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The novel melphalan prodrug J1 inhibits neuroblastoma growth in vitro and in vivo
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical Pharmacology. (Cancer Pharmacology and Informatics)
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2007 (English)In: Molecular Cancer Therapeutics, ISSN 1535-7163, E-ISSN 1538-8514, Vol. 6, no 9, 2409-2417 p.Article in journal (Refereed) Published
Abstract [en]

Neuroblastoma is the most common extracranial solid tumor of childhood. The activity of J1 (l-melphalanyl-p-l-fluorophenylalanine ethyl ester), an enzymatically activated melphalan prodrug, was evaluated in neuroblastoma models in vitro and in vivo. Seven neuroblastoma cell lines with various levels of drug resistance were screened for cytotoxicity of J1 alone or in combination with standard cytotoxic drugs, using a fluorometric cytotoxicity assay. J1 displayed high cytotoxic activity in vitro against all neuroblastoma cell lines, with IC50 values in the submicromolar range, significantly more potent than melphalan. The cytotoxicity of J1, but not melphalan, could be significantly inhibited by the aminopeptidase inhibitor bestatin. J1 induced caspase-3 cleavage and apoptotic morphology, had additive effects in combination with doxorubicin, cyclophosphamide, carboplatin, and vincristine, and synergistically killed otherwise drug-resistant cells when combined with etoposide. Athymic rats and mice carrying neuroblastoma xenografts [SH-SY5Y, SK-N-BE(2)] were treated with equimolar doses of melphalan, J1, or no drug, and effects on tumor growth and tissue morphology were analyzed. Tumor growth in vivo was significantly inhibited by J1 compared with untreated controls. Compared with melphalan, J1 more effectively inhibited the growth of mice with SH-SY5Y xenografts, was associated with higher caspase-3 activation, fewer proliferating tumor cells, and significantly decreased mean vascular density. In conclusion, the melphalan prodrug J1 is highly active in models of neuroblastoma in vitro and in vivo, encouraging further clinical development in this patient group.

Place, publisher, year, edition, pages
2007. Vol. 6, no 9, 2409-2417 p.
Keyword [en]
alkylating prodrug, neuroblastoma, melphalan, aminopeptidase, synergy
National Category
Medical and Health Sciences
URN: urn:nbn:se:uu:diva-96362DOI: 10.1158/1535-7163.MCT-07-0156ISI: 000249716200005PubMedID: 17876040OAI: oai:DiVA.org:uu-96362DiVA: diva2:170913
Available from: 2007-11-09 Created: 2007-11-09 Last updated: 2011-01-26Bibliographically approved
In thesis
1. Preclinical Studies of the Melphalan Prodrug J1 for Cancer Therapy
Open this publication in new window or tab >>Preclinical Studies of the Melphalan Prodrug J1 for Cancer Therapy
2007 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

J1 (L-melphalanyl-L-p-fluorophenylalanyl ethyl ester) is a dipeptide derivative of the alkylating agent melphalan with increased cytotoxicity. In this thesis the preclinical pharmacology of J1 has been characterized.

Our results show that J1 rapidly enters the cells, where melphalan is released by hydrolysis. The maximum concentration (Cmax) of melphalan was detected 15 min after exposure to J1 in human cancer cell lines. In comparison, melphalan exposure resulted in a 10-fold lower Cmax that was shifted to later time points. J1 induced more DNA damage and apoptosis than melphalan. The cytotoxic activity and release of melphalan from J1 were inhibited by preincubating cells with the aminopeptidase inhibitor bestatin. In accordance with these results, we showed that J1 is a substrate for aminopeptidase N (APN), which may result in increased tumor selectivity.

J1 effectively inhibited cell growth in a set of neuroblastoma cell lines. Athymic mice carrying neuroblastoma xenografts were treated either with equimolar doses of melphalan or J1. J1 inhibited the tumor growth more effectively than melphalan and the untreated control, and was associated with higher caspase-3 activation, fewer proliferating tumor cells and decreased mean vascular density.

J1 and melphalan showed similar activity profiles when tested in 176 primary tumor cell cultures from patients, but J1 exhibited 50- to 100-fold higher potency. The difference was greater in some diagnoses (e.g. breast cancer, NHL and AML), and was exceptionally large in some breast cancer samples with aggressive phenotypes. A combination screening of J1 and standard chemotherapeutics yielded mostly additive interactions, except for etoposide which induced synergy in all tested cell lines.

In conclusion, the melphalan prodrug J1 is effectively transported into the cells, where aminopeptidases (for example APN) catalyze the formation of melphalan. J1 shows promising preclinical potential in the diagnoses neuroblastoma and breast cancer.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2007. 59 p.
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 285
Pharmacology, Pharmacology, Chemotherapy, Melphalan prodrug, Cancer, Farmakologi
urn:nbn:se:uu:diva-8285 (URN)978-91-554-7005-0 (ISBN)
Public defence
2007-11-30, Rudbecksalen, Rudbecklaboratoriet, Dag Hammarskjölds väg 20, Uppsala, 09:15
Available from: 2007-11-09 Created: 2007-11-09Bibliographically approved

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