Chemical genetic discovery of targets and anti-targets for cancer polypharmacology.

The complexity of cancer has led to recent interest in polypharmacological approaches for developing kinase-inhibitor drugs; however, optimal kinase-inhibition profiles remain difficult to predict. Using a Ret-kinase-driven Drosophila model of multiple endocrine neoplasia type 2 and kinome-wide drug profiling, here we identify that AD57 rescues oncogenic Ret-induced lethality, whereas related Ret inhibitors imparted reduced efficacy and enhanced toxicity. Drosophila genetics and compound profiling defined three pathways accounting for the mechanistic basis of efficacy and dose-limiting toxicity. Inhibition of Ret plus Raf, Src and S6K was required for optimal animal survival, whereas inhibition of the 'anti-target' Tor led to toxicity owing to release of negative feedback. Rational synthetic tailoring to eliminate Tor binding afforded AD80 and AD81, compounds featuring balanced pathway inhibition, improved efficacy and low toxicity in Drosophila and mammalian multiple endocrine neoplasia type 2 models. Combining kinase-focused chemistry, kinome-wide profiling and Drosophila genetics provides a powerful systems pharmacology approach towards developing compounds with a maximal therapeutic index.

Transient partial response to sorafenib treatment in an adolescent patient with MEN2B syndrome and end-stage medullary thyroid carcinoma.

Metastatic medullary thyroid carcinoma (MTC) is an aggressive malignancy with an extremely poor prognosis. Currently no effective conventional systemic therapies exist to treat pediatric MTC. We describe an adolescent female with newly diagnosed MEN2B syndrome who presented with advanced stage metastatic MTC and demonstrated a partial transient response to sorafenib monotherapy. This clinical result supports further research into the use of sorafenib in the treatment of pediatric MTC.

Proteomics study of medullary thyroid carcinomas expressing RET germ-line mutations: identification of new signaling elements.

Proteomics may help to elucidate differential signaling networks underlying the effects of compounds and to identify new therapeutic targets. Using a proteomic-multiplexed analysis of the phosphotyrosine signaling together with antibody-based validation techniques, we identified several candidate molecules for RET (rearranged during transfection) tyrosine kinase receptor carrying mutations responsible for the multiple endocrine neoplasia type 2A and 2B (MEN2A and MEN2B) syndromes in two human medullary thyroid carcinoma (MTC) cell lines, TT and MZ-CRC-1, which express the RET-MEN2A and RET-MEN2B oncoproteins, respectively. Signaling elements downstream of these oncoproteins were identified after treating cells with the indolinone tyrosine kinase inhibitor RPI-1 to knock down RET phosphorylation activity. We detected 23 and 18 affinity-purified phosphotyrosine proteins in untreated TT and MZ-CRC-1 cells, respectively, most of which were shared and sensitive to RPI-1 treatment. However, our data clearly point to specific signaling features of the RET-MEN2A and RET-MEN2B oncogenic pathways. Moreover, the detection of high-level expression of minimally phosphorylated epidermal growth factor receptor (EGFR) in both TT and MZ-CRC-1 cells, together with our data on the effects of EGF stimulation on the proteomic profiles and the response to Gefitinib treatment, suggest the relevance of EGFR signaling in these cell lines, especially since analysis of 14 archival MTC specimens revealed EGFR mRNA expression in all samples. Together, our data suggest that RET/EGFR multi-target inhibitors might be beneficial for therapy of MTC.

Diagnosis of multiple endocrine neoplasia type 2B and management of its ocular features.

BACKGROUND: To describe ocular and vascular findings in a patient with multiple endocrine neoplasia type 2B. MATERIALS AND METHODS: Case report of a 31-year-old male who was referred for ocular assessment following diagnosis of a carotid artery dissection and multiple vascular anomalies. RESULTS: Clinical examination revealed a marfanoid habitus, myelinated corneal nerve fibers, neuromas in the perilimbal area, conjunctival hyperemia with peripheral corneal neovascularization, and posterior blepharitis. Diagnosis of multiple endocrine neoplasia type 2B was confirmed by genetic testing of the RET proto-oncogene. Genetic screening for hereditary aortic and peripheral vasculopathies failed to reveal an underlying cause for the vascular findings. We noted improvement of the ocular surface disease with topical corticosteroids and oral tetracyclines. CONCLUSIONS: Ophthalmologists play a vital role in recognizing this rare but lethal malignancy. We report on a patient with apart from characteristic ocular findings also staphylococcal hypersensitivity and widespread systemic vasculopathy.

ZD6474 suppresses oncogenic RET isoforms in a Drosophila model for type 2 multiple endocrine neoplasia syndromes and papillary thyroid carcinoma.

Patients with hereditary medullary thyroid carcinoma (MTC) associated with multiple endocrine neoplasia (MEN) types 2A and 2B and familial MTC (FMTC) have mutations in the RET proto-oncogene. Approximately 40 percent of patients with papillary thyroid carcinoma (PTC) typically have either intrachromosomal or extrachromosomal rearrangements that join the promoter and NH(2)-terminal domains of unrelated genes to the COOH-terminal fragment of RET. The RET point mutations associated with MEN2A, MEN2B, or FMTC, or the chromosomal breakpoints and translocations associated with PTC, typically activate the RET receptor tyrosine kinase (RTK). RET kinase inhibitors are likely to be beneficial for patients with hereditary MTC, where currently there is no effective chemotherapy or radiation therapy. Recently, the low molecular weight tyrosine kinase inhibitor ZD6474 was found to block the enzymatic activity of RET-derived oncoproteins in cultured cell lines. We have developed a Drosophila model for MEN2A and MEN2B diseases by targeting oncogenic forms of RET to the developing Drosophila eye. Here we show that, when fed orally, ZD6474 suppressed RET-mediated phenotypes within the context of this in vivo model. Importantly, ZD6474 showed high efficacy and very low toxicity. This compound failed to significantly suppress an activated form of another RTK, the Drosophila epidermal growth factor receptor, nor did it suppress the activity of downstream components of the RET/Ras pathway. Our results support the view that targeting chemical kinase inhibitors such as ZD6474 to tissues with oncogenic forms of RET is a useful treatment strategy for RET-dependent carcinomas.

Vandetanib in children and adolescents with multiple endocrine neoplasia type 2B associated medullary thyroid carcinoma.

PURPOSE: Medullary thyroid carcinoma (MTC) is a manifestation of multiple endocrine neoplasia type 2 (MEN2) syndromes caused by germline, activating mutations in the RET (REarranged during Transfection) proto-oncogene. Vandetanib, a VEGF and EGF receptor inhibitor, blocks RET tyrosine kinase activity and is active in adults with hereditary MTC. EXPERIMENTAL DESIGN: We conducted a phase I/II trial of vandetanib for children (5-12 years) and adolescents (13-18 years) with MTC to define a recommended dose and assess antitumor activity. The starting dose was 100 mg/m(2) administered orally, once daily, continuously for 28-day treatment cycles. The dose could be escalated to 150 mg/m(2)/d after two cycles. Radiographic response to vandetanib was quantified using RECIST (v1.0), biomarker response was measured by comparing posttreatment serum calcitonin and carcinoembryonic antigen (CEA) levels to baseline, and a patient-reported outcome was used to assess clinical benefit. RESULTS: Sixteen patients with locally advanced or metastatic MTC received vandetanib for a median (range) 27 (2-52) cycles. Eleven patients remain on protocol therapy. Diarrhea was the primary dose-limiting toxicity. In subjects with M918T RET germline mutations (n = 15) the confirmed objective partial response rate was 47% (exact 95% confidence intervals, 21%-75%). Biomarker partial response was confirmed for calcitonin in 12 subjects and for CEA in 8 subjects. CONCLUSION: Using an innovative trial design and selecting patients based on target gene expression, we conclude that vandetanib 100 mg/m(2)/d is a well-tolerated and highly active new treatment for children and adolescents with MEN2B and locally advanced or metastatic MTC.

Drosophila as a novel therapeutic discovery tool for thyroid cancer.

BACKGROUND: Multiple endocrine neoplasia type II (MEN2) is a rare but aggressive cancer for which no effective treatment currently exists. A Drosophila model was developed to identify novel genetic modifier loci of oncogenic RET, as well as to provide a whole animal system to rapidly identify compounds that suppressed RET-dependent MEN2. ZD6474 (Vandetanib), currently in phase III trials, suppressed tumorigenesis in MEN2 model flies, demonstrating for the first time the effectiveness of a Drosophila-based whole animal model for identifying therapeutically useful compounds. SUMMARY: Clinical data suggest that drug mono-therapy for MEN2 and other cancers typically yield only moderate benefits as patients develop drug resistance and suffer from drug-induced pathway feedback. Combinations of drugs that target different nodes of the oncogenic pathway are an effective way to prevent resistance as well as feedback. Identifying the optimal drug-dose combinations for therapy poses a significant challenge in existing mouse models. Fly models offer a means to quickly and effectively identify drug combinations that are well tolerated and potently suppress the MEN2 phenotype. This approach may also identify differences in therapeutic responses between the two subtypes of MEN2--MEN2A and MEN2B--providing additional therapeutic insights. CONCLUSIONS: Fly models have proven useful for identifying known drugs as well as novel compounds that, as single agents or in combinations, effectively suppress the MEN2 syndrome. These findings validate the use of fly models for both drug discovery as well as identification of useful drug combinations. In the future, rapid pairing of new genomic information with increasingly complex fly models will aid us in efforts to further tailor drug treatments toward personalized medicine.