Overexpression of the PSMB5 gene contributes to bortezomib resistance in T-lymphoblastic lymphoma/leukemia cells derived from Jurkat line.
OBJECTIVE
To study the mechanism of bortezomib resistance in JurkatB lines derived from T-lymphoblastic lymphoma/leukemia Jurkat line.


METHODS
Cytotoxicities of popular chemotherapeutic drugs to JurkatB cells were analyzed by trypan blue assay. Functional drug efflux in JurkatB cells was determined by flow cytometry utilizing daunorubicin and the expression of P-glycoprotein (P-gp) was detected by Western blot. mRNA expression levels of proteasome beta5 subunit (PSMB5) were measured by quantitation real-time reverse transcription polymerase chain reaction. In situ hybridization was performed to detect the amplification of PSMB5 gene. The chymotrypsin-like activities were assayed by measuring the release of the fluorescent 7-amido-4-methylcoumarin (AMC) from the substrate N-succinyl-Leu-Leu-Val-Tyr-AMC. Cytogenetic studies were performed using R-banded metaphases and fluorescence in situ hybridization (FISH) analysis. IkappaB-alpha levels were detected by Western blot.


RESULTS
No cross-resistance to daunorubicin, adriamycin, vindesine, and etoposide was found in JurkatB cells. No evidence of drug efflux was found in JurkatB cells and the expression of P-gp was negative. The PSMB5 mRNA was overexpressed in highly resistant JurkatB5 and JurkatB1 lines compared with parental Jurkat, corresponding well with the increase of chymotrypsin-like activity and a karyotype of i(14q). Amplification of PSMB5 gene was demonstrated by in situ hybridization and FISH. The decreased IkappaB-alpha level in JurkatB5 cells after bortezomib treatment indicating an upregulation of nuclear factor-kappaB (NF-kappaB) activity.


CONCLUSIONS
The mechanism of bortezomib resistance is different from that of multidrug resistance. Overexpression of PSMB5 is an important mechanism for bortezomib resistance in JurkatB lines. NF-kappaB may play a critical role in evading the apoptotic effects.
Lü
Shuqing
S
Department of Hematology, Changhai Hospital, Second Military Medical University, Shanghai, China.
Chen
Zhilong
Z
Yang
Jianmin
J
Chen
Li
L
Gong
Shenglan
S
Zhou
Hong
H
Guo
Lieping
L
Wang
Jianmin
J
D000970:Antineoplastic Agents / Q000627:therapeutic use*; D001897:Boronic Acids / Q000627:therapeutic use*; D000069286:Bortezomib; D002470:Cell Survival / Q000187:drug effects; D002918:Chymotrypsin / Q000187:drug effects / Q000378:metabolism; D019008:Drug Resistance, Neoplasm / Q000187:drug effects / Q000235:genetics*; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D019169:Jurkat Cells; D020014:K562 Cells / Q000187:drug effects; D015458:Leukemia, T-Cell / Q000188:drug therapy / Q000235:genetics; D016399:Lymphoma, T-Cell / Q000188:drug therapy / Q000235:genetics; D054198:Precursor Cell Lymphoblastic Leukemia-Lymphoma / Q000188:drug therapy / Q000235:genetics; D046988:Proteasome Endopeptidase Complex / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D011719:Pyrazines / Q000627:therapeutic use*; D025801:Ubiquitin / Q000187:drug effects / Q000378:metabolism
D000970:Antineoplastic Agents; D001897:Boronic Acids; D011719:Pyrazines; D025801:Ubiquitin; D000069286:Bortezomib; D002918:Chymotrypsin; C088874:PSMB5 protein, human; D046988:Proteasome Endopeptidase Complex