Melphalan transport, glutathione levels, and glutathione-S-transferase activity in human medulloblastoma.
Melphalan transport, glutathione levels, and glutathione-S-transferase activity were measured in two continuous human medulloblastoma cell lines and transplantable xenografts in athymic nude mice, TE-671 and Daoy. In vitro mean glutathione levels were 10.06 nmol/10(6) cells in TE-671 and 2.96 nmol/10(6) cells in Daoy. In vitro mean glutathione-S-transferase values were 91.52 nmol/min/mg protein in TE-671 and 50.31 nmol/min/mg protein in Daoy. Transport studies revealed kinetic parameters of Km = 108.3 microM, Vmax = 363.1 pmol/10(6) cells/min in TE-671 and Km = 111.7 microM, Vmax = 180.6 pmol/10(6) cells/min in Daoy. Melphalan transport was inhibited by both DL-alpha-2-aminobicyclo[2.2.1]heptane-2- carboxylic acid and sodium ion depletion in TE-671 and Daoy cells in vitro, indicating that both systems of amino acid transport are functional in these medulloblastoma lines. In vivo s.c. xenograft glutathione values were lower (7.79 nmol/mg protein) in TE-671 than in Daoy (13.68 nmol/mg protein). The mean plasma concentration in mice given a 10% lethal dose (71.3 mg/m2) of melphalan i.p. was 50.3 microM at 10 min, with the half-life of 29.9 min. At this dose, s.c. xenograft levels were 2- to 3-fold higher in TE-671 than in Daoy tumors for the 3-h period measured. These studies demonstrate transport parameters confirming facilitated transport of melphalan in human medulloblastoma, a mean murine plasma melphalan concentration (following treatment with melphalan) above the in vitro drug dose at which there is a 90% reduction in the number of colonies in comparison to controls for TE-671 and Daoy for 2 h, and glutathione and glutathione-S-transferase levels in the same range previously reported in other melphalan-sensitive and melphalan-resistant human tumors. Future work with spontaneous and acquired melphalan-resistant human medulloblastoma cell lines and xenografts will define the role of these mechanisms in mediating drug resistance.
Department of Pediatrics, Duke University Medical Center, Durham, North Carolina 27710.
H S
HS
Friedman
S X
SX
Skapek
O M
OM
Colvin
G B
GB
Elion
M R
MR
Blum
P M
PM
Savina
J
J
Hilton
S C
SC
Schold
J
J
Kurtzberg
D D
DD
Bigner
D000818:Animals; D001932:Brain Neoplasms / Q000201:enzymology / Q000378:metabolism*; D002460:Cell Line; D005978:Glutathione / Q000378:metabolism*; D005982:Glutathione Transferase / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008527:Medulloblastoma / Q000201:enzymology / Q000378:metabolism*; D008558:Melphalan / Q000493:pharmacokinetics*; D051379:Mice; D009368:Neoplasm Transplantation; D012964:Sodium / Q000494:pharmacology; D013997:Time Factors
D012964:Sodium; D005982:Glutathione Transferase; D005978:Glutathione; D008558:Melphalan