Neuroblastoma is the most common extracranial solid tumor in children and is a high-risk tumor that causes death in the majority of cases. Alkylating agents are the primary chemotherapeutic agents used in neuroblastoma and resistance contributes to relapse mortality. Recently, we have demonstrated that 1) intracellular depletion of glutathione (GSH) by buthionine sulfoximine (BSO) has a cytotoxic synergy with the alkylator melphalan (L-PAM) that is antagonized by hypoxia and 2) that retinoids improve survival in high-risk neuroblastoma but may antagonize alkylator activity. Our long term goals are to define in neuroblastom: 1) the molecular determinants of alkylator resistance and hypoxia antagonism and to identify agents that overcome them; 2) identify alkylators with the greatest single agent activity and tailor patient therapy to them; 3) identify patterns of alkylator cross-resistance and avoid these combinations clinically; 4) identify alkylators with the highest BSO synergism; 5) with L-PAM and other agents.
Specific Aims. 1) Determine the single agent activity of clinical alkylators in normoxia and hypoxia, their patterns of cross-resistance, and synergism with BSO. 2) Determine, for BSO/L-PAM, if hypoxia effects endogenous reactive oxygen species (ROS), GSH depletion, DNA damage/p53 induction, and if bioreductive agents reverse the effects of hypoxia. 3) Determine in clinical trials the toxicity and efficacy of myeloablative BSO/L-PAM, for neuroblastoma. Research Design and Methods. We Will use our unique panel of 140 cell lines, whose sole drug exposure has been in vivo, and that includes matched cell line pairs from patients at diagnosis and relapse, including relapse after bone marrow transplant. We will expose selected lines to clinically relevant doses of alkylators, BSO, and bioreductive agents (tirapazamine and misonidazole) in normoxia, and in hypoxia using isolation chambers, and assay cytotoxicity using our custom, semi-automated DIMSCAN quantitative imaging system. Because our cell panel is large, we uniquely have the statistical power to detect patterns of alkylator cross-resistance in both normoxia and hypoxia for these agents, the first study of its kind in any tumor system. For BSO/L-PAM hypoxia antagonism studies, we will measure total and nuclear GSH by glutathione-reductase-DTNB recycling assay, ROS by carboxy-DCFDA flow cytometry, ROS-induced 8-oxodeoxyguanosine DNA damage by avidin binding technique, DAN single strand breaks and interstrand crosslinking by alkaline elution, p3/p21 levels by immunoblotting and cytotoxicity by DIMSCAN, apoptotic DNA laddering. TdT-labeling/flow cytometry and caspase activation. We anticipate that this research will contribute to the understanding of alkylator resistance in neuroblastoma and will result in more effective treatment modalities.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA082830-03
Application #
6377430
Study Section
Special Emphasis Panel (ZRG1-CONC (01))
Program Officer
Wu, Roy S
Project Start
1999-09-01
Project End
2003-08-31
Budget Start
2001-09-01
Budget End
2003-08-31
Support Year
3
Fiscal Year
2001
Total Cost
$234,734
Indirect Cost
Name
Children's Hospital of Los Angeles
Department
Type
DUNS #
094878337
City
Los Angeles
State
CA
Country
United States
Zip Code
90027
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
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
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
Anderson, Clarke P; Matthay, Katherine K; Perentesis, John P et al. (2015) Pilot study of intravenous melphalan combined with continuous infusion L-S,R-buthionine sulfoximine for children with recurrent neuroblastoma. Pediatr Blood Cancer 62:1739-46
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

Showing the most recent 10 out of 37 publications