# Carbazole Derivatives as STAT Inhibitors: An Overview
## 3 STATs Inhibitors
### Carbazoles as STAT Inhibitors
Since position 3 of carbazole is close to the side chain of Arg595, they explored the C-3 position to ameliorate the binding to the protein. Therefore, in light of the simple introduction of a nitro group able to interact  
Figure 12: 1,4-Dimethyl-carbazole derivatives (**17–26**). with Arg595, compounds **23** and **24** have been designed. Furthermore, compounds **25** and **26** attempted to raise the contacts with STAT3 through van der Waals interactions by introducing alkyl groups on the nitrogen to the detriment of the hydrogen bond. At the end, the designed compounds **18-26** were docked into the binding cavity of STAT3. The docked poses of **18-26** accommodated well into the protein binding cavity and seem to respect the predicted interactions [115]. Since JAK and STAT proteins (as previously stated) are connected to a pathway that involves many cytokines and hormones for intracellular signaling with potential involvement in cancer, [PERSON] et al. [116] designed and synthesized a series of 9\(H\)-carbazole-1-carboxamides (Figure 13) in order to evaluate their potentiality as selective ATP-competitive inhibitors of Janus kinase 2 (JAK2). By means of in silico studies, they tried to optimize their lead compound **27**, showing a good JAK2 inhibitory potency but poor selectivity toward JAK family. The (S)-dimethylamino-pyrrolidine amide **28** exhibited an enhanced selectivity against other members of the JAK kinase family. Moreover, **28** was tested in vitro for its antiproliferative effect on SET-2 cells (adult acute megakaryoblastic leukemia) reaching an IC\({}_{50}\) value of 80 nM. Further studies were aimed to assess its in vitro ADME properties. Data suggested that **28** possessed a greater solubility than compound **27** (>1.9 mg/mL at pH 1.0; 0.41 mg/mL at pH 6.5; compared to <0.001 mg/mL across pH range for 1). Another compound with a significant selectivity on JAKs was **29**, particularly active against JAK2 (IC\({}_{50}\) amounting to 5.5 nM), which also displayed an antiproliferative activity against SET-2 cells with an IC\({}_{50}\) value amounting to 130 nM. Concerning the metabolic stability of **29**, it is similar to **28**, except its solubility at acidic pH was enhanced when compared to the initial screening (0.044 mg/mL at pH 1.0) [108]. Based on the knowledge that phosphorylation of STAT3 at Y705 induces the dimerization and translocation into the nucleus to relay the oncogenic signals by expressing the genes implicated in proliferation, antiapoptosis, angiogenesis and tumor evasion in 2016, [PERSON] et al. [117] reported the synthesis and evaluation of substituted carbazole derivatives (Figure 14) using nano-cuprous oxide as a catalyst via intramolecular C-N bond forming reactions. Among them, (3-acetyl-6-chloro-9\(H\)-carbazol-9-yl)methyl)-[1,10-biphenyl]-2-carbonitrile (**30**) exhibited the greater antiproliferative activity towards two lung cancer cell lines: A549 and LLC, establishing itself as a lead compound. The calculated IC\({}_{50}\) values amounted to 13.6 and 16.4 \(\upmu\)M, respectively for the two cell lines. Figure 13: Structure of 9\(H\)-carbazole-1-carboxamides (27–29). Paclitaxel, used as a reference molecule, instead achieved IC\({}_{50}\) of 0.0044 uM against A549 cells. Furthermore, they performed their studies on A549, HCC-2279 and H1975 cells, treating them with 10 uM of ACB for 6 h to examine the effect on the persistent activation of STAT3 by Western blot analysis using antibodies that recognize phosphorylation of STAT3 at Tyr-705. They found that **30** inhibited STAT-3 phosphorylation in A549, HCC-2279 and H1975 cells at a concentration of 10 uM. However, **30** did not affect total STAT3 and -actin levels, confirming the inhibitory effects of carbazoles on STAT3 phosphorylation. Moreover, they analyzed the levels of phospho-STAT3 and lysine demethylylase (LSD1) in the nuclear extract of HCC-2279 cells, at different concentrations of **30** (0, 1.25, 2.5, 5, 10, 20, 40 and 80 uM).