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There are no officially approved therapies for metastatic pheochromocytomas apart from ultratrace 131I-metaiodbenzylguanidine therapy, which is approved only in the United States. We have, therefore, investigated the antitumor potential of molecular-targeted approaches in murine pheochromocytoma cell lines [monocyte chemoattractant protein (MPC)/monocyte chemoattractant protein/3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT)], immortalized mouse chromaffin Sdhb-/- cells, three-dimensional pheochromocytoma tumor models (MPC/MTT spheroids), and human pheochromocytoma primary cultures. We identified the specific phosphatidylinositol-3-kinase α inhibitor BYL719 and the mammalian target of rapamycin inhibitor everolimus as the most effective combination in all models. Single treatment with clinically relevant doses of BYL719 and everolimus significantly decreased MPC/MTT and Sdhb-/- cell viability. A targeted combination of both inhibitors synergistically reduced MPC and Sdhb-/- cell viability and showed an additive effect on MTT cells. In MPC/MTT spheroids, treatment with clinically relevant doses of BYL719 alone or in combination with everolimus was highly effective, leading to a significant shrinkage or even a complete collapse of the spheroids. We confirmed the synergism of clinically relevant doses of BYL719 plus everolimus in human pheochromocytoma primary cultures of individual patient tumors with BYL719 attenuating everolimus-induced AKT activation. We have thus established a method to assess molecular-targeted therapies in human pheochromocytoma cultures and identified a highly effective combination therapy. Our data pave the way to customized combination therapy to target individual patient tumors., (Copyright © 2019 Endocrine Society.)

Metastatic pheochromocytoma continues to be an incurable disease, and treatment with conventional cytotoxic chemotherapy offers limited efficacy. In the present study, we evaluated a novel topoisomerase I inhibitor, LMP-400, as a potential treatment for this devastating disease. We found a high expression of topoisomerase I in human metastatic pheochromocytoma, providing a basis for the evaluation of a topoisomerase 1 inhibitor as a therapeutic strategy. LMP-400 inhibited the cell growth of established mouse pheochromocytoma cell lines and primary human tumor tissue cultures. In a study performed in athymic female mice, LMP-400 demonstrated a significant inhibitory effect on tumor growth with two drug administration regimens. Furthermore, low doses of LMP-400 decreased the protein levels of hypoxia-inducible factor 1 (HIF-1α), one of a family of factors studied as potential metastatic drivers in these tumors. The HIF-1α decrease resulted in changes in the mRNA levels of HIF-1 transcriptional targets. In vitro, LMP-400 showed an increase in the growth-inhibitory effects in combination with other chemotherapeutic drugs that are currently used for the treatment of pheochromocytoma. We conclude that LMP-400 has promising antitumor activity in preclinical models of metastatic pheochromocytoma and its use should be considered in future clinical trials.

Currently, there are no reliably effective therapeutic options for metastatic pheochromocytoma (PCC) and paraganglioma. Moreover, there are no therapies that may prevent the onset or progression of tumors in patients with succinate dehydrogenase type B mutations, which are associated with very aggressive tumors. Therefore, we tested the approved and well-tolerated drugs lovastatin and 13-cis-retinoic acid (13cRA) in vitro in an aggressive PCC mouse cell line, mouse tumor tissue-derived (MTT) cells, and in vivo in a PCC allograft nude mouse model, in therapeutically relevant doses. Treatment was started 24 hours before sc tumor cell injection and continued for 30 more days. Tumor sizes were measured from outside by caliper and sizes of viable tumor mass by bioluminescence imaging. Lovastatin showed antiproliferative effects in vitro and led to significantly smaller tumor sizes in vivo compared with vehicle treatment. 13cRA promoted tumor cell growth in vitro and led to significantly larger viable tumor mass and significantly faster increase of viable tumor mass in vivo over time compared with vehicle, lovastatin, and combination treatment. However, when combined with lovastatin, 13cRA enhanced the antiproliferative effect of lovastatin in vivo. The combination-treated mice showed slowest tumor growth of all groups with significantly slower tumor growth compared with the vehicle-treated mice and significantly smaller tumor sizes. Moreover, the combination-treated group displayed the smallest size of viable tumor mass and the slowest increase in viable tumor mass over time of all groups, with a significant difference compared with the vehicle- and 13cRA-treated group. The combination-treated tumors showed highest extent of necrosis, lowest median microvessel density and highest expression of α-smooth muscle actin. The combination of high microvessel density and low α-smooth muscle actin is a predictor of poor prognosis in other tumor entities. Therefore, this drug combination may be a well-tolerated novel therapeutic or preventive option for malignant PCC.

Several lines of evidence, including the recent discovery of novel susceptibility genes, point out an important role for the mammalian target of rapamycin (mTOR) signaling pathway in the development of pheochromocytoma. Analyzing a set of pheochromocytomas from patients with different genetic backgrounds, we observed and confirmed a significant overexpression of key mTOR complex (mTORC) signaling mediators. Using selective ATP-competitive inhibitors targeting both mTORC1 and mTORC2, we significantly arrested the in vitro cell proliferation and blocked migration of pheochromocytoma cells as a result of the pharmacological suppression of the Akt/mTOR signaling pathway. Moreover, AZD8055, a selective ATP-competitive dual mTORC1/2 small molecular inhibitor, significantly reduced the tumor burden in a model of metastatic pheochromocytoma using female athymic nude mice. This study suggests that targeting both mTORC1 and mTORC2 is a potentially rewarding strategy and supports the application of selective inhibitors in combinatorial drug regimens for metastatic pheochromocytoma.