Staurosporine, an inhibitor of hormonally up-regulated neu-associated kinase

Abstract

HUNK is a protein kinase that is implicated in HER2-positive (HER2+) breast cancer progression and resistance to HER2 inhibitors. Though prior studies suggest there is therapeutic potential for targeting HUNK in HER2+ breast cancer, pharmacological agents that target HUNK are yet to be identified. A recent study showed that the broad-spectrum kinase inhibitor staurosporine binds to the HUNK catalytic domain, but the effect of staurosporine on HUNK enzymatic activity was not tested. We now show that staurosporine inhibits the kinase activity of a full length HUNK protein. Our findings further suggest that inhibiting HUNK with staurosporine has a strong effect on suppressing cell viability of HER2/neu mammary and breast cancer cells, which express high levels of HUNK protein and are dependent on HUNK for survival. Significantly, we use in vitro and in vivo methods to show that staurosporine synergizes with the HER2 inhibitor lapatinib to restore sensitivity toward HER2 inhibition in a HER2 inhibitor resistant breast cancer model. Collectively, these studies indicate that pharmacological inhibition of HUNK kinase activity has therapeutic potential for HER2+ breast cancers, including HER2+ breast cancers that have developed drug resistance.

HUNK phosphorylates EGFR to regulate breast cancer metastasis

“…In addition, we have previously shown that STU inhibited HUNK's kinase activity, and can therefore be used as an experimental tool to study HUNK activity [26]. We found that STU treatment of animals orthotopically transplanted with 4T1 cells reduced the phosphorylation of EGFR at T654 in primary mammary tumors, which corresponded to a decrease in EGFR expression and lung metastases.…”

“…Our data clearly shows that pharmacological inhibition of HUNK supports this mechanism, which also has not been previously determined. While a prior publication demonstrates that pharmacological targeting of HUNK can impede tumor growth, those studies were performed in a resistant HER2-positive (HER2+) breast cancer cell model and not a metastatic breast cancer model [26]. This point is important to note because the fundamental mechanism for HUNK in HER2+ breast cancer is different from metastasis since tumor growth is not affected by HUNK targeting in metastatic models [18].…”

“…The peptides were incubated with ATP either in the absence or in the presence of HUNK. We also used staurosporine (STU) as a pharmacological tool to inhibit HUNK kinase activity, since prior studies show STU to bind and inhibit HUNK [25,26]. The kinase reactions were dotted onto nitrocellulose membrane and blotted with a phospho-specific antibody for T654 EGFR (pT654).…”

“…Prior studies show STU to bind and inhibit HUNK [25,26]. Therefore, we performed a dose-response analysis of STU in 4T1 cells to test the effect on phosphorylation of EGFR at T654.…”

HUNK Signaling in Metastatic Breast Cancer

“…We took an approach to deplete Hunk/HUNK in these cell lines using shRNA targeting. We also used a pharmacological inhibitor of HUNK as a tool compound to modulate kinase activity [17]. In all three of these cell lines, HUNK depletion or inhibition led to an impairment of cell migration and invasion that was concomitant with a loss of EGFR phosphorylation at T654.…”