scholarly journals Identification and validation of expressed HLA-binding breast cancer neoepitopes for potential use in individualized cancer therapy

2021 ◽  
Vol 9 (6) ◽  
pp. e002605
Author(s):  
Hannah Reimann ◽  
Andrew Nguyen ◽  
J Zachary Sanborn ◽  
Charles J Vaske ◽  
Stephen C Benz ◽  
...  
Keyword(s):  
Breast Cancer ◽  
T Cells ◽  
T Cell ◽  
Cell Line ◽  
Tumor Infiltrating Lymphocytes ◽  
The Cancer Genome Atlas ◽  
High Avidity ◽  
Binding Prediction ◽  
Hla Binding ◽  
Infiltrating Lymphocytes

BackgroundTherapeutic regimens designed to augment the immunological response of a patient with breast cancer (BC) to tumor tissue are critically informed by tumor mutational burden and the antigenicity of expressed neoepitopes. Herein we describe a neoepitope and cognate neoepitope-reactive T-cell identification and validation program that supports the development of next-generation immunotherapies.MethodsUsing GPS Cancer, NantOmics research, and The Cancer Genome Atlas databases, we developed a novel bioinformatic-based approach which assesses mutational load, neoepitope expression, human leukocyte antigen (HLA)-binding prediction, and in vitro confirmation of T-cell recognition to preferentially identify targetable neoepitopes. This program was validated by application to a BC cell line and confirmed using tumor biopsies from two patients with BC enrolled in the Tumor-Infiltrating Lymphocytes and Genomics (TILGen) study.ResultsThe antigenicity and HLA-A2 restriction of the BC cell line predicted neoepitopes were determined by reactivity of T cells from HLA-A2-expressing healthy donors. For the TILGen subjects, tumor-infiltrating lymphocytes (TILs) recognized the predicted neoepitopes both as peptides and on retroviral expression in HLA-matched Epstein-Barr virus–lymphoblastoid cell line and BC cell line MCF-7 cells; PCR clonotyping revealed the presence of T cells in the periphery with T-cell receptors for the predicted neoepitopes. These high-avidity immune responses were polyclonal, mutation-specific and restricted to either HLA class I or II. Interestingly, we observed the persistence and expansion of polyclonal T-cell responses following neoadjuvant chemotherapy.ConclusionsWe demonstrate our neoepitope prediction program allows for the successful identification of neoepitopes targeted by TILs in patients with BC, providing a means to identify tumor-specific immunogenic targets for individualized treatment, including vaccines or adoptively transferred cellular therapies.

scholarly journals Identification of shared TCR sequences from T cells in human breast cancer using emulsion RT-PCR

2016 ◽  
Vol 113 (29) ◽  
pp. 8272-8277 ◽  
Author(s):  
Daniel J. Munson ◽  
Colt A. Egelston ◽  
Kami E. Chiotti ◽  
Zuly E. Parra ◽  
Tullia C. Bruno ◽  
...  
Keyword(s):  
Breast Cancer ◽  
T Cells ◽  
T Cell ◽  
Peripheral Blood ◽  
Bilateral Breast Cancer ◽  
Tumor Infiltrating Lymphocytes ◽  
Specific Response ◽  
Rt Pcr ◽  
Primary Tumors ◽  
Infiltrating Lymphocytes

Infiltration of T cells in breast tumors correlates with improved survival of patients with breast cancer, despite relatively few mutations in these tumors. To determine if T-cell specificity can be harnessed to augment immunotherapies of breast cancer, we sought to identify the alpha–beta paired T-cell receptors (TCRs) of tumor-infiltrating lymphocytes shared between multiple patients. Because TCRs function as heterodimeric proteins, we used an emulsion-based RT-PCR assay to link and amplify TCR pairs. Using this assay on engineered T-cell hybridomas, we observed ∼85% accurate pairing fidelity, although TCR recovery frequency varied. When we applied this technique to patient samples, we found that for any given TCR pair, the dominant alpha- or beta-binding partner comprised ∼90% of the total binding partners. Analysis of TCR sequences from primary tumors showed about fourfold more overlap in tumor-involved relative to tumor-free sentinel lymph nodes. Additionally, comparison of sequences from both tumors of a patient with bilateral breast cancer showed 10% overlap. Finally, we identified a panel of unique TCRs shared between patients’ tumors and peripheral blood that were not found in the peripheral blood of controls. These TCRs encoded a range of V, J, and complementarity determining region 3 (CDR3) sequences on the alpha-chain, and displayed restricted V-beta use. The nucleotides encoding these shared TCR CDR3s varied, suggesting immune selection of this response. Harnessing these T cells may provide practical strategies to improve the shared antigen-specific response to breast cancer.

773 Adoptive Cell Therapy Response in Melanoma is Mediated by Stem-like CD8 T cells

2020 ◽  
Vol 8 (Suppl 3) ◽  
pp. A822-A822
Author(s):  
Sri Krishna ◽  
Frank Lowery ◽  
Amy Copeland ◽  
Stephanie Goff ◽  
Grégoire Altan-Bonnet ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Therapy ◽  
Single Cell ◽  
Cd8 T Cells ◽  
Tumor Infiltrating Lymphocytes ◽  
High Dimensional ◽  
Tumor Control ◽  
Intrinsic Factors ◽  
Infiltrating Lymphocytes

BackgroundAdoptive T cell therapy (ACT) utilizing ex vivo-expanded autologous tumor infiltrating lymphocytes (TILs) can result in complete regression of human cancers.1 Successful immunotherapy is influenced by several tumor-intrinsic factors.2 3 Recently, T cell-intrinsic factors have been associated with immunotherapy response in murine and human studies.4 5 Analyses of tumor-reactive TILs have concluded that anti-tumor neoantigen-specific TILs are enriched in subsets defined by the expression of PD-1 or CD39.6 7 Thus, there is a lack of consensus regarding the tumor-reactive TIL subset that is directly responsible for successful immunotherapies such as ICB and ACT. In this study, we attempted to define the fitness landscape of TIL-enriched infusion products to specifically understand its phenotypic impact on human immunotherapy responses.MethodsWe compared the phenotypic differences that could distinguish bulk ACT infusion products (I.P.) administered to patients who had complete response to therapy (complete responders, CRs, N = 24) from those whose disease progressed following ACT (non-responders, NRs, N = 30) by high dimensional single cell protein and RNA analysis of the I.P. We further analyzed the phenotypic states of anti-tumor neoantigen specific TILs from patient I.P (N = 26) by flow cytometry and single cell transcriptomics.ResultsWe identified two CD8+ TIL populations associated with clinical outcomes: a memory-progenitor CD39-negative stem-like TIL (CD39-CD69-) in the I.P. associated with complete cancer regression (overall survival, P < 0.0001, HR = 0.217, 95% CI 0.101 to 0.463) and TIL persistence, and a terminally differentiated CD39-positive TIL (CD39+CD69+) population associated with poor TIL persistence post-treatment. Although the majority (>65%) of neoantigen-reactive TILs in both responders and non-responders to ACT were found in the differentiated CD39+ state, CR infusion products also contained a pool of CD39- stem-like neoantigen-specific TILs (median = 8.8%) that was lacking in NR infusion products (median = 23.6%, P = 1.86 x 10-5). Tumor-reactive stem-like T cells were capable of self-renewal, expansion, and persistence, and mediated superior anti-tumor response in vivo.ConclusionsOur results support the hypothesis that responders to ACT received infusion products containing a pool of stem-like neoantigen-specific TILs that are able to undergo prolific expansion, give rise to differentiated subsets, and mediate long-term tumor control and T cell persistence, in line with recent murine ICB studies mediated by TCF+ progenitor T cells.4 5 Our data also suggest that TIL subsets mediating ACT-response (stem-like CD39-) might be distinct from TIL subsets enriched for anti-tumor-reactivity (terminally differentiated CD39+) in human TIL.6 7AcknowledgementsWe thank Don White for curating the melanoma patient cohort, and J. Panopoulos (Flowjo) for helpful discussions on high-dimensional analysis, and NCI Surgery Branch members for helpful insights and suggestions. S. Krishna acknowledges funding support from NCI Director’s Innovation Award from the National Cancer Institute.Trial RegistrationNAEthics ApprovalThe study was approved by NCI’s IRB ethics board.ReferencesGoff SL, et al. Randomized, prospective evaluation comparing intensity of lymphodepletion before adoptive transfer of tumor-infiltrating lymphocytes for patients with metastatic melanoma. J Clin Oncol 2016;34:2389–2397.Snyder A, et al. Genetic basis for clinical response to CTLA-4 blockade in melanoma. N Engl J Med 2014;371:2189–2199.McGranahan N, et al. Clonal neoantigens elicit T cell immunoreactivity and sensitivity to immune checkpoint blockade. Science 2016;351:1463–1469.Sade-Feldman M, et al. Defining T cell states associated with response to checkpoint immunotherapy in melanoma. Cell 2019;176:404.Miller BC, et al. Subsets of exhausted CD8 T cells differentially mediate tumor control and respond to checkpoint blockade. Nat. Immunol 2019;20:326–336.Simoni Y, et al. Bystander CD8 T cells are abundant and phenotypically distinct in human tumour infiltrates. Nature 2018;557:575–579.Gros A, et al. PD-1 identifies the patient-specific CD8+ tumor-reactive repertoire infiltrating human tumors. J Clin Invest 2014;124:2246–2259.

scholarly journals Predicted CD4+ T cell infiltration levels could indicate better overall survival in sarcoma patients

2021 ◽  
Vol 49 (1) ◽  
pp. 030006052098153
Author(s):  
Qing Bi ◽  
Yang Liu ◽  
Tao Yuan ◽  
Huizhen Wang ◽  
Bin Li ◽  
...  
Keyword(s):  
Overall Survival ◽  
T Cell ◽  
Survival Data ◽  
Tumor Infiltrating Lymphocytes ◽  
The Cancer Genome Atlas ◽  
Cell Infiltration ◽  
T Cell Infiltration ◽  
Whole Exome ◽  
Cancer Genome Atlas ◽  
Infiltrating Lymphocytes

Objective The role of tumor-infiltrating lymphocytes (TILs) has not yet been characterized in sarcomas. The aim of this bioinformatics study was to explore the effect of TILs on sarcoma survival and genome alterations. Methods Whole-exome sequencing, transcriptome sequencing, and survival data of sarcoma were obtained from The Cancer Genome Atlas. Immune infiltration scores were calculated using the Tumor Immune Estimation Resource. Potential associations between abundance of infiltrating TILs and survival or genome alterations were examined. Results Levels of CD4+ T cell infiltration were associated with overall survival of patients with pan-sarcomas, and higher CD4+ T cell infiltration levels were associated with better survival. Somatic copy number alterations, rather than mutations, were found to correlate with CD4+ T cell infiltration levels. Conclusions This data mining study indicated that CD4+ T cell infiltration levels predicted from RNA sequencing could predict sarcoma prognosis, and higher levels of CD4+ T cells infiltration indicated a better chance of survival.

scholarly journals Tumor microenvironment in triple-negative breast cancer: the correlation of tumor-associated macrophages and tumor-infiltrating lymphocytes

2021 ◽  
Author(s):  
H. Kuroda ◽  
T. Jamiyan ◽  
R. Yamaguchi ◽  
A. Kakumoto ◽  
A. Abe ◽  
...  
Keyword(s):  
Breast Cancer ◽  
T Cells ◽  
Tumor Microenvironment ◽  
Triple Negative Breast Cancer ◽  
Triple Negative ◽  
Cytotoxic T Cells ◽  
Tumor Infiltrating Lymphocytes ◽  
Tumor Associated Macrophages ◽  
Free Survival ◽  
Infiltrating Lymphocytes

Abstract Purpose Immune cells such as cytotoxic T cells, helper T cells, B cells or tumor-associated macrophages (TAMs) contribute to the anti-tumor response or pro-tumorigenic effect in triple negative breast cancer (TNBC). The interrelation of TAMs, T and B tumor-infiltrating lymphocytes (TILs) in TNBC has not been fully elucidated. Methods We evaluated the association of tumor-associated macrophages, T and B TILs in TNBC. Results TNBCs with a high CD68+, CD163+ TAMs and low CD4+, CD8+, CD20+ TILs had a significantly shorter relapse-free survival (RFS) and overall survival (OS) than those with low CD68+, CD163+ TAMs and high CD4+, CD8+, CD20+ TILs. TNBCs with high CD68+ TAMs/low CD8+ TILs showed a significantly shorter RFS and OS and a significantly poorer prognosis than those with high CD68+ TAMs/high CD8+ TILs, low CD68+ TAMs/high CD8+ TILs, and low CD68+/low CD8+. TNBCs with high CD163+ TAMs/low CD8+, low CD20 + TILs showed a significantly shorter RFS and OS and a significantly poorer prognosis than those with high CD163+ TAMs/high CD8+ TILs and high CD163+ TAMs /high CD20+ TILs. Conclusions Our study suggests that TAMs further create an optimal tumor microenvironment (TME) for growth and invasion of cancer cells when evasion of immunoreactions due to T and B TILs occurs. In TNBCs, all these events combine to affect prognosis. The process of TME is highly complex in TNBCs and for an improved understanding, larger validation studies are necessary to confirm these findings.

scholarly journals Estrogen Receptor Positive Breast Cancer with High Expression of Androgen Receptor has Less Cytolytic Activity and Worse Response to Neoadjuvant Chemotherapy but Better Survival

2019 ◽  
Vol 20 (11) ◽  
pp. 2655 ◽  
Author(s):  
Maiko Okano ◽  
Masanori Oshi ◽  
Ali Linsk Butash ◽  
Mariko Asaoka ◽  
Eriko Katsuta ◽  
...  
Keyword(s):  
Breast Cancer ◽  
T Cells ◽  
Estrogen Receptor ◽  
Androgen Receptor ◽  
Tumor Infiltrating Lymphocytes ◽  
Cytolytic Activity ◽  
Er Positive ◽  
Infiltrating Lymphocytes ◽  
Response To Neoadjuvant Chemotherapy ◽  
Positive Breast Cancer

Estrogen receptor (ER) positive breast cancer (BC), the most abundant BC subtype, is notorious for poor response to neoadjuvant chemotherapy (NAC). The androgen receptor (AR) was reported to support estradiol-mediated ER activity in an in vitro system. Recently, ER-positive BC with fewer tumor infiltrating lymphocytes (TILs) was shown to have a better prognosis, opposite to the trend seen with ER-negative BC. We hypothesized that ER-positive BC with high expression of AR will have fewer TILs and an inferior response to NAC, but with a better prognosis. In both TCGA and METABRIC cohorts, AR expression was significantly higher in ER-positive BCs compared to ER-negatives (p < 0.001, p < 0.001, respectively) and it correlated with ER expression (R = 0.630, R = 0.509, respectively). In ER-positive tumors, AR high tumors enriched UV response down (NES = 2.01, p < 0.001), and AR low tumors enriched DNA repair (NES = −2.02, p < 0.001). AR high tumors were significantly associated with procancer regulatory T-cells, and AR low tumors were associated with anticancer immune cells, such as CD4, CD8, and Gamma-Delta T-cells and memory B-cells in ER-positive BC (p < 0.01). Further, cytolytic activity was significantly lower in AR high BC in both cohorts. Finally, AR high tumors had a significantly lower rate of attaining pathological complete response to NAC (GSE22358), but better survival. In conclusion, our results demonstrated that high AR has fewer tumor infiltrating lymphocytes as well as cytolytic activity and an inferior response to NAC, but better survival in ER-positive BC.

scholarly journals Qualitative Analysis of Tumor-Infiltrating Lymphocytes across Human Tumor Types Reveals a Higher Proportion of Bystander CD8+ T Cells in Non-Melanoma Cancers Compared to Melanoma

Cancers ◽  
2020 ◽  
Vol 12 (11) ◽  
pp. 3344
Author(s):  
Aishwarya Gokuldass ◽  
Arianna Draghi ◽  
Krisztian Papp ◽  
Troels Holz Borch ◽  
Morten Nielsen ◽  
...  
Keyword(s):  
Ovarian Cancer ◽  
T Cells ◽  
T Cell ◽  
Tumor Infiltrating Lymphocytes ◽  
Autologous Tumor ◽  
Epithelial Cancers ◽  
Single Cell Rna Sequencing ◽  
Tumor Types ◽  
Infiltrating Lymphocytes

Background: Human intratumoral T cell infiltrates can be defined by quantitative or qualitative features, such as their ability to recognize autologous tumor antigens. In this study, we reproduced the tumor-T cell interactions of individual patients to determine and compared the qualitative characteristics of intratumoral T cell infiltrates across multiple tumor types. Methods: We employed 187 pairs of unselected tumor-infiltrating lymphocytes (TILs) and autologous tumor cells from patients with melanoma, renal-, ovarian-cancer or sarcoma, and single-cell RNA sequencing data from a pooled cohort of 93 patients with melanoma or epithelial cancers. Measures of TIL quality including the proportion of tumor-reactive CD8+ and CD4+ TILs, and TIL response polyfunctionality were determined. Results: Tumor-specific CD8+ and CD4+ TIL responses were detected in over half of the patients in vitro, and greater CD8+ TIL responses were observed in melanoma, regardless of previous anti-PD-1 treatment, compared to renal cancer, ovarian cancer and sarcoma. The proportion of tumor-reactive CD4+ TILs was on average lower and the differences less pronounced across tumor types. Overall, the proportion of tumor-reactive TILs in vitro was remarkably low, implying a high fraction of TILs to be bystanders, and highly variable within the same tumor type. In situ analyses, based on eight single-cell RNA-sequencing datasets encompassing melanoma and five epithelial cancers types, corroborated the results obtained in vitro. Strikingly, no strong correlation between the proportion of CD8+ and CD4+ tumor-reactive TILs was detected, suggesting the accumulation of these responses in the tumor microenvironment to follow non-overlapping biological pathways. Additionally, no strong correlation between TIL responses and tumor mutational burden (TMB) in melanoma was observed, indicating that TMB was not a major driving force of response. No substantial differences in polyfunctionality across tumor types were observed. Conclusions: These analyses shed light on the functional features defining the quality of TIL infiltrates in cancer. A significant proportion of TILs across tumor types, especially non-melanoma, are bystander T cells. These results highlight the need to develop strategies focused on the tumor-reactive TIL subpopulation.

Therapeutic effect of local Interleukin-15 with radiotherapy in breast cancer.

2017 ◽  
Vol 35 (7_suppl) ◽  
pp. 158-158
Author(s):  
Elena Garcia Martinez ◽  
Karsten A Pilones ◽  
Joseph Aryankalayil ◽  
Silvia Formenti ◽  
Sandra Demaria
Keyword(s):  
Breast Cancer ◽  
T Cells ◽  
T Cell ◽  
Nk Cells ◽  
Therapeutic Effect ◽  
Tumor Immunity ◽  
Tumor Infiltrating Lymphocytes ◽  
Tumor Control ◽  
Complete Regression ◽  
Growth And Survival

158 Background: Interleukin (IL)-15 is a key regulator of T cell homeostasis with activity in cancer and a favorable toxicity profile compared to IL-2. IL-15 stimulates the proliferation and effector differentiation of CD8+T cells, and the proliferation and activation of natural killer (NK) cells. We observed IL-15 upregulation by gene arrays in radiotherapy (RT)-treated TSA mouse breast cancer, suggesting that it may play a role in RT-induced anti-tumor immunity. However, the upregulation was modest prompting us to test the hypothesis that administration of IL-15 may enhance in situ vaccination by RT. Methods: BALB/c mice with established poorly immunogenic TSA tumors were sham-treated, treated with tumor-targeted RT (8GyX3 days), IL-15 given peri-tumorally (2 ug/mouse/day for 10 days) starting on the first day of RT, and RT+IL-15, and monitored for tumor growth and survival. Tumor infiltrating lymphocytes (TIL) were analyzed by flow cytometry and immunostaining. In some experiments, Batf3-/-mice were used as tumor recipient. Results: IL-15 by itself was ineffective, but it significantly increased tumor control by RT (p=0.0007, RT versus RT+IL-15) leading to complete responses in 50% of the mice, most of them durable. Analysis of TILs showed significantly increased NK cells (CD45+ CD3- DX5+) in tumors treated with RT+IL-15 (p<0.0004 versus sham-treated; p<0.02 versus RT). NK cells were also more activated as indicated by expression of CD122 and CD137. Depletion of NK cells completely abrogated the therapeutic effect of the combination, while CD8 T cell depletion reduced tumor control and rate of complete regression. Interestingly, Batf3-/- mice, which lack CD103+ DCs, showed reduced response to RT+IL-15 compared to WT mice. Conclusions: Data suggest that local IL-15 with RT is an effective strategy to induce anti-tumor immunity to poorly immunogenic breast cancer. NK cells are critical mediators of the response, and may act by both killing tumor cells and promoting priming of CD8 T cells. Experiments are ongoing to determine the mechanisms of durable complete responses. <footer>Acknowledgments: IL-15 was provided by NCI BRB. Garcia-Martinez E was supported by GEICAM grant.</footer>

scholarly journals The mutational load and a T-cell inflamed tumor phenotype identify ovarian cancer patients rendering tumor-reactive T cells from PD-1+ tumor-infiltrating lymphocytes

2020 ◽  
Author(s):  
Diego Salas-Benito ◽  
Enrique Conde ◽  
Ibon Tamayo-Uria ◽  
Uxua Mancheño ◽  
Edurne Elizalde ◽  
...  
Keyword(s):  
Ovarian Cancer ◽  
T Cells ◽  
T Cell ◽  
Predictive Biomarkers ◽  
Tumor Infiltrating Lymphocytes ◽  
Ovarian Tumors ◽  
Autologous Tumor ◽  
Tumor Mutational Burden ◽  
Nanostring Technology ◽  
Infiltrating Lymphocytes

Abstract Background: Adoptive immunotherapy with tumor-infiltrating lymphocytes (TIL) may benefit from the use of selective markers, such as programmed cell death protein 1 (PD-1), for tumor-specific T-cell enrichment, as well as predictive biomarkers that help identify those patients capable of rendering tumor-reactive TIL products. We have investigated this in ovarian cancer (OC) patients as candidate for TIL therapy implementation. Methods: PD-1- and PD-1+ CD8 TILs were isolated from resected ovarian tumors and, after polyclonal expansion, TIL products were tested against autologous tumor cells. Reactivity was assessed by IFNg production (ELISPOT) and upregulation of CD137. Baseline tumor samples were examined using flow cytometry, multiplexed quantitative immunofluorescence, Nanostring technology, for gene expression profile (GEP) analyses, as well as a next generation sequencing gene panel, for tumor mutational burden (TMB) calculation, to identify those features that distinguished patients with tumor-reactive and non-tumor-reactive TIL products.Results: Tumor-reactive TILs were detected in half of patients and were exclusively present in cells derived from the PD-1+ fraction. Flow-cytometric studies revealed that fresh tumors from patients rendering tumor-reactive TILs presented a significantly higher frequency of CD137+ cells within the PD1+CD8+ subset. Multiplexed immunofluorescence supported this finding, which was particularly striking in intraepithelial CD8 TILs. Baseline GEP analysis showed that patients rendering tumor-reactive TILs exhibited a significantly higher T-cell inflamed signature. Despite no correlation between TMB and GEP, both parameters stratified tumors, with patients with higher TMB and/or T-cell inflamed signature score rendering tumor-reactive TILs. Conclusion: We have demonstrated that PD-1 identifies autologous-tumor specific CD8 T cells infiltrating ovarian tumors and have uncovered predictive factors that identify OC patients who are likely to render tumor-reactive cells from PD-1+ TILs. These findings have important implications for improving the efficacy of TIL therapy in OC.

scholarly journals 660 Targeting T cell fates: converting exhaustion to memory to improve immunotherapeutic responses to cancer

2021 ◽  
Vol 9 (Suppl 3) ◽  
pp. A688-A688
Author(s):  
Nicole Scharping ◽  
Allison Cafferata ◽  
Maximilian Heeg ◽  
Quynhanh Nguyen ◽  
Ananda Goldrath
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Function ◽  
Tumor Infiltrating Lymphocytes ◽  
Cell Fates ◽  
Gene Target ◽  
Ubiquitin Protein Ligase ◽  
Improved Outcomes ◽  
Immunosuppressive Tumor Microenvironment ◽  
Infiltrating Lymphocytes

BackgroundIn cancer, CD8+ T cells have the power to target and kill tumor cells with precision, but often fail due to chronic activation from the immunosuppressive tumor microenvironment (TME). T cells that experience prolonged activation in the TME differentiate into a severely dysfunctional cell state known as exhaustion. In healthy tissues, T cells differentiate into tissue-resident memory cells (TRM) in response to infection, which remain lodged in tissues to provide protection from reinfection. When TRM-like cells are found in patient tumors, they are correlated with improved outcomes and responses to immunotherapy. Understanding how to manipulate T cell fates in an effort to prevent exhaustion and favor TRM-characteristics could benefit cancer immunotherapy.MethodsTo explore differences between these cell states, we screened the core TRM gene-expression signatures for genes downregulated as T cells undergo terminal exhaustion. Targets were then overexpressed in antigen-specific T cells and adoptively transferred into tumor-bearing mice for analysis.ResultsInterestingly, many genes related to protein regulation and processing were identified, including a novel gene called Neuralized E3 Ubiquitin Protein Ligase 3 (Neurl3). Neurl3's function is not well described, however, experimentally mutating the RING domain suggests Neurl3 transfers ubiquitin to target proteins for degradation. When Neurl3 was overexpressed in tumor-specific T cells, we found tumor infiltrating lymphocytes still upregulated inhibitory receptors PD1 and Tim3, but showed enhanced anti-tumor function. Neurl3-overexpressing T cells had increased accumulation in the TME, upregulated canonical TRM markers CD69 and CD103, produced more cytokines, controlled tumor growth, and increased mouse survival in B16 melanoma.ConclusionsThese results highlight the understudied field of negative regulation of T cell function by protein degradation in T cell differentiation fate and uncover a potential gene target for immunocellular therapies to favor functional T cell fates in cancer.Ethics ApprovalThe study was approved by UCSD's Institutional Animal Care and Use Committee, protocol number S04105.

scholarly journals S-15 in combination of Akt inhibitor promotes the expansion of CD45RA−CCR7+ tumor infiltrating lymphocytes with high cytotoxic potential and downregulating PD-1+Tim-3+ cells as well as regulatory T cells

2019 ◽  
Vol 19 (1) ◽  
Author(s):  
Benling Xu ◽  
Long Yuan ◽  
Guangyu Chen ◽  
Tiepeng Li ◽  
Jinxue Zhou ◽  
...  
Keyword(s):  
T Cells ◽  
Cell Differentiation ◽  
T Cell ◽  
Hepatocellular Cancer ◽  
Tumor Infiltrating Lymphocytes ◽  
T Cell Differentiation ◽  
Great Promise ◽  
Autologous Tumor ◽  
Akt Inhibitor ◽  
Infiltrating Lymphocytes

Abstract Background Autologous tumor-infiltrating lymphocytes (Tils) immunotherapy is a promising treatment in patients with advanced hepatocellular cancer. Although Tils treatment has shown great promise, their persistence and the efficacy after adoptive-transfer are insufficient and remain a challenge. Studies have demonstrated that IL-15 and Akt inhibitor can regulate T cell differentiation and memory. Here, we constructed S-15 (Super human IL-15), a fusion protein consisting of human IL-15, the sushi domain of the IL-15 receptor α chain and human IgG-Fc. Herein we compared the effects of S-15 with IL-2 or in combination with Akti on the expansion and activation of Tils. Methods Hepatocellular cancer tissues were obtained from 6 patients, Tils were expanded using IL-2, IL-2/S-15, IL-2/Akti or in combination IL-2/S-15/Akti. At day 10, anti-CD3 antibody was added to the culture media and expanded to day 25. The composition, exhaustion and T-cell differentiation markers (CD45RA/CCR7) were analyzed by flow cytometry. Results We found that IL-2/S-15/Akti expanded Tils and showed the highest percentage of central memory CD45RA−CCR7+ phenotype prior to anti-CD3 antibody activation and after anti-CD3 antibody activation. T cells cultured with IL-2/S-15/Akti exhibited a mixture of CD4+, CD8+, and CD3+CD4−CD8− T cells; S-15 in combination with Akt inhibitor downregulated the expression of PD-1+Tim-3+ on Tils and decreased the Tregs in Tils. Additionally, the Tils expanded in the presence of the Akt inhibitor and S-15 showed enhanced antitumor activity as indicated by the increase in IFN-γ producing tumor infiltrating CD8+ T cells and without comprising the Tils expansion. Conclusion Our study elucidates that IL-2/S-15/Akti expanded Tils and represent a viable source for the cellular therapy for patients with hepatocellular cancer.

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