Ufuk GÜNDÜZ, Rouhollah KHODADUST, Aktan ALPSOY, Gözde ÜNSOY

Poly (I:C)- and doxorubicin-loaded magnetic dendrimeric nanoparticles affect the apoptosis-related gene expressions in MCF-7 cells

Use of nanoparticles as drug carrier vectors has great potential to circumvent the limitations associated with chemotherapy, including drug resistance and destructive side effects. For this purpose, magnetic generation 4 dendrimeric nanoparticles were prepared to carry chemotherapeutic agent doxorubicin (G 4-DOX) and immune modulator polyinosinic:polycytidylic acid [Poly(I:C)]. As previously reported, DOX and Poly(I:C) was loaded onto G 4 nanoparticles (PIC-G 4-DOX). Cellular internalization study using confocal microscopy demonstrated high levels of cellular internalization of PIC-G 4-DOX nanoparticles by MCF-7 cells. This resulted in higher efficacy of PIC-G 4-DOX nanoparticles in killing MCF-7 breast cancer cells. Alteration in the expression levels of selected genes was determined by RT-qPCR analyses. Proapoptotic NOXA, PUMA, and BAX genes were upregulated, and SURVIVIN, APOLLON, and BCL-2 genes were downregulated, indicating the cell-killing effectiveness of PIC-G 4-DOX nanoparticles. Gene expression analysis provided some insights into the possible molecular mechanisms on cytotoxicity of DOX and Poly(I:C) delivered through G 4 magnetic nanoparticles. The results demonstrated that PIC-G 4-DOX can be useful for targeted delivery affecting apoptotic pathways, resulting in an advanced degree of cancer-cell-killing. They are promising for targeting cancer-cells because of their stability, biocompatibility, higher internalization, and toxicity.

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Abedi-Gaballu F, Dehghan G, Ghaffari M, Yekta R, AbbaspourRavasjani S et al. (2018). PAMAM dendrimers as efficient drug and gene delivery nanosystems for cancer therapy. Applied Materials Today 12: 177-190. doi: 10.1016/j.apmt.2018.05.002
Alessandrini F, Pezzè L, Menendez D, Resnick MA, Ciribilli Y (2018). ETV7-Mediated DNAJC15 repression leads to doxorubicin resistance in breast cancer cells 1. Neoplasia 20 (8): 857-70. doi: 10.1016/j.neo.2018.06.008
An W, Lai H, Zhang Y, Liu M, Lin X et al. (2019). Apoptotic pathway as the therapeutic target for anticancer traditional chinese medicines. Frontiers in Pharmacology 10: 758. doi: 10.3389/ fphar.2019.00758
Asnani A, Zheng B, Liu Y, Wang Y, Chen HH et al. (2018). Highly potent visnagin derivatives inhibit Cyp1 and prevent doxorubicin cardiotoxicity. JCI Insight 3 (1): e96753. doi: 10.1172/jci.insight.96753
Bansal M, Kumar A, Malinee M, Sharma TK (2020). Nanomedicine: Diagnosis, Treatment, and Potential Prospects. Nanoscience in Medicine Vol. 1. Cham, Switzerland: Springer, pp. 297-331.
Bello M, Rodríguez-Fonseca RA, Correa-Basurto J (2020). Complexation of peptide epitopes with G4-PAMAM dendrimer through ligand diffusion molecular dynamic simulations. Journal of Molecular Graphics and Modelling 96: 107514. doi: 10.1016/j.jmgm.2019.107514
Besch R, Poeck H, Hohenauer T, Senft D, Häcker G et al. (2009). Proapoptotic signaling induced by RIG-I and MDA-5 results in type I interferon–independent apoptosis in human melanoma cells. The Journal of Clinical Investigation 119 (8): 2399-2411. doi: 10.1172/JCI37155
Chanphai P, Bekale L, Sanyakamdhorn S, Agudelo D, Bérubé G et al. (2017). PAMAM dendrimers in drug delivery: loading efficacy and polymer morphology. Canadian Journal of Chemistry 95 (9), 891-896. doi: 10.1139/cjc-2017-0115
Chen Y, Wan Y, Wang Y, Zhang H, Jiao Z (2011). Anticancer efficacy enhancement and attenuation of side effects of doxorubicn with titanium dioxide nanoparticles. International Journal of Nanomedicine 6: 2321. doi: 10.2147/IJN.S25460
Chorna I, Datsyuk L, Stoika RS (2005). Expression of BAX, Bad and BCL-2 proteins under x-radiation effect towards human breast carcinoma MCF-7 cells and their doxorubicin-resistant derivatives. Experimental Oncology 27 (3): 196-201.
Da Silva C, Peters GJ, Ossendorp F, Cruz LJ (2017). The potential of multi-compound nanoparticles to bypass drug resistance in cancer. Cancer Chemotherapy and Pharmacology 80 (5): 881- 894. doi: 10.1007/s00280-017-3427-1
Davis JM, Navolanic PM, Weinstein-Oppenheimer CR, Steelman LS, Hu W et al. (2003). Raf-1 and BCL-2 induce distinct and common pathways that contribute to breast cancer drug resistance. Clinical Cancer Research 9 (3): 1161-1170.
De Moraes GN, Vasconcelos FC, Delbue D, Mognol GP, Sternberg C et al. (2013). Doxorubicn induces cell death in breast cancer cells regardless of SURVIVIN and XIAP expression levels. European Journal of Cell Biology 92 (8-9): 247-256. doi: 10.1016/j.ejcb.2013.08.001
Dudgeon C, Peng R, Wang P, Sebastiani A, Yu J et al. (2012). Inhibiting oncogenic signaling by sorafenib activates PUMA via GSK3β and NF-κB to suppress tumor cell growth. Oncogene 31 (46): 4848. doi: 10.1038/onc.2011.644
Faversani A, Vaira V, Moro G. P, Tosi D, Lopergolo A et al. (2014). SURVIVIN family proteins as novel molecular determinants of doxorubbbbicin resistance in organotypic human breast tumors. Breast Cancer Research 16 (3): R55. doi: 10.1186/ bcr3666
Ghavami S, Hashemi M, Ande SR, Yeganeh B, Xiao W et al. (2009). Apoptosis and cancer: mutations within caspase genes. Journal of Medical Genetics 46 (8): 497-510. doi: 10.1136/ jmg.2009.066944
Kato H, Takeuchi O, Sato S, Yoneyama M, Yamamoto M et al. (2006). Differential roles of MDA5 and RIG-I helicases in the recognition of RNA viruses. Nature 441 (7089): 101. doi: 10.1038/nature04734
Kaur D, Jain K, Mehra NK, Kesharwani P, Jain N (2016). A review on comparative study of PPI and PAMAM dendrimers. Journal of Nanoparticle Research 18 (6): 146. doi: 10.1007/s11051-016- 3423-0
Kawai T, Takahashi K, Sato S, Coban C, Kumar H et al. (2005). IPS-1, an adaptor triggering RIG-I-and Mda5-mediated type I interferon induction. Nature Immunology 6 (10): 981. doi: 10.1007/s11051-016-3423-0
Khandare J, Kolhe P, Pillai O, Kannan S, Lieh-Lai M et al. (2005). Synthesis, cellular transport, and activity of polyamidoamine dendrimer− methylprednisolone conjugates. Bioconjugate Chemistry 16 (2): 330-337. doi: 10.1021/bc058064s
Khodadust R, Unsoy G, Gunduz U (2014). Development of poly (I: C) modified doxorbicin loaded magnetic dendrimer nanoparticles for targeted combination therapy. Biomedicine & Pharmacotherapy 68 (8): 979-987. doi: 10.1016/j. biopha.2014.10.009
Khodadust R, Unsoy G, Yalcın S, Gunduz G, Gunduz U (2013). PAMAM dendrimer-coated iron oxide nanoparticles: synthesis and characterization of different generations. Journal of Nanoparticle Research 15 (3). doi: 10.1007/s11051-013-1488-6
Kim SH, Kim J (1972). Lethal effect of adriamycin on the division cycle of HeLa cells. Cancer Research 32 (2): 323-325. doi
Ling W, Wang M, Xiong C, Xie D, Chen Q et al. (2019). Synthesis, surface modification, and applications of magnetic iron oxide nanoparticles. Journal of Materials Research 34 (11): 1828-44. doi: 10.1557/jmr.2019.129
Liu F, Xie ZH, Cai GP, Jiang YY, Bulletin P (2007). The effect of SURVIVIN on multidrug resistance mediated by P-glycoprotein in MCF-7 and its adriamycin resistant cells. Biological and Pharmaceutical Bulletin 30 (12): 2279-2283. doi: 10.1248/ bpb.30.2279
Livak KJ, Schmittgen TD (2001). Analysis of relative gene expression data using real-time quantitative PCR and the 2−ΔΔCT method. Methods 25 (4): 402-408. doi:10.1006/meth.2001.1262
Mirski SE, Gerlach JH, Cole SP (1987). Multidrug resistance in a human small cell lung cancer cell line selected in adriamycin. Cancer Research 47 (10): 2594-2598.
Palmerston Mendes L, Pan J, Torchilin VP (2017). Dendrimers as nanocarriers for nucleic acid and drug delivery in cancer therapy. Molecules 22 (9): 1401. doi: 10.3390/ molecules22091401
Parker-Esquivel B, Flores KJ, Louiselle D, Craig M, Dong L et al. (2012). Association of poly I: C RNA and plasmid DNA onto MnO nanorods mediated by PAMAM. Langmuir 28 (8): 3860- 3870. doi: 10.1021/la203998r
Pedziwiatr-Werbicka E, Fuentes E, Dzmitruk V, Sánchez-Nieves J, Sudas M et al. (2013). Novel ‘SiC’carbosilane dendrimers as carriers for anti-HIV nucleic acids: studies on complexation and interaction with blood cells. Colloids and Surfaces B: Biointerfaces 109: 183-189. doi: 10.1016/j.colsurfb.2013.03.045
Rouhollah K, Pelin M, Serap Y, Gozde U, Ufuk G (2013). Dexorubicin loading, release, and stability of polyamidoamine dendrimercoated magnetic nanoparticles. Journal of Pharmaceutical Sciences 102 (6): 1825-1835. doi: 10.1002/jps.23524
Salvador A, Igartua M, Hernández RM, Pedraz J (2012). Combination of immune stimulating adjuvants with poly (lactide-co-glycolide) microspheres enhances the immune response of vaccines. Vaccine 30 (3): 589-596. doi: 10.1016/j. vaccine.2011.11.057
Seth RB, Sun L, Ea CK, Chen ZJ (2005). Identification and characterization of MAVS, a mitochondrial antiviral signaling protein that activates NF-κB and IRF3. Cell 122 (5): 669-682. doi:10.1016/j.cell.2005.08.012
Sheehan J, Cruickshank P, Boshart G (1961). A convenient synthesis of water-soluble carbodiimides. The Journal of Organic Chemistry 26 (7): 2525-2528. doi: 10.1021/jo01351a600
Singal P, Li T, Kumar D, Danelisen I, Iliskovic N (2000). Adriamycininduced heart failure: mechanisms and modulation. Molecular and Cellular Biochemistry 207 (1-2): 77-86. doi: 10.1023/A:1007094214460
Strasser A, Cory S, Adams JM (2011). Deciphering the rules of programmed cell death to improve therapy of cancer and other diseases. The EMBO Journal 30 (18): 3667-3683. doi: 10.1038/ emboj.2011.307
Sun R, Zhang Y, Lv Q, Liu B, Jin M et al. (2011). Toll-like receptor 3 (TLR3) induces apoptosis via death receptors and mitochondria by up-regulating the transactivating p63 isoform α (TAP63α). Journal of Biological Chemistry 286 (18): 15918-15928.
Sui B, Xu H, Jin J, Gou J, Liu J et al. (2014). Self-assembled micelles composed of doxorubicin conjugated Y-shaped PEG-poly (glutamic acid) 2 copolymers via hydrazone linkers. Molecules 19 (8): 11915-11932. doi: 10.3390/molecules190811915
Svenson S, Tomalia DA (2012). Dendrimers in biomedical applications—reflections on the field. Advanced Drug Delivery Reviews 64: 102-115. doi: 10.1016/j.addr.2012.09.030
Vince JE, Tschopp J (2010). IRF‐3 partners BAX in a viral‐induced dance macabre. The EMBO Journal 29 (10): 1627-1628. doi: 10.1038/emboj.2010.79
Vivek R, Babu VN, Thangam R, Subramanian K, Kannan S (2013). pH-responsive drug delivery of chitosan nanoparticles as Tamoxifen carriers for effective anti-tumor activity in breast cancer cells. Colloids and Surfaces B: Biointerfaces 111: 117- 123. doi: 10.1016/j.colsurfb.2013.05.018
Vivek R, Thangam R, Nipunbabu V, Ponraj T, Kannan SJ (2014). Oxaliplatin-chitosan nanoparticles induced intrinsic apoptotic signaling pathway: a “smart” drug delivery system to breast cancer cell therapy. International Journal of Biological Macromolecules 65: 289-297. doi: 10.1016/j. ijbiomac.2014.01.054
Yuan MM, Xu YY, Chen L, Li XY, Qin J et al. (2015). TLR3 expression correlates with apoptosis, proliferation and angiogenesis in hepatocellular carcinoma and predicts prognosis. BMC Cancer 15 (1): 245. doi: 10.1186/s12885-015-1262-5
Yacoub TJ, Reddy AS, Szleifer I (2011). Structural effects and translocation of doxorubicin in a DPPC/Chol bilayer: the role of cholesterol. Biophysical Journal 101 (2): 378-385. doi: 10.1016/j.bpj.2011.06.015
Yoneda K, Sugimoto K, Shiraki K, Tanaka J, Beppu T et al. (2008). Dual topology of functional Toll-like receptor 3 expression in human hepatocellular carcinoma: differential signaling mechanisms of TLR3-induced NF-κB activation and apoptosis. International Journal of Oncology 33 (5): 929-936. doi: 10.3892/ijo_00000080
Yoneyama M, Kikuchi M, Natsukawa T, Shinobu N, Imaizumi T et al. (2004). The RNA helicase RIG-I has an essential function in double-stranded RNA-induced innate antiviral responses. Nature Immunology 5 (7): 730. doi: 10.1038/ni1087
Zhang S, Tang W, Weng S, Liu X, Rao B et al. (2014). APOLLON modulates chemosensitivity in human esophageal squamous cell carcinoma. Oncotarget 5 (16): 7183. doi: 10.18632/ oncotarget.2293