Myeloablative chemoradiotherapy supported by autologous bone marrow transplantation (ABMT) and followed by 13-cis-retinoic acid improves survival in high-risk neuroblastoma, yet over 50% of such patients eventually die from disease progression after ABMT. Neuroblastoma therapy relies heavily upon alkylating agents, suggesting that alkylator resistance plays a key role in neuroblastoma relapse. Alkylator resistance involves tumor glutathione (GSH), a ubiquitous tri-peptide antioxidant that aids in detoxifying alkylating agents, and tumor cell hypoxia may also contribute to alkylator resistance. Buthionine Sulfoximine (BSO), a selective inhibitor of gamma-glutamylcysteine synthetase (gamma-GCS), the rate-limiting enzyme in GSH synthesis, can decrease intracellular levels of GSH and enhance the cytotoxic effect of alkylating agents. We have established neuroblastoma cell lines at time of relapse after ABMT and we have shown that they have stable resistance to alkylating agents, and that one mechanism of resistance is a loss of p53 function, often via mutation. We will detect TP53 mutations (by Affymetrix GeneChip) and measure expression of p53/MDM2 protein in tumor and marrow metastases (by flow cytometry) from recurrent neuroblastomas (treated on COG and NANT protocols) to identify the frequency of p53 loss-of-function (p53 LOF) in recurrent neuroblastoma and to determine if response to salvage therapy correlates with p53 status. We propose to determine the degree of cross-resistance in a panel of 25 neuroblastoma cell lines of cyclophosphamide, carboplatin, melphalan (L-PAM) alone and in combination with topoisomerase inhibitors or BSO under standard culture conditions (20% O2) and physiological hypoxia (2% O2). We will identify the alkylating agents (+/- a topoisomerase inhibitor) with the greatest activity (+/- BSO in hypoxia) against multi-drug resistant, p53 LOF neuroblastomas in pre-clinical models. We will correlate enhanced response to alkylating agents when combined with topoisomerase inhibitors or BSO with the degree of tumor cell DNA damage and will explore down-stream mechanisms of tumor cell death occurring in the absence of p53 function. We will complete ongoing phase I clinical trials of BSO/L-PAM or pyrazoloacridine (PZA) + stem cell support. Based on laboratory and clinical data from this project we will develop additional phase I trials to define the tolerability of new combinations of agents that show promise against recurrent neuroblastoma, and also phase II trials to define the activity of agents and combinations of agents that show promise in phase I studies. This project will lead to greater understanding of drug resistance in neuroblastoma, will define a marker (p53 LOF) to identify drug-resistant neuroblastomas, and will test novel approaches to overcome alkylating agent resistance, especially resistance associated with a loss of p53 function.

National Institute of Health (NIH)
National Cancer Institute (NCI)
Research Project (R01)
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Clinical Oncology Study Section (CONC)
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Wu, Roy S
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Texas Tech University
Schools of Medicine
United States
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Villablanca, Judith G; Volchenboum, Samuel L; Cho, Hwangeui et al. (2016) A Phase I New Approaches to Neuroblastoma Therapy Study of Buthionine Sulfoximine and Melphalan With Autologous Stem Cells for Recurrent/Refractory High-Risk Neuroblastoma. Pediatr Blood Cancer 63:1349-56
Chen, Nancy E; Maldonado, N Vanessa; Khankaldyyan, Vazgen et al. (2016) Reactive Oxygen Species Mediates the Synergistic Activity of Fenretinide Combined with the Microtubule Inhibitor ABT-751 against Multidrug-Resistant Recurrent Neuroblastoma Xenografts. Mol Cancer Ther 15:2653-2664
Seidel, Diana; Shibina, Anastasia; Siebert, Nikolai et al. (2015) Disialoganglioside-specific human natural killer cells are effective against drug-resistant neuroblastoma. Cancer Immunol Immunother 64:621-34
Kang, Min H; Wang, Jing; Makena, Monish R et al. (2015) Activity of MM-398, nanoliposomal irinotecan (nal-IRI), in Ewing's family tumor xenografts is associated with high exposure of tumor to drug and high SLFN11 expression. Clin Cancer Res 21:1139-50
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Farooqi, Ahsan S; Dagg, Rebecca A; Choi, L Mi Rim et al. (2014) Alternative lengthening of telomeres in neuroblastoma cell lines is associated with a lack of MYCN genomic amplification and with p53 pathway aberrations. J Neurooncol 119:17-26
Tagde, A; Singh, H; Kang, M H et al. (2014) The glutathione synthesis inhibitor buthionine sulfoximine synergistically enhanced melphalan activity against preclinical models of multiple myeloma. Blood Cancer J 4:e229
Shibina, Anastasia; Seidel, Diana; Somanchi, Srinivas S et al. (2013) Fenretinide sensitizes multidrug-resistant human neuroblastoma cells to antibody-independent and ch14.18-mediated NK cell cytotoxicity. J Mol Med (Berl) 91:459-72
Goldsmith, Kelly C; Gross, Michelle; Peirce, Susan et al. (2012) Mitochondrial Bcl-2 family dynamics define therapy response and resistance in neuroblastoma. Cancer Res 72:2565-77
Fang, Hua; Harned, Theresa M; Kalous, Ondrej et al. (2011) Synergistic activity of fenretinide and the Bcl-2 family protein inhibitor ABT-737 against human neuroblastoma. Clin Cancer Res 17:7093-104

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