A novel combination treatment for breast cancer cells involving BAPTA-AM and proteasome inhibitor bortezomib

Yerlikaya,Azmi; Erdoğan,Elif; Okur,Emrah; Yerlikaya,Şerife; Savran,Bircan

doi:10.3892/ol.2016.4597

A novel combination treatment for breast cancer cells involving BAPTA-AM and proteasome inhibitor bortezomib

Authors:
- Azmi Yerlikaya
- Elif Erdoğan
- Emrah Okur
- Şerife Yerlikaya
- Bircan Savran
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Affiliations: Department of Medical Biology, Faculty of Medicine, Dumlupınar University, Kütahya 43100, Turkey, Department of Biology, Faculty of Arts and Sciences, Dumlupınar University, Kütahya 43100, Turkey, Department of Biochemistry, Faculty of Arts and Sciences, Dumlupınar University, Kütahya 43100, Turkey, Department of Pediatrics, Faculty of Medicine, Dumlupınar University, Kütahya 43100, Turkey
Published online on: May 17, 2016 https://doi.org/10.3892/ol.2016.4597
Pages: 323-330

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Abstract

Glucose-regulated protein 78 kDa/binding immunoglobulin protein (GRP78/BIP) is a well-known endoplasmic reticulum (ER) chaperone protein regulating ER stress by facilitating protein folding, assembly and Ca2+ binding. GRP78 is also a member of the heat shock protein 70 gene family and induces tumor cell survival and resistance to chemotherapeutics. Bortezomib is a highly specific 26S proteasome inhibitor that has been approved as treatment for patients with multiple myeloma. The present study first examined the dose‑ and time‑dependent effects of bortezomib on GRP78 expression levels in the highly metastatic mouse breast cancer 4T1 cell line using western blot analysis. The analysis results revealed that GRP78 levels were significantly increased by bortezomib at a dose as low as 10 nM. Time‑dependent experiments indicated that the accumulation of GRP78 was initiated after a 24 h incubation period following the addition of 10 nM bortezomib. Subsequently, the present study determined the half maximal inhibitory concentration of intracellular calcium chelator BAPTA‑AM (13.6 µM) on 4T1 cells. The combination effect of BAPTA‑AM and bortezomib on the 4T1 cells was investigated using 3‑(4,5‑dimethylthiazol‑2‑yl)‑2,5‑diphenyltetrazolium bromide and WST‑1 assays and an iCELLigence system. The results revealed that the combination of 10 nM bortezomib + 5 µM BAPTA‑AM is more cytotoxic compared with monotherapies, including 10 nM bortezomib, 1 µM BAPTA‑AM and 5 µM BAPTA‑AM. In addition, the present results revealed that bortezomib + BAPTA‑AM combination causes cell death through the induction of apoptosis. The present results also revealed that bortezomib + BAPTA‑AM combination‑induced apoptosis is associated with a clear increase in the phosphorylation of stress‑activated protein kinase/Jun amino‑terminal kinase SAPK/JNK. Overall, the present results suggest that bortezomib and BAPTA‑AM combination therapy may be a novel therapeutic strategy for breast cancer treatment.

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1	Hendershot LM: The ER function BiP is a master regulator of ER function. Mt Sinai J Med. 71:289–297. 2004.PubMed/NCBI
2	Hendershot LM, Valentine VA, Lee AS, Morris SW and Shapiro DN: Localization of the gene encoding human BiP/GRP78, the endoplasmic reticulum cognate of the HSP70 family, to chromosome 9q34. Genomics. 20:281–284. 1994. View Article : Google Scholar : PubMed/NCBI
3	Murphy ME: The HSP70 family and cancer. Carcinogenesis. 34:1181–1188. 2013. View Article : Google Scholar : PubMed/NCBI
4	Roller C and Maddalo D: The molecular chaperone GRP78/BiP in the development of chemoresistance: Mechanism and possible treatment. Front Pharmacol. 4:102013. View Article : Google Scholar : PubMed/NCBI
5	Qiu W, Kohen-Avramoglu R, Mhapsekar S, Tsai J, Austin RC and Adeli K: Glucosamine-induced endoplasmic reticulum stress promotes ApoB100 degradation: Evidence for Grp78-mediated targeting to proteasomal degradation. Arterioscler Thromb Vasc Biol. 25:571–577. 2005. View Article : Google Scholar : PubMed/NCBI
6	Zhang Y, Tseng CC, Tsai YL, Fu X, Schiff R and Lee AS: Cancer cells resistant to therapy promote cell surface relocalization of GRP78 which complexes with PI3K and enhances PI(3,4,5)P3 production. PLoS One. 8:e800712013. View Article : Google Scholar : PubMed/NCBI
7	Li B, Cheng XL, Yang YP and Li ZQ: GRP78 mediates radiation resistance of a stem cell-like subpopulation within the MCF-7 breast cancer cell line. Oncol Rep. 30:2119–2126. 2013.PubMed/NCBI
8	Dong D, Ko B, Baumeister P, Swenson S, Costa F, Markland F, Stiles C, Patterson JB, Bates SE and Lee AS: Vascular targeting and antiangiogenesis agents induce drug resistance effector GRP78 within the tumor microenvironment. Cancer Res. 65:5785–5791. 2005. View Article : Google Scholar : PubMed/NCBI
9	Zhang J, Jiang Y, Jia Z, Li Q, Gong W, Wang L, Wei D, Yao J, Fang S and Xie K: Association of elevated GRP78 expression with increased lymph node metastasis and poor prognosis in patients with gastric cancer. Clin Exp Metastasis. 23:401–410. 2006. View Article : Google Scholar : PubMed/NCBI
10	Wang HQ, Du ZX, Zhang HY and Gao DX: Different induction of GRP78 and CHOP as a predictor of sensitivity to proteasome inhibitors in thyroid cancer cells. Endocrinology. 148:3258–3270. 2007. View Article : Google Scholar : PubMed/NCBI
11	Mozos A, Roué G, López-Guillermo A, Jares P, Campo E, Colomer D and Martinez A: The expression of the endoplasmic reticulum stress sensor BiP/GRP78 predicts response to chemotherapy and determines the efficacy of proteasome inhibitors in diffuse large b-cell lymphoma. Am J Pathol. 179:2601–2610. 2011. View Article : Google Scholar : PubMed/NCBI
12	Chen LY, Chiang AS, Hung JJ, Hung HI and Lai YK: Thapsigargin-induced grp78 expression is mediated by the increase of cytosolic free calcium in 9l rat brain tumor cells. J Cell Biochem. 78:404–416. 2000. View Article : Google Scholar : PubMed/NCBI
13	Freshney RI: Culture of Animal Cells: A Manual of Basic Technique. Wiley-Liss. Hoboken, NJ: 2005. View Article : Google Scholar
14	Savran B, Yerlikaya A, Erdoğan E and Genç O: Anticancer agent ukrain and bortezomib combination is synergistic in 4T1 breast cancer cells. Anticancer Agents Med Chem. 14:466–472. 2014. View Article : Google Scholar : PubMed/NCBI
15	Yerlikaya A and Erin N: Differential sensitivity of breast cancer and melanoma cells to proteasome inhibitor velcade. Int J Mol Med. 22:817–823. 2008.PubMed/NCBI
16	Oerlemans R, Franke NE, Assaraf YG, Cloos J, van Zantwijk I, Berkers CR, Scheffer GL, Debipersad K, Vojtekova K, Lemos C, et al: Molecular basis of bortezomib resistance: Proteasome subunit beta5 (PSMB5) gene mutation and overexpression of PSMB5 protein. Blood. 112:2489–2499. 2008. View Article : Google Scholar : PubMed/NCBI
17	Chauhan D, Li G, Shringarpure R, Podar K, Ohtake Y, Hideshima T and Anderson KC: Blockade of Hsp27 overcomes bortezomib/proteasome inhibitor PS-341 resistance in lymphoma cells. Cancer Res. 63:6174–6177. 2003.PubMed/NCBI
18	Ni M, Zhang Y and Lee AS: Beyond the endoplasmic reticulum: Atypical GRP78 in cell viability, signalling and therapeutic targeting. Biochem J. 434:181–188. 2011. View Article : Google Scholar : PubMed/NCBI
19	Wang Y, Wang W, Wang S, Wang J, Shao S and Wang Q: Down-regulation of GRP78 is associated with the sensitivity of chemotherapy to VP-16 in small cell lung cancer NCI-H446 cells. BMC Cancer. 8:3722008. View Article : Google Scholar : PubMed/NCBI
20	Dhanasekaran DN and Reddy EP: JNK signaling in apoptosis. Oncogene. 27:6245–6251. 2008. View Article : Google Scholar : PubMed/NCBI
21	Dai Y, Rahmani M and Grant S: Proteasome inhibitors potentiate leukemic cell apoptosis induced by the cyclin-dependent kinase inhibitor flavopiridol through a SAPK/JNK- and NF-kappaB-dependent process. Oncogene. 22:7108–7122. 2003. View Article : Google Scholar : PubMed/NCBI
22	Dai Y, Rahmani M, Pei XY, Dent P and Grant S: Bortezomib and flavopiridol interact synergistically to induce apoptosis in chronic myeloid leukemia cells resistant to imatinib mesylate through both Bcr/Abl-dependent and -independent mechanisms. Blood. 104:509–518. 2004. View Article : Google Scholar : PubMed/NCBI
23	Roué G, Pérez-Galan P, Mozos A, López-Guerra M, Xargay-Torrent S, Rosich L, Saborit-Villarroya I, Normant E, Campo E and Colomer D: The Hsp90 inhibitor IPI-504 overcomes bortezomib resistance in mantle cell lymphoma in vitro and in vivo by down-regulation of the prosurvival ER chaperone BiP/Grp78. Blood. 117:1270–1279. 2011. View Article : Google Scholar : PubMed/NCBI
24	Kern J, Untergasser G, Zenzmaier C, Sarg B, Gastl G, Gunsilius E and Steurer M: GRP-78 secreted by tumor cells blocks the antiangiogenic activity of bortezomib. Blood. 114:3960–3967. 2009. View Article : Google Scholar : PubMed/NCBI
25	Kardosh A, Golden EB, Pyrko P, Uddin J, Hofman FM, Chen TC, Louie SG, Petasis NA and Schönthal AH: Aggravated endoplasmic reticulum stress as a basis for enhanced glioblastoma cell killing by bortezomib in combination with celecoxib or its non-coxib analogue, 2,5-dimethyl-celecoxib. Cancer Res. 68:843–851. 2008. View Article : Google Scholar : PubMed/NCBI