MECHANISM OF METHOTREXATE RESISTANCE IN NON-HODGKIN LYMPHOMA CELL
LINES IN VITRO
Chen Jing, Ying DM, Wu HJ, Gu LJ
Department of Hematology/Oncology,
Xinhua Hospital/Shanghai Children’s Medical Center,
Shanghai Second Medical University, Shanghai, China
Objective: To know the mechanisms of methotrexate
(MTX) resistance in Non-Hodgkin lymphoma.
Methods: MTX resistant cell lind-Namalwa-12 were
developed by repeatedly (12 times) exposing it to 1uM MTX for 24 hours.
Then used MTT drug sensitivity test to compare its MTX sensitivity with
those of the Sup-m2 (large Cell anaplastic lymphoma) and the Namalwa
(Burkitt’s lymphoma) cell lines. We also compared between the three cells
with respect to the level of dihydrofolate reductase (DHFR) mRNA
expression, the uptake function of reduced folate carrier (RFC) and the
quantity of MTX polyglutamate (MTXPG) assayed by RT-PCR, 3H-MTX
cell culture and HPLC separation.
Results: The results showed that the Sup-m2 cell line
was more sensitive to MTX than the Namalwa cell line (IC50
13.82nM±1.67 to 37.67±3.23nM, P<0.05), which in turn was
more sensitive than the Namalwa-12 cell line. Comparing to the Namalwa cell
line, the Sum-m2 cell line not only accumulated more MTXPG (64.45±10.4pmol/107
cells to 44.53±2.36 pmol/107 cells p<0.05), but the accumulation
reached saturation earlier (the percentages of MTXPG4-6 at 8
hours accounted for 78.36% and 45.19% of the MTXPG4-6 at 24h for
the two cell lines, respectively). Increased expression of DHFR mRNA was
found in the Namalwa-12 Cell Line.
Conclusion: Low accumulation of MTXPG and
over-expression of DHFR mRNA are associated with MTX resistance in
Namalwa-12 cell line. Whether high-dose methotrexate can overcome its
resistance remains to be determined.