Summary
Objective: The dipeptide J1 acts as a prodrug of melphalan with a significant increased potency in vitro resulting from activation by cellular aminopeptidases. The current study was performed to evaluate the ex vivo profile of J1 using 176 primary tumor cell cultures from patients. In addition, the activity of J1 in combination with eight standard drugs, representing different mechanistic classes, was studied in nine different human tumor cell lines of different histopathological origin. Methods: Ex vivo evaluation of tumor type selectivity, was performed using the established fluorometric microculture cytotoxicity assay (FMCA). Combinations between J1 and eight standard chemotherapeutic drugs were analyzed using the median-effect method. Results: The prodrug J1 expressed approximately 50- to 100-fold higher potency but similar activity profile as that of its metabolite, melphalan. The difference was greater in some diagnoses (e.g. breast cancer, NHL and AML), and exceptionally high in some breast cancer samples with aggressive phenotypes. Combination analysis of J1 and standard chemotherapeutics yielded several potentially additive and synergistic interactions, most striking for etoposide with significant synergism in all studied cell lines. Conclusions: In conclusion, the ex vivo profile suggests that further evaluation of J1 as the alkylating agent in for example aggressive breast cancer might be of particular interest, preferentially in combination with DNA-topoisomerase II inhibitors like etoposide.
Similar content being viewed by others
References
Furner RL, Brown RK (1980) L-phenylalanine mustard (L-PAM): the first 25 years. Cancer Treat Rep 64:559–74
Teicher BA (1997) (405–418) Antitumor alkylating agents. In: De Vita Hellman Rodenberg VTSSA (ed) Cancer principals and practice of oncology. Lippincott-Raven, Philadelphia, USA
Jones RB (2002) Clinical pharmacology of melphalan and its implications for clinical resistance to anticancer agents. Cancer Treat Res 112:305–322
Donato ML, Gershenson DM, Wharton JT, Ippoliti CM, Aleman AS, Bodurka-Bevers D, Bevers MW, Burke TW, Levenback CF, Wolf JK, Freedman RS, Bast RC Jr, Gajewski JL, Champlin RE (2001) High-dose topotecan, melphalan, and cyclophosphamide (TMC) with stem cell support: a new regimen for the treatment of advanced ovarian cancer. Gynecol Oncol 82:420–426
Demirer T, Uysal VA, Ayli M, Genc Y, Ilhan O, Koc H, Dagli M, Arat M, Gunel N, Fen T, Dincer S, Ustael N, Yildiz M, Ustun T, Seyrek E, Ozet G, Muftuoglu O, Akan H (2003) High-dose thiotepa, melphalan and carboplatin (TMCb) followed by autologous stem cell transplantation in patients with advanced breast cancer: a retrospective evaluation. Bone Marrow Transplant 31:755–761
Gullbo J, Dhar S, Luthman K, Ehrsson H, Lewensohn R, Nygren P, Larsson R (2003) Antitumor activity of the alkylating oligopeptides J1 (L-melphalanyl-p-L-fluorophenylalanine ethyl ester) and P2 (L-prolyl-m-L-sarcolysyl-p-L-fluorophenylalanine ethyl ester): comparison with melphalan. Anti-cancer Drugs 14:617–624
Gullbo J, Tullberg M, Vabeno J, Ehrsson H, Lewensohn R, Nygren P, Larsson R, Luthman K (2003) Structure–activity relationship for alkylating dipeptide nitrogen mustard derivatives. Oncol Res 14:113–132
Gullbo J, Lindhagen E, Bashir-Hassan S, Tullberg M, Ehrsson H, Lewensohn R, Nygren P, De La Torre M, Luthman K, Larsson R (2004) Antitumor efficacy and acute toxicity of the novel dipeptide melphalanyl-p-L-fluorophenylalanine ethyl ester (J1) in vivo. Invest New Drugs 22:411–420
Wickström M, Johnsen JI, Ponthan F, Segerström L, Sveinbjörnsson B, Lindskog M, Lövborg H, Viktorsson K, Lewensohn R, Kogner P, Larsson R, Gullbo J (2007) The novel melphalan prodrug J1 inhibits neuroblastoma growth in vitro and in vivo. Mol Cancer Ther 6:2409–2417
Gullbo J, Wickstrom M, Tullberg M, Ehrsson H, Lewensohn R, Nygren P, Luthman K, Larsson R (2003) Activity of hydrolytic enzymes in tumour cells is a determinant for anti-tumour efficacy of the melphalan containing prodrug J1. J Drug Target 11:355–363
van Hensbergen Y, Broxterman HJ, Hanemaaijer R, Jorna AS, van Lent NA, Verheul HM, Pinedo HM, Hoekman K (2002) Soluble aminopeptidase N/CD13 in malignant and nonmalignant effusions and intratumoral fluid. Clin Cancer Res 8:3747–3754
Suganuma T, Ino K, Shibata K, Nomura S, Kajiyama H, Kikkawa F, Tsuruoka N, Mizutani S (2004) Regulation of aminopeptidase A expression in cervical carcinoma: role of tumor–stromal interaction and vascular endothelial growth factor. Lab Invest 84:639–648
Martinez JM, Prieto I, Ramirez MJ, Cueva C, Alba F, Ramirez M (1999) Aminopeptidase activities in breast cancer tissue. Clin Chem 45:1797–1802
Bosanquet AG, Bell PB (2004) Ex vivo therapeutic index by drug sensitivity assay using fresh human normal and tumor cells. J Exp Ther Oncol 4:145–154
Fridborg H, Jonsson E, Nygren P, Larsson R (1999) Relationship between diagnosis-specific activity of cytotoxic drugs in fresh human tumour cells ex vivo and in the clinic. Eur J Cancer 35:424–432
Chou TC, Talalay P (1984) Quantitative analysis of dose–effect relationships: the combined effects of multiple drugs or enzyme inhibitors. Adv Enzyme Regul 22:27–55
Frickhofen N, Berdel WE, Opri F, Haas R, Schneeweiss A, Sandherr M, Kuhn W, Hossfeld DK, Thomssen C, Heimpel H, Kreienberg R, Hinke A, Mobus V (2006) Phase I/II trial of multicycle high-dose chemotherapy with peripheral blood stem cell support for treatment of advanced ovarian cancer. Bone Marrow Transplant 38:493–499
Larsson R, Kristensen J, Sandberg C, Nygren P (1992) Laboratory determination of chemotherapeutic drug resistance in tumor cells from patients with leukemia, using a fluorometric microculture cytotoxicity assay (FMCA). Int J Cancer 50:177–185
Csoka K, Larsson R, Tholander B, Gerdin E, de la Torre M, Nygren P (1994) Cytotoxic drug sensitivity testing of tumor cells from patients with ovarian carcinoma using the fluorometric microculture cytotoxicity assay (FMCA). Gynecol Oncol 54:163–170
Nygren P, Kristensen J, Jonsson B, Sundstrom C, Lonnerholm G, Kreuger A, Larsson R (1992) Feasibility of the fluorometric microculture cytotoxicity assay (FMCA) for cytotoxic drug sensitivity testing of tumor cells from patients with acute lymphoblastic leukemia. Leukemia 6:1121–1128
Rickardson L, Fryknas M, Haglund C, Lovborg H, Nygren P, Gustafsson MG, Isaksson A, Larsson R (2006) Screening of an annotated compound library for drug activity in a resistant myeloma cell line. Cancer Chemother Pharmacol 58:749–758
Dixon J, Kaklamanis L, Turley H, Hickson ID, Leek RD, Harris AL, Gatter KC (1994) Expression of aminopeptidase-n (CD 13) in normal tissues and malignant neoplasms of epithelial and lymphoid origin. J Clin Pathol 47:43–47
Pulido-Cejudo G, Miranda H, El Abdaimi K, Wang C, Kar B, Medina Acevedo J, Cardenas JM, Sarti Gutierrez E, Perez Palacios G (2004) A monoclonal antibody driven biodiagnostic system for the quantitative screening of breast cancer. Biotechnol Lett 26:1335–1339
Tokuhara T, Hattori N, Ishida H, Hirai T, Higashiyama M, Kodama K, Miyake M (2006) Clinical significance of aminopeptidase N in non-small cell lung cancer. Clin Cancer Res 12:3971–3978
Surowiak P, Drag M, Materna V, Suchocki S, Grzywa R, Spaczynski M, Dietel M, Oleksyszyn J, Zabel M, Lage H (2006) Expression of aminopeptidase N/CD13 in human ovarian cancers. Int J Gynecol Cancer 16:1783–1788
Shibata K, Kajiyama H, Mizokami Y, Ino K, Nomura S, Mizutani S, Terauchi M, Kikkawa F (2005) Placental leucine aminopeptidase (P-LAP) and glucose transporter 4 (GLUT4) expression in benign, borderline, and malignant ovarian epithelia. Gynecol Oncol 98:11–18
Yamashita M, Kajiyama H, Terauchi M, Shibata K, Ino K, Nawa A, Mizutani S, Kikkawa F (2007) Involvement of aminopeptidase N in enhanced chemosensitivity to paclitaxel in ovarian carcinoma in vitro and in vivo. Int J Cancer 120:2243–2250
Ishii K, Usui S, Yamamoto H, Sugimura Y, Tatematsu M, Hirano K (2001) Decreases of metallothionein and aminopeptidase N in renal cancer tissues. J Biochem (Tokyo) 129:253–258
Varona A, Blanco L, Lopez JI, Gil J, Agirregoitia E, Irazusta J, Larrinaga G (2007) Altered levels of acid, basic, and neutral peptidase activity and expression in human clear cell renal cell carcinoma. Am J Physiol Renal Physiol 292:F780–F788
Birkenkamp-Demtroder K, Christensen LL, Olesen SH, Frederiksen CM, Laiho P, Aaltonen LA, Laurberg S, Sorensen FB, Hagemann R, Orntoft TF (2002) Gene expression in colorectal cancer. Cancer Res 62:4352–4363
Hashida H, Takabayashi A, Kanai M, Adachi M, Kondo K, Kohno N, Yamaoka Y, Miyake M (2002) Aminopeptidase N is involved in cell motility and angiogenesis: its clinical significance in human colon cancer. Gastroenterology 122:376–386
Kawamura J, Shimada Y, Kitaichi H, Komoto I, Hashimoto Y, Kaganoi J, Miyake M, Yamasaki S, Kondo K, Imamura M (2007) Clinicopathological significance of aminopeptidase N/CD13 expression in human gastric carcinoma. Hepatogastroenterology 54:36–40
Buzdar AU, Legha SS, Blumenschein GR, Hortobagyi GN, Yap HY, Schell FC, Barnes BC, Fraschini G, Bodey GP (1982) Peptichemio versus melphalan (L-PAM) in advanced breast cancer. Cancer 49:1767–1770
Kaufmann SH, Peereboom D, Buckwalter CA, Svingen PA, Grochow LB, Donehower RC, Rowinsky EK (1996) Cytotoxic effects of topotecan combined with various anticancer agents in human cancer cell lines. J Natl Cancer Inst 88:734–741
Hirota H, Gosky D, Berger NA, Chatterjee S (2002) Interference with topoisomerase IIalpha potentiates melphalan cytotoxicity. Int J Oncol 20:311–318
Valteau-Couanet D, Vassal G, Pondarre C, Bonnay M, Benhamou E, Couanet D, Plantaz D, Hartmann O (1996) Phase I study of high-dose continuous intravenous infusion of VP-16 in combination with high-dose melphalan followed by autologous bone marrow transplantation in children with stage IV neuroblastoma. Bone Marrow Transplant 17:485–489
Berenson JR, Yang HH, Sadler K, Jarutirasarn SG, Vescio RA, Mapes R, Purner M, Lee SP, Wilson J, Morrison B, Adams J, Schenkein D, Swift R (2006) Phase I/II trial assessing bortezomib and melphalan combination therapy for the treatment of patients with relapsed or refractory multiple myeloma. J Clin Oncol 24:937–944
Acknowledgement
Christina Leek and Lena Lenhammar are gratefully acknowledged for skilful technical assistance and handling of primary tumor samples.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Wickström, M., Haglund, C., Lindman, H. et al. The novel alkylating prodrug J1: diagnosis directed activity profile ex vivo and combination analyses in vitro . Invest New Drugs 26, 195–204 (2008). https://doi.org/10.1007/s10637-007-9092-1
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s10637-007-9092-1