Purpose/Objectives: Small cell lung cancer (SCLC) is a highly aggressive disease representing 12-13% of all lung cancers, with 5 year survival rate of only 6%. While most patients respond initially to cytotoxic chemotherapies such as irinotecan, etoposide or carboplatin, resistance rapidly emerges and response to second line agents such as topotecan is limited, and in contrast to non-small cell lung cancer (NSCLC), few targetable oncogenes occur in SCLC. In this study, we tested new compound, STA-12-8666, which binds tumor-concentrated active form of shock protein 90 (HSP90) and has cleavable linker attached to SN-38, the active metabolite of irinotecan. Cleavage of the linker within the tumor provides time-release of SN-38 at high local concentration, while significantly limiting drug exposure and toxicity in non-transformed issue. The goal for this work was to evaluate STA-12-8666 for potential use as a new second line monotherapy, or as adjuvant in the frontline setting. Materials/Methods: Three dose levels of STA-12-8666 were evaluated in comparison to irinotecan, ganetespib, carboplatin, etoposide, and chemotherapy combinations in 4 independent SCLC xenograft models, including parental and cisplatin-resistant derivative cell lines (SCLC1, SR2), and a patient-derived xenograft (PDX). STA-12-8666 was also evaluated in drug combinations. Intratumoral responses were profiled using a mass spectrometry based approach to evaluate kinase pathway activation, and results confirmed by immunohistochemistry and western blot analysis. Pharmacokinetic analysis was performed to benchmark retention of STA-12-8666 to isomolar irinotecan in lung tumors. Results: In all three models, high dose (150 mg/kg) STA-12-8666 was well tolerated. In most cases, three doses administered at weekly intervals caused complete regression of established tumors, with response durable for > 2 months. Those tumors that regrew were responsive to re-dosing with STA-12-8666, and were subsequently eliminated. STA-12-8666 was also effective in limiting or eliminating tumors growth that had progressed following initial treatment on standard first and second line agents for SCLC. Low dose (50 mg/kg) STA-12-8666 controlled but did not eliminate tumors: however, it strongly enhanced the action of 30 mg/kg carboplatin, resulting in tumor elimination. Pharmacokinetic and proteomic analysis confirmed STA-12-8666 concentration in tumors, and identified a signature of DNA damage response biomarkers in STA-12-8666-treated tumors that strongly contrasted with the pattern induced by irinotecan. Conclusions: Together, these results indicate that STA-12-8666 may be a promising therapy in both frontline and second line settings for SCLC and strongly support the evaluation of this compound in Phase I/II clinical trials. Note: This abstract was not presented at the meeting. Citation Format: Anna Gaponova, AS Nikonova,A Deneka,BL Egleston,S Litwin,JS Duncan,K Duncan,H Borghaei,R Mehra,DA Proia,Y Boumber, Erica Golemis. Preclinical testing demonstrates strong activity of STA-12-8666, an HSP90 inhibitor-SN-38 conjugate, in small cell lung cancer (SCLC). [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1731. doi:10.1158/1538-7445.AM2015-1731

Background: Few promising molecular targets exist for chemoresistant triple negative breast cancer (TNBC). Our laboratory has previously demonstrated that a subset of TNBCs express high glucocorticoid receptor (GR) levels. In breast epithelial cells, GR transcriptional activity directly upregulates the expression of several anti-apoptotic target genes including serum/glucocorticoid regulated kinase 1 (SGK1). High GR and/or SGK1 expression are both associated with decreased chemotherapy-induced cell death in TNBC models. A meta-analysis of patients with early-stage TNBC found that high primary tumor GR (NR3C1) expression significantly associates with earlier relapse compared to GR-non expressing TNBC. Inhibiting GR/SGK1 activity is therefore a testable strategy for improving TNBC responsiveness to chemotherapy. Both GR and SGK1 require Hsp90 for their activity; therefore, Hsp90 inhibition is predicted to disrupt both GR and SGK1 cell survival signaling in TNBC. Employing a second-generation Hsp90 inhibitor, ganetespib (Synta Pharmaceuticals), we determined the effectiveness of Hsp90 inhibition on GR/SGK1 activity and chemotherapy sensitivity using in vitro and in vivo TNBC models. Hypothesis: Hsp90 inhibition will decrease GR and SGK1 activity in TNBC in association with improved chemotherapy effectiveness. Results: Using TNBC cell lines treated with ganetespib at clinically relevant concentrations, we found that total GR and the relative proportion of phosphorylated Ser211 GR were significantly reduced beginning 2 hours following treatment. Reduced glucocorticoid-induced (SGK1) mRNA expression was also seen when cells were pre-treated with ganetespib. We next used fluorescent microscopy and live cell imaging to measure TNBC cell death over a 96 hour time course following chemotherapy +/- ganetespib therapy. Addition of ganetespib to paclitaxel treatment of MDA-MB-231 cells led to significant in vitro cytotoxic synergy; as hypothesized, this synergy was diminished in GR-depleted MDA-MB-231 cells. Cytotoxic synergy following ganetespib and chemotherapy treatment was also seen in vivo in MDA-MB-231 xenografts. Interestingly, tumor GR expression following Hsp90 inhibitor and paclitaxel treatment was significantly reduced compared to tumors from mice treated with paclitaxel alone. Using GR-depleted or control MDA-MB-231 xenografts, studies are currently underway to further characterize the role of GR/SGK1 activity as a target of Hsp90 inhibition to reverse TNBC chemoresistance. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr P4-16-05.

Background: Ganetespib is a fully synthetic and selective inhibitor of heat shock protein 90 (HSP90), a molecular chaperone recognized as a key facilitator of breast cancer initiation, progression and metastasis. Methods: Preclinical activity of ganetespib across the four major breast cancer subtypes and inflammatory breast cancer was assessed in vitro and in vivo. Modulation of cell proliferation and viability was determined both in monolayer and three-dimensional cultures. HSP90 client protein expression and activity was monitored by Western blot and protein array. To recapitulate clinical dosing, kinetics of client protein destabilization were measured following short exposures to drug in vitro. Anticancer activity of ganetespib was further investigated in vivo using breast cancer xenografts. Results: Ganetespib displayed potent, low nanomolar activity in luminal (A and B), basal (A and B) and inflammatory breast cancer cell lines grown as monolayers in vitro. BT-474 (HER2 amplified) luminal cells grown as mammospheres in 3D were equally as sensitive to ganetespib as those grown in monolayer. In luminal cells, ganetespib simultaneously disrupted multiple signaling components including the estrogen and progesterone receptor, several receptor and non-receptor tyrosine kinases, as well as the MAPK pathway. Further, ganetespib effectively inhibited AKT, PDK1 and SGK3 activity in PIK3CA mutant cells suggesting that HSP90 is essential for both AKT-dependent and AKT-independent signaling. Clinically relevant exposure times to ganetespib in vitro resulted in potent, long term destabilization of HER2. In the basal-like breast cancer cell line MDA-MB-231, enriched in CD44+CD24- stem like cells that commonly display chemotherapeutic resistance and activated JAK2/STAT3 signaling, ganetespib (50 nM) induced significant degradation of JAK2 concordant with loss of both tyrosine and serine phosphorylation of STAT3, followed by cell death. The potent anticancer activity in vitro translated in vivo, where ganetespib was effective in modulating breast cancer xenograft growth as a single agent in both luminal and basal-like breast cancer models. Finally, ganetespib has demonstrated encouraging signs of clinical activity in breast cancer patients, including confirmed partial responses in both a triple negative breast cancer patient and a HER2 positive breast cancer patient. Conclusions: Ganetespib is a highly potent HSP90 inhibitor that displays preclinical activity in breast cancer due to its ability to simultaneously perturb multiple oncogenic signaling pathways. Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P3-17-05.

Background: Hsp90 is a molecular chaperone protein required for the stabilization and activation of many proteins, referred to as Hsp90 ‘clients’, such as HER2, HIF1-a, EGFR, ER, PI3K, AKT, P53 and VEGFR. The drug candidate, ganetespib is a novel triazolone inhibitor of Hsp90, with over 700 patients treated to date. Ganetespib has shown activity in preclinical models of HER2+, ER+/PR+ and TNBC. Early clinical trials documented ganetespib single agent activity in heavily pretreated HER2+ and TNBC patients. Ganetespib has been well tolerated in clinical trials with a favorable safety profile. This efficacy-screening study is designed to provide further evidence of ganetespib activity and identify potentially predictive biomarkers in metastatic breast cancer (BC). Methods: The ENCHANT-1 Trial is an international, first-line 2-cohort Phase 2 study in BC patients: Cohort A, HER2 amplified (n = 35), and Cohort B, TNBC (n = 35). Patients who present with previously untreated metastatic disease are eligible for treatment with ganetespib at 150 mg/m2 twice weekly on 3 out of 4 wks, for a total of up to 12 wks. Primary endpoint: ORR assessed using RECIST1.1 criteria. Key secondary endpoints include metabolic response as assessed by PET/CT at wk 3 utilizing modified EORTC criteria. Disease progression (PD) at wk 3 by PET imaging indicates discontinuation of study therapy, and is performed to quickly offer patients with metabolic PD a standard of care treatment. The study is designed as Simon 2-stage requiring at least one OR in 15 patients for the respective cohort to expand to 35 patients. A Steering Committee is established to oversee the overall study and review the interim results. Results: The study was initiated in 23 centers globally. At the time of submission, a total of 17 patients had been enrolled; TNBC (n = 15) and HER2 (n = 2). Here we report the interim analysis in the TNBC cohort. The median age was 54 years (range 30 -77) with ECOG PS 0 (n = 7/15). Most patients (n = 9) presented with de novo metastatic disease. 5 patients were not evaluable for PET assessment (3 had not yet reached wk 3 and 2 withdrawn before wk 3 for clinical progression), and 9 patients were not evaluable for objective response at wk 6 (3 withdrawn before or at wk 3 for clinical progression and 6 had not yet reached wk 6 evaluation). In the 10 patients with evaluable PET imaging, 9 patients achieved metabolic (m) response (2 mPR, 4 mSD with dominant tumor shrinkage and 3 SD) and one patient with mPD. In the 6 patients evaluable for OR at wk 6, one patient achieved PR, 2 SD and 3 PD. Treatment with ganetespib was well tolerated; the most common AEs were mild or moderate diarrhea (8/15, 53%), fatigue (5/15, 33%), decreased appetite (4/15, 27%), insomnia (4/15, 27%), and nausea (4/15, 27%). Conclusion: Ganetespib single agent was generally well tolerated and showed anti-tumor activity TNBC patients as early as 3 weeks following treatment. PET seems to be a good tool to screen antitumor activity of new agents in early settings rather that in heavily pretreated patients. The TNBC cohort has met the protocol criteria for proceeding to stage 2. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr P2-16-23.

Background: While the incidence of triple negative breast cancer (TNBC) is only 10-20%, these tumors show a disproportionate mortality for breast cancer patients. Due to a lack of effective molecular targets in this disease, therapeutic options are largely limited to systemic chemotherapeutic approaches which have shown disappointing efficacy in the metastatic setting. Here we undertook a comprehensive evaluation of the activity of the drug candidate, ganetespib, a potent inhibitor of heat shock protein 90 (Hsp90), in this malignancy. Methods: The sensitivity of TNBC cells to ganetespib was determined in viability assays using a panel of tumor lines. The effects of ganetespib exposure on client proteins and their effector pathways were examined by Western blot and reverse phase protein array analysis. The anti-metastatic activity of ganetespib was evaluated using a 4T1 metastasis model. Combinatorial drug analyses were performed with chemotherapeutic agents. DNA damage and cell cycle disruption were assessed using the comet assay, Western blotting and fluorescence microscopy. The in vivo efficacy of the compound, both as a single agent and in combination, was established using MDA-MB-231 xenograft models. Computed tomography scans were obtained for a metastatic TNBC patient undergoing ganetespib treatment. Results: Ganetespib reduced cell viability in TNBC cell lines with low nanomolar potency. Ganetespib treatment induced robust destabilization of multiple client proteins and oncogenic signaling pathways and suppressed lung metastasis in the experimental model. Ganetespib potentiated the cytotoxic activity of doxorubicin via enhanced DNA damage and mitotic arrest, conferring superior efficacy to the doxorubicin + cyclophosphamide (AC) regimen in MDA-MB-231 xenograft models. Ganetespib also promoted mitotic catastrophe and apoptosis in combination with taxanes in vitro, and these effects translated to significantly improved combinatorial activity in vivo. Marked tumor shrinkage of metastatic lung lesions was seen in the patient while on ganetespib monotherapy. Conclusion: The preclinical activity profile and clinical evidence of tumor regression suggest that ganetespib offers considerable promise as a new therapeutic candidate to target TNBC. In particular, the capacity of ganetespib to potentiate the activity of standard of care chemotherapeutics provides a rationale for the exploration of this agent in novel combinatorial treatment strategies for this disease. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr PD5-3.