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Frontiers of Medicine

ISSN 2095-0217

ISSN 2095-0225(Online)

CN 11-5983/R

Postal Subscription Code 80-967

2018 Impact Factor: 1.847

Front. Med.    2019, Vol. 13 Issue (6) : 646-657
NES1/KLK10 and hNIS gene therapy enhanced iodine-131 internal radiation in PC3 proliferation inhibition
Jiajia Hu1, Wenbin Shen1, Qian Qu1, Xiaochun Fei2, Ying Miao1, Xinyun Huang1, Jiajun Liu1, Yingli Wu3(), Biao Li1()
1. Department of Nuclear Medicine, Ruijin Hospital Affiliated to Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
2. Department of Pathology, Ruijin Hospital Affiliated to Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
3. Hongqiao International Institute of Medicine, Shanghai Tongren Hospital/Faculty of Basic Medicine, Chemical Biology Division of Shanghai Universities E-Institutes, Key Laboratory of Cell Differentiation and Apoptosis of the Chinese Ministry of Education, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
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NES1 gene is thought to be a tumor-suppressor gene. Our previous study found that overexpression of NES1 gene in PC3 cell line could slow down the tumor proliferation rate, associated with a mild decrease in BCL-2 expression. The BCL-2 decrease could increase the sensitivity of radiotherapy to tumors. Thus, we supposed to have an “enhanced firepower” effect by combining overexpressed NES1 gene therapy and 131I radiation therapy uptake by overexpressed hNIS protein. We found a weak endogenous expression of hNIS protein in PC3 cells and demonstrated that the low expression of hNIS protein in PC3 cells might be the reason for the low iodine uptake. By overexpressing hNIS in PC3, the radioactive iodine uptake ability was significantly increased. Results of in vitro and in vivo tumor proliferation experiments and 18F-fluorothymidine (18F-FLT) micro-positron emission tomography/computed tomography (micro-PET/CT) imaging showed that the combined NES1 gene therapy and 131I radiation therapy mediated by overexpressed hNIS protein had the best tumor proliferative inhibition effect. Immunohistochemistry showed an obvious decrease of Ki-67 expression and the lowest BCL-2 expression. These data suggest that via inhibition of BCL-2 expression, overexpressed NES1 might enhance the effect of radiation therapy of 131I uptake in hNIS overexpressed PC3 cells.

Keywords androgen-independent prostate cancer      normal epithelial cell-specific 1/kallikrein 10      sodium/iodide symporter      radiation therapy      proliferation     
Corresponding Authors: Yingli Wu,Biao Li   
Just Accepted Date: 20 March 2019   Online First Date: 24 May 2019    Issue Date: 16 December 2019
 Cite this article:   
Jiajia Hu,Wenbin Shen,Qian Qu, et al. NES1/KLK10 and hNIS gene therapy enhanced iodine-131 internal radiation in PC3 proliferation inhibition[J]. Front. Med., 2019, 13(6): 646-657.
Fig.1  Expression of NES1 and hNIS protein in four kinds of PC3 cell lines and verification of function of overexpressed hNIS protein in PC3 cell lines by radioactive iodine uptake related studies in vitro and in vivo. (A) Western blot confirmed the obvious expression of NES1 and hNIS protein in PC3-NES1-hNIS cell line and hNIS protein in PC3-hNIS cell line. Interestingly, weak expression of hNIS protein in PC3-NES1 and PC3-CON cell lines were also detected. (B) The function of overexpressed hNIS protein in PC3-NES1-hNIS and PC3-hNIS cell lines was verified by 125I uptake study. Less iodine uptake capacity could be detected in PC3-CON and PC3-NES1 cell lines. (C) In vitro iodine uptake NaClO4 inhibition study showed that the iodine uptake capacity of PC3-NES1-hNIS and PC3-hNIS cell lines could be completely inhibited by 30 mmol/L NaClO4. (D) In vitro iodine efflux study showed that Na125I was rapidly effluxed from both PC3-NES1-hNIS and PC3-hNIS cell lines, maintaining for approximately 1 h. (E–H) Animal SPECT imaging further confirmed the iodine uptake ability of hNIS protein in vivo. This observation showed that the radioactive iodine uptake of subcutaneous PC3-NES1-hNIS and PC3-hNIS xenografts (red circle and yellow circle) was significantly higher than that of PC3-NES1 and PC3-CON xenografts, respectively (green circle and purple circle). Radioiodine accumulation in PC3-NES1 and PC3-CON xenografts was almost invisible. (I) Quantitative analysis by the automatic g-counter demonstrated a significantly high radioactivity in PC3-hNIS and PC3-NES1-hNIS xenograft tissue compared with that in muscle tissue (17.8-fold and 11.8-fold, P<0.001), a low radioactivity in PC3-NES1, and PC3-CON xenograft tissue compared with that in muscle tissue (3.0-fold respectively, P<0.01). The iodine uptake capacity of PC3-hNIS and PC3-NES1-hNIS xenograft was much higher than that of PC3-CON and PC3-NES1 xenograft (P<0.01), and the uptake capacity of PC3-hNIS was 1.5-fold higher than that of PC3-NES1-hNIS (P<0.05).
Fig.2  Best therapeutic effect of the combined treatment of NES1 and 131I on PC3 cell lines and xenograft tumors. (A) CCK-8 cell proliferation assay showed an obvious inhibitive effect in the single NES1 gene therapy group PC3-NES1 (P<0.001), single 131I treatment absorbed by the hNIS protein group PC3-hNIS with 131I (P<0.01), and the combined treatment group PC3-NES1-hNIS cell line with 131I (P<0.001). The combined treatment group grew significantly slower than the single NES1 gene treatment group PC3-NES1 cell line (P<0.001) and significantly slower than the PC3-hNIS cell line treated with 131I (P<0.001). (B) Cell clone formation assay, which lasted for 16 days, showed the most significant proliferative inhibition in PC3-NES1-hNIS cells treated with 131I. A significant inhibition effect was also found in the PC3-NES1 group and the PC3-hNIS group with single 131I treatment. The finding was consistent with the result of CCK-8 cell proliferation assay. (C) Quantitative analysis of cell clone formation assay by Image-Pro Plus software showed the same result. Compared with the PC3-CON group, the combined treatment group significantly grew the slowest (P<0.001). A significant inhibitive effect was also observed in the PC3-NES1 group (P<0.001) and the PC3-hNIS group treated with 131I (P<0.01). Compared with the PC3-NES1 group and the PC3-hNIS group treated with 131I, the combined treatment group grew significantly the slowest (P<0.001). (D) Tumor volume line graph showed a proliferative inhibition effect on PC3-NES1 and PC3-NES1-hNIS xenograft from the beginning. At days 32 and 46, 131I systematic therapy was performed twice, with an injection of 37 MBq each. An obvious therapeutic effect could be found in PC3-hNIS-NES1 with the 131I groups (P<0.01). The growth of PC3-NES1 xenograft was always slower than that of the PC3-CON group (P<0.05). The growth of PC3-hNIS xenograft with 131I treatment was also slower than that of the PC3-CON group, but the difference was not significant enough. (E) Nude mice body weight line graph showed that the body weight change of each group of nude mice had no obvious significance. However, those loaded with PC3-NES1 and PC3-hNIS-NES1 xenografts increased slightly faster than those loaded with PC3-CON and PC3-hNIS. After day 24, the weight of each group decreased. After 131I treatment, the weight of PC3-hNIS-NES1 with 131I group declined the slowest (P<0.01). (F) General images showed that the tumor volume of PC3-hNIS-NES1 with the 131I groups was the smallest. Single NES1 gene therapy and single 131I of radiation therapy mediated by the hNIS protein overexpressed groups also had an inhibitive effect compared with the control groups. (G) Tumor inhibition rate (TIR) (mean%±SD%) was as follows: 83.54%±8.79%, 51.91%±6.90%, and 32.46%±17.97% for PC3-CON with PBSvs. PC3-hNIS-NES1 with 131I, PC3-CON with PBS vs. PC3-NES1 with PBS, PC3-hNIS with PBS vs. PC3-hNIS with 131I, respectively. Compared with single NES1 gene therapy and single radioactive iodine-131 treatment, combined treatment had the best tumor inhibition rate (P<0.01 and P<0.05, respectively).
Fig.3  Combined therapy of NES1 and 131I in PC3 xenograft inhibited the tumor proliferation associated with downregulation of BCL-2 expression. (A)18F-FLT micro-PET/CT imaging, which could evaluate the tumor proliferation in vivo, showed that the combined therapy group (PC3-hNIS-NES1 with 131I) obviously decreased the volume of xenograft and the 18F-FLT uptake. The volume of PC3-CON xenograft with and without 131I treatment was large. The uptake was also high. (B) Bar chart of 18F-FLT uptake in xenograft showed that the SUVmax of PC3-NES1-hNIS xenograft tumors with 131I treatment was the lowest in these eight groups. Compared with the PC3-CON group and two single treatment groups, the difference was significant (P<0.01, P <0.05, and P <0.05, respectively). Single radioactive iodine-131 treatment could also decrease the SUVmax of 18F-FLT uptake in xenograft tissue (P<0.05). Single NES1 gene therapy could decrease the 18F-FLT uptake, but the difference was not so significant. (C) Immunohistochemistry showed an obvious decrease of Ki-67 expression in two single-treated groups and PC3-NES1-hNIS with the 131I group. The latter showed the lowest expression. In addition, the BCL-2 expression in the xenograft tissue of both single-treated groups and PC3-NES1-hNIS with the 131I group decreased obviously. The latter was the lowest.
1 RLMK Siegel, A Jemal. Cancer Statistics, 2017. CA Cancer DJ Clin 2017; 67: 23 pmid: 28055103
2 R Chen, DD Sjoberg, Y Huang, L Xie, L Zhou, D He, AJ Vickers, Y Sun; Chinese Prostate Cancer Consortium; Prostate Biopsy Collaborative Group. Prostate specific antigen and prostate cancer in Chinese men undergoing initial prostate biopsies compared with western cohorts. J Urol 2017; 197(1): 90–96 pmid: 27593477
3 JL Mohler, PW Kantoff, AJ Armstrong, RR Bahnson, M Cohen, AV D’Amico, JA Eastham, CA Enke, TA Farrington, CS Higano, EM Horwitz, CJ Kane, MH Kawachi, M Kuettel, TM Kuzel, RJ Lee, AW Malcolm, D Miller, ER Plimack, JM Pow-Sang, D Raben, S Richey, M Roach 3rd, E Rohren, S Rosenfeld, E Schaeffer, EJ Small, G Sonpavde, S Srinivas, C Stein, SA Strope, J Tward, DA Shead, M Ho; National Comprehensive Cancer Network. Prostate cancer, version 2.2014. J Natl Compr Canc Netw 2014; 12(5): 686–718 pmid: 24812137
4 A Heidenreich, G Aus, M Bolla, S Joniau, VB Matveev, HP Schmid, F Zattoni; European Association of Urology. EAU guidelines on prostate cancer. Eur Urol 2008; 53(1): 68–80 pmid: 17920184
5 SB Williams, J Huo, K Chamie, MC Smaldone, CD Kosarek, JE Fang, LA Ynalvez, SP Kim, KE Hoffman, SH Giordano, BF Chapin. Discerning the survival advantage among patients with prostate cancer who undergo radical prostatectomy or radiotherapy: the limitations of cancer registry data. Cancer 2017; 123(9): 1617–1624 pmid: 28099688
6 V Pagliarulo, S Bracarda, MA Eisenberger, N Mottet, FH Schröder, CN Sternberg, UE Studer. Contemporary role of androgen deprivation therapy for prostate cancer. Eur Urol 2012; 61(1): 11–25 pmid: 21871711
7 M Bolla, TM de Reijke, G Van Tienhoven, AC Van den Bergh, J Oddens, PM Poortmans, E Gez, P Kil, A Akdas, G Soete, O Kariakine, EM van der Steen-Banasik, E Musat, M Piérart, ME Mauer, L Collette; EORTC Radiation Oncology Group and Genito-Urinary Tract Cancer Group. Duration of androgen suppression in the treatment of prostate cancer. N Engl J Med 2009; 360(24): 2516–2527 pmid: 19516032
8 TM Amaral, D Macedo, I Fernandes, L Costa. Castration-resistant prostate cancer: mechanisms, targets, and treatment. Prostate Cancer 2012; 2012: 327253 pmid: 22530130
9 E Basch, DA Loblaw, TK Oliver, M Carducci, RC Chen, JN Frame, K Garrels, S Hotte, MW Kattan, D Raghavan, F Saad, ME Taplin, C Walker-Dilks, J Williams, E Winquist, CL Bennett, T Wootton, RB Rumble, SB Dusetzina, KS Virgo. Systemic therapy in men with metastatic castration-resistant prostate cancer: American Society of Clinical Oncology and Cancer Care Ontario clinical practice guideline. J Clin Oncol 2014; 32(30): 3436–3448 pmid: 25199761
10 T Zhang, J Zhu, DJ George, AJ Armstrong. Enzalutamide versus abiraterone acetate for the treatment of men with metastatic castration-resistant prostate cancer. Expert Opin Pharmacother 2015; 16(4): 473–485 pmid: 25534660
11 D Robinson, EM Van Allen, YM Wu, N Schultz, RJ Lonigro, JM Mosquera, B Montgomery, ME Taplin, CC Pritchard, G Attard, H Beltran, W Abida, RK Bradley, J Vinson, X Cao, P Vats, LP Kunju, M Hussain, FY Feng, SA Tomlins, KA Cooney, DC Smith, C Brennan, J Siddiqui, R Mehra, Y Chen, DE Rathkopf, MJ Morris, SB Solomon, JC Durack, VE Reuter, A Gopalan, J Gao, M Loda, RT Lis, M Bowden, SP Balk, G Gaviola, C Sougnez, M Gupta, EY Yu, EA Mostaghel, HH Cheng, H Mulcahy, LD True, SR Plymate, H Dvinge, R Ferraldeschi, P Flohr, S Miranda, Z Zafeiriou, N Tunariu, J Mateo, R Perez-Lopez, F Demichelis, BD Robinson, M Schiffman, DM Nanus, ST Tagawa, A Sigaras, KW Eng, O Elemento, A Sboner, EI Heath, HI Scher, KJ Pienta, P Kantoff, JS de Bono, MA Rubin, PS Nelson, LA Garraway, CL Sawyers, AM Chinnaiyan. Integrative clinical genomics of advanced prostate cancer. Cell 2015; 161(5): 1215–1228 pmid: 26000489
12 Cancer Genome Atlas Research Network. The molecular taxonomy of primary prostate cancer. Cell 2015; 163: 1011–1025 pmid: 26544944
13 XL Liu, DE Wazer, K Watanabe, V Band. Identification of a novel serine protease-like gene, the expression of which is down-regulated during breast cancer progression. Cancer Res 1996; 56(14): 3371–3379
pmid: 8764136
14 J Hu, H Lei, X Fei, S Liang, H Xu, D Qin, Y Wang, Y Wu, B Li. NES1/KLK10 gene represses proliferation, enhances apoptosis and down-regulates glucose metabolism of PC3 prostate cancer cells. Sci Rep 2015; 5(1): 17426 pmid: 26616394
15 PE Czabotar, G Lessene, A Strasser, JM Adams. Control of apoptosis by the BCL-2 protein family: implications for physiology and therapy. Nat Rev Mol Cell Biol 2014; 15(1): 49–63 pmid: 24355989
16 D Zhang, Y Cui, L Niu, X Xu, K Tian, CY Young, H Lou, H Yuan. Regulation of SOD2 and b-arrestin1 by interleukin-6 contributes to the increase of IGF-1R expression in docetaxel resistant prostate cancer cells. Eur J Cell Biol 2014; 93(7): 289–298 pmid: 24939178
17 M Rajecki, M Sarparanta, T Hakkarainen, M Tenhunen, I Diaconu, V Kuhmonen, K Kairemo, A Kanerva, AJ Airaksinen, A Hemminki. SPECT/CT imaging of hNIS-expression after intravenous delivery of an oncolytic adenovirus and 131I. PLoS One 2012; 7(3): e32871 pmid: 22412937
18 KN Barton, H Stricker, MA Elshaikh, J Pegg, J Cheng, Y Zhang, KC Karvelis, M Lu, B Movsas, SO Freytag. Feasibility of adenovirus-mediated hNIS gene transfer and 131I radioiodine therapy as a definitive treatment for localized prostate cancer. Mol Ther 2011; 19(7): 1353–1359 pmid: 21587209
19 X Chen, JY Wong, P Wong, EH Radany. Low-dose valproic acid enhances radiosensitivity of prostate cancer through acetylated p53-dependent modulation of mitochondrial membrane potential and apoptosis. Mol Cancer Res 2011; 9(4): 448–461 pmid: 21303901
20 D Ezekwudo, R Shashidharamurthy, D Devineni, E Bozeman, R Palaniappan, P Selvaraj. Inhibition of expression of anti-apoptotic protein Bcl-2 and induction of cell death in radioresistant human prostate adenocarcinoma cell line (PC-3) by methyl jasmonate. Cancer Lett 2008; 270(2): 277–285 pmid: 18573594
21 J Goyal, KM Smith, JM Cowan, DE Wazer, SW Lee, V Band. The role for NES1 serine protease as a novel tumor suppressor. Cancer Res 1998; 58(21): 4782–4786
pmid: 9809976
22 B Li, J Goyal, S Dhar, G Dimri, E Evron, S Sukumar, DE Wazer, V Band. CpG methylation as a basis for breast tumor-specific loss of NES1/kallikrein 10 expression. Cancer Res 2001; 61(21): 8014–8021
pmid: 11691827
23 KA Ahmed, BJ Davis, TM Wilson, GA Wiseman, MJ Federspiel, JC Morris. Progress in gene therapy for prostate cancer. Front Oncol 2012; 2: 172 pmid: 23181221
24 C Spitzweg, AB Dietz, MK O’Connor, ER Bergert, DJ Tindall, CY Young, JC Morris. In vivo sodium iodide symporter gene therapy of prostate cancer. Gene Ther 2001; 8(20): 1524–1531 pmid: 11704812
25 MA Trujillo, MJ Oneal, S McDonough, R Qin, JC Morris. A steep radioiodine dose response scalable to humans in sodium-iodide symporter (NIS)-mediated radiovirotherapy for prostate cancer. Cancer Gene Ther 2012; 19(12): 839–844 pmid: 23037808
26 LM Zhao, AX Pang. Iodine-131 treatment of thyroid cancer cells leads to suppression of cell proliferation followed by induction of cell apoptosis and cell cycle arrest by regulation of B-cell translocation gene 2-mediated JNK/NF-kB pathways. Braz J Med Biol Res 2017; 50(1): e5933 pmid: 28099584
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