ID: 2251

• Title:
  The immune response in electroporation-based therapies may be controlled by pulse characteristics
  
  Authors:
  

  Polajžer, Tamara - PT
  Miklavcic, Damijan - MD

  
  
  Abstract:
  Background: Traditionally, electroporation-based therapies, such as gene electrotransfer (GET), electrochemotherapy (ECT), and irreversible electroporation (IRE), are performed with a typical pulse parameter. Millisecond-long pulses are used in GET and microsecond-long pulses with different number or amplitude are used in ECT and IRE. GET, ECT, and IRE have also been performed with high-frequency bipolar pulses known as HFIRE and nanosecond pulses, indicating that virtually any pulse duration (millisecond, microsecond, nanosecond) and any type of pulse (monopolar, bipolar) can be used for therapy. During therapy, an immune response can also be triggered. However, inflammation and activation of the immune system is not always desirable, as it may affect the outcome of the therapy. While the immune response is favorable during cancer treatment, it is undesirable during ablation of cardiac tissue and gene therapies because it can destroy therapeutic material. Overall, any electroporation-based therapy can be performed with different pulses, but it is not yet known how different pulse durations affect the activation of the immune response. Therefore, we investigated immunogenic response to different pulses by determining DAMP molecule release following electroporation in vitro.
  
  Methods: Activation of the immune response was determined by the detection of specific signaling molecules. These molecules are called damage-associated molecular patterns (DAMP molecules) and are recognized by the cells of the immune system, triggering an immune response. The best known DAMPs are ATP, HMGB1 and calreticulin. Five different pulses were used for electroporation: a) 5 ms, 8 pulses, 1 Hz, ΔV; b) 100 µs, 8 pulses, 1 Hz, ΔV; c) 2-2-2-2 µs, 32 pulses, 100 burst, 1 Hz (HFIRE), ΔV; d) 200 ns, 100 pulses, 10 Hz, ΔV; and e) 4 ns, ΔN, 500 Hz, 12 kV. After electroporation, ATP, HMGB1, and calreticulin were detected at different time points after electroporation.
  
  Results: All ATP is released into the extracellular space immediately after pulse delivery and is rapidly degraded because the signal is undetectable after one hour. The amount of released ATP increases with intensity of electroporation, i.e. increasing the amplitude and/or pulse number, with different pulses releasing different amounts of ATP. Interestingly, HMGB1 was detected only when 200-nanosecond pulses were used, while calreticulin was detected 24 hours after electroporation and only when high amplitude pulses were used, with all but the longest millisecond pulse used.
  
  Conclusion: DAMP release varies with pulse durations and electroporation intensity used. The most immunogenic pulses appear to be nanosecond pulses, and the least immunogenic are millisecond pulses. DAMP release also depends on the intensity of electroporation.
  
  This research was funded by Slovenian Research Agency and Medtronic
  
  
  
  Keywords:
  immune response, damage/danger-associated molecular pattern molecules, DAMPs, different pulse durations, different pulse types
  
  Refs:
  
  
  Topic 1:
  1. Biological responses (molecular, subcellular, cellular and intercellular)
  
  Topic 2:
  6. Cancer treatment and tumor ablation