Pool Of Antigens Research Articles

Polysaccharide, Conjugate, and mRNA-based Vaccines are Immunogenic in Patients with Netherton Syndrome

BackgroundNetherton syndrome (NS) is a rare, severe genetic skin disorder, currently classified as an inborn error of immunity (IEI) due to previously reported immune dysregulation. We recently reported the results of an immunological evaluation showing no evidence for a relevant B- and/or T-cell mediated immunodeficiency, but immune responses after vaccination were not evaluated in that study. Therefore, we evaluated immune responses to three vaccine platforms in adult NS patients to further investigate the presence of a clinically relevant B- and/or T-cell immunodeficiency.MethodsVaccination responses in eight adult NS patients were assessed in a cross-sectional study performed between January and August 2022. Clinical patient data were retrospectively retrieved from electronic patient files. Immune responses to a polysaccharide Streptococcus pneumoniae vaccine (PPV23) and conjugate Haemophilus influenzae type b vaccine (ActHiB) were measured. SARS-CoV-2-specific (functional) antibody and T-cell responses following booster vaccination with an mRNA-based COVID-19 vaccine were compared to controls.ResultsNone of the included patients suffered from recurrent and/or severe infections that could be attributed to a B- and/or T-cell immunodeficiency. ActHiB induced immune responses were normal in 7/7 NS patients. PPV23 induced responses were absent in 1/7, diminished in 2/7, and normal in 4/7 patients. Levels of SARS-CoV-2-specific binding and neutralizing antibodies after mRNA-based COVID-19 booster vaccination in NS patients were comparable to controls. SARS-CoV-2-specific CD4 + T-cell responses were detectable in all NS patients. In contrast, SARS-CoV-2-specific CD8 + T-cell responses were detectable in only 2/6 NS patients. T-cell responses to a positive control antigen pool were comparable to controls.ConclusionsVaccine-induced immune responses were detectable after polysaccharide, conjugate and mRNA-based vaccination in our cohort of NS patients. A spectrum of responsiveness to vaccine challenges was found, with the ranges of vaccine responses overlapping those demonstrated in healthy control populations.

A T cell receptor specific for an HLA-A*03:01-restricted epitope in the endogenous retrovirus ERV-K-Env exhibits limited recognition of its cognate epitope

Transposable elements (TEs) are often expressed at higher levels in tumor cells than normal cells, implicating these genomic regions as an untapped pool of tumor-associated antigens. In ovarian cancer (OC), protein from the TE ERV-K is frequently expressed by tumor cells. Here we determined whether the targeting of previously identified epitope in the envelope gene (env) of ERV-K resulted in target antigen specificity against cancer cells. We found that transducing healthy donor T cells with an ERV-K-Env-specific T cell receptor construct resulted in antigen specificity only when co-cultured with HLA-A*03:01 B lymphoblastoid cells. Furthermore, in vitro priming of several healthy donors with this epitope of ERV-K-Env did not result in target antigen specificity. These data suggest that the T cell receptor is a poor candidate for targeting this specific ERV-K-Env epitope and has limited potential as a T cell therapy for OC.

CAR T-cells for pediatric solid tumors: where to go from here?

Despite the great success that chimeric antigen receptor (CAR) T-cells have had in patients with B-cell malignancies and multiple myeloma, they continue to have limited efficacy against most solid tumors. Especially in the pediatric population, pre- and post-treatment biopsies are rarely performed due to ethical reasons, and thus, our understanding is still very limited regarding the mechanisms in the tumor microenvironment by which tumor cells exclude effectors and attract immune-suppressive cells. Nevertheless, based on the principles that are known, current T-cell engineering has leveraged some of these processes and created more potent CAR T-cells. The recent discovery of new oncofetal antigens and progress made in CAR design have expanded the potential pool of candidate antigens for therapeutic development. The most promising approaches to enhance CAR T-cells are novel CAR gating strategies, creative ways of cytokine delivery to the TME without enhancing systemic toxicity, and hijacking the chemokine axis of tumors for migratory purposes. With these new modifications, the next step in the era of CAR T-cell development will be the clinical validation of these promising preclinical findings.

Antigen presentation for central tolerance induction.

The extent of central T cell tolerance is determined by the diversity of self-antigens that developing thymocytes 'see' on thymic antigen-presenting cells (APCs). Here, focusing on insights from the past decade, we review the functional adaptations of medullary thymic epithelial cells, thymic dendritic cells and thymic B cells for the purpose of tolerance induction. Their distinct cellular characteristics range from unconventional phenomena, such as promiscuous gene expression or mimicry of peripheral cell types, to strategic positioning in distinct microenvironments and divergent propensities to preferentially access endogenous or exogenous antigen pools. We also discuss how 'tonic' inflammatory signals in the thymic microenvironment may extend the intrathymically visible 'self' to include autoantigens that are otherwise associated with highly immunogenic peripheral environments.

Identification and validation of anti-protein arginine methyltransferase 5 (PRMT5) antibody as a novel biomarker for systemic sclerosis (SSc)

ObjectivesIn the complex panorama of autoimmune diseases, the characterisation of pivotal contributing autoantibodies that are involved in disease progression remains challenging. This study aimed to employ a global antibody profiling...

Limited Immunogenicity of an HLA-A*03:01-restricted Epitope of Erv-k-env in Non-hiv-1 Settings: Implications for Adoptive Cell Therapy in Cancer.

Repetitive elements (REs) are often expressed at higher levels in tumor cells than normal cells, implicating these genomic regions as an untapped pool of tumor-associated antigens. In ovarian cancer (OC), protein from the RE ERV-K is frequently expressed by tumor cells. Here we determined whether the targeting of a previously identified immunogenic epitope in the envelope gene (env) of ERV-K resulted in target antigen specificity in non-HIV-1 settings. We found that transducing healthy donor T cells with an ERV-K-Env-specific T cell receptor construct resulted in antigen specificity only when co-cultured with HLA-A*03:01 B lymphoblastoid cells. Furthermore, these transduced T cells were not specific for HLA-A*03:01 + OC cells nor for the cognate peptide in HLA-matched systems from multiple healthy donors. These data suggest that the ERV-K-Env epitope recognized by this T cell receptor is of low immunogenicity and has limited potential as a T cell target for OC.

Metal-Organic Framework-Based Nanovaccine for Relieving Immunosuppressive Tumors via Hindering Efferocytosis of Macrophages and Promoting Pyroptosis and Cuproptosis of Cancer Cells.

Current cancer vaccines face challenges due to an immunosuppressive tumor microenvironment and their limited ability to produce an effective immune response. To address the above limitations, we develop a 3-(2-spiroadamantyl)-4-methoxy-4-(3-phosphoryloxy)-phenyl-1,2-dioxetane (alkaline phosphatase substrate) and XMD8-92 (extracellular signal-regulated kinase 5 inhibitor)-codelivered copper-tetrahydroxybenzoquinone (Cu-THBQ/AX) nanosized metal-organic framework to in situ-generate therapeutic vaccination. Once inside the early endosome, the alkaline phosphatase overexpressed in the tumor cells' membrane activates the in situ type I photodynamic effect of Cu-THBQ/AX for generating •O2-, and the Cu-THBQ/AX catalyzes O2 and H2O2 to •O2- and •OH via semiquinone radical catalysis and Fenton-like reactions. This surge of ROS in early endosomes triggers caspase-3-mediated proinflammatory pyroptosis via activating phospholipase C. Meanwhile, Cu-THBQ/AX can also induce the oligomerization of dihydrolipoamide S-acetyltransferase to trigger tumor cell cuproptosis. The production of •OH could also trigger the release of XMD8-92 for effectively inhibiting the efferocytosis of macrophages to convert immunosuppressive apoptosis of cancer cells into proinflammatory secondary necrosis. The simultaneous induction of pyroptosis, cuproptosis, and secondary necrosis effectively converts the tumor microenvironment from "cold" to "hot" conditions, making it an effective antigen pool. This transformation successfully activates the antitumor immune response, inhibiting tumor growth and metastasis.

Integrating Multisector Molecular Characterization into Personalized Peptide Vaccine Design for Patients with Newly Diagnosed Glioblastoma.

Outcomes for patients with glioblastoma (GBM) remain poor despite multimodality treatment with surgery, radiation, and chemotherapy. There are few immunotherapy options due to the lack of tumor immunogenicity. Several clinical trials have reported promising results with cancer vaccines. To date, studies have used data from a single tumor site to identify targetable antigens, but this approach limits the antigen pool and is antithetical to the heterogeneity of GBM. We have implemented multisector sequencing to increase the pool of neoantigens across the GBM genomic landscape that can be incorporated into personalized peptide vaccines called NeoVax. In this study, we report the findings of four patients enrolled onto the NeoVax clinical trial (NCT0342209). Immune reactivity to NeoVax neoantigens was assessed in peripheral blood mononuclear cells pre- and post-NeoVax for patients 1 to 3 using IFNγ-ELISPOT assay. A statistically significant increase in IFNγ producing T cells at the post-NeoVax time point for several neoantigens was observed. Furthermore, a post-NeoVax tumor biopsy was obtained from patient 3 and, upon evaluation, revealed evidence of infiltrating, clonally expanded T cells. Collectively, our findings suggest that NeoVax stimulated the expansion of neoantigen-specific effector T cells and provide encouraging results to aid in the development of future neoantigen vaccine-based clinical trials in patients with GBM. Herein, we demonstrate the feasibility of incorporating multisector sampling in cancer vaccine design and provide information on the clinical applicability of clonality, distribution, and immunogenicity of the neoantigen landscape in patients with GBM.

Abstract 7474: T-cell responses across heart, blood, and tumor in patients with immune checkpoint inhibitor-related myocarditis (irMyocarditis)

Abstract Introduction: Immune checkpoint inhibitor (ICI)-related myocarditis (irMyocarditis) is a potentially lethal complication of ICI use that is characterized by the presence of clonally expanded T cells in the heart. The antigens recognized by these expanded intracardiac T-cell clones and their relationships to T-cell clones in the blood and tumors of irMyocarditis patients are poorly understood. Methods: Paired single-cell RNA sequencing (scRNA-seq) and T-cell receptor (TCR) sequencing was performed on heart tissue (n = 13) and peripheral blood from patients with irMyocarditis (n = 25) and ICI-treated controls (n = 28) using the 10X Chromium (10X Genomics) system. TCR-β chain sequencing was performed (Adaptive Biotechnologies) on four autopsy cases of patients with available scRNA-seq data and irMyocarditis heart, tumor, and histologically normal tissue. An established high-throughput protocol was used to screen expanded intracardiac TCRs against candidate antigens (Oliveira G, et al. Nature 2021). Briefly, full-length TCRs were cloned, transduced into donor T cells, and screened against autologous or major histocompatibility complex (MHC) Class I-matched antigen presenting cell lines pulsed with peptide pools that covered the full length of the ⍺-myosin, troponin-I, and troponin-T proteins alongside pools of common viral antigens and appropriate controls. Results: scRNA-seq data showed that TCRs shared between heart and blood are predominantly found in circulating CD8 T cell subsets as compared to circulating CD4 T cell subsets. The gene expression patterns of these shared T-cell clones in circulation appear distinct in fatal and non-fatal irMyocarditis patients, with shared T-cell clones in fatal cases expressing cycling markers (MKI67, STMN1) and the chemokine receptor CXCR3. TCR-β chain sequences most enriched in irMyocarditis tissue relative to control tissues were distinct from those enriched in tumor tissues, and their full-length TCR sequences could be recovered from scRNA-seq data. In total, 52 cardiac-expanded TCRs across eight donors were screened against candidate antigens. None of the screened TCRs recognized the putative cardiac autoantigens. Conclusions: T-cell clones expanded in irMyocarditis are shared in circulation, where the gene expression of these clones may help to distinguish fatal from non-fatal irMyocarditis. TCRs enriched in irMyocarditis appear to be largely distinct from those enriched in tumor and likely recognize currently unknown cardiac autoantigens. Citation Format: Steven Blum, Daniel A. Zlotoff, Neal P. Smith, Isabela J. Kernin, Swetha Ramesh, Giacomo Oliveira, Leyre Zubiri, Joshua Caplin, Nandini Samanta, Sidney Martin, Mike Wang, Alice Tirard, Pritha Sen, Yuhui Song, Katherine Xu, Jaimie L. Barth, Kamil Slowikowski, Mazen Nasrallah, Jessica Tantivit, Kasidet Manakongtreecheep, Benjamin Y. Arnold, John McGuire, Alexander B. Afeyan, Christopher J. Pinto, Daniel McLoughlin, Monica Jackson, PuiYee Chan, Aleigha Lawless, William A. Michaud, Tatyana Sharova, Linda T. Nieman, Justin F. Gainor, Dejan Juric, Mari Mino-Kenudsen, Ryan J. Sullivn, Genevieve M. Boland, James R. Stone, Catherine J. Wu, Molly F. Thomas, Tomas G. Neilan, Kerry L. Reynolds, Alexandra-Chloé Villani. T-cell responses across heart, blood, and tumor in patients with immune checkpoint inhibitor-related myocarditis (irMyocarditis) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 7474.

Abstract 1174: Incorporation of multisector analysis into the design of personalized DNA vaccines for patients with newly diagnosed glioblastoma

Abstract Glioblastoma (GBM) is the most common malignant central nervous system (CNS) tumor in adults. Despite multimodality treatment including surgery, radiation, and chemotherapy, patients with GBM have a median survival of less than 2 years. Recent clinical trials have reported promising results using cancer vaccines to stimulate a tumor-directed immune response in several histologies. The majority of trials targeted tumor neoantigens derived from a single tumor sample, which limits the antigen pool in spatially heterogeneous cancers like GBM. Our group previously implemented multisector sequencing into the design of personalized peptide vaccines to increase the candidate pool of targetable neoantigens (NCT03422094). Although the peptide vaccine treatment was successful in stimulating an effector T cell response, it was limited by a long turnaround time from vaccine design to administration and a low peptide production rate. Therefore, we incorporated multisector sampling into the design of a personalized DNA-based GBM vaccine (NCT04015700). An average of 33% of the candidate targetable neoantigens were spatially restricted to a single sampled region, which would have been missed without multisector sequencing. DNA-based vaccines are potentially advantageous over peptide-based vaccines because they enable a higher neoantigen payload and faster manufacturing time with potent immunogenicity. Here, we report the results from 9 subjects enrolled onto the study. Spatially distinct tumor regions were subjected to whole exome sequencing (WES) and RNA-sequencing (RNA-seq), data from which were used to identify neoantigens through the pVACseq algorithm (http://pvactools.org). An average of 32 neoantigens were included in each DNA vaccine, compared to an average of only 9 for the peptide vaccine, and the turnaround time from date of surgery to administration of the DNA vaccine was about half the time. Three subjects had radiographic evidence of tumor progression prior to vaccination, highlighting the aggressive nature of GBM and the need for short manufacturing timeframes. PBMCs pre- and post-DNA vaccination were collected from 7/9 subjects. All subjects presented detectable responses and 6/7 tested revealed a sustained increment in T cell reactivity against tumor neoantigens post-treatment by IFN-γ ELISpot. Intracellular staining showed a neoantigen-specific antitumor CD8/CD4 T cell cytolytic (CD69+, CD107a+) and proliferative (Ki67+) profile. Post-vaccination tumor resections were performed for 2 subjects who also were treated with PD1 blockade upon progression, providing an opportunity to explore vaccine-induced changes in the tumor microenvironment. Herein, we demonstrate the advantages of incorporating multisector sequencing into the development of DNA-based GBM vaccines and provide insights into the resulting immune responses. Citation Format: Elizabeth A. Garfinkle, Katherine E. Miller, Alexandra J. Livingstone, Renzo Perales-Linares, Neil Cooch, Alfredo Perales-Puchalt, Sarah Rochestie, Joann Peters, Niranjan Y. Sardesai, William E. Gillanders, Elaine R. Mardis, Gavin P. Dunn, Tanner M. Johanns. Incorporation of multisector analysis into the design of personalized DNA vaccines for patients with newly diagnosed glioblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1174.

Using long-read CAGE sequencing to profile cryptic-promoter-derived transcripts and their contribution to the immunopeptidome.

Recent studies have shown that the noncoding genome can produce unannotated proteins as antigens that induce immune response. One major source of this activity is the aberrant epigenetic reactivation of transposable elements (TEs). In tumors, TEs often provide cryptic or alternate promoters, which can generate transcripts that encode tumor-specific unannotated proteins. Thus, TE-derived transcripts (TE transcripts) have the potential to produce tumor-specific, but recurrent, antigens shared among many tumors. Identification of TE-derived tumor antigens holds the promise to improve cancer immunotherapy approaches; however, current genomics and computational tools are not optimized for their detection. Here we combined CAGE technology with full-length long-read transcriptome sequencing (long-read CAGE, or LRCAGE) and developed a suite of computational tools to significantly improve immunopeptidome detection by incorporating TE and other tumor transcripts into the proteome database. By applying our methods to human lung cancer cell line H1299 data, we show that long-read technology significantly improves mapping of promoters with low mappability scores and that LRCAGE guarantees accurate construction of uncharacterized 5' transcript structure. Augmenting a reference proteome database with newly characterized transcripts enabled us to detect noncanonical antigens from HLA-pulldown LC-MS/MS data. Lastly, we show that epigenetic treatment increased the number of noncanonical antigens, particularly those encoded by TE transcripts, which might expand the pool of targetable antigens for cancers with low mutational burden.

Conformation-selective rather than avidity-based binding to tumor associated antigen derived peptide-MHC enables targeting of WT1-pMHC low expressing cancer cells by anti-WT1-pMHC/CD3 T cell engagers.

T cell engagers, a category of T cell-retargeting immunotherapy, are rapidly transforming clinical cancer care. However, the lack of tumor-specific targets poses a significant roadblock for broad adaptation of this therapeutic modality in many indications, often resulting in systemic on-target off-tumor toxicity. Though various tumor-derived intracellular mutations provide a massive pool of potential tumor-specific antigens, targeting them is extremely challenging, partly due to the low copy number of tumor associated antigen (TAA)-derived pMHC on tumor cell surface. Further, the interplay of binding geometry and format valency in relation to the capacity of a T cell engager to efficiently target low density cell-surface pMHC is not well understood. Using the Wilms' tumor 1 (WT1) oncoprotein as a proof-of-principle TAA, combined with an array of IgG-like T cell engager modalities that differ in their anti-TAA valency and binding geometry, we show that the ability to induce an immunological synapse formation, resulting in potent killing of WT1 positive cancer cell lines is primarily dependent on the distinct geometrical conformations between the Fab arms of anti-WT1-HLA-A*02:01 and anti-CD3. The augmented avidity conferred by the binding of two anti-WT1-HLA-A*02:01 Fab arms has only minimal influence on cell killing potency. These findings demonstrate the need for careful examination of key design parameters for the development of next-generation T cell engagers targeting low density TAA-pMHCs on tumor cells.

How mothers tolerate their children.

Shifting pools of antigen can influence pregnancy-induced immune tolerance.

Well-established immunotherapy with R837-loaded boron neutron capture-shocked tumor cells

Although boron neutron capture therapy (BNCT) has been regarded as a curative therapy for aggressive tumors, the frequency of tumor recurrence and metastasis remains high mainly due to the adventitious nature of the immune response. Here, we explore the process of the immune response cascade after BNCT and find that the rate-limiting step is antigen presentation. To overcome this barrier, we develop a carrier-free nano-boron agent (BN-R837 @PVP), comprising hydrophilic 10B-boron nitride nanostructures loading with an immune agonist imiquimod (R837). The nano-boron agent can be internalized and cloaked into tumor cells and then following neutron irradiation, obtain the R837-loaded BNCT-shocked tumor cells (BTCs). When the BTCs are injected subcutaneously, the antigen pool and R837 are co-delivered into the draining lymph nodes, boosting immune responses as well as preventing R837 from entering the systemic circulation. This eventually results in significant inhibition of distal tumor growth and metastasis with a high rate of complete tumor regression in vivo. Moreover, memory T lymphocytes exist in these immunized mice and exhibit potent long-term anti-tumor.

Revealing the impact of CD70 expression on the manufacture and functions of CAR-70T-cells based on single-cell transcriptomics.

Chimeric antigen receptor-modified T cells (CAR T-cells) have shown exhilarative clinical efficacy for hematological malignancies. However, a shared antigen pool between healthy and malignant T-cells remains a concept to be technically and clinically explored for CAR T-cell therapy in T-cell cancers. No guidelines for engineering CAR T-cells targeting self-expressed antigens are currently available. Based on anti-CD70 CAR (CAR-70) T-cells, we constructed CD70 knock-out and wild-type CAR (CAR-70KO and CAR-70WT) T-cells and evaluated their manufacturing and anti-tumor capability. Single-cell RNA sequencing and TCR sequencing were performed to further reveal the underlying differences between the two groups of CAR T-cells. Our data showed that the disruption of target genes in T-cells before CAR transduction advantaged the expansion and cell viability of CAR T-cells during manufacturing periods, as well as the degranulation, anti-tumor efficacy, and proliferation potency in response to tumor cells. Meanwhile, more naïve and central memory phenotype CAR+ T-cells, with higher TCR clonal diversity, remained in the final products in KO samples. Gene expression profiles revealed a higher activation and exhaustion level of CAR-70WT T-cells, while signaling transduction pathway analysis identified a higher level of the phosphorylation-related pathway in CAR-70KO T-cells. This study evidenced that CD70 stimulation during manufacturing process induced early exhaustion of CAR-70T-cells. Knocking-out CD70 in T-cells prevented the exhaustion and led to a better-quality CAR-70T-cell product. Our research will contribute to good engineering CAR T-cells targeting self-expressed antigens.

Modified vaccinia Ankara–Bavarian Nordic vaccine against mpox in children

Modified vaccinia Ankara–Bavarian Nordic vaccine against mpox in children

Generation of repetitive element-specific T cells to target ovarian cancer

Abstract The immunosuppressive tumor microenvironment in ovarian cancer (OC) enables these tumors to evade the immune system, resulting in late diagnosis and poor therapeutic outcomes for patients. This immunosuppression can be partially reversed with epigenetic inhibitors, which stimulate an immunogenic interferon response in OC cells by inducing the transcription of repetitive elements (REs). However, epigenetic therapy alone is not curative in clinical trials and new treatments that further augment the immune response to OC are needed. While REs are epigenetically silent in terminally differentiated cells, their increased expression in OC cells implicate these genomic elements as an unexplored pool of tumor antigens. As epigenetic therapy also upregulates antigen processing and presentation in OC cells, the generation of T cells specific for over-expressed RE-derived antigens may boost the immune recognition of OC cells. Transducing healthy donor T cells with a T cell receptor construct that recognizes the RE ERV-K-Env resulted in antigen specificity only when co-cultured with B lymphoblastoid cells derived from the same donor that the T cell receptor was isolated from. Transduced T cells were not specific for ERV-K-Env in autologous systems. As previous reports have shown that ERV-K-Env-specific T cells can only be expanded from patients with OC, current studies are underway to expand T cells from OC patients. Upon successful expansion of ERV-K-Env-specific T cells from multiple OC patients, we will assess if specific recognition is enhanced by treating autologous tumor cells with epigenetic therapy. These efforts serve as a novel approach to enhance the immune response to OC and improve the overall OC patient survival rate. NIH F31 CA271788-01 DOD W81XWH2010273

Abstract 5114: Development of personalized RNA-based immunotherapeutics for glioblastom

Abstract Background: The development of successful immunotherapy targeting antigens in glioblastoma multiforme (GBM) remains a challenge owing to antigen heterogeneity and the low mutation burden in GBM. Cancer immunogenomics represents a complementary approach to the application of genomics in developing novel immunotherapies for GBM. Our goal is to develop a personalized RNA-based therapeutic vaccine that simultaneously targets a selected pool of tumor rejection antigens including all neoantigens and tumor-associated antigens (TAAs) as identified using cancer immunogenomics. We evaluated the treatment efficacy of these RNA therapeutics in combination with adoptive cellular therapy and checkpoint inhibitors. Methods: We identified immunogenic neoantigens and TAAs that are aberrantly overexpressed in KR158-luc and GL261 murine GBM tumors using our cancer immunogenomics pipeline called the Open Reading Frame Antigen Network (O.R.A.N.). A tumor antigen-specific RNA library was created for each tumor using a novel gene enrichment strategy and validated by RNAseq. Tumor-bearing animals were treated with adoptively transferred ex vivo expanded lymphocytes and dendritic cells loaded with the tumor antigen-specific RNA as vaccines. Additionally, we treated animals with the tumor antigen-specific RNA vaccines in combination with anti-PD-1 checkpoint-blockade therapy. Tumor progression and survival outcomes were analyzed to evaluate the treatment efficacies. We determined the peripheral and tumor-infiltrating lymphocyte responses using flow cytometry, TCR sequencing, and single cell RNAseq. Results: The tumor antigen-specific RNA vaccines were significantly effective in slowing the progression of murine GBM tumors in combination with both the adoptive cellular therapy and the checkpoint blockade therapy and improved the outcome of these therapies as compared to monotherapy alone. Additionally, we identified antigen-specific T cells targeting several of our predicted antigens and an increase in tumor-infiltrating lymphocytes and memory T cells in the treated animals. Conclusion: Here, we developed an RNA-based immunotherapy that simultaneously targets numerous tumor-specific antigens and demonstrated its therapeutic efficacy in preclinical models of GBM. Citation Format: Vrunda Trivedi, Changlin Yang, Oleg Yegorov, Kelena Klippel, Graeme Fenton, Christina von Roemeling, Lan Hoang-Minh, Elizabeth Ogando-Rivas, Paul Castillo, Kyle Dyson, Ginger Moore, Duane Mitchell. Development of personalized RNA-based immunotherapeutics for glioblastom. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5114.

COVID-19 spike polypeptide vaccine reduces the pathogenesis and viral infection in a mouse model of SARS-CoV-2.

The SARS-CoV-2 coronavirus, which causes a respiratory disease called COVID-19, has been declared a pandemic by the World Health Organization (WHO) and is still ongoing. Vaccination is the most important strategy to end the pandemic. Several vaccines have been approved, as evidenced by the ongoing global pandemic, but the pandemic is far from over and no fully effective vaccine is yet available. One of the most critical steps in vaccine development is the selection of appropriate antigens and their proper introduction into the immune system. Therefore, in this study, we developed and evaluated two proposed vaccines composed of single and multiple SARS-CoV-2 polypeptides derived from the spike protein, namely, vaccine A and vaccine B, respectively. The polypeptides were validated by the sera of COVID-19-vaccinated individuals and/or naturally infected COVID-19 patients to shortlist the starting pool of antigens followed by in vivo vaccination to hACE2 transgenic mice. The spike multiple polypeptide vaccine (vaccine B) was more potent to reduce the pathogenesis of organs, resulting in higher protection against the SARS-CoV-2 infection.

IMMU-42. ADOPTIVE HEMATOPOIETIC STEM CELL AND APD1 THERAPY CONVERGE IN THE THYMUS TO PROMOTE ANTI-GLIOBLASTOMA IMMUNITY

Abstract Our lab previously published that adoptive hematopoietic stem cell (HSC) therapy administered in conjunction with aPD1 increases median survival in multiple models of CNS malignancies. Tumor-infiltrating lymphocytes (TIL) abundance is independently associated with better outcomes in many cancers. Impressively, approximately 25% of TILs are HSC-derived in early disease. Importantly, we show HSC therapy generates de novo TILs in a thymus-dependent fashion and offer evidence of altered central tolerance following aPD1 therapy, which may permit immune activation against a larger pool of tumor-associated antigens. Therapeutic HSCs were prepared from constitutive GFP-expressing (CD45.2+) and PD1KO (CD45.2+) mice and transferred into CD45.1+ hosts. Sublethal 5Gy total body irradiation was administered day 5 post-intracranial tumor implantation. The next day, 1E6 HSCs from a 1:1 – GFP:PD1KO cocktail were injected intravenously. Descriptive statistics are reported as mean ± SD. Transferred HSCs engraft, expand, differentiate, and migrate to form approximately 25% of the TIL compartment in only 15 days. We found that, 56.2% ± 20.3 of CD3+ TILs display a thymocyte-like CD4+CD8+ (DP) phenotype in early disease, and 43.8% ± 17.4 of these DP-TILs are HSC-derived. Interestingly, DP-TILs were virtually absent in advanced disease 29 days after treatment. At this stage, HSC-derived TILs most commonly fell within the single-positive (SP) TIL subsets, comprising 9.8% ± 5.6 of SP-CD8+ TILs. Notably, HSCPD1KO-derived cells were 4x more abundant amongst SP-CD8 TILs when compared to HSCGFP-derived cells (6.5% vs 1.6%, p < 0.05). SP-CD8 thymocytes were similarly enriched for HSCPD1KO progeny (41.1% vs 24.7%, p < 0.05) which may suggest PD-1 blockade directly promotes thymopoiesis. The privileged maturation of PD1-deficient HSCs is intriguingly reversed when GFP:PD1KO HSCs are co-transferred into non-tumor-bearing mice. Thus, we show the thymus is an active site where PD-1-blocking agents function, and that HSC + aPD1 therapy is uniquely positioned to leverage thymus-dependent anti-tumor immunity.