T Stem Cell Memory Research Articles

Memory CD4+ T cell subsets in tumor draining lymph nodes of breast cancer patients: A focus on T stem cell memory cells

The compartments of memory T cells play a fundamental role in the immune system by substantiating specific and acquired immunity. A new subset of memory cells, T stem cell memory (TSCM) cells, with stem cell-like properties, a high capacity to proliferate, a long survival, and an ability to differentiate into all effector and memory cells has recently been introduced. In the present study, we aimed to determine the frequency of CD4+ TSCM and other T memory cell subsets in tumor draining lymph nodes of breast cancer patients. Mononuclear cells were obtained from axillary lymph nodes of 52 untreated patients with breast cancer (BC) and stained with fluorochrome conjugated anti-CD4, -CCR7, -CD45RO and -CD95 antibodies to detect different subtypes of memory cells in CD4+ lymphocyte populations. Data were acquired using a four-color FACSCalibur flow cytometer and analyzed using CellQuest Pro software. We found that >70% of CD4+ lymphocytes in draining lymph nodes of BC patients exhibited a memory phenotype of which 7.04±1.04% had a TSCM phenotype (CD4+CCR7+CD45RO-CD95+). The frequency of TSCM cells was significantly higher in tumor positive lymph nodes compared to tumor negative lymph nodes (p=0.026) as well as among those patients who had at least one affected lymph node (p=0.012). Moreover, we found that the total frequency of central memory T cells (TCM) with a low expression of CD45RO was significantly higher among these patients. The percentage of CD45ROLow TCM cells was also found to increase with tumor progression from stage I to stage III (p=0.020). On the other hand, we found that the percentage of CD95Hi effector memory T cells (TEM) was significantly decreased in involved lymph nodes (p=0.009). Our data suggest that following long-term exposure to putative tumor antigens, TSCM cells proliferate to generate a pool of committed memory and effector T cells. As the tumor progresses, the immunosuppressive milieu induced by tumor cells may slow down the differentiation of CD45ROLow TCM cells to more functional sub-populations.

A novel mechanism linking memory stem cells with innate immunity in protection against HIV-1 infection

HIV infection affects 37 million people and about 1.7 million are infected annually. Among the phase III clinical trials only the RV144 vaccine trial elicited significant protection against HIV-1 acquisition, but the efficacy and immune memory were inadequate. To boost these vaccine functions we studied T stem cell memory (TSCM) and innate immunity. TSCM cells were identified by phenotypic markers of CD4+ T cells and they were further characterised into 4 subsets. These expressed the common IL-2/IL-15 receptors and another subset of APOBEC3G anti-viral restriction factors, both of which were upregulated. In contrast, CD4+ TSCM cells expressing CCR5 co-receptors and α4β7 mucosal homing integrins were decreased. A parallel increase in CD4+ T cells was recorded with IL-15 receptors, APOBEC3G and CC chemokines, the latter downmodulating CCR5 molecules. We suggest a novel mechanism of dual memory stem cells; the established sequential memory pathway, TSCM →Central →Effector memory CD4+ T cells and the innate pathway consisting of the 4 subsets of TSCM. Both pathways are likely to be activated by endogenous HSP70. The TSCM memory stem cell and innate immunity pathways have to be optimised to boost the efficacy and immune memory of protection against HIV-1 in the clinical trial.

Abstract 1302: High complexity flow cytometry panels to monitor target expression, T-cell activation and suppresssion by novel immunotherapies in hematomalignancy clinical trials

Abstract The renaissance of cancer immunotherapies and the positive clinical responses observed with chimeric antigen receptor modified T cells in the hematomalignancy setting has stimulated substantial interest in monitoring immune cell activation and suppression to determine efficacy, prognosis and safety in new agent investigational trials. However, immunophenotyping capacity has been limited and required multi-test tube panels. To address the growing needs of clinical trials, we have developed three fit-for-purpose high complexity (10 or more markers) T-cell phenotyping flow cytometry panels on a qualified LSR FortessaTM platform that enables detection of up to 15 markers in a single test tube. The first panel allows identification of multiple phenotypes along the T-cell differentiation pathway, namely, T-naive (TN), T-stem cell memory (TSCM), T-effector memory (TEM) and T-central memory (TCM) and T-effector memory RA+ (TEMRA). The second panel identifies the most common helper T-cell phenotypes such as Th1, Th2, Treg, and Th17. A third panel determines the functional status of T-cells (e.g., activation vs. suppression) but also enables quantitation of important checkpoint inhibitor expression (e.g., PD-1) on T-cells of interest. These high complexity flow cytometry panels can serve as powerful tools for comprehensive examination of T-cells in a small volume of patient specimen. We believe these new flow cytometry panels will have a substantial impact on the determination of efficacy and safety correlates of immunomodulating agents administered alone or in combination to patients with leukemia. Citation Format: Ghanashyam Sarikonda, Devika Ashok, Anil Pahuja, Jelveh Lameh, Shabnam Tangri, Naveen Dakappagari. High complexity flow cytometry panels to monitor target expression, T-cell activation and suppresssion by novel immunotherapies in hematomalignancy clinical trials. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1302. doi:10.1158/1538-7445.AM2015-1302

Increased expression of anti-oxidant molecule Thioredoxin-1 on T cells improves immunotherapeutic control of tumors (VAC7P.1039)

Abstract Adoptive transfer of tumor epitope reactive T cells is a promising strategy to control tumor growth. However, chronically stimulated T cells expanded for adoptive cell transfer (ACT) are susceptible to cells death in an oxidative tumor microenvironment. Since oxidation of cell surface thiols (c-SH) also alters functionality of proteins, we hypothesized that increased level of thioredoxin, an anti-oxidant molecule that facilitates reduction of proteins by cysteine thiol-disulfide exchange, in T cells will result in sustained anti-tumor function. We thus crossed Trx1 transgenic mice with gp100 reactive TCR (Pmel) to generate Pmel/Trx mice. The Trx overexpressing transgenic T cells expressed higher thiols that inversely correlated with ROS, and susceptibility to TCR restimulation or H2O2 mediated cell death. These Trx1 expressing T cells showed CD62Lhi central memory-like (TCM) phenotype with reduced effector function (IFNglo2-NBDGlo), and were less effective in controlling tumor upon adoptive transfer. However, adoptive transfer of Pmel cells mixed with Pmel/Trx1 cells in 1:1 ratio markedly improved tumor control as compared to the mice that were transferred Pmel or Pmel/Trx T cells alone. It is likely that Pmel immediate effectors and Pmel/Trx cells with T-stem cell memory phenotype (i.e. CD62LhiCD44loCD122+Sca1hi), contributed to the improved tumor control. Thus, strategies to increase anti-oxidant capacity of anti-tumor T cells could have immunotherapeutic implications.