Optimization of Anti-CXCL10 Nanobody Expression Using Response Surface Methodology and Evaluation of its Anti-metastatic Effect on Breast Cancer cells

  • Tahereh Sadeghian-Rizi
  • Mahdi Behdani
  • Fateme Naghavi-al-hosseini
  • Seyedeh Simin Dakhilpour
  • Hossein Khanahmad
  • Ali Jahanian-NajafabadiEmail author


Some chemokines and chemokine receptors play important roles in various types of autoimmune diseases, infectious diseases and cancer metastasis. Hence, production of neutralizing antibodies against them are under active investigation. We previously developed a nanobody against CXCL10, designated as 3NB12, which can be expressed by E. coli cells. In the present study, we carried out a detailed study to optimize its expression using response surface methodology based on manipulation of three variables, including temperature, IPTG concentration, and post-induction time. In addition, upon expression and purification of the nanobody, it was also used to evaluate its inhibitory effects on migration of CXCR3 overexpressing MDA-MB-231 breast cancer cells. Seventeen experiments were designed. Total protein of the designed experiments was assayed by SDS-PAGE, followed by size exclusion chromatography to qualify and quantify the relative concentration of the nanobody in the optimized expression condition. The model designed according to the Box Behnken method predicted maximum 3NB12 expression at 28.5 °C, a post-induction time length of 15 h, and 0.9 mM IPTG. Chemotaxic assessment results showed that 3NB12 potently inhibits migration of the cells which has an important role in methastasis of breast cancer. Taken together, a reasonable amount of the nanobody could be produced according to the present study for being used in later in vitro and in vivo studies to further evaluate its anti metastatic and also anti-inflamatory effects.


Nanobody Chemokine CXCL10 CXCR3 Optimization of expression Response surface methodology 



The content of this paper is extracted from the Ph.D thesis NO. 394942 and research project NO. 195151 which was financially supported by the Research Department of Isfahan University of Medical Sciences, Isfahan, I.R. Iran.


The content of this paper is extracted from the Ph.D thesis NO. 394942 and research project NO. 195151 which was financially supported by the Research Department of Isfahan University of Medical Sciences, Isfahan, I.R. Iran.

Compliance with Ethical Standards

Conflict of interest

Tahereh Sadeghian-Rizi, Mahdi Behdani, Fateme Naghavi-al-hosseini, Seyedeh Simin Dakhilpour, Hossein Khanahmad, and Ali Jahanian-Najafabadi declare that they have no conflict of interest.

Ethical Approval

This article does not contain any studies with human participants or animals performed by any of the authors.


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Copyright information

© Springer Nature B.V. 2019

Authors and Affiliations

  • Tahereh Sadeghian-Rizi
    • 1
  • Mahdi Behdani
    • 2
  • Fateme Naghavi-al-hosseini
    • 3
  • Seyedeh Simin Dakhilpour
    • 4
  • Hossein Khanahmad
    • 5
  • Ali Jahanian-Najafabadi
    • 1
    Email author
  1. 1.Department of Pharmaceutical Biotechnology, School of Pharmacy and Pharmaceutical SciencesIsfahan University of Medical SciencesIsfahanIran
  2. 2.Biotechnology Research Center, Medical Biotechnology Research Center, Pasteur Institute of IranTehranIran
  3. 3.Student Research Committee, School of Pharmacy and Pharmaceutical SciencesIsfahan University of Medical SciencesIsfahanIran
  4. 4.Traditional Medicine and Medicinal PlantsIranian Academy Center for Education, Culture and Research (ACECR), Medicinal Herbal Center, Ardabil BranchArdabilIran
  5. 5.Department of Genetics and Molecular BiologyIsfahan University of Medical ScienceIsfahanIran

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