The Effect of Recombinant Killer Factor (LF) and Protective Factor (mPA) Proteins on Human Cancer Cells

Subject Areas : Journal of Animal Biology

Mahboubeh Gholami ¹ (Department of Biology, Jahrom Branch, Islamic Azad University, Jahrom, Iran)
Majid Moghbeli ² (Department of Biology, Damghan Branch, Islamic Azad University, Damghan, Iran)
Farshid Kafilzadeh ³ (Department of Biology, Jahrom Branch, Islamic Azad University, Jahrom, Iran)
Mohammad Kargar ⁴ (Department of Biology, Jahrom Branch, Islamic Azad University, Jahrom, Iran)
Maryam Bikhof Torbati ⁵ (Department of Biology, Yadegar Imam Branch, Islamic Azad University, Tehran, Iran)

Received: 2021-01-03 Accepted : 2021-04-14 Published : 2022-02-20

Keywords: cancer, Bacillus subtilis, Recombinant proteins, Lethal factor, Protective factor,

Abstract :

Cancer causes about 13% of human deaths and has not yet been properly treated. Conventional cancer treatment methods such as surgery, chemotherapy, and radiotherapy have many side effects and in some cases have no effect on cancer treatment, so creating appropriate and targeted therapies can revolutionize cancer treatment. New methods include gene therapy and using immunotoxins, which can act up to a high percentage. This study aimed to investigate the effect of modified PA protein that binds only to cancer cells along with lethal factor (LF) on cancer cells. In this regard, recombinant mPA and LF proteins were isolated from recombinant Bacillus subtilis and after examining the presence of protein bands and their concentrations using SDS-PAGE and Bradford methods, respectively, the combined effect of different concentrations of these proteins on lung cancer cells, Breast, prostate and pancreas were examined by MTT method. The results showed that 75 ng of recombinant LF protein and 50 ng of recombinant mPA protein had the greatest effect on cancer cell death and caused the death of more than 98% of the studied cancer cells. With these results, it is hoped that a new dimension can be added to cancer treatment by producing targeted toxins such as mPA and LF.

References:

1. Abi-HabibJ., Singh R., Liu S., Bugge T.H., Leppla S.H., Frankel A.E.(2006), A Urokinase-Activated recombinant anthrax toxin is selectively cytotoxic to many human tumor cell types. Molecular Cancer Therapy, 5(10): 2556-2562.


2. Abrami L., Kunz B., Vander Goot F.G. (2010), Anthrax toxin triggers the activation of src-like kinases to mediate its own uptake. Proceedings of the National Academy of Sciences of the United States of America, 107: 1420-1424.


3. Bradford M.M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Annals in Biochemistry, 72: 248-254


4. Bromberg-White, Lee C.S., Duesbery N. 2010. Consequences and utility of the Zinc-Dependent metalloprotease activity of anthrax lethal toxin. Toxins, 5: 1038-1053.


5. Chen K.H., Liu S., Leysath C.E., Miller-Randolph S., Zhang Y., Fattah R., Bugge T.H., Leppla S.H. 2016. Anthrax toxin protective antigen variants that selectively utilize either the CMG2 or TEM8 receptors for cellular uptake and tumor targeting. Journal of Biology and Chemistry, 291: 22021-22029.


6. Duesbery N.S., Webb C.P., Leppla S.H. 1998. Proteolytic inactivation of MAP-Kinase-Kinase by Anthrax Lethal Factor. Science, 280:734-737.


7. Baillie L.W., Huwar T.B., Moore S. 2010. An anthrax subunit vaccine candidate based on protective region of B. anthracis protective antigen and lethal factor. Vaccine, 28: 6740-6748


8. Liu S., Schubert R.L., Bugge T.H., Leppla S.H. 2003. Anthrax toxin: structures, functions and tumour targeting. Expert Opinion on Biological Therapy, 5: 843-853.


9. Gholami M., Moghbeli M., Kafilzadeh, F., Kargar M., Torbati M.B., Tavizi A., Eslami Z. 2021. Production of recombinant lethal factor of Bacillus anthracis in Bacillus subtilis. Preparative Biochemistry and Biotechnology, 51(1): 9-15.


10. Liu S., Aaronson H., Mitola D.J., Leppla S.H., Bugge T.H. 2003. Potent antitumor activity of a urokinase-activated engineered anthrax toxin. PNAS, 2: 657-662.


11. Liu S., Moayeri M., Leppla S.H. 2014. Anthrax lethal and edema toxins in anthrax Trends in Microbiology, 6:317-325.


12. Martin EW, Buzza MS, Driesbaugh KH, Liu S, Fortenberry YM, Leppla SH, Antalis TM. 2015. Targeting the membrane-anchored serine protease testis in with a novel engineered anthrax toxin prodrug to kill tumor cells and reduce tumor burden. Oncotarget, 32: 33534-33553.


13. Minchinton A.I., Tannock I.F. 2006. Drug penetration in solid tumors. National Review of Cancer, 6:583–592.


14. Neil S. 2010. Engineering the perfect (bacterial) cancer therapy. Natural Review of Cancer, 10(11): 785-794.


15. Patyar S., Joshi R., Prasad Byrav D.R., Prakash A., Medhi B., Das B.K. 2010. Bacterial in cancer therapy: anovel experimental strategy. Journal of Biomedical Sciences, 17(1): 21.


16. Ryan R.M., Green J., Lewis C.E. 2006. Use of bacteria in anticancer therapies. Bioassays, 28(1): 84-94.


17. Sai L., Xiaoping X., Xin Z., Longjiang L., Qianmimng C., Jing L. 2014. Tumor‑targeting bacterial therapy: A potential treatment for oral cancer. Oncology Letters, 8(6): 2359-2366.


18. Sambrook J.F., Russell D.W. 2001. Molecular cloning: A Laboratory Manual, Cold spring harbor Laboratory Press, 2100 pp.


19. Smith H., Keppie J., Stanley J.L. 1995. The Chemical Basis of the Virulence of Bacillus anthracis. V. The Specific Toxin Produced by Bacillus anthracis in vivo. British Journal of Experimental Pathology, 5: 460-472.


20. St Jean A.T., Zhang M.M., Forbes N.S., 2008. Bacterial therapies: completing the cancer treatment toolbox. Current Opinion in Biotechnology, 19(5): 511-517.


21. Sun Z., Fu Y.X. and Peng H. 2018. Targeting tumor cells with antibodies enhances anti-tumor immunity. Biophysics Reports, 4(5): 243-253.


22. Taherianfard A., Hasan F., Bandehpour M., Mosaffa N., Mashhadi Abbas F., Hameed A. 2010. Cloning and expression of C-terminal of Clostridium perfringens type A enterotoxin and its biological activity. African Journal of Microbiology Research, 4(14): 1469-1474.


23. WeinN., Liu S., Zhang Y., McKenzie A.T., Leppla S.H. 2013. Tumor therapy with a urokinase plasminogen activator-activated anthrax lethal toxin alone and in combination with paclitaxel. Invest New Drugs. 31:206-212.


24. Williams S.A., Merchant R.F., Garrett-Mayer E., Isaacs J.T., Buckley J.T., Denmeade S.R. 2007. A prostate-specific antigen-activated channel-forming toxin as therapy for prostatic disease. Journal of National Cancer Institute, 99(5):376-385.


25. Xu J., Liu X.S., Zhou S.F., Wei M.Q. 2009. Combination of immunotherapy with anaerobic bacteria for immunogene therapy of solid tumours. Gene Therapy and Molecular Biology, 13: 36-52.


26. Yaghoobi H., Bandehpour M., Kazemi B. 2015. Designing and Cloning of cytolethal distending toxin B as Biological Tool against cancer. 10th International Breast Cancer Congress, p: 113