Acquired Resistance to Peloruside A and Laulimalide is Associated with Downregulation of Vimentin in Human Ovarian Carcinoma Cells

ABSTRACT

Purpose

Acquired β-tubulin alterations in human ovarian carcinoma 1A9 cells were previously shown to confer resistance to the microtubule stabilizing agents peloruside A (PLA) and laulimalide (LAU). We examined the proteome of resistant cells to see what other protein changes occurred as a result of the acquired drug resistance.

Methods

Two-dimensional differential in-gel electrophoresis was performed to explore differentially expressed proteins in the resistant 1A9-R1 (R1) and 1A9-L4 (L4) cells. The proteins on the gels were identified by MALDI-TOF MS, and altered protein abundance was confirmed by Western blotting and immunocytochemistry. Vimentin expression was restored in vimentin-deficient L4 cells by transfecting a full-length human vimentin cDNA, and sensitivity to PLA and LAU were tested using an MTT cell proliferation assay.

Results

Proteomic analysis identified several proteins that were significantly altered in the resistant cells relative to the parental 1A9 cells. Using Western blotting and immunocytochemistry, a decreased vimentin abundance in the L4 cells was validated. Vimentin levels were unchanged in PLA-resistant R1 cells and paclitaxel/epothilone-resistant derivatives of 1A9 cells. Vimentin cDNA transfection into L4 cells partially restored PLA and LAU sensitivity.

Conclusions

Downregulation of vimentin contributes to the resistance of 1A9 cells to the microtubule stabilizing agents, PLA and LAU.

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Abbreviations

2D-DIGE:

two-dimensional differential in-gel electrophoresis

BVA:

biological variation analysis

DAPI:

4′,6-diamidino-2-phenylindole

DOX:

doxorubicin

IXA:

ixabepilone

LAU:

laulimalide

MALDI-TOF:

matrix assisted laser desorption/ionization time of flight

P-gp:

P-glycoprotein

PLA:

peloruside A

PTX:

paclitaxel

VDP:

vimentin degradation products

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ACKNOWLEDGMENTS AND DISCLOSURES

The authors thank Dr. Paraskevi Giannakakou for kindly providing the 1A9 and the resistant L4, PTX-10, and A8 cell lines. This research was supported by grants to J.H.M from the Cancer Society of New Zealand, the Wellington Medical Research Foundation, and Victoria University of Wellington.

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Correspondence to John H. Miller.

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Kanakkanthara, A., Rawson, P., Northcote, P.T. et al. Acquired Resistance to Peloruside A and Laulimalide is Associated with Downregulation of Vimentin in Human Ovarian Carcinoma Cells. Pharm Res 29, 3022–3032 (2012). https://doi.org/10.1007/s11095-012-0773-x

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KEY WORDS

  • cancer resistance
  • laulimalide
  • ovarian carcinoma cells
  • peloruside A
  • vimentin