Histological and biochemical analysis of DNA damage after BNCT in rat model

To understand the mechanism of tumor cell death induced by boron neutron capture therapy (BNCT) and to optimize BNCT condition, we used rat tumor graft models and histological and biochemical analyses were carried out focusing on DNA damage response. Rat lymphosarcoma cells were grafted subcutaneously into male Wister rats. The rats with developed tumors were then treated with neutron beam irradiation 45 min after injection of 330 mg/kg bodyweight boronophenylalanine ((10)BPA) (+BPA) or saline control (-BPA). BNCT was carried out in the National Nuclear Center of the Republic of Kazakhstan (neutron flux: 1 x 10(9) nvt/s, fluence: 6 x 10(11) nvt) with the presence of background gamma-irradiation of 33 Gy. 6 and 20 h after BNCT treatment, tumors were resected, fixed and subjected to immunohistochemistry and biochemical analyses. Immunostaining of nuclei showed that double strand break (DSB) marker gamma H2AX staining was high in 20 h/+ BPA sample but not in 20 h/-BPA samples. Poly(ADP-ribose), DSB and single strand break markers of DNA, also demonstrated this tendency. These two markers were observed at low levels in unirradiated tissues or 6 h after BNCT either under BPA and +BPA conditions. HMGB1 level increased in 6 h/+BPA but not in 6 h/- BPA or 20 h/+ BPA samples. The persistent staining of gamma H2AX and poly(ADP-ribose) in +BPA group suggests accumulated DSB damage after BNCT. The early HMGB1 upregulation and gamma H2AX and poly(ADP-ribose) observed later might be the markers for monitoring the DNA damage induced by BNCT. (C) 2014 Elsevier Ltd. All rights reserved.