with metastatic NSCLC were enrolled in one of the two arms, Arm A: BI1361849 + durvalumab and Arm B: BI1361849 + durvalumab+ tremelimumab, to determine Recommended Combination Dose (RCD) for durvalumab according to a standard 3 + 3 design with a starting dose of 1500 mg and possible de-escalation to 750 mg, and a fixed total dose of BI1361849 at 960 μg and tremelimumab at 75 mg. Primary endpoints were safety and tolerability. Secondary endpoints were progression free survival (PFS), objective response rate (ORR), disease control rate (DCR), duration of response (DOR), and overall survival (OS), based on irRECIST and RECIST. Statistical analysis included descriptive statistics, Fisher exact test, and Kaplan-Meier method. Results. A total of 57 patients were treated with 23 patients in Arm A and 34 patients in Arm B for safety analysis. Among them, 36 patients received at least 75% of the scheduled doses of durvalumab and tremelimumab and at least 4 vaccine doses in the first 2 cycles to meet efficacy analysis requirement with 17 patients in Arm A and 19 patients in Arm B. The RCD was durvalumab at 1500 mg, BI 1361849 at 960 μg, and tremelimumab at 75 mg. Both arms had comparable treatment related adverse events (56-57%) with generally low grade and manageable within current guidelines. Both arms had a comparable rate of treatment discontinuation (22-24%). For efficacy analysis, patients in Arm A had 29% ORR and 71% DCR, median months of 10, 7.3 5.7, and not reached for DOR, irPFS, PFS, and OS, respectively. In contrast, Arm B yielded 11% ORR and 53% DCR, median months of 6, 2.5, 2.5 and 10 for DOR, irPFS, PFS, and OS, respectively. Conclusions. Overall, the doublet and triplet regimens were well tolerated. In comparison to recent published studies treated with durvalumab and/or tremelimumab, incorporation of BI1361849 to durvalumab yielded comparable or better treatment response, PFS, and OS than durvalumab alone or with chemotherapy. However, tremelimumab did not add clinical benefit. Citation Format: Dung-Tsa Chen, Joshua Sabari, Jonathan Thompson, Jiaxin Niu, Hirva Mamdani, Ram Thapa, Zachary Thompson, Thanmai Posina, Aileen Ryan, Ralph Venhaus, Toni Ricciardi, Kristen Ramirez, Tawee Tanvetyanon, Jhanelle Gray. A phase IB study of mRNA-based active cancer vaccine, BI1361849, combined with durvalumab and tremelimumab immunotherapy in patients with non-small cell lung cancer (NSCLC) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr CT052.

Abstract LB352: Unraveling therapeutic potentials of mRNA-based cancer vaccine concurrently targeting heterogeneous KRAS mutant cancer

Abstract The utilization of messenger RNA (mRNA)-based cancer vaccines represents a compelling therapeutic strategy, providing distinct advantages over alternative modalities such as peptide vaccines or small-molecule-based cancer therapies. This innovative approach enables the concurrent targeting of heterogeneous cancer populations characterized by multiple mutations while preserving the integrity of normal cells. The Kristen rat sarcoma viral oncogene (KRAS) mutation is prevalent in lung, pancreatic, and colon cancer, and patients harboring KRAS mutations face a particularly dismal prognosis. This study focuses on the optimization of an mRNA-based cancer vaccine designed to target various KRAS mutations, demonstrating its efficacy inducing potent anti-tumor immune responses in both ex vivo immune cell and cancer cell co-culture, as well as in vivo syngeneic mouse cancer models. For the ex vivo evaluation of the multi-targeting capabilities of the vaccine, human peripheral blood mononuclear cells (PBMCs) primed with our mRNA cancer vaccine were subsequently co-incubated with human normal and cancer cell lines harboring KRAS mutations to assess antigen-specific immune responses and the normal cell-sparing effect of the cancer vaccine. Four types of KRAS mutant cancer cells co-cultured with the vaccine-activated PBMCs showed substantially decreased cell viability. We found enrichment of cytotoxic and pro-inflammatory cytokines including ganzyme B, IFNγ, IL-6 and IL-10 in co-culture system with KRAS vaccine transfected monocyte and PBMCs. This suggests that multiple KRAS mutations can be targeted with a single type of mRNA vaccine candidate. In the LL/2 syngeneic model (murine Lewis lung carcinoma, KRAS G12C mutant), the in vivo efficacy of KRAS cancer vaccine candidates was evaluated, revealing a notable reduction in tumor growth in response to the vaccine treatment. Also, during the treatment period, there were no significant clinical symptoms in the mouse between groups indicating the vaccine doses were tolerable. These results indicate that the KRAS vaccine effectively inhibits the growth of an LL/2 tumor which has the immunosuppressive tumor microenvironment. The ability to address the challenges posed by diverse KRAS mutations underscores the promising potential of this multiple-KRAS mutation targeting mRNA-based cancer vaccine in the pursuit of enhanced therapeutic outcomes for patients afflicted with KRAS-mutant cancer. Citation Format: Chang Gyu Lim, Seung-Hyun Shin, Youngjin Han, Yong Ho Heo, Innah Kim, Ji Hee Lee, Miyoung Lee, Seongju Jeong, Yunjae Kim, Jooyun Byun, Daejin Kim, In Young Choi. Unraveling therapeutic potentials of mRNA-based cancer vaccine concurrently targeting heterogeneous KRAS mutant cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr LB352.

Developmental self-reactivity determines pathogenic Tc17 differentiation potential of naive CD8+ T cells in murine models of inflammation

The differentiation of naive CD8+ T cells into effector cells is important for establishing immunity. However, the effect of heterogeneous naive CD8+ T cell populations is not fully understood. Here, we demonstrate that steady-state naive CD8+ T cells are composed of functionally heterogeneous subpopulations that differ in their ability to differentiate into type 17 cytotoxic effector cells (Tc17) in a context of murine inflammatory disease models, such as inflammatory bowel disease and graft-versus-host disease. The differential ability of Tc17 differentiation is not related to T-cell receptor (TCR) diversity and antigen specificity but is inversely correlated with self-reactivity acquired during development. Mechanistically, this phenomenon is linked to differential levels of intrinsic TCR sensitivity and basal Suppressor of Mothers Against Decapentaplegic 3 (SMAD3) expression, generating a wide spectrum of Tc17 differentiation potential within naive CD8+ T cell populations. These findings suggest that developmental self-reactivity can determine the fate of naive CD8+ T cells to generate functionally distinct effector populations and achieve immense diversity and complexity in antigen-specific T-cell immune responses.

A measles-vectored vaccine candidate expressing prefusion-stabilized SARS-CoV-2 spike protein brought to phase I/II clinical trials: protection of African green monkeys from COVID-19 disease.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) emerged at the end of 2019 and is responsible for the largest human pandemic in 100 years. Thirty-four vaccines are currently approved for use worldwide, and approximately 67% of the world population has received a complete primary series of one, yet countries are dealing with new waves of infections, variant viruses continue to emerge, and breakthrough infections are frequent secondary to waning immunity. Here, we evaluate a measles virus (MV)-vectored vaccine expressing a stabilized prefusion SARS-CoV-2 spike (S) protein (MV-ATU3-S2PΔF2A; V591) with demonstrated immunogenicity in mouse models (see companion article [J. Brunet, Z. Choucha, M. Gransagne, H. Tabbal, M.-W. Ku et al., J Virol 98:e01693-23, 2024, https://doi.org/10.1128/jvi.01693-23]) in an established African green monkey model of disease. Animals were vaccinated with V591 or the control vaccine (an equivalent MV-vectored vaccine with an irrelevant antigen) intramuscularly using a prime/boost schedule, followed by challenge with an early pandemic isolate of SARS-CoV-2 at 56 days post-vaccination. Pre-challenge, only V591-vaccinated animals developed S-specific antibodies that had virus-neutralizing activity as well as S-specific T cells. Following the challenge, V591-vaccinated animals had lower infectious virus and viral (v) RNA loads in mucosal secretions and stopped shedding virus in these secretions earlier. vRNA loads were lower in these animals in respiratory and gastrointestinal tract tissues at necropsy. This correlated with a lower disease burden in the lungs as quantified by PET/CT at early and late time points post-challenge and by pathological analysis at necropsy.IMPORTANCESevere acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is responsible for the largest human pandemic in 100 years. Even though vaccines are currently available, countries are dealing with new waves of infections, variant viruses continue to emerge, breakthrough infections are frequent, and vaccine hesitancy persists. This study uses a safe and effective measles vaccine as a platform for vaccination against SARS-CoV-2. The candidate vaccine was used to vaccinate African green monkeys (AGMs). All vaccinated AGMs developed robust antigen-specific immune responses. After challenge, these AGMs produced less virus in mucosal secretions, for a shorter period, and had a reduced disease burden in the lungs compared to control animals. At necropsy, lower levels of viral RNA were detected in tissue samples from vaccinated animals, and the lungs of these animals lacked the histologic hallmarks of SARS-CoV-2 disease observed exclusively in the control AGMs.