Simultaneous targeting of COX-2 and AKT using selenocoxib-1-GSH to inhibit melanoma

Abstract

Melanoma is a highly metastatic and deadly disease. An agent simultaneously targeting the COX-2, PI3K/Akt, and mitogen-activated protein kinase (MAPK) signaling pathways that are deregulated in up to 70% of sporadic melanomas might be an effective treatment, but no agent of this type exists. To develop a single drug inhibiting COX-2 and PI3K/Akt signaling (and increasing MAPK pathway activity to inhibitory levels as a result of Akt inhibition), a selenium-containing glutathione (GSH) analogue of celecoxib, called selenocoxib-1-GSH was synthesized. It killed melanoma cells with an average IC(50) of 7.66 μmol/L compared with control celecoxib at 55.6 μmol/L. The IC(50) range for normal cells was 36.3 to 41.2 μmol/L compared with 7.66 μmol/L for cancer cells. Selenocoxib-1-GSH reduced development of xenografted tumor by approximately 70% with negligible toxicity by targeting COX-2, like celecoxib, and having novel inhibitory properties by acting as a PI3K/Akt inhibitor (and MAPK pathway activator to inhibitory levels due to Akt inhibition). The consequence of this inhibitory activity was an approximately 80% decrease in cultured cell proliferation and an approximately 200% increase in apoptosis following 24-hour treatment with 15.5 μmol/L of drug. Thus, this study details the development of selenocoxib-1-GSH, which is a nontoxic agent that targets the COX-2 and PI3K/Akt signaling pathways in melanomas to inhibit tumor development.

Figure 1. COX-2 expression increased in melanomas, regulated through the MAP kinase pathway

1A (left panel). Elevated levels of COX-2 expression in melanoma patient tumors and cell lines. 1A (right panel). COX-2 expression increased in a cell line based melanoma tumor progression model. 1B. Targeting COX-2 using siRNAs decreased melanoma cell viability. 1C. SiRNA-mediated inhibition of the MAP kinase pathway decreased COX-2 expression in melanomas. 1D (upper panel). PLX-4032 targeting of ^V600EB-Raf decreased COX-2 expression. 1D (bottom panel). U0126 targeting of Mek1/2 decreased COX-2 expression.

Figure 2. Development of selenocoxib-1-GSH

2A. Structure of celecoxib, selenocoxib-1, and selenocoxib-1-GSH. 2B. Selenocoxib-1-GSH kills cancer more effectively that normal cells. 2C. Selenocoxib-1-GSH retained COX-2 inhibitory activity.

Figure 3. Selenocoxib-1-GSH inhibited melanoma cell growth by reducing cellular proliferation, triggering apoptosis, and arresting melanoma cells in the G₀-G₁ phase of the cell cycle

3A, 3B & 3C. Selenocoxib-1-GSH, but not celecoxib, inhibited melanoma cell proliferation and induced apoptosis. 3D. Selenocoxib-1-GSH arrested melanoma cells in the G₀-G₁ phase of the cell cycle.

Figure 4. Selenocoxib-1-GSH inhibited Akt signaling to reduce the proliferative potential and promote apoptotic signaling in melanoma cells

4A. Selenocoxib-1-GSH inhibits the PI3K/Akt signaling pathway. 4B. Selenocoxib-1-GSH activates the MAPK signaling pathway. 4C. Selenocoxib-1-GSH decreased cyclin D1 protein levels indicating a reduction in cellular proliferation. 4D. Selenocoxib-1-GSH increased levels of cellular apoptosis.

Figure 5. Targeting COX-2 inhibited melanoma tumor development with negligible toxicity

5A. SiRNA-mediated reduction of COX-2 protein levels decreased melanoma tumor development in mice. 5B & 5C. Selenocoxib-1-GSH treatment decreased xenografted melanoma tumor development. No significant difference was observed in body weight of mice treated with the drug indicating negligible toxicity (Figs. 5B & 5C; inset). 5D. Selenocoxib-1-GSH does not affect blood biomarkers indicative of major organ related toxicity.