The best results were obtained using a fivefold molar excess of benzimidazole with respect to quinobenzothiazinium salts 2. It may be assumed that the other reaction product are benzimidazolium salts 5, the structure of which can be stabilized via delocalization
SBE-��-CD cell line of positive charge among the benzimidazole nitrogen atoms. Scheme. 3 Synthesis of compounds 4 Benzimidazolium salts 5 were neither isolated from the reaction mixture nor identified in the course of this study, as the primary objective here was to obtain quinobenzothiazine 4 derivatives as free quinoline bases. Excess benzimidazole and benzimidazolium salts 5 that form during the reaction were separated from quinobenzothiazines 4 by pouring post-reaction mixtures into water. Both benzimidazole and salts 5 are well-soluble in water, whereas
compounds 4 fall out of solution as solids. In order to obtain quinobenzothiazine derivatives 7 containing aminoalkyl substituents at the thiazine nitrogen atom, compounds 4 were transformed, in the presence of sodium hydroxide, buy LY411575 into salts 6, which were then alkylated using aminoalkyl chlorides (Scheme 4). The reaction occurred as N-alkylation at the thiazine nitrogen atom and led to compounds 7. The structure of compounds 7 was confirmed with 1H NMR spectroscopy by performing NOE 1H–1H Epacadostat price homonuclear experiment. By irradiating methylene group protons at the thiazine nitrogen atom an enhancement of H1 and H11 proton signals from compounds 7 was obtained (Scheme 5). Scheme. 4 Synthesis of compounds 7 Scheme. 5 NOE 1H–1H homonuclear experiment for compound 7a Antiproliferative activity The activity of the obtained compounds 4 and 7 was investigated in vitro using cultured SNB-19 and C-32 cell lines and cisplatin as a reference. The examined quinobenzothiazines 4 had various substituents (CH3, F, Cl, Br) introduced into 9- and 11-positions of the quinobenzothiazine ring. In Dipeptidyl peptidase addition, they also contain a nitrogen atom in the 8-position
of the quinobenzothiazine ring. Compounds 7 contains aminoalkyl substituents: 2-(N-piperidyl)ethyl (compounds 7(a–d)) and 3-(N,N-dimethylamino)propyl (compound 7e) at the thiazine nitrogen atom. One of the mechanisms involved in antiproliferative effects of chemotherapeutics is DNA intercalation. This mode of action is typical for antiproliferative anthracycline antibiotics (e.g., doxorubicin) that feature planar tetracyclic (aromatic or heteroaromatic) fused rings. This mode of action, affecting cancer cells’ DNA, has been indeed suggested in reports concerning antiproliferative properties of phenothiazine and benzo[a]phenothiazine derivatives (Motohashi et al., 2000; Hossain et al., 2008; Hossain and Kumar, 2009). Structurally, compounds 4 and 7 studied herein are their analogs. The experiments demonstrated that the majority of the investigated compounds 4 and 7 showed antiproliferative activity toward examined cell lines within the 5.6–12.