Our present data clearly demonstrate that the overexpression of Mcl-1 in coordination with Bcl-2/Bcl-xL expression protects cancer cells from apoptosis. Mitochondria are the main ATP producers in cells and are therefore essential for all cellular processes. Furthermore, mitochondria play a pivotal role in life or death decisions in the cell by regulating the apoptosis pathway. The release of cytochrome C from mitochondria leading to the activation of caspases is a hallmark of the apoptotic response. Concomitantly, resistance to apoptosis can arise from a reduction in mitochondrial outer membrane permeabilization. Akt kinase, autophagy, and elevated Bcl-xL and Mcl-1 can cooperate to protect tumor cells against chemotherapy-induced apoptosis by maintaining mitochondrial stability
[24, 25]. The NIH Developmental Therapeutics Program has determined that Bcl-xL may play a unique role in the general resistance of cancer cells to cytotoxic agents by showing that a variety of cancer cell lines that demonstrate resistance to 70,000 cytotoxic agents are characterized by high Bcl-xL expression
. Mcl-1 overexpression has also been reported to contribute to chemoresistance in multiple tumors
[15, 27] and, notably, has been implicated in the chemoresistance of certain types of malignancies to the first of a new class of Bcl-2-family targeting compounds, ABT-737
Because of the overexpression and overlapping functions of the Bcl-2 family proteins, it will be important to develop an inhibitor of both Bcl-2/xL and Mcl-1. It has been shown previously that either Mcl-1 downregulation or NOXA overexpression, an Mcl-1 specific BH3-only protein, strongly sensitizes melanoma cells to ABT-737 in vitro
. Hence, developing BH3 mimetics could be a feasible approach to inhibit Mcl-1 function. Unfortunately, none of the BH3 mimetics under current development are potent and specific Mcl-1 antagonists
. Indeed, many “pan- Bcl2 inhibitors” suffer from a lack of specificity or are simply too weak to compete with native high-affinity BH3-only proteins for pro-survival BH3 binding pockets. Further, such pan-Bcl2 family protein inhibitors might well damage normal tissues. Hence, BH3 mimetics specific for single pro-survival targets could have greater clinical utility
. Pertinently, GDC-0199, a novel BH3 mimetic developed by Abbott and Genentech that is specific for Bcl-2, and which is now entering clinical trials for lymphoid malignancies, should avoid the dose-limiting thrombocytopenia associated with the navitoclax
. For these reasons, designing an Mcl-1 specific inhibitor or searching for alternative targets for Mcl-1 antagonism has become “popular”.
Our current research suggests that USP9X regulates Mcl-1 expression in cancer cells. Deubiquitinases have been demonstrated previously to antagonize specific oncogenic and tumor suppressive E3-ligases and are viewed as emerging targets for cancer therapeutics
. USP9X can now be added to this list due to its role in deubiquitination and in stabilizing Mcl-1, a bona fide oncogene. In our current analyses, USP9X expression was found to be strongly associated with Mcl-1 expression in the human cancer tissue samples we tested. Recent reports have suggested also that USP9X enhances Mcl-1 stability by preventing its proteasomal destruction through de-ubiquitination
. The balance between ubiquitination and deubiquitination determines Mcl-1 stability and expression. Ubiquitination of Mcl-1 promotes USP9X-Mcl-1 binding leading to Mcl-1 deubiquitination and disassociation of these two proteins. Hence, and as shown from our current data, increasing Mcl-1 ubiquitination via PS341 promotes the association of USP9X with Mcl-1. Since Mcl-1 proteins are constantly ubiquitinated, their association with USP9X appears to be a “steady-state” condition. This activity and upregulation of USP9X as well as Mcl-1 have been associated with a poor prognosis and with chemoresistance in a number of cancers. To determine the impact of USP9X inhibition on cancer cell survival in our present experiments, we used its inhibitor WP1130 and found that the treated cells showed Mcl-1 downregulation which increased their sensitivity to ABT-737 as well as to other chemotherapeutic agents. In light of the importance of USP9X in the control of Mcl-1 levels, compounds such as WP1130 or other more specific inhibitors may be useful in overcoming the apoptotic resistance associated with USP9X activity and Mcl-1 protection. WP1130 may therefore have utility as a chemosensitizer in a combinational chemotherapy regimen as it can inhibit several USPs including USP9X, USP5, USP14, and UCH37, which are known to regulate cell survival, protein stability, and 26S proteasomal function
. Furthermore, USP9X is a deubiquitinase that targets multiple proteins involved in cell growth and survival
[35, 36]. Hence, the design of a specific inhibitor that targets the USP9X and Mcl-1 interaction could also be a viable and possibly even a better approach to reducing the impact of chemoresistance in different tumors.