Memory Subsets Research Articles

Mucosal Vaccination With Recombinant Tm-WAP49 Protein Induces Protective Humoral and Cellular Immunity Against Experimental Trichuriasis in AKR Mice.

Trichuriasis is one of the most common neglected tropical diseases of the world’s poorest people. A recombinant vaccine composed of Tm-WAP49, an immunodominant antigen secreted by adult Trichuris stichocytes into the mucosa of the cecum to which the parasite attaches, is under development. The prototype is being evaluated in a mouse model of Trichuris muris infection, with the ultimate goal of producing a mucosal vaccine through intranasal delivery. Intranasal immunization of mice with Tm-WAP49 formulated with the adjuvant OCH, a truncated analog of alpha-GalCer with adjuvanticity to stimulate natural killer T cells (NKT) and mucosal immunity, induced significantly high levels of IgG and its subclasses (IgG1 and IgG2a) in immunized mice. This also resulted in a significant reduction of worm burden after challenge with T. muris-infective eggs. The addition of QS-21 adjuvant to this vaccine formulation further reduced worm counts. The improved protection from the dual-adjuvanted vaccine correlated with higher serum antibody responses (IgG, IgG1, IgG2a, IgA) as well as with the induction of antigen-specific IgA in the nasal mucosa. It was also associated with the robust cellular responses including functional subsets of CD4 T cells producing IL-4, and cytotoxic CD8 T cells expressing granzyme B. The worm reduction achieved by mucosal immunization was higher than that induced by subcutaneous immunization. Intranasal immunization also induced a significantly higher nasal mucosa-secreted antigen-specific IgA response, as well as higher functional cellular responses including CD4+IL4+ (Th1) and CD8+GnzB+ (Th2) T cells, and antigen-specific INFγ-producing T cells in both spleen and MLNs and antibody-producing B cells (CD19+B220+/B220+GL7+). Mucosal immunization further induced long-term T lymphocyte memory with increased central (CD62L+CD44+) and effector (CD62L-CD44+) memory subsets of both CD4 and CD8 T cells at 60 days after the last immunization. In summary, intranasal immunization with recombinant Tm-WAP49 protein induced strong protection versus murine trichuriasis. It represents a promising vaccination approach against intestinal nematodes.

Immune Reconstitution Following Autologous Hematopoietic Stem Cell Transplantation for Multiple Sclerosis: A Review on Behalf of the EBMT Autoimmune Diseases Working Party

Multiple sclerosis (MS) is a central nervous system (CNS) disorder, which is mediated by an abnormal immune response coordinated by T and B cells resulting in areas of inflammation, demyelination, and axonal loss. Disease-modifying treatments (DMTs) are available to dampen the inflammatory aggression but are ineffective in many patients. Autologous hematopoietic stem cell transplantation (HSCT) has been used as treatment in patients with a highly active disease, achieving a long-term clinical remission in most. The rationale of the intervention is to eradicate inflammatory autoreactive cells with lympho-ablative regimens and restore immune tolerance. Immunological studies have demonstrated that autologous HSCT induces a renewal of TCR repertoires, resurgence of immune regulatory cells, and depletion of proinflammatory T cell subsets, suggesting a “resetting” of immunological memory. Although our understanding of the clinical and immunological effects of autologous HSCT has progressed, further work is required to characterize the mechanisms that underlie treatment efficacy. Considering that memory B cells are disease-promoting and stem-like T cells are multipotent progenitors involved in self-regeneration of central and effector memory cells, investigating the reconstitution of B cell compartment and stem and effector subsets of immunological memory following autologous HSCT could elucidate those mechanisms. Since all subjects need to be optimally protected from vaccine-preventable diseases (including COVID-19), there is a need to ensure that vaccination in subjects undergoing HSCT is effective and safe. Additionally, the study of vaccination in HSCT-treated subjects as a means of evaluating immune responses could further distinguish broad immunosuppression from immune resetting.

Development of Tbet- and CD11c-expressing B cells in a viral infection requires T follicular helper cells outside of germinal centers

Tbet+CD11c+ B cells arise during type 1 pathogen challenge, aging, and autoimmunity in mice and humans. Here, we examined the developmental requirements of this B cell subset. In acute infection, T follicular helper (Tfh) cells, but not Th1 cells, drove Tbet+CD11c+ B cell generation through proximal delivery of help. Tbet+CD11c+ B cells developed prior to germinal center (GC) formation, exhibiting phenotypic and transcriptional profiles distinct from GC B cells. Fate tracking revealed that most Tbet+CD11c+ B cells developed independently of GC entry and cell-intrinsic Bcl6 expression. Tbet+CD11c+ and GC B cells exhibited minimal repertoire overlap, indicating distinct developmental pathways. As the infection resolved, Tbet+CD11c+ B cells localized to the marginal zone where splenic retention depended on integrins LFA-1 and VLA-4, forming a competitive memory subset that contributed to antibody production and secondary GC seeding upon rechallenge. Therefore, Tbet+CD11c+ B cells comprise a GC-independent memory subset capable of rapid and robust recall responses.

Effectiveness of working memory on visual-spatial working memory performance of pre-school children with learning problem at risk

Introduction: Specific learning difficulties are the existence of disorder in one or more of the basic psychological processes which play a role in the process of understanding and using spoken or written language. Aim: The current study aimed to investigate the effect of working memory intervention on visual-spatial working memory performance among the preschool children at risk of learning disabilities. Method: This study was a quasi-experimental study including pretest and posttest design with a control group. The statistical population comprised all children who were studying in preschool centers and kindergartens (2016-2017) in Tehran (District 14). A sample of 20 preschool students who were selected through purposive sampling participated in this study. They were randomly divided into two experimental and control groups (10 participants in each group). The experimental group participated in working memory program to 60-minute 12 sessions during the three months. In order to measure the performance of working memory, the working memory subset of IQ scale, Tehran-Stanford-Binet test and N-Back test were used. The data were analyzed by SPSS-22 and t-Test. Results: The mean and standard deviation of the subjects age are 6.2 and 0.45, respectively. The results showed that the working memory intervention caused the improvement on visual-spatial working memory performance of children (P<0.001). Conclusion: Considering the findings of the present study, it seems that visual-spatial working memory intervention can improve the working memory of preschool children and it can be used to enhance the visual-spatial working memory function of children and prevent the learning problems.

Update on the etiopathogenesis of psoriasis (Review).

Psoriasis is one of the most common immune-mediated chronic inflammatory skin disorders, involving hyperproliferative keratinocytes and infiltration of T cells, dendritic cells, macrophages, and neutrophils. Multiple factors appear to play important roles in the pathogenesis of psoriasis. These environmental (e.g., infectious agents and trauma), genetic, and immunologic factors are reviewed in this article. Although the pathogenesis of psoriasis remains to be established, data suggesting immune cell dysregulation in the skin are available. The involvement of the immune system, particularly T cells, in the etiopathogenesis of psoriasis is discussed in this review, indicating a potential justification for innovative treatment intervention. Besides describing pathogenic T cells, the aim of the review was to assess the function of newly identified antimicrobial peptides (AMPs), interleukin (IL)-23, IL-17, and tissue resident memory cells (TRMs), and their role in psoriasis. Furthermore, new insights were presented regarding TRMs, a recently identified subset of memory T cells, and the role they play in the local memory of disease, making them a potential new therapeutic target in psoriasis. Finally, current developments in T-cell research and cytokine-targeted therapy for psoriasis treatment are reviewed.

TGF-β Signaling Supports HIV Latency in a Memory CD4+ T Cell Based In Vitro Model.

During antiretroviral therapy (ART), HIV-1 persists as a latent reservoir in CD4+ T cell subsets in central (TCM), transitional (TTM) and effector memory (TEM) CD4+ T cells. Understanding the mechanisms that support HIV-1 latency in each of these subsets is essential to the identification of cure strategies to eliminate them. Due to the very low frequency of latently infected cells in vivo, model systems that can accurately reflect the heterogenous population of HIV-1 infected cells are a critical component in HIV cure discoveries. Here, we describe a novel primary cell-based model of HIV-1 latency that recapitulates the complex dynamics of the establishment and maintenance of the latent reservoir in different memory T cell subsets. The latency and reversion assay (LARA ) culture conditions uniquely retain phenotypically and transcriptionally distinct memory CD4+ T cell subsets that allow in a single assay to assess LRA activity in each memory subset and differential examination of the dynamics of HIV latency reversal.

Free Papers Compiled

Background–Schizophrenia is a chronic mental illness that affects various domains of socio-occupational functioning.Cognitive dysfunction is seen early in the illness and gradually increases with time .Substance abuse is common in Schizophrenia.The lifetime prevalence of any drug abuse(other than tobacco) is often>50% & is associated with poorer outcome.Aim– To explore the possible additive effect of alcohol on cognitive function among patients with Schizophrenia.Methods – A cross-sectional observational study was carried out in the inpatient department of a psychiatric tertiary care centre. The study consisted of 40 participants (36 males and 4 females).All participants between the ages of 18 –60 years, who met diagnostic criteria of schizophrenia (ICD 10) and those with AUDIT score>8 (for dual diagnosis group) ,had at least 2 years of illness duration. Persons who had a co-morbid psychiatric illness, medical illness were excluded. Patients were allocated into 2 groups - schizophrenia & schizophrenia plus alcohol use disorder.Symptom severity was assessed using PANSS ,cognitive assessment was done using PGI memory subsets (Verbal retention for similar & dissimilar pairs,Visual retention and recognition) Trail A &Trail B.Results– Verbal retention for dissimilar pairs (t= -5.14 ,p=0.02) was better in Schizophrenia group as compared to dual diagnosis group.Differences between mean values were present in other tests among both the groups but was not statistically significant.Conclusion -Cognitive decline is more prominent in dual diagnosis group (schizophrenia plus alcohol use disorder ) as compared to Schizophrenia group.

Off-the-Shelf Partial HLA Matching SARS-CoV-2 Antigen Specific T Cell Therapy: A New Possibility for COVID-19 Treatment.

BackgroundImmunological characteristics of COVID-19 show pathological hyperinflammation associated with lymphopenia and dysfunctional T cell responses. These features provide a rationale for restoring functional T cell immunity in COVID-19 patients by adoptive transfer of SARS-CoV-2 specific T cells.MethodsTo generate SARS-CoV-2 specific T cells, we isolated peripheral blood mononuclear cells from 7 COVID-19 recovered and 13 unexposed donors. Consequently, we stimulated cells with SARS-CoV-2 peptide mixtures covering spike, membrane and nucleocapsid proteins. Then, we culture expanded cells with IL-2 for 21 days. We assessed immunophenotypes, cytokine profiles, antigen specificity of the final cell products.ResultsOur results show that SARS-CoV-2 specific T cells could be expanded in both COVID-19 recovered and unexposed groups. Immunophenotypes were similar in both groups showing CD4+ T cell dominance, but CD8+ and CD3+CD56+ T cells were also present. Antigen specificity was determined by ELISPOT, intracellular cytokine assay, and cytotoxicity assays. One out of 14 individuals who were previously unexposed to SARS-CoV-2 failed to show antigen specificity. Moreover, ex-vivo expanded SARS-CoV-2 specific T cells mainly consisted of central and effector memory subsets with reduced alloreactivity against HLA-unmatched cells suggesting the possibility for the development of third-party partial HLA-matching products.DiscussionIn conclusion, our findings show that SARS-CoV-2 specific T cell can be readily expanded from both COVID-19 and unexposed individuals and can therefore be manufactured as a biopharmaceutical product to treat severe COVID-19 patients.One Sentence SummaryEx-vivo expanded SARS-CoV-2 antigen specific T cells developed as third-party partial HLA-matching products may be a promising approach for treating severe COVID-19 patients that do not respond to previous treatment options.

New Approaches to Multi-Parametric HIV-1 Genetics Using Multiple Displacement Amplification: Determining the What, How, and Where of the HIV-1 Reservoir.

Development of potential HIV-1 curative interventions requires accurate characterization of the proviral reservoir, defined as host-integrated viral DNA genomes that drive rebound of viremia upon halting ART (antiretroviral therapy). Evaluation of such interventions necessitates methods capable of pinpointing the rare, genetically intact, replication-competent proviruses within a background of defective proviruses. This evaluation can be achieved by identifying the distinct integration sites of intact proviruses within host genomes and monitoring the dynamics of these proviruses and host cell lineages over longitudinal sampling. Until recently, molecular genetic approaches at the single proviral level have been generally limited to one of a few metrics, such as proviral genome sequence/intactness, host-proviral integration site, or replication competency. New approaches, taking advantage of MDA (multiple displacement amplification) for WGA (whole genome amplification), have enabled multiparametric proviral characterization at the single-genome level, including proviral genome sequence, host-proviral integration site, and phenotypic characterization of the host cell lineage, such as CD4 memory subset and antigen specificity. In this review, we will examine the workflow of MDA-augmented molecular genetic approaches to study the HIV-1 reservoir, highlighting technical advantages and flexibility. We focus on a collection of recent studies in which investigators have used these approaches to comprehensively characterize intact and defective proviruses from donors on ART, investigate mechanisms of elite control, and define cell lineage identity and antigen specificity of infected CD4+ T cell clones. The highlighted studies exemplify how these approaches and their future iterations will be key in defining the targets and evaluating the impacts of HIV curative interventions.

Abstract P239: Safety and efficacy of copanlisib in combination with nivolumab: A phase Ib study in patients with advanced solid tumors

Abstract Introduction: Copanlisib (C) is a pan-class I PI3K inhibitor, with predominant activity against the PI3K-α and -δ isoforms, approved for patients (pts) with relapsed follicular lymphoma. The PD-1 inhibitor nivolumab (N) is approved for several advanced or metastatic solid tumors. Following preclinical demonstration of the immunomodulatory activity of C (Glaeske et al. AACR 2018), we report Phase Ib results evaluating the safety and efficacy of C+N in pts with advanced solid tumors (NCT03735628). Methods: PD-1 inhibitor-naive adult pts with advanced solid tumors received C 45 mg or 60 mg i.v. (days 1, 8, and 15; 28-day cycle) and N 240 mg (day 15 of cycle 1 and days 1 and 15 of subsequent cycles). The primary objective was determination of the recommended Phase II dose (RP2D) of C in combination with N. Secondary endpoints were safety/tolerability, pharmacokinetics (PK), and efficacy. Exploratory real-time evaluation of 77 pharmacodynamic and predictive immune cell biomarkers by flow cytometry on whole blood was performed. Results: 16 pts were treated (C 45 mg + N 240 mg, n=5; C 60 mg + N 240 mg, n=11). Median age was 65 years (range 37–89), 12 pts (75%) were male, and 8 pts (50%) had stage IV disease at diagnosis; the most common tumor types were head and neck squamous cell carcinoma (HNSCC; 7 pts) and bladder cancer (BC; 4 pts). No dose-limiting toxicities were reported. The RP2D of C+N 240 mg was 60 mg. As of 13 May 2020, 4 pts remain on treatment. The most common treatment-emergent adverse events (TEAEs) of any grade were hypertension and diarrhea (7 pts [44%] each, ≤ grade [G] 3) and maculo-papular rash and fatigue (6 pts [38%] each, ≤G3). C-related TEAEs were reported in 88% of pts, all ≤G3. AEs leading to C dose interruption/reduction were reported in 31%/19% of pts; TEAEs led to C discontinuation in 1 pt (60 mg; hematuria). Serious AEs occurred in 5 pts (31%). One G5 TEAE occurred (45 mg; general physical health deterioration, unrelated to C or N). No PK interactions were observed between C and N. Two pts had a partial response: 1 in the C 45 mg group (HNSCC) and 1 in the 60 mg group (BC; benefit sustained after 19 cycles). Stable disease was seen in 10 pts and disease progression in 3 pts; disease control rate (DCR) was 75%. Maximum decrease in circulating monocytic myeloid-derived suppressor cells (M-MDSCs; p<0.05) from baseline occurred on day 2 after C, returning to baseline on day 8. A significant increase in activated (HLA-DR+ and CD38+) natural killer and CD8+ T cells was seen 2 weeks post-treatment with C+N. Lower baseline levels of CD8+ Teffector memory (TEM) subset CD45RA-/CCR7- (CD3+/CD8+) seemed to associate with higher DCR. Conclusions: C+N showed acceptable safety and preliminary efficacy in pts with advanced solid tumors. The immunomodulatory effect of C on M-MDSCs was seen 2 days post-treatment, and lower TEM subset levels seemed to associate with better disease control. These results support further investigation of C+N in pts with advanced solid tumors. Funding: Bayer AG. Writing support: Complete HealthVizion. Citation Format: Benedito A. Carneiro, Robert Jotte, Nashat Gabrail, Omid Hamid, Funan Huang, Shalini Chaturvedi, Matthias Herpers, Lidia Mongay Soler, Barrett H. Childs, Aaron Hansen. Safety and efficacy of copanlisib in combination with nivolumab: A phase Ib study in patients with advanced solid tumors [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2021 Oct 7-10. Philadelphia (PA): AACR; Mol Cancer Ther 2021;20(12 Suppl):Abstract nr P239.

Immune Response to SARS-CoV-2 Vaccine in 2 Men

Introduction: In the trials of corona virus vaccines, detailed analyses of subsets of lymphocytes were not carried out. We present perhaps the most comprehensive immunological analysis of 29 subsets of B and T cells in 2 healthy subjects receiving 2 doses of the Pfizer SARS-CoV-2 (COVID-19) vaccine. Methods: Analyses were performed prior to vaccination, 3 weeks following the 1st dose, and 4 weeks following the 2nd dose. Total, naïve (T_N), and different memory and effector subsets (T_CM, T_EM, and T_EMRA) of CD4+ and CD8+ T cells; SARS-CoV-2 spike protein-specific tetramer+, and cytotoxic CD8+ T; subsets of T follicular cells (T_FH, T_FH1, T_FH2, T_FH1/T_FH17, and T_FH17); B-cell subsets (mature B cells, naive B cells, transitional B cells, marginal zone B cells, class-switched memory B cells, germinal center B cells, and CD21^low B cells), and plasmablasts; and regulatory lymphocytes (CD4+ Treg, CD8+ Treg, Breg, and T_FR cells) were evaluated with specific monoclonal antibodies by flow cytometry. Results: A lack of COVID-19 IgG antibodies after the 1st dose in one of 2 subjects was associated with increased regulatory lymphocytes and decreased plasmablasts. Seroconversion after the 2nd dose in this subject was associated with decreased T_FR cells and increased plasmablasts. In both subjects, CD4 T_EM and CD8 T_CM were markedly increased following the 2nd dose. T_FH1 and regulatory lymphocytes were increased (except Breg) following the 1st dose. A striking increase in SARS-CoV-2-specific CD8+ T cells was observed following the 2nd dose. Conclusion: Our data support the need for 2nd dose of vaccine to induce strong SARS-CoV-2 CD8 T-cell specific response and generation of memory subsets of CD4+ and CD8+ T cells. Regulatory lymphocytes appear to play a role in the magnitude of response.

IL-6 receptor blockade does not slow β cell loss in new-onset type 1 diabetes.

BackgroundIL-6 receptor (IL-6R) signaling drives development of T cell populations important to type 1 diabetes pathogenesis. We evaluated whether blockade of IL-6R with monoclonal antibody tocilizumab would slow loss of residual β cell function in newly diagnosed type 1 diabetes patients.MethodsWe conducted a multicenter, randomized, placebo-controlled, double-blind trial with tocilizumab in new-onset type 1 diabetes. Participants were screened within 100 days of diagnosis. Eligible participants were randomized 2:1 to receive 7 monthly doses of tocilizumab or placebo. The primary outcome was the change from screening in the mean AUC of C-peptide collected during the first 2 hours of a mixed meal tolerance test at week 52 in pediatric participants (ages 6–17 years).ResultsThere was no statistical difference in the primary outcome between tocilizumab and placebo. Immunophenotyping showed reductions in downstream signaling of the IL-6R in T cells but no changes in CD4 memory subsets, Th17 cells, Tregs, or CD4+ T effector cell resistance to Treg suppression. A DC subset decreased during therapy but regressed to baseline once therapy stopped. Tocilizumab was well tolerated.ConclusionTocilizumab reduced T cell IL-6R signaling but did not modulate CD4+ T cell phenotypes or slow loss of residual β cell function in newly diagnosed individuals with type 1 diabetes.Trial RegistrationClinicalTrials.gov NCT02293837.FundingNIH National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK) and National Institute of Allergy and Infectious Diseases (NIAID) UM1AI109565, UL1TR000004 from NIH/National Center for Research Resources (NCRR) Clinical and Translational Science Award (CTSA), NIH/NIDDK P30DK036836, NIH/NIDDK U01DK103266, NIH/NIDDK U01DK103266, 1UL1TR000064 from NIH/NCRR CTSA, NIH/National Center for Advancing Translational Sciences (NCATS) UL1TR001878, UL1TR002537 from NIH/CTSA; National Health and Medical Research Council Practitioner Fellowship (APP1136735), NIH/NIDDK U01-DK085476, NIH/CTSA UL1-TR002494, Indiana Clinical and Translational Science Institute Award UL1TR002529, Vanderbilt Institute for Clinical and Translational Research UL1TR000445. NIH/NCATS UL1TR003142, NIH/CTSA program UL1-TR002494, Veteran Affairs Administration, and 1R01AI132774.

Evolving Exhaustion of T Cells during the Course of the Disease in AML Can be Abrogated By CD33 BiTE ® Construct Mediated Cytotoxicity

Evolving Exhaustion of T Cells during the Course of the Disease in AML Can be Abrogated By CD33 BiTE ® Construct Mediated Cytotoxicity

The Impact of the Immunophenotyping Characteristics of Patients' Peripheral Blood on the Manufacturing and Clinical Outcome of CD7-Targeted Chimeric Antigen Receptor T Cells

BackgroundT cell acute lymphoblastic leukemia and T cell acute lymphoblastic lymphoma (T-ALL/LBL) is a highly aggressive hematological tumor characterized by immature lymphoblasts invading the bone marrow. Treatment options for patients with T-ALL/ LBL are currently limited. Chimeric antigen receptor (CAR)- T cell therapy has opened an era in the treatment of B-cell malignancies. However, the development of CAR-T therapy for T-ALL/LBL faces many challenges. One of them is that therapeutic targets are usually expressed on both tumor and normal T cells, which causes the potential risk of “cell fratricide”. Therefore, the difficulty of manufacturing CAR T cells for T-ALL/LBL is dramatically increased. CD7, is 40kD membrane-bound glycoprotein majorly expressed on peripheral T-cells and NK cells and their precursors. CD7 is highly expressed in almost all T-ALL/LBL and considered to be one of the most promising targets for T-ALL/LBL treatment.Patients and MethodsThis study is based on a phase I clinical trial (NCT04004637) for patients with relapse/refractory CD7 + NK/T cell lymphoma and T-ALL/LBL. To manufacture CAR-T cells, the peripheral blood mononuclear cells (PBMC) were collected from the patients who met the enrollment criteria. The proportion of viability and the ratio of the T cell markers were analyzed. Subsequently, the isolated T cells were co-transduced with CD7 protein expression blocker (PEBL) and CD7-CAR lentiviruses to obtain CD7-CAR-T cells, which can avoid the fratricide of CD7-CAR-T cells. Before the infusion, the phenotypic characteristics and cytotoxicity of CD7-CAR-T products were analyzed. Then peripheral blood (PB) of patients was collected regularly after receiving treatment to analyze the immunophenotyping of T cells.ResultsFrom August 2019 to June 2021, 24 leukopaks from patients with CD7-positive T-ALL/LBL were collected, and a total of 32 batches of CD7-CAR T cells were manufactured, with a 78.13% (25/32) successful rate. Among the 7 batches of failure cases, one patient had undergone blood collection twice and CAR-T preparation for three times, but all of three attempts failed (brown icon). Another four patients failed to prepare once. Eight patents were recruited for CD7-CAR-T treatment and 87.5% of complete remission (CR) rate was achieved (7/8), of which a patient (P4, blue icon) has been maintaining CR for more than 15 months. Two other patients, P7 (red icon) and P8 (light red icon), had CD7 - relapse at the time of 6th month and 3rd month after CR, respectively.We divided all samples into successful preparation group (GS), infusion group (GI) and preparation failure group (GF). As shown in Fig. 1A, all three groups exhibited good viability of PBMC. There was no significant difference between GS and GF, but GI was higher than that of GF. The proportion of CD3 + cells in PBMC of GS was significantly lower than that of GF, and GI also showed this feature. Meanwhile, GS and GI both have a higher CD4 +/CD8 + ratio compared with GF.The immunophenotyping results showed CD7-CAR-T products had a majority of the central memory subsets (T CM; 69.41 ± 10.71%) and effect memory subsets (T EM; 28.56 ± 10.19%), with limited number of effector T cell (T E) and naive T cells (T N) (Fig. 1B). The percentage of CAR +CD8 +CD27 + and CD4 +CD25 +CD127 - subsets associated with T cells activation and proliferation, as well as CD223 + and CD279 + subsets related to T cells suppression and exhaustion were lower, except for CD366 + subgroup that also indicated depletion signal (Fig. 1B). In addition, CD7-CAR-T cells showed strong cytotoxicity against CEM (CD7 +) tumor cells accompanied by the release of cytokines, in which the level of IL-2 is extremely low (Fig. 1C).Subsequently, we performed statistics on the proportion of CD3 + and CD4 +/CD8 + cells in the PB of patients after infusion. The proportion of CD3 + cells in the PB of the P4 has been maintained at a high level, and the ratio of CD4 +/CD8 + keeps low (Fig. 1D). P7 showed a significant decrease in the amount of T cells on the 60th day after CAR T infusion, while the ratio of CD4 +/CD8 + showed an upward trend.ConclusionThe results indicate that the success rate of CD7-CAR-T manufacturing is positively correlated with higher viability, lower CD3 + and higher CD4 + of PBMC. There was no significant difference among P4 (CR more than 15 months), P7 (CD7 - relapse at 6 th month after CR) and P8 (CD7 - relapse at 3rd month after CR). [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Pharmacodynamic Analysis of CAR-T Cell Persistence in Patients with Hematologic Malignancies Treated with NKTR-255, an IL-15 Receptor Agonist That Enhances CD8 + T-Cells: Preliminary Results from a Phase 1 Study

Pharmacodynamic Analysis of CAR-T Cell Persistence in Patients with Hematologic Malignancies Treated with NKTR-255, an IL-15 Receptor Agonist That Enhances CD8 + T-Cells: Preliminary Results from a Phase 1 Study

The Combination of Anti-Tigit and Lenalidomide Promotes Synergistic Myeloma-Specific Immunity after ASCT

Multiple myeloma is a largely incurable bone marrow (BM) resident plasma cell malignancy that is increasing in incidence. Autologous stem cell transplantation (ASCT) is the current standard consolidation therapy and a subset of patients achieve durable progression free survival that is suggestive of long-term immune control. Utilizing novel preclinical models, we have provided definitive evidence that this is largely mediated by T cell-dependent myeloma-specific immunity. In both patients and preclinical models, myeloma progression is associated with T cell dysfunction and expression of multiple inhibitory receptors suggesting a loss of immunosurveillance. In mice, we have demonstrated potent anti-myeloma efficacy of TIGIT blockade in both ASCT and non-transplant settings. Here we utilized identical TIGIT Abs that do or do not Fc bind to demonstrate that immunological efficacy after ASCT was absolutely dependent on ADCC (median survival was unreached (>110 days) in Fc-binding vs 73 days in Fc-dead and 71 days in control Ig (cIg)-treated mice). Since TIGIT inhibition does not protect against myeloma relapse in all mice, it is apparent that combinational approaches are required to target non-responders. Therefore, we hypothesized that TIGIT blockade could be combined with immunomodulatory drugs (IMiDs) to provide synergistic anti-myeloma activity after ASCT.To that end, we utilized CRBN transgenic mice to investigate the efficacy of TIGIT blockade in combination with lenalidomide, the standard of care IMiD used in maintenance therapy after clinical ASCT. Briefly, B6 Vk*MYC myeloma-bearing (MM-bearing) mice were lethally irradiated and transplanted with B6 bone marrow (BM) and a suboptimal dose of T cells followed by anti-TIGIT or control Ig (100 mg twice weekly) for 5 weeks with lenalidomide (50 mg/kg daily gavage) or control diluent from D+14 for 3 weeks (Figure 1A). The combination of anti-TIGIT and lenalidomide provided synergistic anti-myeloma efficacy evidenced by prolonged median survival (109 days in combination vs < 60 days in monotherapy/control-treated mice, p<0.01; Figure B). Myeloma M bands were also suppressed in the combination treated mice relative to monotherapy or cIg-treated mice (p<0.01; Figure 1). Analysis of BM CD8 T cells 6 weeks after ASCT demonstrated that combination therapy significantly decreased terminal exhaustion (TOX + TIM3 + CD101 + PD-1 + DNAM-1 ─) with an average of only 20% of CD8 T cells with an exhausted phenotype in the combination group compared to greater than 50% exhausted CD8 T cells in monotherapy or cIg-treated mice (p<0.05; Figure 1C-D). The combination also increased the frequency of central memory and tissue-resident memory subsets (CD49b +CD69 +; p<0.05; Figure 1C-D), and increased IFNγ production from activated (PD-1 +; p<0.05; Figure 1E) cells compared to monotherapy or control Ig-treated mice. Importantly, these phenotypic changes were specific to the BM tumor microenvironment as we observed no effect of combination or monotherapy treatment on CD8 or CD4 T cells in peripheral blood.In sum, these data provide a strong rationale for combining TIGIT inhibition with immunomodulatory drugs to prevent the progression of myeloma. [Display omitted] DisclosuresDriessens: iTeos Therapeutics: Current Employment, Current equity holder in publicly-traded company. Holmberg: Up-To-Date: Patents & Royalties; Bristol Myers Squibb: Research Funding; Janssen: Research Funding; Merck: Research Funding; Millennium-Takeda: Research Funding; Sanofi: Research Funding; Seattle Genetics: Research Funding. Hill: NeoLeukin Therapeutics: Consultancy; Compass Therapeutics: Research Funding; NapaJen Pharma: Consultancy; Generon Corporation: Consultancy; Roche: Research Funding; iTeos Therapeutics: Consultancy, Research Funding; Syndax Pharmaceuticals: Research Funding; Applied Molecular Transport: Research Funding.

The cell-surface 5'-nucleotidase CD73 defines a functional T memory cell subset that declines with age.

SUMMARYMemory T cells exhibit considerable diversity that determines their ability to be protective. Here, we examine whether changes in T cell heterogeneity contribute to the age-associated failure of immune memory. By screening for age-dependent T cell-surface markers, we identify CD4 and CD8 memory T cell subsets that are unrelated to previously defined subsets of central and effector memory cells. Memory T cells expressing the ecto-5ʹ-nucleotidase CD73 constitute a functionally distinct subset of memory T cells that declines with age. They resemble long-lived, polyfunctional memory cells but are also poised to display effector functions and to develop into cells resembling tissue-resident memory T cells (TRMs). Upstream regulators of differential chromatin accessibility and transcriptomes include transcription factors that facilitate CD73 expression and regulate TRM differentiation. CD73 is not just a surrogate marker of these regulatory networks but is directly involved in T cell survival.

Interplay between IL-10, IFN-γ, IL-17A and PD-1 Expressing EBNA1-Specific CD4+ and CD8+ T Cell Responses in the Etiologic Pathway to Endemic Burkitt Lymphoma.

Simple SummaryEndemic Burkitt lymphoma (eBL) is a common pediatric cancer in sub-Saharan Africa. The incidence of this aggressive B-cell cancer is linked to Plasmodium falciparum (Pf) malaria and Epstein–Barr virus (EBV) co-infections during childhood. Most eBL tumors contain EBV and are characterized by the Epstein–Barr Nuclear Antigen 1 (EBNA1) latency I pattern of viral gene expression. The aim of our study was to compare the phenotypes and functions of CD4+ and CD8+ T cell responses to EBNA1 in children diagnosed with eBL and in healthy EBV-seropositive children to highlight differences that contribute to the balance between anti-viral immunity and eBL pathogenesis.Children diagnosed with endemic Burkitt lymphoma (eBL) are deficient in interferon-γ (IFN-γ) responses to Epstein–Barr Nuclear Antigen1 (EBNA1), the viral protein that defines the latency I pattern in this B cell tumor. However, the contributions of immune-regulatory cytokines and phenotypes of the EBNA1-specific T cells have not been characterized for eBL. Using a bespoke flow cytometry assay we measured intracellular IFN-γ, IL-10, IL-17A expression and phenotyped CD4+ and CD8+ T cell effector memory subsets specific to EBNA1 for eBL patients compared to two groups of healthy children with divergent malaria exposures. In response to EBNA1 and a malaria antigen (PfSEA-1A), the three study groups exhibited strikingly different cytokine expression and T cell memory profiles. EBNA1-specific IFN-γ-producing CD4+ T cell response rates were lowest in eBL (40%) compared to children with high malaria (84%) and low malaria (66%) exposures (p < 0.0001 and p = 0.0004, respectively). However, eBL patients did not differ in CD8+ T cell response rates or the magnitude of IFN-γ expression. In contrast, eBL children were more likely to have EBNA1-specific CD4+ T cells expressing IL-10, and less likely to have polyfunctional IFN-γ+IL-10+ CD4+ T cells (p = 0.02). They were also more likely to have IFN-γ+IL-17A+, IFN-γ+ and IL-17A+ CD8+ T cell subsets compared to healthy children. Cytokine-producing T cell subsets were predominantly CD45RA+CCR7+ TNAIVE-LIKE cells, yet PD-1, a marker of persistent activation/exhaustion, was more highly expressed by the central memory (TCM) and effector memory (TEM) T cell subsets. In summary, our study suggests that IL-10 mediated immune regulation and depletion of IFN-γ+ EBNA1-specific CD4+ T cells are complementary mechanisms that contribute to impaired T cell cytotoxicity in eBL pathogenesis.

Dynamic SARS-CoV-2-specific B-cell and T-cell responses following immunization with an inactivated COVID-19 vaccine

ObjectiveThe dynamic adaptive immune responses elicited by the inactivated virus vaccine CoronaVac remain elusive. MethodsIn a prospective cohort of 100 healthcare professionals naïve for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) who received two doses of CoronaVac, we analysed SARS-CoV-2-specific humoral and cellular responses at four different timepoints, including before vaccination (T1), 2 weeks after the first dose (T2), 2 weeks after the booster dose (T3), and 8–10 weeks after the booster dose (T4). SARS-CoV-2-specific antibodies, serum neutralizing activities, peripheral B cells, CD4+ and CD8+ T cells and their memory subsets were simultaneously measured in this cohort. ResultsSARS-CoV-2 spike-specific IgG responses reached a peak (geometric mean titre (GMT) 54827, 30969–97065) after two doses and rapidly declined (GMT 502, 212–1190) at T4, whereas suboptimal IgA responses were detected (GMT 5, 2–9). Spike-specific circulating B cells (0.60%, 0.46–0.73% of total B cells) and memory B cells (1.18%, 0.92–1.44% of total memory B cells) were effectively induced at T3 and sustained over time (0.33%, 0.23–0.43%; 0.87%, 0.05–1.67%, respectively). SARS-CoV-2-specific circulating CD4+ T cells (0.57%, 0.47–0.66%) and CD8+ T cells (1.29%, 1.04–1.54%) were detected at T3. At T4, 0.78% (0.43–1.20%) of memory CD4+ T cells and 0.68% (0.29–1.30%) of memory CD8+ T cells were identified as SARS-CoV-2-specific, while 0.62% (0.51–0.75%) of CD4+ T cells and 0.47% (0.38–0.58%) of CD8+ T cells were SARS-CoV-2-specific terminally differentiated effector memory cells. Furthermore, age and interval between doses affected the magnitude of CoronaVac-induced immune responses. SARS-CoV-2 memory CD4+ T cells were strongly associated with both receptor binding domain (RBD)-specific memory B cells (r 0.87, p <0.0001) and SARS-CoV-2-specific memory CD8+ T cells (r 0.48, p <0.0001). ConclusionsCoronaVac induced robust circulating and memory B cell and T cell responses. Our study offers new insight into the underlying immunobiology of inactivated virus vaccines in humans and may have implications for vaccine strategies in the future.

COVID-19 in multiple-myeloma patients: cellular and humoral immunity against SARS-CoV-2 in a short- and long-term view

Multiple myeloma patients are often treated with immunomodulatory drugs, proteasome inhibitors, or monoclonal antibodies until disease progression. Continuous therapy in combination with the underlying disease frequently results in severe humoral and cellular immunodeficiency, which often manifests in recurrent infections. Here, we report on the clinical management and immunological data of three multiple-myeloma patients diagnosed with COVID-19. Despite severe hypogammaglobulinemia, deteriorated T cell counts, and neutropenia, the patients were able to combat COVID-19 by balanced response of innate immunity, strong CD8+ and CD4+ T cell activation and differentiation, development of specific T-cell memory subsets, and development of anti-SARS-CoV-2 type IgM and IgG antibodies with virus-neutralizing capacities. Even 12 months after re-introduction of lenalidomide maintenance therapy, antibody levels and virus-neutralizing antibody titers remained detectable, indicating persisting immunity against SARS-CoV-2. We conclude that in MM patients who tested positive for SARS-CoV-2 and were receiving active MM treatment, immune response assessment could be a useful tool to help guide decision-making regarding the continuation of anti-tumor therapy and supportive therapy.Key messagesImmunosuppression due to multiple myeloma might not be the crucial factor that is affecting the course of COVID-19.In this case, despite pre-existing severe deficits in CD4+ T-cell counts and IgA und IgM deficiency, we noticed a robust humoral and cellular immune response against SARS-CoV-2.Evaluation of immune response and antibody titers in MM patients that were tested positive for SARS-CoV-2 and are on active MM treatment should be performed on a larger scale; the findings might affect further treatment recommendations for COVID-19, MM treatment re-introduction, and isolation measures.