## 3 Results and Discussion
### Biological Part--Analysis of Cytotoxicity towards Cancer and Normal Cells  
Taking into account the results of the in silico simulations and the previously promising anticancer activities and low cytotoxicity of the obtained dipirydothiazines **1-4**[21] and dimers with a lutiidine linker **6a-9a**[25], the new derivatives **6-9** were tested for their anticancer activity. The cytotoxicity and antiproliferative activity of new dimers were determined in vitro in relation to the following five cancer cell lines: human primary colon cancer (SW480), human metastatic colon cancer (SW620), human breast adenocarcinoma (MDA-MB-231), human lung carcinoma (A-549), and human glioblastoma (LN-229), and to the normal line human immortal keratinocyte cell line from adult human skin (HaCaT) using the MTT assay [30]. The study used doxorubicin and cisplatin as reference drugs. The data are expressed as the mean SD, IC\({}_{50}\) (uM)--the half-maximal growth inhibitory concentration after culturing the cells for 72 h with the individual compound--and the SI (Selectivity Index), which was calculated using the following formula: SI = IC\({}_{50}\) for normal cell line/IC\({}_{50}\) cancer cell line. The results are summarized in Table 3, and show that compounds **6-9** were not toxic to the normal keratinocyte HaCaT cell line in the range of the tested concentration. The analysis of the anticancer activity showed that the tested derivatives were mostly inactive (IC\({}_{50}\) > 100 uM). Dimer **6** had poor activity in relation to SW480, MDA-MB-231, and A-549 cell lines (IC\({}_{50}\) = 81-92 uM). Dimer **7**, which contains two 1,8-dipyridylothiazine units in its structure, was the most active in the group of tested compounds in relation to the MDA-MB-231 cell line (IC\({}_{50}\) = 67.3 uM). The results of the  
\begin{table}
\begin{tabular}{c c c c c c} \hline \hline \multicolumn{2}{c}{**No.**} & \multicolumn{3}{c}{**Probability of Cytotoxicity towards Cancer Cell Lines**} \\ \hline
**6** & Breast cancer & Melanoma & Non-small-cell & Colon carcinoma & Pancreatic \\ MDA-MB-468 (62\%) & UACC-257 (51\%) & Lung carcinoma & HCT-116 (42\%) & carcinoma YAPC (43\%) \\ \hline
**7** & Breast cancer & Melanoma & Renal carcinoma & Colon carcinoma & Pancreatic \\ MDA-MB-468 (59\%) & UACC-257 (55\%) & 786 (45\%) & HCT-116 (39\%) & carcinoma YAPC (44\%) \\ \hline
**8** & Breast cancer & Melanoma & Renal carcinoma & Colon carcinoma & Ovarian \\ MDA-MB-468 (39\%) & UACC-257 (54\%) & 786-0 (49\%) & HCT-116 (55\%) & adenocarcinoma \\  & & & & & \\
**9** & Breast cancer & Melanoma & Renal carcinoma & Colon carcinoma & Ovarian \\ MDA-MB-468 (44\%) & UACC-257 (49\%) & 786-0 (44\%) & HCT-116 (54\%) & VOC-4 (37\%) \\ \hline \hline \end{tabular}
\end{table}
Table 2: The probable cytotoxic effect (%) on various cancer cell lines of dimers (**6–9**).  
biological assay were very surprising when analyzed in relation to the previously obtained results of the anticancer activity of structural analogs **6a-9a**, as shown in Figure 1 [25].  
Ludidine derivatives **6a-9a** showed activity in the range of IC\({}_{50}\) = 0.1-11 uM towards MCF 7 and IC\({}_{50}\) = 4.5-25.9 uM towards the SW480 cell lines [25]. This led us to consider the reason for the deactivation of the obtained dimers **6-9**. To explain this phenomenon, quantum chemical calculations and preliminary molecular docking analyses were used.