Cancer chemopreventive effect of phenothiazines and related tri-heterocyclic analogues in the 12-O-tetradecanoylphorbol-13-acetate promoted Epstein-Barr virus early antigen activation and the mouse skin two-stage carcinogenesis models

In continuation of our search for novel agents, we have investigated 29 phenothiazines and related tri-heterocyclic compounds as potential cancer chemopreventive agents in a short-term in vitro assay of Epstein-Barr virus early antigen (EBV-EA) activation induced by the tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA). Among the evaluated compounds, chlorpromazine, phenoxazine, ethylpropazine, 9-oxo-9H-thioxanthene-3-carbonitrile-10,10-dioxide, thiothixene and phenothiazine showed profound inhibition of EBV-EA in the in vitro assay. This activity was influenced by a modification of the phenothiazine ring. Replacement of nitrogen in the phenothiazine ring with sulfur atoms decreased the anti-tumor activity. Overall analysis showed that the simple tri-cyclic compound phenoxazine was the most active anti-tumor promoting compound in the test system. Therefore, we assessed the anti-tumor promoting effect of phenoxazine in vivo in two different chemical carcinogen-induced-promotion experimental models in mice namely the 7,12-dimethylbenz(a)anthracene (DMBA) initiated and TPA-promoted ICR mouse skin two-stage carcinogenesis protocol and the peroxynitrite (PN)-induced and TPA-promoted skin carcinogenesis in HOS:HR-1 mouse. Following tumor initiation with DMBA, topical application of 0.0025% phenoxazine to the dorsal initiated mouse skin resulted in a highly significant inhibition of TPA tumor promotion. The compound exhibited remarkable inhibitory effects on the mouse skin tumor promotion in terms of a reduction in tumor multiplicity (>50%) and incidence, accompanied by an extension of the tumor latency. In the PN-induced and TPA-promoted two-stage mouse skin carcinogenesis, oral administration of phenoxazine (0.0025%) for 2 weeks showed profound decrease in both the tumor incidence and burden by more than 20 and 80%, respectively, at 10 weeks of treatment. This was also accompanied by a 20% delay in the tumor latency period. In all the treatment groups, there was no toxicity due to phenoxazine in the treatment groups as compared to the control animals. These significant anti-tumor potentials of phenoxazine either via topical or oral administration might be due to the inherent cytotoxicity of these classes of compounds, which can be utilized in the prevention of development of overt tumors, immunopotentiation, induction of differentiation and apoptosis. In addition, since phenoxazine derivatives and other related phenothiazine compounds in use, as anti-psychotic agents without any reported adverse effect are known to pass the blood-brain barrier, they represent a new class of cancer chemopreventive agents with greater implication in the prevention of brain cancers.