ID: 2234

• Title:
  Distinct Tumor Immune Responses to Nanopulse Stimulation Determine Immunity
  
  Authors:
  

  Beebe, Stephen J - Frank Reidy Research Center for Bioelectrics
  Nanajian, Anthony - Frank Reidy Research Center for Bioelectrics
  Ruedlinger, Brittney - Frank Reidy Research Center for Bioelectrics
  Asadipour, Kamal - Frank Reidy Research Center for Bioelectrics and Department of Electrical and Computer Engineering 
  Guo, Siqi - Frank Reidy Research Center for Bioelectrics

  
  
  Abstract:
  Background and Objectives: Nanopulse stimulation (NPS) induces immune-mediated vaccine effects (in situ vaccination) after ablation of orthotopic rat liver (75-80%) and mouse breast tumors (80-95%). These studies are designed to determine immune mechanism in cancer models that do or do not readily induce immunity with in situ vaccination.
  
  Methods: At various NPS post-treatment days, the remaining tumor microenvironment (TME), draining lymph nodes, spleen, and blood are analyzed for immune cell characteristics and functions.
  
  Results: 4T1-luc breast cancer immunome mechanisms showed that NPS selectively targeted activated T-regulatory cells (Tregs, TGFβ+, 4-1BB+) by apoptosis with a decrease in activated/naïve T-reg ratio from 2.0 (day 0, control) to 0.5 (day 3 post-treatment). NPS also rapidly eliminated tumor-associated macrophages (TAMs, CD11b+ F4/80+) by apoptosis through days 1-7. Surprisingly, NPS reduced the apoptosis of myeloid-derived suppressor cells (MDSC, CD11b+ Gr1+), which nevertheless showed a delayed decrease of quantity at post-treatment days 3 to 7. These changes relieved immunosuppression in the TME. There was also a stronger NPS bias for apoptotic reduction of suppressive Tregs vs. T-effector cells leading to a 2.7-fold increase in the ratio of resident memory CD8+ CD103+ T-cells to CD4 Tregs. This increased ratio coincided with significant increases in CD11c+ dendritic cell (DC) numbers expressing costimulatory receptors (CD80+ CD86+ and MHC II+). These findings define immunome changes that empower immunity and identify fragile Tregs and other immunosuppressor cells as therapeutic NPS targets in the 4T1-luc model.
  
  Ongoing B16f10 melanoma studies with conditions that induce immune responses in orthotopic rat liver and mouse breast cancer models, NPS show that only about 25-30 % of NPS-treated mice were in situ vaccinated. Like the 4T1-luc tumors, TAM and MDSCs were increased after NPS on day1. There were increases in TME DCs expressing costimulatory receptors and increases in M1 and APCs indicted the the first step of antigen presentation in the immune response but an immune response is likely blocked due to an overwhelming induction of TAM&MDSCs. CD8+ cells increased in the TME, but central and effector memory cell numbers were low, and many were PD1+, suggesting a more anergic than an activated TME. In addition, MDSC and TAMs were increased again on day 7. Unlike immunity in the liver rat model, TME NK cell responses were meager in the B16f10 model. Overall, these responses allude to a narrow potential for immunity in the B16f10 melanoma model in response to NPS. These studies require functional analyses for the activated TAMs, a possible means to eliminate them, and determination of mechanisms of regulated cell death in the TME of both tumor models.
  
  Preliminary studies indicate that NPS-treated tumors co-treated with carbon nanotubes decreased the charging effect required for effective tumor elimination by at least 2-fold, but had no effect on immunity. In contrast NPS-tumors treated with losartan provided mice that were in situ vaccinated. Losartan, which is an angiotensin II receptor (ARIIR) inhibitor used to control blood pressure, also acts on monocyte recruitment receptors CxC2Rs, which inhibit monocyte migration into inflammatory sites, suggesting that losartan may have acted in an ARIIR-independent manner, acting on the CXC2R prevented the return of immunosuppressive cells to the TME allowing an environment for T-cells to inhibit tumor growth. Studies to test this hypothesis are in progress.
  
  Conclusions: To allow NPS to act as an immunotherapy and in situ vaccinate mice against the treated tumor, tumor elimination must be followed by an exclusion of immunosuppressive cells and inclusion of activated T-cells into the TME, like that shown in the rat liver cancer and 4T1-luc breast cancer models.
  
  
  
  Keywords:
  
  
  Refs:
  Lassiter BP, Guo S, Beebe SJ. Nano-Pulse Stimulation Ablates Orthotopic Rat Hepatocellular Carcinoma and Induces Innate and Adaptive Memory Immune Mechanisms that Prevent Recurrence. Cancers (Basel). 2018 Mar 13;10(3):69. Guo S, Burcus NI, Hornef J, Jing Y, Jiang C, Heller R, Beebe SJ. Nano-Pulse Stimulation for the Treatment of Pancreatic Cancer and the Changes in Immune Profile. Cancers (Basel). 2018 Jun 27;10(7):217. Beebe SJ, Lassiter BP, Guo S. Nanopulse Stimulation (NPS) Induces Tumor Ablation and Immunity in Orthotopic 4T1 Mouse Breast Cancer: A Review. Cancers (Basel). 2018 Mar 30;10(4):97.
  
  Topic 1:
  6. Cancer treatment and tumor ablation
  
  Topic 2:
  1. Biological responses (molecular, subcellular, cellular and intercellular)