Characterization of bortezomib-adapted I-45 mesothelioma cells

Our plain-English summary of Characterization of bortezomib-adapted I-45 mesothelioma cells, by L. Zhang, J.E. Littlejohn, Y. Cui, et al (Molecular Cancer, 2010).

The study

• Authors: L. Zhang, J.E. Littlejohn, Y. Cui, X. Cao, C. Peddaboina, W.R. Smythe
• Journal: Molecular Cancer
• Published: 2010;9:110
• Cited by: 24 articles, as counted by NIH iCite
• Read it: PubMed · Publisher · Free full text

Abstract

Background: Bortezomib, a proteasome-specific inhibitor, has emerged as a promising cancer therapeutic agent. However, development of resistance to bortezomib may pose a challenge to effective anticancer therapy. Therefore, characterization of cellular mechanisms involved in bortezomib resistance and development of effective strategies to overcome this resistance represent important steps in the advancement of bortezomib-mediated cancer therapy.

Results: The present study reports the development of I-45-BTZ-R, a bortezomib-resistant cell line, from the bortezomib-sensitive mesothelioma cell line I-45. I-45-BTZ-R cells showed no cross-resistance to the chemotherapeutic drugs cisplatin, 5-fluorouracil, and doxorubicin. Moreover, the bortezomib-adapted I-45-BTZ-R cells had decreased growth kinemics and did not over express proteasome subunit beta5 (PSMB5) as compared to parental I-45 cells. I-45-BTZ-R cells and parental I-45 cells showed similar inhibition of proteasome activity, but I-45-BTZ-R cells exhibited much less accumulation of ubiquitinated proteins following exposure to 40 nm bortezomib. Further studies revealed that relatively low doses of bortezomib did not induce an unfolded protein response (UPR) in the bortezomib-adapted cells, while higher doses induced UPR with concomitant cell death, as evidenced by higher expression of the mitochondrial chaperone protein Bip and the endoplasmic reticulum (ER) stress-related pro-apoptotic protein CHOP. In addition, bortezomib exposure did not induce the accumulation of the pro-apoptotic proteins p53, Mcl-1S, and noxa in the bortezomib-adapted cells.

Conclusion: These results suggest that UPR evasion, together with reduced pro-apoptotic gene induction, accounts for bortezomib resistance in the bortezomib-adapted mesothelioma cell line I-45-BTZ-R.

In plain English

This laboratory study used cells grown in dishes, not people, and did not examine asbestos. The authors exposed I-45, a human sarcomatoid mesothelioma cell line, to the cancer drug bortezomib at step-wise increasing concentrations up to 40 nM over 6 months. This produced a drug-resistant line called I-45-BTZ-R. The dose that halved cell viability was 185.5 nM for the resistant cells and 12.1 nM for the original cells. At 40 nM, bortezomib triggered signs of programmed cell death in the original cells but not in the resistant cells. The resistant cells grew more slowly; both lines responded similarly to the chemotherapy drug cisplatin. The authors found no difference between the lines in PSMB5, a part of the proteasome (the cell's protein-disposal machinery). The drug cut proteasome activity in both lines, leaving about 25 to 40% of the starting level. The resistant cells built up fewer ubiquitinated (disposal-tagged) proteins and did not switch on the stress markers Bip and CHOP at 40 nM. They also showed little or no build-up of p53, Mcl-1S or noxa. The authors concluded these changes may account for this line's resistance.

We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

What the study did and found

• Design: Laboratory cell-culture study: development and characterization of a bortezomib-resistant mesothelioma cell line
• Who was studied: The human sarcomatoid type mesothelioma cell line I-45, expressing wild type p53 and obtained from Dr. J. Testa (Fox Chase Institute, Philadelphia, PA), and the bortezomib-resistant line I-45-BTZ-R derived from it; no human subjects
• Exposure: The proteasome-inhibiting cancer drug bortezomib, given at step-wise increasing concentrations reaching 40 nM over 6 months to create the resistant line. No asbestos exposure was studied.
• Main results:
  • The bortezomib concentration that halved cell viability (IC50) was 185.5 plus or minus 0.3 nM for resistant I-45-BTZ-R cells and 12.1 plus or minus 0.4 nM for parental I-45 cells
  • At 40 nM, bortezomib caused caspase-3 and PARP cleavage (markers of apoptosis) in parental I-45 cells but not in I-45-BTZ-R cells
  • In parental I-45 cells, 25 nM bortezomib for 72 hours raised the share of cells in the G2/M phase from 4% to 70.9%. In I-45-BTZ-R cells, the share stayed at 10.5% compared with a basal 10.1%.
  • I-45-BTZ-R cells grew significantly more slowly than I-45 cells (p less than 0.05 at day 2; p less than 0.01 at days 4, 6 and 8)
  • No difference in PSMB5 protein expression was detected between the two cell lines
  • Basal chymotrypsin-like proteasome activity was about 30% lower in I-45-BTZ-R cells than in I-45 cells. Both lines kept about 25 to 40% of their original activity after bortezomib treatment.
  • After 40 nM bortezomib, ubiquitinated proteins rose sharply in I-45 cells. I-45-BTZ-R cells showed lower accumulation, which returned to basal level by 48 or 72 hours.
  • Both cell lines showed similar sensitivity to cisplatin. The authors reported similar sensitivity to 5-fluorouracil and doxorubicin as data not shown.
  • At 40 nM, bortezomib raised Bip and CHOP (ER stress markers) in I-45 cells but not in I-45-BTZ-R cells
  • In I-45-BTZ-R cells, bortezomib at 100 to 400 nM increased Bip and CHOP together with increased apoptosis
  • In bortezomib-treated I-45-BTZ-R cells, silencing CHOP raised cell viability from 53.9 plus or minus 5.4% (control siRNA) to 72.3 plus or minus 2.6% (CHOP-specific siRNA)
  • After bortezomib, noxa was strongly expressed in I-45 cells but barely detectable in I-45-BTZ-R cells
  • At 40 nM, bortezomib caused strong p53 accumulation in parental I-45 cells but only weak accumulation in I-45-BTZ-R cells
  • Mcl-1S rose about 150% at 24 hours in I-45 cells after bortezomib; no change in Mcl-1S was detected in I-45-BTZ-R cells
  • The TPPII inhibitor AAF-CMK at 5, 10 or 20 μM for 48 hours did not reduce I-45-BTZ-R cell viability. Combined with bortezomib, viability remained above 85%.
• The authors' conclusion: The authors concluded that their results support a resistance mechanism independent of PSMB5 expression and proteasome activity. In their view, reduced accumulation of ubiquitinated proteins prevents ER stress and the UPR and reduces stabilization of p53, Mcl-1S and noxa, and these changes may account for bortezomib resistance in this cell line.
• Limitations the authors state:
  • The authors could not exclude that other, unidentified proteases compensated for reduced proteasome function in the resistant cells

Topics

The National Library of Medicine indexes this study under subjects our site covers: Mesothelioma.

Medical terms in this study, explained in our Lexicon: Mesothelioma, Cisplatin, Fluorouracil (5-FU), Doxorubicin.

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