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Fusion of gelonin and anti-insulin-like growth factor-1 receptor (IGF-1R) affibody for enhanced brain cancer therapy

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Abstract

Owing to the extraordinary potency in inhibiting protein translation that could eventually lead to apoptosis of tumor cells, ribosome-inactivating proteins (RIPs) such as gelonin have been considered attractive drug candidates for cancer therapy. However, due to several critical obstacles (e.g., severe toxicity issues caused by a lack of selectivity in their mode of action and the low cytotoxicity via poor cellular uptake, etc.), clinical application of RIPs is yet far from being accomplished. To overcome these challenges, in the present study, we engineered gelonin fusion proteins with anti-insulin-like growth factor-1 receptor (IGF-1R) affibody (“IAFF”) via the genetic recombinant method and the SpyCatcher/SpyTag-mediated conjugation method. To this end, recombinant gelonin-anti-IGF-1R affibody (rGel-IAFF), gelonin-SpyCatcher (Gel-SpyCatcher) and SpyTag-IAFF fusion proteins were produced from the E. coli expression system, and gelonin-IAFF conjugate was synthesized by mixing Gel-SpyCatcher and SpyTag-IAFF. After preparation of both rGel-IAFF and Gel-IAFF conjugate, their components’ functionality was characterized in vitro. Our assay results confirmed that, while both Gel-IAFF and Gel-SpyCatcher retained equipotent N-glycosidase activity to that of gelonin, IAFF was able to selectively bind to IGF-1R overexpressed U87 MG brain cancer cells over low expression LNCaP cells. The results of cellular analyses showed that rGel-IAFF and Gel-IAFF conjugate both exhibited a greater cell uptake in the U87 MG cells than gelonin, but not in the LNCaP cells, yielding a significantly augmented cytotoxicity only in the U87 MG cells. Remarkably, rGel-IAFF and Gel-IAFF conjugate displayed 22- and 5.6-fold lower IC50 values (avg. IC50: 180 and 720 nM, respectively) than gelonin (avg. IC50: 4000 nM) in the U87 MG cells. Overall, the results of the present research demonstrated that fusion of gelonin with IAFF could provide an effective way to enhance the anti-tumor activity, while reducing the associated toxicity of gelonin.

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References

  • Allen TM (2002) Ligand-targeted therapeutics in anticancer therapy. Nat Rev Cancer 2:750–763

    Article  CAS  PubMed  Google Scholar 

  • Atkinson SF, Bettinger T, Seymour LW, Behr JP, Ward CM (2001) Conjugation of folate via gelonin carbohydrate residues retains ribosomal-inactivating properties of the toxin and permits targeting to folate receptor positive cells. J Biol Chem 276:27930–27935

    Article  CAS  PubMed  Google Scholar 

  • Bashyal S, Noh G, Keum T, Choi YW, Lee S (2016) Cell penetrating peptides as an innovative approach for drug delivery; then, present and the future. J Pharm Investig 46:205–220

    Article  CAS  Google Scholar 

  • Bertrand N, Wu J, Xu X, Kamaly N, Farokhzad OC (2014) Cancer nanotechnology: the impact of passive and active targeting in the era of modern cancer biology. Adv Drug Deliv Rev 66:2–25

    Article  CAS  PubMed  Google Scholar 

  • Chong LD (2017) Brain cancer therapy. Science 355:258

    PubMed  Google Scholar 

  • Domeradzka NE, Werten MW, Wolf FA, de Vries R (2016) Protein cross-linking tools for the construction of nanomaterials. Curr Opin Biotechnol 39:61–67

    Article  CAS  PubMed  Google Scholar 

  • Frejd FY, Kim KT (2017) Affibody molecules as engineered protein drugs. Exp Mol Med 49:e306

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Ham SH, Min KA, Shin MC (2017) Molecular tumor targeting of gelonin by fusion with F3 peptide. Acta Pharmacol Sin 38:897–906

    Article  CAS  PubMed  Google Scholar 

  • Huang X, Peng X, Wang Y, Wang Y, Shin DM, El-Sayed MA, Nie S (2010) A reexamination of active and passive tumor targeting by using rod-shaped gold nanocrystals and covalently conjugated peptide ligands. ACS Nano 4:5887–5896

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Kim CH, Lee SG, Kang MJ, Lee S, Choi YW (2017a) Surface modification of lipid-based nanocarriers for cancer cell-specific drug targeting. J Pharm Investig 47:203–227

    Article  CAS  Google Scholar 

  • Kim MC, Lin MM, Sohn Y, Kim JJ, Kang BS, Kim DK (2017b) Polyethyleneimine-associated polycaprolactone-Superparamagnetic iron oxide nanoparticles as a gene delivery vector. J Biomed Mater Res B Appl Biomater 105:145–154

    Article  CAS  PubMed  Google Scholar 

  • Lee TY, Park YS, Garcia GA, Sunahara RK, Woods JH, Yang VC (2012) Cell permeable cocaine esterases constructed by chemical conjugation and genetic recombination. Mol Pharm 9:1361–1373

    CAS  PubMed  PubMed Central  Google Scholar 

  • Lee JH, Sahu A, Jang C, Tae G (2015) The effect of ligand density on in vivo tumor targeting of nanographene oxide. J Control Release 209:219–228

    Article  CAS  PubMed  Google Scholar 

  • Li L, Fierer JO, Rapoport TA, Howarth M (2014) Structural analysis and optimization of the covalent association between SpyCatcher and a peptide Tag. J Mol Biol 426:309–317

    Article  CAS  PubMed  Google Scholar 

  • Luo HC, Li N, Yan L, Mai KJ, Sun K, Wang W, Lao GJ, Yang C, Zhang LM, Ren M (2017) Comparison of the cellular transport mechanism of cationic, star-shaped polymers and liposomes in HaCat cells. Int J Nanomedicine 12:1085–1096

    Article  PubMed  PubMed Central  Google Scholar 

  • Malik A, Sutantyo ML, Hapsari I, Sinurat AV, Purwati EM, Jufri M, Suryadi H (2016) Simultaneous identification and verification of gelatin type in capsule shells by electrophoresis and polymerase chain reaction. J Pharm Investig 46:475–485

    Article  CAS  Google Scholar 

  • Min KA, He H, Yang VC, Shin MC (2016) Construction and characterization of gelonin and saporin plasmids for toxic gene-based cancer therapy. Arch Pharm Res 39:677–686

    Article  CAS  PubMed  Google Scholar 

  • Olson JJ, Nayak L, Ormond DR, Wen PY, Kalkanis SN, Committee ACJG (2014) The role of cytotoxic chemotherapy in the management of progressive glioblastoma: a systematic review and evidence-based clinical practice guideline. J Neurooncol 118:501–555

    Article  CAS  PubMed  Google Scholar 

  • Reddington SC, Howarth M (2015) Secrets of a covalent interaction for biomaterials and biotechnology: SpyTag and SpyCatcher. Curr Opin Chem Biol 29:94–99

    Article  CAS  PubMed  Google Scholar 

  • Reiss K, Wang JY, Romano G, Furnari FB, Cavenee WK, Morrione A, Tu X, Baserga R (2000) IGF-I receptor signaling in a prostatic cancer cell line with a PTEN mutation. Oncogene 19:2687–2694

    Article  CAS  PubMed  Google Scholar 

  • Rock K, McArdle O, Forde P, Dunne M, Fitzpatrick D, O’Neill B, Faul C (2012) A clinical review of treatment outcomes in glioblastoma multiforme–the validation in a non-trial population of the results of a randomised Phase III clinical trial: has a more radical approach improved survival? Br J Radiol 85:e729–e733

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Sathornsumetee S, Reardon DA, Desjardins A, Quinn JA, Vredenburgh JJ, Rich JN (2007) Molecularly targeted therapy for malignant glioma. Cancer 110:13–24

    Article  PubMed  Google Scholar 

  • Schardt JS, Oubaid JM, Williams SC, Howard JL, Aloimonos CM, Bookstaver ML, Lamichhane TN, Sokic S, Liyasova MS, O’Neill M, Andresson T, Hussain A, Lipkowitz S, Jay SM (2017) Engineered multivalency enhances affibody-based HER3 inhibition and downregulation in cancer cells. Mol Pharm 14:1047–1056

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Schrot J, Weng A, Melzig MF (2015) Ribosome-inactivating and related proteins. Toxins (Basel) 7:1556–1615

    Article  CAS  Google Scholar 

  • Shin MC, Zhang J, David AE, Trommer WE, Kwon YM, Min KA, Kim JH, Yang VC (2013) Chemically and biologically synthesized CPP-modified gelonin for enhanced anti-tumor activity. J Control Release 172:169–178

    Article  CAS  PubMed  Google Scholar 

  • Shin MC, Zhao J, Zhang J, Huang Y, He H, Wang M, Min KA, Yang VC (2015) Recombinant TAT-gelonin fusion toxin: synthesis and characterization of heparin/protamine-regulated cell transduction. J Biomed Mater Res A 103:409–419

    Article  PubMed  Google Scholar 

  • Shin MC, Min KA, Cheong H, Moon C, Huang Y, He H, Yang VC (2016) Preparation and characterization of gelonin-melittin fusion biotoxin for synergistically enhanced anti-tumor activity. Pharm Res 33:2218–2228

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Shin MC, Min KA, Cheong H, Moon C, Huang Y, He H, Yang VC (2017) Tandem-multimeric F3-gelonin fusion toxins for enhanced anti-cancer activity for prostate cancer treatment. Int J Pharm 524:101–110

    Article  CAS  PubMed  Google Scholar 

  • Singh P, Alex JM, Bast F (2014) Insulin receptor (IR) and insulin-like growth factor receptor 1 (IGF-1R) signaling systems: novel treatment strategies for cancer. Med Oncol 31:805

    Article  PubMed  Google Scholar 

  • Smith PK, Krohn RI, Hermanson GT, Mallia AK, Gartner FH, Provenzano MD, Fujimoto EK, Goeke NM, Olson BJ, Klenk DC (1985) Measurement of protein using bicinchoninic acid. Anal Biochem 150:76–85

    Article  CAS  PubMed  Google Scholar 

  • Son JH, Hwang EC, Kim J (2016) Systematic analyses of the ultraviolet radiation resistance-associated gene product (UVRAG) protein interactome by tandem affinity purification. Arch Pharm Res 39:370–379

    Article  CAS  PubMed  Google Scholar 

  • Srinivasarao M, Galliford CV, Low PS (2015) Principles in the design of ligand-targeted cancer therapeutics and imaging agents. Nat Rev Drug Discov 14:203–219

    Article  CAS  PubMed  Google Scholar 

  • Stirpe F (2013) Ribosome-inactivating proteins: from toxins to useful proteins. Toxicon 67:12–16

    Article  CAS  PubMed  Google Scholar 

  • Stirpe F, Olsnes S, Pihl A (1980) Gelonin, a new inhibitor of protein synthesis, nontoxic to intact cells. Isolation, characterization, and preparation of cytotoxic complexes with concanavalin A. J Biol Chem 255:6947–6953

    CAS  PubMed  Google Scholar 

  • Su X, Cheng K, Liu Y, Hu X, Meng S, Cheng Z (2015) PET imaging of insulin-like growth factor type 1 receptor expression with a 64Cu-labeled Affibody molecule. Amino Acids 47:1409–1419

    Article  CAS  PubMed  Google Scholar 

  • Sudo K, Niikura K, Iwaki K, Kohyama S, Fujiwara K, Doi N (2017) Human-derived fusogenic peptides for the intracellular delivery of proteins. J Control Release 255:1–11

    Article  CAS  PubMed  Google Scholar 

  • Tolmachev V, Orlova A, Nilsson FY, Feldwisch J, Wennborg A, Abrahmsen L (2007) Affibody molecules: potential for in vivo imaging of molecular targets for cancer therapy. Expert Opin Biol Ther 7:555–568

    Article  CAS  PubMed  Google Scholar 

  • Ul Ain Q, Lee JH, Woo YS, Kim YH (2016) Effects of protein transduction domain (PTD) selection and position for improved intracellular delivery of PTD-Hsp27 fusion protein formulations. Arch Pharm Res 39:1266–1274

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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Acknowledgements

This work was supported by the grants from Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education, Science and Technology (NRF-2015R1C1A1A02036781 and NRF-2017R1C1B5015491). This study was also supported by the Inje University Research Fund, Republic of Korea (Grant #: 20160539). We thank Dr. Wolfgang E. Trommer (University of Kaiserslautern, Germany) for the gelonin expression vector (pET28a-Gel).

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  1. Songhee Ham and Kyoung Ah Min have contributed equally to this work.

Authors and Affiliations

  1. College of Pharmacy and Research Institute of Pharmaceutical Sciences, Gyeongsang National University, 501 Jinju Daero, Jinju, Gyeongnam, 660-751, Republic of Korea

    Songhee Ham & Meong Cheol Shin

  2. College of Pharmacy and Inje Institute of Pharmaceutical Sciences and Research, Inje University, 197 Injero, Gimhae, Gyeongnam, 621-749, Republic of Korea

    Kyoung Ah Min

  3. Department of Ophthalmology, Inje University College of Medicine, 75 Bokji-ro, Busanjin-gu, Busan, 47392, Republic of Korea

    Jae Wook Yang

  4. T2B infrastructure center for ocular diseases, Inje University Busan Paik Hospital, 75 Bokji-ro, Busanjin-gu, Busan, 47392, Republic of Korea

    Kyoung Ah Min & Jae Wook Yang

  5. College of Pharmacy and Research Institute of Pharmaceutical Sciences, Gyeongsang National University, 501 Jinju Daero, Jinju, Gyeongnam, 52828, Republic of Korea

    Meong Cheol Shin

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  1. Songhee Ham
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Correspondence to Meong Cheol Shin.

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Ham, S., Min, K.A., Yang, J.W. et al. Fusion of gelonin and anti-insulin-like growth factor-1 receptor (IGF-1R) affibody for enhanced brain cancer therapy. Arch. Pharm. Res. 40, 1094–1104 (2017). https://doi.org/10.1007/s12272-017-0953-7

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