Alginate microspheres encapsulating hox transcript antisense RNA siRNA regulate the Hedgehog-Gli1 pathway to alleviate epidermal growth factor receptor tyrosine kinase inhibitors resistance

Abstract

The long non-coding RNA HOTAIR and the Hedgehog-Gli1 signaling pathway are closely associated with tumor occurrence and drug resistance in various cancers. However, their specific roles in the development of EGFR-TKIs resistance in non-small cell carcinoma remain unclear. To address the issue of EGFR-TKIs resistance, this study utilized the electrospray method to prepare sodium alginate microspheres encapsulating HOTAIR siRNA (SA/HOTAIR siRNA) and investigated its effects on RNA interference (RNAi) in the gefitinib-resistant cell line PC9/GR. Furthermore, the study explored whether HOTAIR could modulate EGFR-TKIs resistance through the Hedgehog-GLi1 signaling pathway. The experimental results showed that sodium alginate (SA) microspheres demonstrated excellent biocompatibility with high encapsulation efficiency and drug-loading capacity, effectively enhancing the silencing efficiency of siRNA. HOTAIR siRNA significantly inhibited the proliferation, migration, and invasion abilities of PC9/GR cells while promoting apoptosis. Additionally, HOTAIR siRNA effectively suppressed tumor growth and downregulated the Hedgehog-GLi1 pathway and anti-apoptotic proteins, which were confirmed in animal experiments. Moreover, SA/HOTAIR siRNA exhibited superior inhibition of cellular and tumor functions compared to using HOTAIR siRNA alone. Clinical research findings indicated that monitoring the expression level of HOTAIR in the serum and urine samples of NSCLC patients before and after receiving EGFR-TKIs treatment can predict the efficacy of EGFR-TKIs to a certain extent. This study provided evidence that HOTAIR siRNA effectively mitigated the development of acquired resistance to EGFR-TKIs by inhibiting the Hedgehog-GLi1 pathway. Furthermore, it introduced a reliable and long-lasting drug delivery system for combating acquired resistance to EGFR-TKIs.

Keywords

Hox transcript antisense RNA epidermal growth factor receptor tyrosine kinase inhibitors gefitinib resistance hedgehog-GLi1 sodium alginate microspheres non-small cell lung cancer

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References

Shen

Xie

, et al. LncRNAs LCETRL3 and LCETRL4 at chromosome 4q12 diminish EGFR-TKIs efficiency in NSCLC through stabilizing TDP43 and EIF2S1. Signal Transduct Targeted Ther 2022; 7(1): 30.

Dong

Zhu

Liu

, et al. EGFR mutation mediates resistance to EGFR tyrosine kinase inhibitors in NSCLC: from molecular mechanisms to clinical research. Pharmacol Res 2021; 167: 105583.

Zhu

Chen

Liu

, et al. EMT-mediated acquired EGFR-TKI resistance in NSCLC: mechanisms and strategies. Front Oncol 2019; 9: 1044.

Wang

Ren

, et al. HOTAIR induces EGFR-TKIs resistance in non-small cell lung cancer through epithelial-mesenchymal transition. Lung Cancer 2020; 147: 99–105.

Oxnard

Arcila

Sima

, et al. Acquired resistance to EGFR tyrosine kinase inhibitors in EGFR-mutant lung cancer: distinct natural history of patients with tumors harboring the T790M mutation. Clin Cancer Res 2011; 17(6): 1616–1622.

Bai

Zhang

Yang

, et al. Blockade of hedgehog signaling synergistically increases sensitivity to epidermal growth factor receptor tyrosine kinase inhibitors in non-small-cell lung cancer cell lines. PLoS One 2016; 11(3): e0149370.

Nurwidya

Takahashi

Murakami

, et al. Acquired resistance of non-small cell lung cancer to epidermal growth factor receptor tyrosine kinase inhibitors. Respir Investig 2014; 52(2): 82–91.

Rotow

Bivona

. Understanding and targeting resistance mechanisms in NSCLC. Nat Rev Cancer 2017; 17(11): 637–658.

Pan

Wang

, et al. Cholesterol promotes EGFR-TKIs resistance in NSCLC by inducing EGFR/Src/Erk/SP1 signaling-mediated ERRalpha re-expression. Mol Cancer 2022; 21(1): 77.

10.

Juchum

Gunther

Laufer

. Fighting cancer drug resistance: opportunities and challenges for mutation-specific EGFR inhibitors. Drug Resist Updates 2015; 20: 12–28.

11.

Alam

Sohoni

Kalainayakan

, et al. Cyclopamine tartrate, an inhibitor of Hedgehog signaling, strongly interferes with mitochondrial function and suppresses aerobic respiration in lung cancer cells. BMC Cancer 2016; 16: 150.

12.

Zhou

Fan

, et al. WW45, a Gli1 binding protein, negatively regulated Hedgehog signaling in lung cancer. Oncotarget 2016; 7(42): 68966–68975.

13.

Bai

Jin

, et al. HOTAIR and androgen receptor synergistically increase GLI2 transcription to promote tumor angiogenesis and cancer stemness in renal cell carcinoma. Cancer Lett 2021; 498: 70–79.

14.

Xin

Fang

, et al. LncRNA HOTAIR: a potential prognostic factor and therapeutic target in human cancers. Front Oncol 2021; 11: 679244.

15.

Liu

Sun

, et al. The long non-coding RNA HOTAIR indicates a poor prognosis and promotes metastasis in non-small cell lung cancer. BMC Cancer 2013; 13: 464.

16.

Zhang

, et al. HOTAIR promotes gefitinib resistance through modification of EZH2 and silencing p16 and p21 in non-small cell lung cancer. J Cancer 2021; 12(18): 5562–5572.

17.

Huang

Liu

Kong

, et al. Synthesis of siRNA nanoparticles to silence plaque-destabilizing gene in atherosclerotic lesional macrophages. Nat Protoc 2022; 17(3): 748–780.

18.

Murata

Takashima

Toyoshima

, et al. Anti-tumor effects of anti-VEGF siRNA encapsulated with PLGA microspheres in mice. J Contr Release 2008; 126(3): 246–254.

19.

Zhang

Shen

, et al. Liver-targeted siRNA lipid nanoparticles treat hepatic cirrhosis by dual antifibrotic and anti-inflammatory activities. ACS Nano 2020; 14(5): 6305–6322.

20.

McMillan

Nguyen

Huynh

, et al. Hydrogel microspheres for spatiotemporally controlled delivery of RNA and silencing gene expression within scaffold-free tissue engineered constructs. Acta Biomater 2021; 124: 315–326.

21.

Dhamecha

Movsas

Sano

, et al. Applications of alginate microspheres in therapeutics delivery and cell culture: past, present and future. Int J Pharm 2019; 569: 118627.

22.

Hua

Zhang

, et al. Poly(lactic-co-glycolic acid) microsphere production based on quality by design: a review. Drug Deliv 2021; 28(1): 1342–1355.

23.

Zhang

Cheng

. Preparation of alginate-based biomaterials and their applications in biomedicine. Mar Drugs 2021; 19(5).

24.

Swastha

Varsha

Aravind

, et al. Alginate-based drug carrier systems to target inflammatory bowel disease: a review. Int J Biol Macromol 2023: 244: 125472.

25.

Arruda

Gonzalez

Finet

, et al. Modifying internal organization and surface morphology of siRNA lipoplexes by sodium alginate addition for efficient siRNA delivery. J Colloid Interface Sci 2019; 540: 342–353.

26.

Luo

, et al. Dental pulp stem cell-derived exosomes alleviate cerebral ischaemia-reperfusion injury through suppressing inflammatory response. Cell Prolif 2021; 54(8): e13093.

27.

Animal care and use guidelines update committee. Guidelines for the Care and use of laboratory animals 8th ed. Washington, DC: National Academy of Sciences Press 2011.

28.

Zhu

Ying

, et al. Alginate self-adhesive hydrogel combined with dental pulp stem cells and FGF21 repairs hemisection spinal cord injury via apoptosis and autophagy mechanisms. Chem Eng J 2021; 426: 130827.

29.

Chen

Liu

Zhang

, et al. LncRNA H19 downregulation confers erlotinib resistance through upregulation of PKM2 and phosphorylation of AKT in EGFR-mutant lung cancers. Cancer Lett 2020; 486: 58–70.

30.

Chen

Cheng

, et al. Long non-coding RNA CASC9 promotes gefitinib resistance in NSCLC by epigenetic repression of DUSP1. Cell Death Dis 2020; 11(10): 858.

31.

Zheng

Wang

, et al. LncRNA APCDD1L-AS1 induces icotinib resistance by inhibition of EGFR autophagic degradation via the miR-1322/miR-1972/miR-324-3p-SIRT5 axis in lung adenocarcinoma. Biomark Res 2021; 9(1): 9.

32.

Dai

Yin

, et al. LncRNA LINC00969 promotes acquired gefitinib resistance by epigenetically suppressing of NLRP3 at transcriptional and posttranscriptional levels to inhibit pyroptosis in lung cancer. Cell Death Dis 2023; 14(5): 312.

33.

Chen

Yang

Wang

, et al. Activation of the Hedgehog pathway mediates resistance to epidermal growth factor receptor inhibitors in non-small cell lung cancer. J Cancer 2022; 13(3): 987–997.

34.

Sun

Bai

. Hedgehog/Gli1 signal pathway facilitates proliferation, invasion, and migration of cutaneous SCC through regulating VEGF. Tumour Biol. 2016. doi:10.1007/s13277-016-5435-x

35.

Wasson

Ross

Wells

, et al. Long non-coding RNA HOTAIR induces GLI2 expression through Notch signalling in systemic sclerosis dermal fibroblasts. Arthritis Res Ther 2020; 22(1): 286.

36.

Han

Lee

Baek

, et al. Hedgehog signaling regulates the survival of gastric cancer cells by regulating the expression of Bcl-2. Int J Mol Sci 2009; 10(7): 3033–3043.

37.

Kong

Zhou

, et al. Targeting HOTAIR induces mitochondria related apoptosis and inhibits tumor growth in head and neck squamous cell carcinoma in vitro and in vivo. Curr Mol Med 2015; 15(10): 952–960.

38.

Dana

Chalbatani

Mahmoodzadeh

, et al. Molecular mechanisms and biological functions of siRNA. Int J Biomed Sci 2017; 13(2): 48–57.

39.

Zhou

Xiong

, et al. MSC based gene delivery methods and strategies improve the therapeutic efficacy of neurological diseases. Bioact Mater 2023; 23: 409–437.

40.

Abid

Uzair

Niazi

MBK

, et al. Bursting the virulence traits of MDR strain of Candida albicans using sodium alginate-based microspheres containing nystatin-loaded MgO/CuO nanocomposites. Int J Nanomed 2021; 16: 1157–1174.

41.

Martin

Calpena

Fernandez

, et al. Development of alginate microspheres as nystatin carriers for oral mucosa drug delivery. Carbohydr Polym 2015; 117: 140–149.

42.

Wells

Sheardown

. Extended release of high pI proteins from alginate microspheres via a novel encapsulation technique. Eur J Pharm Biopharm 2007; 65(3): 329–335.

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