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BMC Cancer
Published in: BMC Cancer 1/2015

Open Access 01-12-2015 | Research article

Combined inhibition of the cell cycle related proteins Wee1 and Chk1/2 induces synergistic anti-cancer effect in melanoma

Authors: Gry Irene Magnussen, Elisabeth Emilsen, Karianne Giller Fleten, Birgit Engesæter, Viola Nähse-Kumpf, Roar Fjær, Ana Slipicevic, Vivi Ann Flørenes

Published in: BMC Cancer | Issue 1/2015

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Abstract

Background

Malignant melanoma has an increasing incidence rate and the metastatic disease is notoriously resistant to standard chemotherapy. Loss of cell cycle checkpoints is frequently found in many cancer types and makes the cells reliant on compensatory mechanisms to control progression. This feature may be exploited in therapy, and kinases involved in checkpoint regulation, such as Wee1 and Chk1/2, have thus become attractive therapeutic targets.

Methods

In the present study we combined a Wee1 inhibitor (MK1775) with Chk1/2 inhibitor (AZD7762) in malignant melanoma cell lines grown in vitro (2D and 3D cultures) and in xenografts models.

Results

Our in vitro studies showed that combined inhibition of Wee1 and Chk1/2 synergistically decreased viability and increased apoptosis (cleavage of caspase 3 and PARP), which may be explained by accumulation of DNA-damage (increased expression of γ-H2A.X) - and premature mitosis of S-phase cells. Compared to either inhibitor used as single agents, combined treatment reduced spheroid growth and led to greater tumour growth inhibition in melanoma xenografts.

Conclusions

These data provide a rationale for further evaluation of the combination of Wee1 and Chk1/2 inhibitors in malignant melanoma.
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Literature
1.
2.
3.
Flaherty KT, Puzanov I, Kim KB, Ribas A, McArthur GA, Sosman JA, et al. Inhibition of mutated, activated BRAF in metastatic melanoma. N Engl J Med. 2010;363:809–19.CrossRefPubMedPubMedCentral
4.
Hodi FS, O’Day SJ, McDermott DF, Weber RW, Sosman JA, Haanen JB, et al. Improved survival with ipilimumab in patients with metastatic melanoma. N Engl J Med. 2010;363:711–23.CrossRefPubMedPubMedCentral
5.
Watanabe N, Broome M, Hunter T. Regulation of the human WEE1Hu CDK tyrosine 15-kinase during the cell cycle. EMBO J. 1995;14:1878–91.PubMedPubMedCentral
6.
Magnussen GI, Holm R, Emilsen E, Rosnes AK, Slipicevic A, Florenes VA. High expression of Wee1 is associated with poor disease-free survival in malignant melanoma: potential for targeted therapy. PLoS One. 2012;7, e38254.CrossRefPubMedPubMedCentral
7.
Magnussen GI, Hellesylt E, Nesland JM, Trope CG, Florenes VA, Holm R. High expression of wee1 is associated with malignancy in vulvar squamous cell carcinoma patients. BMC Cancer. 2013;13:288.CrossRefPubMedPubMedCentral
8.
Mir SE, De Witt Hamer PC, Krawczyk PM, Balaj L, Claes A, Niers JM, et al. In silico analysis of kinase expression identifies WEE1 as a gatekeeper against mitotic catastrophe in glioblastoma. Cancer Cell. 2010;18:244–57.CrossRefPubMedPubMedCentral
9.
PosthumaDeBoer J, Wurdinger T, Graat HC, van Beusechem V, Helder MN, van Royen BJ, et al. WEE1 inhibition sensitizes osteosarcoma to radiotherapy. BMC Cancer. 2011;11:156.CrossRefPubMedPubMedCentral
10.
Slipicevic A, Holth A, Hellesylt E, Trope CG, Davidson B, Florenes VA. Wee1 is a novel independent prognostic marker of poor survival in post-chemotherapy ovarian carcinoma effusions. Gynecol Oncol. 2014;135:118–24.CrossRefPubMed
11.
Murrow LM, Garimella SV, Jones TL, Caplen NJ, Lipkowitz S. Identification of WEE1 as a potential molecular target in cancer cells by RNAi screening of the human tyrosine kinome. Breast Cancer Res Treat. 2010;122:347–57.CrossRefPubMed
12.
Rajeshkumar NV, De OE, Ottenhof N, Watters J, Brooks D, Demuth T, et al. MK-1775, a potent Wee1 inhibitor, synergizes with gemcitabine to achieve tumour regressions, selectively in p53-deficient pancreatic cancer xenografts. Clin Cancer Res. 2011;17:2799–806.CrossRefPubMedPubMedCentral
13.
Davies KD, Cable PL, Garrus JE, Sullivan FX, von Carlowitz I, Huerou YL, et al. Chk1 inhibition and Wee1 inhibition combine synergistically to impede cellular proliferation. Cancer Biol Ther. 2011;12:788–96.CrossRefPubMed
14.
Hirai H, Arai T, Okada M, Nishibata T, Kobayashi M, Sakai N, et al. MK-1775, a small molecule Wee1 inhibitor, enhances anti-tumour efficacy of various DNA-damaging agents, including 5-fluorouracil. Cancer Biol Ther. 2010;9:514–22.CrossRefPubMed
15.
16.
Aarts M, Sharpe R, Garcia-Murillas I, Gevensleben H, Hurd MS, Shumway SD, et al. Forced mitotic entry of S-Phase cells as a therapeutic strategy induced by inhibition of WEE1. Cancer Discov 2012;2:524–539.
17.
Bartek J, Lukas J. Chk1 and Chk2 kinases in checkpoint control and cancer. Cancer Cell. 2003;3:421–9.CrossRefPubMed
18.
19.
O’Connell MJ, Raleigh JM, Verkade HM, Nurse P. Chk1 is a wee1 kinase in the G2 DNA damage checkpoint inhibiting cdc2 by Y15 phosphorylation. EMBO J. 1997;16:545–54.CrossRefPubMedPubMedCentral
20.
Carrassa L, Chila R, Lupi M, Ricci F, Celenza C, Mazzoletti M, et al. Combined inhibition of Chk1 and Wee1: in vitro synergistic effect translates to tumour growth inhibition in vivo. Cell Cycle. 2012;11:2507–17.CrossRefPubMed
21.
Chaudhuri L, Vincelette ND, Koh BD, Naylor RM, Flatten KS, Peterson KL, et al. CHK1 and WEE1 inhibition combine synergistically to enhance therapeutic efficacy in acute myeloid leukemia ex vivo. Haematologica. 2014;99:688–96.CrossRefPubMedPubMedCentral
22.
Florenes VA, Lu C, Bhattacharya N, Rak J, Sheehan C, Slingerland JM, et al. Interleukin-6 dependent induction of the cyclin dependent kinase inhibitor p21WAF1/CIP1 is lost during progression of human malignant melanoma. Oncogene. 1999;18:1023–32.CrossRefPubMed
23.
Herlyn M, Balaban G, Bennicelli J, Guerry D, Halaban R, Herlyn D, et al. Primary melanoma cells of the vertical growth phase: similarities to metastatic cells. J Natl Cancer Inst. 1985;74:283–9.PubMed
24.
Fodstad O, Kjonniksen I, Aamdal S, Nesland JM, Boyd MR, Pihl A. Extrapulmonary, tissue-specific metastasis formation in nude mice injected with FEMX-I human melanoma cells. Cancer Res. 1988;48:4382–8.PubMed
25.
Lai F, Jiang CC, Farrelly ML, Zhang XD, Hersey P. Evidence for upregulation of Bim and the splicing factor SRp55 in melanoma cells from patients treated with selective BRAF inhibitors. Melanoma Res. 2012;22:244–51.CrossRefPubMed
26.
Chou TC. Drug combination studies and their synergy quantification using the Chou-Talalay method. Cancer Res. 2010;70:440–6.CrossRefPubMed
27.
Chou TC. Theoretical basis, experimental design, and computerized simulation of synergism and antagonism in drug combination studies. Pharmacol Rev. 2006;58:621–81.CrossRefPubMed
28.
Krutzik PO, Nolan GP. Fluorescent cell barcoding in flow cytometry allows high-throughput drug screening and signaling profiling. Nat Methods. 2006;3:361–8.CrossRefPubMed
29.
Hirai H, Iwasawa Y, Okada M, Arai T, Nishibata T, Kobayashi M, et al. Small-molecule inhibition of Wee1 kinase by MK-1775 selectively sensitizes p53-deficient tumour cells to DNA-damaging agents. Mol Cancer Ther. 2009;8:2992–3000.CrossRefPubMed
30.
Zabludoff SD, Deng C, Grondine MR, Sheehy AM, Ashwell S, Caleb BL, et al. AZD7762, a novel checkpoint kinase inhibitor, drives checkpoint abrogation and potentiates DNA-targeted therapies. Mol Cancer Ther. 2008;7:2955–66.CrossRefPubMed
31.
Chitcholtan K, Sykes PH, Evans JJ. The resistance of intracellular mediators to doxorubicin and cisplatin are distinct in 3D and 2D endometrial cancer. J Transl Med. 2012;10:38.CrossRefPubMedPubMedCentral
32.
Mehta G, Hsiao AY, Ingram M, Luker GD, Takayama S. Opportunities and challenges for use of tumour spheroids as models to test drug delivery and efficacy. J Control Release. 2012;164:192–204.CrossRefPubMedPubMedCentral
33.
Vinci M, Gowan S, Boxall F, Patterson L, Zimmermann M, Court W, et al. Advances in establishment and analysis of three-dimensional tumour spheroid-based functional assays for target validation and drug evaluation. BMC Biol. 2012;10:29.CrossRefPubMedPubMedCentral
34.
Mah LJ, El-Osta A, Karagiannis TC. gammaH2AX: a sensitive molecular marker of DNA damage and repair. Leukemia. 2010;24:679–86.CrossRefPubMed
35.
Kreahling JM, Gemmer JY, Reed D, Letson D, Bui M, Altiok S. MK1775, a selective Wee1 inhibitor, shows single-agent antitumour activity against sarcoma cells. Mol Cancer Ther. 2012;11:174–82.CrossRefPubMed
36.
Haarberg HE, Paraiso KH, Wood E, Rebecca VW, Sondak VK, Koomen JM, et al. Inhibition of Wee1, AKT, and CDK4 underlies the efficacy of the HSP90 inhibitor XL888 in an in vivo model of NRAS-mutant melanoma. Mol Cancer Ther. 2013;12:901–12.CrossRefPubMedPubMedCentral
37.
Guertin AD, Martin MM, Roberts B, Hurd M, Qu X, Miselis NR, et al. Unique functions of CHK1 and WEE1 underlie synergistic anti-tumour activity upon pharmacologic inhibition. Cancer Cell Int. 2012;12:45.CrossRefPubMedPubMedCentral
38.
McNeely S, Conti C, Sheikh T, Patel H, Zabludoff S, Pommier Y, et al. Chk1 inhibition after replicative stress activates a double strand break response mediated by ATM and DNA-dependent protein kinase. Cell Cycle. 2010;9:995–1004.CrossRefPubMed
39.
Russell MR, Levin K, Rader J, Belcastro L, Li Y, Martinez D, et al. Combination therapy targeting the Chk1 and Wee1 kinases shows therapeutic efficacy in neuroblastoma. Cancer Res. 2013;73:776–84.CrossRefPubMed
40.
Moser R, Xu C, Kao M, Annis J, Lerma LA, Schaupp CM, et al. Functional kinomics identifies candidate therapeutic targets in head and neck cancer. Clin Cancer Res. 2014;20:4274–88.CrossRefPubMedPubMedCentral
41.
Zenvirt S, Kravchenko-Balasha N, Levitzki A. Status of p53 in human cancer cells does not predict efficacy of CHK1 kinase inhibitors combined with chemotherapeutic agents. Oncogene. 2010;29:6149–59.CrossRefPubMed
42.
Box NF, Vukmer TO, Terzian T. Targeting p53 in melanoma. Pigment Cell Melanoma Res. 2014;27:8–10.CrossRefPubMed
43.
Brooks K, Oakes V, Edwards B, Ranall M, Leo P, Pavey S, et al. A potent Chk1 inhibitor is selectively cytotoxic in melanomas with high levels of replicative stress. Oncogene. 2013;32:788–96.CrossRefPubMed
44.
Vriend LE, De Witt Hamer PC, Van Noorden CJ, Wurdinger T. WEE1 inhibition and genomic instability in cancer. Biochim Biophys Acta. 1836;2013:227–35.
45.
46.
47.
Sausville E, Lorusso P, Carducci M, Carter J, Quinn MF, Malburg L, et al. Phase I dose-escalation study of AZD7762, a checkpoint kinase inhibitor, in combination with gemcitabine in US patients with advanced solid tumours. Cancer Chemother Pharmacol. 2014;73:539–49.CrossRefPubMedPubMedCentral
48.
Seto T, Esaki T, Hirai F, Arita S, Nosaki K, Makiyama A, et al. Phase I, dose-escalation study of AZD7762 alone and in combination with gemcitabine in Japanese patients with advanced solid tumours. Cancer Chemother Pharmacol. 2013;72:619–27.CrossRefPubMed
49.
Chen T, Stephens PA, Middleton FK, Curtin NJ. Targeting the S and G2 checkpoint to treat cancer. Drug Discov Today. 2012;17:194–202.CrossRefPubMed
50.
Goto H, Izawa I, Li P, Inagaki M. Novel regulation of checkpoint kinase 1: Is checkpoint kinase 1 a good candidate for anti-cancer therapy? Cancer Sci. 2012;103:1195–200.CrossRefPubMed
51.
Takai H, Tominaga K, Motoyama N, Minamishima YA, Nagahama H, Tsukiyama T, et al. Aberrant cell cycle checkpoint function and early embryonic death in Chk1(−/−) mice. Genes Dev. 2000;14:1439–47.PubMedPubMedCentral
52.
Nakanishi M, Ando H, Watanabe N, Kitamura K, Ito K, Okayama H, et al. Identification and characterization of human Wee1B, a new member of the Wee1 family of Cdk-inhibitory kinases. Genes Cells. 2000;5:839–47.CrossRefPubMed
53.
Sanchez Y, Wong C, Thoma RS, Richman R, Wu Z, Piwnica-Worms H, et al. Conservation of the Chk1 checkpoint pathway in mammals: linkage of DNA damage to Cdk regulation through Cdc25. Science. 1997;277:1497–501.CrossRefPubMed
54.
Beck H, Nahse-Kumpf V, Larsen MS, O’Hanlon KA, Patzke S, Holmberg C, et al. Cyclin-Dependent kinase suppression by WEE1 kinase protects the genome through control of replication initiation and nucleotide consumption. Mol Cell Biol. 2012;32:4226–36.CrossRefPubMedPubMedCentral
55.
Shimada M, Niida H, Zineldeen DH, Tagami H, Tanaka M, Saito H, et al. Chk1 is a histone H3 threonine 11 kinase that regulates DNA damage-induced transcriptional repression. Cell. 2008;132:221–32.CrossRefPubMed
56.
Beck H, Nahse V, Larsen MS, Groth P, Clancy T, Lees M, et al. Regulators of cyclin-dependent kinases are crucial for maintaining genome integrity in S phase. J Cell Biol. 2010;188:629–38.CrossRefPubMedPubMedCentral
57.
Syljuasen RG, Sorensen CS, Hansen LT, Fugger K, Lundin C, Johansson F, et al. Inhibition of human Chk1 causes increased initiation of DNA replication, phosphorylation of ATR targets, and DNA breakage. Mol Cell Biol. 2005;25:3553–62.CrossRefPubMedPubMedCentral
58.
Sorensen CS, Syljuasen RG. Safeguarding genome integrity: the checkpoint kinases ATR, CHK1 and WEE1 restrain CDK activity during normal DNA replication. Nuck Acids Res. 2011;40:477–86.
Metadata
Title
Combined inhibition of the cell cycle related proteins Wee1 and Chk1/2 induces synergistic anti-cancer effect in melanoma
Authors
Gry Irene Magnussen
Elisabeth Emilsen
Karianne Giller Fleten
Birgit Engesæter
Viola Nähse-Kumpf
Roar Fjær
Ana Slipicevic
Vivi Ann Flørenes
Publication date
01-12-2015
Publisher
BioMed Central
Published in
BMC Cancer / Issue 1/2015
Electronic ISSN: 1471-2407
DOI
https://doi.org/10.1186/s12885-015-1474-8

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