| Abstract |
A continuonsly maintained cell line, RED-3, was successfully isolated from
the peripheral blood blasts of a 24 yo man with a TdT(+) acute leukemia in
relapse. The patient presented with a mediastinal mass and a WBC count of
30,000/mm3 with 24% blasts, which were lymphoblasts by morphology (FAB
class L2), TdT(+), polar block acid phosphatase (+), PAS (-), peroxidase
(-), nonspecific esterase (-), and 90% positive for E-rosette formation,
leading to a diagnosis of T-ALL. An initial complete remission was
achieved with vincristine, prednisone, daunomycin, and L-asparaginase. A
subsequent remission was achieved with the same drugs when the patient
relapsed 17 months after diagnosis. During a second relapse 9 mos later,
leukemic blasts were obtained for tissue culture after an unsuccessful
attempt to induce remission with mitoxantrone. Blasts were separated from
blood by Hypaque-Ficoll gradients and cultured at 10^6 cells/ml in RPMI
1640 supplemented with insulin, transferrin, selenium (ITS), 20% fetal
calf serum (FBS), and 10% PHA stimulated human lymphocyte conditioned
media (LCM). After 3 weeks' undisturbed incubation at 37 Celsius, 7% CO2,
additional media was added and then renewed weekly by demi-depletion with
gradual withdrawal of FBS and LCM. By 8 wks the cells were established in
RPMI and ITS alone with an average doubling time of 26 hrs and have been
maintained continuously since that time (>10 mos). Like the blast cells in
vivo, RED-3 cells in culture were found to be TdT(+) and polar block acid
phospatase(+). FACS analysis also showed them to be 86-96% positive for
the T-cell antigens, Leu 3a and Leu 3b. However, at the same time, these
cells were found to be histochemically positive for PAS, peroxidase, and
Sudan black and to be 85% positive for the myeloid antigen, Leu M1.
Moreover, when incubated with retinoic acid (10^-6M) or dimethylformamide
(6x10^-2M), the cells underwent myeloid maturation as assessed by
morphology, NBT reduction, and acquisition of the OKM1 antigen (78%+).
They also partially lost Leu 3 reactivity. The characteristics of this new
human leukemia cell line support previous reports that T-ALL lymphoblasts
or their precursors may retain a potential for myloid differentiation.
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