## Results and Discussion
### Pharmacological Studies  
In order to screen the cancer cell-specific antiproliferative effect of the synthesized D-ring-fused steroidal 5-aminopyrazoles (**6a-h**), various human cancerous cell lines, namely MCF-7, PC-3, A549, HeLa and U2 Os cells and MRC-5 noncancerous fibroblasts were treated with the test compounds in 5 uM concentration for 72 h. A heat map (Figure 3) was constructed based on the obtained viability data (Supplementary Material, Table S1). It was found that compound **6g** showed the most prominent antiproliferative activity and proved to be efficient on all the tested cancer cells. The other compounds exerted minor or fair cytotoxic effects and their cancer-specific antiproliferative features were much less pronounced than that of **6g**. Based on the primary screen, compound **6g** was selected from the compound library. Then its cytotoxicity was evaluated more precisely on all aforementioned noncancerous and cancerous cell lines. The IC\({}_{50}\) values of **6g**, especially those obtained on PC-3, HeLa, and U2 Os cells are notably lower compared to that on MRC-5 noncancerous fibroblasts, despite falling in the same order of magnitude (Table 3, Supplementary Material, Figure S1a). Among all cancer cell lines, A549 showed the lowest, whereas U2 Os displayed the highest sensitivity to **6g** treatment. As a positive control, the same set of cell lines was treated with the 'gold standard' cisplatin for 72 h, and the IC\({}_{50}\) values were determined likewise (Table 3, Supplementary Material, Figure S1b). As expected, cisplatin exhibited strong antiproliferative activity at fairly low concentrations on the tested cell types, whereas compound **6g** was effective at concentrations in the uM range. It is well-known that a number of cancer cells exhibit resistance towards cisplatin treatment. In line with previous reports [32], we also observed that PC-3 cell lines displayed resistance to the applied cisplatin, indicated by the high IC\({}_{50}\) value. Interestingly, compound **6g** effectively inhibited the proliferation of PC-3 cells, and the IC\({}_{50}\) value of compound **6g** obtained on this particular cell line was comparable to its IC\({}_{50}\) values on other tested cancer cells. No resistance toward molecule **6g** could be observed upon our experiments. \begin{table}
\begin{tabular}{c c c c c c c} \hline \hline
**Entry** & **Compd.** & **R** & **pK\({}_{\text{a}}\)** & **\(\lambda_{\text{max}}\) (nm) of HL\({}^{+}\)** & **\(\lambda_{\text{max}}\) (nm) of L** & **logD\({}_{7.4}\) (Predicted)** \\ \hline
1 & **6a** & H & 3.97 \(\pm\) 0.03 & 243 & 249 & +4.31 \\
2 & **6b** & CH\({}_{3}\) & 3.37 \(\pm\) 0.03 & 243 & 252 & +4.77 \\
3 & **6c** & OMe & 3.78 \(\pm\) 0.06 & 240 & 244 & +4.05 \\
4 & **6d** & F & 3.56 \(\pm\) 0.04 & 242 & 2212 1 & +4.45 \\
5 & **6e** & Cl & 2.00 \(\pm\) 0.03 & 245 & - 1 & +4.83 \\
6 & **6f** & Br & 2.07 \(\pm\) 0.06 & 247 & - 1 & +5.10 \\
7 & **6g** & CN & \(<\)1.8 & \(\sim\)260 & \(\sim\)276 & +4.12 \\
8 & **6h** & NO\({}_{2}\) & 2.13 \(\pm\) 0.09 & 248, 289 & 236, 334 & +4.26 \\ \hline \hline \end{tabular}  
* The spectrum of L has no characteristic absorption band in the studied wavelength range.  
\end{table} 
Table 2: Proton dissociation constants (pK\({}_{\text{a}}\)) of the compounds determined by UV-visible titrations, \(\lambda_{\text{max}}\) values (nm) of the ligand species in HL\({}^{+}\) and L forms and logD\({}_{7.4}\) (= logP) values predicted by the MarvinSketch software [31]. {t = 25 °C; I = 0.1 M (KCI); 1% (_v/v_) EtOH/water}. Apart from cancer cell resistance, the sensitivity of noncancerous cells can also cause complications. In fact, noncancerous MRC-5 cells were more sensitive to cisplatin treatment than cancerous cells. This was clearly demonstrated by the IC\({}_{50}\) value for cisplatin on fibroblasts, being one magnitude smaller (37 nM) than any of its IC\({}_{50}\) values on cancerous cells.