## 4 Discussion  
For BNCT, a high concentration of boron within the cancer cells is mandatory as, theoretically, only those cells will be destroyed after neuron irradiation, leaving surrounding normal cells undamaged. However, even if trans-stimulation seems to increase boron uptake in HNSCC cells with increasing BPA concentrations, no distinct effect of trans-stimulation for the uptake of boron in HNSCC cells after BPA incubation in comparison to non-stimulated cells could be shown. For HNSCC, adjuvant radiation therapy and/or chemotherapy are recommended for advanced stages. Known adverse effects of radiation such as pain and xerostomia have a negative impact on speech, chewing and swallowing [22]. BNCT might reduce these side effects because of its principle to target tumour cells. Here, one major concern is that boron uptake in tumour cells has to be increased, as reported disadvantages of BNCT include inhomogeneous BPA distribution in the tumour and an insufficient coverage of the clinical target volume [23]. The expression of LAT-1, which is proposed to be one of the most relevant amino acid transporters for BPA uptake, increases from normal to precursor lesions to tumour tissue, but seems to decrease for higher dedifferentiation levels of HNSCC [9]. Therefore, the HNSCC cell lines used in the present study are described as moderately and badly differentiated [21] and the presence of LAT-1 in both HNSCC cell lines was confirmed by Western blotting. The BPA uptake of HNSCC cells in this analysis was 1.10 times higher after trans-stimulation with a BPA concentration of 50 ppm when compared to the non-trans-stimulated samples, but without a significant difference. In the literature, the boron content of B16F1 melanoma cells was increased after trans-stimulation to 1.8-2.7 times higher [19] and in GS-9L gliosarcoma cells to approximately two times higher [17]. A reason for the non-existent effect of trans-stimulation in HNSCC cells could be the dedifferentiation of the tumour cells, which leads to a lower LAT-1 expression [9]. Overall, beneath the trans-stimulation effect, it is also possible that the lack of leucin, an amino acid similar to BPA, leads to an activation of system L [24; 25]. Subsequently, the observed minor increase in boron content \begin{table}
\begin{tabular}{c c c c c c} \hline \hline \multirow{2}{*}{**Trans-Stinulation**} & **BPA Concentration** & **Mean Boron Concentration** & & & \(p\) \\ \cline{3-6}  & **(ppm)** & **(ppp per One Million Cells)** & **SD** & **5 ppm vs.** & **5 ppm vs.** & **20 ppm vs.** \\  & & & **20 ppm** & **50 ppm** & **50 ppm** \\ \hline \multirow{3}{*}{no} & 5 & 50.31600 & 63.72422 & \multirow{3}{*}{1.000} & \multirow{3}{*}{0.002} \\  & 20 & 29.25684 & 13.29931 & & \\  & 50 & 48.92545 & 30.77410 & & \\ \cline{1-1}  & 5 & 51.18492 & 26.31162 & \multirow{3}{*}{0.000} & \multirow{3}{*}{0.0032} \\ \cline{1-1}  & 20 & 98.6308 & 49.07148 & & \\ \cline{1-1}  & 50 & 82.42652 & 43.24766 & & \\ \hline \hline \end{tabular}
\end{table} 
Table 2: Results of the Games-Howell post hoc test—significant differences were detected within the trans-stimulated groups. could be justified by this activation, because the trans-stimulation medium was free of amino acids except L-tyrosine. It is also remarkable that the overexpression of system L transporter LAT-1 in different cells such as hepatocytes and fibroblasts did lead to increased leucin uptake in hepatocytes but not in fibroblasts [26]. It appears likely that miscellaneous cell lines with their proteomes show different uptake behaviour. For example, in human hepatocellular carcinoma cells, no trans-stimulation effect was seen [27]. Likewise, melanoma cells showed no significantly increased boron content after trans-stimulation, and a lower boron uptake after 90 min of trans-stimulation was observed for adenocarcinoma cells and human hepatocellular carcinoma cells [28]. Most commonly, the concentration of the used L-tryrosine solution was 5 mmol/L, as this concentration is known to be non-toxic in vitro.