Induction of Tertiary Lymphoid Structures With Antitumor Function by a Lymph Node-Derived Stromal Cell Line

Tertiary lymphoid structures (TLSs) associate with better prognosis in certain cancer types, but their underlying formation and immunological benefit remain to be determined. We established a mouse model of TLSs to study their contribution to antitumor immunity. Because the stroma in lymph nodes (sLN) participates in architectural support, lymphogenesis, and lymphocyte recruitment, we hypothesized that TLSs can be created by sLN. We selected a sLN line with fibroblast morphology that expressed sLN surface markers and lymphoid chemokines. The subcutaneous injection of the sLN line successfully induced TLSs that attracted infiltration of host immune cell subsets. Injection of MC38 tumor lysate-pulsed dendritic cells activated TLS-residing lymphocytes to demonstrate specific cytotoxicity. The presence of TLSs suppressed MC38 tumor growth in vivo by improving antitumor activity of tumor-infiltrating lymphocytes with downregulated immune checkpoint proteins (PD-1 and Tim-3). Future engineering of sLN lines may allow for further enhancements of TLS functions and immune cell compositions

Cardiomyocyte Specific Ablation of p53 Is Not Sufficient to Block Doxorubicin Induced Cardiac Fibrosis and Associated Cytoskeletal Changes

Doxorubicin (Dox) is an anthracycline used to effectively treat several forms of cancer. Unfortunately, the use of Dox is limited due to its association with cardiovascular complications which are manifested as acute and chronic cardiotoxicity. The pathophysiological mechanism of Dox induced cardiotoxicity appears to involve increased expression of the tumor suppressor protein p53 in cardiomyocytes, followed by cellular apoptosis. It is not known whether downregulation of p53 expression in cardiomyocytes would result in decreased rates of myocardial fibrosis which occurs in response to cardiomyocyte loss. Further, it is not known whether Dox can induce perivascular necrosis and associated fibrosis in the heart. In this study we measured the effects of acute Dox treatment on myocardial and perivascular apoptosis and fibrosis in a conditional knockout (CKO) mouse model system which harbours inactive p53 alleles specifically in cardiomyocytes. CKO mice treated with a single dose of Dox (20 mg/kg), did not display lower levels of myocardial apoptosis or reactive oxygen and nitrogen species (ROS/RNS) compared to control mice with intact p53 alleles. Interestingly, CKO mice also displayed higher levels of interstitial and perivascular fibrosis compared to controls 3 or 7 days after Dox treatment. Additionally, the decrease in levels of the microtubule protein α-tubulin, which occurs in response to Dox treatment, was not prevented in CKO mice. Overall, these results indicate that selective loss of p53 in cardiomyocytes is not sufficient to prevent Dox induced myocardial ROS/RNS generation, apoptosis, interstitial fibrosis and perivascular fibrosis. Further, these results support a role for p53 independent apoptotic pathways leading to Dox induced myocardial damage and highlight the importance of vascular lesions in Dox induced cardiotoxicity

The Endoplasmic Reticulum Stress Response in Neuroprogressive Diseases: Emerging Pathophysiological Role and Translational Implications

The endoplasmic reticulum (ER) is the main cellular organelle involved in protein synthesis, assembly and secretion. Accumulating evidence shows that across several neurodegenerative and neuroprogressive diseases, ER stress ensues, which is accompanied by over-activation of the unfolded protein response (UPR). Although the UPR could initially serve adaptive purposes in conditions associated with higher cellular demands and after exposure to a range of pathophysiological insults, over time the UPR may become detrimental, thus contributing to neuroprogression. Herein, we propose that immune-inflammatory, neuro-oxidative, neuro-nitrosative, as well as mitochondrial pathways may reciprocally interact with aberrations in UPR pathways. Furthermore, ER stress may contribute to a deregulation in calcium homoeostasis. The common denominator of these pathways is a decrease in neuronal resilience, synaptic dysfunction and even cell death. This review also discusses how mechanisms related to ER stress could be explored as a source for novel therapeutic targets for neurodegenerative and neuroprogressive diseases. The design of randomised controlled trials testing compounds that target aberrant UPR-related pathways within the emerging framework of precision psychiatry is warranted

Abstract 5417: Expansion of effector regulatory T-cells represents a novel and independent prognostic factor marking escape from immune surveillance in Myleodysplastic Syndrome

Abstract Myelodysplastic syndromes (MDS) refer to a group of pathophysiologically diverse hematopoietic neoplasms associated with cytopenias, myeloid dysplasia, and variable acute myeloid leukemia risk. Response to immunosuppressive therapies (IST) in a subset of younger patients classified as lower-risk by the International Prognostic Scoring System (IPSS) suggests that inflammation and immune reactivity in the bone marrow likely plays a role in early MDS pathogenesis. Conversely, higher-risk patients with more established disease do not respond to IST. Previous reports of increased regulatory T-cells (Tregs) in these patients suggest that immune escape may be required for progression to higher-risk disease. The distinct changes in the role of the immune system between lower and higher-risk disease suggests that MDS may progress according to established principles of cancer immuno-editing involving elimination, equilibrium and escape phases. Because Treg suppressive activity is dependant upon auto-antigen presentation, we hypothesized that analyzing the activation state of Tregs may better reflect the onset of tumor-induced immune suppression in MDS and have better prognostic utility than measuring total Treg numbers alone. Here, we use a panel of surface markers on Tregs that are normally employed to distinguish effector and memory populations among conventional T-cells and analyzed the resulting subpopulations for associations with clinical features including overall survival (OS) among patients predominantly classified as lower-risk. Abnormal numbers of Treg subtypes were seen in 18 (34.6%) of 52 patients compared to age-matched controls. The most prominent and unique change in MDS patients was an expansion of effector Tregs (TregEff) which was significantly associated with anemia (p=0.046) and reduced hemoglobin (p=0.038). This group of patients had increased blast percentage (p=0.006) and displayed worse OS by Cox-regression (HR 4.3, 95% CI 1.6-11.6, p=0.004) independent of IPSS (p=0.002) and the MD Anderson Scoring System (MDAS) (p=0.047). Increased TregEff numbers were not dependant on total Treg numbers. Rather, they came at the expense of central memory Tregs suggesting that Treg “class switching” may reflect active presentation of auto-antigens. When isolated, TregEff cells displayed increased suppressive capacity compared to other Treg subpopulations and heightened TregEff numbers were limited to patients with elevated blasts (tumor burden), suggesting a mechanistic link between immune-evasion and MDS progression. The MDAS significantly refined OS estimates (p<0.001) compared to IPSS. Using the MDAS, OS in higher-risk patients could be further stratified using TregEff numbers (p=0.018), suggesting that enumeration of TregEff cells may improve prognostication and impact therapeutic selection in MDS. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 5417. doi:1538-7445.AM2012-541

Expansion of Effector Memory Regulatory T Cells Represents a Novel Prognostic Factor in Lower Risk Myelodysplastic Syndrome

Myelodysplastic syndromes are premalignant diseases characterized by cytopenias, myeloid dysplasia, immune dysregulation with association to autoimmunity, and variable risk for acute myeloid leukemia transformation. Studies of FOXP3(+) regulatory T cells (Tregs) indicate that the number and/or activation state may influence cancer progression in these patients. Focusing on patients with a lower risk for leukemia transformation, 18 (34.6%) of 52 patients studied displayed an altered Treg compartment compared with age-matched controls. Delineation of unique Treg subsets revealed that an increase in the absolute number of CD4(+)FOXP3(+)CD25(+)CD127(low)CD45RA(-)CD27(-) Tregs (effector memory Tregs [Treg(EM)]) was significantly associated with anemia (p = 0.046), reduced hemoglobin (p = 0.038), and blast counts >= 5% (p = 0.006). In healthy donors, this Treg(EM) population constitutes only 2% of all Tregs (one to six Tregs per microliter) in peripheral blood but, when isolated, exhibit greater suppressive activity in vitro. With a median follow-up of 3.1 y (range 2.7-4.9 y) from sample acquisition, increased numbers of Treg(EM) cells proved to have independent prognostic importance in survival estimates, suggesting that enumeration of this Treg subset may be a more reliable indicator of immunological escape than FOXP3(+) T cells as a whole. Based on multivariate analyses, Treg(EM) impacted survival independently from myeloblast characteristics, cytopenias, karyotype, and comorbidities. Based on these findings, Treg(EM) cell expansion may be synonymous with human Treg activation and indicate microenvironmental changes conducive to transformation in myelodysplastic syndromes. The Journal of Immunology, 2012, 189:3198-3208