Tubulin-based antimitotic mechanism of E7974, a novel analogue of the marine sponge natural product hemiasterlin

Abstract

E7974 is a synthetic analogue of the marine sponge natural product hemiasterlin. Here, we show that E7974, such as parental hemiasterlin, acts via a tubulin-based antimitotic mechanism. E7974 inhibits polymerization of purified tubulin in vitro with IC 50 values similar to those of vinblastine. In cultured human cancer cells, E7974 induces G 2 -M arrest and marked disruption of mitotic spindle formation characteristic of tubulin-targeted anticancer drugs. Extensive hypodiploid cell populations are seen in E7974-treated cells, indicating initiation of apoptosis after prolonged G 2 -M blockage. Consistent with this observation, E7974 induces caspase-3 activation and poly ADP ribose polymerase cleavage, typical biochemical markers of apoptosis. Only a short cellular exposure to E7974 is sufficient to induce maximum mitotic arrest, suggesting that E7974's antitumor effects in vivo may persist even after blood levels of the drug decrease after drug administration. Interactions of E7974 with purified tubulin were investigated using two synthetic tritiated photoaffinity analogues incorporating a benzophenone photoaffinity moiety at two different positions of the E7974 scaffold. Both analogues preferentially photolabeled α-tubulin, although minor binding to β-tubulin was also detected. E7974 thus seems to share a unique, predominantly α-tubulin-targeted mechanism with other hemiasterlin-based compounds, suggesting that, unlike many tubulin-targeted natural products and related drugs, the hemiasterlins evolved to mainly target α-tubulin, not β-tubulin subunits.

Marine Pharmaceuticals: A New Wave of Anti-angiogenic Drugs

“…It had reached the phase II clinical trials but was discontinued due to lack of objective responses along with observance of toxicities in patients [28]. In 2009, another synthetic hemiasterlins analogue E7974 was found which showed strong in vivo anti-tumour efficacy in many human xenograft cancer models along with decreased drug resistance in cancer cells [29]. It has cleared the phase I clinical trial for the treatment of advanced solid tumours in colorectal, pancreatic and liposarcoma cancer patients and is currently undergoing human clinical trial [30].…”

A phase 1 trial of E7974 administered on day 1 of a 21‐day cycle in patients with advanced solid tumors

“…8,[11][12][13] E7974 induces marked disruption of mitotic spindle formation and a G2/M arrest characteristic of tubulin-acting anticancer drugs. 14,15 The mitotic block induced by E7974 is relatively irreversible. Extensive hypodiploid cell populations are detected in E7974-treated cells, indicating apoptosis after prolonged G2/M blockage.…”

“…Consistent with this observation, E7974 induces caspase-3 activation and poly-ADP-ribose polymerase cleavage, both characteristic biochemical markers of apoptosis. 14,[16][17][18][19][20][21][22][23][24] Its unique mechanism of interaction with the tubulin molecule has been shown to overcome drug resistance by at least 2 different mechanisms. First, unlike taxanes and vincas, E7974 is a poor substrate for the P-glycoprotein drug efflux pump, a key contributor to both intrinsic and acquired multidrug resistance.…”

“…First, unlike taxanes and vincas, E7974 is a poor substrate for the P-glycoprotein drug efflux pump, a key contributor to both intrinsic and acquired multidrug resistance. 14 Second, unlike paclitaxel and vinblastine, E7974 binds predominantly to the a subunit, with minor binding to the b subunit. 25 Resistance to both paclitaxel and vinblastine is based on mutations in b-tubulin.…”

“…Moreover, in several CRC models, E7974 showed superior efficacy to standard therapies for CRC (oxaliplatin, 5-fluorouracil, and irinotecan). 14,[27][28][29][30] Significant preclinical in vivo activity was also seen in models of breast cancer, pancreatic cancer, and melanoma. [31][32][33][34][35][36][37][38] E7974 thus offers significant promise of activity in taxane-resistant tumor types, regardless of whether the mechanism driving resistance is based on P-glycoprotein or tubulin mutations.…”