Keizo Koya

Ganetespib, a unique triazolone-containing Hsp90 inhibitor, exhibits potent antitumor activity and a superior safety profile for cancer therapy

Targeted inhibition of the molecular chaperone Hsp90 results in the simultaneous blockade of multiple oncogenic signaling pathways and has, thus, emerged as an attractive strategy for the development of novel cancer therapeutics. Ganetespib (formerly known as STA-9090) is a unique resorcinolic triazolone inhibitor of Hsp90 that is currently in clinical trials for a number of human cancers. In the present study, we showed that ganetespib exhibits potent in vitro cytotoxicity in a range of solid and hematologic tumor cell lines, including those that express mutated kinases that confer resistance to small-molecule tyrosine kinase inhibitors. Ganetespib treatment rapidly induced the degradation of known Hsp90 client proteins, displayed superior potency to the ansamycin inhibitor 17-allylamino-17-demethoxygeldanamycin (17-AAG), and exhibited sustained activity even with short exposure times. In vivo, ganetespib showed potent antitumor efficacy in solid and hematologic xenograft models of oncogene addiction, as evidenced by significant growth inhibition and/or regressions. Notably, evaluation of the microregional activity of ganetespib in tumor xenografts showed that ganetespib was efficiently distributed throughout tumor tissue, including hypoxic regions >150 μm from the microvasculature, to inhibit proliferation and induce apoptosis. Importantly, ganetespib showed no evidence of cardiac or liver toxicity. Taken together, this preclinical activity profile indicates that ganetespib may have broad application for a variety of human malignancies, and with select mechanistic and safety advantages over other first- and second-generation Hsp90 inhibitors. Mol Cancer Ther; 11(2); 475–84. ©2011 AACR.

Multifaceted intervention by the Hsp90 inhibitor ganetespib (STA-9090) in cancer cells with activated JAK/STAT signaling.

There is accumulating evidence that dysregulated JAK signaling occurs in a wide variety of cancer types. In particular, mutations in JAK2 can result in the constitutive activation of STAT transcription factors and lead to oncogenic growth. JAK kinases are established Hsp90 client proteins and here we show that the novel small molecule Hsp90 inhibitor ganetespib (formerly STA-9090) exhibits potent in vitro and in vivo activity in a range of solid and hematological tumor cells that are dependent on JAK2 activity for growth and survival. Of note, ganetespib treatment results in sustained depletion of JAK2, including the constitutively active JAK2V617F mutant, with subsequent loss of STAT activity and reduced STAT-target gene expression. In contrast, treatment with the pan-JAK inhibitor P6 results in only transient effects on these processes. Further differentiating these modes of intervention, RNA and protein expression studies show that ganetespib additionally modulates cell cycle regulatory proteins, while P6 does not. The concomitant impact of ganetespib on both cell growth and cell division signaling translates to potent antitumor efficacy in mouse models of xenografts and disseminated JAK/STAT-driven leukemia. Overall, our findings support Hsp90 inhibition as a novel therapeutic approach for combating diseases dependent on JAK/STAT signaling, with the multimodal action of ganetespib demonstrating advantages over JAK-specific inhibitors.

Synergistic activity of the Hsp90 inhibitor ganetespib with taxanes in non-small cell lung cancer models

SummarySystemic chemotherapy using two-drug platinum-based regimens for the treatment of advanced stage non-small cell lung cancer (NSCLC) has largely reached a plateau of effectiveness. Accordingly, efforts to improve survival and quality of life outcomes have more recently focused on the use of molecularly targeted agents, either alone or in combination with standard of care therapies such as taxanes. The molecular chaperone heat shock protein 90 (Hsp90) represents an attractive candidate for therapeutic intervention, as its inhibition results in the simultaneous blockade of multiple oncogenic signaling cascades. Ganetespib is a non-ansamycin inhibitor of Hsp90 currently under clinical evaluation in a number of human malignancies, including NSCLC. Here we show that ganetespib potentiates the cytotoxic activity of the taxanes paclitaxel and docetaxel in NSCLC models. The combination of ganetespib with paclitaxel, docetaxel or another microtubule-targeted agent vincristine resulted in synergistic antiproliferative effects in the H1975 cell line in vitro. These benefits translated to improved efficacy in H1975 xenografts in vivo, with significantly enhanced tumor growth inhibition observed in combination with paclitaxel and tumor regressions seen with docetaxel. Notably, concurrent exposure to ganetespib and docetaxel improved antitumor activity in 5 of 6 NSCLC xenograft models examined. Our data suggest that the improved therapeutic indices are likely to be mechanistically multifactorial, including loss of pro-survival signaling and direct cell cycle effects resulting from Hsp90 modulation by ganetespib. Taken together, these findings provide preclinical evidence for the use of this combination to treat patients with advanced NSCLC.

Indole- and indolizine-glyoxylamides displaying cytotoxicity against multidrug resistant cancer cell lines

We report herein the SAR studies of a series of indole- and indolizine-glyoxylamides that demonstrate substantial in vitro anti-proliferative activities against cancer cell lines, including multidrug resistance (MDR) phenotypes. The in vitro cytotoxic effects have been demonstrated across a wide array of tumor types of various origins (e.g., breast, colon, uterine).

The oncology drug elesclomol selectively transports copper to the mitochondria to induce oxidative stress in cancer cells.

Elesclomol is an investigational drug that exerts potent anticancer activity through the elevation of reactive oxygen species (ROS) levels and is currently under clinical evaluation as a novel anticancer therapeutic. Here we report the first description of selective mitochondrial ROS induction by elesclomol in cancer cells based on the unique physicochemical properties of the compound. Elesclomol preferentially chelates copper (Cu) outside of cells and enters as elesclomol-Cu(II). The elesclomol-Cu(II) complex then rapidly and selectively transports the copper to mitochondria. In this organelle Cu(II) is reduced to Cu(I), followed by subsequent ROS generation. Upon dissociation from the complex, elesclomol is effluxed from cells and repeats shuttling elesclomol-Cu complexes from the extracellular to the intracellular compartments, leading to continued copper accumulation within mitochondria. An optimal range of redox potentials exhibited by copper chelates of elesclomol and its analogs correlated with the elevation of mitochondrial Cu(I) levels and cytotoxic activity, suggesting that redox reduction of the copper triggers mitochondrial ROS induction. Importantly the mitochondrial selectivity exhibited by elesclomol is a distinct characteristic of the compound that is not shared by other chelators, including disulfiram. Together these findings highlight a unique mechanism of action with important implications for cancer therapy.

A rat retinal damage model predicts for potential clinical visual disturbances induced by Hsp90 inhibitors

In human trials certain heat shock protein 90 (Hsp90) inhibitors, including 17-DMAG and NVP-AUY922, have caused visual disorders indicative of retinal dysfunction; others such as 17-AAG and ganetespib have not. To understand these safety profile differences we evaluated histopathological changes and exposure profiles of four Hsp90 inhibitors, with or without clinical reports of adverse ocular effects, using a rat retinal model. Retinal morphology, Hsp70 expression (a surrogate marker of Hsp90 inhibition), apoptotic induction and pharmacokinetic drug exposure analysis were examined in rats treated with the ansamycins 17-DMAG and 17-AAG, or with the second-generation compounds NVP-AUY922 and ganetespib. Both 17-DMAG and NVP-AUY922 induced strong yet restricted retinal Hsp70 up-regulation and promoted marked photoreceptor cell death 24h after the final dose. In contrast, neither 17-AAG nor ganetespib elicited photoreceptor injury. When the relationship between drug distribution and photoreceptor degeneration was examined, 17-DMAG and NVP-AUY922 showed substantial retinal accumulation, with high retina/plasma (R/P) ratios and slow elimination rates, such that 51% of 17-DMAG and 65% of NVP-AUY922 present at 30 min post-injection were retained in the retina 6h post-dose. For 17-AAG and ganetespib, retinal elimination was rapid (90% and 70% of drugs eliminated from the retina at 6h, respectively) which correlated with lower R/P ratios. These findings indicate that prolonged inhibition of Hsp90 activity in the eye results in photoreceptor cell death. Moreover, the results suggest that the retina/plasma exposure ratio and retinal elimination rate profiles of Hsp90 inhibitors, irrespective of their chemical class, may predict for ocular toxicity potential.

Pharmacokinetic analysis and antitumor efficacy of MKT-077, a novel antitumor agent

Abstract MKT-077 (1-ethyl-2-{[3-ethyl-5-(3-methylbenzothiazolin-2-yliden)]-4-oxothiazolidin-2-ylidenemethyl} pyridinium chloride), a novel rhodacyanine dye in phase I/II clinical trials, may provide a new approach to cancer therapy based on the accumulation in the mitochondria of the cells of certain carcinomas, for example, those of the colon, breast and pancreas. To support the development of MKT-077 for clinical application as an intravenous (i.v.) therapy, we investigated the metabolic fate of [14C]MKT-077 in BDF1 mice as well as the distribution of MKT-077 in experimental LS174T tumor-bearing mice using a high-performance liquid chromatography (HPLC) method. The plasma levels of 14C after i.v. administration of [14C]MKT-077 declined in a triphasic manner. In the first distribution phase, the levels of 14C decreased with a T1/2 of ∼5 min. In the second and terminal phase, the T1/2 of 14C was 2.8–4.6 h and 16.2 h, respectively. Cmax (1 min after injection) increased from 0.3 to 1.5 μg/ml linearly, but less than proportionately between the doses. The AUC(0–∞) at 0.3, 1 and 3 mg/kg were 0.030 ± 0.002, 0.60 ± 0.12 and 1.73 ± 0.25 μg · h/ml, respectively. Plasma clearance was ∼1.8 l/h per kg (at doses of 1 and 3 mg/kg). The steady state volume of distribution (6.8 and 25.1 l/kg) indicated that MKT-077 distributed as a lipid-soluble molecule. The mean residence time (MRT) was 4.1 (at a dose of 1 mg/kg) and 14.1 h (at a dose of 3 mg/kg). In the first rapid phase (5 min after dosing), 14C radioactivity was detected in most of the tissues and organs, most strongly in the kidney cortex, and not in the central nervous system and testes. In the terminal phase (24 h after dosing), 14C contents increased in the intestinal tract, and in the kidney and liver were nearly to the background level. After i.v. bolus administration at a dose of 3 mg/kg of [14C]MKT-077, the predominant route of elimination of the radioactivity was via the feces, and recoveries of total radioactivity in urine and feces corresponded to 33.5% and 61.1%, respectively. More than 60% was recovered within 24 h and 95% within 1 week. MKT-077 was primarily excreted in unmetabolized form with five unidentified metabolites found in the urine and plasma. Intact MKT-077 was retained in the tumor tissue longer than in plasma and kidney in LS174T tumor-bearing mice receiving MKT-077 at an i.v. therapeutic dose (10 mg/kg). This accumulation decreased very slowly, suggesting that the high membrane potentials of tumor cell mitochondria may help retain the drug in tumors.

Novel indolizine compounds as potent inhibitors of phosphodiesterase IV (PDE4): structure–activity relationship

A series of novel indolizine 2-oxoacetamides were designed and synthesized as PDE4 inhibitors. Preliminary SAR of this new class of compounds revealed key structural features required for high potency. Compounds 1ab and 2a are among the most potent inhibitors of PDE4 with low single nM IC50. Cellular activity was demonstrated by the inhibition of TNFα production from human PBMC with IC50 ranging from 14 to 72 nM. Docking analyses suggest the OH group in 1ab enhance the binding via an H-bond interaction with the PDE4 enzyme.

Abstract C212: A critical role for the tissue distribution profile in heat shock protein (Hsp) 90 inhibitor-induced ocular toxicity in rats.

In addition to regulating a number of oncogenic client proteins, the Hsp90 molecular chaperone also controls the folding of key signaling molecules required to maintain normal cell function in many organs, including the retina. In human clinical trials Hsp90 inhibition has been associated with visual disorders including blurred vision, flashes, delayed light/dark accommodation, and photophobia. These adverse effects involving injury to the retina may be attributable to photoreceptor degeneration and cell death, as previously reported in dogs following repeated doses of PF-04929113. In contrast, ganetespib, a potent Hsp90 inhibitor currently in phase II/III trials, has demonstrated promising clinical activity without manifesting ocular toxicity. This difference in vision deficits between ganetespib and other Hsp90 inhibitors likely depends on a number of contributing factors. In this study, we examined the relationship between retinal drug distribution profiles and photoreceptor degeneration in male SD rats treated with17-DMAG, 17-AAG, and STA-9056 (an Hsp90 inhibitor with comparable in vitro activity to 17-DMAG). All compounds were tested in short-term studies at 1–3 dose levels administered i.v., At necropsy, eyes were dissected and processed for histopathological examination. In subsets of animals, the retinal tissues, along with plasma and cerebrospinal fluid (CSF) samples, were collected for analysis. Our results indicate that all compounds evaluated showed greater exposure in the retinal tissue compared to plasma and CSF. 17-DMAG, for which visual changes have been reported in clinical subjects, produced marked photoreceptor cell death and was associated with a slow elimination rate (at 6 hrs post-dose, 50% of the drug present at 30 min remained in the retina) and a high retina/plasma (R/P) ratio. In contrast, and consistent with the absence of clinically-reported visual changes, 17-AAG at the maximum tolerated dose did not produce detectable photoreceptor injury. At 6 hr post dose, 94% of 17-AAG had been eliminated from the retina resulting in a low R/P ratio. Finally, STA-9056 showed 79% drug elimination at 6 hrs and an R/P ratio that was moderately low. Photoreceptor degeneration was not observed at doses of STA-9056 that are active in animal tumor models, and only minimal degeneration was seen at a higher dose. Our findings suggest that the R/P exposure ratio and elimination rate profiles play crucial roles in ocular toxicity and can be used as indicators of potential Hsp90 inhibitor-induced damage in rats. In summary, Hsp90 plays an important role in the retina and prolonged Hsp90 inhibition can lead to vision disorders. However, ocular toxicity may be successfully minimized by administration of Hsp90 inhibitors with favorable drug properties that include, although not necessarily limited to, lower retina/plasma exposure ratios and faster retinal elimination. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2011 Nov 12-16; San Francisco, CA. Philadelphia (PA): AACR; Mol Cancer Ther 2011;10(11 Suppl):Abstract nr C212.

Synthesis, crystallographic characterization and electrochemical property of a copper(II) complex of the anticancer agent elesclomol.

Elesclomol is a novel anticancer agent that has been evaluated in a number of late stage clinical trials. A new and convenient synthesis of elesclomol and its copper complex is described. X-ray crystallographic characterization and the electrochemical properties of the elesclomol copper(II) complex are discussed. The copper(II) cation is coordinated in a highly distorted square-planar geometry to each of the sulphur and amide nitrogen atoms of elesclomol. Electrochemical measurements demonstrate that the complex undergoes a reversible one-electron reduction at biologically accessible potentials. In contrast the free elesclomol is found electrochemically inactive. This evidence is in strong support of the mechanism of action we proposed for the anticancer activity of elesclomol.