Abstract
Purpose
The retinoic acid signaling pathway, crucial for differentiation, is silenced by epigenetic mechanisms in many cancers. Epigenetically active, chromatin-modifying agents offer a novel treatment approach, by reactivating aberrantly silenced genes in tumor cells and by sensitizing them to subsequent treatments. We hypothesized that the treatment of non-small cell lung cancer (NSCLC) cells with a histone deacetylase (HDAC) inhibitor may prime them to the antiproliferative and differentiating activity of all-trans retinoic acid.
Methods
The NSCLC cell lines A549, NCI-H460 and HCC827 were treated with ATRA (2 µM) and the pan-HDAC inhibitor panobinostat (LBH589; 10–35 nM).
Results
While treatment with ATRA alone showed only very modest effects, panobinostat reduced cellular proliferation by at least 50 %. Notably, the combination of panobinostat and ATRA had additive and synergistic effects, respectively, on growth inhibition and differentiation, with almost no cytotoxicity. Effects were strongest in A549, followed by the EGFR-mutant HCC827, and least pronounced in NCI-H460. Global histone H3 acetylation was strongly induced by panobinostat; interestingly, ATRA alone had also an effect on histone acetylation, which was synergistically enhanced when the HDAC inhibitor was added. The combination of the two drugs additively decreased expression of phospho-ERK and phospho-AKT, whereas p53 and p21CIP1/WAF1 proteins were both induced.
Conclusion
Panobinostat sensitized, to varying degrees, all three cell lines to the antiproliferative and differentiating effects of ATRA, with synergistic histone H3 acetylation. Combination therapy with an epigenetic drug and ATRA may offer an alternative to aggressive chemotherapy even in primary ATRA-insensitive tumors, such as adenocarcinomas of the lung.
Similar content being viewed by others
References
Aberle H, Schwartz H, Kemler R (1996) Cadherin–catenin complex: protein interactions and their implications for cadherin function. J Cell Biochem 61:514–523. doi:10.1002/(SICI)1097-4644(19960616)61:4<514:AID-JCB4>3.0.CO;2-R
Arrieta O, la Rosa CHG-D, Aréchaga-Ocampo E et al (2010) Randomized phase II trial of all-trans-retinoic acid with chemotherapy based on paclitaxel and cisplatin as first-line treatment in patients with advanced non-small-cell lung cancer. J Clin Oncol 28:3463–3471. doi:10.1200/JCO.2009.26.6452
Blagitko-Dorfs N, Jiang Y, Duque-Afonso J et al (2013) Epigenetic priming of AML blasts for all-trans retinoic acid-induced differentiation by the HDAC class-I selective inhibitor entinostat. PLoS ONE 8:e75258. doi:10.1371/journal.pone.0075258
Brabender J, Metzger R, Salonga D et al (2005) Comprehensive expression analysis of retinoic acid receptors and retinoid X receptors in non-small cell lung cancer: implications for tumor development and prognosis. Carcinogenesis 26:525–530. doi:10.1093/carcin/bgi006
Bremnes RM, Veve R, Gabrielson E et al (2002) High-throughput tissue microarray analysis used to evaluate biology and prognostic significance of the E-cadherin pathway in non-small-cell lung cancer. J Clin Oncol 20:2417–2428. doi:10.1200/JCO.2002.08.159
Costa DB, Halmos B, Kumar A et al (2007) BIM mediates EGFR tyrosine kinase inhibitor-induced apoptosis in lung cancers with oncogenic EGFR mutations. PLoS Med 4:e315. doi:10.1371/journal.pmed.0040315
Dauphin M, Barbe C, Lemaire S et al (2013) Vimentin expression predicts the occurrence of metastases in non small cell lung carcinomas. Lung Cancer 81:117–122. doi:10.1016/j.lungcan.2013.03.011
Degos L (1992) All-trans-retinoic acid treatment and retinoic acid receptor alpha gene rearrangement in acute promyelocytic leukemia: a model for differentiation therapy. Int J Cell Cloning 10:63–69. doi:10.1002/stem.5530100202
Dmitriev AA, Kashuba VI, Haraldson K et al (2012) Genetic and epigenetic analysis of non-small cell lung cancer with NotI-microarrays. Epigenetics 7:502–513. doi:10.4161/epi.19801
Edwards A, Li J, Atadja P et al (2007) Effect of the histone deacetylase inhibitor LBH589 against epidermal growth factor receptor-dependent human lung cancer cells. Mol Cancer Ther 6:2515–2524. doi:10.1158/1535-7163.MCT-06-0761
Friday BB, Anderson SK, Buckner J et al (2012) Phase II trial of vorinostat in combination with bortezomib in recurrent glioblastoma: a north central cancer treatment group study. Neuro-Oncol 14:215–221. doi:10.1093/neuonc/nor198
Garcia-Manero G, Jabbour E, Borthakur G et al (2013) Randomized open-label phase II study of decitabine in patients with low- or intermediate-risk myelodysplastic syndromes. J Clin Oncol 31:2548–2553. doi:10.1200/JCO.2012.44.6823
Gazdar AF, Carney DN, Russell EK et al (1980) Establishment of continuous, clonable cultures of small-cell carcinoma of the lung which have amine precursor uptake and decarboxylation cell properties. Cancer Res 40:3502–3507
George P, Bali P, Annavarapu S et al (2005) Combination of the histone deacetylase inhibitor LBH589 and the hsp90 inhibitor 17-AAG is highly active against human CML-BC cells and AML cells with activating mutation of FLT-3. Blood 105:1768–1776. doi:10.1182/blood-2004-09-3413
Grignani F, De Matteis S, Nervi C et al (1998) Fusion proteins of the retinoic acid receptor-alpha recruit histone deacetylase in promyelocytic leukaemia. Nature 391:815–818. doi:10.1038/35901
Guo A, Villen J, Kornhauser J et al (2008) Signaling networks assembled by oncogenic EGFR and c-Met. Proc Natl Acad Sci 105:692–697. doi:10.1073/pnas.0707270105
Jemal A, Bray F, Center MM et al (2011) Global cancer statistics. CA Cancer J Clin 61:69–90. doi:10.3322/caac.20107
Jones DR, Moskaluk CA, Gillenwater HH et al (2012) Phase I trial of induction histone deacetylase and proteasome inhibition followed by surgery in non-small cell lung cancer. J Thorac Oncol Off Publ Int Assoc Study Lung Cancer 7:1683–1690. doi:10.1097/JTO.0b013e318267928d
Kantarjian HM, Thomas XG, Dmoszynska A et al (2012) Multicenter, randomized, open-label, phase III trial of decitabine versus patient choice, with physician advice, of either supportive care or low-dose cytarabine for the treatment of older patients with newly diagnosed acute myeloid leukemia. J Clin Oncol 30:2670–2677. doi:10.1200/JCO.2011.38.9429
Lee H-L, Chung T-S, Ting L-L et al (2012) EGFR mutations are associated with favorable intracranial response and progression-free survival following brain irradiation in non-small cell lung cancer patients with brain metastases. Radiat Oncol 7:181. doi:10.1186/1748-717X-7-181
Lieber M, Smith B, Szakal A et al (1976) A continuous tumor-cell line from a human lung carcinoma with properties of type II alveolar epithelial cells. Int J Cancer 17:62–70
Lo-Coco F, Avvisati G, Vignetti M et al (2013) Retinoic acid and arsenic trioxide for acute promyelocytic leukemia. N Engl J Med 369:111–121. doi:10.1056/NEJMoa1300874
Manna SK, Aggarwal BB (2000) All-trans-retinoic acid upregulates TNF receptors and potentiates TNF-induced activation of nuclear factors-kappaB, activated protein-1 and apoptosis in human lung cancer cells. Oncogene 19:2110–2119. doi:10.1038/sj.onc.1203547
Munker M, Munker R, Saxton RE, Koeffler HP (1987) Effect of recombinant monokines, lymphokines, and other agents on clonal proliferation of human lung cancer cell lines. Cancer Res 47:4081–4085
Nouzova M, Holtan N, Oshiro MM et al (2004) Epigenomic changes during leukemia cell differentiation: analysis of histone acetylation and cytosine methylation using CpG Island microarrays. J Pharmacol Exp Ther 311:968–981. doi:10.1124/jpet.104.072488
Nowak D, Stewart D, Koeffler HP (2009) Differentiation therapy of leukemia: 3 decades of development. Blood 113:3655–3665. doi:10.1182/blood-2009-01-198911
Ohira T, Gemmill RM, Ferguson K et al (2003) WNT7a induces E-cadherin in lung cancer cells. Proc Natl Acad Sci USA 100:10429–10434. doi:10.1073/pnas.1734137100
Ramalingam SS, Maitland ML, Frankel P et al (2010) Carboplatin and paclitaxel in combination with either vorinostat or placebo for first-line therapy of advanced non-small-cell lung cancer. J Clin Oncol 28:56–62. doi:10.1200/JCO.2009.24.9094
Reimer P, Chawla S (2013) Long-term complete remission with belinostat in a patient with chemotherapy refractory peripheral t-cell lymphoma. J Hematol Oncol 6:69. doi:10.1186/1756-8722-6-69
Renshaw J, Taylor KR, Bishop R et al (2013) Dual blockade of the PI3K/AKT/mTOR (AZD8055) and RAS/MEK/ERK (AZD6244) pathways synergistically inhibits rhabdomyosarcoma cell growth in vitro and in vivo. Clin Cancer Res 19:5940–5951. doi:10.1158/1078-0432.CCR-13-0850
Soltermann A, Tischler V, Arbogast S et al (2008) Prognostic significance of epithelial–mesenchymal and mesenchymal–epithelial transition protein expression in non-small cell lung cancer. Clin Cancer Res 14:7430–7437. doi:10.1158/1078-0432.CCR-08-0935
Tarhini AA, Zahoor H, McLaughlin B et al (2013) Phase I trial of carboplatin and etoposide in combination with panobinostat in patients with lung cancer. Anticancer Res 33:4475–4481
Virmani AK, Fong KM, Kodagoda D et al (1998) Allelotyping demonstrates common and distinct patterns of chromosomal loss in human lung cancer types. Genes Chromosomes Cancer 21:308–319. doi:10.1002/(SICI)1098-2264(199804)21:4<308:AID-GCC4>3.0.CO;2-2
Wang S, Han X, Wang J et al (2014) Antitumor effects of a novel chromosome region maintenance 1 (CRM1) inhibitor on non-small cell lung cancer cells in vitro and in mouse tumor xenografts. PLoS ONE 9:e89848. doi:10.1371/journal.pone.0089848
Yamashita F, Azuma K, Yoshida T et al (2013) Prognostic value of EGFR mutation and ERCC1 in patients with non-small cell lung cancer undergoing platinum-based chemotherapy. PLoS ONE 8:e71356. doi:10.1371/journal.pone.0071356
Zhang Z, Lee JC, Lin L et al (2012) Activation of the AXL kinase causes resistance to EGFR-targeted therapy in lung cancer. Nat Genet 44:852–860. doi:10.1038/ng.2330
Acknowledgments
We thank Tobias Ma for technical assistance and Gregor Klaus, Dr. Nadja Blagitko-Dorfs, Dr. Milena Pantic, Dr. Ralph Fritsch and Dr. Dietmar Pfeifer for helpful discussions. This work was supported by the German Research Foundation (DFG, CRC 992 MEDEP, Project C-04).
Conflict of interest
The authors have no potential conflicts of interest.
Author information
Authors and Affiliations
Corresponding author
Electronic supplementary material
Below is the link to the electronic supplementary material.
Rights and permissions
About this article
Cite this article
Greve, G., Schiffmann, I. & Lübbert, M. Epigenetic priming of non-small cell lung cancer cell lines to the antiproliferative and differentiating effects of all-trans retinoic acid. J Cancer Res Clin Oncol 141, 2171–2180 (2015). https://doi.org/10.1007/s00432-015-1987-1
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00432-015-1987-1