Therapeutic targets and novel anticancer agents for endocrine cancers

Division Of Basic Sciences - Nci

Investigators

Linked publications, trials & patents

Abstract

There are three main projects: I. Novel synergistic drug combination therapy for adrenocortical cancer (ACC) 1.1 Synergy of MELK and CDK inhibitors targeting multiple clinically-relevant molecules in ACC 1.2 Novel synergistic combination of PI3K and HSP90 inhibitors in ACC 1.3 Drug repurposing study identified auranofin as a novel radiosensitizer in ACC II. Targeted therapy focusing on the effects of TNF-a on tumor microenvironment to enhance drug delivery III. Quantitative high-throughput drug screening in BRAF V600E WT anaplastic thyroid cancer (ATC) 1.1 Synergy of MELK and CDK inhibitors targeting multiple clinically-relevant molecules in ACC Aim: To identify the effective novel synergistic drug combinations against ACC of OTS167 (MELK inhibitor) and RGB286638 (multi-CDK inhibitor) and to study the molecular mechanisms of synergy. Summary: The in vitro synergistic efficacy of OTS-167 and RGB286638 was confirmed by our laboratory. The combination effectively caused G2M cell cycle arrest, induced caspase-dependent apoptosis, inhibited b-catenin via FOXM1, and pSTMN1, resulting in increased depolymerized microtubule and decreased cell invasion. The additional data from the last report showed that the combination treatment downregulated cyclins A2, B1, and B2, which are the partnering proteins of CDKs 1 and 2 with similar prognostic significance in patients with ACC to MELK and CDKs 1 and 2. In vivo studies showed the anti-tumor efficacy of the combination treatment is higher than that of single-drug and control groups. The analyses of xenografts from mice in the combination treatment group confirmed the effects seen in vitro. 1.2 The novel synergistic combination of PI3K and HSP90 inhibitors in ACC Aim: To identify and validate the effective novel synergistic drug combinations using PI3K inhibitor and Heat-shock protein 90 inhibitors and to study the molecular mechanisms of synergy Summary: The most effective synergistic combinations identified by the computerized drug combination matrix screening in ACC cells are one of the three HSP90 inhibitors and PIK-75, a selective p110a inhibitor of PI3K. We analyzed publicly available databases and found that ACC overexpressed several isoforms of HSP90 and HSP-90 clients involving in ACC initiation and progression. We validated the synergy in two ACC cell lines in a cell proliferation assay. The ACC tumor spheroid assays showed increased efficacy in the combination treatment group as compared to the single-drug and control groups. Because PIK-75 is not yet available for a clinical trial, we studied clinically available GDC-0077 which selectively targets p110a isoform of PI3K in combination with STA9090 (HSP90 inhibitor). We did not find synergistic activity at clinically achievable doses. Next, we are testing GDC-0032 which is a new clinically available selective p110a isoform inhibitor of PI3K with a much higher potency than GDC-0077. We found that the combination treatments (colony formation, cell migration) and reduced PI3K pathway. 1.3 Drug repurposing study identified auranofin as a novel radiosensitizer in ACC Aim: To evaluate the effects and mechanism of action of auranofin in combination with radiotherapy (RT) in ACC preclinical models Summary: Despite a complete surgery, 50-80% of patients with ACC develop locoregional recurrence with or without distant metastasis. Mitotane is still the only FDA-approved systemic therapy for ACC, and it imparts no survival benefit but has significant toxicities. Thus, the role of adjuvant or neoadjuvant treatment using in mitotane, chemotherapy, and/or RT in ACC is not known. Because an effective treatment regimen is urgently needed to reduce ACC recurrence and the associated morbidity, we analyzed the data on a quantitative high-throughput drug screening in two ACC cell lines and identified auranofin as a novel radiosensitizer that also exhibited a potent antiproliferative effect in ACC. In addition to locoregional control when used with RT, we anticipate that auranofin can control the systemic disease at a clinically achievable dose. Auranofin was selected because of its high anti-neoplastic efficacy, compared to the current cytotoxic agents used in ACC (Figure 7), and radiosensitization effect by inhibiting TXNRD. Auranofin is an FDA-approved, orally-administered, gold particle-based drug with low toxicity for rheumatoid arthritis. The analyses of the independent, publicly available databases showed several enzymes in the TXNRD pathway, targetable by auranofin, are differentially expressed in ACC, suggesting that the combination of auranofin and RT can be effective. TXNRD is the pro-survival reducing enzyme of TXN, which cleans up ROS and oxygen free radicals created in abundance in both the cytoplasm and mitochondria during external beam radiation therapy. Although the radiosensitizing effects were promising, we found that the doses of auranofin and RT were higher than expected. We discovered that selenium supplement in the culture medium was likely the culprit that partially reversed auranofin-induced TXNRD inhibition. Thus, the current experiments are performed without selenium supplements. II. Targeted therapy focusing on the effects of TNF-a on tumor microenvironment to enhance drug delivery Aim: to elucidate the mechanism of action of TNF-a in reducing tumor interstitial pressure leading to improved drug delivery efficiency. I. Summary: Because the effects of TNF-a on expression and functions of several intratumoral extracellular matrix proteins and cytokines and the roles of these proteins and cytokines on intratumoral vascular permeability are not fully understood. The insight of this process can be used to improve drug delivery in TNF-a insensitive tumors by targeting the downstream targets of TNF-a. To understand the molecular mechanism of TNF-a-induced vascular leakage which results in decreased tumor IFP and increased intratumoral drug delivery, we performed ECM, human matrix metalloproteinase (MMP) arrays, and gene expression profiling of iodine-resistant thyroid cancer cells treated with TNF-a, we identified and validated that TNF-a down-regulated TGFB and LOX, and upregulated MMPs, TIMP1, and ITGB3 which have roles in vascular permeability in TME. siLOX treated iodine-refractory thyroid cancer xenograft recapitulated the intratumoral vascular leakage seen in xenografts treated with nanomedicine carrying TNF-a. Next, we validated the effects of TNF-a on vascular endothelium using vascular endothelial cells and found that TNF-a treatment downregulated TGFB1, Lox, and VEGF-R (a key regulator of vascular permeability). III. Quantitative high-throughput drug screening in BRAF V600E WT anaplastic thyroid cancer (ATC) Aim: To identify novel synergistic combinations of drugs in BRAF V600E WT ATC and to study the mechanism of synergy. Summary: ATC is a rare and one of the most aggressive malignancy. The median survival of patients with ATC is 6 months prior to the discovery of the combination of trametinib and dabrafenib in BRAF V600E mutated ATC that effectively improves treatment outcomes. Unfortunately, patients with BRAF V600E WT ATC remain without effective treatment options. Thus, it is critically important to identify the effective drug combinations that can be readily translated to a clinical trial. We performed a quantitative high-throughput drug screening using a pharmaceutical library of over 5,000 drugs on three ATC cell lines with RAS, BRAF V600E, and non-RAS, non-BRAF mutations. Ninety drugs were effective in RAS a *TRUNCATED*