The most interesting finding from the present study was that the small molecule tyrosine kinase inhibitor Sorafenib reverses DHA-induced suppression of nuclear Bach1 expression, thereby attenuating HO-1 induction by DHA. Consequentially, the combination of DHA and Sorafenib led to a synergistic interaction in suppressing cancer cell viability. Since Sorafenib is a well-established anticancer drug [
40], and DHA is a dietary compound that possesses great health benefits and is also used clinically [
7‐
9], our findings indicate that the combination of Sorafenib and DHA is an attractive new strategy for more effective cancer therapy.
DHA’s anticancer activity and its mechanisms of action have been extensively investigated over the last several decades [
11,
41,
42]. Although the combination of long chain n-3 PUFA, including DHA, and chemotherapeutics for cancer therapy has been described in recent years in various experimental model systems [
30,
41,
42] and in clinical trials [
11], the potential mechanisms of how DHA might interact with other therapeutics to achieve high therapeutic efficacy remains poorly understood. This lack of understanding limits the further development of combination therapies using DHA and other anticancer drugs. We have previously reported that DHA induces apoptosis of cancer cells primarily through enhanced lipid peroxidation [
13], indicating the importance of the cellular antioxidant enzyme system in mediating DHA’s anticancer action. It is well established that cancer cells are more vulnerable to oxidative stress than normal cells. This is thought to be due to a constant increase in cellular levels of reactive oxygen species and an already stressed antioxidant response, thus providing a strategy to selectively kill cancer cells by further enhancing cellular oxidative stress [
43‐
45]. In our recent report, we demonstrated that DHA induces HO-1 gene transcription by promoting Bach1 protein nuclear exportation and degradation. The loss of Bach1 protein allowed an increase in Nrf2 binding to the AREs in the HO-1 gene promoter and activation of HO-1 gene transcription [
23]. HO-1 is an antioxidant enzyme that is coupled by the Nrf2 signaling pathway [
19], and has been shown to be over-expressed in cancer tissues [
22] and contribute to chemo resistance [
46,
47]. Thus targeting HO-1 is a potential strategy for cancer therapy [
21,
22]. Because Bach1 protein nuclear exportation and subsequent degradation is controlled by tyrosine phosphorylation [
26], we assumed that tyrosine kinase inhibitors would reverse DHA-induced Bach1 degradation and block HO-1 induction by DHA, thus leading to enhanced cytotoxicity upon cancer cells. The results from the present study confirmed our hypothesis by testing the combination of the tyrosine kinase inhibitor Sorafenib and DHA in our model systems. Both western blot and reporter gene assay revealed that Sorafenib is able to reverse DHA-induced suppression of nuclear Bach1 expression and induction of HO-1 gene transcription. Furthermore, Sorafenib was found to enhance DHA’s cytotoxicity in various human cancer cell lines and further suppressed xenograft tumor growth in vivo in fish oil fed mice. The combined drug interaction of DHA and Sorafenib was synergistic as evidenced by Isobologram analysis. These findings clearly indicate that targeting Bach1/Nrf2-mediated HO-1 gene expression enhances DHA’s cytotoxicity. This study supports further development of a new combination cancer therapy using DHA and Sorafenib, both being well tolerated in patients and approved by the FDA for clinical applications.
It should be noted that Sorafenib has been described as an inhibitor to the soluble epoxide hydrolase, an enzyme that functions in converting active lipid epoxides to inactive diols [
48]. The possibility that Sorafenib inhibition of the soluble epoxide hydrolase results in more active DHA-derived epoxydocosapentaenoic acids thereby enhancing DHA’s cytotoxicity in our model system cannot be excluded. It is indeed plausible that the combination of DHA and Sorafenib could weaken the cellular antioxidant forces (targeting Bach1/Nrf2-mediated HO-1 gene expression) on the one hand and potentially enhance the oxidative potential (inhibiting soluble epoxide hydrolase to elevate DHA-derived epoxydocosapentaenoic acids) on the other hand, thereby leading to synergistic cytotoxicity against cancer cells. The fact that Sorafenib is a multi tyrosine kinase inhibitor [
49] suggests that future studies could reveal even more mechanistic insight in the synergistic anticancer action of DHA and Sorafenib.
While DHA has been shown to enhance the anticancer effect of various cancer therapies, including chemotherapy [
11] and radiotherapy [
50,
51], it remains unclear whether DHA universally enhances the efficacy of all anticancer drugs, or whether it may antagonize the anticancer effect of certain chemotherapeutics. By determining the combined effect of DHA and the 101 anticancer compounds in the Oncology Drug Set IV, the present study for the first time has shown that DHA selectively enhances the cytotoxicity of certain cancer therapeutics, while antagonizing or having little to no effect on the cytotoxicity of other anticancer compounds. These results underline the importance of better understanding the mechanism of action when DHA is used in combination with other cancer therapeutics. In this regard, the present study has raised a critical issue in DHA-based combination therapy: that not every anticancer drug is suitable in combination with DHA for cancer therapy and drugs should be individually evaluated in the appropriate model system if they are intended to be used in combination with DHA for cancer therapy.